Masanori Nishi

Congenital amegakaryocytic thrombocytopenia iPS cells exhibit defective MPL-mediated signaling

Congenital amegakaryocytic thrombocytopenia (CAMT) is caused by the loss of thrombopoietin receptor-mediated (MPL-mediated) signaling, which causes severe pancytopenia leading to bone marrow failure with onset of thrombocytopenia and anemia prior to leukopenia. Because Mpl(-/-) mice do not exhibit the human disease phenotype, we used an in vitro disease tracing system with induced pluripotent stem cells (iPSCs) derived from a CAMT patient (CAMT iPSCs) and normal iPSCs to investigate the role of MPL signaling in hematopoiesis. We found that MPL signaling is essential for maintenance of the CD34+ multipotent hematopoietic progenitor (MPP) population and development of the CD41+GPA+ megakaryocyte-erythrocyte progenitor (MEP) population, and its role in the fate decision leading differentiation toward megakaryopoiesis or erythropoiesis differs considerably between normal and CAMT cells. Surprisingly, complimentary transduction of MPL into normal or CAMT iPSCs using a retroviral vector showed that MPL overexpression promoted erythropoiesis in normal CD34+ hematopoietic progenitor cells (HPCs), but impaired erythropoiesis and increased aberrant megakaryocyte production in CAMT iPSC-derived CD34+ HPCs, reflecting a difference in the expression of the transcription factor FLI1. These results demonstrate that impaired transcriptional regulation of the MPL signaling that normally governs megakaryopoiesis and erythropoiesis underlies CAMT.

Mutations of syntaxin 11 and SNAP23 genes as causes of familial hemophagocytic lymphohistiocytosis were not found in Japanese people

AbstractAlthough mutations of perforin, MUNC13-4 and syntaxin 11 genes have been found in children with familial hemophagocytic lymphohistiocytosis (FHL), the incidence of each genetic subtype varies in different ethnic groups. We evaluated mutations of syntaxin 11 and SNAP23 genes in 30 Japanese FHL patients. The patients had no mutations and 10% had one polymorphism (146G>A) of syntaxin 11, while no mutation of SNAP23 was observed. Our results indicate that aberrations in the SNARE system may not cause FHL in Japanese families.

Reduced-intensity conditioning in unrelated donor cord blood transplantation for familial hemophagocytic lymphohistiocytosis†

Familial hemophagocytic lymphohistiocytosis (FHL) is a disorder of immune homeostasis characterized by fever, cytopenias, hepatosplenomegaly, and coagulopathy. We studied the outcomes of 13 FHL patients who underwent the first unrelated cord blood transplantation (UCBT) after non‐myeloablative conditionings. The major regimen consisted of fludarabine (FLU; n = 12)+melphalan (MEL; n = 11)±low‐dose total body irradiation (TBI 2‐4 Gy; n = 6). The median age at presentation and period to UCBT were 6 and 5 months, respectively. Central nervous system (CNS) disease developed in one infant at diagnosis, and in two others until UCBT. HLH activity was controlled in all but one at the time of UCBT. Ten patients had early engraftment on median day 21 with no grade >2 treatment‐related toxicity and two controllable grade >2 acute GVHD. Two patients with early rejection successfully underwent subsequent UCBT after myeloablative conditioning. Two others had late graft failure following mixed donor chimerism. Two deaths occurred from HLH; early liver failure and late CNS disease. Of 11 FLU+MEL‐conditioned patients, the frequency of disease‐free complete engraftment was higher for MEL (≥120 mg/m2)+TBI, or high‐dose MEL (180 mg/m2) than for others (83% vs. 25%, p = 0.036). The FLU+MEL‐based non‐myeloablative regimen was acceptable for FHL infants undergoing UCBT, although further studies will be needed for confirmation. Am. J. Hematol. 87:637–639, 2012.

Expression and production of aberrant PAX5 with deletion of exon 8 in B-lineage acute lymphoblastic leukaemia of children

We investigated PAX5 expression in childhood B‐lineage acute lymphoblastic leukaemia (ALL). Seven of 21 children with B‐lineage ALL had multiple PAX5 variants, while 14 children and healthy controls showed full‐length (FL) and one variant PAX5. By Western blotting, healthy controls displayed Pax5‐FL, while one short Pax5, derived from the deletion of exon 8 (Pax5‐ΔE8) was produced in 90% of ALL samples, as well as in ALL cell lines. PAX5‐ΔE8 lacked more than 50% of the transactivation domain, indicating that aberrant Pax5 production might lead to the arrest of B‐cell differentiation, contributing to the pathogenesis of B‐lineage ALL.

