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Journal of Neuropathology & Experimental Neurology:
doi: 10.1097/NEN.0b013e3182227917
Abstracts

American Association of Neuropathologists, Inc. Abstracts of the 87th Annual Meeting June 23-26, 2011 Seattle, WA American Association of Neuropathologists, Inc. Abstracts of the 87th Annual Meeting June 23-26, 2011 Seattle, WA

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PLATFORM SESSION 1: TUMORS - GLIAL Friday 8-10 a.m. June 24, 2011

1 Integrated Analysis of Necrosis, Angiogenesis, and Gene Expression in Glioblastoma

Daniel Brat, Lee Cooper, Carlos Moreno, Candace Chisolm, Christina Appin, David Gutman, Jun Kong, Tahsin Kurc, Joel Saltz. Emory University

We analyzed gene expression and genetic correlates of angiogenesis and necrosis in glioblastoma (GBM) using both histology and molecular data from The Cancer Genome Atlas (TCGA). Frozen sections of 88 GBM samples used for molecular studies were manually marked to identify the surface area occupied by necrosis and angiogenesis as a percentage of total tissue area. Gene expression from the the HT-HGU133A platform was analyzed using Significance Analysis of Microarrays (SAM) Cox Proportional Hazards modeling to identify mRNAs significantly associated with extent of necrosis and/or angiogenesis using a false discovery rate cutoff of < 5%. Significant genes were submitted to Ingenuity Pathway Analysis (IPA) to identify pathways enriched with mRNA correlates. SAM analysis of necrosis identified 2184 correlated genes. Among those positively correlated were factors identified as master regulators of the mesenchymal tumor subtype, including C/EBP-B, C/EBP-D, STAT3, FOSL2, and RUNX1 (Carro MS, et al. Nature 263: 318-25, 2010). IPA analysis of genes correlated with necrosis identified significantly enriched canonical pathways including HIF-1a (p = 3.0e-7), NFκB (p = 1.4e-3), IL-6 (p = 6.9e-6), FGF (p = 2.7e-5), ERK/MAPK (p = 1.2e-4), Protein Kinase A signaling (p = 1.9e-4), Thrombin signaling (p = 5.2e-3), and HGF (p = 0.023) signaling. SAM analysis of degree of angiogenesis identified 188 significantly correlated genes, none of which were mesenchymal master regulators. IPA analysis of genes correlated with angiogenesis identified significantly enriched canonical pathways including HIF1a signaling (p = 2.3e-4).The mesenchymal tumor subtype was significantly enriched with high percent-necrosis samples (29% samples with > 25% necrosis) compared to the other three tumor subtypes (3% samples with > 25% necrosis)(p = 0.025). Analysis of extent of angiogenesis and tumor subtype did not reveal statistically significant associations, but we noted that all extreme outliers (percent-angiogenesis > 8%) were mesenchymal subtype samples.

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2 Genome-Wide Chromatin Landscape of Glioblastoma Cancer Stem Cells Highlights an Aberrant Network of Transcription Factors

Mario Suvà. Department of Pathology, Andrew Chi. Department of Neurology; Esther Rheinbay. Department of Pathology; Hiro Wakimoto. Department of Neurosurgery; Miguel Rivera, Ozgur Oksuz. Department of Pathology; Samuel Rabkin. Department of Neurosurgery; Robert Martuza. Department of Neurosurgery; David Louis, Bradley Bernstein. Department of Pathology Massachusetts General Hospital

Glioblastoma (GBM) is the most common malignant adult brain tumor and despite concomitant chemo- and radiotherapy, its prognosis remains poor. GBM has recently been shown to contain a primitive population of cancer stem cells (CSC) with exclusive tumor-initiating potential. These cells show high chemotherapy and radiation resistance, raising the urgent need for alternative therapeutic strategies. Epigenetic mechanisms play an important role in stem cell maintenance, but little is known on their role in GBM CSC. We have used chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) to profile a panel of histone modifications in GBM CSCs and matched traditional GBM cell lines from different individuals, as well as in human neural stem cells and primary human astrocytes. Global comparative analysis of enrichment patterns for active and repressive promoter marks together with transcriptional elongation marks reveals aberrant activation of a set of transcription factors (TFs) in CSC. Overexpression experiments in primary astrocytes establish a functional network of glioma TFs and highlight a connection to the Wnt signaling pathway. Knock-down experiments of TFs in GBM CSC followed by in vitro and in vivo functional assays confirm the importance of these TFs for CSC maintenance in GBM. Immunohistochemistry on tumor samples shows expression of these TFs by cancer cells co-expressing stem cells markers, underscoring TFs role in specific subpopulations of cancer cells. In summary, we present the chromatin-landscape of GBM CSC, a novel approach that allows detection of gene dysregulation independently of expression level and demonstrate its power in identifying aberrant TFs activation. We further demonstrate that some of these genes are putative CSC markers, are functionally important for GBM CSC and are potential targets for therapeutic intervention.

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3 Correlates of Mass Spectrometric Measurements of Gadolinium in Gliomas Undergoing Fluorescence Guided Resection

Brent Harris. Georgetown University School of Medicine; Pablo Valdes. Dartmouth Medical School; Ziev Moses. Dartmouth Medical School; Anthony Kim, Carolyn Niu, Marco Brantsch, Brian Wilson. University of Toronto; Keith Paulsen. Dartmouth Thayer School of Engineering; David Roberts. Dartmouth Medical School

Gliomas are diffusely infiltrating and highly heterogeneous tumors. Achieving maximal tumor removal is a major goal in resection of gliomas. The use of 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PpIX) fluorescence-guided resection (FGR) is increasing in clinical interest for resection of gliomas. Previous studies have shown a correlation between levels of contrast-enhancement on MR imaging in gliomas and intraoperative, qualitative PpIX fluorescence. In this study we evaluated whether highly sensitive and quantitative ex vivo tissue measurements of PpIX concentrations identify regions of blood-brain barrier (BBB) breakdown in low- and high-grade gliomas beyond the capabilities of current fluorescence imaging in patients undergoing FGR. Specimens were collected and processed for ex vivo neuropathological analysis, PpIX fluorimetry to measure PpIX concentrations (CPpIX), and for inductively coupled plasma mass spectrometry (ICP-MS) to quantify gadolinium concentrations (CGd). CGd serves as a quantitative surrogate marker of BBB breakdown, which as expected, correlates with the degree of tissue malignancy in gliomas (i.e., strong correlation with histopathological score based on WHO grading criteria (r=0.54, p < 0.0001)). Intraoperative qualitative levels of fluorescence showed significantly higher levels of CPpIX and CGd. We observed a strong correlation between BBB breakdown (i.e., using CGd as a biomarker) and CPpIX (r=0.58, p < 0.0001). There were also high correlations of microvascular density (as assessed by blinded CD31 immunohistochemistry counts) with CGd (r= 0.71, p<0.0001) and CPpIX (r=0.55, p < 0.0001). Previous studies have shown a correlation between contrast-enhancement on MRI (i.e., BBB break-down) and qualitative levels of PpIX fluorescence. To date, no study has performed a quantitative assessment associating BBB breakdown and PpIX tissue levels. These results corroborate the association between PpIX fluorescence and BBB breakdown. More importantly, they demonstrate that CGd, used as a quantitative marker of malignant changes associated with BBB breakdown, correlates with increasing tissue levels of quantitative CPpIX.

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4 BRAF-KIAA1549 Fusion Predicts Better Clinical Outcome in Pediatric Low Grade Astrocytoma

Cynthia Hawkins, Erin Walker, Nequesha Mohamed, Iris Fried. The Hospital for Sick Children; Nada Jabado. Montreal Children's Hospital; Uri Tabori. The Hospital for Sick Children

Purpose: Recent studies have revealed that the majority of pediatric low grade astrocytomas (PLGA) harbor the BRAF-KIAA1549 (B-K) fusion gene resulting in constitutive activation of the RAS/MAPK pathway. However, the clinical significance of this genetic alteration is yet to be determined. We aimed to test the prognostic role of the B-K fusion in progression of incompletely resected PLGA.

Patients and Methods: We retrospectively identified 150 PLGA patients with biopsy material and adequate clinical follow diagnosed at our institution between 1985 and 2010. We examined Braf alterations by RT-PCR, FISH and SNP array analysis. The main outcome measure was progression-free survival (PFS).

Results: Overall, 61% of tumors were B-K fusion positive. All patients with B-K fused PLGA are still alive. Five-year PFS was 61+/-8% and 18+/-8% for fusion positive and negative patients, respectively (p=0.0004). B-K fusion resulted in similarly significant favorable PFS for patients who received chemotherapy. Multivariate analysis revealed that B-K fusion was the most significant favorable prognostic-factor in incompletely resected PLGA and was independent of location, pathology and age. In vitro, Braf overexpression resulted in growth arrest associated with DNA damage (γH2AX expression). Five-year PFS was 68+/-15% and 0% for patients with B-K fused and γH2AX expressing PLGA vs. negative tumors (p=0.001).

Conclusion: These data suggest that B-K fusion confers a less aggressive clinical phenotype on PLGA and may explain their tendency to growth arrest. Combined analysis of B-K fusion and γH2AX expression can determine prognosis and may be a powerful tool to tailor therapy for these patients.

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5 BRAF Alterations in Primary Glial and Glioneuronal Neoplasms of the Central Nervous System: Clinicopathologic Correlations

Fausto Rodriguez, Alex Lin, Susan Williams. Johns Hopkins University; Matthias Karajannis, David Zagzag. New York University; Peter Burger, Eric Raabe, Kenneth Cohen, Charles Eberhart, Eli Bar. Johns Hopkins University

Recent studies have highlighted the importance of BRAF alterations resulting in MAPK/ERK pathway activation in low grade CNS tumors, in particular pilocytic astrocytomas (PA). We studied 118 primary CNS neoplasms using a large cohort of primarily pediatric patients to identify the prevalence and clinicopathologic significance of these alterations. Testing for BRAF fusion transcripts identified KIAA1549:BRAF fusions in 52/105(50%) of low grade gliomas and glioneuronal tumors overall, including 40 (56%) PA; 2 (50%) pilomyxoid astrocytomas/astrocytomas with pilomyxoid features, 4 (57%) low grade glioneuronal tumors, 0 (of 3) diffuse astrocytomas (WHO grade II), 6 (40%) unclassifiable low grade gliomas, and 0 (of 4) pleomorphic xanthoastrocytomas. The following KIAA1549:BRAF gene exon fusions were confirmed by sequencing: 1-16/9-18 (n=26), 1-15/9-18 (n=17), 1-16/11-18(n=4), 1-15/11-18 fusion (n=3), and 1-17/10-18 (n=1). Quantitative RT-PCR analysis indicated that the fusions were expressed at levels as high or higher than KIAA1549. In addition DNA sequencing identified BRAFV600E mutations in 7 of 102 (7%) low grade tumors (3 low grade astrocytomas, 2 PA, 1 pleomorphic xanthoastrocytoma, and 1 low grade glioneuronal tumor). KIAA1549:BRAF fusions were absent in all high grade tumors tested (5 astrocytomas and 4 medulloblastomas), but BRAFV600E mutation was identified in two high grade astrocytomas. BRAFG468A mutations were absent. KIAA1549:BRAF fusions were more frequent in posterior fossa (65% fused, 35% non-fused) than optic pathways (42% fused, 58% non-fused), or supratentorial (28% fused, 72% non-fused) tumors. Median age for fusion positive and fusion negative tumors was 10.8 vs. 12.6 years, respectively. Additional clinical correlations are ongoing. In summary, KIAA1549:BRAF fusions predominate in PA and low grade glioneuronal tumors, and are also present in a subset of low grade unclassifiable gliomas, but not in the limited number of WHO grade II diffuse astrocytomas tested. In addition, the frequency of these alterations may vary by anatomic location.

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6 Diffuse Astrocytomas Can Arise From Either GFAP Astrocytes or Subventricular Zone (SVZ) Neural Stem/Progenitor Cells

C. Miller, Ryan Bash, Mark Vitucci, Andrea Werneke, Ralf Schmid. University of North Carolina

Diffuse astrocytomas, including glioblastomas (GBM), are characterized by cellular, molecular, and morphological heterogeneity. CD133 cells from human GBM have been shown to phenotypically resemble neural stem cells (NSC), displaying self-renewal and multi-lineage differentiation potential in vitro, and to function as tumor initiating cells (TIC) upon injection into the immunodeficient mouse brain. However, the relationship between stem-like TIC and true NSC remains unclear. Using conditional, inducible genetically-engineered mouse models (GEMM) with inactivated RB (GFAP-driven transgenic expression of N-terminal SV40 large T mutant (T121), T) and/or PTEN (P) and/or constitutively activated KRAS (R, KRASG12D), we show that tumorigenesis is initiated in both GFAP astrocytes as well as GFAP NSC/progenitor cells within two weeks of induction in adult mice. At two months post-induction, diffuse astrocytoma (A2, WHO grade II) burden increased with increasing genotypic complexity (T< TP-/-<TP+/-< TR<TRP+/-<TRP-/-, range 6.7-26.9% brain area) and varied by region (brain stem = diencephalon < cortex ≪ olfactory bulb (OFB)). In all six genotypes tested, A2 burden was greatest in the OFB (P<0.05 for TP+/-, TP-/-, and TRP+/-). In addition, T121 A2 was histopathologically identified within the rostral migratory stream in multiple GEMM. High-grade astrocytomas (HGA) with (GBM) and without (anaplastic astrocytoma) pseudopalisading necrosis developed as invasive OFB (22-69%), cortical (22-36%), brainstem (8-24%), and/or diencephalic (23-50%) masses in 4 of 6 genotypes after 3-6 months. Array CGH analysis showed recurrent gains of chromosome 6 (MET, BRAF, KRAS), but no loss of chromosome 11 (TP53) in 7/7 HGA. Primary astrocytes from TRP-/- mice cultured in vitro under NSC conditions displayed self-renewal and multi-lineage differentiation potential and expressed multiple NSC markers, including GFAP, nestin, CD133, and Sox2. These results suggest that GFAP NSC within the SVZ, as well as GFAP astrocytes elsewhere in the neuraxis, may serve as the cell-of-origin for diffuse astrocytomas in the adult murine brain.

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7 In-vivo and Immediate Ex-vivo Confocal Laser Endomicroscopy for Intraoperative Histopathological Imaging of Brain Tumors

Jennifer Eschbacher. Barrow Neurological Institute/St. Joseph's Hospital and Medical Cente; Nikolay Martirosyan, Nader Sanai, Kris Smith, Mark Preul, Stephen Coons. Barrow Neurological Institute/S.J.H.; Peter Nakaji, Robert Spetzler. Barrow Neurological Institute

Frozen section analysis is the current standard for intraoperative diagnosis. Confocal laser endomicroscopy is an emerging technology increasingly used in the clinical setting, including the fields of gastroenterology, gynecology, and urology. Using an endoscopic probe with an attached confocal lens and contrast dye, clinicians can view histopathologic features and cell morphology in real time. We report our findings and experience with this new intra-operative technology in the neurosurgical setting in vivo. A confocal laser endomicroscope with a 6.3 mm distal probe tip was used to image abnormal areas of brain tissue. Using sodium fluorescein for contrast, we visualized 127 regions from 62 brain tumors, 2 spinal cord tumors and 6 nontumor lesions. 5 additional regions from two brain tumors were imaged ex vivo due to intricate tumor locations. Corresponding biopsies from imaged areas were submitted for routine traditional pathologic processing and interpretation by a neuropathologist. Tumors included astrocytomas (grade II-IV), oligodendrogliomas (grade II, III), oligoastrocytomas (grade III), ependymomas (grade I, II), craniopharynigomas (adamantomitous and papillary variants), chordoma (recurrent), subependymal giant cell astrocytoma, and meningiomas (grades I-III). Nontumor lesions included radiation necrosis, sarcoidosis and reactive/ inflammatory processes. Of the 132 total virtual biopsies, 11 (8.3%) did not demonstrate useful or interpretable images due to blood and/or motion artifact. Confocal evaluation of the remainder of lesions produced images which demonstrated architectural, cytostructural and morphologic detail which often corresponded with histological images. We describe and compare the histological features between the two modalities - routine histopathology and intra-operative confocal micrscopy. Our preliminary findings suggest that this new technology will have great implications in shaping the future of intraoperative brain tumor diagnosis and in defining tumor margins.

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8 The Impact of IDH1 and IDH2 Mutations on Progression-Free and Overall Survival in Low Grade Gliomas

Sidney Croul. University of Toronto, Dept. of Lab Medicine and Pathobiology; Christiane Knobbe. Campbell Family Institute at Princess Margaret Hospital; Nesrin Sabha. Arthur and Sonia Labatts Brain Tumor Centre, U of Toronto; Andreas von Deimling. Dept.Neuropathology Ruprecht-Karls-University, Heidelberg, Germany; Abhijit Guha. Division of Neurosurgery, University of Toronto, Toronto, Ontario

Adults presenting with a low grade glioma have an unpredictable time to recurrence and tumor progression, making management decisions and patient counseling difficult. Currently several clinical and molecular characteristics are utilized to guide treatment. However, molecular marker(s) to augment prediction reliability are urgently needed, especially for astrocytic tumors. Recently mutations in IDH1, a key metabolic enzyme, have been identified in the vast majority of low grade gliomas as well as in anaplastic gliomas and secondary GBMs, but not in primary GBMs. Since the first description of mutations in IDH1 numerous additional studies for mutations in IDH1 and its homologue IDH2 have been carried out and several studies have reported that mutations in these genes may confer a better prognosis. Therefore we asked the question whether mutational analysis of IDH1 and IDH2 might be a helpful marker for time to progression and overall survival. Using both immunohistochemistry for the R132H mutation in IDH1 as well as sequencing of the hot spot codons in IDH1 and IDH2 we could show that there was a strong correlation between IDH mutations and longer progression-free and overall survival. This correlation was strongest for anaplastic oligodendrogliomas and oligoastrocytomas as well as for diffuse astrocytomas.Concluding from our data as well as from already published data we propose to introduce screening for IDH1 mutations by immunohistochemistry (R132H), and if negative, for apply sequencing to detect possibly missed other mutations in IDH1 or IDH2 to help in personalizing glioma treatment and patient counseling.

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PLATFORM SESSION 2: NEURODEGENERATIVE - ALZHEIMER DISEASE Friday 8-10 a.m. June 24, 2011

9 Differences in Clinical and Neuropathological Concepts of Alzheimer's Disease at NIA Alzheimer's Disease Centers 2005-2010

Thomas Beach. Banner Sun Health Research Institute; Leslie Phillips. National Alzheimer's Coordinating Center, University of Washington; Sarah Monsell, Walter Kukull. National Alzheimer's Coordinating Center, University of Washington

Clinical diagnostic accuracy heavily impacts epidemiological studies, economic impact studies and especially treatment and prevention trials. We compared the clinical diagnosis of Alzheimer's disease (AD) with neuropathological 'gold standard' criteria, using data collected by the National Alzheimer's Coordinating Center (NACC) from the network of US Alzheimer's Disease Centers (ADCs) since 2005. The initial NACC database query included all 1199 subjects that had at least one Uniform Data Set clinical assessment, had died and were autopsied. Subjects were excluded for whom there was critical missing data, leaving 1043 subjects. Clinical diagnosis was that given at the last assessment during life. Both the clinical diagnosis and the neuropathological diagnosis were stratified by level of confidence. For the clinical diagnosis of AD, NINDS-ADRDA criteria were used, stratified by considering 'probable AD' alone as well as 'probable' plus 'possible' AD. The neuropathological diagnosis of AD was stratified using a 2 × 2 matrix that defined multiple combinations of CERAD neuritic plaque density scores and Braak neurofibrillary tangle stages. The sensitivity of the clinical diagnosis of AD ranged from 65% to 87% while specificity ranged from 64% to 78%. Sensitivity was increased with more permissive clinical criteria while specificity was increased with more restrictive criteria while the opposite was true for neuropathological criteria. The highest combined diagnostic performance (sensitivity of 79% and specificity of 71%) was achieved when the clinical diagnosis was defined as including both probable and possible AD and the neuropathological diagnosis was defined as being inclusive of the following two situations: 1) frequent neuritic plaque density with Braak stage III-VI 2) moderate neuritic plaque density with Braak stage V or VI. In conclusion, the level of agreement between the clinical and neuropathological definitions of AD, based on data from NIA ADC's, vary depending on the levels of clinical and neuropathological confidence.

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10 Towards a Novel Therapeutic Strategy for Alzheimer Disease(AD) Based on Modified Aß Peptides

Michela Morbin, Giulia Mazzolelni, Andrea Uggetti. IRCSS Foundation Istituto Neurologico 'Carlo Besta', Milan - Italy; Laura Colombo, Marco Gobbi. Mario Negri Institute for Pharmachological Research, Milan, Italy; Matteo Salvalglio, Politecnico di Milano. University of Milan - Italy; Giuseppe Di Fede. IRCSS Foundation Istituto Neurologico 'Carlo Besta', Milan - Italy; Marcella Catania. IRCSS Foundation Istituto Neurologico 'Carlo Besta', Milan - Italy; Mario Salmona. Mario Negri Institute for Pharmachological Research, Milan - Italy; Fabrizio Tagliavini. IRCSS Foundation Istituto Neurologico 'Carlo Besta', Milan - Italy

AD is the most common form of degenerative dementia for which there are not significant therapies. Recently we have found an APP mutation at codon 673 (A673V) corresponding to position 2 of Aβ, that causes early-onset AD only in the homozygous while the heterozygous are not affected. In vitro studies showed that the A673V mutation shifts APP processing towards the amyloidogenic pathway and increases Aß aggregation in amyloid fibrils. However, co-incubation of A673V and wild-type peptides inhibits amyloidogenesis and Aß-mediated neurotoxicity. This finding is consistent with the observation that the A673V heterozygous do not develop the disease, and offers grounds for the development of a therapeutic strategy based on A673V-modified Aß peptides. As a first step, we have generated short synthetic peptides homologous to residues 1-6 of Aß carrying the A2V substitution and tested their ability to hinder amyloidogenesis using, circular dichroism, electron and atomic force microscopy. Moreover, a comparative conformation analysis of wt and mutated Aß1-6 was carried out using an all-atom molecular dynamics approach. These studies showed that the interaction between the mutated six-mer peptide and Aß1-40wt antagonizes the elongation of fibrils, inhibits ß-sheet secondary structure, and prevents amyloid-fibril formation. Noteworthy, the anti-amyloidogenic capacity was higher when the mutated Aß1-6 contains only D-amino acids. Molecular dynamics showed that the wild-type peptide is characterized by a "closed" configuration in which the N and the C termini are strongly interacting. Conversely, the structure of the mutated peptide is marked by a stable interaction between the Val2 and Phe4 side chains, resulting in higher flexibility of the peptide. It is conceivable that the structural mobility of the mutated hexapeptide facilitates the interaction with Aß and hinders Aß assembly. Although preliminary, these data provide a foundation for the rational design of therapeutic agents based on Aß fragments with the A673V mutation.

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11 Intraneuronal APP, Not Free Aß Peptides In 3xTg-AD Mice: Implications For Tau Versus Aß Mediated Alzheimer Neurodegeneration

Edward Lee, Matthew Winton, Eveline Sun, Margaret Wong, Susan Leight, Bin Zhang, John Trojanowski, Virginia Lee. University of Pennsylvania

Alzheimer's disease (AD) is characterized by the accumulation of intraneuronal tau and extracellular amyloid-ß (Aß) peptide. A triple transgenic (Tg) mouse (3xTg-AD) was reported to develop Aß plaques and tau inclusions as well as remarkable accumulations of intracellular Aß that were suggested to be the initiators of AD pathogenesis. However, it was unclear whether the anti-Aß antibodies were able to distinguish Aß peptide from the same Aß epitopes within the amyloid precursor protein (APP). To further elucidate the identity of the immunoreactive intraneuronal material in 3xTg-AD mice, we conducted immunohistochemical, biochemical and ultrastructural studies using a well characterized panel of antibodies that distinguish Aß within APP from cleaved Aß peptides. We found that the intraneuronal material shared epitopes with full-length APP but not free Aß. To demonstrate unequivocally that this intraneuronal material was not free Aß peptide, we generated 3xTg-AD mice deficient for ß-secretase (BACE), the protease required for Aß generation from APP. In the absence of Aß production, robust intraneuronal APP immunostaining was detected in the 3xTg-AD/BACE(-/-) mice. Finally, we found that the formation of tau lesions was not different between 3xTg-AD versus 3xTg-AD/BACE(-/-) mice, thereby demonstrating that tau pathology forms independently from Aß peptide generation in this mouse model. Although we cannot corroborate the presence of intraneuronal Aß peptide in 3xTg-AD mice, our findings warrant further study as to the role of aberrant APP accumulation in this unique model of AD.

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12 Ocular Dominance Plasticity Defect in Alzheimer Disease Model Mice

Christopher William. Neuropathology Service, Massachusetts General Hospital, Boston, MA; Carla Shatz. Bio-X, Stanford University, Stanford, CA; Matthew Frosch. Neuropathology Service, Massachusetts General Hospital, Boston, MA; Bradley Hyman. MassGeneral Institute for Neurodegenerative Disease, MGH, Boston, MA

Soluble amyloid-beta has been implicated in the synaptic dysfunction of Alzheimer disease, however, tests of synaptic function in transgenic mice engineered to express human amyloid precursor protein (APP) are typically performed after months of transgene expression. To test the hypothesis that synaptic defects in APP-expressing transgenic mice may be present at much earlier ages, we turned to the developing visual system. In juvenile mice, a critical period exists during which the loss of vision from one eye triggers an expansion in cortical territory driven by the open eye, as well as a shift in cortical responsiveness in favor of visual input from the open eye - a process termed ocular dominance plasticity (ODP). ODP was tested in APP/PS1 mice which co-express the Swedish mutant allele of APP (APPswe) and an exon 9-deleted allele of Presenilin 1 (PSEN1dE9). In non-transgenic mice, a four-day period of monocular deprivation during the peak of the critical period for ODP (P28-32) is sufficient to produce an increase in the width of the domain of primary visual cortex responsive to the ipsilateral, non-deprived eye, as assessed by induction of mRNA for the immediate early gene, Arc, in layer 2/3 cells. This shift in favor of the ipsilateral, non-deprived eye does not occur in APP/PS1 transgenic mice, even following a stronger challenge of a 10-day period (P22-P32) of monocular enucleation. This plasticity defect was also present in an independent, transgenic line that expresses APPswe alone. ODP was intact in a transgenic line which expresses PS1dE9 alone. These data suggest that basic defects in synaptic plasticity are present in transgenic mice expressing APP well in advance of plaque accumulation. The developing mouse visual system may provide a useful system in which to study the effects of APP and amyloid-beta on synaptic function and plasticity in vivo.

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13 Dementia Without AD in Advanced Aging: Many Diseases, Much Confusion

Peter Nelson. University of Kentucky

Correlation between Alzheimer's disease (AD) neuropathology and cognitive impairment is not simplistic. Like most chronic diseases, there is an expected prodrome phase (i.e., preclinical disease) and imperfect clinical-pathological correlation at autopsy. Cognitive impairment in advanced aging is the norm although the brains of many aged individuals lack appreciable AD pathology. Two key non-AD brain pathologies are cerebrovascular disease (CVD) and hippocampal sclerosis (HS-Aging). Rubrics are still evolving for clinical-pathological correlation in these brain diseases. The prevalence of CVD pathology in the National Alzheimer's Coordinating Center (NACC) dataset gives strong indication that CVD pathology is practically ubiquitous in extreme old age. The clinical impact of CVD - although difficult to predict confidently for any individual patient - is correlated with systematic changes in pathological outcomes. Persons with CVD tend to have lower AD-related pathology with a given degree of cognitive impairment. Correspondingly, as people become older (with more CVD), a larger percentage of AD patients die with more moderate AD pathology-Braak stage V rather than Braak stage VI. We are also increasingly impressed by the prevalence and impact of HS-Aging. Based on data from the largest series of HS-Aging cases to date, it seems as though prior low estimates of HS-Aging prevalence were partially biased by inclusion of younger patients. The average age at death for HS-Aging in this large cohort was over 90 years. In this group, each year after age 95 brought higher risk for HS-Aging pathology, but lower risk for AD pathology. Since HS-Aging shares a key biomarker with AD (shrunken hippocampi on MRI), there is likely to be clinical diagnostic uncertainty in some cases. Analyses of CVD and HS-Aging neuropathology from large autopsy cohorts provides key insights of direct relevance to Alzheimerologists: some high-morbidity brain diseases, unlike AD, peak in extreme old age.

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14 Striatal Plaque Density Predicts Braak Tangle Stage and ClinicoPathological Alzheimer's Disease: Implications for Amyloid Imaging

Thomas Beach, Lucia Sue, Monica Mariner, Kim Yantos, Leslie Souders, Glenn Chiarolanza, Jonette Henry-Watson, Geidy Serrano, Marwan Sabbagh, Douglas Walker. Banner Sun Health Research Institute

Amyloid imaging may revolutionize Alzheimer's disease (AD) research and clinical practice but is critically limited by the lack of correlation between cerebral cortex amyloid plaques and dementia. Also, amyloid imaging does not indicate the extent of neurofibrillary tangle spread throughout the brain. Autopsy studies suggest that the presence of striatal amyloid plaques correlates with dementia and Braak neurofibrillary tangle (NFT) stage. Striatal amyloid plaques, which are readily identified by amyloid imaging, might therefore be used to predict, in living subjects, NFT stage and the likelihood that dementia is due to AD. The sensitivity and specificity of striatal plaque density scores, assigned on a 0-3 semi-quantitative scale using Campbell-Switzer and thioflavine S stains, for indicating a Braak stage V or VI and the presence of clinicopathological AD (dementia with NIA-Reagan "intermediate" or "high" rating) was determined in a postmortem series of 229 elderly subjects. Subjects were clinicopathologically classified as 90 non-demented elderly controls, 92 with AD, 18 with AD and dementia with Lewy bodies and 18 with AD and vascular dementia. Subjects with conditions primarily considered to be movement disorders, including Parkinson's disease, progressive supranuclear palsy and corticobasal degeneration were excluded. A higher striatal plaque density score (2 or 3) predicted a higher Braak NFT stage (Braak V or VI) with 96% sensitivity and 79% specificity. A higher striatal plaque density score (2 or 3) predicted the presence of dementia and clinicopathological AD with 86% sensitivity and 89% specificity. Amyloid imaging of the striatum may be useful as a predictor, in living subjects, of Braak NFT stage and the presence or absence of dementia and clinicopathological AD. Validation of this hypothesis will require autopsy studies of subjects that had amyloid imaging during life.

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15 Characterization of Human Brains Resilient to ß-amyloid and Tau Pathologies

Thor Stein, Oriol Dols, Thomas Scotton, Bibiana Da Rocha-Souto, Serrano-Pozo Alberto, Frosch Matthew, Growdon John, Hyman Bradley, Gomez-Isla Teresa. Massachusetts General Hospital, Harvard University, Boston

A significant proportion of cognitively intact elderly individuals have numerous amyloid plaques (SPs) matching the regional distribution seen in Alzheimer Disease (AD). This suggests that some people may tolerate the insult of Aß accumulation in their brains and remain cognitively normal. From the MGH ADRC Brain Bank we identified subjects older than 60 years that were clinically classified as non-demented individuals. Thirteen percent of them met NIA-RI criteria for high likelihood of AD (mismatched). In five of them enough tissue was available for detailed neuropathological assessments, and they were compared to age-matched controls (N=8) and age-matched demented individuals that met pathological criteria for definite AD (N=12). Stereologically-based counts of neurons, NFTs, and astrocytes were performed in immunostained sections in the region that forms the superior temporal sulcus. The total amyloid burden was quantified using Aß immunostaining (10D5), and oligomeric Aß was quantified with the conformational specific antibody NAB61. The mismatch group had similar amounts of SPs and NFTs as the AD group. However, the burden of oligomeric Aß was significantly decreased within the mismatch group as compared to the AD group (p<0.001). As expected, the number of neurons was dramatically decreased in the AD group in comparison to controls (p<0.01). However, no significant neuronal loss was found in the mismatch group. Moreover, while there was a significant increase in the number of activated astrocytes and microglia within the AD group compared to controls (p<0.001), the numbers of these reactive cells in the mismatch brains did not differ from control brains. In conclusion, the neuronal populations in the temporal neocortex of some subjects appear to be resistant to the insult of fibrillar Aß deposition. The low level of oligomeric Aß in these subjects may explain the low numbers of reactive astrocytes and microglia as well as the lack of neuronal loss.

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16 Relationship Between Apolipoprotein E Genotype and Blood-Brain Barrier Permeability

John Donahue, Junghyun Kim. Dept. of Pathology, Rhode Island Hospital/Alpert Medical School; Jill Dennis. Central Research Facilities, Rhode Island Hospital/Lifespan; Michelle Reyzer, Richard Caprioli. Vanderbilt Mass Spectrometry Research Center

There is increasing evidence for blood-brain barrier (BBB) compromise in Alzheimer disease (AD). The presence of the epsilon-4 (e4) allele of the apolipoprotein E (APOE) gene is a risk factor for sporadic AD. APOE is essential for maintenance of BBB integrity. Thus, it is hypothesized that the presence of an apoE4 allele will lead to increased BBB permeability, as compared to the apoE3 allele. To test this hypothesis, sodium flourescein tracer (molecular weight 376 daltons) was injected into transgenic mice that expressed either human apoE3 or apoE4 proteins, as well as APOE knockout (KO) mice, and allowed to circulate for an hour prior to sacrifice to enable it to potentially cross the BBB. The presence and location of tracer in the brain tissue was assessed using imaging mass spectrometry, and the results were quantified for prefrontal cortex, hippocampus, and cerebellum. Results demonstrated that the most tracer in the brain was seen in the APOE KO brains, indicating that they had the leakiest BBB's. Brains that expressed apoE4 showed increased amounts of tracer, as compared to those that expressed apoE3, thus indicating an increase in BBB permeability in the apoE4 brains. These results suggest that apoE4 is not able to maintain BBB integrity as effectively as apoE3, and this may be one of the mechanisms whereby the presence of apoE4 confers an increased risk of developing sporadic AD. [Supported by National Institute on Aging grant 5K08AG029147-03.]

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PLATFORM SESSION 3: INFLAMMATION/INFECTION/MYELIN Friday 2-4 p.m. June 24, 2011

17 Paradoxical in Vivo Versus in Vitro Expression of YKL-40 by Macrophages and Astrocytes

Clayton Wiley, Julia Kofler, Adam Starkey, Guoji Wang, Dafna Bonneh-Barkay. University of Pittsburgh Medical Center

In systemic inflammatory diseases, macrophages express abundant YKL-40. Since neuroinflammatory conditions are characterized by elevated cerebrospinal fluid levels of YKL-40, we expected CNS macrophages to express high levels of YKL40 during inflammatory diseases. To our surprise in situ hybridization and immunohistochemistry have shown that astrocytes are the predominant source of YKL40 in vivo. To elucidate this paradox we attempted a series of in vitro experiments to characterize mediators of YKL40 transcription in pure and mixed cultures of human astrocytes and macrophages. As expected monocytes differentiated in vitro into macrophages produced abundant YKL-40. Classical activation with IFNgamma and LPS further induced YKL40-expression while alternative activation with IL-4 inhibited YKL40-expression. Infection with HIV or treatment with anti-inflammatory drugs did not modulate YKL40 expression. Untreated astrocyte cultures expressed little YKL-40, however, when stimulated with macrophage supernatant showed marked induction of YKL-40 transcription. IL-1beta and to a lesser extent TNFalpha induced astrocyte YKL-40 transcription in a dose-dependent manner, however, blocking IL-1beta and TNFαlpha action inhibited only a portion of the stimulatory effect of macrophage supernatant. Paradoxically co-culturing astrocytes with macrophages neither induced astrocyte expression of YKL40 nor inhibited macrophage expression of YKL-40 as seen in vivo. Presumably some key aspect of the in vivo CNS milieu is not recapitulated in vitro.

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18 MiRNA Expression Profiles of Classically and Alternatively Activated Human Adult Microglia

Julia Kofler, Stephanie Bissel. Department of Pathology, Uma Chandran, Department of Biomedical Informatics, Mark Stauffer. Department of Pathology, Clayton Wiley. Department of Pathology, University of Pittsburgh

Similar to macrophages, microglial cells can be induced into classical (M1) and alternative (M2) activation states, each associated with a distinct gene expression profile. miRNAs are small non-coding RNAs involved in the regulation of gene expression at the posttranscriptional level. Recently, miRNA-124 has been shown to be an important regulator of microglial quiescence in the mouse brain, but little is known about the contribution of miRNAs to the regulation of classical and alternative activation states in human microglia. The goal of our study was to perform an array analysis to identify miRNAs differentially expressed between M1 and M2 stimulated microglial cells. Microglial cultures isolated from postmortem adult human brain tissue were either classically stimulated by IFN-γ and LPS or alternatively stimulated by IL-4. After 48 hours, chemokine secretion into the culture supernatant was measured by ELISA to confirm adequate polarization as indicated by strong release of CXCL10 after M1 and of CCL17 after M2 stimulation. RNA was collected at 48 hours and miRNA expression profile determined by TaqMan quantitative RT-PCR arrays. Of the 754 distinct human miRNAs covered by the array, 50% were not detected in any of the samples, whereas 149 (19.8%) were at least 2 fold differentially expressed between the two activation states. Of the differentially expressed miRNAs, 13 (8.7%) were higher expressed in M1 polarized cells and 136 (91.3%) were preferentially expressed in M2 activated cells. In conclusion, we have identified a subset of miRNAs that are differentially expressed in M1 and M2 stimulated human microglia. These miRNAs warrant further investigation as they may act as regulators of microglial polarization and may serve as potential therapeutic targets to shift neuroinflammatory states in the brain.

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19 Automated Identification of Myelin in Paraffin Sections

Andrew Dwork. NY State Psychiatric Inst/Columbia University

The superficial layers of the human subiculum, presubiculum, parasubiculum, and entorhinal cortex reportedly myelinate during adolescence and adulthood. To verify the progress of myelination and to determine whether it is altered in psychiatric disease, we sought to develop an automated procedure to detect myelin in stained paraffin sections. Myelin was stained by Verhoeff's method, which is relatively resistant to adverse effects of fixation. Although this method yields good visual contrast, intensity of staining can vary within and between slides. While myelin is stained grey or black and is easily detected subjectively, as with most staining procedures, the targeted substance cannot be satisfactorily detected by a density threshold, even if the threshold is set individually for each case. However, to quantify the regional extent of myelination by manual outlining is prohibitively slow and labor-intensive. We thus sought to segment images of the slides by texture, rather than density. Using Visiopharm (Medicon Valley, Denmark) software, we acquired montages (∼ 100 images each) a resolution of 1.3 microns per pixel. We then applied filters for standard deviation, gradient, and linearity. The results of these filters (about the size of individual units of axons and myelin) were then averaged over areas 3-4 times wider, and the results of these averages were used for segmentation by a linear Bayesian process, after "training" the software by marking sample areas of myelin in fornix and deep white matter (not in the region to be analyzed). Identification of myelin in the superficial layers was virtually identical to subjective identification by a neuropathologist. Furthermore, in 8 test slides, it made little difference whether the "training" was performed on each slide separately, or on one slide for the whole group. We conclude that regions of myelination can be detected by image analysis aimed at the microstructural architecture of myelinated axons.

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20 Myelin Loss Despite Normal Oligodendroglial Density in Autosomal Dominant Leukodystrophy With Duplication of the LMNB1 Gene

Michael Barnes. UCSF Dept of Pathology; Shu-Ting Lin, Ying-Hui Fu. UCSF Dept of Neurology; Louis Ptácek. UCSF Dept of Neurology and Howard Hughes Medical Institute; Eric Huang. UCSF Dept of Pathology

Autosomal dominant leukodystrophy (ADLD) is an adult-onset demyelinating disease with manifestations of a progressive loss of autonomic function and fine motor skills at the earliest stage of disease. In later stages, patients experience complete loss of voluntary movements. Molecular genetic analyses have shown that ADLD is associated with duplication of the lamin B1 (LMNB1) gene on chromosome 5q31, which encodes a protein that supports nuclear membrane architecture and DNA functions. Currently six families have been described each with duplications of LMNB1, but the neuropathology of ADLD duplication has not been thoroughly characterized. Here we report a patient with both clinical and genetic features of classical ADLD. Consistent with the autosomal dominant inheritance, multiple family members show similar clinical symptoms. Neuropathology examinations in the proband show a severe loss of subcortical white matter in both hemispheres, particularly affecting the frontal, parietal, and occipital lobes. This myelin loss ranges from a complete absence of myelin in the long tracts to focal areas of disruption and partial loss in the fibers at the grey-white junction. Despite the severe loss of myelin, the density of oligodendroglia is surprisingly well preserved and there is no significant presence of astrogliosis or inflammation. The general architecture and density of neuronal axons are intact, but many axons show disorganization and focal swelling. Ultrastructural analyses confirm a spectrum of myelin loss in axons, with a severity that is proportional to the diameter of the axons. Taken together, our results indicate that the underlying pathology in ADLD is distinctly different from other demyelinating diseases, such as multiple sclerosis, and is likely to affect the production and/or maintenance of myelin without affecting the survival of oligodendroglia.

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21 Central Nervous System Demyelination Following Hepatitis A and DTaP Vaccinations Clinically Mimicking High-Grade Glioma

Sarah Martin. Department of Pathology; Joel Boaz. Department of Pediatric Neurosurgery, Mandy Harris. Department of Child Neurology, Eyas Hattab. Indiana University School of Medicine, Dept. of Pathology and Laboratory Medicine

Central nervous system demyelination has been described in association with vaccination, particularly the hepatitis B vaccine; however, whether the demyelinating process is caused by the vaccine or simply coincidental is still a controversial issue. We describe a case of a previously healthy 12-year-old girl who presented with headache and nausea two days after receiving hepatitis A and DTaP vaccinations. She then developed ataxia, myalgias, tongue deviation and head tilt. MRI showed a 1.1 × 9 × 7 mm enhancing lesion in the left inferior cerebellar peduncle/medulla. Despite steroid treatment, the lesion enlarged and her symptoms worsened to the point of requiring intubation. The lesion was not in a location that could be biopsied, and given its rapid growth and lack of response to steroids, it was assumed to be a high-grade glioma and the patient was treated with radiation therapy. Follow-up MRI showed interval decrease in size of the medullary lesion and multiple new FLAIR hyperintense lesions in the subcortical white matter. Radiation was discontinued, as a demyelinating process was now favored. Several months later, a follow-up MRI showed interval enlargement and increased enhancement of the multiple lesions. At this time, a stereotactic-guided biopsy of a right parietal lesion was performed. Microscopic examination revealed a reactive, non-neoplastic process with patchy loss of myelin and relative preservation of axons amid a background of reactive gliosis and perivascular lymphohistiocytic inflammatory infiltrate, consistent with a demyelinating process. Given the patient's clinical course, clinically isolated syndrome is likely, and her propensity to go on to develop multiple sclerosis is uncertain at this time. We discuss the controversy surrounding post-vaccination demyelinating processes and emphasize that the differential diagnosis of high-grade glioma may include a demyelinating process. Caution should therefore be used when treating an undiagnosed lesion with radiation therapy.

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22 Neuroprotection Provided by IV Transplanted Human Fetal Cerebral Endothelial Cells in an Experimental Brain Ischemia Model

Min-Cheol Lee, Kyung-Hwa Lee. Department of Pathology, Hyung-Seok Kim. Department of Forensic Medicine, Chonnam Natl Univ Med School; Seung U Kim. College of Medicine, Chung Ang University

A stable clonal cell line of hCEC, HEN7, has been generated from human fetal telencephalon using a retroviral vector encoding human telomerase reverse transcriptase (hTERT). HEN7 cells were transplanted intravenously in rat brain with photochemically induced focal cerebral ischemia, and the clinical effects for infarct size, edema volume, and clinical outcome were evaluated. FACS analysis showed HEN7 cells express the characteristics of cerebral endothelial cells. Matrigel assay showed HEN7 cells are capable of good tube formation. HEN7 showed positive immunoreactivity for vascular, stem cell, and tight junction proteins. HEN7 transplanted group showed reduced infarct lesion as identified by bioluminescence and X-gal staining located in the infarcted lesion border area. HEN7 transplanted group showed markedly reduced edema volume associated with MMP-9 expression reduction, and markedly increased nestin-positive cells around infarcted area. HEN7 transplanted group showed earlier recovery from the neurological deficit. Intravenously transplanted hCECs selectively migrated and integrated into cerebral ischemic lesion area and accelerate neurological functional recovery. This new hCEC-based cellular therapy is applicable for clinical trial in ischemic stroke patients.

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23 Antenatal and Perinatal Coxsackie B4 Brain Infection: Case Series

Marc Del Bigio. Pathology - University of Manitoba; Jennifer Hunt. Obstetrics and Gynaecology, University of Manitoba

Enteroviruses are a common cause of neonatal infection. In particular, Coxsackie B viruses are often associated with severe, fatal disease. Antenatal diagnosis of Coxsackie B viral infection is uncommon. Following identification of a fetal case, we undertook a retrospective study. In the index case, maternal fever occurred at 17-18 weeks gestation. Fetal ultrasound examination at 21 weeks showed edema involving the scalp, neck and abdomen, abdominal ascites, tachycardia, and placentomegaly. PCR identified Coxsackie virus B4 from the amniotic fluid. Fetal hydrops worsened and stillbirth occurred at 24 weeks gestation. Examination of the brain showed severe multifocal ventriculitis characterized by loss of ependymal cells, T lymphocyte infiltrate, microglial activation, small foci of hemorrhage, and dystrophic calcification, along with mild leptomeningitis. Lymphocytic infiltrates were also in the myocardium. Two children, born at 37 and 40 weeks mothers who had fever in the preceding week, presented at 4 days age with fever. Both had Coxsackie B4 cultured. One child with lymphocytes in the CSF died at 8 days and was found to have inflammation involving the medulla, spinal cord, and leptomeninges. The other developed congestive heart failure, seizures and died at 28 days. Autopsy revealed myocarditis and multifocal brain hemorrhage with hemosiderin and microcalcification. Two male infants who died at 2 and 3 days age from congenital heart defects had elevation of antibodies titers to Coxsackie B2 and B4 but neither had evidence of heart or brain inflammation and virus was not isolated. A 2.5 year male died of Coxsackie A9 bronchopneumonia; the brain and heart were normal. We conclude that transplacental acquisition of Coxsackie infection, perhaps especially B4, carries a poor prognosis. Affinity of the virus for the CAR receptor, which is highly expressed in developing brain, may play a role in the pathogenesis.

