Akihiko Yokohama

Evidence for discrete stages of human natural killer cell differentiation in vivo

Human natural killer (NK) cells originate from CD34(+) hematopoietic progenitor cells, but the discrete stages of NK cell differentiation in vivo have not been elucidated. We identify and functionally characterize, from human lymph nodes and tonsils, four NK cell developmental intermediates spanning the continuum of differentiation from a CD34(+) NK cell progenitor to a functionally mature NK cell. Analyses of each intermediate stage for CD34, CD117, and CD94 cell surface expression, lineage differentiation potentials, capacity for cytokine production and natural cytotoxicity, and ETS-1, GATA-3, and T-BET expression provide evidence for a new model of human NK cell differentiation in secondary lymphoid tissues.

The Usefulness of 18 F-fluorodeoxyglucose Positron Emission Tomography ( 18 F-FDG-PET) and a Comparison of 18 F-FDG-PET With 67 Gallium Scintigraphy in the Evaluation of Lymphoma Relation to Histologic Subtypes Based on the World Health Organization Classification

Although studies comparing conventional imaging modalities with 18F‐fluorodeoxyglucose positron emission tomography (18F‐FDG‐PET) for the detection of lymphoma and although the relations between 18F‐FDG‐PET and histologic types were reported previously, most studies were not systematic and involved relatively small numbers of patients.

Steroid‐refractory chronic idiopathic thrombocytopenic purpura associated with hepatitis C virus infection

Abstract: Objectives: Hepatitis C virus infection has often been suggested as a possible cause of various kinds of autoimmune diseases. The aim of this study was to determine the relationship between chronic idiopathic thrombocytopenic purpura (ITP) and hepatitis C virus infection and to characterize the clinical features of anti‐HCV antibody (HCVab) positive chronic ITP patients. Subjects and methods: We studied HCVab in 79 patients with chronic ITP (25 males, 54 females, mean age 42.3 yr, range 11–86 yr) using the third‐generation ELISA method. Results: HCVab was detected in 11 of the 79 patients (13.9%). Quantitative HCV‐RNA studies showed a high serum concentration of HCV‐RNA in these patients. The platelet counts in these 11 HCVab‐positive patients (Group 1) were lower than in the 68 HCVab‐negative patients (Group 2)[(2.6 ± 0.9) versus (4.9 ± 3.0) × 1010/L, respectively; p<0.02]. Significantly more patients in Group 1 required prednisolone therapy (10/11, 90.9%) than in Group 2 (31/68, 45.6%) (P < 0.005). The response rate to prednisolone treatment was significantly higher in Group 2 (19/31, 61.3%) than in Group 1(0/10, 0%) (P < 0.001). There was no difference in the response to splenectomy between Groups 1 (4/7, 57.1%) and 2 (3/5, 60%). Conclusion: Given these findings, we recommend that HCVab is measured upon diagnosis of chronic ITP, and that splenectomy is planned in patients with HCVab in the event that prednisolone treatment is ineffective.

Expression of ABO blood-group genes is dependent upon an erythroid cell-specific regulatory element that is deleted in persons with the B(m) phenotype.

The ABO blood group is of great importance in blood transfusion and organ transplantation. However, the mechanisms regulating human ABO gene expression remain obscure. On the basis of DNase I-hypersensitive sites in and upstream of ABO in K562 cells, in the present study, we prepared reporter plasmid constructs including these sites. Subsequent luciferase assays indicated a novel positive regulatory element in intron 1. This element was shown to enhance ABO promoter activity in an erythroid cell-specific manner. Electrophoretic mobility-shift assays demonstrated that it bound to the tissue-restricted transcription factor GATA-1. Mutation of the GATA motifs to abrogate binding of this factor reduced the regulatory activity of the element. Therefore, GATA-1 appears to be involved in the cell-specific activity of the element. Furthermore, we found that a partial deletion in intron 1 involving the element was associated with B(m) phenotypes. Therefore, it is plausible that deletion of the erythroid cell-specific regulatory element could down-regulate transcription in the B(m) allele, leading to reduction of B-antigen expression in cells of erythroid lineage, but not in mucus-secreting cells. These results support the contention that the enhancer-like element in intron 1 of ABO has a significant function in erythroid cells.

JAK2-V617F mutation analysis of granulocytes and platelets from patients with chronic myeloproliferative disorders: advantage of studying platelets

There have been conflicting reports over the JAK2‐V617F mutation status of platelets in chronic myeloproliferative diseases (CMPDs). The aim of this study was to analyse JAK2‐V617F status, not only in granulocytes but also in platelets. The JAK2‐V617F mutation was analysed in both granulocytes and platelets in 115 patients with CMPDs using direct sequencing. JAK2‐V617F was detected in granulocytes from 71 of those patients, all 71 of whom also had platelet JAK2‐V617F expression. The remaining 44 patients showed negative JAK2‐V617F expression on granulocytes, but positive JAK2‐V617F expression was detected on the platelets from nine of the 33 essential thrombocythaemia (ET) patients, one of the eight polycythaemia vera patients, and two of the three primary myelofibrosis patients. When ET patients were divided into three groups according to granulocyte and platelet JAK2‐V617F status (both‐positive, platelets‐only positive and both‐negative), the both‐positive and platelets‐only positive groups shared the clinical features of higher white blood cell count and frequent thrombosis. These results suggest that analysis of platelets is a more sensitive approach for detecting JAK2‐V617F in CMPD patients than analysis of granulocytes. They also suggest that previous reports of the incidence of JAK2‐V617F in CMPD patients, obtained using only analysis of granulocytes, could be underestimations.

