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Immunohistochemical Expression of Stem Cell, Endothelial Cell, and Chemosensitivity Markers in Primary Glioma Spheroids Cultured in SerumContaining and SerumFree Medium

Christensen, Karina PhD; Aaberg-Jessen, Charlotte MS; Andersen, Claus DMSc; Goplen, Dorota MD, PhD; Bjerkvig, Rolf Professor; Kristensen, Bjarne Winther MD, PhD

doi: 10.1227/01.NEU.0000368393.45935.46
Experimental Studies

OBJECTIVE: To investigate the influence of serum-free medium (SFM) supplemented with epidermal growth factor and basic fibroblast growth factor compared with conventional serum-containing medium (SCM) on the phenotype of organotypic primary spheroids from seven gliomas.

METHODS: Paraffin sections of the original surgical specimens, primary glioma spheroids, and U87 derived spheroids were stained immunohistochemically with the stem cell markers CD133, podoplanin, Sox2, Bmi-1, and nestin; the endothelial cell markers CD31, CD34, and Von Willebrand Factor (VWF); the chemosensitivity markers P-glycoprotein and tissue inhibitor of metalloproteinases-1 (TIMP-1); and glial fibrillary acidic protein, neural cell adhesion molecule CD56, and the proliferation marker Ki67.

RESULTS: Scoring of the immunohistochemical stainings showed that the expression of CD133 and all other markers included was preserved in primary spheroids, confirming the in vivo-like nature of these spheroids. Spheroids in SFM better mimicked the in vivo phenotype with significantly more CD133, CD34, VWF, P-glycoprotein, TIMP-1, and Ki67 compared with SCM.

CONCLUSION: In this first study of the influence of SFM on primary glioma spheroids, the conditions favored an in vivo-like phenotype with increased expression of CD133. More vascular structures were found in SFM, suggesting that the close relationship between blood vessels and tumor stem-like cells was better preserved in this medium.

Department of Pathology, Odense University Hospital, Odense, Denmark (Christensen) (Aaberg-Jessen) (Kristensen)

Department of Neurosurgery, Odense University Hospital, Odense, Denmark (Andersen)

Department of Biomedicine, University of Bergen, Bergen, Norway (Goplen) (Bjerkvig)

NorLux Neuro-Oncology, Centre de Recherche Public Santé, Luxembourg, Luxembourg (Bjerkvig)

Reprint requests: Bjarne Winther Kristensen, MD, PhD, Department of Pathology, Odense University Hospital, Winsløwparken 15, 5000 Odense C, Denmark. E-mail:

Received, July 20, 2009.

Accepted, December 27, 2009.

Copyright © by the Congress of Neurological Surgeons