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24 Neurosurgical Presentation of Inflammatory Demyelinating Disease: Clinical, Neuroimaging and Neuropathologic Correlation

Suash Sharma, Mitzi Williams. Georgia Health Sciences University, Medical College of Georgia; Nicole DeMers-Riddle, Pushpa Allam-Nandyala. University of South Florida, Moffitt Cancer Center; James Crownover. Georgia Health Sciences University, Medical College of Georgia; Steven Brem, Reed Murtagh. University of South Florida, Moffitt Cancer Center; Cargyle Allen, Samuel Macomson, Amyn Rojiani. Georgia Health Sciences University, Medical College of Georgia.

The occurrence of large demyelinating lesions clinically simulating brain masses such as tumors or abscesses (demyelinating pseudotumors) is rare but well recognized, with occurrences estimated to occur at 1-2/1000 cases of multiple sclerosis (MS), with a suggested prevalence of 3/million inhabitants per year. Demyelinating pseudotumors frequently present at any age, but more frequently in the second and third decades. They are rare in the pediatric population, and when present tend to be monophasic. Clinical presentations vary depending on their location and size, and include headache, cognitive abnormalities, mental confusion, aphasia, apraxia, and seizures. Modern magnetic resonance imaging is the most sensitive method of detecting the white matter lesions of multiple sclerosis. The neuroimaging differential diagnosis of mass lesions in the central nervous system especially differentiating between inflammatory demyelinating disease and malignant glioma however can be challenging. Potential indications for biopsy include atypical clinical presentations (e.g. encephalopathy, seizures, aphasia), older age, the presence of at least one large lesion (more than 2.0 cm), associated mass effect, edema and/or atypical enhancement patterns. Here we describe the clinical, radiological and pathological findings in 14 cases of inflammatory demyelinating disease (7 MS, 1 neuromyelitis optica, 1 Marburg variant, 3 progressive multifocal leukoencephalopathy, 2 acute disseminated encephalomyelitis) who presented with clinical and neuroimaging findings strongly suggestive of malignant glioma. Novel findings in selected cases included spectroscopic evidence of elevated choline and lactate, a case of neuromyelitis optica with significant clinical recovery, and a case of primary progressive MS demonstrating significant axonal loss on histology. These cases illustrate the importance of considering an inflammatory demyelinating lesion in the differential diagnosis of a mass lesion within the central nervous system and the critical need for expert neuropathologic interpretation on adequate tissue.

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PLATFORM SESSION 4: NEURODEGENERATIVE - FTD/LEWY BODY/PARKINSON Friday 2-4 p.m. June 24, 2011

25 Mitochondrial Fragmentation Mediates 1-methyl-4-phenylpyridinium Toxicity in Neurons: Implication for Parkinson Disease (PD)

Xiongwei Zhu, Bo Su, Yuan Gao. Department of Pathology, Case Western Reserve University; Rudy Castellani. Department of Pathology, University of Maryland; George Perry. Dept. of Path Case Western Reserve Univ & College of Sciences UTSA; Mark Smith, Xinglong Wang. Department of Pathology, Case Western Reserve University

MPP causes selective degeneration of nigrostriatal dopaminergic neurons and Parkinsonism in human. As such, both MPTP and MPP have been extensively used in a variety of in vivo mammalian species and in vitro paradigms as experimental models of PD, respectively. Given the parallels with PD, further understanding the mechanisms by which MPTP/MPP lead to dopaminergic neuronal death could provide insights into therapeutic targets for PD. Compelling evidence suggest that mitochondrial dysfunction could represent a critical event in the pathogenesis of PD and recent studies suggest that abnormal mitochondrial dynamics likely contributes to mitochondrial dysfunction in PD. However, to date, the impact of MPP on mitochondrial dynamics remains unknown. To address this, in this study, we determined whether MPP induced abnormal changes in mitochondrial dynamics and determined the temporal and causal relationship between changes in mitochondrial dynamics and MPP+-induced mitochondrial abnormalities and neuronal death. In SH-SY5Y cells, MPP causes a rapid increase in mitochondrial fragmentation followed by a second wave of dramatic increase in mitochondrial fragmentation, along with increased DLP1 expression and mitochondrial translocation. Genetic inactivation of DLP1 completely blocked MPP+-induced mitochondrial fragmentation. While blockage of mitochondrial fragmentation could partially rescues ATP decline and increased [Ca2]i, it could almost completely prevent MPP+-induced increased ROS production, loss of mitochondrial membrane potential, enhanced autophagy and cell death. Thiol antioxidant NAC or glutamate receptor antagonist D-AP5 could also partially alleviate MPP+-induced mitochondrial fragmentation. We further validated our findings in primary rat midbrain dopaminergic neurons that 0.5 μM MPP induced mitochondrial fragmentation only in TH-positive dopaminergic neurons in a similar pattern to that in SH-SY5Y cells. Overall, these findings suggest that DLP1-dependent mitochondrial fragmentation plays a crucial role in mediating MPP+-induced mitochondria abnormalities and cellular dysfunction and may represent a novel therapeutic target for PD. Supported by NIH R21NS071184 and American Parkinson Disease Association.

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26 Hierarchical Cluster Analysis of Cortical Pathology Suggests Pathologic Heterogeneity of Dementia in Parkinson's Disease

Dennis Dickson, Carolyn Orr, Melissa Murray. Mayo Clinic; Rajesh Pahwa. University of Kansas; Kelly Lyons. University of Kansas; Samuel Goldman, William Langston. The Parkinson Institute; Zbigniew Wszolek, Ryan Uitti. Neill Graff-Radford; Tanis Ferman. Mayo Clinic

The relationship of dementia with Lewy bodies (DLB) to Parkinson disease dementia (PDD) beyond the temporal sequence of cognitive dysfunction relative to motor dysfunction (early in DLB and late in PDD) remains an area of active investigation. We used image analysis to assess phospho-tau, Aß and a-synuclein burden in midfrontal and superior temporal gyri of 19 cases of PD (11M;8W; 76 y) and 20 cases of PDD (16M:4W; 77 y) who had been followed in life by movement disorder experts. They were compared to 15 cases of DLB (12M:3W; 79y) enrolled in a longitudinal study of DLB. The average Braak NFT stages were similar: II, II and III; while MMSE scores were greater in PD than PDD and DLB: 29, 15 and 18, respectively. Tau, Aß and a-synuclein burdens were entered into hierarchical cluster analysis to define subtypes independent of clinical features. Cluster analysis defined four groups: group 1 with cortical tau, Aß and a-synuclein; group 2 with cortical Aß and a-synuclein; group 3 with cortical a-synuclein; and group 4 with minimal cortical tau, Aß and a-synuclein. Almost all of the PD cases (89%) were found in the latter two groups, characterized by a paucity of Alzheimer type pathology. All but one of the DLB cases fell in groups 1 (53%), 2 (17%) or 3 (27%) characterized by significant cortical tau, Aß and a-synuclein pathology. PDD was more heterogeneous, including nearly equal numbers of cases in the four groups (1 - 15%, 2 - 35%, 3 - 30%, 4 - 20%). The results indicate that dementia in PDD includes a subset of cases with a paucity of cortical pathology, while significant cortical a-synuclein pathology and variable Aß and tau pathologies defines DLB. This novel classification of Lewy body disorders based upon unbiased statistical methods awaits further validation and genetic correlates.

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27 Causal Therapy of prion and Parkinson's Disease With Novel Inhibitors of Protein Aggregation

Hans Kretzschmar, Jens Wagner. Institut für Neuropathologie Universität München; Sergey Ryazanaov, Andrei Leonov. Max-Planck-Institut für biophysikalische Chemie; Johannes Levin. Neurologische Klinik Universität München; Song Shi, Catharina Prix. Institut für Neuropathologie Universität München; Martin Groschup. Friedrich-Löffler-Institut; Christian Griesinger. Max-Planck-Institut für biophysikalische Chemie; Armin Giese. Institut für Neuropathologie Universität München

The hallmark feature of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease or prion disease is the ability of disease-specific cellular proteins to fold into stable alternative conformations and to form neurotoxic aggregates that accumulate in the brain. These deposits share similar morphological, structural, and staining characteristics. Small molecules that inhibit pathological protein aggregation, therefore, may provide a disease-modifying therapy for these diseases, for which only symptomatic treatment is available so far. We performed a systematic high-throughput screening campaign employing in-vitro-aggregation and cell-culture assays. Based on primary screening data, we synthesized secondary focused chemical libraries, and tested candidate compounds using in vivo models of prion diseases and Parkinson's disease. We identified a new lead structure (3,5-Di-Phenyl-Pyrazole derivatives) with anti-prion and anti-synuclein aggregation activity in vitro. Interestingly, similar results regarding structure-activity relationships were obtained for both targets. In prion-infected mice, compound treatment inhibited PrPSc accumulation, blocked neuronal cell death and significantly prolonged survival, even when treatment was started after disease onset. Similarly, in a transgenic Parkinson mouse model expressing human A30P-a-synuclein, both inhibition of protein deposition and of disease-associated behavioural deficits was observed with a significantly prolonged disease-free survival. We observed no toxicity at therapeutic doses and found an excellent oral bioavailability and blood-brain-barrier penetration. Our in vivo experiments show that inhibition of protein aggregation provides a potent disease-modifying therapy, which corroborates the key role of protein aggregation in disease pathogenesis. The compounds identified by us hold promise for the treatment of neurodegenerative diseases in humans and support the relevance of common molecular features in protein aggregation disorders.

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28 Overexpression of Mutant LRRK2 in Cultured Cortical Neurons is Associated With Excitotoxic Neurodegeneration

Edward Plowey. Division of Neuropathology, Jon Johnson. Department of Neuroscience, Charleen Chu. Division of Neuropathology, University of Pittsburgh Medical Center

Mutations in leucine-rich repeat kinase 2 (LRRK2) underlie PARK8-linked autosomal dominant Parkinsonism and are found in approximately 2-3% of patients with sporadic Parkinson's disease (PD). Furthermore, genome wide association studies have identified variations in LRRK2 as important risk factors in sporadic PD. In vitro and in vivo studies have shown that neuronal expression of mutant LRRK2 induces progressive neurite degeneration with inclusion formation and neurite autophagy. However, little is known about the upstream events that lead to neurite degeneration. We hypothesized that an alteration in synaptic function underlies mutant-LRRK2 induced neurite degeneration. We performed in vitro patch clamp studies of glutamatergic synaptic function in high-density rat cortical neuronal cultures that were co-transfected with wild-type or mutant-LRRK2 cDNA expression plasmids and a GFP-expression vector. Transfection conditions were optimized for LRRK2 expression in recorded neurons without LRRK2 expression in the afferent pre-synaptic terminals. Voltage clamp recordings of miniature excitatory post-synaptic potentials demonstrated elevated event frequency, but not amplitude, in neurons expressing Parkinson's associated G2019S-LRRK2 or 1441C-LRRK2 compared to vector-transfected control neurons and kinase-dead 1906M-LRRK2 transfected neurons. An underlying increase in excitatory synapse density (per unit length of dendrite) in mutant-LRRK2 expressing neurons was demonstrated via confocal microscopic examination of immunofluorescence-labeled excitatory synapse protein puncta. G2019S-LRRK2 or 1441C-LRRK2 expression was associated with enhanced whole-cell current responses to direct application of AMPA and NMDA receptor agonists, suggesting elevated post-synaptic glutamate receptor expression. Mutant-LRRK2 transfected neurons demonstrated enhanced vulnerability to synaptic glutamate stress induced by cell culture depolarization with potassium. Furthermore, treatment of 2019S-LRRK2 expressing neurons with the NMDA receptor antagonist memantine significantly restored neurite length and branching complexity. These experiments suggest that toxic elevations in NMDA receptor activity contribute to neurite degeneration in mutant LRRK2 overexpressing neurons.

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29 Ultrastructural Localization of FUS/TLS in Neuronal Intermediate Filament Inclusion Disease

Wen-Lang Lin, Dennis Dickson. Mayo Clinic

Neuronal intermediate filament disease (NIFID) is a rare neurodegenerative disease, usually presenting as dementia, sometimes with motor or extrapyramidal signs. Its neuropathology is characterized by frontotemporal lobar degeneration (FTLD) with neuronal inclusions (NCI) that are immunoreactive with intermediate filaments (IF), including neurofilament (NF) and a-internexin (INX). Ultrastructural studies have shown that the NCIs in NIFID are heterogeneous, with different types of filaments and variable immunolabeling for NF, INX, ubiquitin and p62 (Neuropathology 2006; 26, 417). A recent immunohistochemical (IHC) study showed FUS (fused in sarcoma), a protein found recently to be the cause of familial amyotrophic lateral sclerosis (FALS), was more abundant than IF in NIFID, suggesting that FUS was the primary pathologic process. Double IHC showed FUS- and IF-immunoreactivity in separate or in the same neurons (Acta Neuropathol 2009; 118, 605). Here we use post-embedding immunogold electron microscopy to study the localization of FUS in NIFID. Formalin-fixed autopsy tissues were processed and embedded in LR White resin. Three rabbit polyclonal antibodies against FUS, recognizing aa 1-50, 52-400, 86-213, 52-400, respectively, as well as antibody to NF were used. Our data showed that FUS was always localized to granule-coated filaments in neuronal cytoplasm whether they were present exclusively, or with IF that were positive for NF. Furthermore, when present in the same neuron, FUS-positive granule-coated filaments tended to surround FUS-negative IF inclusions. Despite their close proximity, the two proteins rarely mixed. Some neurons had only NF-positive inclusions. In conclusion, FUS-positive granulofilamentous and IF-positive inclusions are distinct structures in their morphology and immunoreactivity in NIFID.

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30 Light and Ultrastructural Analysis of FUS Immunoreactivity in Neuronal Intermediate Filament Inclusion Disease

Keith Josephs. Mayo Clinic, Department of Neurology, Rochester, MN; Wen-Lang Lin. Mayo Clinic, Department of Neuroscience, Jacksonville, FL; Joseph Parisi. Mayo Clinic, Department of Laboratory Medicine and Pathology; Neil Graff-Radford. Mayo Clinic, Department of Neurology, Jacksonville, FL; Ronald Petersen. Mayo Clinic, Departement of Neurology, Rochester, MN; Dennis Dickson. Mayo Clinic, Department of Neuroscience, Jacksonville, FL

Neuronal intermediate filament inclusion disease (NIFID), also known as neurofilament inclusion body disease, is a rare neurodegenerative disorder characterized pathologically by the presence of alpha-internexin and neurofilament immunoreactive inclusions. Clinicopathological features of NIFID overlap with two other rare neurodegenerative diseases: atypical FTLD with ubiquitin-immunoreactive inclusions (aFTLD-U) and basophilic inclusion body disease (BIBD). It has recently been reported that all three entities have fused in sarcoma (FUS) immunoreactive inclusions. The aim of this study was to assess the immunohistochemical profile of our 5 cases of NIFID, at the light and electron microscopic level, and compare the profile with that of our cases of aFTLD-U (n=8) and BIBD (n=2). All 10 aFTLD-U and BIBD cases were FUS immunoreactive. Of the five NIFID cases, only three showed FUS immunoreactivity. One NIFID case had a striking abundance of IF immunoreactive inclusions and no FUS-immunoreactive inclusions. The other FUS-negative NIFID case had few IF immunoreactive inclusions and a moderate number of TDP-43 immunoreactive inclusions. Electron and immunoelectron microscopic examinations revealed FUS-immunoreactive granulofilamentous cytoplasmic inclusions in the three FUS-positive NIFID cases, similar to the FUS-immunoreactive granulofilamentous inclusions in the aFTLD-U and BIBD. Granulofilamentous inclusions in the TDP-43-positive NIFID case were FUS negative, yet TDP-43-immunoreactive. Compact hyaline inclusions in all 5 NIFID cases were FUS-negative. No compact inclusions were identified in aFTLD-U or BIBD. These results demonstrate that FUS immunoreactivity in NIFID is variable and unlike in aFTLD-U and BIBD, is limited to only certain types of neuronal cytoplasmic inclusions. The lack of FUS in one NIFID case argues against the notion that FUS is the primary pathological process in NIFID. Given the ratio of TDP-43 to IF immunoreactive inclusions, the other TDP-43 positive NIFID case is likely an example of FTLD-TDP with IF immunoreactivity, instead of, NIFID with TDP-43 immunoreactive inclusions.

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31 Neurodegeneration With Brain Iron Accumulation and Lewy Body Disease With Mutations in a Novel Mitochondrial Membrane Protein

Hans Kretzschmar, Sigrun Roeber. Institut für Neuropathologie Universität München; Gabor Kovacs. University of Vienna; Herbert Budka. University of Vienna; Wolfgang Feiden. Universität des Saarlandes; Howard Hurtig, John Trojanowski. University of Pennsylvania; Wouter Kamphorst. Netherlands Brain Bank; Monika Hartig. Thomas Meitinger; Holger Prokisch. Helmholtz Zentrum München

The common denominator of neurodegeneration with brain iron accumulation (NBIA) is iron deposition and neuroaxonal dystrophy in the basal ganglia; recent neuropathologic and genetic research has shown changes typically associated with Parkinson disease, i.e. widespread Lewy body (and tau) pathology in some of the NBIA cases with mutations in a number of different genes. NBIA comprises two major types of recessively inherited diseases, (1) NBIA 1, which is caused by mutations in the pantothenate kinase 2 gene (PANK2) and is therefore also known as panthotenate kinase associated neurodegeneration (PKAN). PANK 2 is a mitrochondrial protein that catalyzes the first step of the CoA pathway. On MR images this disease is characterized by the 'eye-of-the-tiger' sign. (2) NBIA 2 with mutations in the phospholipase A2 gene (PLA2G6), a mitochondrial calcium-independent A2 phospholipase that catalyzes the release of fatty acids from phospholipids. Widespread a-synuclein-positive Lewy body pathology has recently been described in the neocortex in these cases while other patients with PLA2G6 mutations show infantile neuroaxonal dystrophy (INAD). (3) Other, rare cases of NBIA are associated with aceruloplasminemia or neuroferritinopathy and due to mutations in CP and FTL. (4) Here we describe two cases with pathological changes typical of NBIA and massive Lewy body as well as tau pathology and mutations in a novel NBIA gene encoding a mitochondrial membrane protein. (5) Additional idiopathic cases show iron deposits of various forms in the basal ganglia, axonal dystrophy and other pathological changes but no mutations in the genes known to be associated with NBIA.

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32 The Relationship Between Mitochondrial Dysfunction and Alpha-Synuclein in a Yeast Model of Alpha-Synucleinopathies

Pavan Auluck. Massachusetts General Hospital; Susan Lindquist. Whitehead Institute for Biomedical Research, HHMI, and MIT

Alpha-synuclein (a-syn) is a small lipid binding protein whose function is poorly characterized. Mutations, duplication, and triplication of the a-syn gene locus (SNCA) have been linked to dominantly inherited forms of Parkinson Disease (PD). Notably, a-syn is also the major constituent of Lewy bodies and Lewy neurites, protein aggregates that are the histopathological hallmark of idiopathic PD. Thus a-syn dysfunction appears to be central to the pathogenesis of both hereditary and sporadic forms of PD. The Lindquist laboratory has previously created a yeast model of a-syn pathobiology that replicates the dose-dependent toxicity associated with the protein. Through a forward genetic screen for modifiers of toxicity, we established that vesicle trafficking, particularly within the ER and Golgi, was a process that was particularly sensitive to a-syn expression. Subsequently, we found that a yeast strain expressing somewhat higher levels of a-syn also exhibits defects in mitochondrial function. We performed an unbiased chemical screen using this high-toxicity strain to identify a class of compounds that were capable of alleviating both the mitochondrial toxicity and the ER-to-Golgi trafficking defects associated with a-syn expression. As we have been unable to detect the presence of a-syn in or on the mitochondria, we posit that mitochondrial dysfunction must be the indirect consequence of a-syn dysfunction within the cytosol and/or ER. We are now examining whether altering the relationship between the ER and mitochondria modifies a-syn toxicity. We will present preliminary data from our ongoing studies.

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POSTER SESSION 1 Friday June 24, 2011, 8 a.m.-6 p.m.

33 Astroblastoma: A Rare Clinicopathological Entity

Derick Aranda, Andrew Parsa, Tarik Tihan, Soonmee Cha. University of California San Francisco

Background: Astroblastoma is a rare and controversial glial neoplasm that is currently considered to be a distinct clinicopathological entity by the WHO classification. The biological behavior and criteria for grading astroblastomas are still uncertain. This is partly due to limited number of reported cases in the literature of these neoplasms. Well documented small series and case reports appear to be the only means of providing a better insight into the cliniocopathological characteristics of this unique neoplasm.

Design: We present the clinicopathological features of four patients (two males and two females) diagnosed with astroblastoma. The mean age was 36 years (range: 9 - 68 years). The tumors were located in the frontoparietal region in three and frontotemporal region in one patient.

Results: Two patients presented with headache and the other two patients presented with seizures. All underwent craniotomy and the extent of resection was recorded as near total in two cases and subtotal in two cases. Adjuvant therapy was given to three patients following initial surgery. Subsequently, three patients underwent additional surgical procedures, two patients underwent radiotherapy and one patient received radioactive seeds and subsequent Gamma Knife treatment. Progression-free survival (PFS) for each patient was 2, 4, 5 and 11 years with a mean value of 5.5 years. At the end of follow-up three patients were alive with no evidence of disease with overall survival of 5, 11 and 21 years (Mean: 12.3 years). One patient (case 4) died of progressive tumor and associated complications 29 years after initial surgery.

Conclusion: Little is known about the origin, true biological nature of astroblastoma and factors that may influence prognosis. Well-documented cases with long-term follow-up such as the ones presented in this study will allow us to better characterize this neoplasm and provide material for future meta analysis of this enigmatic entity.

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34 MIB-1 Labeling Index (MLI) Estimation Using Reference Images in Gliomas

Soma Karak, Xianyuan Song, Srinivas Mandavilli. Hartford Hospital

Background: MIB-1 labelling index (MLI) is used routinely in diagnosis of gliomas. This has both a diagnostic and prognostic implication. The estimation of MLI is labor intensive and confounded by interobserver variability (per recent literature). Some expert neuropathologists use a cartoon drawing as estimates to MLI. Using that as an inspiration, we wanted to evaluate the application of digital images of MIB-1 IHC (immunohistochemical staining) to estimate labeling indices in gliomas of the brain.

Design: Gliomas were retrieved from pathology files for the time period 2003 to 2010. Forty cases of gliomas (grades 2, 3 and 4) were reviewed including the MIB-1 IHC slides. Numerous MIB-1 positive areas were photographed at 400× to demonstrate variable labeling indices (1-2%, 3-4%, 5-6 %, 7-9%, 10-14%, 15-19%) in foci of variably cellularity. The actual MLI was calculated by manually counting MIB-1 positive nuclei: total number of nuclei in any given photograph. Care was taken to exclude inflammatory/ endothelial cells with MIB-1 positivity whenever possible. Foci more than 20% MLI were excluded from the study.

Results: Pictures are arranged in an ascending order according to MIB-1 positivity, with different cellularity, and posted as reference. See figure 1 (MIB-1 3-4% in 2 cases with variable cellularity).

Conclusions: Variability in cellularity can confound a visual estimation of MIB-1 labeling index and counting MLI in every case can be tedious and often not reproducible. Using a reference set of 36 images, this is an attempt to create a practical tool for more accurate and quick assessment of MLI.

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35 Dysembryoplastic Neuroepithelial Tumor-Useful Diagnostic Features and Importance of Radiographic Correlation

Knarik Arkun, William Gomes. Neuroradiology; Patrick LaSala. Neurosurgery; Ira Abbott. Neurosurgery; Christian Keller. Neuropathology; James Goodrich. Neurosurgery; Karen Weidenheim. Montefiore Medical Center, Neuropathology

Dysembryoplastic neuroepithelial tumors (DNT) are rare primary WHO grade I CNS neoplasms of glioneuronal type. DNT are typically found in the temporal lobe and show a nodular, non-enhancing, often cystic neuroimaging pattern. Tumor patterns include simple, complex and glioma-like variants. We reviewed clinical data, radiologic appearance of nine DNT seen at our institution over a six-year period,and correlated those findings with neurohistology of the neoplasms. 4/9 patients had two tumor resections. Our patients presented at a mean age of 15.6yr (5-38yr) with a M:F ratio of 3.5:1. Eight of nine patients presented with medically intractable epilepsy; one presented with headache. MRI study showed that all patients had bubble-like nodular intracortical masses varying in size from 0.5-4.2 cm, and four of nine showed focal radiographic enhancement. Neurohistology in all cases showed a nodular pattern predominantly composed of oligodendrocyte-like cells (OLC). Two cases had focal knotting of cells. A very scanty specific glioneuronal element with floating neurons was present in 6 complex cases/9 cases. None of the cases were simple DNT. The stroma of both the oligodendrocyte-like areas and the specific glioneuronal element stained only focally with Alcian blue. Six cases (three cases also had enhancement on MRI) had minimal endothelial proliferation and one case had a prominent glomeruloid pattern. The MIB-1 proliferation index was 4-8%. Two cases showed tumor regrowth after 1 and 3 years respectively, but only one of these enhanced. In 4/9 patients, disabling neurological symptoms persisted after surgery. Study of these nine cases indicated that the specific glioneuronal element and floating neurons were scanty and difficult to identify. More useful diagnostic features, present in all cases, were multinodularity and small round OLC, which could be identified even in fragmented specimens. Clinical and radiographic correlation is critical in preventing errors in diagnosis of tumors with OLC.

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36 Adult-Onset Angiocentric Glioma in the Mesial Temporal Lobe

Hajime Miyata, Masae Ryufuku. Research Institute for Brain and Blood Vessels - Akita; Taku Ochiai. Department of Neurosurgery, Tokyo Women's Medical University; Kaku Niimura, Tomokatsu Hori. Department of Neurosurgery, Moriyama Memorial Hospital

Angiocentric glioma (AG) is an epilepsy-associated stable or slowly growing cerebral tumor primarily affecting children and young adults, histologically consisting mainly of monomorphic, bipolar spindle-shaped cells and occasional round epithelioid cells, showing angiocentric growth pattern and features of ependymal differentiation. We have encountered two cases of adult-onset AG locating in the mesial temporal lobe predominantly composed of epithelioid cells showing marked vascular polarity and conspicuous ependymal differentiation. Case 1 is a 54-year-old woman with a 10-year history of complex partial seizure. MRI revealed T1-low/T2-high intensity change without enhancement in the left hippocampus and amygdala with partial cystic change. After selective amygdalohippocampectomy, she has rare non-disabling seizure under medication for over 50 months (Engel's class I). Case 2 is a 37-year-old man with a 3-year history of convulsion. MRI revealed a tumor in the left uncus and amygdala, showing T1-low/T2-high intensities. FDG-PET study demonstrated lateral temporal hypometabolism. After the combined amygdalohippocampectomy and anterior temporal lobectomy, he has been seizure-free for over 3 months as of February 1, 2011. Histologically the tumors in both cases consisted mainly of round to polyhedral epithelioid cells with EMA-positive paranuclear eosinophilic dot-like structure, showing conspicuous single- and multi-layered angiocentric arrangements. These cells were variably immunoreactive for GFAP but not S-100. There was also a partial solid area containing GFAP-positive bipolar spindle-shaped tumor cells showing Schwannoma-like fascicular arrangement and rare EMA-positive dots. These cells were aligned radially and longitudinally along parenchymal blood vessels. There were no histological features of malignancy. Electron microscopic investigation demonstrated ependymal differentiation of tumor cells showing microlumens filled with microvilli, and intermediate junctions, cilia and basal body. Severe hippocampal sclerosis was observed in case 1 but not in case 2. Similar cases have rarely been described in the literature. These cases may constitute a rare subset of adult-onset AG of epithelioid cell-predominant type.

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37 Cerebellar Low Grade Astrocytomas With Diffuse Growth Pattern

Cristiane Ida. Mayo Clinic; Fausto Rodriguez. Johns Hopkins University; Jesse Voss, Brooke Mc Cann, Sarah Jenkins, Kevin Halling, Caterina Giannini. Mayo Clinic

Cerebellar low grade astrocytomas with diffuse pattern of growth include the exceedingly rare diffuse astrocytomas (WHO grade II) (DA) and a minority of pilocytic astrocytomas (PA) (WHO grade I), so called "diffuse PA". To characterize the clinical, pathological and molecular features of these tumors, we reviewed 106 cerebellar low grade astrocytomas (WHO grade I & II) operated at our institution (1984 to 2010). We identified 19 cases: 5 DA (WHO grade II), 8 "diffuse PA" (WHO grade I) and 6 low grade astrocytomas, subtype indeterminate (LGSI). Eighty-six classic PA and 1 DA centered in the brainstem, but diagnosed through a cerebellar peduncle biopsy, were excluded. Patients included 13 M and 6 F, median age at diagnosis 14 yrs (2-63), with 26 yrs (8-63) in DA, 10 yrs (2-34) in "diffuse PA" and 13 yrs (2-26) in LGSI, respectively. All tumors were centered in the cerebellum, with brain stem extension in 3 (of 5) DA and 2 (of 6) LGSI. Contrast enhancement was noted in 1 (of 5) DA, 6 (of 8) "diffuse PA" and 5 (of 6) LGSI. Median follow-up after surgery was 5.3 yrs (0.4-14.6): 4 DA patients are alive (1 died at 3.6 yrs); 7 "diffuse PA" are alive, 2 with recurrence (1 died at 3.3 years); and 6 LGSI are alive, 1 with recurrence. IDH1-R132H immunostain was negative in 19 (of 19) cases. IDH1-2 pyrosequencing mutation analysis disclosed IDH1-R132G mutation in 1 (of 16) case, a DA. Studies to identify BRAF V600E mutation & KIAA1549- BRAF fusion are in progress. Morphologically, cerebellar low grade astrocytomas with diffuse growth pattern encompass 3 groups: DA, "diffuse PA" and LGSI. IDH1 mutations are rare, and exclusively found in 1 DA (20%), a lower frequency than their cerebral counterpart.

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38 Next Generation Sequencing of Oligodendroglioma - A Work in Progress

Stephen Yip, Yaron Butterfield, Olena Morozova. Genome Sciences Centre, BC Cancer Agency; Michael Blough. Department of Clinical Neurosciences, University of Calgary; Jennifer Chan. Department of Pathology and Laboratory Medicine, University of Calgary; Alexandra Maslova, Suganthi Chittaranjan. Genome Sciences Centre, BC Cancer Agency; J Greg Cairncross. Department of Clinical Neurosciences, University of Calgary; Marco Marra. Genome Sciences Centre, BC Cancer Agency

Rapid advances in sequencing technology have revolutionized the field of molecular oncology through identification of novel, biologically and clinically relevant genes. This is achieved via a massively parallel approach that allows for relatively unbiased examination of the cancer exome, transcriptome, and genome. In this study we performed whole exome sequencing to 100X haploid exon coverage of fourteen 1p19q co-deleted oligodendrogliomas/normal pairs using the Illumina platform. In addition, we completed whole genome shotgun sequencings to 30X haploid coverage of two oligodendroglioma brain tumor initiating cell (BTIC) lines derived from two of the above tumours. We have identified 1274 candidate somatic non-synonymous single nucleotide variations (SNVs) in our exome sequencing discovery cohort. The number of SNV per tumor ranged from 13 to 716 with a median of 37.5. Strikingly, 12/14 oligodendrogliomas harbor somatic R132H mutation in IDH1. The remaining two tumors have R172H mutation in IDH2. Somatic changes in TP53 are not detected in all samples except one. We are currently verifying and validating the other candidate SNVs using Sanger sequencing and deep Illumina amplicon sequencing in a larger cohort of co-deleted cases and other gliomas. Analysis of WGSS data of two BTIC lines also revealed IDH1 mutation and absence of TP53 mutation in one. BT054 has 192 and BT088 has 2390 somatic non-synonymous SNVs. The latter fulfills the hypermutation phenotype with more than 50% of SNVs being transitions in CpC context; moreover, it harbors a somatic heterozygous SNV in MSH6 that results in Thr to Ile amino acid change in codon 521. Lastly, BT088 is derived from a tumour that was heavily treated with alkylating agents which is consistent with known association between somatic mutations in MSH6, exposure to alkylating agents, and genome-wide hypermutation. In summary, next generation sequencing has the potential to offer an unprecedented view of the oligodendroglioma genome.

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39 Malignant Transformation of Clear Cell Ependymoma: A Case Report

Mark Smethurst. Dept of Pathology, Alexis Demopoulos, Susan Morgello. The Mount Sinai Medical Center

Clear cell ependymoma is usually a supratentorial tumor of children and young adults. Histological features include clear cells with round nuclei arranged in a honeycomb pattern, which can resemble oligodendroglioma and central neurocytoma. Considered a WHO grade II tumor, the natural history is poorly documented. Some investigators suggest a predilection for recurrence. We are aware of only one published report documenting malignant progression to anaplastic glioma (Neuropathology 2009, 29:299). Herein, we present a 14 year old boy who underwent subtotal resection of a complex, partially cystic 8 cm left temporal lobe mass. The tumor was centered in the temporal horn, with extensive parenchymal involvement. Initial histological features were felt to represent oligodendroglioma; further workup demonstrated clear cell ependymoma. Following radiation therapy, the tumor remained radiographically stable for four years. The patient was lost to radiographic follow-up for a further three years. Seven years after initial presentation, a generalized seizure prompted neuroimaging, revealing recurrent, enhancing left temporal tumor. Re-resection demonstrated glioblastoma multiforme. He received chemoradiation with temozolomide, followed by temozolomide monotherapy. Seven months after completing radiation therapy, a third resection was performed for progression of disease. Histological evaluation revealed a primitive neuroepithelial neoplasm consistent with small cell glioblastoma, WHO Grade IV. The tumor exhibited marked heterogeneity with divergent histological and immunohistochemical phenotypes. Further post-operative rapid tumor growth occurred two months after the last surgery and the patient expired 9.5 years after initial presentation. The nature of supratentorial ependymomas of childhood, and in particular, clear cell variants, is unclear. More experience is needed in the long term follow up of these lesions.

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40 Anaplastic Astrocytomas With Abundant Rosenthal Fibers in Elderly Patients

Yasuo Sugita, Yousuke Okada, Yoshihiko Nakamura, Koichi Ohshima. Department of Pathology, Mizuhiko Terasaki. Department of Pathology, Department of Neurosurgery, Kurume University School of Medicine

To investigate the clinicopathological features of anaplastic astrocytoma with abundant Rosenthal fibers (RFs), this study assessed four cases of anaplastic astrocytoma (AA) (elderly patients; age>70 years). Histologically, these tumors were diffusely infiltrating astrocytomas with brightly eosinophlic cytoplasm. In these cases, pleomorphism, bizarre giant cells and lymphocytic collection were observed, but not necrosis. The cytoplasmic granules were PAS-negative, and deep purple with PTAH. Immunohistochemically, the tumor cells with cytoplasmic granular cells showed a positive reaction for glial fibrillary acidic protein (GFAP). The cytoplasmic eosinophlic granules were positive for aΒ-crystallin, ubiqutin and HSP 27. The MIB-l labeling index of these cases ranged 8-10%, respectively. Ultrastructurally, the tumor cells had electron dense amorphaus structures in the cytoplasm and in the processes. The structures were accompanied by glial filaments. Based on these microscopic, immunohistochemical and ultrastructural findings, the cytoplasmic eosinophlic granules of these cases were considered to be RFs. Considering the duration from the onset of the resection of the tumor, and the postoperative course, two of present cases had relatively good prognoses compared with ordinary AA. These results indicate that the presence of RFs in astrocytic tumors does not necessarily exclude a diagnosis of high grade astrocytomas, and pathologists should keep AA with abundant RFs in mind, particularly when they diagnose astrocytic tumors in a small sample or an intraoperative frozen section. In addition, AA with abundant RFs in elderly patients may be classified as a peculiar variant of anaplasic astrocytomas.

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41 Papillary Tumor of the Pineal Region: Ultrastructural Study of a Case

Matthew Cykowski. Dept. of Pathology, Oklahoma University Health Sciences Center; Eric Wartchow, Gary Mierau. Dept. of Pathology, The Children's Hospital, Denver; Ethan Stolzenberg. Dept. of Pathology, Mary Kay Gumerlock. Dept. of Neurosurgery, Kar-Ming Fung. Dept. of Pathology, Oklahoma University Health Sciences Center

Papillary tumor of the pineal region (PTPR) is a neuroepithelial tumor with unique histologic and immunohistochemical features and a wide differential diagnosis, ranging from pineal parenchymal tumors to metastatic carcinoma. Reports of PTPR to date have emphasized radiologic, morphologic, and/or immunohistochemical features, with few cases reporting electron microscopy (EM) findings. Here, we report the ultrastructural features of PTPR in a 70-year-old man presenting with features of Parinaud syndrome and obstructive hydrocephalus. Post-contrast MRI revealed a heterogeneously enhancing, cystic pineal region mass and subsequent endoscopic biopsy via the third ventricle procured both diagnostic and EM-dedicated tissue. Microscopic evaluation of the tissue revealed a papillary arrangement of columnar cells alternating with solid tumor, a low mitotic rate with focal tumor necrosis, vessel hyalinization, positivity with CK-8/ 18, pan-keratin, NSE, and synaptophysin, and negativity with EMA and GFAP. EM showed the epithelioid tumor cells to have abundant glycogen, coarsely granular chromatin, and irregular microvilli. Intercellular junctions between epithelioid cells included an apical zonula occludens and zonulae adherentes at both the apical and lateral cell membrane. Histiocyte-like cells were focally present underlying these cells. The solid tumor component featured a "mosaic"-type arrangement of organelle- and heterochromatin-poor cells with ovoid nuclei admixed with microtubule-rich cell processes and conspicuous concentric lamellae. Ependymal rosettes had zonulae adherentes organized in series and lumina with microvilli and cilia. Perivascular rosettes had cellular processes abutting capillary basement membranes. Dense-core vesicles, clear secretory vesicles, and perinuclear filaments were infrequent. This description confirms and extends key elements described by Jouvet and colleagues in their EM study of 2 cases (2003, American Journal of Surgical Pathology) and supports their assertion of a tumor with combined ependymal, secretory, and neuroendocrine features.

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42 Stem Cell Marker Expression and Genetic Mutations in Glioblastoma: A Comparison of Primary and Recurrent Tumors

Christina Appin, Matthew Schniederjan. Dept. of Pathology, Erwin Van Meir. Dept. of Neurosurgery, Gena Mastrogianakis. Dept. of Neurosurgery, Daniel Brat. Dept. of Pathology, Emory University School of Medicine

Background: Glioblastoma (GBM) carries a dismal prognosis, with an average survival of one year after standard therapy. Following surgical resection, radiation therapy and chemotherapy, recurrence is nearly inevitable, yet varies in time. Animal models have suggested that biological aggressiveness may be related to the stem cell compartment and have also indicated that the stem cell component is resistant to radiation therapy and enriched in GBM recurrences. We investigated whether the immunohistochemical (IHC) expression of stem cell markers was increased in recurrent GBM as compared to the primary tumor.

Methods: Paraffin-embedded tissue sections of primary and recurrent GBMs from 24 patients were stained by immunohistochemistry for Nestin, Sox-2, c-Myc and CD133, as well as p53. FISH was performed for EGFR status. Immunoexpression of stem cell markers was analyzed by examining 10 HPFs (40x) per slide and calculating an H-score. Differences between H-scores in primary and recurrent GBMs were evaluated with a rank sum test. EGFR amplification and p53 immunoexpression were correlated with expression of stem cell markers.

Results: Analysis of Nestin, Sox-2, c-Myc and CD133 revealed no statistically significant difference in IHC H-score in primary and recurrent GBMs. Nuclear p53 immunoreactivity was present in 11/23 (48%) primary GBMs and EGFR amplification was detected in 11/24 (46%). A slightly higher H-score for Sox-2 was noted in primary GBMs with EGFR amplification (mean = 2.8) compared to non-amplified cases (mean = 2.3; p < 0.05).

Conclusion: GBM recurrence was not associated with an increased immunoexpression of the stem cell markers used in this investigation. We did observe a significant association between Sox-2 expression and EGFR amplification in primary GBM. No association was noted with p53 and stem cell marker immunoexpression.

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43 Immunohistochemical Analysis of Actinin-4 in Malignant Gliomas

Shintaro Fukushima. Division of Pathology, National Cancer Center Hospital

Actinin-4 is an isoform of non-muscular a-actinin and actin-bundling protein, and plays an important role in cancer invasiveness and metastasis by enhancing cell motility. It has been reported that variable clinicopathological implications of actinin-4 has been demonstrated in some human malignancies such as breast cancers, colorectal cancers, ovarian cancers, and pancreatic cancers. In this study, we investigated the expression of actinin-4 with immunohistochemical analyses against malignant glioma cases, which have aggressive infiltrative ability. 37 histological diagnosed as malignant glioma among surgical resections registered. These included from December 2009 to January 2011 in our institute which include 5 diffuse astrocytoma(A), 1 oligoastrocytoma(OA), 5 anaplastic oligodendroglioma(AO), 3 anapl- astic astrocytoma(AA), 2 anaplastic oligoastrocyotma(AOA), 17 glioblastoma(GBM), 4 gliosarcoma(GS). Mean age was 51.6 years old ranging from 34 to 75, and male/female ratio was 2:1.The eligible tumors were subdivided into four groups according to the number of actinin-4 positive cells as follows: -(0%), +(<10%), ++(10-50%), +++(50%<). As a result, all 6 WHO grade II cases (A/OA) showed -, while many positive cells were demonstrated in grade III and IV cases (In 10 cases of AO/AA/AOA group, 5 and 2 were + and ++, respectively. In 21 cases of GBM/GS group, 2 and 18 were + and ++, respectively). It was indicated the expression of actinin-4 have a correlation with the WHO grade. We plan to further analyze more available cases, and report correlations of actinin-4 expression level with MIB-1 labeling index and patients' clinical outcome.

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44 Supratentorial Extension of Diffuse Intrinsic Pontine Glioma (DIPG)

Terri Haddix, Michelle Monje, Marilyn Masek, Morgan Freret. Stanford University Medical Center; Paul Fisher. Lucile Packard Children's Hospital; Hannes Vogel. Stanford University Medical Center

Diffuse intrinsic Pontine Gliomas (DIPG) account for approximately 80% of all pediatric brainstem tumors. DIPG carries a dismal prognosis, with median survival of 9 months and limited treatment options. Frustratingly little is known about the biology of this devastating pediatric tumor. The biologic behavior of these tumors has largely been followed radiographically with few cases undergoing biopsy or presenting for thorough postmortem characterization. We performed a systematic evaluation of tumor dissemination in an autopsy series of five patients with DIPG. The patients ranged in age from 3-14 years (mean 8.6 years); three patients were male. Time from diagnosis to death was 3 to 21 months (mean 11.6 months). The brains were examined and sectioned per routine and subjected to routine processing and staining. The tumors in the pontine region were WHO grade II-IV astrocytic neoplasms. Tumor infiltration into the midbrain was identified in all five cases. Tumor was identified in the medulla in only one case. Infiltration into the cerebellum was identified in every case. In four cases, tumor infiltrated supratentorially to involve the thalamus. Tumor infiltration as far as the frontal lobe structures was observed in two cases. The primary tumor in all cases showed a variable amount of nestin immunoreactivity; the greater anaplasia of the primary tumor and the aggressiveness of the infiltration correlated with greater nestin immunoreactivity. This small, but thoroughly sampled, autopsy series suggests that DIPGs are more widely disseminated than previously recognized. The frequent presence of supratentorial infiltration is previously unrecognized. These observations may help to elucidate DIPG pathogenesis and ultimately new treatment strategies.

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45 Atypical Teratoid/Rhabdoid Tumor Arising in a Ganglioglioma

Bette Kleinschmidt-DeMasters. University of Colorado Anschutz Medical Campus; Diane Birks. MS; Todd Hankinson. The Children's Hospital Dept of Neurosurgery; Marc Rosenblum. Sloan Kettering Dept of Pathology

Atypical teratoid/rhabdoid tumor (AT/RT) is an uncommon, aggressive, embryonal pediatric brain tumor that almost always develops de novo and does not arise within, or evolve from, other brain tumor types. While rhabdoid morphology can be seen in extrarenal tumor types including meningioma and glioblastoma, these are phenotypic mimics and, with only rare exceptions, do not manifest the INI-1 deletion at the 22q11.2 locus, monosomy 22, or INI-1 nuclear protein loss that characterizes conventional AT/RT. One of the authors (MKR) and colleagues previously reported a case of an optic pathway pilocytic astrocytoma that 9 years later, after extensive interim therapy, evolved into a mixed ganglioglioma-AT/RT. In that case, molecular genetic studies demonstrated a mutation in exon 9 of the INI1 gene in the tumor and selective loss of INI-1 nuclear protein only in the AT/RT portion. We now report a similar case in a 6-year-old male with acute onset of symptomatology, large right parietal mass, and mixed ganglioglioma-AT/RT morphology at onset, with relatively distinct separation between the two components. Numerous neoplastic neurons, Rosenthal fibers, low MIB-1 rate, and retention of INI-1 nuclear immunostaining characterized the ganglion cell portion, while rhabdoid cells, polyphenotypic IHC expression, high MIB-1 rate and loss of INI-1 nuclear expression were found in the AT/RT portion. The two areas were separately assessed for genetic features by fluorescence in situ hybridization and monosomy 22 was found only in the AT/RT component and not the ganglioglioma. These rare cases suggest a post-clonal acquisition of the genetic INI-1 signature is possible.

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46 Immunohistochemical Staining Profile of Seventeen Subependymal Giant Cell Tumors

Dana Altenburger, Alexander Bottini, Spencer Tung, Harry Vinters. University of California Los Angeles

Subependymal giant cell tumors (SGCTs, also known as subependymal giant cell astrocytoma/SEGA) are benign, non-infiltrating ventricular and periventricular tumors that arise almost exclusively in patients with tuberous sclerosis complex (TSC). Immunohistochemical (IHC) characteristics of these tumors are rather poorly described, sometimes providing conflicting data. We examined 17 SGCTs from 14 different patients (7 female, 7 male), ages 1-35, and performed a battery of immunohistochemical studies on the tumors, including use of primary antibodies against GFAP, Neu-N, Neurofilament Protein (NF), synaptophysin, and Ki-67. Glial fibrillary acidic protein (GFAP) immunohistochemical staining was seen in all 17 tumors at varying intensities from faint and focal, to strong and diffuse. Five cases showed variable staining within the same tumor. Eleven of 17 cases were negative for synaptophysin. Of the remaining six, the rare positive cells were felt to be most consistent with entrapped neurons, though synaptophysin immunoreactivity within tumor cells could not be ruled out. Nearly all tumors showed at least rare scattered tumor cells immunoreactive for neurofilament protein. Diffuse NF staining was not seen and one tumor was completely negative. The proliferation index, using Ki-67 IHC, was generally less than 5%. One tumor had a focally increased Ki-67 labeling index of 5-10%, while two other tumors had ranges from 5-15% and 10-20%.

