Tamotsu Miyazaki

Cytokine gene expression in peripheral blood mononuclear cells during graft-versus-host disease after allogeneic bone marrow transplantation

Summary. Cytokine gene expression in peripheral blood mononuclear cells during the development of graft‐versus‐host disease (GVHD) in patients who underwent allogeneic bone marrow transplantation (allo BMT) was analysed using a semiquantitative reverse‐transcriptase polymerase chain reaction (RT‐PCR). The expression of interleukin (IL)‐lβ, IL‐6, and tumour necrosis factor (TNF)‐α mRNA was increased during the development of GVHD and the degree of this increment depended on the severity of the disease. IL‐2 expression was not detected at all and interferon‐γ expression was not much changed during GVHD. In patients with hepatic veno‐occlusive disease (VOD), another transplantation‐related complication, the expression of IL‐1β and TNF‐a mRNA was increased but IL‐6 mRNA expression showed little increase. These findings suggest that IL‐lβ, IL‐6 and TNF‐α produced by peripheral blood mononuclear cells play an important role in the development of GVHD. Furthermore, liver dysfunction due to GVHD or VOD may be distinguishable by this type of cytokine analysis. Analysis of cytokine mRNA expression in peripheral blood mononuclear cells after allogeneic bone marrow transplantation may provide important information concerning the immune response and the cytokine network system in marrow transplant patients.

Efficacy of granulocyte colony‐stimulating factor in the treatment of acute myelogenous leukaemia: a multicentre randomized study

Summary. To investigate the efficacy and safety of granulocyte colony‐stimulating factor (G‐CSF) in patients with acute myelogenous leukaemia, a multicentre randomized study was performed. From October 1993 to September 1996, 270 patients with newly diagnosed acute myelogenous leukaemia were randomized to G‐CSF or control groups after remission induction therapy. The G‐CSF group received G‐CSF (Filgrastim) from 48 h after the completing chemotherapy until the absolute neutrophil count exceeded 1·5 × 109/l. The control group did not receive G‐CSF unless severe infection occurred. There were 245 evaluable patients (120 and 125 in the G‐CSF and control groups respectively). The complete remission rate was similar in the G‐CSF and control groups (80·8% versus 76·8%), as was the 5‐year probability of disease‐free survival (34·5% versus 33·6%) and overall survival (42·7% versus 35·6%). Neutrophil recovery was significantly faster in the G‐CSF group than in the control group (12 d versus 18 d, P = 0·0001). The median duration of febrile neutropenia was significantly shorter in the G‐CSF group than in the control group (3 d versus 4 d, P = 0·0001). In conclusion, prophylactic administration of G‐CSF after remission induction therapy for acute myelogenous leukaemia is safe and useful even in patients without infection on completing chemotherapy.

Long-term Results of a Multicenter Randomized, Comparative Trial of Modified CHOP versus THP-COP versus THP-COPE Regimens in Elderly Patients with Non-Hodgkin's Lymphoma

In treating elderly non-Hodgkin’s lymphoma (NHL) patients, it is particularly important to use drugs that have a low incidence of adverse events and high efficacy. In this multicenter study, THP (pirarubicin)-COP (cyclophosphamide, vincristine, and prednisolone) was compared to two thirds dosage of full CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisolone) regimen with regard to both adverse events and efficacy. For a third group, etoposide (E) was added to the THP-COP regimen (THP-COPE) in order to achieve high dose-intensity. Subjects were 486 previously untreated patients, aged 65 or older (range, 65–92 years; median, 74 years), with NHL. Subjects were randomly assigned to receive THP-COP, two thirds CHOP, or THP-COPE. Four hundred and forty-three patients were assessed for response and followed for 8 years after the last subject registered. The complete remission rates for the THP-COP, CHOP, and THP-COPE groups were 42.5%, 41.4%, and 48.0%, respectively. There was no difference in overall survival or progression-free survival among these 3 groups. In aggressive lymphoma, there was also no difference in complete response (CR) rate (45.3% in THP-COP, 44.9% in CHOP, 48.0% in THP-COPE), overall survival, and progression-free survival among these groups. The 5- and 8-year survival rates for all patients were 29.4% and 18.7%, respectively. The 5- and 8-year survival rates for patients with aggressive lymphoma were 27.4% and 17.4%, respectively. Although long-term survival for patients with aggressive lymphoma on our regimens was not worse compared to previous reports, the CR rate was lower. Because severe adverse events were not observed, higher dose chemotherapy may be directed to achieve better CR rates. In patients with T-cell-type lymphoma, the CR rate was greater after treatment with THP-COP (51.4%) or THP-COPE (57.7%) compared to treatment with CHOP (19.4%). Pirarubicin may be more useful for T-cell lymphoma than doxorubicin. Because adverse cardiac events were reported only in CHOP, adverse cardiac events might be low in the THP group.

