Haemophagocytic lymphohistiocytosis (HLH) is an uncommon entity comprising a constellation of symptoms and laboratory findings that together form the clinical inflammatory syndrome. HLH encompasses two separate conditions: a primary hereditary form (familial haemophagocytic lymphohistiocytosis, FHL) and a secondary/acquired form (secondary HLH, sHLH).1 sHLH may follow a variety of stimuli including viral, bacterial, and fungal infections, as well as a number of malignant diseases, notably T-cell lymphomas. FHL is generally inherited in an autosomal recessive or x-linked manner, presents during infancy or early childhood, and is uniformly fatal if left untreated. Here we present an interesting case of FHL presenting in a pair of identical twins in the immediate post-natal period.
The patients were diamnionic-dichorionic identical twins born at 35 weeks gestation. Twin 1 was born via normal spontaneous vaginal delivery, while twin 2 was born via emergent caesarean section due to placental abruption. The initial post-natal clinical course was uncomplicated, and the patients were discharged home without complication. At 2 months of age both twins were hospitalised for fever with neutropenia, and severe thrombocytopenia. Liver ultrasound showed hepatomegaly. Laboratory studies (Table 1) showed many abnormalities, including low fibrinogen, elevated D-dimers, and prolonged coagulation times, indicating likely disseminated intravascular coagulation. Blood cultures at this time were negative in both twins. Simultaneous bone marrow aspirates were obtained on both twins. While the bone marrow aspirate from twin 2 showed non-specific findings due to markedly haemodilute aspirate smears (i.e., predominantly mature lymphocytes similar to peripheral blood), the bone marrow aspirate from twin 1 showed significantly increased haemophagocytic histiocytes on the smear (Fig. 1) in addition to erythroid hyperplasia and relative decrease in myeloid cells. Megakaryocytes were unremarkable. Additional laboratory studies (Table 2) showed elevated alanine aminotransferase (ALT), elevated aspartate aminotransferase (AST), increased triglycerides and increased soluble interleukin-2 receptor (CD25) in both twins. Studies for NK-cell functional activity were non-contributory due to technical reasons in twin 1 and not decreased significantly in twin 2.
Twin 1 met the following criteria for the diagnosis of haemophagocytic lymphohistiocytosis: (1) fever, (2) fibrinogen level less than 1.5 g/L, (3) increased haemophagocytic lymphohistiocytosis on bone marrow aspirate smear, (4) ferritin level more than 500 ng/mL, and (5) elevated soluble interleukin-2 receptor (soluble CD25) level of more than 2400 U/mL. Twin 2 met the following diagnostic criteria for the diagnosis of haemophagocytic lymphohistiocytosis: (1) fever, (2) cytopenias (haemoglobin less than 9 g/dL and platelet count less than 100 k/μL), (3) fibrinogen level less than 1.5 mg%, (4) ferritin level more than 500 ng/mL, and (5) soluble interleukin-2 receptor (soluble CD25) level more than 2400 U/mL. A diagnosis of haemophagocytic lymophohistiocytosis was made on both the twins, based on these findings (see Table 3).2 Due to the fact that these were identical twins, FHL was considered and subsequent perforin-1 (PRF1) gene mutation analysis showed identical mutations, i.e., homozygous for 50delT (L17fsX50), confirming the diagnosis of FHL type 2 in both the twins. Both patients were started on chemotherapy consisting of dexamethasone, etoposide, and cyclosporine according to the HLH-2004 protocol. Twin 2 progressively deteriorated, developing respiratory and hepatic failure, fungaemia, and worsening coagulopathy, with death occurring 8 days after the start of chemotherapy. Twin 1 did well on chemotherapy and was eventually discharged 4 months after admission.
FHL is a rare condition, most often presenting in infancy, with a peak age of presentation between 1 and 6 months.1 The reported incidence ranges from 0.12 per 100 000 cases in studies conducted in Sweden3 to 7.5 per 10 000 among hospitalised patients in Turkish studies.4 The high rate of FHL in the Turkish population may be due to the high rate of consanguineous marriage, fitting with the autosomal recessive inheritance patterns seen in most genetically elucidated types of FHL. In addition, as the incidence of FHL varies amongst people of different ethnic backgrounds, so too do the genetic defects that give rise to the HLH phenotype.