CONCLUSION: Immunohistochemical features of SGCTs are variable, and suggest predominant astrocytic, less prominently neuronal differentiation. Ki-67 labeling indices are variable and probably not predictive of tumor recurrence.

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47 The Role of Metabotropic Glutamate Receptor Allosteric Modulators in the Investigation and Treatment of Gliomas

Hilary Nickols. Vanderbilt University; P Conn. Vanderbilt University Department of Pharmacology

Pharmacological therapy for the treatment of human gliomas, specifically glioblastoma, is severely limited. Antagonists of the Group II metabotropic glutamate receptors (mGluR2/3) inhibit glioma cell growth in vivo and in vitro, demonstrating their potential as therapeutic agents for malignant gliomas. The mGluR3 metabotropic glutamate receptor is expressed in primary human glioblastoma cultures, human glioma cell lines, including glioma initiating cells, and is the predominant mGluR subtype expressed in human astrocytes. Both mGluR2 and mGluR5 are variably expressed in glioma cell lines. Gliomas release glutamate, which results in both excitotoxic neuronal cell death and serves as a motogen, allowing for tumor expansion within the limited confines of the cranial cavity. Gliomas with reduced cell surface glutamate transporter expression grow more rapidly. mGluR3 and mGluR5 differentially regulate glutamate transporter expression in vitro, with mGluR5 agonists resulting in the downregulation of glutamate transporter expression. We are investigating the expression of metabotropic glutamate receptors in human glioma cell lines and primary human gliomas. Allosteric agents, which act at sites separate from the orthosteric glutamate binding site, provide potential novel therapeutic tools for human gliomas. The allosteric modulators function in conjunction with the endogenous ligand glutamate to either potentiate or inhibit the glutamate response. The availability of novel mGluR3 negative allosteric modulators (NAMs) as well as mGluR5 allosteric agents provides exciting tools in the investigation and potential therapy of human gliomas.

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48 Clonal 8q Amplification Detected in a Multicentic Anaplastic Astrocytoma by Array CGH

Michelle Madden Felicella. University of California San Francisco; Jill Hagenkord, Shera Kash. Creighton University; Martin Powers, Mitchel Berger, Arie Perry. University of California San Francisco

We report a case of a 27-year old right-handed man who presented with a nocturnal seizure. Imaging studies showed two radiologically separate and distinct non-enhancing infiltrative mass lesions in the anterior frontal lobe and the posterior temporoparietal regions, respectively. No intervening disease was evident on any of the magnetic resonance imaging (MRI) modalities and the lesions were relatively stable over a period of many months. He underwent two separate resections a few months apart which revealed histologically identical tumors that were both diagnosed as anaplastic astrocytoma, WHO grade III. Given the question of whether these two tumors represented two de novo primary tumors or just one multifocal tumor, comparative genomic hybridization via SNP array karyotyping was performed on formalin-fixed paraffin embedded tissue from both tumors. The two tumor profiles looked remarkably similar, with both tumors having a common 33Mb duplication of 8q23.30q24.3, suggesting that they originated from a single clone, rather than representing two synchronous glial neoplasms. The larger temporal neoplasm showed additional genetic alterations implying that it underwent secondary alterations not present at the other site. This case illustrates that even with today's advanced neuroimaging modalities, extensive radiologically invisible tumor may be present between seemingly separate sites of glioma involvement. Thus, modern global genomic studies of such tumors can help distinguish whether a multifocal glioma represents one extensive neoplasm with microscopically invasive disease in between radiologically discernable sites of involvement or multiple genetically distinct primary tumors.

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49 Anaplastic Oligodendroglioma in a Patient With Neurofibromatosis Type I

Ronnie Self Self. Tulane University; Liqiong Liu, John Gauchiani, Arthur Ulm. Louisiana State University; Zhenggang Xiong. Neuropathology Section, Department of Pathology, Tulane University

Oligodendroglioma is not documented in a patient with neurofibromatosis type 1 (NF-1). We present an unusual case of a 19 year old female patient with NF-1 and intraventricular anaplastic oligodendroglioma of the fourth ventricle. MRI identified a multi-lobated and multi-cystic mass expanding into the fourth ventricle with resultant hydrocephalus of all four ventricles and the cerebral aqueduct, extending slightly into the cerebellar vermis. Microscopically, the tumor showed moderate cellularity with diffusely infiltrating monomorphic neoplastic glial cells, with some cells showing a variable degree of nuclear atypia and some showing perinuclear halos. Vascular endothelial hyperplasia was present. Occasional mitoses were seen. Immunohistochemical studies revealed a Ki67 labeling index of approximate 5%. These histopathological and immunohistochemical findings are consistent with anaplastic oligodendroglioma. While glial neoplasms are well documented in NF-1 patients, these tumors are typically astrocytic and most commonly affect children. Additionally, astocytomas and oligodendrogliomas are known to follow different genetic pathways. Our case is contributory to understanding the tumorigenesis of glioma and neurofibromatosis.

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50 pStat3 Immunoreactivity in Glioblastomas and Atypical Teratoid/Rhabdoid Tumors

Jian-Qiang Lu. Department of Laboratory Medicine & Pathology, Beverly Wilson. Department of Pediatrics, Lothar Resch. Department of Laboratory Medicine and Pathology, Jeffrey Pugh. Department of Surgery, Vivek Mehta. Department of Surgery, University of Alberta

Signal transducer and activator of transcription (Stat3) has been suggested to be crucial in biology of glioblastomas and also a useful marker for inflammatory activity, as well as a potential target for novel treatments of glioblastomas and inflammatory activity. Phosphorylated Stat3 (pStat3) has been reported in many non-CNS cancers, but rarely reported in human gliomas and not yet reported in other CNS tumors. We examined pStat3 immunostaining in 5 cases of glioblastomas (age range: 30 ∼ 72 years; 3 male and 2 female) and 4 cases of infratentorial atypical teratoid/rhabdoid tumors (AT/RTs; ages: ∼ 2 years; 3 male and 1 female). All glioblastomas showed pStat3-immunoreactive nuclei which percentage ranged from 10% to 90% of the cells, likely depending on the genetic background of the tumor (de la Iglesia et al. Curr Mol Med. 2009; 9: 580-590). In AT/RTs, only 10 - 50% of the nuclei were immunoreactive for pStat3. The immunoreactivity for pStat3 appeared not to be correlated with that for glial fibrillary acidic protein, epithelial membrane antigen, cytokeratins, smooth muscle actin, synaptophysin, neurofilament protein or MIB-1 labelling index of proliferation. The pStat3-immunoreactive cells were similar to the cells with retained INI-1 immunoreactivity in distribution and morphology. The pStat3 immunoreactivity was also identified in the cells of blood vessels within and adjacent to the tumors but not consistently in the adjacent cerebellar tissues. All AT/RTs exhibited moderate infiltration of CD68-immunoreactive macrophages/microglia and CD4/CD8-immunoreactive T-cells, which may be related to the innate immunosuppressive properties of the brain cancer stem cells and their mediation by pStat3 (Wu et al. Neuro-Oncology 2010; 12:1113-1125). The findings of this study suggest that pStat3 may be a useful marker for diagnosis of glioblastomas and AT/RTs, and its immunoreactivity in AT/RTs may share some common characteristics with the INI-1 immunoreactivity.

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51 Mutant Isocitrate Dehydrogenase 1 Immunohistochemistry: Not Useful in the Diagnosis of Low Grade Diffuse Astrocytomas in NF1

Anat Stemmer-Rachamimov. Massachusetts General Hospital; Harvard Medical School; Sandra Camelo-Piragua. Massachusetts General Hospital; Scott Plotkin. Massachusetts General Hospital

Background: Gliomas are common malignancies in neurofibromatosis type 1 (NF1) patients. Although most NF1-associated gliomas are pilocytic astrocytomas (PAs), diffuse astrocytomas also occur, and the distinction between PA and diffuse gliomas may at times be challenging. In addition, the differential diagnosis between reactive gliosis and low grade astrocytomas may be more difficult because of the presence of scattered atypical glial cells in the white matter of NF1 patients. IDH1 was shown to be a reliable marker in distinguishing low grade astrocytomas from reactive astrocytosis in sporadic (non-NF1) patients. Isocitrate dehydrogenase 1 (IDH1) mutations have been shown in 40%-70% of low grade and high grade infiltrating sporadic astrocytomas and the most common IDH1 mutation, a change of arginine to histidine in codon 132 (R132H), can be detected by immunohistochemistry. In addition, due to the low incidence of IDH1 mutations in PAs, IDH1 immunohistochemistry may be useful in the distinction between low grade diffuse astrocytomas and PAs when histology alone is not definitive.

Aim: To evaluate the utility of the mutant IDH1 immunohistochemistry as a marker for diffuse low grade astrocytoma in NF1 patients.

Methods: Six surgical specimens of diffuse low grade astrocytomas from NF1 patients were retrieved from the pathology archives at the Massachusetts General Hospital. PAs were excluded. The slides were reviewed by two pathologists (SCP & ASR) to confirm the diagnosis. Mutant IDH1 immunohistochemistry was performed following previously published protocol with positive and negative controls.

Results: All six diffuse infiltrating NF1-associated astrocytomas were negative for the mutant IDH1 antibody.

Conclusion: Our study shows that in contrast to previously published studies in sporadic (non NF1) gliomas, mutant IDH1 expression is not present in NF1-associated low grade astrocytomas. Therefore in the NF1 patient, mutant IDH1 expression is not helpful for distinguishing low grade gliomas from reactive astrocytosis or from PAs.

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52 Pathology of 'Pseudoprogression' in a Phase II Study of PPX, TMZ, and RT for Newly Diagnosed High-Grade Gliomas

John Donahue. Dept. of Pathology, Rhode Island Hospital/Alpert Medical School; Suriya Jeyapalan. Brown University Oncology Group, Rhode Island Hospital; Marc Goldman. Dept. of Neurosurgery, Rhode Island Hospital/Alpert Medical School; Heinrich Elinzano. Brown University Oncology Group, Rhode Island Hospital; Jerrold Boxerman. Dept. of Radiology, Rhode Island Hospital/Alpert Medical School; Thomas DiPetrillo, Howard Safran. Brown University Oncology Group, Rhode Island Hospital

Paclitaxel poliglumex (PPX) is a drug conjugate that links paclitaxel to a polyglutamic acid polymer, yielding a radiosensitization index of 4-8, as compared to 1.5-2 for paclitaxel alone. Phase I/II studies of PPX in esophageal cancer established that 50 mg/m2/week of PPX can be safely administered with concurrent chemotherapy and radiation therapy (RT). The primary objective of this study is to determine the safety of PPX with standard temozolamide (TMZ) and RT for high-grade gliomas. Patients received weekly PPX 50 mg/m2 and daily TMZ 75 mg/m2 for 6 weeks with concomitant RT (60 Gy). Adjuvant chemotherapy with TMZ (200 mg/m2/d x 5 d), every 28 days, was started 1 month after completion of chemoradiation. This study completed planned accrual of 25 patients (median age, 60 years; 48% male; 60% glioblastomas [GBM]). The 6-month progression-free survival (PFS) of GBM patients was 66.7% (10/15). Median PFS was 12.5 months. Pseudoprogression (PsdP) was prominent. 16 patients had post-PPX enhancement, and 5 were classified with progressive disease via radiologic analysis of regional cerebral blood volume (rCBF). (Mean time to progression was 4.6 months.) The 5 patients with apparent progression by MRI and rCBF analysis had surgical resection (2) or stereotactic needle biopsy (3). Pathology in all cases showed high-grade glioma admixed with nonradiation necrosis. However, the proliferative index, as measured using immunohistochemical staining for Ki-67, was less than 5%. Thus, the tumors were rendered `quiescent' by the therapy. In conclusion, PPX is a promising radiation enhancer in high grade gliomas, and early, apparent progression of tumor when treated with PPX/TMZ/RT likely represents 'pseudoprogression,' with the tumor being rendered 'quiescent' by treatment. [Supported by Cell Therapeutics, Inc., Seattle, WA.]

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53 p53 Deletion Synergizes With Oncogenic K-ras in a Transgenic and Orthotopic Transplant Model of Giant Cell Glioblastoma

Ty Abel, Sabah Ghazi, Ping Li. Vanderbilt University Medical Center; Harold Moses. Vanderbilt University; Michelle Stark. Vanderbilt University Medical Center

Glioblastoma multiforme (GBM) is one of the most aggressive human cancers, and the most common primary, adult CNS neoplasm. We recently reported a novel, transgenic mouse model in which conditional activation of oncogenic K-ras (K-rasG12D) in the brain resulted in infiltrating glioma. The glioma showed histopathologic features that closely mimicked intermediate-grade, human tumors, and glioma cells expressed markers associated with neural stem/progenitor cells (NSPC). In addition, the mice showed marked expansion of the subventricular zone (SVZ), a NSPC population in the brain. We have now developed a mouse model of glioblastoma multiforme. p53 mutation is a common genetic alteration in human glioma specimens, particularly in giant cell GBM. Again using hGFAP-Cre mice, we deleted both p53 alleles in concert with activation of K-rasG12D. The intermediate-grade gliomas induced by activation of K-rasG12D alone are converted, with concomitant deletion of p53, to high-grade, lethal gliomas. The infiltrating, GFAP-immunoreactive, neoplasms are histologically reminiscent of giant cell glioblastoma, showing large, grotesque cells, many of which are multinucleated. Microscopic examination of the brains of two-week-old, hGFAP-Cre/K-rasG12D/p53fl/fl pups shows early tumor formation around the SVZ. Cultured astrocytes harvested from hGFAP-Cre/K-rasG12D/p53fl/fl pups and injected into the brains of adult mice formed lethal tumors at approximately 3 weeks post-injection. This model may be useful as a pre-clinical tool for studying novel glioma therapies. In addition, it will be possible to use this model to address basic questions about glioma biology, including the role of NSPC, and tumor-microenvironment interactions.

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54 Gliosarcoma Arising in an Oligodendroglioma (Oligosarcoma)

Annie Hiniker, Arie Perry. University of California San Francisco, Department of Pathology

A 57-year-old man with familial thrombocytopenia presented with altered mental status and was found to have a large right frontal hemorrhagic mass extending to the corpus callosum with hemorrhage into the right lateral ventricle. Stereotactic biopsy and subsequent subtotal resection revealed a WHO grade II oligodendroglioma with classic histological features and chromosome 1p19q codeletion by FISH analysis. The patient subsequently received adjuvant radiation therapy, but developed recurrent tumor fifteen months after initial presentation. A biopsy of the recurrent tumor showed minimal residual features of classical oligodendroglioma, but instead appeared hypercellular, relatively solid and composed of intersecting fascicles of spindled cells with associated necrosis and high mitotic rate. The majority of tumor cells were smooth muscle actin-positive and demonstrated intercellular reticulin deposition; a smaller subset was GFAP immunoreactive, consistent with the diagnosis of gliosarcoma, a biphasic tumor more typically considered a variant of glioblastoma, WHO grade IV. Further testing for 1p19q codeletion, EGFR amplification, chromosome 10 loss, and IDH1-R132H expression in the recurrent tumor is pending. There have been only rare reports of sarcomatous transformation of oligodendroglioma (`oligosarcoma') and most were published prior to the development of modern genetic modalities. Although rare in comparison to astrocytic neoplasms, mesenchymal metaplasia with sarcomatous transformation is possible in classic oligodendroglial neoplasms with 1p19q codeletions. Published data suggest that patient prognosis is markedly worsened when this occurs.

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55 Quantum Dot Probes for Glioma Cell Detection

Cody Weston, Achuthamangalam Madhankumar, Jennifer Baccon, Michael Glantz, James Connor. Penn State M. S. Hershey Medical Center

Abstract: Identification of circulating cancer cells has broad clinical relevance. In the cerebrospinal fluid (CSF), free cancer cells are associated with leptomeningeal metastasis and neoplastic meningitis, both carrying a dire prognosis. Currently, these phenomena are diagnosed by CSF cytology, magnetic resonance imaging, or neurologic exam, which are all limited by low sensitivity. Improved detection methods could allow for early treatment and better prognosis. We have designed a strategy to use quantum dots as high-intensity, photostable fluorophores to identify cancer cells in CSF. We first sought to determine if the quantum dots could be used to specifically target free glioma cells in vitro.

Methods: Human U251 cells were used to model free glioma cells, and were cultured at 37 degrees C using ATCC-recommended media. Carboxylic acid modified quantum dots (Ocean Nanotech) were conjugated to plasmid-derived IL-13. To demonstrate effective conjugation, conjugated and unconjugated quantum dots were compared using agarose gel electrophoresis. To assess the binding properties of the IL-13 bound quantum dots, cells at a density of 20,000 cells/well in chamber slides were subjected to a binding protocol.

Results: Agarose gel electrophoresis demonstrated successful conjugation of functionalized quantum dots to targeting ligands. Fluorescence microscopy demonstrated that an IL-13 conjugated quantum dot bound to fixed U251 cells, but did not bind to U251 cells following receptor blocking with 100-fold excess of IL-13, demonstrating IL-13 Receptor dependent binding.

Conclusions: Conjugation of quantum dots to antibodies and specific targeting ligands yields a fluorescent probe for in-vitro immunocytochemistry and immunohistochemistry applications. The use of targeted quantum dots has potential applications as a mechanism to use flow cytometry and other cell-based diagnostic procedures for higher throughput and increased sensitivity.

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56 Utility of Spectral Image Analysis For Prediction of 1p/19q Deletion Status in Gliomas

Kimberly Stogner-Underwood, Colleen Jackson-Cook, Carleton Garrett, Hope Richard, Catherine Dumur, Christine Fuller. Virginia Commonwealth University Health System

Divergent diagnostic approaches and subjective morphologic criteria have resulted in considerable inter-observer variability in the routine classification of infiltrative gliomas. 1p/19q deletion, implicated as a marker of "oligodendroglial phenotype," is present in most oligodendrogliomas (O) and few oligoastrocytomas (OA); importantly, its presence strongly correlates with prolonged survival and increased chemoresponsiveness. Spectral image analysis (SIA) provides rapid accurate detection/classification of objects by shape, size, and spectral composition. To evaluate the utility of SIA for identifying gliomas harboring 1p/19q deletion, we assembled a study set of 50 gliomas with known 1p/19q status; original diagnoses included 21 O, 3 OA, and 17 "glioma with O features". H&E stained slides from each case were subjected to 1) histological review by a single neuropathologist applying strict morphologic criteria for O classification, and 2) spectral image analysis of nuclear area, roundness, perimeter, and intensity. Statistically significant differences for the means (M), standard deviations (SD), and coefficients of variation (CoV) for the Area and Perimeter variables relative to 1p/19q deletion status were detected. Interestingly, while mean nuclear area was smaller in 1p/19q deleted tumors (p<0.0002), the area SD and CoV measures were likewise significantly smaller (p<.00093), indicating one potential morphological correlate with 1p/19q deletion is nuclear uniformity. Analysis using compound covariate predictor and linear diagonal discriminant analysis classification rules indicated sensitivities and specificities of 78% and 82% respectively for both classifiers, improving somewhat on the levels of sensitivity in comparison with the initial O diagnoses. However, expert histological reclassification as O gave results of 89% sensitivity and 96% specificity (agreement with 1p/19q; Kappa = 0.87; P= 8.0E-10). Though accurate histological classification of O remains the strongest predictor of 1p/19q deletion, SIA holds promise in discriminating those gliomas likely to harbor 1p/19q deletion. Additional studies are necessary to determine the most appropriate use of this technology.

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57 Is Polysomy of Chromosome 1p and/or 19q in Oligodendrogliomas Prognostically Significant?

Andrea Wiens. Indiana University School of Medicine; Karen Johnson. University of Wisconsin-Madison; Liang Cheng, Eyas Hattab. Indiana University School of Medicine

Background: It is well established that codeletion of 1p and 19q (del1p/19q) is a positive prognostic molecular event in oligodendrogliomas (OG). One recent report suggested that polysomy of chromosomes 1 and 19 in the presence of del1p/19q correlates with earlier recurrence in anaplastic oligodendrogliomas (AOG)(Snuderl 2009). The purpose of this study was to elucidate whether polysomies of chromosomes 1 and 19 are of prognostic value in oligodendrogliomas with respect to tumor grade, tumor recurrence, and overall survival. Materials and methodsOur cohort consisted of 90 consecutive cases (69 WHO grade II OGs, 18 WHO grade III AOGs and 3 glioblastomas with oligodendroglial features (GBM-O)) diagnosed at Indiana University from 2005-2010. All cases were previously analyzed for 1p32/19q13 deletion by FISH using dual color break-apart probes. 1p/19q deletion status was recorded with accompanying polysomy status, WHO grade, recurrence free survival and overall survival. Statistical analysis using Fisher's Exact Test and Kaplan-Meier survival curves was performed. ResultsDel1p/19q was detected in 42 of 90 (46.7%) cases (36/69 OGs, 6/18 AOGs, and 0/3 GBM-Os), showing better overall survival (p=0.009), as expected. 33 of 90 cases (36.7%) showed polysomy of chromosome 1, while 34 of 90 cases (37.8%) showed polysomy of chromosome 19, and 30 of 90 cases (33.3%) showed polysomies of both chromosomes 1 and 19. We found no statistically significant effect of polysomy of chromosome 1 alone, chromosome 19 alone, or both chromosomes 1 and 19 with regards to WHO grade (p=0.460, p=0.245, p=0.402 respectively), recurrence free survival (p = 0.283, p = 0.434, p = 0.613), or overall survival (p = 0.524, p=0.294, p=0.379). Conclusions In our series, polysomies of chromosomes 1 and/or 19 do not correlate with tumor grade, recurrence free survival, or overall survival. Conforming to previous studies, the presence of del1p/19q is a positive prognostic indicator in oligodendrogliomas.

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58 Orbital Glioma With Focal Rhabdoid Morphology

Andrea Wiens, Richard Burgett. Dept. of Ophthalmology; Mitesh Shah. Dept. of Neurosurgery; Eyas Hattab, Jose Bonnin. Indiana University School of Medicine, Dept. of Pathology

Orbital tumors are relatively uncommon in young children. They include a range of benign and malignant conditions including dermoid cyst, hemangioma, and rhabdomyosarcoma. Optic nerve gliomas and metastatic neuroblastomas may also be observed. We report a case of an orbital tumor in a girl who presented with a two- to three-week history of swelling in the right eye. The swelling resolved spontaneously but a bluish black discoloration remained. Neuroimaging studies revealed a heterogeneously enhancing 29×26×24 mm retrobulbar mass with apparent central necrosis and involvement of the superior rectus, superior oblique and medial rectus muscles. The anterior portion of the optic nerve and the eye were displaced anteriorly and inferiorly. The tumor extended to the bony portion of the optic canal which was expanded. A right orbitotomy revealed a circumscribed tumor mass, and a biopsy through the tense capsule revealed very soft yellowish and apparently necrotic tissue. No mass reduction was achieved after a course of chemotherapy, and the tumor was resected. Pathologic examination showed a tumor mass within the confines of a greatly expanded optic nerve sheath. Necrosis was not observed, but there were large areas with a prominent myxoid background. Cytologically, there were areas with oligodendroglial-like features and scattered individual and small whorled clusters of cells with rhabdoid morphology. Degenerative cellular atypia, eosinophilic granular bodies, rare mitoses, and proliferation of the vascular endothelium were observed, but there were no Rosenthal fibers. The tumor cells were positive for glial fibrillary acidic protein, and the Ki-67 labeling index was low. Reactive meningothelial cells were noted in the periphery. This is an example of an optic nerve glioma with unsual cytologic features that may be misinterpreted as other forms of more aggressive orbital neoplasms.

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59 Determination of RB Mutation Status in Glioblastoma and Correlation With Data From The Cancer Genome Atlas

Patricia Goldhoff. Department of Pathology, UCSF; Ivan Smirnov, C. James. Department of Neurological Surgery, UCSF; Arie Perry, Joanna Phillips. Department of Neurological Surgery and Pathology, UCSF

Glioblastoma (GBM) is a heterogeneous disease with a poor prognosis. Therapeutic targeting of signaling pathways critical in GBM may lead to improvements in patient care. The success of targeted therapy, however, depends on the identification of signaling pathway alterations and thus potential susceptibility to specific therapies. The CDKN2/p16-CDK4/6-RB pathway is commonly altered in GBM and includes deletions of p16, deletions of RB, and CDK4 amplification. A recent study, using a preclinical model for GBM, identified RB status as an important determinant of potential efficacy for CDK4/6-specific inhibitors (Michaud K. et al., 2010). To determine if RB status could be assessed in human clinical tumor samples, we analyzed 33 GBM tumors by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) and compared these results to paired array comparative genomic hybridization (array CGH) data and expression array data from The Cancer Genome Atlas (TCGA). To do this we generated tissue microarrays from tumors analyzed by TCGA and performed FISH analysis and IHC for RB. There was 100% (11/11) concordance between FISH analysis for RB status and TCGA array CGH data. Similarly, all tumors with absent RB protein by IHC had low-level expression of RB transcript based on TCGA expression data. In a total of 33 tumors, 3 (9.1%) had loss of RB protein expression by IHC, and these tumors had either homozygous deletion of RB by FISH or hemizygous deletion of RB by FISH combined with a novel nonsense mutation in RB. We are analyzing RB status by IHC in an independent set of GBMs. In this study we will address the advantages and disadvantages of IHC and FISH for detecting RB alterations and the potential utility of these approaches in the clinical stratification of patients.

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60 Ependymosarcoma: Light and Electron Microscopic Study of a Case

Kael Mikesell, Cathy Housman. Dept of Pathology, Penn State - Hershey Medical Center; Jonas Sheehan. Dept of Neurosurgery; Charles Specht. Penn State - Hershey Medical Center

Ependymosarcoma is a rare tumor that contains areas of ependymoma and sarcomatous change. Our patient is a 69 year old male who presented with a six week history of dizziness and unsteady gait. Imaging studies revealed a 4.5 × 3.5 × 3.2 cm enhancing multilobulated mass in the left cerebellar hemisphere; this mass had solid and small cystic/necrotic components. The tumor effaced the fourth ventricle. Gross total resection was performed. Histological sections were stained with hematoxylin-eosin (H&E) and a method for reticulin, and with immunohistochemistry for glial fibrillary acidic protein (GFAP), epithelial membrane antigen (EMA), synaptophysin, neurofilament protein (NFP), chromogranin, smooth muscle actin (SMA), vimentin, and MIB-1 (Ki-67). The tumor was also evaluated with transmission electron microscopy (TEM). The H&E-stained sections show neoplastic ependymal cells and sarcomatous spindle cells with nuclear atypia and scattered mitotic figures. The Ki-67 nuclear labeling index ranges focally up to 25%. Perivascular pseudorosettes are seen in the more GFAP-positive ependymomatous areas; these areas show some dot-like cytoplasmic reactivity for EMA. The sarcomatous areas consist of GFAP-negative neoplastic spindle cells that have abundant pericellular reticulin. These spindle cells exhibit patchy positive reactivity for SMA. Vimentin is positive throughout the tumor. The tumor cells are negative for synaptophysin, NFP, and chromogranin. Lack of entrapped NFP-positive axons through much of the tumor is consistent with a solid architecture. TEM reveals intracellular microlumens, many microvilli, intermediate filaments, and elongated intercellular junctions among the neoplastic cells. This is the first report of the ultrastructure of ependymosarcoma. Findings consistent with an ependymal neoplasm are seen. In follow-up, the patient was treated with local radiation therapy. Recurrence in the surgical bed was not seen during one year post-surgery.

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61 Massive Gliosis of the Retina - A Müller Cell Proliferation or an Intraocular Ependymoma?

Dana Timek, Cathy Housman. Dept of Pathology, Penn State - Hershey Medical Center; Michael Wilkinson. Dept of Ophthalmology, Charles Specht. Penn State - Hershey Medical Center

Massive gliosis of the retina is an uncommon lesion that may occur with various forms of retinal injury. Recently, cases of retinal ependymoma have been reported. We have studied a case of massive retinal gliosis with ependymoma-like features. The literature was reviewed. Our patient is a 33 year old male with proliferative diabetic retinopathy, recurrent vitreous hemorrhage, and blind painful left eye. This eye was enucleated; sections were stained with routine histochemical methods and with immunohistochemistry for glial fibrillary acidic protein (GFAP) and epithelial membrane antigen (EMA). Transmission electron microscopy (TEM) was also performed. By light microscopy, reactive and degenerative changes include iris neovascularization, fibrovascular organization of vitreous, and total retinal detachment with organizing subretinal hemorrhage. The detached retina is thickened by a mass containing variably GFAP-positive cells with long processes; perivascular pseudorosettes are present and dot-like cytoplasmic reactivity for EMA is demonstrated. By TEM, intracytoplasmic lumens, microvilli, cytoplasmic intermediate filaments, and elongated intercellular junctions are seen. Such findings may occur in ependymoma. However, ependymal cells are not normally found in the retina and sustained reactive proliferation of ependymal cells is not seen in the post-fetal central nervous system (CNS). A proliferative lesion of ependymal cells in the post-fetal CNS is designated ependymoma, a neoplasm. The Müller cell, a glial cell normally found in the retina, can proliferate in the adult in response to various types of injury. Indeed, all the ultrastructural and immunohistochemical findings in our case of massive retinal gliosis can be explained by a reactive proliferation of Müller cells. It is proposed that, although lesions of massive retinal gliosis may resemble ependymoma, the histological, immunohistochemical, and ultrastructural findings can be explained by a proliferation of Müller cells, a normal retinal cell capable of reactive change. Intraocular ependymoma is thus an unlikely designation for this type of lesion.

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62 Mitotic Counts and the Accuracy of Tumor Grading

Jeffrey Joseph. University of Calgary

While important for grading, the methodology for counting mitoses has not been standardized. Accurately grading an individual tumor is fundamentally different from the methodology used to derive statistically significant mitotic thresholds in large population studies. Assuming that mitoses occur at random places brings mathematical limitations on the confidence of mitotic counts. Using this assumption, this presentation examines three questions: 1. Given an actual tumor mitotic index and assuming mitotic frequencies follow a Poisson distribution, what is the probability that a measured mitotic count is in error (defined as a pathologist's count that undergrades a higher-grade tumor or overgrades a lower-grade tumor)? 2. Given a pathologist-measured mitotic count and again assuming a Poisson distribution, what are the confidence windows on the actual tumor mitotic index?3. Given a measured mitotic count and a known grading mitotic threshold, again assuming mitoses follow a Poisson distribution and using the gamma function for the calculation, what is the probability that an error has occurred in grading? (What is the probability that the measured count is above the threshold when the real tumor index is below the threshold? What is the probability that the count is below the threshold when the real tumor average lies above the threshold?)Grading accuracy, especially near a threshold mitotic count, requires either counting of numerous fields to reduce stochastic noise, use of markers that provide greater sensitivity to proliferation, or modification of grading criteria to include more tumor characteristics than mitotic counts alone.

LEARNING OBJECTIVES: 1. To understand the mathematical limits imposed by the Poisson distribution on the confidence of a pathologist's tumor mitotic count and hence on the accuracy of grading a tumor.

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63 Primary Diffuse Leptomeningeal Gliomatosis - A Case Study

Maria Martinez-Lage, Zissimos Mourelatos. Dept of Pathology and Laboratory Medicine, University of Pennsylvania

Involvement of the leptomeninges by a primary glial neoplasm has an almost invariably ominous prognosis, but an early diagnosis may confer the possibility of receiving chemoradiation, offering a potential increase in life expectancy. A 76 year old female presented with 2-4 weeks of occipital headache, nausea, vomiting, confusion and ataxia. Examination revealed lethargy with no focal neurological deficits, and in the following days she became increasingly unresponsive. An MRI of the brain showed diffuse meningeal enhancement around the cervical cord and brainstem, and a second MRI showed multifocal dural based, leptomeningeal and cranial nerve enhancement. A body PET scan showed diffuse uptake in cervical and upper thoracic spinal cord. Multiple microbiological tests were negative. Three cytologic examinations of CSF were negative for malignancy, although there was pleocytosis (up to 62 WBC/uL) and increasingly high protein content (up to 361 mg/dL). A blind brain biopsy showed focal mild leptomeningeal inflammation and bacterial, mycobacterial and fungal cultures were negative. The patient developed worsening hydrocephalus with no improvement of neurological status despite optimal treatment and expired 8 weeks after onset of symptoms. Postmortem examination revealed extensively opacified and thickened meninges with diffuse nodularity, predominantly in the basal surface. Upon serial sectioning, there was no evidence of a mass-forming lesion. Microscopic examination demonstrated extensive leptomeningeal involvement by a GFAP positive, focally p53 positive malignant neoplasm with perivascular distribution and encasing of cranial nerve roots. There was minimal focal invasion with pseudopalisading necrosis at the basis pontis. A diagnosis of primary diffuse leptomeningeal gliomatosis was made. To address the role of CSF immunocytochemistry with novel antibodies in improving the diagnosis of this entity, studies on retrospective CSF samples from this case and a second archival case will be reported.

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64 Characterization of R132H Mutant Form of Isocitrate Dehydrogenase 1 and Olig2 Expression Patterns in Glioblastoma Variants

Nancy Joseph, Michelle Madden, Joanna Phillips, Arie Perry. UCSF Department of Pathology

Background: Recent work has demonstrated that the bHLH transcription factor, Olig2, is nearly universally expressed in human gliomas and that the R132H IDH1 mutant protein is expressed in secondary rather than primary forms of glioblastomas (GBM). However, these diagnostic and prognostic markers have not been widely examined in rarer glioblastoma variants, such as small cell GBM, gliosarcoma, GBM with a primitive neuroectodermal (PNET)-like component, and GBM with an oligodendroglial component.

Design: IDH-R132H and Olig2 immunohistochemistry was performed in 30 small cell GBMs and 21 GBMs with PNET-like foci. Additional glioblastoma variants including gliosarcoma and glioblastoma with an oligodendroglial component are currently being analyzed.

Results: IDH1-R132H expression was seen in 0/30 (0%) small cell GBMs and in 3/21 (14%) GBMs with PNET-like foci, with 1/3 demonstrating positivity in both the glial and PNET-like components and 2/3 showing expression limited to the glial component. Olig 2 expression was seen in all 30 (100%) small cell GBMs and 18/21 (86%) GBMs with PNET-like foci, with 15/18 (83%) showing nuclear expression in both components, and 3/18 (16.7%) cases showing immunoreactivity only in the glial component.

Conclusion: The small cell variant of GBM typically expresses Olig2, but is negative for the R132H IDH1 mutation, a pattern consistent with primary glioblastomas in general. In sharp contrast to CNS PNETs which often enter into the differential diagnosis, GBMs with PNET-like foci usually retain Olig2 expression in both the glial and primitive components,. Although many of these tumors have features suggesting the secondary form of GBM, IDH1-R132H expression was surprisingly rare, though other forms of IDH1 and IDH2 mutations were not tested. Application of new markers to rare glioma subtypes provides new insights and may be of diagnostic assistance, particularly in ruling out important diagnostic considerations such as CNS PNET.

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65 Deep White Matter Venous Infarction in Infants: A Neuroradiologic-Neuropathologic Correlation

Christopher Pierson. Department of Laboratory Medicine, Jerome Rusin. Department of Laboratory Medicine,Department of Radiology, Nationwide Children's Hospital

Deep white matter venous infarction is thought to be rare in infants; however, it is commonly recognized at our institution due to recent advances in neuroimaging which has demonstrated a spectrum of injury from punctate periventricular white matter infarction to diffuse hemorrhagic white matter venous infarction and subsequent white matter liquefaction. We have identified two infants with venous infarction on MR imaging with correlative neuropathologic autopsy findings. Both infants were term-born males. At 5 weeks of age, Case 1 developed severe gastroenteritis-induced dehydration, resulting in diffuse superficial dural venous sinus and deep intramedullary venous thrombosis. Gross examination showed bilateral hemorrhagic necrosis of cerebral white matter with extensive intraventricular hemorrhage and hemorrhagic necrosis of the thalamus. Histopathology showed edema and diffuse venous congestion accompanied by extensive acute white matter hemorrhagic necrosis with numerous ring hemorrhages, some of which had centrally located veins containing occlusive thrombi. The thalamus showed extensive hemorrhagic injury with numerous ring hemorrhages and thrombi. Case 2 had hypoplastic left heart syndrome and died at 4 weeks of age due to multiple organ failure. Two premorbid MRI scans had demonstrated swelling and hemorrhage of the atrial choroid plexus; several punctuate hemorrhagic venous infarcts in the periventicular white matter, and superficial venous infarcts in the left occipital lobe and left Rolandic cortex. Gross autopsy examination showed multifocal white matter necrosis, periventricular venous congestion, and engorged, hemorrhagic atrial choroid plexus. Histology showed edema, engorged veins, and periventricular leukomalacia (focal white matter necrosis with diffuse white matter gliosis). We believe these cases are important because they demonstrate rare neuroradiologic-neuropathologic correlation of two cases along the spectrum of deep white matter venous infarction in infants and in addition illustrate that thrombosis within the deep venous system can be associated with deep white matter as well as thalamic infarction.

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66 Unusual Low-Grade Neuroepithelial Neoplasm With Pseudopapillary Growth Pattern and Perivascular Collagen Deposition

Nguyen Vo. Department of Pathology; Eduardo Gonzalez-Toledo. Department of Neuroradiology; Shashikant Patil. Department of Neurosurgery; Jon Wilson. Louisiana State University Health Sciences Center Shreveport

We present the clinicopathologic features of an unusual low-grade neuroepithelial neoplasm arising in the left parietal cortex of a 25 year-old female. This small, well-vascularized and circumscibed lesion demonstrated monontonous oligodendroglial-like cells, a pseudopapillary growth pattern, Rosenthal fibers and sparse eosinophilic granular bodies. At the periphery of the lesion the neoplastic cells were arranged around blood vessels in an almost `meningioangiomatous-like' pattern. Small intratumoral blood vessels demonstrated variably thick cuffs of perivascular collagen which contained neoplastic glial cells. The possibilities of papillary glioneuronal tumor or pilocytic astrocytoma were considered, but the morphologic and immunophenotypic features did not support these differential diagnoses.

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67 Tissue Factor Expression in Glioblastoma Does Not Correlate With Clinical Development of Venous Thromboembolism

Carrie Mohila, Stephen Cook, Kaanchan Gangal, Dawit Aregawi, Nigel Key. University of Virginia M. Lopes, David Schiff. University of Virginia

Patients with glioblastomas have a high risk of venous thromboembolism (VTE) that extends beyond the immediate postoperative period. Tissue factor (TF) is a potent initiator of coagulation protease cascades and is expressed by neoplastic cells in a variety of cancers. TF protein is expressed in gliomas, and expression directly correlates with glioma grade. To determine if TF expression in glioblastomas (GBMs) correlates with the clinical development of VTE, a total of 90 GBMs in patients with and without VTE were selected from the University of Virginia Neurooncology database. VTE cases were matched to non-VTE controls on potential clinical risk factors for VTE. After initial review of the pathology, 17 GBMs were eliminated because of insufficient tissue. Retrospective immunohistochemical studies using anti-TF antibodies were performed. In patients with VTE, 28 out of 39 (71.8%) GBMs were positive for TF; in patients without VTE, 29 out of 33 (87.9%) GBMs expressed TF. Most GBMs showed heterogenous staining primarily in a fibrillary pattern. The frequency and pattern of tumoral staining did not differ in patients with and without clinical VTE. These findings suggest that tumoral TF immunostaining does not appear to be a risk factor for VTE in GBM patients and that other prothrombotic mechanisms contribute to the increased VTE risk.

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68 Glioblastoma Occurring at the Site of a Previous Medulloblastoma Following a Five Year Remission Period

Sarah Martin. Indiana University School of Medicine, Department of Pathology; Daniel Brat. Dept. of Pathology, Emory University School of Medicine; Annette Douglas-Akinwande. Dept of Neuroradiology; Eyas Hattab. Indiana University School of Medicine, Dept. of Path. and Lab. Med.

We describe a case of a 14-year-old boy who developed a cerebellar and brainstem glioblastoma five years following treatment for a medulloblastoma. The patient first presented in 2003 with nine months of vomiting and a 20-pound weight loss. A head MRI showed a heterogeneously enhancing posterior fossa mass with hydrocephalus. Gross total resection was performed and the tumor was consistent with a classic medulloblastoma. Postoperative chemotherapy and craniospinal radiation was administered. The patient remained tumor-free until 2008 when he presented with right-sided weakness and numbness, left eye pain, vomiting, and weight loss. Imaging showed abnormalities within the posterior pons, medulla, inferior cerebellar peduncles, cerebellar hemispheres and cervicomedullary junction with expansion of the medulla and cervical spinal cord. Due to the location of the lesion, a biopsy was felt to be too risky and was avoided. Despite receiving chemotherapy, his symptoms continued to worsen, and he died four months later. Post-mortem examination limited to the brain and spinal cord confirmed the radiographic extent of tumor. Microscopic examination showed a highly cellular infiltrative glial neoplasm with hyperchromatic, pleomorphic tumor cells including large bizarre and multinucleated cells. Extensive palisading necrosis was also present. The tumor cells were immunoreactive for GFAP but not synaptophysin. A diagnosis of glioblastoma was rendered. It is uncertain whether this second tumor represents a tumor recurrence with unique histology or chemotherapy-induced differentiation along glial lines, or a radiation-induced tumor. Additional immunohistochemical and molecular studies are being performed on both samples to further elucidate the relationship between the two tumors. A handful of similar cases have also been reported, but few have examined the molecular characteristics of the tumors. By studying such cases, we hope to gain a better understanding of the biology of such rare isolated events.

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69 Locus Ceruleus Neurofibrillary Degeneration in Patients With High Braak Stage Pathology But Relative Cognitive Preservation

Maryam Pezhouh. University of Minnesota; Gang Chentree. Dept of Neuroscience; Joyce Meints. Dept. of Laboratory Medicine and Pathology; Laura Hemmy. Dept of Psychiatry; Michael Lee. Dept of Neuroscience; Karen SantaCruz. Dept. of Laboratory Medicine and Pathology

Locus Ceruleus (LC) neurofibrillary degeneration is an early event in Alzheimer's disease (AD). However, the relationships between cortical pathology and brain stem (LC) pathology is unknown. Herein, we examined whether LC pathology reflects the extent of cortical AD pathology or the functional status of individuals. We compared histological and immunohistochemical features of the LC in the Sisters of Notre Dame from the Nun study. The groups were age and education matched sisters that exhibitedhigh Braak stage Alzheimer1s disease pathology but were either classified as demented (AD) or nondemented (NDAP). We used immunohistochemistry for tyrosine hydroxylase as a marker for adrenergic neurons; AT8 for phospho-tau associated neurofibrillary tangles, pretangles and dystrophic neuritis; 4G8 as a marker for amyloid deposits; and IBA-1 as a marker for microglial cells. TH immunostained sections suggest greater preservation of neurons in the NDAP group compared to AD group. Further, the percentage of neurons and neuritic processes in LC containing AT8 was higher in the AD group compared to the NDAP group. There were no significant differences between the LC cells that stained beta amyloid, extracellular amyloid deposits, and IBA-1. These results indicate that the extent of brain stem (LC) pathology can be dissociated from extent of cortical AD pathology and may reflect cognitive preservation in the NDAP group.

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70 Neurogenesis in Alzheimer's Disease Neocortex

Stefanie Marquez, Li Zhou, Celia Williams, Carol Miller. University of Southern California Keck School of Medicine

In the adult human, neurogenesis occurs throughout life in selected areas of the CNS, specifically the hippocampus and olfactory bulb. Acute insults, such as hypoxia/ischemia, may stimulate further activation of neural stem cells (NSCs) and neural progenitor cells (NPCs) of limited potential. Less is known about the response of these cell types to chronic stimuli present during specific neurodegenerative processes. As the toxic species, oligomeric amyloid beta (oAb) has the potential as an oxidant stressor to affect neurogenesis. However, neurogenesis has been found to be altered in the hippocampus of patients with Alzheimer's Disease (AD) and in rodent models of AD. Here we present evidence of neurogenesis in several unexpected areas of the AD brain: frontal cortex and subcortical white matter. Using immunohistochemistry on formalin-fixed tissue, we detected neurogenesis with antinestin, anti-epidermal growth factor receptor (EGFR), antidoublecortin (DCX) and antibeta III tubulin (Tuj1) antibodies. Nestin and DCX-immunoreactive cells were found in the brains of 12 individuals diagnosed with either AD or incipient Alzheimer's Disease (IAD), but to a much lesser extent in age-matched controls. The diagnoses were confirmed neuropathologically using NIA-Reagan criteria. These immunopositive cells often appear as part of a pair, with one or both of the cells expressing DCX. A small fraction of the stem/progenitors differentiated into S-100b positive glial cells, while others displayed markers of immature neurons, illustrated by DCX/Tuj1 double labeling. Cells expressing EGFR, a marker for transit amplifying cells, are also present in the AD PVZ1. Homogenates of frontal cortex snap-frozen at autopsy were examined for oAb content by Western blot. We detected an 8 KDa band appearing in patients with IAD and AD, but not in normal controls. These results suggest ongoing neurogenesis occurring throughout the progression of AD in an affected cortical region. It occurs concurrently with oAb accumulation and cognitive decline.

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71 Biochemical Analysis of ß-Amyloid and Tau in Tangle-Only Dementia

John Crary, Ismael Santa-Maria Perez, Aya Haggiagi, Jean Paul Vonsattel, Michael Shelanski. Columbia University Medical Center, Department of Pathology

Almost 6% of patients diagnosed with late-onset Alzheimer disease (AD) exhibit neuropathological changes remarkable for the absence of ß-amyloid (Aß)-positive neuritic plaques that are associated with AD. Patients with this limbic predominant dementia subtype, termed "tangle-only dementia" (TOD), display neurofibrillary tangles (NFTs) that are morphologically and regionally identical to those found in moderate AD. TOD is controversial as it must be reconciled with the hypothesis that increased levels of toxic Aß cause AD. To determine whether there are differences in soluble Aß in TOD, similar to those implicated in classical "plaque and tangle" AD, we performed enzyme-linked immunosorbent assays (ELISAs) to compare the levels of soluble Aß40 and Aß42 isolated by both diethylamine (DEA) and formic acid (FA) extraction from TOD, AD and age-matched controls. This analysis failed to uncover increased Aß40 or Aß42 levels or alterations in the Aß42/40 ratio in TOD compared to controls. To determine whether differences in tau biochemistry or assembly occur in TOD, Western blot and electron microscopy was performed on purified sarkosyl-insoluble tau from TOD patients. This analysis revealed similar tau isoform composition and paired-helical filament ultrastructure in TOD and AD. These data suggest that TOD may develop through a non-amyloid dependent mechanism, arguing that it is best classified as a primary tauopathy. The intriguing overlap of biochemical, ultrastructural and regional characteristics of NFT in TOD and AD suggest convergent pathophysiologic mechanisms.

