Yoshiyuki Niho

Primary Chronic Myelofibrosis: Clinical and Prognostic Evaluation in 336 Japanese Patients

We retrospectively analyzed 336 patients with primary chronic myelofibrosis from 203 medical institutes in Japan. Notwithstanding their heterogeneous treatments, the median survival in 298 patients that could be evaluated was 10.0 years. Average age at onset was 60.7 years. Men were affected 1.4 times more frequently than women. The factors associated with shorter survival included anemia, leukocytosis/leukocytopenia, thrombocytopenia, and increased blasts in the peripheral blood, and sex (male), age (>60), and the presence of symptoms. A new scoring system based on the peripheral blood findings (hemoglobin [Hb] level, platelet count, and rate of blast formation) at initial diagnosis clearly correlated with survival rate. Accordingly, patients could be categorized into 3 groups by severity grading. Through stepwise multivariate survival analysis, the significant prognostic factors were identified as the severity grading based on the new scoring system (P = .0002), age (P = .0024), sex (P = .0153), and Hb (P = .0198). Chromosomal abnormalities were found in 58 of 154 patients (38%), although these did not influence survival.

Hair dye use and occupational exposure to organic solvents as risk factors for myelodysplastic syndrome

To investigate the relationships of personal hair dye use and environmental factors to myelodysplastic syndromes (MDS), we conducted a case-control study in Japan. A total of 111 MDS cases and 830 controls randomly selected from the residents in the same prefecture of cases using telephone directories responded to a health questionnaire. The odds ratio (OR) for ever having used hair dye was 1.99 (95% confidence interval (CI) 1.17-3.38) and there were statistically significant trends in risk with increasing duration and number of hair dye use. Occupational exposure to organic solvents was marginally associated with the risk of MDS (OR = 1.99; 95% CI 0.97-4.10).

Association of the interferon-γ receptor variant (Val14Met) with systemic lupus erythematosus

Abstract Genetic factors seem to play a significant role in susceptibility to systemic lupus erythematosus (SLE). The purpose of this study was to investigate whether the amino acid polymorphism (Val14Met) found within the IFN-γ receptor gene (IFNGR1) plays a prominent role in susceptibility to SLE. We found Val14Met located at the COOH terminal of the signal peptide of the IFN-γ receptor. There was a significant difference in this polymorphism frequency between SLE patients and healthy populations. To clarify whether this amino acid substitution resulted in the alteration of the receptor function, we evaluated the induction of HLA-DR antigen expression on B cells by IFN-γ stimulation. There was also a significant difference in the induction of HLA-DR by IFN-γ stimulation between B cells. Furthermore, an intracellular cytokine assay indicated that the Th1/Th2 balance of Th cells bearing the variant receptor shifted to Th2. The genetic polymorphism found within the IFN-γ receptor gene (Val14Met) may result in a shift to Th2, and this shift may increase susceptibility to SLE.

Role of IL-10 in the Crossregulation of Prostaglandins and Cytokines in Monocytes

In the present study we have focused mainly on the role of IL (interleukin)-10 in the crossregulation of prostaglandins and cytokines in human monocytes. We first determined the effects of tumor necrosis factor-α (TNF-α) and IL-10 on monocyte prostaglandin E2 (PGE2) production. Unstimulated monocytes constitutively produced a small but significant amount of PGE2 in the culture supernatants. Both TNF-α and lipopolysaccharide (LPS) caused a remarkable increase in monocyte PGE2 production. On the other hand, IL-10 alone was without effect on constitutive PGE2 production but drastically inhibited LPS-induced PGE2 production in monocytes. Moreover, this inhibitory effect of IL-10 was not simply attributable to its inhibition of TNF-α production in LPS-stimulated monocytes. Next, we determined the effect of PGE2 on TNF-α mRNA expression in monocytes. Treatment of monocytes with or without PGE2 showed no detectable TNF-α mRNA. Activation of monocytes by LPS resulted in a remarkable accumulation of TNF-α mRNA and PGE2 efficiently inhibited this expression. Finally, we determined the effect of PGE2 on IL-10 mRNA expression in monocytes. Similar to TNF-α mRNA, unstimulated monocytes showed no detectable IL-10 mRNA. Interestingly, PGE2 alone drastically induced IL-10 mRNA. Besides, activation of monocytes by LPS resulted in a remarkable accumulation of IL-10 mRNA, and PGE2 further enhanced this expression. These results indicate that TNF-α and PGE2 are key molecules for the induction of IL-10 in monocytes, and that IL-10, in turn, plays a crucial role in terminating the inflammatory cascade via downregulation of production of proinflammatory molecules including TNF-α and PGE2.