Down-regulation of CD5 expression on activated CD8+ T cells in familial hemophagocytic lymphohistiocytosis with perforin gene mutations

Hemophagocytic lymphohistiocytosis (HLH) is characterized by uncontrolled activation of T cells and macrophages with overproduction of cytokines. Familial HLH type 2 (FHL2) is the most common form of primary HLH and is caused by mutations in PRF1. We have recently described a significant increase in the subpopulation of CD8(+) T cells with clonal expansion and CD5 down-regulation in Epstein-Barr virus associated-HLH, which represented a valuable tool for its diagnosis. However, this unusual phenotype of CD8(+) T cells has not been investigated fully in patients with FHL2. We performed immunophenotypic analysis of peripheral blood and measured serum pro-inflammatory cytokines in five patients with FHL2. All patients showed significantly increased subpopulations of activated CD8(+) T cells with down-regulation of CD5, which were negligible among normal controls. Analysis of T-cell receptor Vβ repertoire suggested the reactive and oligoclonal expansion of these cells. The proportion of the subset declined after successful treatment concomitant with reduction in the serum levels of cytokines in all patients except one who continued to have a high proportion of the subset and died. These findings suggest that down-regulation of CD5 on activated CD8(+) T cells may serve as a useful marker of dysregulated T cell activation and proliferation in FHL2.

Diagnostic challenge of Diamond–Blackfan anemia in mothers and children by whole-exome sequencing

Diamond–Blackfan anemia (DBA) is a pure red cell aplasia that arises from defective ribosomal proteins (RPs). Patients with this rare ribosomopathy present with neonatal anemia and occasional dysmorphism. Clinical heterogeneity and clusters of causative RP genes hamper the diagnosis and perinatal management. We report three mother-and-child pairs of anemia who were finally diagnosed by whole-exome sequencing. Each pair showed distinct disease severity and response to anemia treatment. Only one mother had the diagnostic dysmorphism, including short stature, webbed neck, and thenar hypoplasia. This mother had a frame-shift mutation of RPL11 (exon 3, c.58_59del). Her infant showed transient neonatal anemia, but had no mutations of RP genes. The other mother–child pairs had a missense mutation of RPS19 (exon 4, c.185G>A), and a splicing error of RPS7 (exon 3, c.76-1G>T), respectively. Other than the reported mutations, there were no variants in genes significantly associated with anemia. Our results suggested that whole-exome sequencing (WES) is effective for achieving a prompt and correct diagnosis of human ribosomopathy.

High-dose Cepharanthin for pediatric chronic immune thrombocytopenia in Japan

A nationwide, multicenter and observational study was retrospectively conducted to evaluate the clinical utility of Cepharanthin (CEP) for pediatric patients with chronic immune thrombocytopenia (ITP).

A case of subacute combined degeneration of the spinal cord due to folic acid and copper deficiency

Subacute combined degeneration of the spinal cord (SACD) is a rare neurologic disorder manifesting progressive symptoms of paresthesia and spastic paralysis. Herein we present an autopsy case of SACD caused by folic acid and copper deficiency. A 16-year-old male presented with gradually worsening unsteady gait, and bladder and rectal dysfunction. He had a medical history of T-cell acute lymphoblastic leukemia (T-ALL), diagnosed 1.5 years previously. The patient had undergone chemotherapy, including methotrexate, as well as allogeneic bone mallow transplantation. Laboratory tests revealed normal vitamin B12 and methylmalonic acid concentration, but reduced serum copper, ceruloplasmin and folic acid concentrations. Magnetic resonance imaging revealed symmetrical T2 signal hyperintensities in the posterior and lateral spinal cord. The patient was treated with oral copper, oral folate, and intravenous vitamin B12. A month after this treatment, the patients symptoms were unchanged, and 2 months later he died of acute adrenal insufficiency. The pathological findings of the spinal cord were compatible with SACD. Because SACD is usually reversible with early treatment, it should be suspected in high-risk patients undergoing chemotherapy or those who are malnourished with characteristic symptoms of SACD, even in young patients.

Anaplastic Lymphoma Kinase-Positive Anaplastic Large Cell Lymphoma with Cardiac Metastasis and Arterial Tumor Embolisms during First-Course Chemotherapy.

We described an 11-year-old boy suffering from pediatric anaplastic lymphoma kinase-positive anaplastic large cell lymphoma with heart metastasis at diagnosis and arterial tumor embolisms during chemotherapy. Both the heart metastasis and pericardial effusion showed improvement with prednisolone, but numbness and pallor sequentially developed in his lower extremities during the first course of chemotherapy. Contrast-enhanced imaging revealed occlusion of the right anterior tibial artery and left popliteal artery. These symptoms were spontaneously remitted due to the compensation of other arteries. Arterial tumor embolism is a rare but possible complication when a lymphoma shows intracardiac infiltration.