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72 Distinctive Patterns of Tau and TDP-43 in a Former Professional Football Player and Marine as Compared to 3 Siblings

Ann McKee. Bedford VAMC/Boston University School of Medicine; Thomas Montine. Univ Washington School of Medicine; Victor Alvarez. Boston Univ School of Medicine; Aimee Schantz, Eileen Steinbart, Thomas Bird. Univ Washington School of Medicine

Four siblings (2F:2M) developed progressive dementia and were followed to autopsy. At death, the siblings ranged in age from 72-84 years, with onset of dementia symptoms at ages 63-76. Unlike the others, one sibling (onset dementia 66, death 72) played 11 years of American football, including 4 years college (All American tackle), 1 year NFL, and 2 years military. He also served 2 years of active duty with the Marine Corps in World War II and was awarded a purple heart. When the microscopic brain sections of the 4 siblings were analyzed blindly, the brain of the former football player and Marine was easily distinguished from the others based on distinctive patterns of tau and TDP-43 immunoreactivities, although all 4 met criteria for NIA-Reagan high likelihood AD (sparse -frequent neuritic plaques, Braak neurofibrillary tangle (NFT) stage VI, and mild-severe amyloid angiopathy). The brain of the former football player and Marine was identified by prominent tau immunoreactive NFT in the superficial layers of the frontal and temporal cortex, patchy glial tangles perivascularly and at the depth of the sulci, and marked tau neurofibrillary involvement of the thalamus, hypothalamus, basal ganglia, brainstem and white matter. TDP-43 immunoreactivity was also distinctive with prominent TDP-43 immunoreactivity in the hypothalamus, thalamus, white matter, neocortex, medial temporal lobe and brainstem, compared to sparse medial temporal lobe TDP-43 immunoreactivity in his siblings. Although all brains fulfilled neuropathological criteria for AD, the brain of the former football player and veteran showed co-existent Chronic Traumatic Encephalopathy (CTE) and AD. These findings suggest that the repetitive mild brain trauma associated with the play of football and/or military combat is associated with distinctive patterns of tau and TDP-43 immunoreactivities that can be distinguished blindly from the brains of genetically similar individuals without athletic or military exposure.

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73 In vivo and In Silico Evidence: Hippocampal Cholesterol Metabolism Decreases With Aging and Increases with Alzheimer Disease

Clyde Phelix. The University of Texas at San Antonio; Sandra Siedlak, Xiongwei Zhu. Case Western Reserve University; George Perry. The University of Texas at San Antonio

Cholesterol metabolism is implicated in the etiology of Alzheimer's Disease (AD). The present study used gene expression profiles on human CA1 in age-matched and severe AD groups to determine enzyme reaction rate constants for metabolic pathways including cholesterol biosynthesis, isoprenoid production, and cholesterol catabolism. Previous studies on adult baboons examined proteins involved in cholesterol and fatty acid synthesis. These studies showed the enzyme targeted by statins was lower in aged baboons, which corresponds to gene expression pattern in human hippocampus. These observations correspond to decreased cholesterol content and synthesis rate of aging human brain. The core metabolic model was used to simulate at 20-39 years of age and compare with aged-controls, mean age of 85.3 years. Flux through this rate limiting step in the simulation for aged was lower by 9.5%. The cholesterol level was 52.3% lower in the simulation and 33.6% lower in the aged brain. This evidence validates the in silico method. Data from the microarray study on AD was used to evaluate sterol regulatory element binding protein 1 and 2 (SREBP1 & SREBP2) showing the levels were increased significantly in the severe AD. Cytochemistry showed robust increase in SREBP2 staining in CA1 of AD versus control. Immunoblot analysis showed a significant increase in SREBP1. Since these are transcription factors driving cholesterol and fatty acid synthesis pathways, we predicted that simulation of severe AD versus age-matched control would show corresponding results. Flux for both cholesterol and fatty acid biosynthesis increased dramatically in severe AD. Also cholesterol level was 92% higher. The sensitivities analyses for incipient and severe AD demonstrated how they differ. Most reactions are insensitive for severe AD and only two sensitive peaks are obvious; cholesterol and ubiquinone levels most sensitive to CYP46a1. No surprise that statins were ineffective in clinical trials for treatment of AD, post-diagnosis.

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74 Embryonic Stem Cell-Derived Choroid Plexus Epithelial Cells: A Potential Therapy for Alzheimer's Disease

Edwin Monuki, Momoko Watanabe, Lauren Davies, Chi-Yeh Chung, Jaymin Kathiriya. UC Irvine

The choroid plexus epithelial cell (CPEC) is a distinctive cell type derived from neuroectoderm of the developing brain. CPECs provide several developmental and homeostatic functions via the production of cerebrospinal fluid (CSF) and formation of the blood-CSF barrier. CPEC dysfunction has also been widely implicated in neurodegenerative disorders, particularly Alzheimer's Disease. However, clinical applications using CPECs have been hindered by the inability to expand or generate these cells in culture. Here we provide molecular, cellular, ultrastructural, and functional evidence for CPEC differentiation from neuroectodermal cells derived from mouse and human embryonic stem cells. These findings expand the repertoire of neural cell derivatives in culture and herald a CPEC-based regenerative medicine for Alzheimer's and other neurodegenerative disorders.

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75 FAM76B, a Novel PGRN Interacting Protein Screened by Yeast Two-Hybrid System

Qinwen Mao. Northwestern University; Lihong Chai, Shunjun Wang, Xiaojing Zheng. Shaanxi Normal University, P.R. China; Eileen Bigio. Northwestern University Feinberg School of Medicine; Haibin Xia. Shaanxi Normal University, P.R. China

Progranulin (PGRN) encodes a 68.5-kDa secreted growth factor that is composed of seven and a half tandem repeats of a 12-cysteine granulin motif. PGRN is expressed in many tissues and has a role in mediating development, wound repair, inflammation, and tumorigenesis. Mutations leading to a loss of function in PGRN are the most common cause of familial frontotemporal lobar degeneration with TDP-43-positive inclusions (FTLD-TDP), which is the second most common form of early-onset dementia after Alzheimer's disease. Correlated with its widespread distribution and diverse functions, PGRN was found to interact with many molecules such as Epstein-Barr Virus RK-BARF0, Perlecan, and most recently identified Sortilin, etc. However, the functions of PGRN in brain and peripheral tissues and the pathways involved are far from clear. In this study, by using the yeast two-hybrid system and for the first time by using human PGRN as bait, we identified a novel PGRN interacting protein FAM76B, which is a nuclear protein of unknown function. We then characterized the precise interaction sites in FAM76B and PGRN with the yeast two-hybrid system, and confirmed their interaction by pull-down assay in vitro and co-immunoprecipitation assays in vivo. Furthermore, to determine whether PGRN colocalizes intracellularly with FAM76B, the fluorescent tagged PGRN, Grns and FAM76B were co-expressed in CHO cells. The results comfirmed that FAM76B localizes to the nucleus, and PGRN and most of the Grns to the cytoplasm. However, in the presence of FAM76B, GrnBACD was found to translocate into the nucleus and colocalize with FAM76B. These data will provide new clues to the function of PGRN in normal and pathological conditions.

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76 Japanese Consortium for Research in Motor Neuron Disease and Frontotemporal Dementia

Shigeo Murayama. Dpt. Neuropath. (BBAR), Tokyo Metro. Geriatr. Hosp. & Inst. Gerontol.; Yuko Saito. Neuropathology, NCNP; Masaki Takao. Neuropathology, Mihara Mermoiral Hospital and TMGHIG; Hiroyuki Hatsuta. Pathology, TMGHIG; Jun Shimizu. Neurology, University of Tokyo; Tameko Kihira. Kansai Medical College; Yasumasa Kokubo. Neurology, Mie University; Haruhiko Akiyama. Neuropathology, Tokyo Institute of Psychiatry (TIP); Kinuko Suzuki. Dpt. Neuropath. (BBAR), Tokyo Metro. Geriatr. Hosp. & Inst. Gerontol.; Masato Hasegawa. Molecular Genetics, TIP

Common etiology has been postulated between motor neuron disease (MND) and frontotemporal dementia (FTD), after discovery of TDP43 and FUS. The purpose of this study is to establish consortium for MND/FTD resource and neuropathological studies, in collaboration of neurological, psychiatric and gerontological insitutes. The Brain Bank for Aging Research (BBAR) Project was employed and combined longitudinal clinical follow ups and detailed neuropathological studies. All the brains and spinal cords were examined with antibodies raised against ubiquitin and TDP 43, and ubiquitin- positive TDP- 43 negative cases were screened with anti- FUS antibodies. Half brains and total length of spinal cords, after sampling specific cervical, thoracic, lumbar segments, a lumbar dorsal root ganglion and pieces of cauda equina for histological studies, were frozen for biochemical and genetic studies. Brains and spinal cords from 33 MND/ FTD and 228 control cases were stored for biobanking, consisting of 27 ALS- TDP 43 cases, one case each with FUS R521C and SOD1 G93S mutation and two cases each with Pick disease and FTLD- TDP43. Neuropathological studies were conducted on familial ALS with SOD1 (3), TDP43 (1) and FUS (3) mutation and 111 sporadic MND cases with a case of juvenile ALS with FUS- immunoreactive basophilic inclusions. Eighty one FTD cases were classified into FTLD- tau (33), TDP43 (36), FUS (9) and without specific features (3). Four cases registered to BBAR were classified into primary TDP 43 proteinopathy without clinical FTD. Spinal neuronal intracytoplasmic TDP43 proteinopathy was only detected in primary or combined (6 cases with ALS/ parkinson dementia complex- Kii) subtypes. Immunoblot with anti- TDP43 antibody confirmed same typing both in the brain and spinal cord from ALS- TDP43 cases. With this MND/ FTD consortium, we can establish research background to pursuit common pathogenesis of these disorders on the basis of neuropathological findings.

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77 Neuroanatomical Distribution of Tau Pathology in a Case of Frontotemporal Dementia Associated With the MAPT P301L Mutation

Bernardino Ghetti, Jill Murrell, Jose Bonnin, Matthew Hagen. Indiana University; Alberto Espay. University of Cincinnati; Michael Keys. Lindner Center of Hope; Clarissa Rentz. Alzheimer's Association

A 52-year-old woman began experiencing dizziness and fatigue followed by personality changes including aggressivity and paranoid thinking. MRI revealed atrophy of the frontal lobes. At age 56, she exhibited a marked decrease in verbal output and at age 58, she could not attend to any of her personal needs. She died at age 62. Her sister is affected with a dementing illness associated with aphasia and behavioral changes. Their mother died with a dementing illness lasting approximately 12 years. A molecular genetic analysis revealed a single nucleotide (C → T) substitution at codon 301 in one allele of the patient's MAPT gene resulting in a leucine for proline substitution (P301L). In addition, she had an H1/H1 MAPT haplotype. At autopsy, the brain weighed 937 grams. The frontal and temporal lobes were severely atrophic. The parietal lobes were atrophic with the left being more severely affected. The occipital lobe was mildly atrophic. There was mild atrophy of the midbrain with a depigmentation of the substania nigra. Tau immunohistochemistry revealed severe involvement of the frontal, cingulate, insular, and temporal cortices. In the temporal lobe, severe tau deposition is noted in the middle and inferior temporal gyri, parahippocampal gyrus and hippocampus. The superior temporal gyrus is remarkably spared. In the hippocampus, the dentate fascia is severely affected as well as the CA1 and CA41. The parietal lobes were relatively spared. Within the cortex, the second, third, fifth and sixth layers were most affected. Tau deposits were within neuronal perikarya and proximal dendrites. Numerous astrocytic plaques and occasional tufted astrocytes were seen. Among the subcortical nuclei, the amygdala was most severely affected while the globus pallidus and the claustrum were moderately involved. This case is of particular interest for the sharp anatomical demarcation and severity of neuronal tau pathology. Supported by NIA P30AG010133.

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78 Diffuse TDP-43 Pathology in a Case of Hereditary Spastic Paraparesis With a NIPA1/SPG6 Mutation

Maria Martinez-Lage, Laura Molina-Porcel, Dana Falcone. Dept of Pathology and Laboratory Medicine, Leo McCluskey. Dept of Neurology, Vivianna Van Deerlin, Virginia Lee, John Trojanowski. Dept of Pathology and Laboratory Medicine, University of Pennsylvania

Hereditary spastic paraparesis (HSP) and amyotrophic lateral sclerosis (ALS) predominantly affect the pyramidal system and can involve other non-motor systems. Both present clinical and genetic similarities that suggest a possible mechanistic commonality. We present a case of HSP with a mutation in NIPA1/SPG6 and postmortem examination revealing a diffuse TDP-43 proteinopathy.A 53-year-old woman with progressive severe spasticity and weakness since age 13 developed progressive upper extremities and bilateral facial weakness, increasing bladder and bowel dysfunction, hoarseness, dysphagia, dyspnea and orthopnea. She demonstrated cognitive decline with poor attention, memory loss and personality change. She died of respiratory failure at age 56. Laboratory evaluation was negative. Nerve conduction/needle studies demonstrated a sensorimotor polyneuropathy, mild chronic denervation proximally and marked chronic partial denervation distally. Family history revealed a father with personality changes, aggressiveness, and language deficits starting at age 61, who developed gait problems and a mild tremor, and died at age 71. DNA sequencing for NIPA1 (SPG6) in the proband demonstrated a nucleotide substitution (c.G341A) resulting in an amino acid change (p.G106R), a well documented HSP-causing mutation. Postmortem examination revealed a diffusely atrophic spinal cord. Microscopic exam showed marked loss of motor neurons with gliosis, and Bunina bodies. Kluver-Barrera stains revealed degeneration of lateral corticospinal tracts and dorsal columns. Axonal loss was concomitant with myelin loss. TDP-43 immunostaining revealed typical skein-like and round cytoplasmic inclusions in motor neurons, substantia nigra, dentate gyrus, entorhinal cortex, amygdala, basal ganglia, and neocortical areas including motor cortex. To our knowledge, this is the first neuropathological description of a patient with a NIPA1 mutation. Frequent TDP-43 inclusions, motor neuron loss and long tract degeneration were identified, and although we cannot rule out the co-existence of two different pathologies, these findings support a possible common pathway for motor neuron degeneration in HSP and ALS.

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79 Blastomycosis Presenting as a Pontine Mass Lesion

Cheryl Palmer, Paula Province, Hassan Fathallah-Shaykh, Barton Guthrie, James Hackney. University of Alabama at Birmingham

A 51-year-old man presented with a three-month history of dizziness, disordered gait, headaches, and episodes of distal right-sided numbness. CT scan demonstrated a pontine mass with extension into the middle cerebellar peduncle. Follow-up MRI of the brain demonstrated a prominently enhancing lesion in the left lateral pons with increased T2 signal in the surrounding area. CT scans of the chest revealed multiple small pulmonary nodules with hilar and mediastinal adenopathy. The presumptive diagnosis was high-grade pontine glioma and the patient was treated with dexamethasone, producing immediate symptomatic improvement. When seen in clinic two weeks later, imaging studies revealed improvement in the size of the mass and the degree of edema. Mediastinoscopy with lymph node biopsy demonstrated noncaseating granulomas with special stains negative for organisms. In clinic two weeks later, the patient had diplopia and worsened coordination. The pontine lesion had enlarged on MRI scan. Consideration was given to initiating radiation and chemotherapy without biopsy due to the high probability of inducing further injury with the procedure. However, MR spectroscopy was normal lateral to the enhancing component of the mass, an unusual finding for an infiltrating glioma. For this reason, it was decided to proceed with stereotaxy. Neuropathologic evaluation demonstrated large fungal organisms with doubly birefringent walls and occasional broad-based budding suggestive of Blastomycosis. Fungal culture was positive for Blastomyces dermatitidis. Blastomycosis predominantly affects men, and is not associated with immunosuppression. CNS involvement is rare, affecting about 2.5% of patients with pulmonary or systemic disease. Usually, the fungus produces meningitis, but occasionally focal abscess develops with localizing symptomatology. Overall mortality in treated, non-immunocompromised patients is 10% or less. CNS Blastomycotic abscesses can mimic malignancy on MRI scan. Careful investigation of all clinical features, including appropriate biopsy even in eloquent brain regions, should lead to the avoidance of potentially inappropriate empirical therapies.

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80 Aspergillus granuloma - A Rare Pseudoneoplasm of the CNS

Maria Martinez-Lage, Sriram Venneti. Dept of Pathology and Laboratory Medicine, Melandee Brown, James Schuster. Dept of Neurosurgery, Zissimos Mourelatos. Dept of Pathology and Laboratory Medicine, University of Pennsylvania

Mass lesions of the central nervous system of non-neoplastic origin frequently pose a diagnostic challenge for clinicians and neuropathologists. A 30-year-old male of asian origin who presented with headaches and right arm dysmetria was found to have a large rim-enhancing right cerebellar hemispheric mass with mild surrounding edema. A primary neoplasm was suspected. Upon surgical resection, the mass appeared very firm and relatively well demarcated from the surrounding tissue. Microscopic examination revealed a parenchymal and leptomeningeal inflammatory process with extensive granulomatous inflammation and abundant multinucleated giant cells. Tissue cultures were positive for Aspergillus flavus, and a Grocott stain revealed numerous septated hyphae with acute angle branching consistent with Aspergillus species. A diagnosis of Aspergillus granuloma was rendered.Fungal infections of the central nervous system are rare, and most frequently found in immunocompromised patients, with cryptococcal meningitis being the most common example. Identification of the causative organism is essential for appropriate treatment. Aspergillus species are ubiquitous filamentous fungi known to cause human disease in a variety of settings. In the central nervous system, aspergillosis can present as a fulminant angioinvasive disease in the immunocompromised host, as a less aggressive form invading the central nervous system directly from the sinonasal cavities, or as an indolent mass-forming lesion known as aspergillus granuloma in the host that is able to amount a competent immune response. A definite diagnosis requires brain tissue for histopathological analysis and cultures. Aspergillus granulomas comprise a very rare pseudoneoplastic entity, most commonly caused by Aspergillus flavus and identified in hot and dry climates where this species thrives. They are characterized by good prognosis when treated with surgical excision and antifungal therapy.We present a well-documented case of aspergillus granuloma as an example of a rare pseudoneoplasm of the central nervous system with specific epidemiological and therapeutical implications.

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81 Quantitation of JC Virus in a Case of Progressive Multifocal Leukoencephalopathy (PML) Caused by Selective Immune Deficiency

Keith Wharton. Biogen Idec; Catherine Quigley, Marian Themeles. Comparative Pathology Laboratory, Jing Wei, Alex Buko. Bioanalytical Chemistry, Carl Reid, Chao Sun, John Carulli. Molecular Discovery, Susan Goelz. Neurology, Biogen Idec; Susan Staugaitis. Departments of Neurosciences and Anatomic Pathology, Robert Fox. Mellen Center for Multiple Sclerosis, Cleveland Clinic

Over half of humanity is seropositive for the human JC polyomavirus (JCV), but only a tiny fraction of individuals develop PML, a destructive JCV infection of CNS white matter. PML is receiving increased attention due to its occurrence in a rare subset of patients on a variety of selective immunomodulatory agents, including natalizumab (Tysabri®, Biogen Idec/Elan) used as a therapy for multiple sclerosis (MS) and Crohn's disease. Fundamental aspects of JCV biology and PML pathology remain mysterious, including the cell type(s) infected, the mechanism of viral spread throughout the CNS, and the abundance and distribution of virus throughout affected brain. To investigate PML without the potential confounding demyelination of MS, we analyzed a case of histologically-confirmed PML that was acquired by a 70-year-old man treated for psoriasis with efalizumab (Raptiva®, Genentech) for four years. Examination of postmortem brain revealed gross and microscopic pathology characteristic of PML, and virions were identified by electron microscopy. We further sampled the center, edge, and adjacent grossly normal-appearing white matter of three different lesions. For each sample, we correlated immunohistochemical findings with the abundance of selected viral and non-viral proteins measured by multiple reaction monitoring (MRM) mass spectrometry, as well as viral DNA copy number determined by qPCR. We observe a positive correlation between the abundance of viral constituents and cytopathic effect but not the extent of demyelination. These studies serve as a foundation to better understand JCV biology and PML pathology, with the goal of improving risk management in patients on selective immunomodulatory therapies.

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82 Chronic Granulomatous Herpes Encephalitis In A 10-Year-Old Boy With Clinically Intractable Epilepsy

Keith Harrison. The Children's Hospital of Alabama

This 10-year-old boy was admitted to our institution for medically intractable epilepsy as part of a presurgical evaluation for epilepsy surgery. His medical history is significant for herpes meningitis at age 4 months. At that time he presented to an outside hospital with fever for 3 days, with acyclovir treatment beginning on day 4 of his 40 day hospital course. He subsequently developed infantile spasms and ultimately a mixed seizure disorder. Recent video electroencephalogram shows a Lennox-Gastaut-type pattern with some focal features of frequent runs of right frontotemporal spikes. His captured clinical seizure semiology shows 2 types of seizures; dialeptic seizures and left arm fencing. Clinically his seizure types also include frequent drop attacks with multiple episodes of status epilepticus. His MRI scan shows an abnormality in the inferior right frontal lobe operculum and anterior insula. Based on these findings, the patient underwent a right frontal craniotomy for partial frontal lobectomy. Histologic sections of the right frontal lesion demonstrate chronic granulomatous inflammation with foci of necrosis and mineralization. Scattered clusters of lymphocytes, microglial nodules and small, discrete non-necrotizing granulomas are present with scattered multinucleate giant cells. Some giant cells have mineralized material within the cytoplasm. These changes are set within gliotic cerebral cortex and superficial white matter. Immunohistochemical staining for herpes simplex virus (HSV I/II) shows focal nuclear and cytoplasmic immunoreactivity. Special staining for fungi and immunohistochemical staining for cytomegalovirus are also performed and both are negative. After undergoing intravenous acyclovir therapy, the patient has returned to baseline with decreased seizure frequency. It is unclear if this histology results from reactivated latent virus or represents residual damage from the initial insult. This rare form of herpes encephalitis has only been reported in children; however, the initial presentation of herpes meningitis and the approximate 10-year time interval are unusual in this case.

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83 Progressive Multifocal Leukoencephalopathy Secondary to Natalizumab With Cortical Involvement and Negative CSF Evaluation

Andrea Wiens, Jose Bonnin. Dept. of Pathology; Scott Shapiro. Dept. of Neurosurgery; Monica Mazda. Dept. of Neurology; Riley Snook. Indiana University School of Medicine, Dept. of Neurology

Progressive multifocal leukoencephalopathy (PML) is a demyelinating disease caused by the JC polyomavirus (JCV). It is most common in immunocompromised patients such as those harboring HIV-1 infection, patients with hematological diseases, or in patients receiving immunosuppressive therapy. The development of PML has also been linked to natalizumab (Tysabri), the first monoclonal antibody treatment for multiple sclerosis which was introduced in 2004. The condition usually appears after 24 months of treatment. In such patients, the diagnosis of PML is most often established by magnetic resonance imaging (MRI) scans demonstrating new lesions not previously identified as multiple sclerosis plaques, together with cerebrospinal fluid (CSF) evaluation for JCV by polymerase chain reaction (PCR). We report a case of PML in a 71-year-old woman who presented with speech difficulties and mild dysarthria. She had been diagnosed with multiple sclerosis 51 years earlier and had been given natalizumab for 23 months prior to the onset of symptoms. Four lumbar punctures in the previous 12 months were all negative for JCV. Worsening of symptoms to the point of inability to ambulate and severe aphasia led to a brain biopsy. This was taken from a new enhancing lesion in the left frontal lobe involving the cortex and subcortical white matter. Diagnosis of PML was readily confirmed by morphology and immunohistochemistry. The viral inclusions were numerous in cortical oligodendrocytes, and there were also changes consistent with immune reconstitution inflammatory syndrome (IRIS). As a significant portion of patients with PML may have a negative PCR for JCV in the CSF, a brain biopsy should be considered without delay in patients who have suspicious neuroimaging findings for the disease.

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84 Bacterial Abscess With Diffuse Sterile Leptomeningeal Calcific "Sugar-Coating'

Nguyen Vo, James Traylor. LSUHSC Shreveport Department of Pathology; Wilson Jon. Louisiana State University Health Sciences Center Shreveport

We present the case of a 51 year-old Caucasian male with unknown past medical history found dead in his house. Postmortem examination revealed bilateral cerebellar purulent abscesses and fine white-tan sandpaper-like granules diffusely over the leptomeninges. The latter was noted following formalin fixation, but in retrospect was noted to have been barely visible in pre-fixation images. Warthin-Starry stain demonstrated numerous filamentous bacteria morphologically compatible with actinomyces or nocardia species within the cerebellar abscesses, but was negative in the nodular granular calcific deposits. The presence of "sugar coating" following fixation in this case in unusual, and on gross inspection raised the differential of necrotic cerebellar metastasis with leptomeningeal dissemination versus rupture of squamous-lined cyst with leptomeningeal seeding. The precise mechanism of leptomeningeal calcific "sugar-coating" in this case is not known with certainty.

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85 Xanthogranulomatous Hypophysitis Occurring in Association With Pituitary Adenoma: A Report of Two Cases

Cara Sedney, Fahad Bafakih, Charles Rosen, Kymberly Gyure. West Virginia University

Xanthogranulomatous hypophysitis (XH) is a rare sellar region lesion which may occur as a primary process mimicking a pituitary adenoma or as a secondary lesion that is most commonly associated with craniopharyngioma or Rathke cleft cyst. We report two cases of XH with coexisting pituitary adenoma. The first patient, a 54-year-old woman, presented with an intractable headache. Endocrine work-up showed hypothyroidism with a normal thyroid-stimulating hormone level and a mildly elevated prolactin level, and no visual field deficits were noted. Magnetic resonance imaging (MRI) revealed a sellar mass which was followed with serial imaging for two years until the lesion enlarged and was subsequently resected. The second patient, also a 54-year-old woman, was being followed for an incidentally discovered cystic pituitary mass when she developed blurred vision. MRI revealed an increase in the size of this mass, which was then resected. Pathologic evaluation of both transsphenoidal resection specimens revealed a loss of normal adenohypophyseal architecture with weakly follicle-stimulating hormone-positive adenoma cells which failed to stain with adrenocorticotropic hormone, growth hormone, luteinizing hormone, prolactin, or thyroid-stimulating hormone. These adenomas were associated with chronic inflammation including xanthoma cells and foreign body-type giant cells, cholesterol clefts, and hemosiderin deposition. In the first patient, the XH was adherent to the diaphragma sellae and was intraoperatively felt to be distinct from the adenoma. In both cases, XH most likely represents a reaction to hemorrhage or necrosis within the adenomas. Although unusual, recognition that XH and pituitary adenoma can occur together is important to avoid misdiagnosis.

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86 Fulminant Demyelinating Process Presenting as Wernicke Encephalopathy in a Non-Alcoholic Patient

Murat Gokden. University of Arkansas for Medical Sciences; Dittana Phoncharoensri. Northwest Mississippi Regional Medical Center, Clarksdale, MS

Wernicke encephalopathy (WE) is a syndrome characterized by altered consciousness, ocular dysfunction and gait disturbances, and results from thiamine deficiency. It is typically associated with chronic alcoholism; however, it can also be seen in many other conditions resulting in thiamine deficiency. Most commonly affected regions in the brain are mamillary bodies, hypothalamus, thalamus and periaqueductal gray matter. Here, we present a case of WE resulting from a demyelinating process.51-year-old black woman with history of hypertension presented with progressive weakness for a week. Right eye deviation, left internuclear ophthalmoplegia and severe hyponatremia were identified. MRI showed hyperintensity in left periaqueductal area and the floor of the 4th. ventricle with no enhancement. Cerebral salt wasting syndrome was treated and she was discharged after one week. She was readmitted 23 days later for lethargy and decreased responsiveness of one week duration and was reported to be of poor nutritional status. MRI showed symmetric hyperintense lesions in mamillary bodies and periaqueductal gray matter. Tests for infectious and paraneoplastic etiologies were negative. Thiamine and methylprednisolone treatments were started for WE and possible demyelinating disease; however, her condition gradually became worse, resulting in death in two weeks. Autopsy examination of the brain showed an active demyelinating process symmetrically involving the hypothalamus, periaqueductal gray matter, pontine tegmentum and medulla oblongata. Congestion, microhemorrhages, necrosis or significant gliosis were not seen. No other lesions, demyelinating or otherwise, were identified in other parts of the brain, including central basis pontis, internal capsule, corpus callosum, periventricular white matter, among others. This case illustrates a WE of unclear, but likely of nutritional, nonalcoholic etiology. The presence of demyelinating process, without the typical pathologic features of WE, in a distribution involved in WE, raises the possibility of an acute fulminant demyelinating process presenting as WE.

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87 Intimal Thickening of Meningeal Arteries After Serial Corticectomies for Rasmussen's Encephalitis

Aaron Wagner, Kristina Biado, Madeline Schwarz, Alexander Bottini, Gary Mathern, Harry Vinters. UCLA

Rasmussen encephalitis (RE) is a rare cause of intractable epilepsy in children. Between 2008 and 2010, 4 patients had additional cortical resections performed after a primary corticectomy for RE. In each of these cases we observed some degree of atherosclerotic change in leptomeningeal arteries, consisting of moderate to moderately severe intimal hyperplasia. The intervals between original resection and revision ranged from 8 months to 10 years. Ages of the patients ranged from 9 - 12 years at their first resection and ranged from 10 - 19 years at the time of revision. Four cases in years 2006 - 2010, with surgical resections diagnosed as Rasmussen's encephalitis (age range 5 - 21 years), as well as 2 surgical revisions for severe cortical dysplasia, one for mild cortical dysplasia and one for recurrent dysembryoplastic neuroepithelial tumor (DNET), did not show significant vascular abnormalities (with surgical intervals of 10 months - 16 years).

Conclusion: Leptomeningeal atherosclerosis appears to develop in the interval between cortical resections for RE, an inflammatory disorder. The pathogenesis of this vascular change may be related to meningeal inflammation in RE. This complication in children undergoing surgical resection for RE may itself lead to "secondary" ischemic change that contributes to worsening of epilepsy.

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88 Radiologic-Pathologic-Proteomics Correlation. High Field 7T MRI and Multiplex Tissue Immunoblotting of Autopsy Brains

Kant Matsuda. Division of Neuropathology, Dept of Pathology, New York University; Joon-Yong Chung. Applied Molecular Pathology Laboratory, Lab of Pathology, NCI/NIH; Kris Ylaya. Tissue Array Research Program, Lab of Pathology, NCI/NIH; Masaki Fukunaga. Laboratory of Functional and Molecular Imaging, NINDS/NIH; Tie-Qiang Li. Laboratory of Functional and Molecular Imaging. NINDS/NIH; Elisabeth Rushing. Institute for Neuropathology, UniversitätsSpital Zürich; Stephen Hewitt. Tissue Array Research Program, Lab of Pathology, NCI/NIH

In the era of "high-tech" medicine, there is an ever increasing variety of non-invasive, radiological imaging modalities. Key to the development of these emerging technologies is the validation of imaging with histopathologic correlation. Frequently, discrepancies are documented with respect to size or extent of abnormalities seen in in vivo radiographic findings when compared to the surgical pathology specimens. Given the challenges of point-to-point correlation of in vivo imaging with histopathology, we investigated the feasibility of using ex vivo tissue imaging to bridge the gap between in vivo imaging and histopathology. Recent advances in high field MRI have achieved resolution at the microscopic level. For example, high field systems such as 7Tesla (7T) are available, which offer 150 micron in-plane resolution in clinical settings. With high resolution, findings obtained by ex vivo imaging are readily correlated with histopathology after the identical plane is obtained. We further explored the feasibility of imaging correlation, with the proteomics profile using a novel technology known as"multiplex tissue immunoblotting" (MTIB). This technique is capable of demonstrating the expression pattern of multiple proteins from a single histologic tissue section and can be reliably correlated with the histopathologic findings. Thus, it is feasible to correlate ex vivo imaging with the protein expression pattern using MTIB. To achieve precise correlation, the identical tissue plane is required for accurate ex vivo imaging of the corresponding histopathology site. Accordingly, we scanned autopsy brains with 7T MRI, which were subsequently processed for whole mount preparation. MTIB was then performed and protein expression profile was correlated with MRI at the identical histogeographic location.

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89 Pituitary Blastoma: An Entity

Bernd Scheithauer. Mayo Clinic; Eva Horvath. St. Michael's Hospital, Toronto, Canada; Cheri Deal. University of Montreal / CHU Sainte-Justine, Montreal, Canada; Yves Robitaille. University of Montreal / CHU Sainte-Justine, Montreal, Canada; Kalman Kovacs. St. Michael's Hospital, Toronto, Canada

Pituitary blastoma is a recently described tumor of the neonatal pituitary exhibiting differentiation to Rathke epithelium and embryonic adenohypophysial cells, both folliculostellate and secretory (Scheithauer, et al, Acta Neuropathol 116: 657, 2008). Exceedingly rare, its cellular composition reflects pituitary development. Herein, we report a second case involving a 9-month-old male presenting with progressive right ophthalmoplegia. An MRI scan showed a large sellar/suprasellar mass with cavernous sinus invasion. Endocrine testing demonstrated secondary hypothyroidism as well as elevated plasma ACTH levels and nonsuppressible hypercortisolemia. Diabetes insipidus was not evident. A subtotal resection was achieved. Histologically, the complex tumor consisted of small undifferentiated cells, rosettes and glandular structures resembling Rathke epithelium, as well as large secretory cells. A scant spindle cell stroma was focally noted. Cell proliferation was evident (mitoses, maximal 3/10 HPF; MIB-1 LI, maximal 50%). The immunophenotype of the numerous larger secretory cells included synaptophysin, chromogranin, various keratins and, to a lesser extent, ACTH, beta-endorphin, and MGMT. Germ cell markers were negative. Ultrastructurally, two cell populations were seen, including small, primitive-appearing, polyhedral epithelial cells with abundant glycogen but with scant cytoplasm and organelles (folliculostellate cells) and a second population of larger corticotrophic cells containing rough endoplasmic reticulum, Golgi membranes, spherical,150-400nm secretory granules and occasional bundles of intermediate filaments (corticotrophic cells). The morphologic features were those of the previously described pituitary blastoma, albeit featuring a greater number of folliculostellate cells and more limited secretory cell differentiation. Our finding of the cellular elements of adenohypophysial development confirms the diagnosis of pituitary blastoma, aligns the lesion with blastomas of other organs, and suggests that specific genetic abnormalities may underlie its genesis.

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90 Collicular Hemorrhages; Metronidazole Toxicity or Wernicke's Encephalopathy?

Chia-Ling Phuah. Brigham and Women's Hospital; Hart Lidov. Children's Hospital Boston

A 58 y.o. woman with metastatic uterine leiomyosarcoma was admitted for failure to thrive and weight loss. She was treated with chemotherapy, IGFR-1 antibody and mTOR inhibitor. On a prior admission for a seizures she had no acute CNS imaging abnormality. She was unable to continue chemotherapy due to inanition. On re-admission no neurologic findings were reported. She had worsening nausea and diarrhea. Stool was positive for C. difficile; IV metronidazole and TPN were started. She became progressively weaker, sleepy, confused and frequently disoriented. She complained of vertigo and exhibited an unsteady gait. No brain imaging was done; she became more somnolent and succumbed after 3 weeks. At autopsy the brain was externally unremarkable and had no metastatic neoplasm. The remarkable gross finding was bilaterally symmetric hemorrhagic lesions of the inferior colliculli. These raised the question of Wernicke's encephalopathy or mitochondrial disorders in the Leigh's spectrum and through microscopic sampling of pertinent structures was undertaken. The midbrain lesions show rarefaction of the neuropil, capillary proliferation, relative sparing of neurons, and at the periphery of these focal lesions, petechial hemorrhages. Smaller petechial hemorrhages which did not appear to be post mortem, where found in the hypothalamus, and bilaterally symmetric vacuolation of the vestibular nuclei and pons. The microscopic lesions are interpreted as precursors to the grossly visible lesions of the colliculi. Lesions were not found in the mammillary bodies or thalamus. Lesions histologically remarkably similar occur in mitochondrial cytopathy and in Wernicke encephalopathy, the latter characteristically involving the mammillary bodies and anterior thalamus. There have been reports in the MRI literature of reversible signal abnormalities in a pattern similar to both of these disorders as an unusual response to treatment with metronidazole, as in this case. Nevertheless the possibility of nutritional deficiency or nutritionally induced increased susceptibility to metronidazole toxicity cannot be discounted.

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91 Pathologic Findings In Sudden Unexpected Death In Epilepsy (SUDEP): A Ten Year Retrospective Review

Janet McNaughton. Orlando Health; Jan Garavaglia. District 9 Medical Examiner's Office; Gary Pearl. Orlando Health

While various risk factors of sudden unexpected death in epilepsy (SUDEP) are well documented in the literature, few studies have focused on the neuropathologic findings associated with this condition. In this study, we performed a retrospective ten-year review of cases from the District 9 Medical Examiner's Office in Orlando, Florida, in an attempt to identify histologic abnormalities in the brain of decedents whose cause of death was documented as SUDEP. We reviewed 103 cases from 2001 through 2010 in which the cause of death was listed as SUDEP, out of a total of 10445 post mortem examinations performed. Of those cases, 75 were excluded due lack of postmortem evaluation or absence of gross pathology in the brain at autopsy. Twenty-eight cases remained in which neuropathology was identified by gross and histologic examination in decedents ranging from 2 to 69 years in age (mean: 41 years; median: 44.5 years). Decedent clinical history was reviewed to identify surgery or trauma related to seizure onset. Eighty two percent of decedents were on antiepileptic medications, although the majority (78%) was found to be subtherapeutic by postmortem blood drug levels. We found 43% of SUDEP cases to be surgical or trauma related, with one surgical case being represented as loss of tissue in left temporal region with associated gliosis. Eleven cases of trauma-related seizure disorder were associated with contusions of the brain and encephalomalacia. Additional findings included hippocampal sclerosis and congenital abnormalities in 25% of cases, cerebral infarction in 18% of cases and infectious causes and vascular malformations in 7% each. While incidence of SUDEP is increased in individuals who are at subtherapeutic drug levels, most cases of SUDEP were not associated with specific histological findings in the brain. The majority of histologic findings were associated with a history of prior surgery or trauma.

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92 Going Green in the Neuropathology Laboratory- A Review of Long-Term Room Temperature Biospecimen Storage

Leili Mirsadraei, Jerry Lou, Sergey Mareninov, William Yong. Brain Tumor Translational Resource, Dept of Pathology

Preserving biospecimens and derivatives in the laboratory has traditionally required freezing at ultralow temperatures. The risk of freezer failure as well as growing space and cost constraints argue for the need to evaluate room temperature storage modalities. In this study, our aim is to review some of the major technologies available and the potential issues with longterm (decades) room temperature storage. Most commercially available technologies commit a biospecimen (tissue, blood, nucleic acids) to preserving DNA only or RNA only. Typically, biospecimens are placed in a small tube or in a well within a microtiter plate. These plates can potentially reduce occupied floor space by up to 90%, condensing the equivalent of an entire freezer of samples into a medium-sized box. However, as most neuropathology laboratories store predominantly tissue rather than nucleic acids, this extent of benefit is theoretical. DNA storage using modified Whatman FTA-based methodologies are well established with long term room-temperature storage data extending decades- as much as18 years in real time. Newer commercial technologies involve proprietary compounds. With stabilization, tissue and blood have yielded DNA after greater than 1 year. RNA has been stored for approximately 3 years at room temperature. Accelerated aging by storing biospecimens at elevated temperatures have been utilized in an attempt to predict long-term stability with claims in the order of decades. Lyophilization has been investigated for room temperature preservation of tissue and enables extraction of DNA, RNA, and protein after one year or more. Mixed data has been reported as to the quality of RNA preservation. In summary, room temperature modalities, with the exception of lyophilization, are limited by the commitment of a specimen to a specific nucleic acid type. Most technologies, with the exception of Whatman FTA paper, have limited real-time long-term stability data though accelerated aging data is promising.

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93 What Constitutes Adequate Histologic Validation for Tumor Presence in a Frozen Glioblastoma Biospecimen?

Desiree Sanchez, Jason De Jesus, Andrew Kay, Bob Shafa, Negar Khanlou, Harry Vinters, William Yong. UCLA Pathology Department

Molecular testing of brain tumor tissue after it has been frozen necessitates histological analysis in order to determine the extent and purity of tumor tissue. The objective of this study was to determine whether the extent of tumor and the tumor cellularity significantly changes while cutting through a frozen glioblastoma fragment to a depth of approximately 0.92 mm. Fragments averaging 106 mg (SD = 0.05 mg; range: 58-914mg) were obtained from 91 frozen glioblastoma specimens. Using a cryostat, four 5-μm sections were cut- each 300 μm apart and H&E stained. The percentage of tumor by area and the degree of cellularity were scored by a neuropathologist. The difference between the highest and lowest tumor percentage within each specimen was calculated. The mean difference in tumor percentage was low (Median = 0, Mean = 4.67, SD = 8.92). Cellularity was classified on a scale of 0-5 where 0 = no tumor present and 5 = high cellularity tumor. Cellularity changes across each consecutive section were categorized as follows: 0 = no change, 1 = change one level, 2 = change two levels. Cellularity for 96.7% of the specimens did not change after sectioning to a depth of 0.92 mm (Median = 0, Mean = 0.55, SD = 0.31). Furthermore, presence of tumor in the first H&E section correctly predicts the presence of tumor in subsequent deeper sections of all cases (n=82). Similarly, absence of tumor in the first histologic section correctly predicts absence of tumor to the full depth assessed (n=9). These findings suggest that obtaining one H&E section of a small brain tumor specimen (∼100 mg) is sufficient to demonstrate the adequacy of tumor when sectioned to a depth of 0.92 mm.

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94 Ideal Warm Ischemia Times for Minimizing RNA Expression and Protein Phosphorylation Changes- A Literature Review

Bijan Ameri, William Yong. Brain Tumor Translational Resource, Dept of Pathology

The National Cancer Institute's Office of Biorepositories and Biospecimen Research (OBBR) has recommended the collection of pre-analytical variables. One important variable is warm ischemia time. We reviewed studies cataloged in the OBBR Biospecimen Research Database and supplemented these with papers identified in PubMed. Studies have shown activation of hypoxia-sensitive genes shortly following ischemia. For example, prostate specimens stored at room temperature upregulate numerous genes in a time-dependent manner. Colon specimen studies have indicated that ischemia also affects the expression of oncogenes and other growth-related genes. However, warm ischemia affects tissues in varying degrees. Some tissues, such as lung, are not as affected by moderate ischemia times. Articles reporting these findings recommend time-frames to decrease gene expression changes: 60 minutes for lung and prostate tissue and 20 minutes for colon tissue. The emergence of RNALater has aided in RNA preservation for gene expression profiling. However, protein-signaling profiles are also an issue and may vary unrelated to RNA transcript stability. Analysis of myometrial phosphoproteins suggests that the maximum time from excision to stabilization should be 20 minutes. Beyond that time phosphorylation profiles change substantially. Other studies of breast, colon, lung, ovary, uterus and melanoma tissues suggest that significant changes in protein-signaling pathways occur in as little as 30 minutes after excision. Furthermore different tissues and tumors in the same organ site may differ. Limited brain tissue studies have shown prevention of protein degradation by storing tissues at 4°C for up to 8 hours. Unfortunately, data on procurement-related ischemia effects for brain tissues is scant. What then should a brain or brain tumor repository aspire to in tissue procurement? Until further data is acquired, zero minutes clearly would be ideal but a maximum of 20 minutes of warm ischemia time may be a practical target in a surgical setting.

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95 Lyophilization as a Brain Tumor Preservation Method for Long-Term Storage at Ambient Temperature

Sergey Mareninov. Brain Tumor Translational Resource, Dept of Pathology; Andrew Kay, Desiree Sanchez, Leili Mirsadraei, Jerry Lou, Bob Shafa, Isaac Yang. UCLA Neurosurgery; Tracie Pham, Aaron Wagner, Harry Vinters, William Yong. UCLA Pathology Department

Long-term and cost effective storage of pathological specimens is of great importance for clinical pathology and tissue banking. Currently, snap-freezing tissue in liquid nitrogen and formalin fixation are generally accepted for clinical tissue preservation. In order to investigate alternatives to conventional tissue preservation methods, we tested freeze-drying (lyophilization) for three types of human brain tumors: glioblastoma, low grade glioma, and meningioma. Comparative molecular and morphological analysis was performed on freeze-dried, snap-frozen and FFPE tissue samples, 24 in total, after 1 year of storage to evaluate the ability of freeze-drying to preserve tissues with sufficient quality for clinical applications. Our data showed that freeze-dried tissue after one year of storage at ambient temperature yielded high quality, high molecular weight genomic DNA acceptable for PCR analysis. RNA extracted from freeze-dried tissue demonstrated suitability for RT-PCR though RIN values showed some degradation. Proteins from freeze-dried specimens fully retained antigenic reactivity and showed strong IHC staining for clinically relevant markers: Ki-67, GFAP and EMA. Moreover, proteins demonstrated no degradation based on Coomassie blue stained Western blots as well as specific blot analyses for GFAP and GAPDH. Lactate dehydrogenase and pyruvate kinase enzymatic activity showed no difference between freeze-dried and snap-frozen tissue. In conclusion, the data demonstrates the practical value of using freeze-drying as a preservation method for clinical samples intended for various types of molecular and morphological analysis. We suggest that freeze-dried tissue specimens are potentially more versatile than other room temeperature storage modalities as well as formalin-fixed materials. In addition, the potential cost, space, and environmental savings by avoiding frozen storage are attractive.

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96 ß-Amyloid Precursor Protein Immunoreactivity in Infant Brains: A Detailed Look at Pattern of Injury

Jocelyn Posthumus, Carrie Mohila. University of Virginia Health System; Virginia Richards, Deborah Kay. The Office of the Chief Medical Examiner Richmond Virginia; Maria Lopes. University of Virginia Health System

The use of ß-amyloid precursor protein (ß-APP) immunohistochemistry in the evaluation of diffuse axonal injury is complicated by similar histomorphologic changes occurring in the setting of trauma and hypoxia-ischemia. The literature suggests that a distinction can be made based on the presence of a "geographic" or "irregular" pattern of ß-APP immunoreactivity, with the former suggestive of vascular and the latter traumatic injury. An additional diagnostic challenge is that traumatic injury is often complicated by a hypoxic insult. In an attempt to expand upon the definitions of vascular and traumatic injury we retrospectively investigated ß-APP reactivity in 52 infant brains including both medical and forensic autopsies. We reviewed autopsy reports for cause of injury and evidence of hypoxia-ischemia to assign cases to a nonaccidental central nervous system injury (NAI), hypoxic-ischemic injury (HI), or undetermined group. The cohort consisted of 4 NAI without ventilator support, 12 NAI with ventilator support, 14 natural/accidental deaths without head trauma, and 22 sudden unexpected deaths in infancy (SUDI). Sections from the corpus callosum, brainstem, upper cervical cord, and hippocampus were blindly reviewed by two neuropathologists for neuronal hypoxic-ischemia and/or axonal damage. Interobserver agreement for the presence of neuronal hypoxia was low at 58.3% based on hematoxylin and eosin-stained sections; however, improved to 90.6% based on interpretation of neuronal perikarya ß-APP reactivity. Axonal spheroids were identified on ß-APP in all 16 NAI cases (100%), 8 HI cases (57.1%), and 13 SUDI cases (59.1%). Geographic and irregular patterns of ß-APP were seen in both NAI and HI cases. We conclude that the majority of infant brains show nonspecific axonal ß-APP reactivity, precluding a definitive diagnosis of traumatic injury. The observed high interobserver agreement in neuronal ß-APP reactivity, suggests a possible role in documenting neuronal hypoxic-ischemic changes.