Immunosuppressive therapy for patients with refractory anemia.

Abstract. Trials of immunosuppressive therapy have been reported in some case reports of hypoplastic myelodysplastic syndrome (MDS). In this study, we gave immunosuppressive therapies to eight patients with normo- or hyperplastic MDS of refractory anemia subtype without karyotypic abnormalities and analyzed the HLA-DRB1 type or the presence of paroxysmal nocturnal hemoglobinuria (PNH) neutrophils in these patients. Cyclosporin A (CyA) therapy was effective for improving cytopenia in four of the eight MDS patients. While the side effects of CyA were mostly mild and transient, one patient demonstrated karyotypic abnormality following CyA therapy and accelerated to refractory anemia with an excess of blasts. Additional antithymocyte globulin (ATG) therapy was effective in one of three nonresponders to CyA therapy. One patient died due to leukemic transformation after ATG therapy. When we analyzed the correlation between the response to CyA therapy and the HLA-DRB1 type, there were more responders with DRB1*1501 (three of four patients) than without (one of four patients), but a statistically significant difference was not evident between the two groups. In addition, the presence of PNH neutrophils was not correlated with the response to CyA and/or ATG therapy. These results indicate the usefulness of immunosuppressive therapies even for normo- or hyperplastic MDS patients. Further trials using more patients with a long follow-up period would be worthwhile in order to clarify the possibility of disease progression and in order to predict the response of patients.

Autoreactive antibodies following autologous peripheral blood stem cell transplantation

A 35-year-old woman diagnosed with multiple myeloma (IgG, λtype, stage IIIA) received an autologous peripheral blood stem cell transplant (PBSCT). She was euthyroid without autoreactive antibodies prior to the transplant. The patient complained of malaise, weight loss and low grade fever 1 month after transplant, despite rapid haematopoietic recovery. Thyroid function tests on day 34 revealed hyperthyroidism associated with anti-thyroid peroxidase antibody. Antinuclear antibody was also detected, and platelet-associated immunoglobulin was increased. These findings disappeared spontaneously by day 62 without treatment. Autoimmune diseases may occur transiently after autologous PBSCT.

Constitutive activation of Jak-2 and Tyk-2 in a v-Src-transformed human gallbladder adenocarcinoma cell line

It is well known that v‐Src phosphorylates various substrates on tyrosine residue and associates with tyrosine‐phosphorylated proteins as well as proline‐rich ligands through its SH2 and SH3 domains, respectively, thereby inducing oncogenic transformation. A signal pathway from the cell surface to genes in the nucleus, the Jak/STAT (signal transducers and activators of transcription) pathway, has been shown to be involved in the signal transduction mechanism mediated by many cytokines and growth factors. Although a member of the STAT family, STAT3 has been reported to be constitutively activated in several v‐Src‐transformed cells, and it still remains unknown whether Jak molecules, which act upstream of STATs, are involved in the v‐Src‐induced activation mechanism of STAT3. In this study, we analyzed activations of both Jak and STAT molecules using v‐Src‐transformed HAG‐1 cells derived from a human gallbladder adenocarcinoma. STAT3 was found to be constitutively activated in v‐Src‐transformed HAG‐1 cells, but not in either non‐transformed mock‐transfected or activated c‐H‐ras‐transfected HAG‐1 cells, even though the other known STAT molecules are expressed. Furthermore, both Jak‐2 and Tyk‐2 were constitutively activated only in v‐Src‐transformed HAG‐1 cells. Association of v‐Src with either STAT3 or the Jak molecules was not observed. No change of this activation was detected by either interferon (IFN)‐α2a or IFN‐γ, which had shown inhibitory effects on the growth of v‐Src‐transformed HAG‐1 cells. These results raise the possibility that Jak‐2 and Tyk‐2 are both activated by v‐Src, thereby contributing to the constitutive activation of STAT3 in the v‐Src‐transformed cells. J. Cell. Physiol. 175:220–228, 1998.