Establishment of a novel granulocytic sarcoma cell line which can adhere to dermal fibroblasts from a patient with granulocytic sarcoma in dermal tissues and myelofibrosis

Summary A novel human myeloid cell line, designated HSM‐1, has been established from the pleural effusion of a patient with granulocytic sarcoma (GS) who had been followed as having primary myeiofibrosis for 10 years. When he was diagnosed as having granulocytic sarcoma in dermal tissues, no evidence of malignant transformation into leukaemia was found in both the peripheral blood and bone marrow. The established cell line was positive for myeloperoxidase, Sudan black B. Naphthol AS‐D chloroacetate esterase. Surface marker analysis revealed that HSM‐1 expressed CD4. CD13, CD11a, CDllb, Leu8. CD49b. CD49d, CD49e, CD29 and HLA‐DR.

Cytokine Gene Expression after Allogeneic Bone Marrow Transplantation

Cytokines produced by T lymphocytes, monocytes/macrophages, and fibroblasts play a central role in the immune response and in the development of graft-versus-host disease (GVHD). Also, it has been reported that dysregulated production of cytokines maybe the primary mediator of clinical manifestation of acute GVHD. Regarding cytokine gene expression after human allogeneic bone marrow transplantation (allo BMT), we have demonstrated increased IL-1 beta, IL-6, and TNF-alpha mRNA expression in peripheral blood mononuclear cells during the development of acute and chronic GVHD and that the degree of the increase was dependent on the severity of the disease. Furthermore, overexpression of these cytokine mRNAs could be detected before the clinical manifestations of GVHD developed. In contrast, IL-2 mRNA expression was not detected in peripheral blood mononuclear cells in GVHD patients. On the other hand, we have reported that increased mRNA expression and protein product of IL-2 and IFN-gamma were evident in the mixed lymphocyte culture of the cases who developed severe lethal transplantation-related complications. Therefore, the detection of increased IL-2 and IFN-gamma gene expression in MLC appeared to be useful for predicting transplantation-related complications in BMT patients. Furthermore, we found increased IL-2 receptor alpha subunit mRNA expression in the peripheral blood mononuclear cells during GVHD. These findings may indicate the important role of inflammatory cytokines such as IL-1 beta, IL-6 and TNF-alpha in the development of the clinical manifestation of GVHD and also may be indicative of the important role of IL-2 and the IL-2 receptor in allo response perhaps mainly as an autocrine effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation of mixed chimaerism and origin of bone marrow derived fibroblastoid cells after allogeneic bone marrow transplantation

Summary We assessed the origin of bone marrow derived fibroblastoid cells (BMF) in long‐term cultures of 13 samples obtained from nine patients after allo‐BMT by polymerase chain reaction (PCR) amplification of MCT118, one of the variable number of tandem repeats regions (VNTR). BMF showed a complete recipient pattern in nine samples obtained from seven patients; however, a recipient‐predominant mixed chimaeric pattern was detected in BMF from four patients. Also, two of the four patients died with bone marrow hypoplasia. These data suggest that mixed chimaeric pattern of BMF may be correlated with bone marrow hypoplasia.