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PLATFORM SESSION 5: MUSCLE AND NERVE Saturday 8-10 a.m. June 25, 2011

97 Expanding Histopathological and Clinical Spectrum of RYR1 Associated Myopathies

Mariarita Santi. Children's Hospital of Philadelphia; Kristen Perkins. Neuromuscular and Neurogenetic Disorders of Childhood Section, Nationa; Amanda Kan. Department of Pathology, Children's Hospital of Philadelphia; Ying Hu. Neuromuscular and Neurogenetic Disorders of Childhood Section, Nationa; Livija Medne. Department of Genetics, Children's Hospital o; Diana Bharrucha. Department of Neurology and Genetics, Children's Hospital of Perry Shieh. Department of Neurology, UCLA David Geffen School of Medicine, Los Ang; Carsten Bönnemann. Department of Neurology Children's Hospital of Philadelphia, Philadelphia

Central core disease (CCD) is a rare nonprogressive myopathy characterized by hypotonia, proximal muscle weakness and presenting in infancy. The disease is caused by mutation in the gene for the skeletal muscle ryanodine receptor (RYR1). The majority of CCD mutations in the RYR1 gene are dominantly acting missense mutations, although small deletions have been detected. Recently the phenotypic spectrum of CCD has broadened to include severe cases with neonatal lethality or precluding independent ambulation. The classical morphology is characterized by the "central core" due to lack of oxidative enzyme activity in the central region of the fibers. However the spectrum has broadened in the past years. Genetically confirmed mutation of the RYR1 gene may present with centrally placed nuclei (CNM), multiple focal areas of myofibrillary disruption (MMC) and in some cases with the coexistence of cores and rods. Other cases fulfill criteria for congenital fiber type disproportion (CFTD), in other cases there is prominent fiber atrophy with fibrosis and adipose tissue infiltration resembling a congenital muscular dystrophy (CMD). We reviewed 36 patients with genetically proven RYR1 mutations with both dominant as well as recessive mutations. The age of patients ranges from 24 days to 48 years. Clinical and morphological presentation includes familial typically mild CCD, severe neonatal CCD, mild congenital myopathy with MMD or CFTD, congenital myopathy with external ophthalmoplegia and CNM, and severe infantile presentation with CMD-like histological features. Presentations with CCD is in general caused by inherited or de novo dominant mutations, whereas the latter three presentations tend to be caused by recessively acting RYR1 mutations. Immunohistochemical analysis to assess the precise distribution of mitochondria, RYR1 and SR associated proteins in the various histological phenotypes will be presented in the 13 cases for which sufficient frozen tissue was available.

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98 Double Null Mice Lacking Dysferlin and Myoferlin Show a Muscular Dystrophy Phenotype

Peter Pytel. Dept. of Pathology; Alexis Demonbreun, Kieran Devaux. Dept. of Medicine; Manuel Alvarez. Dept. of Pathology; Elizabeth McNally. University of Chicago - Dept. of Medicine

INTRODUCTION: Loss of dysferlin function has been linked to LGMD2B in humans and to a muscular dystrophy phenotype in mouse models. This phenotype is thought to be the result of a defect in membrane fusion events essential for repairing small defects in the sarcolemma of mature skeletal muscle fibers. The dysferlin homologue myoferlin has been shown to be important for membrane fusion events during skeletal muscle development.

METHODS: The dysferlin null allele found in the A/J mice was bread into the 129SVJ background over 6 generations to generate dysferlin null 129SVJ mice. These mice were then also bread with 129SVJ mice carrying a myoferlin null allele. The phenotype of 129SVJ wild-type mice, dysferlin null mice (dysf), myoferlin null mice (MKO) and double null mice (fer) was analyzed.

RESULTS: All four genotypes are viable and have been aged to over 14months. Internalized nuclei are found in 3.6%, 11.4%, 21.6% and 23.4% of quadriceps myofibers in 6-months WT, MKO, dysf and fer animals respectively. The mean cross-sectional size of quadriceps myofibers is 96%, 91% and 90% of WT in 6-month MKO, dysf and fer animals respectively. Overall comparison of muscle morphology at different ages shows that dysf and the fer animals display progressive muscle damage with myofiber necrosis, internalized nuclei and at older ages chronic remodeling. These changes are most prominent in proximal muscles of the hind limb, paraspinal muscles and abdominal muscles.

CONCLUSIONS: (1) Double null animals lacking dysferlin as well as myoferlin exhibit progressive muscle injury. (2) Double null fer animals tend to have more severe disease than dysferlin null animals. This finding would support a role of myoferlin in the repair of adult skeletal muscle.

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99 Amyloid Myopathy Associated With Dysferlinopathy: Comparison With Light Chain Amyloidosis

Syed Kazmi, Rodney McComb. Dept. of Pathology and Microbiology, Univ. of Nebraska Med Ctr Omaha; Gokden Murat. Dept. of Pathology, Univ. of Arkansas for Med. Sci, Little Rock, AR; Simone Spuler. Experimental and Clinical Research Ctr at the Charité, Berlin, Germany; J. Americo Fernandes Filho. Dept. of Neurological Sciences, Univ. of Nebraska Med Ctr. Omaha, NE; Kent Huston. The Center for Rheumatic Disease, Kanasa City, MO; Betul Gundogdu. Dept. of Neurology, Univ. of Arkansas for Med Sci. Little Rock AR; Steven Moore. Dept. of Pathology, Univ. of Iowa Hospitals and Clinics, Iowa City, IA

Amyloid myopathy is a rare disorder that may be histologically subtle. Biochemical forms include immunoglobulin light chain (AL type), amyloid associated with infectious/inflammatory disease (AA type), and familial amyloid. A recent addition is amyloid associated with dysferlinopathy, an autosomal recessive myopathy caused by mutations in DYSF that typically manifests as Miyoshi myopathy or limb-girdle muscular dystrophy. Here we describe three patients with amyloid myopathy. The first two patients presented with chronic proximal weakness and the third with distal weakness. All of them had myopathic electromyography and high creatinine kinase (2500-7500 IU/L). Amyloid in all biopsies was confirmed by Congo red. A deltoid biopsy from the first patient showed mild perivascular and endomysial lymphocytic inflammation. Amyloid was present in blood vessel walls and in the endomysium cuffing myofibers, and was immunoreactive for lambda light chain. Subsequent evaluation showed high serum lambda light chain (13.9 mg/dL; reference range: 0.5-2.6) and a slight increase in bone marrow plasma cells (5%). The quadriceps biopsy from the second patient showed severe myopathic changes with perivascular amyloid deposition. Further workup for plasma cell dyscrasia and light chain disease was negative. A repeat muscle biopsy for amyloid typing by liquid chromatography tandem mass spectrometry failed to detect peptides associated with AA, ATTR or AL amyloid. Dysferlin was absent by immunofluorescence and western blot. DYSF sequencing is in progress. The gastrocnemius biopsy from the third patient showed end-stage muscle with perivascular and interstitial deposits of amyloid. Immunostaining for dysferlin (Hamlet antibody) was negative in the myofibers and amyloid deposits. DYSF sequencing revealed two heterozygous mutations. Although dysferlinopathy is a rare cause of amyloid deposition in muscle, it should be considered along with other forms of amyloid. Mononuclear endomysial or perivascular inflammation may be seen in amyloid myopathy, potentially drawing attention away from the amyloid.

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100 Modeling the Human C205T Mutation in Murine Mtm1 Results in Exon 4 Skipping and a Less Severe Myotubular Myopathy Phenotype

Christopher Pierson, Ashley Dulin-Smith, Ashley Durban, Jordan Marshall, Andrew Snyder, Jordan Gladman, Dawn Chandler. The Research Institute at Nationwide Children's Hospital, Columbus, OH; Anna Buj-Bello. Généthon, Evry, France; Michael Lawlor. Children's Hospital Boston, Boston, MA; James Dowling. University of Michigan, Ann Arbor, MI.; Alan Beggs. Children's Hospital Boston, Boston, MA

MTM1 mutations cause myotubularin deficiency and underlie the pathogenesis of myotubular myopathy (MTM). The short life span (about 8 weeks) of the Mtm1 knockout mouse limits the scope of preclinical trials that can be performed, so we sought to generate an MTM model with a milder phenotype. Based on human genotype-phenotype correlation studies, we chose to model the C205T point mutation in exon 4, which is predicted to introduce the R69C missense change in the catalytically inactive PH-GRAM domain of myotubularin. Mtm1C205T mice exhibit early muscle atrophy, but appear clinically unaffected until mild weakness develops at 2 months of age. At 2 months, Mtm1C205T mice generate 70% of the grip strength force of wild type mice and the weakness remains stable over 12 months. The median survival period is 60 weeks. Histopathology shows many small myofibers with central nuclei and necklace fibers. Myotubularin protein is not detectable in muscle, which prompted a study of mRNA structure. C205T induced exon 4 skipping in mouse and in patient muscle. Bioinformatic analysis suggested that C205T perturbs an exonic splicing silencer in Mtm1 mRNA. Four other human exon 4 missense mutations (T202G, T205A, G206C, and C208T), modeled in vitro using a minigene system, also showed exon 4 skipping; suggesting that this region may indeed, contain a regulatory sequence. Exon 4 skipping alters the reading frame of Mtm1 mRNA and creates a premature stop codon, limiting the amount of myotubularin that is translated. Myotubularin dephosphorylates the 3'-phosphate group from PI3P and PIP2 phosphoinositides, so myotubularin phosphatase activity was tested by determining PI3P levels using an ELISA method in Mtm1C205T and Mtm1 knockout muscles. Both models showed PI3P levels that were elevated to similar degrees compared to age-matched controls, suggesting that PI3P levels alone cannot account for the difference in phenotype between these MTM models.

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101 Distribution and Extent of Weakness Is Dependent on Fiber Type and Mutation in Murine Models of Myotubularin Deficiency

Michael Lawlor, Marissa Viola. Children's Hospital Boston/Harvard Medical School, Boston, MA; Jeffrey Widrick. Spaulding Rehabilitation Hospital/Harvard Medical School, Boston, MA; Robert Grange. Virginia Polytechnic Institute and State University, Blacksburg, VA; Anna Buj-Bello. Genethon, Evry, France; Christopher Pierson. Nationwide Children's Hospital/Ohio State University, Columbus, OH; Alan Beggs. Children's Hospital Boston/Harvard Medical School, Boston, MA

X-linked myotubular myopathy (XLMTM) is a congenital myopathy caused by deficiency of the lipid phosphatase, myotubularin. XLMTM patients often present with severe perinatal weakness that requires mechanical ventilation to prevent death from respiratory failure. Muscle biopsies from XLMTM patients display small myofibers with characteristic central nuclei and central aggregations of organelles in many cells. To better understand the role of myotubularin in specific fiber types, we have investigated whether the weakness seen in severely- (Mtm1d4) and moderately (Mtm1C205T) symptomatic myotubularin-deficient mice is dependent on fiber type. Preliminary studies suggest that the fiber type composition of muscles in Mtm1C205T mice differs dramatically from that seen in Mtm1d4 mice, with a greater number of oxidative fibers seen in Mtm1C205T mice. Contractile studies of untreated Mtm1d4 and Mtm1C205T mice also revealed marked differences in the observed physiological deficits in these mice. While ex vivo field stimulation of soleus and extensor digitorum longus (EDL) muscles of Mtm1d4 mice showed marked decreases in isometric force in comparison to WT counterparts, Mtm1C205T mice showed marked decreases in isometric force in the EDL and higher than normal strength in the soleus. Ultrastructural examination of these muscles in several pilot animals revealed a larger number of normal-appearing triad structures in Mtm1C205T muscles compared to Mtm1d4 muscles. Maximum Ca2 activated force of skinned fibers from Mtm1d4 mice was 25% lower than normal, suggesting that the majority of the force deficit observed in the whole muscle was due to processes occurring before cross-bridge activation. These findings suggest that the effects of myotubularin deficiency are fiber type specific, and that the different mutations in these two models may determine disease severity primarily through their differential effects on oxidative fibers.

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102 Immunohistochemistry for LC3 as a Diagnostic Marker for Drug-Induced Autophagic Myopathy

Han Lee. University of California San Francisco; Brianne Daniels. Touro University California; Eduardo Salas, Andrew Bollen, Jayanta Debnath, Marta Margeta. University of California San Francisco

Chloroquine, its analog hydroxychloroquine, and colchicine are commonly used for the treatment of malaria, rheumatologic diseases, and gout, respectively. In a subset of patients, these treatments lead to development of vacuolar myopathy, the pathologic diagnosis of which currently requires identification of autophagic vacuoles by electron microscopy. Microtubule-associated protein light chain 3 (LC3), a protein required for autophagy, has emerged as a robust marker of autophagosomes. In this study, we evaluated the utility of LC3 immunohistochemistry in establishing the diagnosis of drug-induced autophagic myopathy. We reviewed UCSF muscle biopsy case records from 1997 to 2010 and retrieved autophagic myopathy cases (myopathy group), cases with no pathologic findings (normal control group), and cases that received treatment with either colchicine or hydroxychloroquine but did not show diagnostic myopathic features (drug-treated control group). Electron microscopic findings for drug-treated control and autophagic myopathy groups were reviewed by 2 blinded neuropathologists (MM, AB) to confirm accurate classification; cases with equivocal diagnoses or lack of consensus were excluded, with 10 normal controls, 7 drug-treated controls, and 9 autophagic myopathy cases used for the final analysis. Immunohistochemistry for LC3 and p62/SQSTM (a protein selectively degraded via autophagy) was performed on formalin-fixed, paraffin-embedded tissue. In all drug-treated cases, but not in normal controls, LC3 and p62 showed a characteristic pattern of punctate linear staining, often in the vicinity of vacuoles. However, the fraction of LC3-positive fibers was significantly higher in autophagic myopathy group (median 0.57, SD 0.31) compared to either the normal control group (median 0, SD 0; p<0.001) or the drug-treated control group (median 0.02, SD 0.05; p<0.05; Kruskal-Wallis one-way ANOVA). Thus, LC3 immunohistochemistry is a diagnostically useful autophagy marker that can replace electron microscopy in the diagnosis of most cases of drug-induced autophagic myopathy.

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103 Patterns of TDP-43 Immunoreactivity in Selected Inflammatory Myopathies

Jeremy Deisch, Dennis Burns. University of Texas Southwestern Medical Center

Dermatomyositis is an inflammatory myopathy characterized by preferential involvement of the perifascicular compartment, manifesting pathologically as predominantly perimysial inflammation accompanied by perifascicular atrophy and myopathic changes. Ultrastructurally, dermatomyositis is associated with characteristic tubuloreticular inclusions, detected most often in endothelial cells. Recently, abnormal sarcoplasmic accumulation of the nucleoprotein TAR DNA-binding protein (TDP-43) has been described in protein-aggregation myopathies, namely sporadic and hereditary inclusion body myositis (IBM) and myofibrillar myopathies associated with desmin and myotilin mutations. In the central nervous system, cytoplasmic TDP-43 accumulation in neurons of the brain and spinal cord is the pathologic substrate of frontotemporal lobar degeneration with ubiquitinated inclusions (FTLD-TDP, formerly FTLD-U) and a subset of amyotrophic lateral sclerosis (ALS) cases, respectively. To investigate the pattern of TDP-43 immunoreactivity in inflammatory myopathies other than IBM, we utilized anti-TDP-43 antibodies (Proteintech Group, Chicago, IL) on paraffin-embedded, formalin-fixed muscle tissue. The study comprised 20 muscle biopsies with dermatomyositis, as well as 2 with polymyositis, 2 with sporadic IBM, and 2 normal muscle controls. The dermatomyositis muscles were from 5 male and 15 female patients ranging in age from 3 to 70 years (mean 28.9). Fifteen (75%) cases of dermatomyositis showed nuclear TDP-43 staining preferentially involving myofibers at the periphery of the fascicle. In both IBM specimens, pathologic sarcoplasmic TDP-43 reactivity was accompanied by scattered reactive nuclei. Diffuse nonspecific nuclear staining was noted in one of two cases of polymyositis; the second case was nonreactive. TDP-43 immunoreactivity was absent in the normal muscle biopsies. Statistically, perifascicular TDP-43 reactivity in dermatomyositis was associated with the presence of perifascicular myofiber atrophy, significant myopathic alterations, and the presence of inflammation. To the best of our knowledge, this is the first report of perifascicular TDP-43 immunoreactivity in dermatomyositis. This finding is intriguing, as the precise etiology of perifascicular atrophy in this disorder remains elusive.

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104 Clinical and Myopathological Manifestations of Filaminopathy in a Newly Identified Pedigree

Juan Bilbao. Sunnybrook Hospital, University of Toronto; Lev Goldfarb. National Institute of Health; Beverley Young, Kester Kong, Sandra Cohen. University of Toronto

Myofibrillar myopathies (MFMs) consist of heterogeneous disorders having in common disruption of myofibrils and disintegration of Z-disks, and sarcoplasmic accumulation of surplus proteins. MFM is known to be associated with mutations in DES, CRYAB, MYOT, ZASP and FLNC. We present a Canadian patient of Chinese ancestry who at age 61 years, against a background of late onset cataracts and peroneal neuropathy, developed proximal muscle weakness in all limbs with slightly elevated CPK. Family history was relevant for muscle weakness in late mother and muscle weakness and heart disease in late older brother. A muscle biopsy revealed pleomorphic sarcoplasmic masses with immunolocalization of multiple proteins, autophagic vacuoles, congophilic sarcoplasmic inclusions, no intranuclear inclusions and, by electron microscopy, spheroid bodies, granulofilamentous material, and aggregates of intermediate filaments with a perihelical configuration associated with autophagic vacuoles. Sequencing of DES, CRYAB, MYOT, and ZASP revealed no mutations. A W2710X mutation in the filamin C (FLNC) gene was identified. This mutation is similar to the Filamin C gene mutation first identified in members of an extended German family in 2005. With the identification of the first family in the western hemisphere the results of our study expands the geographical distribution of filaminopathy, and corroborates that the MFM caused by mutation of FLNC shares the histological, immunohistological and ultrastructural profile of the MFM's caused by mutations in DES, CRYAB and MYOT.To establish the population frequency of this type of MFM, molecular genetic analysis for Filamin C should be recommended for all muscle biopsy specimens showing multiple protein aggregates in pleomorphic sarcoplasmic regions, autophagic vacuoles, Congophilia in hyaline deposits and ultrastructural evidence of sarcomeric disintegration, and accumulation of degraded filamentous material.

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PLATFORM SESSION 6: TUMORS - NONGLIAL Saturday 8-10 a.m. June 25, 2011

105 Molecular Subgroups In Medulloblastoma

David Ellison. St. Jude Children's Research Hospital; James Dalton. St. Jude Children's Research Hospital; Dan Brat. Emory University School of Medicine; Arie Perry. UCSF; William Yong. UCLA; Mehmet Kocak. St. Jude Children's Research Hospital

Subgroups of medulloblastoma are separated by histopathologic features, molecular genetic abnormalities, or gene expression profiles, and some subgroups are associated with distinct outcomes. We examined associations between molecular subgroup and clinical, histopathologic and cytogenetic variables among 235 medulloblastomas from patients aged 0.4-52 years. FFPE tumors were available for analysis using immunohistochemistry and iFISH with probes to MYC, MYCN, PTCH1 and loci on chromosomes 6 and 17. WNT, SHH, and non-SHH/WNT molecular subgroups were defined by four immunohistochemical markers: ß-catenin, GAB1, filamin A, and YAP1. Immunoreactivity for GAB1 characterized only SHH tumors and nuclear IR for ß-catenin only WNT tumors. IRs for filamin A and YAP1 identified SHH and WNT tumors. SHH, WNT, and non-SHH/WNT tumors contributed 31%, 14%, and 55% to the series. All desmoplastic / nodular (D/N) medulloblastomas were SHH tumors, while most WNT tumors (94%) had a classic phenotype. Monosomy 6 was strongly associated with WNT tumors, while PTCH1 loss occurred almost exclusively among SHH tumors. MYC or MYCN amplification and chromosome 17 imbalance occurred predominantly among non-SHH/WNT tumors. Among patients aged 3-16 years on the SIOP PNET3 trial, outcome was significantly better for children with WNT tumors, when compared to SHH or non-SHH/WNT tumors, which showed similar survival curves. However, high-risk factors (M disease, LC/A pathology, MYC amplification) significantly influenced survival in both SHH and non-SHH/WNT groups. In corroborating other studies that indicate the value of combining clinical, pathological, and molecular variables in therapeutic stratification schemes for medulloblastoma, we also provide a novel method for detecting WNT, SHH, and non-SHH/WNT subgroups using immunohistochemistry, the first outcome data based on a clinical trial cohort, and novel data on how molecular subgroups are distributed across the range of disease.

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106 Prospective Comparison of FISH Versus PCR-Based Microsatellite LOH in the Evaluation of Gliomas for 1p/19q Codeletion

Craig Horbinski. University of Kentucky, Department of Pathology; Marina Nikiforova. University of Pittsburgh, Department of Pathology; Jonathan Hobbs. University of Kentucky; Kathy Cieply, Ronald Hamilton. University of Pittsburgh, Department of Pathology

1p/19q codeletion has been a robust marker of oligodendrogliomas for years, and has repeatedly been shown to be a favorable prognostic factor, correlating with improved response to adjuvant therapy. The most common tool for evaluation of 1p/19q status is fluorescence in situ hybridization (FISH), although other techniques for interrogating larger regions of 1p and 19q are sometimes used, including microsatellite-based polymerase chain reaction (PCR) for loss of heterozygosity (LOH). Herein we describe patient outcomes from a prospective cohort of 675 gliomas (including 109 grade II and III oligodendrogliomas) in which both FISH and LOH were done to evaluate 1p/19q status at the time of original diagnosis. 93% of all gliomas showed FISH-LOH concordance; in discrepant cases, classic astrocytic morphology tended to be a better predictor of survival than either test alone. Neither test was able to significantly stratify grade II oligodendrogliomas by survival, but in grade III oligodendrogliomas, LOH correlated with survival better than FISH (P < 0.0001 for LOH versus P = 0.05 for FISH). Thus, although most cases show concordance between the two modalities, LOH appears to be a slightly more specific test for true 1p/19q codeletion. The implications of this, including technical details (e.g. choice of FISH probes) and the future of 1p/19q analysis (e.g. array-based technology), are discussed.

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107 Dissecting the Role of PTEN in Astrocytoma Invasion Using Genetically-Engineered Mouse Models

Byron Huff, Ryan Bash, Natalie Karpinich, Ralf Schmid, C Miller. University Of North Carolina

Astrocytomas are characterized by diffuse invasion, precluding their complete surgical resection. PTEN, a negative PI3 kinase (PI3K) pathway regulator, is altered in 40-80% of high-grade astrocytomas (HGA), including glioblastoma (GBM). However, its role in astrocytoma invasion remains unclear. PI3K pathway signaling, proliferation, migration, and invasion of primary astrocytes from six genetically-engineered mouse models with conditional alleles that inactivate RB (T) and/or PTEN (P) and/or constitutively activate KRAS (R, KRASG12D) upon Cre recombination were analyzed in vitro by immunoblot, cell counting, wound healing and time-lapse video microscopy, and collagen invasion, respectively. Tumorigenicity and survival were determined in vivo in orthotopic allograft models. Invasion was assessed by morphometric analysis. PTEN deletion increased levels of phospho-AKT and phospho-S6. In cells with abnormal RB and KRAS (TR), loss of both PTEN alleles shortened doubling time (DT) 78% (P<0.01) and increased single cell migration velocity and wound healing closure 12 and 66%, respectively (P<0.0001), but had no significant effect on collagen invasion (P=0.89). Both proliferation and migration effects were rescued by transient transfection with wild-type PTEN. In cells lacking both RB and PTEN (TP), activated KRAS shortened DT 53-67% (P<0.01), increased velocity 36-76% and wound healing 91-180% (P<0.0001), and increased invasion 2.4-57 fold (P=0.25). RNAi-based screening of select PI3K pathway effectors identified Rac1 as significantly affecting migration. In vivo, 10E5 TRP-/- cells produced GBM in 5/5 animals at 32 d, while TR cells produced HGA in 2/6 animals at 207 d (P<0.0001). TRP-/- tumors were 2.2 fold more invasive than TR tumors (P=0.066). PTEN deletion significantly affects proliferation and migration of astrocytoma cells, both in vitro and in vivo. Its effects may be mediated via Rac1. In future studies, the orthotopic allograft model system and morphometric analyses described herein will be useful for further dissection of the genetics of astrocytoma invasion in vivo.

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108 Glioneuronal Tumor With Neuropil-Like Islands (GTNI) is Characterized by IDH1 R132H Mutations

Jason Huse, Khedoudja Nafa, Marc Ladanyi, Cyrus Hedvat, Marc Rosenblum. Memorial Sloan-Kettering Cancer Center

Glioneuronal tumor with neuropil-like islands (GTNI) is a recently discovered primary brain neoplasm exhibiting synaptophysin-rich micronodules admixed with diffusely infiltrating astroglial elements. While the precise lineage of GTNI remains unclear, recent work has demonstrated aberrant p53 expression by immunohistochemistry and a notable absence of 1p 19q chromosomal co-deletion, both findings suggestive of a fundamental biological relationship with diffuse astrocytoma. Point mutations in isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) have recently been associated with diffuse gliomas of either astrocytic or oligodendroglial lineage along with secondary glioblastoma, and appear to define a specific mode of oncogenic evolution in primary brain tumors. To establish whether IDH mutations occur in GTNI, we analyzed 9 histologically confirmed cases by Sequenom-based sequencing for all known mutations in IDH1 and IDH2. We also employed a recently optimized immunostain for the IDH1 R132H mutation. We found that, without exception, every case of GTNI in our sample set harbored the IDH R132H mutation. Our data provides further molecular evidence supporting a pathogenic link between GTNI and diffusely infiltrating gliomas, specifically diffuse astrocytoma.

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109 Identification of Driver Mutations Promoting Tumorigenesis in a Transgenic Mouse Model of Neurofibroma-MPNST Progression

Steven Carroll. University of Alabama at Birmingham; Syed Kazmi. University of Nebraska; Stephanie Byer, Nicole Brossier, William Grizzle, Fady Mikhail. University of Alabama at Birmingham

Although comprehensive analyses of somatic mutations in human neoplasms have provided important new insights into the mechanisms mediating tumorigenesis, accumulating sufficient numbers of rare tumor types such as malignant peripheral nerve sheath tumors (MPNSTs) for such studies is challenging. The availability of animal models of neurofibroma-MPNST progression that accurately recapitulate essential somatic mutations promoting tumorigenesis could provide an alternative means of identifying key driver mutations. We have previously shown that transgenic mice expressing the Schwann cell mitogen neuregulin-1 (NRG-1) in Schwann cells (P0-GGFß3 mice) develop MPNSTs. We examined these mice to determine whether their MPNSTs arise from pre-existing neurofibromas and found that they developed neurofibromas in virtually every spinal nerve root, cranial nerves and the sympathetic nervous system; a subset of these animals developed MPNSTs, with small MPNST-like foci identified within some neurofibromas. Examination of the core p14ARF-MDM2-p53 and p16INK4A-cyclin D/CDK4-Rb pathways in early passage MPNST cultures demonstrated multiple mutated molecules in both pathways, although the specific proteins affected varied widely between different tumors. These mutations did not render the tumor cells growth factor-independent as both erbB inhibitors and RNAi-mediated ablation of specific erbB receptors inhibited their proliferation. High density array comparative genomic hybridization (aCGH) was performed to identify unbalanced regions of chromosomal gain and loss in the early passage MPNST cultures. We found that these neoplasms commonly and reproducibly demonstrated loss or amplification of multiple chromosomal regions; some of these included genes previously implicated in MPNST pathogenesis, while others were novel. We conclude that P0-GGFß3 mice represent a bona fide model of neurofibroma-MPNST progression in which tumorigenesis is driven by somatic mutations analogous to those seen in their human counterparts. Given this similarity, neurofibromas and MPNSTs arising in P0-GGFß3 mice can potentially be used to globally identify somatic mutations promoting the pathogenesis of human neurofibromas and MPNSTs.

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110 Dye-Enhanced Multimodal Confocal Imaging as a Novel Approach to Intraoperative Diagnosis of Brain Tumors

Matija Snuderl. Department of Pathology, Sameer Sheth. Department of Neurosurgery, Massachusetts General Hospital; Dennis Wirth. Department of Physics and Applied Physics,University of Massachusetts; Polina Ogas, Churl-Su Kwon, William Curry. Department of Neurosurgery, Massachusetts General Hospital; Matthew Frosch. Department of Pathology, Massachusetts General Hospital; Anna Yaroslavsky. Wellman Center for Photomedicine, Massachusetts General Hospital

Background and Significance: Intraoperative diagnosis plays an important role in accurate sampling of brain tumors, limiting the number of biopsies required and maximizing the extent of resection. Improved detection may enhance both quality of life and overall survival. The goal of this study was to evaluate dye-enhanced multimodal confocal imaging for discriminating gliomas from non glial brain tumors and from normal brain tissue as a potential approach to intravital diagnostic use.

Materials and Methods: Fresh specimens were obtained from craniotomies performed at Massachusetts General Hospital following approval by the IRB. We investigated total of 30 samples including glioblastomas(9), anaplastic astrocytomas(2), low grade gliomas(1), meningiomas(3), ependymoma(1), pilocytic astrocytoma(1), metastatic tumors(6) and normal brain removed for nontumoral indication(7). Tissue was stained in 0.2 mg/ml aqueous solution of methylene blue (MB) for 45-60 seconds. Excess dye was rinsed off and multimodal confocal images were acquired using a specially built microscope, equipped with a 40X Olympus LUCPlan FL N objective (0.70 NA), capable of lateral resolution of 0.6 μm and axial resolution of 7 μm. Reflectance and fluorescence signals of MB were excited at 658 nm. Fluorescence emission and polarization were collected from 670 nm to 710 nm. After imaging, tissue was formalin-fixed and paraffin-embedded for standard neuropathologic evaluation. Four pathologists (2 with and 2 without subspecialty training in neuropathology) were then to provide diagnoses based on the multimodal confocal images.

Results and Conclusions: The investigated tumor types exhibited distinctive characteristics in both the reflectance and fluorescence responses. Images showed distinct morphological features similar to standard histology. Pathologists were able to distinguish gliomas from the normal brain tissue and non glial brain tumors and render diagnoses from the images in a manner comparable to H&E slides. These results confirm the feasibility of multimodal confocal imaging for intravital intraoperative diagnosis.

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111 Malignant Rhabdoid Tumors Express Stem Cell Factors, Which Relate With the Expression of EZH2 and Id Proteins

Sriram Venneti, Paul Le, Daniel Martinez. Dept of Pathology, CHOP, Sharon Xie. Dept of Biostatistics and Epidemiology, Lisa Sullivan, Lucy Rorke-Adams, Bruce Pawel, Alexander Judkins. Department of Pathology, Children's Hospital of LA

Malignant rhabdoid tumors (MRT) are highly aggressive pediatric tumors associated with loss of expression of SMARCB1, occurring in the central nervous system (referred to as atypical teratoid/rhabdoid tumors (AT/RT)) and in the kidney and soft tissues. Histologically, MRT are characterized by immunohistochemical evidence of primitive neuroectodermal, mesenchymal and epithelial differentiation. The ability of MRT to differentiate along multiple lines as evidenced both by histologic features and polyphenotypic immunohistochemical staining, along with the proliferative nature of MRT cells, are characteristics shared with the self-renewal and plasticity of embryonic stem cells (ES). To test the hypothesis that MRT share similarities with ES we used immunohistochemistry to evaluate the expression of various stem cell markers in a tissue microarray (TMA) containing 42 MRT (26 AT/RT) and 16 non-central nervous system MRT (NCMRT)). Expression ranging from strong to weak was noted with Sall4 (24/42, 57%), Glypican-3 (28/42, 67%), TCL-1 (34/42, 81%), and UTF-1 (28/42, 67%). Weak expression was noted with Sox2 (9/42, 21%), Nanog (11/42, 26%), Klf4 (14/42, 33%), Zfp206 (16/42, 38%) and musashi-1 (29/42, 69%). Markers that regulate the expression of stem cell transcription factors were also expressed in MRT. Strong to weak expression was noted with EZH2 (26/42, 62%), Id1 (30/42, 71%), Id2 (38/42, 90%), Id3 (36/42, 86%) and Id4 (33/42, 79%). Regression analysis revealed a significant relationship between the expression of stem cell markers and EZH2 (p=0.0066), Id1 (p=0.0299), Id2 (p=0.0118), Id3 (p=0.013) and Id4 (p=0.0002). These data suggest that MRT express many stem cell associated transcription factors, which may be regulated by the expression of the EZH2 and the Id family of proteins. This study underscores similarities between MRT and stem cells and may help elucidate common biologic pathways that could serve in advancing more effective therapeutic strategies to treat MRT.

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112 Integrative Pathogenomic Analysis of Primary Human Glioblastoma Cell Line Library Reveals Novel Subclass Distinctions

Shakti Ramkissoon. Brigham and Women's Hospital; Karl Olausson, Matthews Theisen, Justin Craig, Malika Hayashi, Sam Haidar, Cecile Maire, Keith Ligon. DFCI-Center for Molecular Oncologic Pathology

Glioblastomas (GBMs) are a heterogeneous group of neoplasms for which several morphologic, genetic and expression subclasses have been previously described. Analysis of patient tumor samples have distinct limitations due to contributions from non-tumor cell types such as normal brain, vascular elements, and inflammation. Here we report the establishment of a library including 28 primary human GBM cell lines grown and propagated under non-adherent stem/progenitor conditions to eliminate non-glioma contributions. Additionally, xenografts were established for each line by stereotactic intracranial injection. Comparative histologic and immunophenotypic characterization of surgical specimens (primary), xenografts and tumor spheres revealed preservation of major W.H.O. histologic subclasses and morphologic correlates (e.g. giant cell glioblastoma). Cell lines and xenografts overall retained excellent correlation of nuclear glial markers (OLIG2, SOX2) and Ki-67 proliferation indices. We next performed whole genome RNA expression profiling analysis of 28 in vitro GBM cell lines (Affymetrix U133 2.0 Plus) followed by unsupervised NMF consensus clustering (GenePattern), which revealed three stable subgroups designated GBM Class A, B, and C. Comparative extraction and Ingenuity Pathway analyses identified distinct biologic pathways associated with each NMF subgroup. Subclass-gene network associations include: Class A - cell cycle and DNA repair, Class B - cell-to-cell signaling and interaction, Class C - cell growth, proliferation and migration. Whole genome array CGH (Agilent) analysis showed excellent preservation of major genomic aberrations characteristic of GBM and further identified subclasses with genomically defined lesions including: Amplifications - PIK3C2B (n=2), MDM4 (n=2), PDGFRA (n=1), EGFR (n=6), CDK6 (n=4), CDK4 (n=1); Homozygous deletions - PTEN (n=2), CDKN2A (n= 15). Ongoing studies will integrate genomic and mRNA expression data in order to refine and/or redefine GBM subclasses. This library will provide the basis for preclinical drug evaluation and contribute to therapeutic decision making by allowing for real-time identification of tumor specific combinatorial drug sensitivities.

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PLATFORM SESSION 7: DEVELOPMENT/PEDIATRIC Saturday 2-4 p.m. June 25, 2011

113 The Application of Layer Specific Immunohistochemistry to Hemimegalencephay

Aaron Wagner, Nicole Yin, Alexander Bottini, Gary Mathern, Harry Vinters. UCLA

Neuropathologic features of hemimegalencephaly (HME) include multiple distinct, but often overlapping, features of abnormal cortical and subcortical architecture. These include: severe cortical dysplasia, polymicrogyria, band heterotopia, nodular heterotopia, lisencephaly and others. Cortical layer specific markers may provide a tool not only to help categorize different variants of HME, but also suggest pathophysiology of the observed architectural abnormalities, their relationship, to other more well described disorders, and to guide future efforts at treatment. We surveyed characteristics of 14 cases of HME seen at UCLA between the years of 1996 and 2010 using several immunohistochemical markers including MAP1b, Pax6, N200, Calbindin, NEU-N and Tau. Abnormal expression patterns were observed with each of these markers, with the exception of Tau. We did not observe significant immunoreactivity for phosphorylated Tau in any of our cases of HME. MAP1b showed derangements of normal cortical layering in each case, including effacement of laminar architecture, crowding of pyramidal neurons, and disordered neuronal polarity. N200 showed abnormal expression in dysmorphic neurons and balloon cells in cases witch exhibited these cells on routine staining. PAX6 showed increased expression across all variants of HME observed. Calbindin showed focal loss of expression among pyramidal neurons in neocortex. Finally, NEU-N highlighted abnormal cortical layering, neuronal crowding and abnormal neuronal orientation in each case of HME.

CONCLUSIONS: These results could support prior postulated mechanisms suggesting derangements in neuronal cell migration, maturation and protein expression in the pathogenesis of hemimegalencephaly.

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114 GABAA Receptor Abnormalities in the Medullary Serotonergic System in the Sudden Infant Death Syndrome

Kevin Broadbelt, David Paterson, Richard Belliveau. Children's Hospital Boston; Felicia Trachtenberg. New England Research Institutes; Elizabeth Haas, Henry Krous. Children's Hospital San Diego and University of California, San Diego; Hannah Kinney. Children's Hospital Boston

GABAergic neurons in the medulla oblongata through their extensive interactions with caudal serotonergic (5-hydroxytryptamine [5-HT]) neurons, i.e., the medullary 5-HT system, help regulate homeostasis. Complex reciprocal GABA/5-HT interactions exist, with each neurotransmitter modulating the other's actions. Previously, abnormalities in multiple 5-HT tissue markers in the medullary 5-HT system of infants dying from sudden infant death syndrome (SIDS) have been identified by us, suggesting that medullary 5-HT dysfunction is involved in its pathogenesis. Here we tested the hypothesis that markers of GABAA receptors are altered in the medullary 5-HT system (critical to homeostatic regulation) in SIDS cases compared to controls, adjusted for postconceptional age and postmortem interval. We used tissue receptor autoradiography with the radioligand 3H-GABA and displacer isoguvacine after preincubation with the GABAB blocker baclofen. We also examined the expression of the GABAAa3 receptor subunit with western blotting. We found 43-55% reductions in GABAA receptor binding density in key cardiorespiratory-related medullary nuclei sampled in the SIDS cases (n=28) compared with age-adjusted controls (n=8) (p< 0.02), and in the same nuclei with altered 5-HT1A receptor binding previously demonstrated by us. The major exception was that there was no difference found in binding in the arcuate nucleus at the ventral medullary surface. We also found a 46.2% reduction in the levels of the GABAAa3 subunit in the gigantocellularis (key component of the medullary 5-HT system) of SIDS cases (n=24) compared with controls (n=8) (56.8% standard in SIDS cases versus 99.35% in controls; p=0.026). No association was found between GABAA receptor binding or GABAAa3 subunit expression and known risk factors for SIDS. These data suggest that the GABAergic, as well as 5-HT, receptors are dysfunctional in SIDS infants in a common homeostatic network in the medulla, and that SIDS is a complex disorder of this network that includes impaired GABA/5-HT interactions.

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115 Persistence Of Apoptotic Debris In Brain-Specific Rac1-Conditional Knockouts: Evidence For A Macroglial Clearance Function

James Mandell, Daniel Heffron. University of Virginia, Dept. of Pathology; Jennifer Sokolowski. Medical Scientist Training Program and Neuroscience Graduate Program

Apoptotic neuronal death plays an important role in the sculpting of developing brain circuits and is aberrantly activated in neuropathological states. Silver degeneration stains performed on human autopsy material indicate that degenerating axons may persist for years after trauma or neurosurgical procedures, suggesting that clearance in the adult brain is very ineffective. Surprisingly little is known about the cellular and molecular mechanisms of the recognition and clearance of apoptotic neurons in the developing or mature central nervous system. Most work has assumed a generic role of professional phagocytes, namely the microglia, in CNS clearance. Because the small GTPase Rac1 is an essential component of the phagocytic machinery, we generated mice lacking Rac1 in the nervous system (Rac1-/-) by crossing mice homozygous for a floxed Rac1 allele (Rac1flox/flox) with mice heterozygous for a floxed Rac1 allele (Rac1flox/wild type) and expressing Cre recombinase under the control of a Nestin promoter. The Nestin promoter is not active in microglia or macrophages, and thus these cell types remain competent for phagocytosis in the knockouts studied. We show that Rac1-/- mice exhibit a massive developmental accumulation of apoptotic neuritic debris (cleaved caspase 3- and fractin-immunoreactive) during the second postnatal week in several brain areas. There was no significant increase in the density of apoptotic cell bodies in most brain regions (detected either by TUNEL or cleaved caspase 3 immunoreactivity), suggesting a specific defect in neuritic clearance. In contrast to the NestinCre:Rac1 knockouts, Pcp-Cre (Purkinje cell-specific) and ChAT-Cre (cholinergic neuron-specific) conditional Rac1 knockouts do not exhibit apoptotic debris accumulation, arguing against an alternative interpretation, that the NestinCre:Rac1 phenotype is due primarily to an elevated rate of neuronal apoptosis. Our results are consistent with a role for a macroglial cell type in the clearance of apoptotic neurites during development.

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116 Mechanisms of Sensorineural Hearing Loss Related to Congenital Infection by the CMV in Human

Natacha Teissier, Anne-Lise Delezoide, Suonavy Khung-Savatovsky. INSERM U676; Bettina Bessiere, Anne-Elizabeth Mas. INSERM; Pierre Gressens, Thierry Van Den Abbeele. INSERM U676; Homa Adle-Biassette. INSERM

Congenital infection by CMV is the first cause of non-hereditary congenital sensorineural hearing loss (SNHL) in humans. SNHL may be present at birth or appear after several years. The management of these congenitally infected infants is not clearly codified. The aim of this study was to investigate the inner ear and CNS lesions in fetuses with CMV infection and to correlate these findings with the viral load in amniotic fluid, placenta and organs. Seven fetuses aged 21 to 35 weeks of gestation were examined. Diffuse lesions were found within the inner ear, the brain, organs and placenta in all the fetuses and suggested a hematogenous dissemination. CMV preferentially infected proliferating cells compared to highly differentiated sensorineural cells. Combination of immunohistochemistry and in situ hybridization did not evidence any viral latency. In the cochlea, the lesions predominated in the stria vascularis. In the vestibule, the dark cells of the saccule and the utricule were constantly and extensively infected. Lesions were also present in the vestibulo-cochlear nerve and in the spiral ganglion. In the brain diffuse micronodular meningoencephalomyelitis, microcephalia and microgyria were present. The severity of the lesions was correlated to the stage of gestation but no obvious parallelism was identified between the viral load in the amniotic fluid, fetal viral load, neurological lesions and inner ear lesions. In conclusion, this study suggests that the major mechanism of SNHL and vestibular disorders is the disruption K homeostasis in the cochlea and in the vestibule leading to a secondary degeneration of the organ of Corti. Direct cytopathic effect and indirect inflammatory mechanisms may be involved. The success of cochlear implantation in children suggests that despite the presence of diffuse CNS lesions, the function of the VIIIth nerve as well as the rest of the auditory pathway is conserved.

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117 Mitochondrial Enzyme Histochemistry Demonstrates Hypermetabolic Neurons in Paediatric Epileptic Foci

Harvey Sarnat, Laura Flores-Sarnat, Walter Hader. University of Calgary

Background. The hypermetabolic state of repeatedly discharging neurons in epileptic foci might be demonstrated histochemically as increased mitochondrial enzymatic activity in brain resections for epilepsy in children.

Materials and Methods. Frozen sections were studied histochemically of 10 brain resections from 7 epileptic children, 2 months to 17 years of age. Three patients had tuberous sclerosis or hemimegalencephy. Tissues included hippocampus and neocortex. Oxidative enzymes were studied for respiratory chain complexes I, II, IV, using the muscle biopsy protocol. Immunoreactivities as neuronal markers and a-B-crystallin, as well as transmission EM, also were performed.

Results. Oxidative activities were variable in adjacent neurons within a field: a minority were strong, next to neurons with much weaker mitochondrial activity and poor contrast of the soma because of similar oxidative activity in surrounding neuropil. Endothelium of vessels uniformly exhibits strong activity.

Conclusions. Histochemistry of mitochondrial oxidative enzymes reveals scattered hypermetabolic neurons at epileptic foci. This activity does not denote a field of the epileptic focus in the same manner as a-B-crystallin, but rather identifies individual hypermetabolic, and probably epileptogenic, neurons within the focus.

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118 GABAA Receptor Binding in the Cerebral Cortex in Periventricular Leukomalacia (PVL)

Gang Xu, Kevin Broadbelt, Robin Haynes. Children's Hospital Boston and Harvard Medical School; Rebecca Folkerth. Brigham and Women's Hospital and Harvard Medical School; Felicia Trachtenberg. New England Research Institutes; Joseph Volpe, Hannah Kinney. Children's Hospital Boston and Harvard Medical School

Cognitive deficits are a major adverse outcome of prematurity associated with PVL, yet their neuroanatomic substrate is poorly understood. We previously showed that GABAergic neurons migrate to the cortex from subependymal germinal zones across the white matter over the second half of gestation, i.e., the peak period of PVL risk, and that PVL is associated with white matter neuronal loss. Pre- and post-synaptic GABAA receptors in the cortex are essential to GABAergic activity in cognitive processing. Here we tested the hypothesis that GABAA receptor binding in PVL cases compared to controls is reduced overall or in specific patterns in the cerebral cortex either overlying or distant from periventricular necrotic foci. We analyzed 3H-GABA binding utilizing the displacer isoguvacine with tissue autoradiography in the parietal associative cortex of 11 PVL cases and 21 controls adjusted for postconceptional age and postmortem interval. The absolute binding density was determined at percent distances from the pial surface, and a repeated measures regression model was applied with an interaction term to test for differences in the spatial distribution and absolute binding levels with increasing age. The developmental pattern of GABAA receptor binding was essentially the same in the PVL cases and controls, with uniformly low binding throughout all layers at midgestation and increasingly elevated binding in the middle layers until approximately 3.5 postnatal years, the oldest control time-point analyzed. There was also no significant difference in the absolute binding levels between the PVL cases and controls with increasing postconceptional age, as confirmed by comparative assessments at 25%, 50%, and 75% distances from the pial surface (p>0.05). These data suggest that a defect in GABAA receptor binding is not present in the cerebral cortex in PVL, and that other GABAergic tissue marker, i.e., cell density, are needed to detect a potential GABAergic defect in PVL.