The mechanism of immune impairment leading to the syndrome is unclear; however, it is clear that the mechanism involves the release of inflammatory cytokines, T-cell and histiocyte activation, and NK-cell impairment. This leads to the defining clinical and laboratory findings, including fever, splenomegaly, multiple cytopenias, hypertriglyceridaemia and/or hypofibrinogenaemia, haemophagocytosis in the bone marrow, spleen, or lymph nodes, low/absent NK-cell activity, hyperferritinaemia, and increased soluble IL-2 receptor (soluble CD25), in the absence of malignancy (Table 3). Five of the eight criteria are necessary for a diagnosis of HLH, though a diagnosis may still be made if there is molecular evidence consistent with HLH.2 Diagnostic work-up includes complete blood count and blood smear analysis, liver function tests, blood chemistry, triglyceride and cholesterol levels, ferritin, and coagulation tests. Bone marrow biopsy and aspirate may be initially negative, and should be repeated over time to see if evidence of haemophagocytosis appears; biopsies from other organs can help demonstrate haemophagocytic histocytes or evidence of chronic persistent hepatitis in more uncommon instances.2
Microscopic features are identical in the familial and acquired forms. The main histological feature is diffuse infiltration by T-lymphocytes and histiocytes. The organs most frequently examined are the bone marrow, spleen, liver, and central nervous system, and so are the most frequently described; however, almost no organ is spared, and the characteristic findings may be seen in almost any organ. Perhaps most frustrating is the fact that the characteristic haemophagocytic histiocytes (showing phagocytosis of nucleated cells as well as red blood cells) may be absent in some organs, or infrequent at best, thus necessitating repeated biopsies for histological confirmation. Histological findings in the liver, which may be the second most common organ biopsied after the bone marrow, shows a pattern of sinusoidal dilatation, congestion and hyperplasia of Kupffer cells. Haemophagocytic histiocytes are not seen in the portal inflammatory infiltrate, but characteristically in the dilated sinusoids.5 The immunophenotype of the phagocytic histiocytes are also unique. They express common macrophage-associated antigens, S100 protein and also may express CD1a (which are more commonly seen in Langerhans cells or interdigitating dendritic cells).6
Five distinct genetic subtypes of FHL have been identified to date. FHL type 1 has been mapped to chromosome 9 (9p21), although the exact protein and gene remain unknown.7 This mutation is believed to account for 10% of all FHL. FHL type 2 has been mapped to the perforin gene located on chromosome 10 (10q21–22).8 Perforin gene mutations cause either a non-functional form or markedly reduced/absent form of the perforin protein (PRF1), resulting in decreased granzyme mediated toxicity by NK cells and cytotoxic T-cells. Missense mutations of PRF1 result in a non-functional form of the protein through conformational changes that inhibit proteolytic processing of the protein precursors; nonsense mutations result in absent protein production.9,10 Further, the type of mutation affects the clinical presentation with missense mutations having a later age of onset than nonsense mutations.11 Mutations in PRF1 account for 20–40% of FHL. While there have been over 50 different PRF1 mutations identified, a subset of mutations have been found to be more prevalent in certain ethnicities. The 1122G→A (W374X) mutation appears to be more prevalent in Turkish cohorts, while the 272C→T (A91 V) mutation appears to dominate in Italian populations.11 The 50delT mutation, also seen in our described patients, is interestingly the single mutation seen in African/African American individuals. FHL type 3 is traced to the hMunc13–4 gene, encoding the Munc13–4 protein, which is involved in vesicle priming.12 The granules containing perforin and granzymes A and B are normal, however priming prior to release is abnormal, resulting in the abnormal FHL phenotype. FHL type 4 results from abnormalities in the Syntaxin 11 (STX11) gene,13 found only in patients of a Turkish/Kurdish background thus far. Syntaxin 11 is also involved in vesicle priming. FHL type 5 has been mapped to chromosome 19p, which encodes the STXBP2 gene encoding Munc18–2 protein (syntaxin binding protein 2).14 This protein is involved in intracellular trafficking and granule exocytosis. Regardless of the mutated protein, or specific type of mutation, patients with FHL tend to have a clinically similar disease.
Prior to the use of cytotoxic chemotherapy FHL was uniformly fatal, with patients succumbing to infection and multi-organ failure. With the use of chemotherapeutic agents, including etoposide, anti-thymocyte globulin, and CSA, as well as steroids, the majority of patients were able to show symptomatic improvement; however, a true cure was not obtained without haematopoietic stem cell transplantation. The regimen currently in use, as produced by the 2004 revision of the Histiocyte Society consensus study protocol (HLH-2004), uses etoposide, dexamethasone, cyclosporine, and intrathecal methotrexate.4
In conclusion, haemophagocytic lymphohistiocytosis is a rare disease that may present in a phenotypically indistinguishable acquired or familial form. The familial form is a genetically heterogeneous and phenotypically homogeneous disease, of which five genetically distinct subtypes have been described. Diagnosis rests on a combination of clinical and histological features, and chemotherapy along with haematopoietic stem cell transplant is the mainstay of therapy.
Conflicts of interest and sources of funding: The authors state that there are no conflicts of interest to disclose.
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