Fas/Fas ligand and hematopoietic progenitor cells.

Fas antigen is a receptor that crosslinks with a ligand or antibody initiating a signal transduction cascade that leads to apoptosis. During normal hematopoiesis, Fas antigen is not expressed on CD34+ cells, including premature hematopoietic progenitor cells. Functional Fas antigen expression is induced by several hematopoietic regulators. These changes may appear not only in the process of differentiation of hematopoietic progenitor cells, but also as a negative feedback mechanism that controls chaotic proliferation of these cells. These findings suggest that the Fas/Fas ligand system is closely related to the maintenance of homeostasis during the process of normal hematopoiesis. Furthermore, increased Fas antigen expression is observed on CD34+ cells from patients with aplastic anemia, suggesting that it might cause bone marrow suppression. The use of Fas-mediated apoptosis of malignant cells as a tool for eliminating hematologic malignancies is promising. Increased Fas ligand expression is observed on natural killer lymphoma cells and may be associated with the pathogenesis of failure of several organs. The Fas/Fas ligand system plays an important role in the physiologic and pathologic processes of hematopoiesis. The development of treatments using this system are forthcoming.

Differential effects of interleukin-4 and interleukin-10 on nitric oxide production by murine macrophages

Abstract.Objective: To study the effect of interleukin (IL)-4 and IL-10 on nitric oxide (NO) production by macrophages.¶Materials and Methods: Elicited or resident peritoneal macrophages (PMO) and a macrophage cell line Raw264.7 were primed by IL-4 or IL-10 for 6 hours, and were further incubated in the presence of interferon (IFN)-γ and/or lipopolysaccharide (LPS) for 48 hours. NO2- accumulation in the supernatant of cultured cells was used as an indicator of NO production and was determined by the standard Griess reaction adapted for microplates. The amount of tumor necrosis factor (TNF)-α in the culture supernatants was determined with a commercially available ELISA kit. The absorbance was measured at 450 nm with a microplate photometer.¶Results: IL-4 inhibited NO production by murine macrophages of different sources and the macrophage cell line Raw264.7. In contrast, different macrophage populations showed differential responses to IL-10. After stimulation with LPS or IFN-γ, IL-10 suppressed NO production by elicited PMO but enhanced NO production by resident PMO or by Raw264.7. Both IL-4 and IL-10 inhibited the production of TNF-α, which has been shown to play a crucial role in NO production. In the presence or the absence of blocking antibody to TNF-α, IL-10 always enhanced NO production by resident PMO. This result suggests that the inhibition of TNF-α production and the enhancement of NO production by resident PMO stimulated with IL-10 are independent, coexisting events.¶Conclusions: Factors other than TNF-α have been suspected to influence NO production by macrophages, and this study indicates that IL-10 may be a candidate cytokine for resident PMO.

Differential gene expression of human telomerase-associated protein hTERT and TEP1 in human hematopoietic cells

The maintenance of telomere length is crucial for the survival of cells. Recently, genes for proteins that consist of human telomerase have been cloned and the results have indicated a close relationship between telomerase activity and its gene expression. We studied the mRNA expression of the telomerase-associated genes, hTERT and TEP1, in hematopoietic cells in order to clarify the relation between them and telomerase activity using semiquantitative RT-PCR. In polymorphonuclear cells and monocytes isolated from peripheral blood, which had no detectable telomerase activity, no hTERT mRNA expression was seen. On the other hand, lymphocytes and CD34-positive cells both demonstrated hTERT mRNA expression. TEP1 mRNA was detected in all samples, showing no differential expression. We then assessed hTERT and TEP1 mRNA expression in CD34-positive cells cultured in vitro with growth factors. After 4 weeks of culture, all the cells showed myeloid differentiation and the telomerase activity was downregulated. hTERT mRNA was expressed in CD34-positive cells, but was downregulated in 4-week-cultured cells. TEP1 showed no apparent differential expression. We conclude that hTERT mRNA expression is downregulated in accordance with telomerase downregulation during the course of myeloid differentiation, which suggests that it plays a crucial role in the expression of enzyme activity, while TEP1 has a much smaller role to play, if any.