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119 Diffuse Astrocytomas of the Childhood Spinal Cord: Correlation Between Incidence and Presence of a Neural Precursor Cell Type

Morgan Freret, Michelle Monje. Departments of Neurology & Developmental Biology; Marilyn Masek. Department of Neuropathology; Paul Fisher. Department of Neurology; Hannes Vogel. Department of Neuropathology; Philip Beachy. Stanford University, Department of Developmental Biology

We previously reported the presence in the ventral brainstem of a Nestin neural precursor cell whose location and frequency correlate remarkably with the incidence of diffuse intrinsic pontine glioma (DIPG), highlighting a candidate cell of origin for DIPG. This cell type is observed diffusely throughout the ventral pons and medulla but never in the midbrain or dorsal pons. We suspected that this cell type might represent a myelinating precursor cell contributing to mid-childhood myelination of a ponto-spinal tract(s), such as the reticulospinal tract. In light of this, we sought to ascertain whether this cell population was also present in the spinal cord during childhood. Archival autopsy samples of the human thoracic and lumbar spinal cord were collected from a cohort (n = 11) of subjects aged 3 months to 17 years with no history of neurodevelopmental or oncological disease, immunostained for the neural stem/precursor cell marker Nestin, and analyzed by light microscopy. Here, we report that this Nestin precursor cell type is present in the postnatal spinal cord white matter at ages <10 and essentially disappears after age 10. This temporal distribution is in contrast to the sharply peaking density of this cell type in the ventral pons. Furthermore, the age distribution of this Nestin precursor cell in the spinal cord correlates closely with the incidence of diffuse Grade II-IV spinal cord astrocytoma gleaned from the Lucile Packard Brain Tumor Database (1997-2008; 11/699 cases). This Nestin cell type thus is present in the ventral pons and below, and its density correlates with the incidence of diffuse astrocytoma in both pons and spinal cord, suggesting a candidate cell of origin. We postulate that the risk of malignant transformation depends on not only the frequency of this susceptible cell type, but also as yet unknown microenvironmental factors that support oncogenesis.

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120 Is Hemimegalencephaly a Fetal Tauopathy?

Harvey Sarnat, Laura Flores-Sarnat. University of Calgary; Peter Crino. University of Pennsylvania; Walter Hader, Luis Bello-Espinosa. University of Calgary

Background: Upregulation of abnormally phosphorylated tau protein is a feature of many adult neurodegenerative diseases, but is not reported in fetuses or infants. Abnormal tau during development potentially can interfere with growth, differentiation and migration of neuroblasts and glioblasts by microtubular disruption, resulting in dysgenesis and hamartoma. Several reports have identified enhanced mTOR cascade signalling in hemimagalencephaly (HME).

Methods: We examined surgical resections for epilepsy of brains of 3 infants with HME. One case died postoperatively of complications and autopsy was performed promptly, providing opportunity to examine other brain structures. Multiple immunocytochemical cell markers, mildly phosphorylated tau antibody and a-B-crystallin were examined, as well as ultrastructural examination. The mTOR pathway also was studied.

Results: Overexpression of tau protein was demonstrated in the hippocampus and neocortex. Antibodies against a-synuclein, ubiquitin and TDP45 were nonreactive, but a-B-crystallin was positive. Many dysmorphic cells showed mixed neuronal/glial lineage and expression of nestin and vimentin. Resident stem cells in the dentate gyrus were proliferated. EM exhibited lipidoic degeneration of many hippocampal neurons. The contralateral hemisphere, by contrast, did not show tau overexpression, except in rare, scattered dysmorphic neurons, and none in subcortical structures. Robust immunolabeling for the phosphorylated isoform of S6 protein, a marker of activated mTOR signalling, was identified in the dysmorphic cells.

Conclusions: Abnormal tau expression may be a factor in the pathogenesis of HME by disrupting microtubule assembly and the mTOR pathway during cellular growth and differentiation.

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PLATFORM SESSION 8: NEURODEGENERATIVE-OTHER Saturday 2-4 p.m. June 25, 2011

121 Treatment of Prion Disease in Brain Aggregate Cultures Predicts Results In Vivo

Stephen DeArmond, Krystyna Bajsarowicz, Misol Ahn. University of California, San Francisco

Our goal was to develop an in vitro model of prion diseases that mimics the composition of the brain, is infectable with prions, and can be used to test drugs. Prion-infected cell lines, such as scrapie N2a cells (ScN2a) have been used for drug screening, but the results are often misleading. Quinacrine (Qa) cleared PrPSc from ScN2a cells but failed to clear PrPSc from scrapie-infected mice or patients with Creutzfeldt-Jakob disease. Also ScN2a cells do not reflect the complexity of the central nervous system. Brain aggregates (BA) are prepared from E15 day mouse embryos. The cells are grown in constantly rotated DeLong flasks for 8 days. The single cell suspension forms into aggregates containing neurons, astrocytes, oligodendrocytes and microglia. At 15 days in culture they are infected with prions followed by drug treatments at 25 days. BAs infected with prions convert PrPC to PrPSc. Treatment of prion-infected BAs with a γ-secretase inhibitor (GSI) plus Qa eliminates PrPSc similar to mice. Early neurodegenerative changes in scrapie are shortening of dendrites and decreased numbers of dendritic branches. Dendritic growth rate was measured in uninfected control and prion-infected BAs. We found that dendritic density in uninfected controls doubled between 15 and 25 days. In contrast, BAs infected with scrapie prions showed a progressive decrease in the length and number of dendrites at 25 and 35 days. The results indicate that dendrites degenerate in brain aggregates early during prion disease just as they do in mice. GSI+Qa treatment prevented dendritic degeneration by 100%. The results in vitro were identical to results found in vivo. We concluded that brain aggregates accurately mimic neuropathology caused by prion disease in animals and humans and can be used for drug treatment studies.

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122 Pathologic Examination of Hippocampal Sclerosis (N=106 Cases and 1,004 controls) in the Aging Population

Janna Neltner. Department of Pathology; Frederick Schmitt. Department of Neurology; Yushun Lin. Statistics; Erin Abner. Department of Statistics; Gregory Jicha. Department of Neurology; Ela Patel, Paula Thomason. University of Kentucky, Sanders-Brown Center on Aging; Charles Smith. Department of Neurology; Karen Santacruz. University of Minnesota Department of Pathology; Linda Van Eldik, Peter Nelson. University of Kentucky, Sanders-Brown Center on Aging

OBJECTIVE: Hippocampal sclerosis (HS-Aging) is a relatively prevalent disease of brain aging with strong impact on cognition. Two proposed mechanisms for the HS- Aging pathology involve aberrant TDP-43 expression and vasculopathy. We examined a large group of autopsy-confirmed HS-Aging cases to determine characteristic clinical and pathologic findings, including TDP-43 immunohistochemistry and vascular pathology.

METHODS: In this case/control cross-sectional (autopsy) study, we combined three different large autopsy cohorts - University of Kentucky Alzheimer's Disease Center, the Nun Study, and the Georgia Centenarian Study - for which the neuropathological analyses were performed at the University of Kentucky. From these groups, we examined cortical and hippocampal sections from 106 cases of hippocampal sclerosis by routine H&E, modified Bielschowsky, and Gallyas stains as well as TDP-43 immunohistochemistry. We then compared those findings, and their corresponding clinical information, to 1,004 cases of controls, and also a subset (N=10) of younger individuals with epilepsy-associated hippocampal sclerosis.

RESULTS: We found that hippocampal sclerosis was typically found in older patients than those with AD type pathology, especially over the age of 95. The majority of HS-Aging cases were positive for TDP-43 (90% vs. 10% in normal controls). The HS-Aging pathology was usually bilateral by routine H&E staining; the percentage of bilaterality was increased by the use of TDP-43 immunohistochemistry. As reported previously by others (Lee et al, 2008), there was no aberrant TDP-43 positivity in the epilepsy-associated HS cases. There was no association between vascular pathology and HS- Aging when comparing cases to age-matched controls.

INTERPRETATION: Our findings support the idea that HS-Aging is linked with aberrant TDP-43 and is a major cause of dementia in patients over the age of 85. Clinicopathological features of HS-Aging are quite different from FTLD-TDP. HS- Aging does not appear to be associated with vascular pathology.

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123 Comparative Study: Tauopathy and TDP-43 Proteinopathy in Amyotrophic Lateral Sclerosis and Chronic Traumatic Encephalopathy

Kyung-Hwa Lee, Qinwen Mao, Nailah Siddique, Lisa Kinsley, Manjari Mishra, Katherine Gasho, Teepu Siddique, Eileen Bigio. Northwestern University Feinberg School of Medicine

There is epidemiologic evidence that head injury is a risk factor for amyotrophic lateral sclerosis (ALS). Chronic traumatic encephalopathy (CTE), originally described in impaired boxers (dementia pugilistica), is a 3R/4R tauopathy characterized by neurofibrillary tangles in specific brain regions, without senile plaques (SPs). Overlapping ALS and CTE pathology has recently been reported. To investigate the relationship between CTE and ALS, we compared the immunohistochemical expression of tau and TDP-43 proteins in ALS and CTE. Of 97 ALS autopsy cases, 7 had CTE-type tauopathy (ALStau) (7.2%) and were compared with two cases of CTE. ALStau showed TDP-43-positive neuronal inclusions in the motor cortex but not in other neocortical regions, and patchy inclusions in the hippocampus, basal ganglia, pontine nuclei and medulla. CTE showed rare TDP-43-positive neuronal inclusions in both the motor and non-motor neocortex. Compared to ALStau, CTE displayed more prominent inclusions in the hippocampus, midbrain and medulla. The tauopathy in ALStau was similar to CTE in distribution but tangle density was much lower. 75 of the 97 ALS cases had retrievable clinical history for presence or absence of head trauma (77%). Four of the seven ALStau cases (57%) had a history of head trauma and the other three (43%) either had accidents or played sports (or both) without known history of head trauma. Of the remaining 68 ALS cases, 24 (35%) had a history of head trauma and an additional 13 (19%) had accidents of played sports (or both) without known history of head trauma. Although only 7.2% of our ALS cases had combined TDP-43 and tau pathology, the tau expression is comparable with that of CTE. The results suggest that a history of head trauma is common in ALS, but resultant tauopathy is uncommon. Whether or not head trauma is conceivably causative of ALS remains to be determined.

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124 The Spectrum of Neuropathology in Patients Dying With Amyotrophic Lateral Sclerosis (ALS) - A Clinicopathologic Study

Marla Gearing. Dept. Pathology, Deborah Cooper. Dept. Neurology, Nichole Costa. Dept. Neurology, Jonathan Glass. Emory Univ. Dept. Neurology, ADRC & Ctr. for Neurodegenerative Disease

A recent report by McKee and colleagues described 12 patients with chronic traumatic encephalopathy (CTE), three of whom also had motor neuron disease. This publication has generated considerable publicity and interest regarding the possible role of head trauma in the pathogenesis of ALS. We addressed this issue by studying a cohort of 32 patients with a clinical diagnosis of ALS who were cared for and came to autopsy at Emory University. Brain and spinal cord tissues were examined extensively for the pathological hallmarks of neurodegenerative disease. Neuropathologic analysis confirmed the clinical diagnosis of ALS in all 32 cases. In three patients, a secondary clinical diagnosis of frontotemporal dementia was confirmed pathologically by widespread cortical staining for TAR DNA binding protein-43 (TDP-43). Six other cases showed widespread cortical TDP-43 pathology, but did not have clinical dementia. Additional neuropathologic findings were present in the majority of cases. In particular, the brains from three patients showed pathological tau deposits resembling those described in CTE; one of these patients had a history of head trauma 33 years previous, the other two had no history of head trauma. Twenty-three others exhibited neurofibrillary tangles in a distribution consistent with Braak & Braak staging of neurofibrillary degeneration (scores ranging from I to IV). Fourteen of the 32 cases had widespread Aß deposition in the neocortex. One case with a clinical history of dementia met CERAD criteria for Alzheimer's disease (AD); four others with no history of dementia met criteria for possible AD. Additional diagnoses that were encountered included hippocampal sclerosis (unilateral), infarcts and Lewy body disease. We conclude that in patients dying with ALS, other neuropathologic findings are often identified, none of which negate the primary clinical and pathological diagnosis of ALS. Supported by NIH grants: P50 AG025688 and P30 NS055077.

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125 Neuropathologic Findings in 32 Nondemented Elderly Subjects

Mark Jentoft. Mayo Clinic College of Medicine; Joseph Parisi, Dennis Dickson, Kris Johnson, Bradley Boeve, David Knopman, Ronald Petersen. Mayo Clinic

Alzheimer disease (AD) pathology is thought to be the substrate for age-related cognitive decline, with little emphasis of coexisting pathologies frequently present in elderly subjects. We describe neuropathologic findings in 32 Mayo Clinic ADRC/MCSA nondemented subjects with <18 months (mean 260 days) follow-up from last evaluation to autopsy. Assessments included Consortium to Establish a Registry for Alzheimer Disease (CERAD) plaque scores, Braak stages, and National Institute on Aging-Reagan (NIA-Reagan) criteria. Subjects included 19 women and 13 men, average age at death 90.25 years (range 81-102). Braak stages ranged from II (10) to III-IV (16) to V (6), while CERAD scores varied from 0 (10) to A (11) to B (10) to C (1). NIA-Reagan assignments were problematic in neurofibrillary tangle-dominant cases with high Braak stages and low to moderate CERAD scores, which were operationalized as Low or Intermediate likelihoods, respectively. By these criteria, most cases were either 'not AD' (10) by absence of neuritic plaques (CERAD 0), or Low likelihood AD (11); about 1/3 were Intermediate likelihood (10), and only one, High likelihood. Lewy body (LB) pathology, identified in 6, ranged from sporadic brainstem/limbic lesions (3) to neocortical Lewy body disease (3). Argyrophilic grain disease (AgD) was present in 40% (13). Hippocampal sclerosis (HS) occurred in 4, all with NIA-Reagan Low likelihood. Vascular pathologies were common, including arteriolosclerosis (29), amyloid angiopathy (20), lacunes (10), and infarcts (5).Although most patients demonstrate some AD neurodegenerative changes, these are insufficient for a pathologic diagnosis of AD. Coexisting pathologies likely cause or contribute to clinical impairment, as only 16% (5/32) were free of vascular, LBs, AgD or HS. It is timely that new clinical criteria for cognitive impairment are being developed to include biomarkers that will likely refine diagnosis and provide new insights in disease pathogenesis and potential therapeutic targets. Supported by AG016574 and AG006786.

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126 The Cerebellum in Friedreich's Ataxia

Arnulf Koeppen, Ashley Davis, Jennifer Morral. VA Medical Center

Friedreich's ataxia (FRDA) causes progressive destruction of the dentate nucleus (DN). The lesion is apparent on T2-weighted magnetic resonance images, which may be used as a biomarker of the disease. Synaptic connections between Purkinje cells, neurons of the dentate nucleus, and nerve cells in the inferior olivary nucleus are tightly organized in a topistic manner. Movement control depends on the proper reciprocal interaction of these gray matter structures in a 'cerebellar module'. In FRDA, the module is impaired only at the level of the dentate nucleus. The main neurohumoral transmitters in the module are glutamate and gamma-aminobutyric acid (GABA). We visualized GABA-ergic and glutamatergic connections in autopsy tissues of 24 FRDA patients by immunocytochemistry with antibodies to glutamic acid decarboxylase (GAD) as an indicator of GABA-ergic transmission, and vesicular glutamate transporters 1 and 2 (VGluT1, VGluT2) as selective markers of glutamatergic transmission. Neuronal loss in the DN spares small GABA-ergic nerve cells that give rise to dentato-olivary connections. The integrity of this tract prevents trans-synaptic degeneration of the inferior olivary nucleus. As expected, glutamatergic input to Purkinje cells by VGluT2-positive climbing fibers also remains normal. Glutamatergic mossy fiber terminals (VGluT1 and VGluT2) in the granular layer and parallel fibers in the molecular layer (VGluT1) are also intact. Mossy and climbing fiber collaterals to the DN, however, are fewer than normal. Grumose degeneration in the DN is due to proliferation of Purkinje cell-derived GABA-ergic terminals. These abnormal synaptic clusters are non-reactive for VGluT1 or VGluT2. The described observations allow the conclusion that cerebellar dysfunction in FRDA is mainly due to lack of GABA-ergic corticonuclear transmission. Neuronal loss in the DN also causes a deficit of glutamatergic transmission arising from mossy and climbing fibers collaterals. Grumose degeneration may represent a reparative response.

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127 Loss of Layer Pre-Alpha Entorhinal Neurons in Huntington Disease and Controls: Age, Braak Stage, and Vonsattel Grade

John Hedreen. Harvard Brain Tissue Resource Center, Dept. Psychiatry, McLean Hospital

Loss of layer Pre-alpha neurons (magnocellular neurons in layer II) in the entorhinal cortex has been described in Huntington disease (HD). The present study explores this neuron loss in relation to the Vonsattel grade of neuropathological severity in HD, age at death, and Braak neurofibrillary degeneration stage, in HD and control brains. Pre-alpha neuron loss was semiquantitatively scored at the anterior hippocampal level in age-matched control and Vonsattel grades 2, 3, and 4 HD (HD-2, HD-3, HD-4) brains (n= 35 each group), all at Braak stages 0/I. Additional age-matched control, HD-2, HD-3, and HD-4 brains at higher Braak stages (II, III) were also scored. Pre-alpha neuron loss in HD increases significantly with Vonsattel grade. In the HD groups it is correlated with age, and with Braak stage. Mean neuron loss in controls is not correlated with age but is correlated with Braak stage. Neuron loss is significant at Braak stage III compared to stage 0/I in both control and HD-2 groups. In conclusion, neuron loss in entorhinal cortex in controls is present at Braak stage III; in HD this neuron loss is increasingly responsible for cognitive disability as disease severity advances, and is more severe with older age and higher Braak stage.

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128 Nrf2 Signaling Modulates Neuronal NMDA Receptor Currents in a Glia-Dependent Manner

Junghyun Hahn. University of California San Francisco; Xianhong Wang. University of California San Francisco; Marta Margeta. University of California San Francisco

Nrf2, a transcription factor that regulates expression of a battery of antioxidant and phase II detoxification enzymes, plays an important role in neuronal susceptibility to injury. Activation of Nrf2 pathway protects neurons from insults such as glutamate excitotoxicity and cerebral ischemia, and promotes survival in mouse models of Parkinson's disease and amyotrophic lateral sclerosis. Little is known, however, about the effects of Nrf2 pathway on synaptic signaling. The NMDA subtype of ionotropic glutamate receptors (NMDAR) plays a central role in normal synaptic function and plasticity, but has also been implicated in neuronal cell death. Here, we investigated how Nrf2 pathway regulates NMDAR currents at baseline and oxidative stress conditions using hippocampal neurons derived from embryonic rats, which were cultured with or without mixed population of glia for 13-17 days prior to whole-cell voltage clamp recording. To probe the modulation of NMDAR by reactive oxygen species (ROS), we applied NMDA together with hypoxanthine/xanthine oxidase (HX/XO), a substrate/enzyme system that mimics oxidative stress by generating a mixture of superoxide anions, hydroxyl radicals, and hydrogen peroxide. When HX/XO and NMDA were applied together, the induced inward current was significantly larger than the sum of currents induced by either NMDA or HX/XO alone; there was no significant difference in the degree of HX/XO-induced potentiation of NMDAR current between the two culture conditions. Interestingly, activation of Nrf2 pathway by overnight pretreatment with sulforaphane abolished HX/XO-induced potentiation of NMDA current in mixed cultures, but had no effect in pure neuronal cultures. Similarly, baseline (ROS-independent) NMDAR currents were significantly attenuated by sulforaphane pretreatment in mixed, but not in neuronal cultures. Thus, Nrf2 pathway regulates NMDAR signaling in a glia-dependent manner, providing a potential mechanism for Nrf2-mediated neuroprotection under oxidative stress conditions and raising a possibility that Nrf2-targeting drugs may modulate learning and memory.

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POSTER SESSION 2 Saturday, June 25, 2011, 8 a.m. - 6 p.m.

129 Sensory Perineuritis: Reappraisal of the Entity

Negar Khanlou, Tracie Pham. Pathology-Neuropathology, Perry Shieh. Neurology - M Verity, Pathology-Neuropathology, UCLA David Geffen School of Medicine

The entity of sensory perineuritis was first described by Asbury et al in 1972 as a relapsing painful migratory neuropathy characterized pathologically by perineurial scarring and inflammation, associated with variable interfascicular Wallerian degeneration considered compressive in etiology. Subsequent case reports have broadened this initial description.We report two non-diabetic cases of sensory perineuritis with clinical features suggestive of a vasculitic neuropathy and propose redefinition of the terminology based on our findings and review of the literature. Patient A was a 46 year-old male with 4 month history of progressive ascending bilateral lower extremity weakness without sensory symptoms and axonal polyneuropathy along peroneal nerve distribution on nerve conduction studies. Patient B was a 79 year-old female with a 6 week history of gradual asymmetrical bilateral lower extremity weakness, footdrop and sensory hypoesthesia bilaterally. Electrodiagnostic testing revealed evidence of left sciatic neuropathy. Both patients responded clinically after treatment with corticosteroids and intravenous immunoglobulin. Superficial peroneal nerve biopsy in both patients revealed perineurial scarring, perineurial lamina widening, and chronic inflammation confined to the perineurium and epineurial perivascular areas. There was no evidence of necrotizing vasculitis and the endoneurium was intact. Epithelioid/giant cells were absent. Special staining for leprobacilli was negative. Active Wallerian degeneration with interfascicular variability, loss of large and small myelinated fibers and diminished unmyelinated fibers were noted.

Conclusion: Our cases confirm the variable sensori-motor nature of this entity which is pathologically defined on nerve biopsy. This broadens the original description of sensory perineuritis and allows for the redefinition of the entity as a fibrosing perineuritis.

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130 Nemaline Rods-like Structures Presented in Colchicine-Induced Myopathy: A Case Report and Literature Review

Jilin Bai, Mark Belsky. University of Connecticut School of Medicine; Daniel Menkes. Department of Neurology, University of Connecticut Health Center; Qian Wu. Dept of Pathology & Laboratory Medicine, UConn Health Center

Colchicine-induced myotoxicity is a rare medication side effect manifested by myopathic changes characteristic of intracytoplasmic lysosomal and autophagic vacuoles without necrosis. This myopathic alteration is related to colchicine-mediated disruption of the microtubule-dependent cytoskeletal network. We report a 66-year-old female who developed subacute onset of lower extremity proximal muscle weakness after taking colchicine 0.6mg by mouth twice a day for four weeks. A serum creatine phosphokinase (CPK) was elevated at 1031 IU/L (reference range 30-135 IU/L). A left quadriceps femoris muscle biopsy showed striking cytoplasmic vacuoles and Nemaline rods-like inclusions affecting both type I and type II fibers with a predominance of type II fibers. Electron microscopy demonstrated autophagic vacules as well as Nemaline rods-like structures in the central region of the myofibers. These rods-like structures were composed of closely packed filaments arranged in a lattice-like pattern and had a periodicity comparable to that seen in Z lines. Five weeks after the colchicine was discontinued, the patient's muscle strength gradually improved along with a concomitant reduction in CPK to 145 IU/L. To our knowledge, this is the first report of a colchicine-induced myopathy to be associated with Nemaline rods-like structures. The quantity and distribution of the rods in colchicine-induced myopathy are different from what is observed in typical Nemaline (rod) myopathy. This case indicates that the subacute myotoxicity induced by colchicine may disrupt the organization of myofibrils especially the sarcomeric cytoskeleton in addition to a disruption of microtubule-dependent cellular cytoskeleton.

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131 Non-Diagnostic Muscle Biopsies in Late Onset Pompe's Disease

Ana Lia Taratuto. Institute for Neurological Research.FLENI.Buenos Aires.Argentina; Alberto Dubrovsky, Jose Corderi. Favaloro Foundation.Neuromuscular Unit.Buenos Aires.Argentina

Late-onset Pompe's disease main symptoms may be progressive muscle weakness, waddling gait,frequent falls, back pain,scoliosis, respiratory insufficiency or difficulty in swallowing. The literature clearly states a mean delay between onset of symptoms and diagnosis ranging from 8.4-10.4 yrs. We report early biopsies in three cases.1: a 29 yrs-old male had weakness in lower limbs for two yrs. Muscle biopsy showed slight variation in fibre sizes with onerimmed vacuole(RV). Immunostainings for limb-girdle muscular dystrophy (LGMD) were nornal. Progessive weakness andventilatory insufficiency developed since 35 yrs-old. A 2nd biopsy at 37yr old was diagnostic for Pompe's disease. Acid Alpha Glucosidase (AAG) was 0.01(normal 0.49-1.27).2: a 29 yr-old male developed respiratory insufficiency requiring nocturnal ventilatory assistance. He had proximal muscleweakness and slight scapular winging but no facial involvement. A biceps muscle biopsy showed slight type 2 fibre hypertrophy and normal histochemistry and ultrastructure as well as immunostainings and WB for LGMD. 1/3 siblings had two unconclusive biopsies in USA. AAG proved abnormal and DNA studies confirmed the disease.3: a 48 year-old female developed right lid ptosis and lower limbs weakness, suspected of having myasthenia gravis. Later shedeveloped waddling gate and left lid ptosis. At 68 yr old, had a 'myopathic'EMG, CK within normal range, normal musclebiopsy with one ragged red fibre. Then had breathing difficulties in supine position, proximal limb as well as axial weakness and scapular winging. Adult Onset Pompe Disease was confirmed with an abnormaly low AAG activity assay at 78 years old. Based on the literature and on our own experience, our strategy for interpretation of muscle biopsy in 'sporadic' or familial cases referred as possible late-onset LGMD with almost normal biopsy or with a single RV is first rule out Pompe's disease.

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132 Myopathy With Atypical Inclusions in a Child With Progressive Hypertonia, Scoliosis and Respiratory Failure

Benjamin Ellezam. The Methodist Hospital; Tim Lotze. Neurology; Chester Brown. Molecular Genetics; Adekunle Adesina. Texas Children's Hospital - Pathology - Baylor College of Medicine

This is a 14 year-old male of African-American descent who presented with a 6 month history of progressive limb stiffness, failure to thrive, and recent onset of shortness of breath, and rapidly deteriorated to a state of cardiac and respiratory failure requiring intubation. He has a previous diagnosis of Shone's complex associated with congenital cardiac malformations and severe scoliosis with secondary restrictive lung disease. He has above average intelligence. There is no family history of neuromuscular diseases or consanguinity. Neurologic examination while intubated was remarkable for diffuse hypertonia. In the face of rapid clinical deterioration and unknown disease etiology, muscle (thigh) and sural nerve biopsies were performed. The nerve biopsy did not show any significant pathologic abnormalities. The muscle biopsy showed a primary myopathy with atypical inclusions. No dystrophic or inflammatory changes were seen. The inclusions were variably sized, peripheral or central, and hyaline-like on H&E. They were also highlighted on Gomori's trichrome and non-specific esterase stains. They showed partial immunoreactivity for muscle specific actin (HHF35), ubiquitin, and dysferlin. They were negative for desmin, alpha-, beta- and gamma-sarcoglycans, alpha- and beta-dystroglycans, laminin, lamin a/c, dystrophin, vimentin, and beta-amyloid. Collagen VI immunostain was normal. On electron microscopy, many large ovoid to elongated finely fibrillar/granular inclusions with entrapped glycogen and occasional lipid droplets were present. DNA sequencing of the desmin gene showed no mutation. The features of these inclusions are atypical and to our knowledge have not been reported in any of the currently known myopathies with inclusions.The patient showed improvement with supportive treatment and was discharged. The 3 month follow-up visit was remarkable for new onset of difficulty swallowing and persisting hypertonia.

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133 Fatal Hydroxychloroquine-Induced Skeletal and Cardiac Myopathy

Bret Mobley. Department of Pathology; Morteza Azimian. Department of Medicine; James Atkinson. Vanderbilt University Department of Pathology

Hydroxychloroquine (HCQ) is an antimalarial agent widely used for long term management of common rheumatologic diseases. Skeletal and cardiac myopathies are under-recognized side effects of HCQ treatment. We report a 64-year-old female on chronic HCQ therapy for rheumatoid arthritis who presented with progressive weakness and weight loss. A diagnosis of polymyositis was rendered on the basis of electromyography studies and prednisone treatment was initiated. The patient subsequently presented with dyspnea, at which time myocardial infarction and diastolic heart failure were identified. Biopsy of the left quadriceps muscle demonstrated a necrotizing vacuolar myopathy. Electron microscopy revealed large numbers of secondary lysosomes containing myelin figures. Endomyocardial biopsy demonstrated cytoplasmic vacuolization with identical ultrastructural features. The clinical and pathologic features were classic for HCQ myotoxicity. The patient died several days later. Myotoxicity due to hydroxychloroquine may be fatal and should be considered in connective tissue disease patients presenting with otherwise unexplained weakness or cardiac complaints.

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134 Unusual Morphological Changes in Skeletal Muscle Biopsy of Children With Drop-Head Syndrome due to LMNA Mutations

Fabiana Lubieniecki. Pediatric Hospital JP Garrahan, Department of Pathology. Argentina; Maria Soledad Monges. Pediatric Hospital J P Garrahan, Neuropaediatrics. Argentina; Maria Saccoliti. FLENI. Department of Neuropathology, Buenos Aires, Argentina; Susana Quijano-Roy. Service de Pediatrie, Hopital Poincare, Garches, France; Pascale Richard. UF Cardiogenetique et Myogenetique Moleculaire et Cellulaire. GHU Piti; Norma Romero. Institut de Myologie, GHU Pitie Salpetriere, AP-HP, Paris, France; Ana Lia Taratuto. Pediatric Hospital, Department of Pathology; FLENI, Argentina

We describe the unusual morphological findings of muscles biopsies from a group of patients with drop-head syndrome affected by LMNA mutations in early years of life. Case 1, male- 2years(yrs) old. Since 11months (mo) he had progressive hyperlordosis and head drop as well as loss of deambulation at 15 mo, proximal upper limbs and distal lower limbs hypotrophy. CK 841 IU/l. A deltoid muscle biopsy showed a dystrophic pattern, increase in central nuclei as well as fibers increase in central oxidative activity in some fascicles. Molecular studies showed a Lys32Glu(K32E) missense heterozygous mutation in exon 1.Case 2, male- 2yrs and 3 mo old, normal siblings. Since 1 yr of age he had generalized weakness and proximal upper limbs hypotrophy, frequent falls and head drop. CK 937 IU/l. A deltoid muscle biopsy showed perimysial fibrosis, relatively preserved structure, only one necrotic fiber could be disclosed and there was increase in oxidative central activity in some fascicles. Ultrastructural study showed abnormal nuclei with fragmentation, invagination and folding membrane. Molecular studies identified a missense heterozygous mutation p.Thr528Arg (c.1583 C> G) in exon 9. Case 3.female- 7 mo of age, generalized hypotonia, weakness, areflexia and drop head. CK 991 IU/l. A deltoid muscle biopsy showed extreme fascicular atrophy, severe perimysial fibrosis, a few hypertrophic rounded fibers of both fiber types, only focal slight endomysial fibrosis. Ultrastructural study showed abnormal nuclei with invagination, fragmentation and seudoinclusions. Molecular studies identified a missense heterozygous mutation p.Arg249Trp (C.745 C> T) in exon 4. Since first recognized as an entity by Quijano et al in 2008, there are only few reports on LMNA-related congenital muscular dystrophy. The spectrum of morphological changes highlights the importance of this particular clinical phenotype in order to direct molecular genetic analysis to LMNA in early onset myopathies.

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135 Responses to Treatment Are Dependent on Fiber Type, Muscle, and Mutation in Murine Models of Myotubularin Deficiency

Michael Lawlor, Marissa Viola, Rachel Edelstein. Children's Hospital Boston/Harvard Medical School, Boston, MA; Christopher Pierson. Nationwide Children's Hospital/Ohio State University, Columbus, OH; Anna Buj-Bello. Genethon, Evry, France; Jennifer Lachey. Acceleron Pharma, Cambridge, MA; Jasbir Seehra. Acceleron Pharma, Cambridge, MA; Alan Beggs. Children's Hospital Boston/Harvard Medical School, Boston, MA

X-linked myotubular myopathy (XLMTM) is a congenital myopathy caused by deficiency of the lipid phosphatase, myotubularin. XLMTM patients often present with severe perinatal weakness that requires mechanical ventilation to prevent death from respiratory failure. Muscle biopsies from XLMTM patients display small myofibers with central nuclei and central aggregations of organelles in many cells. We proposed that therapeutically increasing muscle fiber size would cause symptomatic improvement in myotubularin deficiency. Recent studies have elucidated an important role for the activin receptor type IIB (ActRIIB) in the regulation of muscle growth, and have shown that ActRIIB inhibition results in significant muscle hypertrophy. We recently reported that treatment with the extracellular domain of the activin type IIB receptor fused to a murine Fc region (ActRIIB-mFC) produced a 17% extension of lifespan in severely symptomatic (Mtm1d4) myotubularin-deficient mice, with transient increases in body mass, and forelimb grip strength. Additionally, pathological analysis of Mtm1d4 mice during treatment revealed that ActRIIB-mFC produced marked hypertrophy that was restricted to type 2b myofibers. Subsequent studies have evaluated ActRIIB-mFC as a treatment for moderately symptomatic (Mtm1C205T) myotubularin-deficient mice, and this trial is currently in progress. In contrast to what was seen using Mtm1d4 mice, preliminary studies suggest that treatment of Mtm1C205T mice results in minimal increases in body mass and no increase in forelimb grip strength. Treatment-induced myofiber hypertrophy in Mtm1C205T mice is restricted to the gastrocnemius muscle, in comparison to the hypertrophy in all muscles that was seen in treated Mtm1d4 mice. Within the gastrocnemius muscle of Mtm1C205T mice, hypertrophy remains restricted to type 2b myofibers. These findings confirm that only type 2b fibers are capable of responding to ActRIIB-mFC treatment in myotubularin-deficient muscle, and that additional factors may determine whether a given muscle is capable of responding to this treatment strategy.

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136 Lymphoid Follicles in a Gastrocnemius Biopsy

Derek Mathis, Jeffrey Elliott, Jack Raisanen, Kimmo Hatanpaa, Dennis Burns, Charles White III, Emily Herndon. University of Texas Southwestern Medical Center at Dallas

Inflammatory myopathies while rare are the cause of significant morbidity and health care cost. Besides evidence of myofiber and connective tissue injury, muscle biopsies typically contain a mixed infiltrate of B-cells, T-cells, and plasma cells. Here we present a 69 year-old woman with painless, progressive right lower extremity weakness and dense perimysial and endomysial inflammation on a gastrocnemius biopsy in the form of lymphoid follicles with B-cell-rich germinal centers encircled by T-cells. Physical examination demonstrated asymmetric leg weakness with prominent plantar flexion weakness and no clear upper motor neuron signs or sensory changes. Her past medical history is significant for hypertension and hyperlipidemia that was treated briefly with statin drugs. Besides the lymphoid follicles, the biopsy also displayed abnormal fiber size variability and both myopathic and neurogenic changes. Although occasional sarcoplasmic vacuoles were present, no tubulofilamentous inclusions characteristic of inclusion body myositis or features of any other disease entity were identified ultrastructurally. These intramuscular lymphoid follicles recapitulate the microarchitecture of normal secondary lymphoid tissue such as lymph nodes, spleen, and tonsil. This phenomenon has been described previously as tertiary lymphoid organ formation and is recognized in the thyroid gland in Hashimoto's thyroiditis and Graves' disease, synovial tissue in rheumatoid arthritis, and salivary gland in Sjogren syndrome. Reports of intramuscular lymphoid follicles are rare however and have included patients with juvenile dermatomyositis, mixed connective tissue disease, and other overlap syndromes. Although an ANA and RF were elevated in this patient, no other clinical or laboratory evidence of a connective tissue disorder has been found. The significance of the patient's history of statin therapy to her muscle pathology also remains unclear. This case adds to the spectrum of intramuscular tertiary lymphoid organ formation and raises important questions about its diagnostic meaning for patients with neuromuscular symptomatology.

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137 Granulomatous Inflammation and Myasthenia Gravis - Is There a Connection?

Michael Kyle, Gordon Peterson, Bryan Tsao, Ravi Raghavan. Loma Linda University Medical Center

Introduction: The peculiar association of non-caseating granulomatous myopathy with eosinophilia, and cardiac arrhythmia in a patient with myasthenia gravis is presented. A concomitance of such features is rare, but has been documented.

History: A 20 year old female presented with worsening myalgia, muscle weakness, and intermittent diplopia. She had normal sensations, cranial nerve function and deep reflexes. Electrophysiology revealed abnormal neuromuscular junction transmission with decrement at low frequency repetitive stimulation and no facilitation with high frequency stimulation. Besides an absolute eosinophilia, and elevated creatinine kinase (1567 U/L), she had increased acetylcholine receptor (AChr) binding antibodies (29.7 nmol/L). An MRI of the thigh showed confluent patches of T2 hyperintensity within muscles, and a biopsy was done.

Results and Course: The muscle demonstrated florid non-caseating granulomatous inflammation, multinucleated giant cells, and myonecrosis. Eosinophils were prominent between muscle fibers. Stains for fungi, mycobacteria, and parasites were negative. Angiotensin-converting-enzyme levels were normal, and there was no hypercalcemia or systemic evidence of sarcoidosis. Based on symptoms, elevated AChr antibodies, and electrophysiology, treatment for myasthenia gravis was started, to which she responded well. A subsequent thymectomy was normal.

Discussion: Granulomatous inflammatory processes, albeit rare, have been reported in association with myasthenia, but myositis, eosinophilia and myalgia are highly unusual presenting features of this disease. The pathogenesis is unclear, but associations with co-existing sarcoidosis and thymoma, and therapy for myasthenia have been implicated. Concomitant myocardial granulomas have been reported in myasthenia, and coincidentally our patient had a cardiac arrhythmia. The findings hint at a peculiar syndrome of immune dysregulation affecting multiple tissue locations. There is one case report of a genetic mutation that may provide a link between myasthenia and granulomatous disease. Neuropathologists may consider myasthenia in the differential diagnosis of granulomatous myopathy (Note: Clinical aspects presented in part at AANEM conference).

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138 Juvenile Xanthogranulomas of the Nervous System: A Report of Two Cases and Review of the Literature

Jeremy Deisch. University of Texas Southwestern Medical Center; Rajankumar Patel, Korgun Koral. Departments of Radiology, Sandy Cope-Yokoyama. Departments of Pathology, UTSW and Childrens Med Center, Dallas, TX

Juvenile Xanthogranulomas (JXG) are uncommon non-Langerhans cell histiocytic proliferations which arise most often in children. While most cases present as solitary cutaneous lesions, occasional cases involve extracutaneous sites. Rare examples of JXGs have been reported involving all levels of the neuroaxis. We present two cases of JXGs involving the nervous system, and review the literature. The first patient was a 14-year-old female with headaches and a mass involving the left trigeminal nerve; pathologic examination showed a JXG. At 11 months follow-up, after administration of systemic chemotherapy, the patient remained stable with residual tumor. The second patient was a 15-year-old female with leg weakness and numbness, who underwent complete surgical resection of a dural JXG. At eight months follow-up, she showed no evidence of tumor, and was able to walk without difficulty. Review of the literature revealed 38 previously published reports of JXGs involving the nervous system. The central nervous system (CNS) was involved in the majority (75%) of cases. The clinical characteristics of JXGs arising in the CNS varied significantly from cases in the peripheral nervous system (PNS); CNS tumors occurred in younger patients, more often males, and were more likely to be associated with concurrent cutaneous and extra-nervous systemic lesions. The clinical outcomes were similar for CNS and PNS lesions, with the caveat that all three lethal JXGs occurred in the CNS. The clinical and radiologic presentation of JXGs is nonspecific, thus necessitating biopsy and pathologic examination to arrive at the diagnosis. The pathologic differential diagnosis includes a heterogeneous group of histiocytic proliferations; immunostaining for histiocytic markers CD68, factor XIIIa, and Fascin, and the absence of Birbeck granules and CD1a immunoexpression suggests the diagnosis of JXG. In many cases, total surgical resection is curative. However, some cases will require additional chemotherapy and/or radiotherapy.

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139 Silent Clivial Pituitary Adenoma

Knarik Arkun, Adam Sandler. Neurosurgery; Patrick Lasala. Neurosurgery; Yvonne Lui, Sophia Rodriguez. Pathology; Christian Keller. Neuropathology; Karen Weidenheim. Montefiore Medical Center, Neuropathology

Ectopic pituitary adenomas usually occur in the sphenoid sinus. Ectopic pituitary adenomas in the clivus are also rare and a source of diagnostic difficulty. We present an ectopic null cell pituitary adenoma arising in the clivus of a 77-year-old woman. The tumor was an incidental finding at the time of head CT performed after head trauma, in an otherwise asymptomatic woman without headaches or changes in vision. MRI of the brain demonstrated a homogeneous contrast-enhancing mass destroying the clivus and sphenoid bone and extending into the petrous apices. Partial dehiscence of the floor of the sella was present. The pituitary infundibulum deviated mildly to the left. Tumor involved the cavernous sinus and Meckel's cave on the right. Based on its aggressive appearance on imaging, malignancies including chordoma, chondrosarcoma, lymphoma, and metastasis were considered preoperatively. At transphenoidal resection of the lesion, a frozen section revealed pituitary adenoma and the mass was subtotally resected. The tumor was composed of uniform polygonal cells with granular amphophilic cytoplasm, arranged in trabeculae and papillae with perivascular pseudorosettes and lacking expression of pituitary hormones, and null cell adenoma was diagnosed. Pituitary hormone levels performed after subtotal resection were all within normal limits. Ectopic pituitary adenomas are thought to arise from remnants of Rathke's pouch and may present with bony destruction. When they are primary in the clivus, this aggressive appearance suggests a malignant neoplasm. Pituitary adenoma was not considered in the present case until the time of frozen section, where the presence of cohesive, uniform polygonal cells with granular to clear cytoplasm in a trabecular and perivascular pattern was diagnostic of pituitary adenoma. Benign adenomas may occur in the clivus despite the appearance of a radiographically aggressive mass. Careful consideration of histological pattern will ensure accurate frozen section diagnosis and optimal surgical treatment for these patients.

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140 Solitary Fibrous Tumour of the Central Nervous System

Jane Cryan, Michael Jansen. Department of Neuropathology, Joanna Ti. Department of Radiology, Beaumont Hospital, Dublin, Ireland; Patrick Buckley. Department of Cancer Genetics, Royal College of Surgeons in Ireland; Irfan Yousef. Department of Surgery, Clare Faul. Department of Radiation Oncology, Francesca Brett. Department of Neuropathology, Beaumont Hospital, Dublin, Ireland; Ray Stallings. Department of Cancer Genetics, Royal College of Surgeons in Ireland; Michael Farrell. Department of Neuropathology, Beaumont Hospital, Dublin, Ireland

Solitary fibrous tumours (SFT) in the CNS are rare, and like haemangiopericytoma (HPC), with which they share histologic features, are of unknown cellular origin. SFT may have a better prognosis than HPC. We report clinical, radiologic and pathologic features of SFT and HPC of the CNS to illustrate diagnostic and management challenges posed by this spectrum. All cases reported as CNS SFT and HPC over 10 years were reviewed for clinical, radiological and histological features. 16 cases retrieved included 9 SFT and 7 HPC. Ages ranged from 27-81 years at presentation (SFT range 27-58years; HPC range 31-81years). Men outnumbered women in both groups (SFT 7/9 M; HPC 4/7 M). Duration of symptoms ranged from days to in excess of 2years. Tumour location was predominantly supratentorial and superficial but intraventricular, infratentorial and spinal tumours were also present, particularly in the SFT group. Radiologically the tumours mimicked meningiomas and glial tumours. Histologic variability was conspicuous, particularly in the SFT group, and overlapping features between SFT and HPC included hypercellularity, oval-cell morphology, branching HPC-like vessels, whereas a biphasic (hypercellular/hypocellular) pattern was seen more frequently in SFTs (6/9SFT; 1/6HPC). Necrosis was infrequent in both groups except where pre-surgical embolisation was performed. Mitotic count ranged from 1-11/10hpf in SFTs and 1-12/10hpf in HPCs. Similar variation was seen in MIB counts. Histological features did not correlate with clinical outcome. Whole-genome DNA copy number profiling using aCGH displayed a far higher frequency of chromosomal losses than gains in tumours, with no evidence for amplifications of disease related genes. In our experience complete surgical excision is the treatment of choice for SFT. Due to the absence of prognostic pathological features; radiation therapy should be given earlier rather than later if surgery is incomplete. Close clinical follow up is recommended until there is further long term data on behaviour.

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141 Primary Burkitt Lymphoma of CNS: A Case Report and Literature Review

Fahad Alghamdi. Department of Pathology, Christopher Gillis, Brian Toyota. Neurosurgery Department, Bijal Patel, Manraj Heran. Department of Radiology, Vancouver General Hospital, Vancouver, Canada; Stephen Yip. Department of Pathology, Adam Smith. Department of Pathology, Laurie Sehn. BC Cancer Agency, Vancouver, Canada; Kenneth Berean. Department of Pathology, Robert O'Connor. Department of Pathology, G Moore. Department of Pathology, Vancouver General Hospital, Vancouver, Canada

Primary CNS Burkitt lymphoma is a rare occurrence, only 9 cases being reported from 1987 to 2010. We present the case of a 73 year-old right-handed female with a two week history of sudden onset of alexia. Computed tomography (CT) and magnetic resonance (MR) imaging demonstrated a left posterior temporal lesion measuring up to 5.8 cm associated with significant vasogenic edema and mild subfalcine and uncal herniation. Biopsy of the tumor showed characteristic histopathological features of Burkitt lymphoma - diffuse proliferation of medium-sized cells with a starry sky appearance and an angiocentric infiltration pattern. The tumor invaded dura mater, leptomeninges and brain parenchyma. Immunohistochemically, the tumor cells were positive for CD20, CD10, and BCL-6, and negative for BCL-2. The MIB-1 proliferative index was virtually 100%. Fluorescence in situ hybridization showed a MYC (8q24) translocation. Post-surgery assessments excluded other intra- or extracranial tumor sites. In contrast to most primary CNS lymphomas, the patient was immunocompetent and in situ hybridization for Epstein-Barr virus was negative, as has been previously reported in other primary CNS Burkitt lymphomas. Our case, the tenth reported in the literature, is unique in that it is the first to demonstrate contiguous involvement of the meninges and the parenchyma and to document the characteristic MYC translocation.