Interleukin-17F gene polymorphism in patients with chronic immune thrombocytopenia

Introduction:  IL‐17F is a novel inflammatory cytokine and plays an important role in some autoimmune diseases. We investigated the association between chronic ITP and the frequency of the single‐nucleotide polymorphism rs763780 (7488T/C), which causes a His‐to‐Arg substitution at amino acid 161.

Clinical significance of granulocytic sarcoma in adult patients with acute myeloid leukemia

To investigate the clinical significance of granulocytic sarcoma (GS) in adults with acute myeloid leukemia (AML), 434 consecutive patients with AML were analyzed retrospectively. Forty‐five patients (10.4%) with GS at diagnosis were younger (P < 0.001), presented with higher white blood cell counts (P = 0.03) and were more likely to conform to French–American–British M4 (P = 0.001) and M5 (P = 0.045) classifications than those without GS. In contrast, no significant difference in frequency of cytogenetic abnormalities was found between the GS and non‐GS groups. Treatment outcomes in 260 patients (40 with GS) who underwent intensive chemotherapy, excluding patients with acute promyelocytic leukemia, were investigated. Complete remission rates did not differ significantly between the GS and non‐GS groups (75.0% vs 79.1%; P = 0.192, respectively) or the 5‐year overall survival (OS) rates (39.9% vs 38.7%; P = 0.749, respectively). However, the GS group had a significantly higher relapse rate than the non‐GS group (74.2% vs 55.3%; P = 0.048) and a significantly lower 5‐year disease‐free survival rate (8.2% vs 25.7%, respectively; P = 0.005). When considered together with the results of multivariate analysis, which identified the presence of GS as an independent predictor for disease‐free survival time, these findings indicate that GS might identify a high‐risk subset of patients with AML. (Cancer Sci, doi: 10.1111/j.1349‐7006.2012.02324.x, 2012)

Analysis of the distribution of CAG repeats and X-chromosome inactivation status of HUMARA gene in healthy female subjects using improved fluorescence-based assay.

We investigated the polymorphic CAG-repeat distribution and the X-inactivation status of the human androgen receptor (HUMARA) gene in 58 female Japanese volunteers. Polymerase chain reaction amplification was performed using a fluorescent-dye-labeled primer under conditions specific for GC-rich targets, and fragments were analyzed. To estimate the length of these fragments, FAM-labeled (blue fluorescent) products were simultaneously compared with ROM-labeled size markers (red) that were created by sequencing various HUMARA fragments. The number of polymorphic CAG repeats of HUMARA in 116 alleles from 58 female subjects ranged from 15 to 28. Of the 58 volunteers, 51 (88.0%) were heterozygous. In 96% of the heterozygous female subjects, the allelic differences were no greater than 6 repeats. X-chromosome inactivation was calculated as the ratio of the area of the smaller peak to the sum of the areas of the smaller and larger peaks. The average ratio was 0.38 (range, 0.09-0.50). Preferential use of 1 allele, by more than 75% (ratio, <0.25), was observed in 5 volunteers (10.9%). The clonal nature of a patient with chronic myelogenous leukemia was easily identified. This method is sensitive enough to discriminate a difference of 1 triplet repeat.

Philadelphia chromosome‐positive mixed phenotype acute leukemia in the imatinib era

Although the introduction of imatinib dramatically improved the outcomes for patients with Philadelphia chromosome‐positive B‐cell precursor acute lymphoblastic leukemia (Ph+BCP‐ALL), the survival benefit of imatinib has not been assessed in the context of Ph+ mixed phenotype acute leukemia (Ph+MPAL). To clarify this important issue, we studied 42 Ph+ acute leukemia (Ph+AL) patients who received intensive chemotherapy and concurrent administration of imatinib. Of the 42 Ph+AL patients, 13 (31%) patients were categorized as Ph+MPAL (positive for both myeloid and B‐cell lineage), 27 (64%) were categorized as Ph+BCP‐ALL, and two (5%) were categorized as Ph+ acute myeloid leukemia. The complete remission rates after the initial induction therapy were not significantly different when comparing Ph+MPAL and Ph+BCP‐ALL patients (100% vs. 85%, respectively, P = 0.14). Likewise, there were no significant differences in the 5‐yr overall survival (OS) or disease‐free survival (DFS) rates when comparing the MPAL and BCP‐ALL groups (OS: 55% vs. 53%, respectively, P = 0.87, DFS: 46% vs. 42%, respectively, P = 0.94). These findings suggest that concurrent imatinib administration with chemotherapy improved the outcomes of Ph+MPAL patients to the level seen in Ph+BCP‐ALL patients and should, therefore, be considered as the standard therapy for these patients.

Circulating plasmacytoid dendritic cells in patients with primary and Helicobacter pylori-associated immune thrombocytopenia.

Objectives:  Immune thrombocytopenia (ITP) is an autoimmune disorder characterized by the production of autoreactive antibodies against platelet antigens. Although dysfunction of multiple aspects of cellular immunity is considered to be important in the pathogenesis of ITP, it has not been clarified which cell types play a principal role.