Cytokine gene expression in the mixed lymphocyte culture in allogeneic bone marrow transplants as a predictive method for transplantation-related complications

SUMMARY. We have investigated cytokine gene expression in the two‐way mixed lymphocyte cultures (MLC) enhanced by concanavalin A to assess whether this is a useful predictive method for severe graft‐versus‐host disease (GVHD) and graft failure in nine allogeneic bone marrow transplantation (allo‐BMT) patients. Our present study revealed that increased IL‐2, IL‐5 and IFN‐γ mRNA expression and IL‐2 and IFN‐γ production in the MLC in two cases with severe lethal transplantation‐related complications (graft failure and grade III acute GVHD). These findings suggest that increased cytokine mRNA expression and cytokine products in this assay may be predictive of the development of transplantation‐related complications.

Dipyridamole potentiates the anti-aggregating effect of endothelium-derived relaxing factor

One effector of the anti-aggregatory property of endothelium is thought to be endothelium-derived relaxing factor. The best characterized of these, nitric oxide, inhibits platelet aggregation by increasing cyclic GMP levels. The effects of nitric oxide and dipyridamole (a cyclic GMP phosphodiesterase inhibitor), alone and in combination, on in vitro platelet aggregation were evaluated. Dipyridamole had no effect per se on platelet aggregation but potentiated the inhibition of aggregation due to nitric oxide. This was concomitant with an increase in platelet cyclic GMP concentration. The author suggests an alternative mechanism for the clinical efficacy of dipyridamole as an antiplatelet agent.

Radioimmunoassay of aldolase A. Determination of normal serum levels and increased serum concentration in cancer patients.

A radioimmunoassay specific for human aldolase A subunits was used to measure human aldolase A (ALD‐A) in human serum. The double antibody competitive inhibition radioimmunoassay technique used radioiodinated purified ALD‐A as ligand, chicken antisera specific for human ALD‐A and rabbit antichicken IgG. The serum levels of ALD‐A in 42 normal healthy subjects ranged from 130 to 210 ng/ml (mean average, 171 ± 39 ng/ml). In 177 hospitalized patients without cancer, muscle diseases, or hemolytic anemia, the ALD‐A serum levels ranged from 125 to 220 ng/ml. In contrast, 82% of 260 patients with various types of malignancy had ALD‐A serum concentrations above the normal range. The CEA levels increased only 44% of the sera of 80 patients with cancer of the digestive tract, whereas the ALD‐A levels were increased in 86% of the patients. The AFP levels were greater than 100 ng/ml in only 70% of the sera of 33 liver cell carcinoma patients, whereas the ALD‐A levels were increased in 94% of these sera. The measurement of serum ALD‐A by radioimmunoassay may be a valuable adjunct in the clinical diagnosis of certain cancer patients.

Components and proteolytic processing sites of arylsulfatase B from human placenta

Previous studies have shown that mature arylsulfatase B purified from human sources is composed of two non-identical chains with apparent molecular masses of 43 kDa and 8 kDa. Arylsulfatase B purified from human placenta in the present study, however, included another 7 kDa component that could be detected only by carbohydrate staining on reducing SDS-PAGE employing the Tris-Tricine system. The 43 kDa and 7 kDa components contained a carbohydrate moiety, but the 8 kDa one did not, as demonstrated by periodic acid-Schiff staining, Con-A lectin blotting, endo-glycosidase treatment and in vitro phosphorylation by UDP-N-acetylglucosamine: lysosomal enzyme N-acetylglucosamine 1-phosphotransferase. The purified arylsulfatase B migrated as a single polypeptide of 58 kDa on non-reducing SDS-PAGE, indicating that the three chains are linked by disulfide bonds. In order to determine the origin of the components, N-terminal sequencing of the isolated polypeptides was performed. As a result, the 43, 7 and 8 kDa components were found to commence with Ala-41, Ala-424 and Asp-466, respectively. These results suggest that after removal of the signal peptide, human arylsulfatase B undergoes proteolytic processing on at least two sites during maturation.