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142 Giant Cell Tumor of Bone With Intradural Extension

Fahad Bafakih, Vijayalakshmi Rajasekaran, Cara Sedney, Jeffery Hogg, Kymberly Gyure. West Virginia University

Giant cell tumor of bone is a generally benign, locally aggressive lesion that typically occurs in the epiphyses of long bones. The peak incidence is in young adult patients with a slight female predominance. Pain and swelling are the most common presenting symptoms. These lesions rarely involve the skull; when they do, the sphenoid bone is the most common location. We report an unusual case of a giant cell tumor involving the temporal bone with intradural extension. The patient, an 81-year-old woman, presented with an approximately one-year history of right-sided hearing loss and tinnitus. Examination revealed sensorineural hearing loss on this side, and imaging studies revealed an expansile lesion in the right temporal bone as well as a left-sided, extra-axial sphenoid wing lesion thought to be a meningioma. Following a CT-guided biopsy of the right temporal bone lesion, the patient underwent a right frontotemporal craniotomy with resection of a large tumor which involved the right middle cranial and infratemporal fossae with intradural extension. The lesion also involved the middle ear and condylar fossa. Microscopic examination revealed a bone-invasive neoplasm composed of numerous multinucleated giant cells with abundant eosinophilic cytoplasm which were uniformly distributed in a background containing mononuclear cells with similar-appearing nuclei. The adjacent brain tissue exhibited gliosis. Although rare, giant cell tumor should be included in the differential diagnosis of a bony lesion of the skull, and it should be distinguished from other tumors containing giant cells which can occur in this region including aneurysmal bone cyst and giant cell reparative granuloma.

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143 Myoepithelial Neoplasm Arising From Pleomorphic Adenoma of the Lacrimal Gland: A Case Report

Sarah Martin, Andrea Wiens. Department of Pathology, Richard Burgett. Dept. of Ophthalmology, Eyas Hattab. Indiana University School of Medicine, Dept. of Path. and Lab. Med

Myoepithelial neoplasms of the lacrimal gland are exceedingly rare, with only five previously reported cases. We discuss an unusual case of a 40-year-old woman with a past medical history significant only for endometriosis, who presented with proptosis and globe displacement secondary to a large superolateral orbital mass measuring approximately 2.7 cm in greatest dimension. Imaging studies suggested a lacrimal gland origin. The mass was resected. Microscopic examination showed a partially encapsulated lesion composed predominantly of a hypercellular spindle cell proliferation replacing a residual pleomorphic adenoma composed of myxoid stroma intermixed with numerous ducts and myoepithelial cells. Immunohistochemical staining for p63 and calponin demonstrated strong and diffuse immunoreactivity in the spindle cell portion, suggesting a myoepithelial origin. The spindle cell lesion did extend to the inked surgical margins. The differential diagnosis included myoepithelioma and myoepithelial carcinoma ex pleomorphic adenoma. While increased mitotic activity (up to 7 per 10 hpfs) and elevated MIB1 proliferative index (10-15%) were present to support a diagnosis of carcinoma, there was little nuclear atypia and no invasive growth pattern. Therefore, an intermediate and somewhat vague diagnosis of myoepithelial neoplasm ex pleomorphic adenoma was rendered, and complete surgical resection was recommended. The present case is unique in that its histopathological features are intermediate between that of a benign and malignant neoplasm of myoepithelial origin. The true nature of this neoplasm will likely be revealed as time progresses. The appropriate course of treatment and prognosis are therefore uncertain at this time. It is important for surgical neuropathologists to be aware of this rare ocular tumor.

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144 Primary Intracranial Leiomyosarcoma in a Young Adult With Hodgkin's Lymphoma

Hidehiro Takei, Benjamin Ellezam, Andreana Rivera. The Methodist Hospital

Although it is well known that Epstein-Barr virus (EBV) infection has been associated with a heterogeneous group of malignancies including Hodgkin's lymphoma (HL), an association with smooth muscle tumors has recently been described. Primary intracranial EBV-associated leiomyosarcomas (EBV-LMS) are extremely rare, and most of the reported cases were immunocompromised (e.g., HIV positive) and/or pediatric. A neurologically asymptomatic, previously healthy 27-year-old man was found to have a positron emission tomography (PET)-positive lesion in the brain during a staging workup for his recently diagnosed HL. A subsequent MRI revealed a 3×4.5×3.5cm contrast-enhancing mass with marked surrounding edema in the right anterior frontal lobe. He was serologically HIV-negative. He underwent right frontal craniotomy with gross total resection of the tumor. Intraoperatively, the tumor had fairly discrete margins and appeared to arise from the anterior falx (i.e., dural based). Microscopically, the tumor was composed of interlacing fascicles of spindled tumor cells with elongated cigar-shaped nuclei and brisk mitotic activity (up to 23 mitoses/10HPFs). There was multifocal tumor necrosis. Immunohistochemically, the tumor cells were positive for caldesmon and smooth muscle actin, and negative for desmin, CD34, CD99, bcl-2, S-100 protein, and GFAP. A Ki-67 labeling index was up to 30% in most active areas. These histologic features as well as immunohistochemical profile are those of a LMS. EBV-encoded RNA in situ hybridization demonstrated strong diffuse positivity with > 90% of the tumor cells staining. Our case is very unique in that very rare EBV-LMS and HL, which is also known to be EBV-related, were seen in an "immunocompetent" adult patient without significant neurological symptoms. We should be aware of smooth muscle tumors (e.g., LMS) as a differential diagnosis of dural-based spindle cell neoplasm even in an immunocompetent patient.

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145 Intraventricular Collision Tumor of Metastatic Lung Adenocarcinoma to Atypical Meningioma: A Case Report

Keyla Kleyser-Sugrue, Srinivas Mandavilli, Xianyuan Song. Hartford Connecticut

Introduction: Collision tumor (tumor-to-tumor metastasis) is a well known but rare phenomenon. Meningiomas are the most common primary intracranial tumors that host metastases, with lung and breast carcinomas being the most common donor. Most reported cases of meningiomas harboring metastasis are dural-based. Intraventricular meningiomas are rare, comprising about 3-5% of all meningiomas. According to our knowledge, no carcinomas metastatic to intraventricular meningiomas have been reported in the literature. Here we report such a case.

Case report: A 71-year-old female patient with heavy smoking history was found to have multiple bilateral lung masses, as well as adrenal and brain lesions. FNA of the lung confirmed the diagnosis of primary lung adenocarcinoma. MRI of the brain revealed a enhancing mass with surrounding edema involving the atria of left lateral ventricle. The patient underwent craniotomy. Intraoperative frozen section revealed a meningioma and gross total resection of a 6 × 4 × 3.6 cm mass was done. Microscopically the tumor was primarily composed of a psammomatous meningioma arising within the choroid plexus with focal brain invasion (W.H.O Grade II). In one section of the entirely submitted tumor, there was a small focus of carcinoma within the meningioma. Further immunohistochemical studies (CK-7 and TTF-1 positive with CK-20 negative) confirmed this to be metastatic adenocarcinoma from the lung primary to this intraventricular meningioma.

Conclusion: This is the first case report of an intraventricular meningioma with metastatic adenocarcinoma. While such a presentation might be rare, pathologists should be cognizant of the phenomenon of tumor to tumor metastasis (in particular when there is a known history of prior malignancy), since the finding of brain metastasis often has a negative prognostic implication.

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146 Analysis of Invasive and Non-Invasive Meningiomas

Sidney Croul. University of Toronto, Dept. of Lab Medicine and Pathobiology; Fateme Salehi, Shahrzad Jalali, Kelly Burrell, Gelareh Zadeh. Division of Neurosurgery, University Health Network, Toronto, Ontario

Background: Specific clinical subtypes of meningiomas, in particular those which occur at the skull base, with extensive bone invasion and hyperostosis, cause significant morbidity due to their intimate involvement with neural elements and tendency to recur despite surgery and radiotherapy.

Design: Clinical and Pathologic data were collected from 62 patients with skull base meningiomas. A TMA was constructed from their surgical specimens and stained immunohistochemically for osteoponin (OPN), integrin-beta-1 (ITGB1) and matrix metalloproteinase-2 (MMP-2). A subset of 4 invasive and 4 non-invasive meningiomas was further investigated for whole genome RNA expression via the Illumina DASL microchip and selected RNA's via quantitative PCR.

Results: MMP-2, OPN, and ITGB1 stained both tumor cells and vessels. The immunoreactivity for MMP-2 and OPN was largely cytoplasmic while ITGB1 was more restricted to the cell surface. Non-invasive trans basal meningiomas exhibited higher MMP-2 immunoexpression in vascular endothelial cells compared to invasive meningiomas. Tumor cell immunoreactivity was significantly higher in invasive trans basal meningiomas than non invasive tumors. OPN expression was significantly higher in invasive trans basal meningiomas compared to non invasive meningiomas. Although it did not reach statistical significance, ITGB1 expression exhibited a striking concentric periivascular pattern in some meningiomas. RNA was extracted from formalin fixed paraffin embedded meningiomas (4 invasive and 4 non-invasive) and hybridized to the Illumina DASL platform. Significant analysis of micro-arrays was used to identify over 300 RNAs which were under and over expressed. Gene set enrichment analysis was employed using DAVID to identify which biological processes are up regulated or down regulated. Combining the list of differentially expressed genes and the DAVID pathway analysis we chose to focus our next set of studies on MATRILIN 4, ADAM TS4,MMP19, MMP16 and PDGFR alpha Over-expression of the selected candidate genes in bone invasive meningiomas was validated using quantitative PCR.

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147 Simultaneous Intrasellar Meningioma and Pituitary Adenoma Discovered at Autopsy: Case Report and Review of the Literature

Jennifer Ross, Alex John, Glenn Sandberg, Luis Sanchez. Harris County Institute of Forensic Sciences

A 47-year-old white female with a past medical history of congenital adrenal hyperplasia (CAH), back pain, multiple back surgeries, and prescription drug abuse was found dead in her residence. External examination revealed a hirsute, muscular female weighing 129 pounds and measuring 65 inches long (body mass index = 21.5 kilograms per square meter). Internal examination revealed enlarged adrenal glands (right-80 grams and left-150 grams) and a small, relatively well-circumscribed nodule at each end of the 1.5 by 0.5 by 0.5 centimeter pituitary gland. Microscopic examination of the pituitary gland revealed a 0.3 centimeter diameter meningothelial meningioma at one end and a 0.2 centimeter diameter pituitary adenoma at the other. Immunohistochemical stains in the adenoma were diffusely positive for chromogranin and had scattered positivity for follicle stimulating hormone (FSH). Epithelial membrane antigen (EMA) was positive in the meningioma. Microscopic examination of the adrenal glands revealed cortical hyperplasia with proliferation of eosinophilic, lipid-depleted cells intermixed with lipid-laden clear cells and scattered foci of chronic inflammation. The cause of death was accidental combined drug toxicity. We discuss the gross, histopathological, and immunohistochemical findings, and the association with CAH. Patients with CAH have hyperactivity of the hypothalamic-pituitary-adrenal axis and are at risk for pituitary tumor formation, most commonly with ACTH production. There are two previously reported cases of meningioma associated with CAH and 24 cases of purely intrasellar meningiomas. The association between a coexistent meningioma and pituitary adenoma has been reported, with a prevalence of suprasellar, parasellar, and sphenoid wing meningiomas. Growth hormone-producing tumors are the secreting adenomas most commonly found in occurrence with meningiomas. Prior to our case, there are only two reported cases with coexistent tumors located below the diaphragma sella. To our knowledge, this is the first account of a simultaneous intrasellar meningioma and pituitary adenoma in a patient with CAH.

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148 DNET-like Tumor of the Septum Pellucidum: Part of an Expanding Category of Rosette-Forming Glioneuronal Tumor

Matthew Schniederjan, Cristina Vincentelli, Daniel Brat. Emory University School of Medicine, Department of Pathology

DNET-like tumor of the septum pellucidum (DNETSP) comprises a small number of low-grade glioneuronal neoplasms arising from the septum near the foramina of Monro. They tend to present with headaches and obstructive hydrocephalus, or are found incidentally. We present four cases of DNETSP and compare their and other published cases' histologic features with those of two cases of rosette-forming glioneuronal tumor from the fourth ventricle (RGNT). All four DNETSP patients presented with headaches, one of whom also had evidence of obstructive hydrocephalus. All of the lesions arose from the septum pellucidum in the vicinity of the foramina of Monro. All patients underwent gross total tumor resection, with one of them receiving post-operative chemotherapy based on an initial diagnosis of oligodendroglioma. Currently, three of the patients are alive with no evidence of disease and one is alive with stable disease. The histologic features of these lesions included oligodendroglia-like cells in a myxoid background with a smaller component of piloid astrocytes, perivascular pseudorosettes, fibrillar neurocytic rosettes and occasional "floating" neurons. Areas of neuropil within rosettes and pseudorosettes were immunoreactive for synaptophysin. This appearance was highly similar to that of both fourth ventricle RGNTs, except that floating neurons were not identified in the latter. A literature review reveals similar age and sex distributions for RGNT and DNETSP (RGNT: M:F = 1:2, mean age 30.3 years; DNETSP: M:F = 2:3, mean age 25.5 years). Based on the histologic similarity of these two diagnostic categories, their similar age and sex distributions, similarly benign behavior, and their common occurrence in the midline, we suggest that they may represent very closely related lesions and may be in the future, as experience and evidence accumulate, shown to form a single diagnostic entity.

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149 Primary Intracranial Atypical Ewing's Sarcoma-Peripheral Primitive Neuroectodermal Tumor: A Case Report and Review Literature

Jantima Tanboon. Department of Pathology, Faculty of Medicine, Siriraj Hospital; Bunphot Sitthinamsuwan, Tewajetsada Paruang. Division of Neurosurgery, Department of Surgery, Siriraj Hospital; Paul Thorner. Division of Pathology, The Hospital for Sick Children

Ewing's sarcoma-peripheral primitive neuroectodermal tumor(ES-pPNET) is an uncommon extraskeletal tumor. The occurrence of intracranial ES-pPNET is very rare. To date, 16 of 28 cases reported in the literature were confirmed by molecular study. We present the clinical, imaging, and pathologic findings of an ES-pPNET with atypical features in a 22-year-old woman. The patient underwent craniotomy and the tumor was totally excised. The diagnosis was confirmed by detection of t(11;22) translocation in the tumor using the polymerase chain reaction from the formalin fixed paraffin embedded tissue. A rearrangement of the EWS gene was detected by fluorescent in situ hybridization. The patient developed multiple metastases at two months and died at 6 months after the operation.Sponsored by Amyn M Rojiani MD, PhD.

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150 Diffuse Low Grade Leptomeningeal Glio-Neuronal Tumor Not an Oligodendrogliomatosis: Clinic-Pathologic Features of 2 Cases

Sarah Alghamdi. Department of Pathology Mount Sinai Medical Center; Amilcar A. Castellano-Sanchez M.D. Department of Pathology Mount Sinai Medical Center, Department of Pathology and Laboratory Medicine, Miami Children's Hospital:, Carole Brathwaite M.D, Steven Melnick M.D. Ph.D. Department of Pathology and Laboratory Medicine; David Sandberg, M.D., Greg Olavarria M.D., Neurosurgery; Ziad Khatib, M.D, Hematology-Oncology, Miami Children's Hospital Miami, FL; Caterina Giannini, M.D Mayo Clinic, Department of Pathology and Laboratory Medicine, Rochester MN; Peter Burger, MD.. Johns Hopkins Department of Pathology. Baltimore, MD

Few cases of leptomeningeal "oligodendrogliomatosis" (LODM) in children have been reported in literature. These cases describe a diffuse leptomeningeal dissemination of CNS neoplasm that has an indolent course and shows "oligodendroglial" morphology. Here, we report two pediatric cases with similar in clinical and pathologic appearance -as those in the literature - however immunohistochemistry (IHC) and electron microscopy (EM) proved unequivocal neuronal differentiation.

Case 1: a previously healthy, two-and-a-half year old girl presented with a history of headaches, vomiting and abnormal posturing. MRI revealed diffuse leptomeningeal enhancement and meningeal thickening in multiple areas of the brain and spinal cord which subsequently progressed.

Case 2: a six-and-a-half year old boy with Chiari type I malformation and previous TB meningitis presented with vomiting and hydrocephalus. Neuroimaging of the brain and the spinal cord showed diffuse areas of abnormal leptomeningeal enhancement extending along the basilar cisterns. CSF from case 1 showed bland, round-to-polygonal cells with abundant, pale eosinophilic slightly granular cytoplasm and round-to-oval nuclei without nucleoli. Similar cells infiltrating a dense fibrotic, well-vascularized background were noted in the meningeal biopsies on both cases. By IHC, the cells in case 1 were positive for GFAP and negative for epithelial membrane antigen (EMA), Synaptophysin (SYN), Neu-N, CAM5.2, KP-1. Cells in case 2 were SYN and GFAP positive. Both cases have low Mib-1 proliferation rate (<1). EM in case 1 showed distinctive EM features of neuronal differentiation, i.e. numerous cellular projections, neurosecretory granules, microtubular network, and neurofibrillary bodies. Most cases of so called LODM have been diagnosed on the bases of morphologic and IHC findings. Morphologically the neoplastic cells display bland oligodendrocyte-like features. However, our cases showed IHC and EM features characteristic of neuronal differentiation. Hence we strongly believe that the term Diffuse Low Grade Leptomeningeal Glio-Neuronal Tumor (DLGLGNT) best characterizes these neoplasms clinically and pathologically.

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151 Giant Pituitary Adenomas: Pathologic-Radiographic Correlations and Lack of Role for p53 and MIB-1 Labeling

Bette Kleinschmidt-DeMasters, Helen Madsen, Thomas Borges. University of Colorado Anschutz Medical Campus Dept of Neuroradiology; Aaron Knox, Katherine Michaelis, Mei Xu. Dept of Endocrinology; Kevin Lillehei. Dept of Neurosurgery; Margaret Wierman. Dept of Endocrinology, University of Colorado Anschutz Medical Campus

Giant pituitary adenomas, with diameter = 4 cm., were formerly considered rare and not surgically approachable. Few U.S.-based series exist. We reviewed our 10-year experience with these tumors and identified 17 patients, 11 males and 6 females, ages 27-65 years. Twelve of 17 were either gonadotroph or null cell adenomas and 5 were giant prolactinomas. By neuroimaging, all invaded the cavernous sinus(es) and tumors in 13 patients invaded skull base. Despite massive size, only 5 showed apoplectic clinical and neuroimaging features; when present, this feature occasionally prompted preoperative consideration of craniopharyngioma. Transsphenoidal surgical excision was possible in all patients, with 3 undergoing planned second-stage reoperations and 2 requiring a second surgery for recurrence (both at 6 year intervals). Despite the aggressive features of massive size and cavernous sinus invasion, mitotic rates and immunohistochemistry (IHC) labelling for p53 and MIB-1, features alleged to be associated with atypical adenomas, were minimally increased. Absence of a role for p53 and cell cycle markers was further verified on a subset of our cases by microarray analyses. Five giant gonadotroph adenomas were compared to 7 non-aggressive, non-giant gonadotroph cell adenomas and no statistically significant changes in transcript levels of MIB-1 (MKI67) or P53 were observed. Differential gene expression was identified, however, for a number of other genes. In conclusion, most giant pituitary adenomas are gonadotroph cell adenomas or giant prolactinomas. Microarray profiling validates the IHC impression that MIB-1 and p53 IHC do not correlate with aggressive features in the most common type of giant adenoma.

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152 The Role of the Inhibitor of Growth-4 (ING-4), a NF-κB Regulatory Molecule, in Astrocytoma's Cells Invasion

Georgios Klironomos. Department of Neurosurgery&Anatony University of Patras; Spyridon Karadimas. Toronto Western Research Institute Krembil Neuroscience Research Centr; Georgios Gatzounis. Department of Neurosurgery, University of Patras; Eleni Papadaki. Department of Anatomy, University of Patras

Introduction: One of the most prominent characteristics of astrocytomas is their ability to invade adjacent normal brain parenchyma by degrading components of extracellular matrix. In vitro studies have revealed a possible role of tumor suppressor gene ING-4 in NF-κB pathway. These data suggest the implication of ING-4 in NF- κB target genes expression, such as Matrix Metalloproteases-2,-9 (MMP-2, MMP-9) and urokinase- Plasminogen Activator (u-PA). These last enzymes are known to contribute in the extracellular matrix degradation during tumor invasion.

Materials and Methods. In this study the expression pattern of ING-4, NF- κB p65 and p50 subunits, MMP-9, MMP-2 and u-PA were examined in a well characterized set of 101 human astrocytomas of all grades and in normal brain. The study was performed in formalin fixed, paraffin embedded tissue samples using immunohistochemistry. The implication of these molecules in the pathogenesis and progression of astrocytic tumors as well as the correlation of their expression levels with clinicopathologic parameters such as age, sex and tumor grade were investigated.

Results. Normal brain tissue and adjacent to tumors normal parenchyma showed strong nuclear expression of ING-4. The expression levels of this protein were found to be significantly negatively correlated with the tumor grade (p<0.001 for all correlations). For example, grade IV astrocytomas showed significantly lower ING-4 expression than grade III. In contrast, the expression levels of NF-κB, MMP-2, MMP-9 and u-PA were revealed significantly positively correlated with the tumor grade (p<0.001 for all correlations). The expression levels for all proteins did not reveal any significant difference between different age groups and sex.

Conclusion. This study suggests a possible role of ING-4 and NF-κB pathway in the regulation of invasiveness of astrocytomas. Additional studies are required in order to evaluate the precise role of this pathway and reveal potential future therapeutic interventions.

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153 Infantile Hemangiopericytomas in Children

Garth Warren. Oregon Health and Sciences University; Sakir Gultekin. Neuropathology/Surgical pathology at OHSU

Hemangiopericytomas are rare vascular tumors. It is primarily a tumor of adults, with only 10% of cases occurring in children. Tumors occurring in the first year of life are even more rare and are referred to as infantile hemangiopericytomas. We present an 8 month old boy who developed a fever and new onset seizures, and imaging revealed a large intracranial, heterogeneously enhancing, partially cystic mass. He was treated with surgery alone, and the tumor ultimately proved to be a hemangiopericytoma. Although histological features of increased cellularity with pleomorphism, necrosis, hemorrhage, and frequent mitoses are indicative of malignant behavior in adult hemangiopericytomas, the infantile form has been recognized as having a benign course despite having these histological features.

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154 Dura-Based High-Grade Large B-Cell Lymphoma Without Systemic Involvement 4 Years After Diagnosis and Successful Therapy

Sverre Mörk, Ellen Berget, Lars Helgeland. Department of Pathology, Anne Lehmann. Oncology Department, Olav Vintermyr. Department of Pathology, Haukeland University Hospital, Bergen, Norway

A 75-year-old woman presented slightly bewildered after an episode of brief confusion and short loss of consciousness in her home the same morning. Initial brain CT scan showed an intracranial occipital (left) tumor with extracranial extension. A few weeks later an MRI revealed a solid lesion (4.6 × 2.2 × 4.2) cm) with a broad dural base, in the left occipital lobe. Surrounding edema and midline shift signalled expansivity. There was an extracranial tumefaction overlying the occipital lesion. Preoperative diagnosis was a large meningioma. A craniotomy with removal of most of the large intracranial lesion was performed in February 2007. The histopathological evaluation was initially that of a poorly differentiated, somewhat pleomorphic tumor. Mitotic figures were >10/1hpf in hot spots. Immunohistochemistry strongly suggested a B-cell lymphoma; PCR evidenced monoclonality, confirming the diagnosis. The patient was treated with radiation and cisplatin-based chemotherapy. She is subjectively well and without recurrence or other pathology at follow-up 3.5 years after initial diagnosis. The present case demonstrates a high-grade large B-cell lymphoma of the dura with extracranial extension with more than 4 years recurrence-free survival per ultimo February 2011.

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155 Primary Leptomeningeal Sarcomatosis: Case Report With Immunohisotchemical Analysis

Janet McNaughton. Orlando Health; Carlos Pardo. Johns Hopkins University School of Medicine; Gary Pearl. Orlando Health

We report the case of a 56 year old Caucasian male who developed severe headaches and a meningeal syndrome. Brain MRI revealed extensive leptomeningeal enhancement that involved the left optic nerve, peri-insular region, interpeduncular cistern and basilar region. Multiple lumbar punctures and two brain biopsies over the next several months were non-diagnostic, with histology of right brain tissue revealing acute and chronic inflammation. A fluorodeoxiglucose PET scan was negative for signs of systemic inflammatory or neoplastic disease. He deteriorated neurologically with persistence of a chronic meningeal syndrome and transitory ischemic attacks in the territory of the left middle cerebral artery despite aggressive treatment with parenteral steroid therapy for possible neurosarcoidosis. He also received subsequent treatment with antituberculosis medications. Six months after the onset of symptoms, the patient became progressively weaker and required ventriculoperitoneal shunt placement, due to significant hydrocephalus. He developed a seizure disorder followed by respiratory insufficiency and expired. Autopsy, limited to the brain, revealed diffuse leptomeningeal thickening localized mostly in the basilar and interpeduncular regions of the brainstem as well as the left hemisphere. Histologically, a malignant spindle cell neoplasm diffusely infiltrated the leptomeninges with extension into Virchow-Robin spaces. Immunohistochemistry was positive for S100 and negative for smooth muscle actin, CD31, CD34, epithelial membrane antigen, CD45, vimentin, myogenin, keratin CAM5.2, keratin AE1/3, Mart-1, HMB-45, MITF, CD30, ALK-1, GFAP, and p53. The overall findings are most consistent with primary leptomeningeal sarcomatosis (PLS). PLS is a rare disorder defined as a diffuse leptomeningeal sarcoma lacking circumscribed masses. The cell of origin is unknown, although the pattern of immunohistochemical reactivity in our case raises the possibility of a nerve sheath origin.

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156 Primary Intracranial Dural Based Ewing's Sarcoma in an Elderly Woman-Case Report

Knarik Arkun, Patrick Lasala. Neurosurgery; Jack Farinhas. Neuroradiology; Christian Keller. Neuropathology; Karen Weidenheim. Montefiore Medical Center, Neuropathology

Extraskeletal Ewing's sarcoma/PNET is extremely rare in the central neuraxis. Both metastatic and primary tumors are rare in this location, especially in elderly individuals, and may present difficulty in diagnosis. The purpose of this report is to present a rare primary intracranial Ewing's sarcoma/PNET in 69-year-old woman who presented with complains of worsening of headache, history of "meningioma" resection and a negative PET scan for neoplasms at other sites. Imaging showed an exraaxial 5 × 6 cm, lobulated, homogeneously enhancing mass filling the entire middle cranial fossa on the right. Subtotal resection of the mass was performed, revealing a friable, hemorrhagic tumor. Frozen and permanent sections showed a poorly differentiated tumor arranged in sheets of uniform small cells with numerous mitoses and ill defined cell borders. Small amounts of eosinophilic cytoplasm contained clear vacuoles. Nuclei were irregular with fine chromatin and prominent nucleoli. The tumor showed strong, diffuse immunostaining for CD99 and vimentin, but was negative using antibodies against GFAP, cytokeratins, CD45, EMA, chromogranin A, synaptophysin, CD34, SMA, desmin, myogenin and MYOD-1. FISH analysis showed rearrangement of the EWS locus on chromosome 22q12, characteristic of ES-PNET. The patient underwent resection, radiation and chemotherapy and five months after resection, there was no evidence of residual tumor in the brain. While meningiomas and metastases are by far the more common entities of dural based mass lesions in this age group, it is important to keep unusual neoplasms in the differential diagnosis of dural based tumors to avoid undertreatment of possibly biologically more aggressive tumors.

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157 Granular Cell Tumor of the Trigeminal Nerve

Dimitri Trembath, Michael Solle. Radiology, Adam Zanation. Radiology, Otolaryngology, University of North Carolina, Chapel Hill, NC

Central nervous system (CNS) granular cell tumors are extremely rare, generally occurring in the infundibulum of the pituitary gland. Cranial nerve involvement is extremely unusual with three reported cases involving the trigeminal nerve. We report a fourth case of a granular cell tumor of the trigeminal nerve that was initially considered a schwannoma based on clinical presentation and radiology. The patient is a 39 year-old female who initially presented in 2003 with a complaint of trigeminal neuralgia. Anti-migraine therapy was unsuccessful and the patient underwent decompression surgery that only temporarily relieved the pain; gamma knife radiation therapy was also unsuccessful. MRI demonstrated an enhancing, T1 and T2 isointense, fusiform soft tissue mass expanding the right foramen rotundum and pteryogopalatine fossa, and extending proximally to Meckel's cave and distally into the inferior orbital fissure. CT imaging demonstrated no evidence of osseous dehiscence. The working diagnosis was schwannoma. The patient underwent a right infratemporal fossa tumor resection with complete removal of the tumor. The patient had an uneventful post-operative course with resolution of her trigeminal pain. Gross examination demonstrated 2.8 cm of pink/tan soft tissue. Microscopic examination demonstrated cells with round, slightly eccentric nuclei surrounded by copious amounts of eosinophilic granular cytoplasm focally positive for PAS. No Antoni A or Antoni B areas were identified. No significant nuclear atypia or mitotic activity was seen. Immunohistochemical stains demonstrated that the tumor cells were diffusely positive for S-100, inhibin and CD68. The tumor cells were negative for neurofilament, GFAP and cytokeratin AE1/AE3. The final diagnosis was granular cell tumor. Granular cell tumors are generally benign neoplasms with a predilection for arising in the oral cavity. CNS involvement usually involves the infundibulum; however they can arise in cranial nerves and should be considered in the differential diagnosis of cranial nerve neoplasms.

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158 Spinal Neurocytoma in a 6-Year-Old Boy Presenting With Hydrocephalus and Diffuse Leptomeningeal Dissemination

Andrea Wiens, Aaron Kamer. Dept. of Radiology; Joel Boaz. Dept. of Neurosurgery; Jose Bonnin. Indiana University School of Medicine, Dept. of Pathology

Extraventricular neurocytomas are exceedingly rare, and only 16 cases have been reported in the spinal cord. Two of such cases occurred in children. In the spinal cord, neurocytomas generally have a good prognosis and do not require post-operative adjuvant therapy. We report a case of spinal cord neurocytoma in a young boy who presented with progressive headaches, nausea, vomiting, fever, and gait disturbances at six years of age. Magnetic resonance imaging (MRI) demonstrated communicating hydrocephalus and a partially cystic and solid intramedullary mass at C3-C4 with a small associated nodule of enhancement. A ventriculoperitoneal shunt was placed to relieve the hydrocephalus, but he developed a blockage requiring shunt revision three months later. At that time, chocolate-brown fluid suggestive of previous hemorrhage was seen in the catheter. An MRI scan two months later again showed the cervical cord lesion as well as diffuse leptomeningeal enhancement over the cerebral convexities and along the cervico-thoracic spinal cord. Pathologic examination of meningeal biopsies from L5-S1 demonstrated a relatively uniform population of tumor cells with round nuclei, sometimes with perinuclear halos, in a delicate fibrillary background. The tumor showed clusters of hyperplastic blood vessels and sparse mitotic activity. Immunohistochemically, tumor cells were positive for synaptophysin and negative for glial fibrillary acidic protein. In the absence of any evidence of an intracranial tumor to suggest metastatic dissemination to the spinal canal, it is felt that this tumor represents a primary neurocytoma arising in the spinal cord. This is the youngest patient reported to date, and he had a unique clinical presentation with hydrocephalus and diffuse leptomeningeal involvement.

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159 Diffusely Infiltrative Plexiform Schwannoma in Two Patients With Neurofibromatosis Type 2

Christian Davidson, Scott Plotkin, Anat Stemmer-Rachamimov. Massachusetts General Hospital

Introduction: Plexiform schwannomas are a histologic variant of schwannoma with a predilection for patients with neurofibromatosis type 2 (NF2). Herein we describe an unusual infiltrative pattern of growth in two plexiform schwannomas with otherwise conventional histology.

Clinical presentation: Case 1: A 23 year-old woman with NF2 had two scalp lesions, clinically presumed to be neurofibromas.

Case 2: A 34 year-old man with NF2 underwent right orbital exenteration for a large multilobulated mass in the right orbit extending subcutaneously into the forehead (presumed to be meningioma). Neuropathologic examination of the scalp lesions (case 1) and the orbital mass (case 2) showed plexiform schwannomas with a diffusely infiltrative growth pattern.

Pathology: The tumors showed classic schwannoma histology with Antoni A/B areas, and Verocay bodies. Tumor cells were bland spindled cells, diffusely positive for S100. In both tumors there was diffuse invasion into adjacent subcutaneous fibroadipose tissue, with tumor cells dissecting between collagen bundles and surrounding adnexal structures. Tumor cells infiltrated as single cells and as small nests. Immunohistochemistry for EMA, Glut-1, and claudin-1 showed breaching of the perineurium in both tumors with extrafascicular tumor invasion, in a pattern similar to that seen in plexiform neurofibromas. Immunohistochemistry for c-kit highlighted numerous mast cells in the peripheral (infiltrative) parts of each tumor.

Conclusions: We report an unusual infiltrative growth pattern in otherwise conventional schwannomas in two NF2 patients. These tumors show features usually seen with plexiform neurofibromas and may exhibit other unique molecular or biologic characteristics. Infiltrative schwannomas are a prominent feature in NF2 mouse models in which malignant transformation may be seen.

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160 Papillary Tumor of Pineal Region (PTPR): Clinico-Pathological Findings and Follow-Up in Three Pediatric Cases

Fabiana Lubieniecki, Silvana Sandrone. Pediatric Hospital JP Garrahan, Pathology Department. Argentina; Daniel Alderete. Pediatric Hospital JP Garrahan. Oncology Department. Argentina; Ana Lia Taratuto. Pediatric Hospital, Pathology Department; FLENI. Argentina

Papillary tumor of the pineal region (PTPR) was recognized as a Grade II/III entity in the 2007 WHO CNS Classification. It is considered a rare tumor of uncertain prognosis mainly described in adults. We report clinical, pathological findings and follow-up in three pediatric patients, accounting for 5.5% (3/57) of the pineal tumors studied between 1988 and 2009.

Case 1: 4-year-old girl with expanding pineal lesion. Biopsy (2002): small papillary tumor with slight GFAP positivity. Subtotal resection and radiotherapy were performed. Local relapse and metastasis (MTS) were twice removed (2007, 2009). PTPR was diagnosed. Radiosurgery and chemotherapy were performed. Patient died 1 year later.

Case 2: 12-year-old boy with intracranial hypertension and visual disturbances. In 2003 neoplastic cells in cytologic studies were disclosed. Local radiotherapy was achieved. 2006: relapse and MTS in posterior fossa. PTPR was diagnosed. Radiosurgery in metastatic tumors was performed. After progression, chemotherapy was done with complete remission.

Case 3: 8-year-old girl with an expanding pineal lesion. 1999: papillary pineocytoma was diagnosed. MRI (2001): cystic lesion with annular enhancement. Diagnosis: PTPR. Patient underwent radiotherapy. Last evaluation: stable pineal residual tumor. Tumor revision disclosed an epithelial aspect with papillary and solid areas, perivascular pseudorosettes, isolated rosettes, focal necrosis, and mitosis. Immunohistochemistry: Vim (3/3 cases), focal GFAP (1/3), slight Syn(2/3), intense and diffuse AE1AE3 (3/3), diffuse CK18 (2/2), focal S100 (2/2). NF and EMA negative (3/3). Ki67 index: 5 (case 2), 7 (case 3) and 15% (case 1). In the past, PTPR was probably misdiagnosed as papillary ependymoma, papillary pineocytoma, or choroid plexus papilloma. Papillary morphology and immunohistochemistry profile support this diagnosis. The outcome is uncertain, with local progression and isolated cases reported with CSF dissemination, suggesting malignancy and the need for adjuvant radiotherapy. PTPR should be considered in the differential diagnosis of pineal tumors even in children.

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161 Central Nervous System Involvement of Inflammatory Myofibroblastic Tumor (IMT) in Two Children

Fabiana Lubieniecki, Silvana Sandrone, Jessica Lopez Marti, Dora Diaz. Pediatric Hospital JP Garrahan, Pathology Department. Argentina; Javier Gonzalez Ramos. Pediatric Hospital JP Garrahan, Neurosurgery Department. Argentina; Maria Teresa Garcia de Davila. Pediatric Hospital JP Garrahan, Pathology Department. Argentina

We present 2 unusual cases from two boys with nodular intracranial lesions diagnosed as IMT, accounting for 0,09% (2/2046) of the CNS tumors collected in the pathology service in the last 20 years. Routine techniques, immunohistochemistry, and detection of the rearrangement of the ALK gene (t (2;5) by FISH were performed.

Case 1: 2-year-old boy with a seizure showing in MRI studies multiple cortico-subcortical nodules with peripheral edema interpreted as an abscess. He had a 1-year history of neumonectomy with diagnosis of IMT with cardiac involvement. Because of progressive growth and the presence of new lesions on consecutive studies, four resections of the intracranial masses were performed.

Case 2: 4-year-old boy with a 1-month history of focal seizures. A CT scan showed a hyperdense left parietal lesion with contrast enhancement and edema, and a calcified lesion in the left lung hilus. There was a familial history of tuberculosis. The intracranial nodule interpreted as tuberculoma was resected. Diagnosis of IMT was made. After that, the lung lesion was removed with the same diagnosis. The multiple intracranial nodules of both cases showed similar macro and microscopic features. The growth pattern was infiltrative with nests of nervous tissue intermixed with tumor. It was composed of a fusocellular component, focal hyaline aspect, mixed inflammatory infiltrates, with a variable amount in different areas. The immunohistochemistry was positive with vimentin, actin and focally with desmin. ALK expression was positive in one out 2 of the CNS tumors and negative in lung tissue (2/2). The rearrangement of the ALK gene by FISH was positive in both CNS tumors and in one lung lesion. IMT arising from the CNS is quite rare, with isolated reports of synchronic or metachronic forms. Detection of the rearrangement of the ALK gene supports the hypothesis of the neoplastic nature of this entity.

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162 Primary Central Nervous System Lymphoma in an Adolescent: A Rare Presentation of Acquired Immunodeficiency Syndrome (AIDS)

Juanita Evans. Dept of Pathology, Robert Greiner. Dept of Pediatrics, Mark Iantosca. Dept of Neurosurgery, Charles Specht. Penn State - Hershey Medical Center

A 13 year old female presented with left sided seizures. She had a history of six to eight months of sinopulmonary infections, shingles, and intermittent confusion. Magnetic resonance imaging showed a single midline bilobed lesion based in the third ventricle and body of the corpus callosum. Histopathologic evaluation after craniotomy resection revealed a poorly differentiated neoplasm composed of pleomorphic lymphocytes associated with necrosis and hemorrhage. Via immunohistochemistry, the lymphocytes were positive for CD20 and BCL2; consistent with a diffuse large B-cell lymphoma (DLBCL). Furthermore, the lymphocytes were Epstein Barr virus (EBV) RNA positive via in-situ immunohybridization. The lymphoma was isolated to the central nervous system (CNS). Given the rare nature of primary CNS lymphoma in children a search for an underlying immunodeficiency was undertaken and discovered the patient to be human immunodeficiency virus (HIV) positive. Thus, the patient's first presentation of HIV infection was an acquired immunodeficiency syndrome (AIDS) defining malignancy. Further review revealed that both parents were HIV positive with perinatal transmission being the source of her infection. After several cycles of chemotherapy, the lymphoma is in remission eight months after her initial diagnosis. Less than 1% of primary CNS lymphomas occur in children, and initial presentation of HIV/AIDS in a child as a primary CNS lymphoma is rare. Immunodeficiency is a known risk factor for the development of these tumors and is typically associated with EBV positivity. For many children with perinatally acquired HIV disease progression is slow with symptoms not appearing until 8-10 years of age. This patient is uncommon in that her syndrome did not present until early adolescence. Although the incidence of HIV in adolescents in the United States is extremely low, the presence of primary CNS lymphoma in a previously healthy child warrants further work-up to rule out HIV amongst other causes of immunodeficiency.

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163 Hybrid Schwannoma/Perineurioma of the VIII Cranial Nerve: Case Report

Facundo Las Heras. University of Chile Clinical Hospital; Anat Stemmer-Rachamimov. Massachusetts General Hospital

Background: Benign peripheral nerve sheath tumors are classically divided into schwannomas, neurofibromas and perineuriomas; each characterized by a unique histological and immunohistochemical profile. In recent years tumors with hybrid features, composed of multiple, discrete areas of different histological types, were described. These tumors may represent a diagnostic challenge, and their pathogenesis is unclear. We present the first case report, to our knowledge, of a hybrid schwannoma/perineurioma tumor in a cranial nerve.

Clinical presentation: 24-year-old woman with multiple sclerosis was found to have a left internal auditory canal mass in a follow-up MRI. She had normal hearing, normal balance and no facial dysfunction. MRI revealed a 1.3 cm TV × 0.7 cm AP T2 isointense lesion within, and possibly minimally expanding, the left auditory canal.

Pathology: The lesion was tan-white, nodular, well circumscribed and firm. Histologic examination demonstrated a well demarcated, cellular, solid neoplasm with a biphasic pattern. Most of the tumor was composed of spindle cells arranged in fascicles with focal Verocay body formation and diffuse S100 positivity; consistent with a conventional schwannoma. A second, minor area showed concentric proliferation of neoplastic spindle cells around one or more axons (highlighted with neurofilament immunostaining). Tumor cells in this area were positive for perineurial markers, claudin-1 and Glut-1, and focally immunopositive for CD34; consistent with a perineurioma.

Conclusion: We present a case of a benign peripheral nerve sheath tumor with histological and immunohistochemical features consistent with a dual pattern of differentiation of schwannoma and perineurioma, in the VIIIth cranial nerve. Hybrid schwannoma/perineurioma tumors usually arise in the dermis and subcutaneous tissue. This is, to our knowledge, the first case of hybrid perineurioma/schwannoma reported in a cranial nerve.

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164 Primary Intracerebral Histiocytic Sarcoma in a 53-year Old Woman

Cristina Vincentelli, Matthew Schniederjan, Stephen Hunter. Emory University

Histiocytic sarcoma (HS) is a rare and aggressive malignancy that shows histologic and immunophenotypic features of mature histiocytes. Most cases present as a single mass, usually in an extranodal site such as the skin, soft tissue or intestine. To our knowledge, only 6 cases of true primary HS of the central nervous system have been reported. Three of these cases were reported in adults, and three were reported in children. Amongst the cases reported in adults, only one presented as an intra-axial lesion. HS of the CNS is poorly responsive to therapy with an estimated median survival of 4.5 months. The diagnosis is based on the histologic and immunophenotypic features of the neoplasm. We present a case of a primary HS of the CNS in a 53-year-old woman with two intraparenchymal masses, the largest located in the right frontal lobe measuring 4.7 cm. Histologic examination showed a highly cellular malignant neoplasm with extensive necrosis and numerous atypical mitotic figures. The malignant cells were large, pleomorphic and discohesive with some cytoplasmic vacuoles and spindle cells. Nuclei were irregularly folded and eccentrically located with numerous wreath-like and multinucleated forms. An inflammatory infiltrate composed predominantly of mature T-lymphocytes was present within the neoplasm. Immunohistochemistry revealed tumor expression of histiocytic markers CD68, HAM56 and lysozyme. Markers of other hematolymphoid cell lineages, including lymphocytic, myeloid, Langerhans cell and other dendritic cells were negative. There was no immunoreactivity for CD30 or ALK1, ruling out the possibility of anaplastic large cell lymphoma. Epithelial and melanoma markers were also negative. GFAP was positive in a few entrapped reactive astrocytes. Despite whole brain and spine radiation, a PET-CT performed 4 months after surgery revealed widespread metastatic disease.

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165 Embryonal Tumor With Abundant Neuropil and True Rosettes (ETANTR)

Laura Denham, Alexander Zouros. Loma Linda University Medical Center; Arie Perry. University of California San Francisco; Ravi Raghavan. Loma Linda University Medical Center

Introduction: We present a primitive embryonal tumor with features consistent with the rare entity, ETANTR.

Case history: A 2 year old male presented with nausea and vomiting, and a 7.6 cm cm heterogeneous, circumscribed right supratentorial mass. No significant enhancement was noted on MRI, but areas of cystic degeneration were seen. MR spectroscopy suggested a rapidly growing, aggressive tumor, and diffusion restriction suggested hypercellularity.

Results: Sections showed a neoplasm of variable cellularity composed of small blue round cells with scant cytoplasm, forming diffuse sheets and small clusters of well-defined `layered' rosettes, against a background of less cellular neuroglial tissue. Immunohistochemical stains revealed no convincing glial differentiation within the small blue round cells or rosettes, but there was evidence of synaptophysin reactivity. Rare Neu-N reactive cells, and focal weak membrane reactivity for CD99 were also seen. The intervening, low cellularity areas were weakly immunoreactive for GFAP, synaptophysin and NFP. High Ki-67 labeling indices were seen highlighting the rosetted areas, but the intervening low grade neuropil showed no nuclear labeling. Flourescent in situ hybridization (FISH) performed using probes against 19p13.3 and 19q13.4 showed normal dosages of chromosome 19. Following surgery and high-dose chemotherapy, patient had a recurrence, but has been clinically stable following radiation, one year after diagnosis.

Discussion: The findings are consistent with an embryonal tumor with abundant neuropil and true rosettes (ETANTR). This is a high grade neoplasm some features overlapping with ependymoblastomas, but is now considered to be morphologically and immunohistochemically more distinct. Less than 50 cases have been reported. A region of gene amplification has been described for chromosome 19q in some cases. In spite of its known dismal prognosis, at least one patient with prolonged survival (7 years) has been documented. Current literature will be reviewed.

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166 Two Cases of Atypical Teratoid Rhabdoid Tumor of the Spinal Cord

Ethan Stolzenberg, Matthew Cykowski, Ryan Rahhal, Timothy Mapstone, Mary Gumerlock, Kalliopi Petropoulou. University of Pittsburgh Medical Center; Kar-Ming Fung. University of Oklahoma Health Sciences Center

Atypical teratoid rhabdoid tumor (AT/RT) is a malignant embryonal tumor typically seen in infants and young children. Although the vast majority of cases are seen in the posterior fossa and cerebral hemispheres, rare cases have been reported in the spinal cord. We report two cases of AT/RT involving the spinal cord that occurred at our institution, one in a 3 year 10 month old female and the second in a 6 year 8 month old male. The first tumor was an intradural mass extending from L1 to L3 while the second tumor was an intradural mass extending from L4 to S1. The histological and immunohistochemical characteristics of the tumors were variable. Histologically, the first tumor was composed of large pleomorphic cells with prominent nucleoli, abundant eosinophilic cytoplasm and occasional cytoplasmic inclusions. The second tumor had a biphasic appearance with a portion of the tumor comprised of cells with a "small blue cell" cytologic appearance with little to moderate eosinophilic cytoplasm, while other areas contained cells with enlarged vesicular nuclei with prominent nucleoli. Immunohistochemically, both tumors were positive for vimentin and negative for desmin, synaptophysin, chromogranin, and LCA. The tumors differed in their staining characteristics for EMA (focally positive versus rare membranous positive), GFAP (negative versus positive), neurofilament (rare positive versus positive) and smooth muscle actin (negative versus positive). Immunohistochemistry for INI-1 was performed on the first tumor and tumor cells were negative; the second tumor had a 22q deletion involving the BCR region, consistent with a diagnosis of AT/RT. These cases add to the few published cases of spinal AT/RT and may help provide a better understanding of the pathologic and immunohistochemical characteristics of this tumor. Although rare, AT/RT should be part of the differential diagnosis of high grade or small blue cell tumors of the spinal cord.

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167 Metastatic Hepatocellular Carcinoma to Brain and Skull

Yuan Shan, Jesse Kresak. Moffitt Cancer Center

A 57-year-old Caucasian male presented with visual disturbances, proptosis and two rapidly enlarging scalp masses located in the left parietal area and left periorbital frontal bone. A skeletal survey revealed lytic lesions in these regions with a negative workup for multiple myeloma. Ophthalmic examination revealed an orbital tumor prompting a neurosurgical consultation. The histological diagnosis obtained from a craniotomy biopsy was metastatic hepatocellular carcinoma. Subsequent imaging studies confirmed infiltrative tumor throughout the liver with metastatic disease of numerous bones, a lung, retroperitoneal lymph nodes, and right adrenal gland. Calvarial metastasis of hepatocellular carcinoma, prior to the diagnosis of the primary tumor, and without known liver disease is a rare presentation, especially in the United States. However, with evolving treatment modalities leading to increased patient survival times, distant metastases are on the rise. Thus, a broader differential diagnosis, including hepatocellular carcinoma, should be considered when faced with a cranial mass.

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168 Angiofibromyxoma of the Falx Cerebri

Rolf Pfannl, Safain Mina, James Kryzanski. Tufts Medical Center, Boston, MA

Myxomatous and angiofibromyxomatous tumors of the falx cerebri and skull base have been previously reported and are extremely rare in the nervous system. We report a 45-year-old woman with a history of headaches and a 2.5cm well-defined, hypodense, extraaxial tumor arising in the anterior falx cerebri. The tumor was completely excised via bifrontal craniotomy and did not adhere to the adjacent meninges. The post-operative course was unremarkable. The tumor was relatively hypocellular and was composed of bland, small, spindle and stellate-shaped cells embedded in a myxoid stroma with prominent thin-walled blood vessels. Scattered thin bundles of collagen were present in the tumor. Tumor cells stained diffusely for Vimentin and were negative for S100, CD34, SMA, Desmin, Pankeratin, EMA and GFAP.

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169 Inflammatory Cerebral Amyloid Angiopathy in a Cognitively Intact Young Subject

Miguel Riudavets. FLENI; Naomi Arakaki, Marcelo Schultz, Ana Lia Taratuto, Gustavo Sevlever. Department of Neuropathology, FLENI. Buenos Aires. Argentina

On one hand, even though the cerebral amyloid angiopathy (CAA) associated with ß-amyloid could be a part of the Alzheimer's disease spectrum, it can present as an acute lobar hemorrhage (intraparenchymal hemorrhage) usually in patients older than 45 years of age. On the other hand, the vasculitis of the Central Nervous System (CNS) are a group of entities that can have diverse clinical patterns depending mainly on the vascular compartment involved, and the location. The association between these two pathologies is not frequent and has been shown in subjects older than 75 years of age. We present a 36 year old subject who, in september 2009, consulted for headache and vomiting. MRI: parietal lesion, hypointense on T1, hyperintense on T2 and FLAIR without mass effect. Starts treatment with steroids and TBC drugs. November 2009: presents with febrile syndrome. Negative PCR for several microorganisms. ADEM is considered as the main diagnosis. Methylprednisolone is administered. December 2009: immunoglobulins are added and a pulse of solumedrol is given. January 2010: presents with headache and cognitive impairment. Dies few days later. The necropsy was performed. The neuropathological assessment demonstrated a necrotizing cerebral lesion in the parietal region with vasculitis, presence of multinuclead giant cells, and thickened and hypereosinophilic leptomeninges and blood vessels (both intraparenchymal and leptomeningeal).The blood vessels and leptomeninges in that area were positive for ß-amyloid immunohistochemistry The case was interpreted as Inflammatory Amyloid Angiopathy and leptomeningeal amyloidosis with inflammation.

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170 Brain Biopsy of Susac's Syndrome: A Case Report

Howard Chang. Pathology, Sparrow Health System

Susac's Syndrome (SS) is an unusual autoimmune disease affecting mainly young to middle-age women. It is diagnosed clinically based on the triad of encephalopathy, branch retinal artery occlusion (BRAO), and hearing loss. Very few brain biopsies from SS patients have been performed or reported. However, when only encephalopathy is evident initially, the differential diagnoses are broad, including demyelinating diseases, infections, lymphoma, and vasculitis. The brain biopsy findings of a SS patient are presented here. A 40-years-old female was treated with azithromycin for flu-like symptoms diagnosed as otitis media. She developed a severe headache several days later and was admitted to hospital with headache and altered mental status. CSF showed elevated protein and WBC. CSF cultures were negative. MRI and CT showed features consistent with meningovasculitis with areas of punctuate infarctions scattered throughout the cerebral and cerebellar hemispheres. Her symptoms stabilized during several weeks on steroids but deteriorated upon tapering of steroids. To rule out vasculitis or atypical infections, brain (cerebellum) and meningeal biopsies were done at about 10 weeks after initial hospital admission. These showed nonspecific focal neuronal loss with accompanying microglial proliferation consistent with subacute/old microscopic infarcts, and focal perivascular lymphocytes in the leptomeninges. Subsequent physical examinations revealed focal BRAO and hearing loss. These clinical features together with the imaging and pathological findings were consistent with Susac's Syndrome. The patient was treated with long-term immunosuppressive therapies and her condition had stabilized. This case illustrates that even though the brain biopsy findings were nonspecific, they were useful to rule out infections, neoplasms, as well as classic vasculitis. Susac's Syndrome is now considered as an autoimmune endotheliopathy affecting the precapillary arterioles of the brain, retina, and inner ear (Rennebohm et al., 2010). Further studies are needed to determine the underlying pathogenetic mechanisms of this disease.

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171 Joubert Syndrome: Neuropathological Update

Gordana Juric-Sekhar. Department of Pathology, Daniel Doherty. Department of Pediatrics, Robert Hevner. Center for Integrative Brain Research, University of Washington

Joubert syndrome (JS) is a rare, primarily autosomal recessive condition characterized by specific malformations of the mid-hindbrain and a broad spectrum of other phenotypic findings caused by defects in the structure and/or function of the primary cilium/basal body organelles. Ten causative genes have been identified, but they account for <50% cases. We report a series of postmortem neuropathological examinations in 5 subjects with JS, 2 with mutations in the OFD1 gene. In addition to HE stains, sections were studied by immunohistochemistry to detect GFAP, myelin basic protein, neurofilament, synaptohysin, calretinin, and MAP2. The most common findings included vermis aplasia, hypoplastic cerebellum, fragmentation of the dentate nucleus, and cerebellar heterotopia. Other features included hypoplastic inferior olivary nuclei, abnormal decussation of the cortico-spinal tract and the superior cerebellar peduncle, fragmented spinal nucleus of V (trigeminal), abnormal arcuate nucleus, decreased celullarity of the reticular formation and basis pontis. In 3 subjects, an accumulation of neural tissue on the dorsal aspect of the caudal medulla showed peculiar dysplastic structure, consisting of disorganized neuronal and glial cells and bundles of axonal fibers. Similar findings were noted in the area of the dorsal funiculi of the cervical spinal cord. This study indicates that CNS abnormalities in JS are heterogeneous and appear to involve multiple developmental mechanisms including patterning, proliferation, cell migration and axon guidance. These abnormalities provide clues to the underlying molecular and developmental mechanisms causing the brain malformations seen in JS.

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172 Neuropathological Homology in True Galloway Mowat Syndrome

Julia Keith. Sunnybrook Health Sciences Centre, University of Toronto; Victoria Fabian. Royal Perth Hospital, Department of Anatomical Pathology; Peter Walsh. Neurology Department, Princess Margaret Hospital for Children, Perth; Adrian Charles. Pathology Department, Princess Margaret Hospital for Children, Perth; Yves Robitaille. Department of Pathology, CHU Sainte-Justine, Universite de Montreal

Galloway Mowat Syndrome is a rare condition that is likely hereditary though the underlying offending gene has not been identified. It is characterized by microcephaly and severe nephrotic syndrome culminating in childhood death. Some of the reported cases have abnormalities in neuronal migration and intractable seizures, but many of the described cases focus on the renal pathology and emphasize a diversity of clinical and neuropathological features, with only 5 reported cases having a thorough neuropathological description. The case described herein is that of an infant with microcephaly, seizures and nephrotic syndrome who lived 11 months. The neuropathology includes extreme microcephaly and disordered neuronal migration including an abnormal cortical ribbon, periventricular nodular heterotopia, absent dentate gyri and aplasia of the cerebellar granular layer. When the neuroradiology and neuropathology of the previously published cases of combined microcephaly and nephrotic syndrome is scrutinized (N of 40) two distinct phenotypes emerge: those with disordered neuronal migration, whose renal disease tends to be worse (true Galloway Mowat Syndrome), and those without disordered neuronal migration, who have a better prognosis. Thorough neuropathological description of cases from the former category has yielded some consistent findings, namely abnormal gyral patterns, disordered cortical lamination with frequent abnormalities in the second cortical layer, absent dentate gyri, and cerebellar hypoplasia with aplasia of the granular layer.

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173 A Six-Week Old Male With Haddad Syndrome: Clinical, Genetic, and Pathological Evaluation

José Otero, Olivier Danhaive, Maria Cilio, Eric Huang, David Rowitch. UCSF

We present the clinical and neuropathological findings of a 6-week-old male with Congenital Central Hypovention Syndrome (CCHS) with severe intestinal aganglianosis (Haddad Syndrome). The CCHS proband showed increasing hypercapnia that was worse during non-REM sleep. Genetic evaluation of this patient showed a frameshift mutation in PHOX2B (Genbank accession number NG_008243) caused by deletion of 8 nucleotides in exon 3 (2911-GGCCCGGG-2918). This frameshift mutation causes a loss of a 20-alanine repeat present in C-terminus of PHOX2B protein. Intestinal aganglianosis was present from 10 cm distal to the Ligament of Treitz to the rectum. No gross abnormalities were noted on external evaluation of the brain. Hypoplasia of the Locus Ceruleus and Median Raphe nuclei were observed in the CCHS proband. No significant pathological abnormalities were noted in the solitary tract nucleus or arcuate nucleus of the CCHS proband. Reevaluation of the CCHS proband's half-sibling that died of Sudden Infant Death Syndrome with co-sleeping showed marked reactive astrogliosis in Pons and Medulla, with a normal Locus Ceruleus.

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174 Asperger's Syndrome With Unusual Cerebral Pathology: Case Report and Literature Review

Liqiong Liu. Louisiana State University Health Science Center; Van Vo. Department of Pathology, Marcus Ware. Department of Neurosurgery, Zhenggang Xiong. Section of Neuropathology, Department of Pathology, Tulane University

Asperger's syndrome is characterized by impaired social communication with normal language skills and intelligence. Although this syndrome is not uncommon in the childhood, the information about neuropathological changes are very limited, which has been barring the understanding of the etiology and pathogenesis of this disorder. Here, we report a case of Asperger's syndrome with unusual cerebral pathological changes. A twenty-two year-old male with history of Asperger's syndrome presented with chronic epilepsy for the past two years. The diagnosis of Asperger's syndrome was made when the patient was eight years old. For the recent two years, the patient also developed episodes of unusual movements, staring spells and headaches. Neuroradiological examinations showed a focal lesion with hyperintensity and contrast-nonenhancing superficially located in the left lateral frontal lobe. Microscopic examination and immunohistochemical studies identified cortical laminar disorganization and scattered dysmorphic cortical neurons as well as scattered dysmorphic or enlarged heterotopic neurons in the subcortical white matter. The only neuropathologic findings that have been previously reported in Asperger's syndrome are the cerebral cortical minicolumnar abnormality and macroscopic gyrial malformation in very few patients. Our case demonstrates different cerebral pathologic abnormalities and expands our neuropathological view into the Asperger's syndrome. The possible etiology and pathogenesis are discussed.

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175 Infantile Sialic Acid Storage Disease (ISSD): Neuropathology of Two Inuit Patients With the Novel c.526-2a>G Mutation

Jean Michaud, Matthew Lines, Michael Geraghty. Children's Hospital of Eastern Ontario and University of Ottawa; Berivan Baskin. Hospital for Sick Children and University of Toronto; David Grynspan, Elizabeth Nizalik. Children's Hospital of Eastern Ontario and University of Ottawa

Infantile sialic acid storage disease is the most severe expression of the lysosomal free sialic acid storage disorders characterized by the accumulation of free sialic acid within the lysosomes of essentially all tissues. We report the neuropathology findings of two Inuit patients with a novel homozygous change in the SLC17A5 (sialin) gene; c.526-2A>G. This splice site mutation introduces a deletion in the mRNA resulting in a frameshift and premature termination of the message. This confirms the pathogenicity of this mutation. Patient 1 was a male born prematurely at 27 week gestation who died at 35 minutes with features of prematurity, hydrops fetalis and pulmonary hypoplasia. Patient 2 was a 26-day-old male born prematurely at 31 weeks with a clinical course dominated by complications of prematurity, ascites and hepatomegaly. The neuropathology findings of both cases are remarkably similar. The histology and selected ultrastructural studies shows features of storage involving all cell types, in the central (CNS) and peripheral nervous system. The storage material is in large single membrane-bound clear vacuoles. There is also a significant number of axonal swellings involving all structures of the CNS. They are very numerous in the deep white matter and the corpus callosum where they form large clusters corresponding to peculiar white infarct-like spots seen grossly in these locations. Generalized reactive astrocytosis and microglial activation are also present. The general autopsy showed storage features in nearly all tissues and organs. ISSD is a rare disorder and the neuropathological descriptions have so far been few and brief. The combination of storage involving most cell types, the clear lysosomal material and the presence of a significant number of axonal swellings in the CNS should raise the possibility of a sialic acid storage disorder and could facilitate the choice of metabolic/molecular testing leading to the characterization of the disorder.

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176 Spinal Muscular Atrophy, Not only a Motor Neuron Disorder: Phenotype-Genotype Correlation

Brian Harding, Sabrina Yum. Childrens Hospital of Philadelphia; Wendy Chung. Columbia University, NY; Richard Finkel. Childrens Hospital of Philadelphia

Spinal muscular atrophy (SMA), the second most common neuromuscular disease of childhood, is caused by a deficiency of survival of motor neuron (SMN) protein resulting from homozygous mutations of the telomeric copy of the gene, SMN1, at chromosome 5q13. A nearly identical centromeric homocopy gene, SMN2, rescues an otherwise embryonic fatal disorder. Progressive motor neuron loss leads to a clinical presentation of skeletal muscle atrophy and weakness. In the classical severe form, Werdnig-Hoffmann disease or SMA type 1, neuronal degeneration is confined to spinal anterior horn cells, bulbar motor neurons and thalamic ventral nuclei, as well as sensory spinal ganglia. In contrast, a miscellaneous group of other genetic motor neuron disorders eg. SMA with pontocerebellar hypoplasia, distal SMA and SMA with Respiratory Distress (SMARD), are not linked to chromosome 5q deletions. This report concerns clinical and postmortem features of a neonate who presented at birth with joint contractures, extensive weakness and requirement for mechanical ventilation. No motor or sensory nerve conduction responses were elicited. Diffuse fasciculations were present on EMG. Brain MRI was normal. A homozygous deletion of exons 7 & 8 of the SMN1 gene was identified, confirming the diagnosis of SMA, and a single copy of the SMN2 gene (most infants with SMA type 1 have 2 or 3 copies of SMN2). Support was withdrawn on day 18 of life. In this case characteristic neuronal degeneration extended far beyond the usual pattern, including motor cortex, nuclei pontis and cerebellar nuclei, substantia nigra, locus ceruleus and Meynert's nucleus. The clinical, electrophysiologic and pathologic findings in this severe conatal form of Werdnig-Hoffmann disease are likely related to the low copy number of the SMN2 gene. The pathology demonstrated here supports animal model evidence for SMA being a disorder of a network of neuronal cells, not just motor neurons.

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177 Atypical Posterior Reversible Encephalopathy With Underlying Multilobar Cortical Dysplasia in an Older Patient

Stephen Saikali, Martin Savard, Steve Jodoin, Myriam Pétrin, Peter Gould. CHA Hôpital de l'Enfant-Jésus

Background: Multilobar cortical dysplasia (MCD) is a rare symptomatic cerebral malformation usually found in young subjects.

Objective: We report the case of a 66 year old patient who developed a posterior reversible encephalopathy syndrome (PRES) with atypical clinical progression over three years, retrospectively attributable at autopsy to the presence of an unsuspected posterior MCD.

Results: Gross examination and histologic study of the brain showed not only diffuse ischemic damage consistent with a PRES, but diffuse areas of cortical dysplasia involving the right parietal lobe and both occipital lobes, consistent with MCD.

Conclusion: MCD may be unusually asymptomatic and compatible with a normal life. However we make the hypothesis that it can become symptomatic with specific clinical signs during a superimposed ischemic event. Despite the exceptional nature of this disease in older adults, decompensation of an underlying quiescent brain malformation should be considered during atypical evolution of an episode of brain ischemia.

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178 Neuropathology of Schinzel-Giedion Syndrome

Boleslaw Lach. McMaster University, Hamilton Health Sciences; Jeorge Arredondo, Jackie Bourgeois. Department of Pathology and Molecular Medicine, McMaster University

Schinzel-Giedion syndrome (SGS) is characterized by severe mental retardation, multiple congenital malformations as well as frequent association with neuroectodermal tumours and agenesis of the corpus callosum (cc). Neuropathological details are limited to a study of a single fetal case of SGS. Here we report findings in a 5-year old boy with SGS and clinically suspected agenesis of corpus callosum (cc).Grossly, the brain showed a cobblestone-like surface of markedly expanded orbital and recti gyri, and milky, thickened meninges over the frontal and parietal convexities. The occipital lobes displayed excessive cortical folding due to very deep sulci on the lateral surface of both convexities. There was only minimal hydrocephalus associated with enlarged glomus of the right choroid plexus (cp) and a slight atrophy of cc and fornix. The brain stem was flattened, but otherwise unremarkable. Microscopic examination of multiple sections from the frontal lobes and abnormal orbital gyri revealed widespread glioneuronal tissue bridges extending from the cortex to the leptomeninges through gaps in the pial basal lamina. Meningeal heterotopias (MH) appeared as patches of disorganized neurons, axons and myelin on the gliotic background. MH occasionally bridged the facing cortical surfaces. The cortical layering was absent in the most affected areas, and capillaries increased in numbers throughout the brain. The occipital cortex was perfectly normal while temporal and parietal lobes showed minimal architectural abnormalities and few minute MH. The Ammon horns contained normal populations of pyramidal neurons and hypocellular dentate gyri. Multiple MH were found around pons, medulla and the cervical cord. The right cp glomus showed stromal degeneration while the remaining cp displayed villous hyperplasia. Our findings suggest that SGS may represent one more pediatric entity within an enlarging spectrum of lysencephalic cortical dysplasia syndromes.

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179 The Pathologic Findings Of An 18q22.2q23 Loss And Its Relationship To Other 18q Terminal Deletions

John Fortune, Jennifer Eschbacher, Stephen Coons. Barrow Neurological Institute/ St. Joseph's Hospital and Medical Center

We present a near term (3.36 kg) female infant born to a 26 year old G3P2A1 mother with apgar scores of 4 and 7. Immediately after birth, the patient became decreasingly responsive requiring intubation. She was twitching intermittently and developed an unusual odor likened to sweaty socks. Blood ammonia levels were noted to be increasingly elevated. The patient developed a severe progressive metabolic acidosis and died at 3 days of age. At autopsy, multiple congenital anomalies were documented. In addition to skeletal and facial dysmorphisms, the cardiovascular system was particularly affected. The brain was normally formed but microscopic examination demonstrated vacuolar leucoencephalopathy. Chromosomal analysis revealed an 18q22.2q23 loss. 18q deletions are present in approximately 1 in 40,000 births worldwide. The terminal portion of the long (q) arm of chromosome 18 is missing in 18q- syndrome, a grouping of chromosomal deletions involving the region from 18q21.1 to the end of the chromosome. There is no common breakpoint involved in 18q deletions, leading to a great deal of variability in clinical presentation. The genetic findings identified after autopsy give some insight to the congenital anomalies identified in this case. We discuss the clinical expression of this unique mutation and its relationship to other 18q- syndromes.

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180 Neuropathology of PEHO-Like Syndrome

Mircea Iftinca. University of Calgary; Ross McLeod. Alberta Children's Hospital; Jeff Joseph. University of Calgary

Progressive encephalopathy with edema, hypsarrhythmia, and optic nerve atrophy (PEHO) syndrome is a rare, autosomal recessive condition in which characteristic dysmorphic features (midface hypoplasia, epicanthic folds, protruding ear lobules, and micrognathia) are associated with non-pitting edema, hypotonia, visual deficit, developmental regression, seizures, and cerebellar atrophy. A condition similar to PEHO syndrome, but without the neuroradiologic or ophthalmologic signs, is known as PEHO-like syndrome. We present the neuropathology of a 10-year-old boy of Korean descent, who presented shortly after birth with hypotonia, contractures of the lower limbs, non-pitting edema in the face and hands, and seizures. Later he showed severe developmental delay, a head circumference in the 50% percentile (90% at birth) and poor visual fixation, nystagmus and pale optic discs. The initial MRI was normal but a later study showed diffuse cerebral atrophy. He had a normal 46, XY karyotype. Although initially thought to have PEHO-syndrome, because he had areflexia and his EEG never showed hypsarrhythmia, he was later considered to have PEHO-like syndrome. At autopsy, his brain showed cerebellar atrophy due primarily to granular neuron loss and a thinned molecular layer, but a relatively normal density of Purkinje neurons. Both the dentate nucleus and inferior olivary nuclei had increased and fragmented gyrations. The basis pontis was thinned, due primarily to a diminished number of corticospinal fibers. Magnocellular neurons were absent in the lateral geniculate body and myelinated fibers were decreased in layer IV of the primary visual cortex, which likely correlate with his inability to fix or follow. His optic nerve and tract had a normal complement of myelinated axons. To our knowledge this is the first description of a PEHO-like syndrome in a child of Korean descent.

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181 Cystic Atypical Choroid Plexus Papilloma With a Markedly Elevated Mitotic Rate in an Infant

Jennifer Bennett, Arabinda Choudhary, Narasimha Jatavallabhula, Mark Dias, Jennifer Baccon. Penn State M. S. Hershey Medical Center

Choroid plexus tumors are rare lesions of the central nervous system that account for 10-20% of brain neoplasms in patients less than one year old. Differentiating between the benign choroid plexus papilloma (CPP) and the malignant choroid plexus carcinoma (CPC) can prove challenging when one or more atypical histological features are present. Here we report a case of an atypical choroid plexus papilloma which radiologically mimicked a benign neuroglial cyst in a 10-week old female. Neurosurgical intervention to drain the cyst resulted in fortuitous removal of the entire cyst wall, and examination by pathology. The histologic features of the lesion were diagnostic of an atypical choroid plexus papilloma, due to the elevated mitotic rate of up to 12 mitotic figures per high power field, but absence of other high grade features in the entirely examined lesion. The exceedingly high mitotic rate is unusual for atypical choroid plexus papillomas. The cystic nature of the lesion is also an uncommon presentation. We describe here the presentation of an atypical choroid plexus papilloma with an unusual radiographic presentation and unique histologic features. The subtle preoperative radiographic features of this lesion are important to recognize so that full excision and pathologic examination of such lesions are performed. The importance of this has recently been emphasized in the literature by studies which have shown that an elevated mitotic rate is the atypical feature that significantly correlates with recurrence, thus necessitating the importance of interval radiological surveillance for these patients.

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182 The Neuropathology of Mucopolysaccharidosis Type VII (Sly Disease): A Case Report and Review of the Literature

Kelly Devers, Marie River-Zengotita, Charles Williams. Department of Pediatrics; Anthony Yachnis. University of Florida College of Medicine

Mucopolysaccharidosis Type VII (MPS VII, Sly Syndrome) is a rare autosomal recessive disorder caused by mutations on chromosome 7q11.21-22. We report a 10-year-old male who had been the 4883 gm product of a 37 week gestation in a 22-year-old female. The child was born with marked ascites and was noted to have coarse facial features and macrocephaly. Laboratory evaluation of lysosomal enzymes revealed reduced beta-glucuronidase levels (patient = 5.8 nmol/mg/hr; normal: 363+/-85.1). The patient had lifelong developmental and neurologic delay and was lost to follow-up in the years preceding death. At autopsy the brain weighed 1270 gm (normal for age: 1290 gm) and on sectioning revealed widening of white matter perivascular spaces that was most prominent in the parietal and occipital lobes. Microscopic examination of H&E and LFB-PAS stained sections revealed extensive neuronal storage with affected cells being distended with accumulation of periodic acid-Schiff (PAS) positive granular material. These neuronal changes were associated with reactive gliosis. Hippocampal pyramidal cells of CA 2-4 and the dentate nucleus were particularly involved while CA1 and the subiculum were relatively spared. Significant neuronal storage was seen in the temporal lobe neocortex with lesser degrees identified in the frontal, parietal, and occipital cortices. The deep nuclei, particularly the caudate and putamen, contained globose neurons, while neuronal loss and gliosis were seen in the thalamus. Tegmental brainstem motor nuclei, especially the hypoglossal nucleus, showed marked neuronal storage while pigmented neurons of the substantia nigra and locus ceruleus were less affected. Multiple granular accumulations of material were present in the cerebellar molecular layer with Bergman gliosis but there was little Purkinje cell storage. White matter perivascular spaces contained a loose, hypocellular, collection of mesenchymal-appearing cells but no storage material. Our findings provide additional insights into the spectrum of MPS VII neuropathology, of which only rare reports exist.

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183 Septo-Optic Dysplasia With Auricular Duplication and Transposition of the Great Vessels: A Case Report

Mark Smethurst, Mary Fowkes. The Mount Sinai Medical Center, Dept. of Pathology

Septo-optic dysplasia is a rare congenital malformation characterized by two of three criteria: optic nerve hypoplasia, midline abnormalities (absence of the corpus callosum or agenesis of the septum pellucidum) and pituitary hypoplasia. Septo-optic dysplasia occurs in 1/10,000 live births with increased frequency in younger and primigravid mothers and decreased socioeconomic status. While the etiology is unknown, some reports suggest the disorder results from vascular insufficiency during the second and third month's gestation. We present autopsy findings of a 42.3 week gestation infant with septo-optic dysplasia associated with fronto-nasal dysplasia that expired 1.4 hours post-partum. At 27.29 weeks gestation ultrasound examination revealed transposition of the great arteries, `holoprosencephaly' and hydrocephalus. Post-mortem genetic testing revealed no karyotypic abnormalities. Autopsy revealed agenesis of the septum pellucidum, the presence of a rostral corpus callosum (ruling out holoprosencephaly), agenesis of the eyes, optic nerves and optic chiasm, and hypoplasia of the anterior pituitary, confirming the diagnosis of septo-optic dysplasia. The finding of cortical dysplasia (polymicrogyria and pachygyria) and cerebral heterotopias (here at the 4th cranial nerve) are common associations with septo-optic dysplasia. The finding of hydrocephalus, foreshortened frontal and occipital lobes, cranial dysraphism, and median facial cleft defects are rare associations. However, duplication of the auricles and dextro-transposition of the great arteries have never previously been described with septo-optic dysplasia. The finding of absent posterior communicating arteries and an atretic right anterior cerebral artery support an etiology of vascular disruption. The failure of median facial structure fusion has been associated only rarely with septo-optic dysplasia, but not to the degree seen here. This case represents a unique confluence of septo-optic dysplasia, transposition of the great arteries and duplication of the auricles suggesting a profound defect in midline embryogenesis.

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184 Brain Abnormalities Associated With Glycogen Storage Disease, Type IV (Glycogen Branching Enzyme Deficiency), A Case Report

Mary Fowkes. Mount Sinai Medical Center and School of Medicine

Glycogen storage disease (GSD) type IV is an autosomal recessive disorder caused by a deficiency in the glycogen branching enzyme referable to mutations in its gene. It is clinically heterogeneous with variable presentations distinguished by age and predominant organ involvement. The `classical' type features hepatic involvement in childhood with progressive liver disease. Other subgroups include a variety of neuromuscular forms with perinatal, congenital, juvenile and adult presentations. To date, there has been limited documentation of the brain abnormalities associated with this disorder. This case is of a male fetus at 35 weeks gestation that developed fetal heart rate decelerations and bradycardia necessitating emergency cesarean section, but was dead upon delivery and could not be resuscitated. Autopsy findings revealed diagnostic large diastase resistant PAS positive intracytoplasmic inclusions typical of GSD in the liver, heart and skeletal muscle. The 364 gram brain had a gyral pattern consistent with just under 36 weeks gestation. However, following fixation the brain was found to be abnormally soft. Microscopically the white matter was poorly myelinated for the gestational age and there were large diastase resistant PAS positive intracytoplasmic inclusions typical of GSD confined to neurons within selected brainstem nuclei. These brainstem nuclei included the interstitial nucleus of the medial longitudinal fasciculus, the abducens nucleus, the vestibular nucleus, and the hypoglossal nucleus. No inclusions were seen within neurons of the cerebral cortex or thalamus. No inclusions were seen within white matter axons. Only finely granular predominantly non-diastase resistant PAS intracytoplasmic staining was seen in neurons of the globus pallidus, substantia nigra, oculomotor nucleus, olivary nucleus, dorsal motor nucleus of vagus, and neurons of the basis pontis, and in Purkinje cells. The selective presentation of intracytoplasmic inclusions suggests targeted sampling for autopsy and may represent groups of neurons with similar and dissimilar physiologic characteristics warranting further investigation.

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185 Propionic Acidemia: Neuropathologic Features and Review of the Literature

Stephen Wilhem, Hannah Coulson, Suash Sharma, David Flannery, Amyn Rojiani. Departments of Pathology and Pediatrics, Georgia Health Sciences University, Medical College of Georgia

Propionic acidemia is an autosomal recessively inherited organic acidemia characterized by a diminished activity of Propionyl co-A Carboxylase (PCC) and resulting excess of the substrate Propionic Co-A produced from the catabolism of isoleucine, valine, methionine, threonine and odd-chain fatty acids. As a result, propionic academia patients may develop metabolic acidosis, ketonuria, hyperammonemia, and hyperuricemia. Patients typically manifest symptoms early in life, commonly as neonates. Presenting signs and symptoms include lethargy, nausea and vomiting, and acute neurologic symptoms as well. This case report details the neuropathologic findings following the autopsy of a seven year old African-American female diagnosed with Propionic acidemia in infancy. The patient initially presented at 3 weeks of age in coma., without acidosis or ketosis. Persistent neutropenia led to urinary organic acid analysis which demonstrated elevated propionate, methyl-citrate, propionyl glycine. PCC activity in leukocytes was low, confirming the diagnosis of propionic acidemia. An initial head CT scan was performed on the 23rd day of life and showed multifocal parenchymal hyperdensities. A corresponding brain MRI without contrast revealed multiple small foci of low signal with both cerebellar hemispheres. She died at 7 years of age from aspiration pneumonia and intra-alveolar pulmonary hemorrhage resulting in hypoxemia and secondary cerebral edema. Significant neuropathologic findings included diffuse vacuolation within the cerebral cortex, the basal ganglia and the cerebellum consistent with previous case reports. Her previous brain CT scans confirmed the presence of advanced cortical involutional changes. Histopathology also revealed white matter changes with pallor and mild gliosis as well as occasional Alzheimer type II astrocytosis. Ultrastructrural studies were performed using formalin fixed material. The patient's neuroimaging studies and pathology are described, along with a review of the literature.

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186 Smurf2 Accumulates in TDP-43 Positive Cytoplasmic Inclusions in Spinal Cord But Not in Hippocampus of Sporadic ALS

Masataka Nakamura, Satoshi Kaneko. Kansai Medical University; Hidefumi Ito. Kyoto University Graduate School of Medicine; Shinya Asayama, Kengo Fujita, Reika Wate, Hirofumi Kusaka. Kansai Medical University

Aims: Recent biochemical and immunohistochemical studies on amyotrophic lateral sclerosis (ALS) spinal cord and hippocampus tissues demonstrated that the composition of the TAR DNA binding protein-43 (TDP-43) inclusions was regionally distinct, suggesting different underlying pathogenic processes. We have previously reported that phosphorylated Smad2/3 (pSmad2/3), the major intracellular mediators of transforming growth factor (TGF)-ß signaling, abnormally accumulated in TDP-43 inclusions of anterior horn cells (AHCs) in sporadic ALS (SALS). Smad ubiquitination regulatory factor-2 (Smurf2) is an E3 ligase in a family of HECT domain ubiquitin ligases, interacts with Smad proteins, and promotes their ubiquitin-dependent degradation thereby controlling TGF-ß signaling. To elucidate regional differences in pathomechanisms and composition of TDP-43 inclusions in relation to Smad2/3 and Smurf2, we immunohistochemically analyzed brain and spinal cord tissue of SALS.

Methods: Spinal cord and hippocampus of eight SALS (four men and four women, ages 66-81years, mean ± SD=71.0 ± 6.1 years) and three control patients (three men, ages 60-76years, mean ± SD = 68.7 ± 8.1 years) without neurological disorders were investigated immunohistochemically using antibodies against Smurf2, pSmad2/3, pTDP-43 (anti-phosphoserine 409/410) and ubiquitin.

Results: Multiple immunofluorescence staining for Smurf2, pSmad2/3, pTDP-43 and ubiquitin revealed co-localization of these four proteins within the neuronal and glial inclusions of spinal cord in SALS patients. On the other hand, TDP-43 inclusions in granular cells of hippocampus of SALS were immunohistochemically negative for Smurf2 or pSmad2/3.

Conclusions: Our data indicate that the pathogenic mechanisms of cytoplasmic inclusion development may be different between AHCs of spinal cord and hippocampus in SALS. Moreover, regionally different pathways that lead to TDP-43 inclusion formation and neurodegeneration may be present in SALS.

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187 Presence of Somatic Dendrites and Reduction of Apical Dendrites of Purkinje Cells in Spinocerebellar Ataxia Type 31

Masaki Takao. Department of Neuropathology, Tokyo Metropolitan Geriatric Hospital; Bernardino Ghetti. Indiana Alzheimer Disease Center; Tatsuru Mihara. Mihara Clinic; Kinya Ishikawa. Department of Neurology, Tokyo Medical and Dental University; Aya Tokumaru. Department of Radiology, Sayaka Funabe. Department of Neuropathology, Tokyo Metropolitan Geriatric Hospital; Hiroaki Kimura. Department of Neurology, Ban Mihara. Department of Neurology, Mihara Memorial Hospital; Masaki Fujita. Department of Legal Medicine, School of Medicine, Keio University; Kinuko Suzuki, Shigeo Murayama. Department of Neuropathology, Tokyo Metropolitan Geriatric Hospital

Spinocerebellar ataxia type 31 (SCA31), an autosomal dominant cerebellar degeneration, is characterized clinically by slowly progressing ataxia and genetically by a 2.5- to 3.8-kb insertion of pentanucleotide repeats (TGGAA)n between the TK2 and BEAN genes located on chromosome 16q21-q22. We present genetic, clinical, neuroradiological and neuropathologic data on a family identified as "SCA31-KGJ." We reconstructed a pedigree consisting of 61 members over five generations. Genetic analysis, carried out on 11 members, revealed the genetic mutation in three symptomatic and three asymptomatic individuals. The affected members developed pure cerebellar ataxia in the sixth decade of life. MRIs of three affected individuals show severe cerebellar atrophy at the level of vermis and anterior lobes.Neuropathologic analysis carried out in the proband, macroscopically showed a severe cerebellar atrophy. Histologically, moderate loss of Purkinje cells and a mild neuronal loss in the inferior olivary and dentate nuclei are evident. Purkinje cells appear disorganized in their arrangement, are atrophic and have a basophilic cytoplasm. Calbindin D28K (CB28) immunohistochemistry shows a variety of pathologic changes in Purkinje cells. Somatic dendrites originate from all around the cell perikaryon. Apical dendrites are reduced in number, thin and extend into the molecular layer for short distances. Purkinje cell axons are immunoreactive with CB28 and appear to form torpedoes within the granule cell layer. In the white matter, axons of Purkinje cells are fewer as compared to a normal control. Antibody SMI31 against phosphorylated neurofilament shows basket-like structures around Purkinje cell perikarya. Synaptophysin immunohistochemistry shows numerous immunopositive profiles around Purkinje cell somata and the proximal portion of the apical dendrites. By electron microscopy, numerous spines and synaptic membranes are present in the vicinity of the Purkinje cell soma.Our findings suggest the neuropathologic changes of SCA31 are the result of an abnormal development of Purkinje cells and their connectivity.

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188 Anti-Yo-Associated Paraneoplastic Cerebellar Degeneration Without Associated Malignancy

Carrie Mohila, T. Bourne. Univeristy of Virginia

Paraneoplastic cerebellar degeneration (PCD) is a rare complication of malignancy, associated with rapid progression of pancerebellar dysfunction. PCD associated with anti-Yo autoantibodies is most commonly associated with gynecological and breast malignancies. Well characterized neuropathologic examination of these patients at autopsy is limited. We present a case of a 61-year-old woman with a recent diagnosis of Lyme disease who presented with a four month history of gait instability, vertigo, and diploplia. She was also found to have a high titer anti-Yo antibody. Extensive search for a primary malignancy was negative. Her neurologic symptoms rapidly progressed and included cognitive changes suggestive of concomitant limbic encephalitis. She died eight months after initial presentation. Microscopic neuropathologic findings in the cerebellum included near complete absence of Purkinje cells with preserved basket cells, marked Bergmann gliosis, and decreased granule cells. The dentate nucleus showed loss of neurons with gliosis and surrounding demyelination. The cerebellar white matter and dentate nucleus showed extensive microglial activation and an exuberant inflammatory reaction composed almost exclusively of macrophages and CD8 T cells. There were no pathologic findings in the cerebral cortex or spinal cord to support an encephelomyelitic process. The findings in this case support other reports that PCD associated with anti-Yo antibodies is mediated, at least in part, by autoantibodies and/or CD8 T cells. Furthermore, this case represents a very rare example of a patient with anti-Yo positive PCD without detectable malignancy clinically or at autopsy.

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189 Atypical Protein Kinase C in Down Syndrome (DS): Preliminary Neuropathological Observations

Jianying Zeng, Charles Shao, Chandrakant Rao, Jenny Libien, Suzanne Mirra. Suny Downstate Medical Center, Department of Pathology

Neuropathological features of Alzheimer's disease (AD) often occur in Down syndrome (DS) brains by age 35 to 40 years. We had the opportunity to study autopsy brains derived from two women with DS, aged 50 and 23 years, both of whom died of complications of congenital heart disease. Previously, we had identified PKMzeta, an atypical protein kinase C isoform, in limbic neurofibrillary tangles (NFT) in AD (Crary et al. J Neuropath Exp Neurol 2006;65:319); PKMzeta has been shown necessary for AMPA glutamate receptor trafficking and the maintenance of long-term memory. We sought to compare the distribution of PKMzeta in DS with that in AD. At autopsy, both DS brains exhibited foreshortening with bilateral atrophy of the superior temporal gyrus and extensive mineralization of the basal ganglia, predominantly involving the globus pallidus. The brain of the 50 year old also displayed frequent neuritic plaques and NFT involving multiple sites. The brain of the 23 year old DS patient exhibited less advanced AD pathology with frequent diffuse plaques in many regions but only rare NFT in the entorhinal cortex and dorsal raphe nucleus. PKMzeta immunohistochemistry revealed diffuse label of cell bodies and apical dendrites in hippocampal neurons in the brain of the 23 year old, identical to that observed in age-matched controls. The brain of the 50 year old also displayed diffuse cytoplasmic immunolabel in hippocampal neurons without NFT, but most tangle-bearing neurons lacked PKMzeta immunolabel. Only small number of NFT-containing neurons and most Hirano bodies labeled with PKMzeta antibody. Thus, the distribution of PKMzeta in the 50 year old DS patient with advanced AD pathology paralleled that which we previously noted in AD. We hypothesize that the apparent loss of PKMzeta in NFT-bearing neurons plays a role in the cognitive decline and memory impairment in DS individuals with advanced AD pathology.

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190 Endothelial-Reactive Antibodies in Susac's Syndrome

W Waldman, Deborah Knight. The Ohio State University Department of Pathology; Cynthia Magro. Cornell University; Martin Lubow. The Ohio State University Department of Opthalmology; Robert Rennebohm. Alberta Children's Hospital Department of Pediatrics; John Susac. Neurology and Neurosurgery Associates

Susac's syndrome (SS) is a rare disorder characterized by a clinical triad of encephalopathy, branch retinal artery occlusion, and hearing loss. The frequent failure of these components to develop concurrently, and the variable pattern of changes observed in the brains of SS patients by MRI often lead to misdiagnosis. Since its identification in the 1970's, little progress has been made in resolving the etiology of SS. However the disorder is postulated to result from microangiopathy affecting the precapillary arterioles of the brain, retina, and inner ear, and the fact SS patients respond in varying degrees to immunosuppressive therapy implies an immunological basis for the disorder. In the current investigation, we have tested the hypothesis that SS patients harbor potentially injurious circulating endothelial-reactive autoantibodies. Immunofluorescent staining of human endothelial cells using SS patient sera as the "primary antibody" demonstrated the presence of endothelial-reactive antibodies in the sera of SS patients. Western blot analysis of endothelial cell protein extracts probed with SS patient sera revealed a relatively consistent pattern of bands at molecular weights of 40, 45, and 70 kD, with most sera generating two of these bands, some generating all three, and all generating at least one. To identify potential targets at a higher resolution, endothelial protein extracts were separated by 2-dimensional gel electrophoresis. Protein spots of the above molecular weights and with demonstrated serum reactivity were tentatively identified by mass spectrophotometry as annexin (40 kD) and non-neuronal enolase (45 kD). Further studies are currently in progress to identify the 70 kD target. Data generated by these studies provide novel insights into the etiology of the enigmatic disease and provide the foundations for the development of a definitive diagnostic test for Susac's syndrome.

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191 FTLD Pathology in Adult Polyglucosan Body Disease

Kyung-Hwa Lee, Qinwen Mao, Manjari Mishra, Katherine Gasho, Darren Gitelman. Northwestern University Feinberg School of Medicine; Charles White, Dennis Burns, Kimmo Hatanpaa. University of Texas Southwestern Medical Center; Hasan Akman. Columbia University Medical Center; Salvatore DiMauro. Columbia University Medical Center; Eileen Bigio. Northwestern University Feinberg School of Medicine

Adult polyglucosan body disease (APBD) is a rare disorder characterized by the diffuse accumulation of insoluble glucose polymers (polyglucosan bodies) in the brain and peripheral nerves. The classic presentation is that of upper and lower motor neuron dysfunction, urinary incontinence, and dementia. Despite the frequent occurrence of dementia, histologically proven cases with FTLD (frontotemporal lobar degeneration) have not previously been described. We report a unique case of APBD with a heterozygous mutation in the gene (GBE1) encoding glycogen branching enzyme and concomitant FTLD-FUS (fused in sarcoma) proteinopathy. This 43-year-old woman diagnosed with frontotemporal dementia died after an 8-year-course of disease. Pathologic examination revealed numerous polyglucosan bodies in many regions of the brain including the neocortex, striatum, basal ganglia, cerebellum and brain stem. In addition, ubiquitin-positive inclusions were identified in the dentate gyrus, which were negative for TDP-43 and positive for FUS protein. Genetic analysis demonstrated a heterozygous intronic point mutation (ac-to-gc) in the splicing donor side of exon 5 in the GBE1 gene on chromosome 3. We investigated three additional APBD cases for presence of concomitant FTLD. One case, a 72-year-old man with an 8-year history of dementia, had TDP-43-positive inclusions in medial temporal regions. Discovering TDP-43 or FUS pathology in two of the four cases suggests that combined FTLD pathology may be common in APBD.

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192 Collagenous Fibrosis: A Rare Tissue Reaction of Deep Brain Stimulation (DBS)

Vinata Vedam-Mai, Michael Ullman. McKnight Brain Institute; Michael Okun. Department of Neurology, Anthony Yachnis. McKnight Brain Institute;Department of Neurology,University of Florida College of Medicine

The University of Florida Movement Disorders Center and DBS-BTN have conducted post-mortem brain examinations on 26 cases cases with the goal of identifying and characterizing pathologic tissue changes associated with DBS lead placement. In most cases, only mild reactive gliosis was observed adjacent to the lead tip, which is typically located in the subthalamic nucleus or the globus pallidus interna. However, we report the unusual finding of prominent collagenous fibrosis that occurred near the lead tip in a 74-year old man with clinical idiopathic Parkinson's disease. This patient had bilateral DBS electrodes placed in both the subthalamic nuclei. At the time of death, the left electrode had been implanted for 13 months, and the right electrode had been implanted for 11 months. A complete, neuropathological examination was performed after the subject died from complications of pneumonia. After fixation, the brain weighed 1460 grams and contained bilateral cortical DBS lead defects. Serial sectioning confirmed the locations of distal lead placement in the left subthalamic nucleus and at the junction between the right subthalamic nucleus and the substantia nigra. The brainstem showed marked depigmentation of the substantia nigra and locus ceruleus but no other significant gross findings. Histopathologic analyses confirmed the loss of pigmented nigral and locus ceruleus neurons with alpha-synuclein-reactive Lewy bodies consistent with idiopathic Parkinson's disease. No cortical alpha-synuclein-reactive Lewy bodies were identified and no significant tau- or A-beta-amyloid pathology was seen. There was striking dense collagenous fibrosis at distal end of the right DBS electrode that was associated with focal hemosiderin deposition, chronic inflammation, and mild gliosis. We are not aware of any prior reports of such a reaction to DBS placement to date and speculate that the cause of this change could be related to local hemorrhage or infection at the lead site.

© 2011 American Association of Neuropathologists, Inc

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