Shigenori Kanazawa

Effects of efonidipine on platelet and monocyte activation markers in hypertensive patients with and without type 2 diabetes mellitus

We compared the levels of microparticles, platelet activation markers, soluble cell adhesion molecules, and soluble selectins between hypertensive patients with and without type 2 diabetes and control subjects. Binding of anti-glycoprotein IIb/IIIa and anti-glycoprotein Ib monoclonal antibodies to platelets did not differ significantly between the hypertensive patients and controls, but platelet expression of activation markers (CD62P, CD63, PAC-1, and annexin V) was higher in the hypertensive patients. Platelet-derived microparticle (PDMP) and monocyte-derived microparticle (MDMP) levels were significantly higher in the hypertensive patients than in the controls. Soluble ICAM-1, VCAM-1, P-selectin, and E-selectin levels were also higher in the hypertensive patients, and they were significantly higher in the hypertensive patients with diabetes. After treatment with efonidipine, the levels of PDMPs, CD62P-, CD63-, PAC-1-, and annexin V-positive platelets, sICAM-1, sVCAM-1, sP-selectin, and sE-selectin all decreased significantly. The MDMP levels decreased, and the decrease was significant in the hypertensive patients with diabetes. These findings suggest that administration of efonidipine to hypertension patients with diabetes may prevent the development of cardiovascular complications caused by cell adhesion molecules or activated platelets and monocytes.

Effects of eicosapentaenoic acid on platelet activation markers and cell adhesion molecules in hyperlipidemic patients with Type 2 diabetes mellitus

We compared the levels of microparticles, platelet activation markers, soluble cell adhesion molecules, soluble selectins, and antioxidized low-density lipoprotein (anti-Ox LDL) antibody between patients with hyperlipidemia and control subjects. Binding of anti-glycoprotein (GP) IIb/IIIa and anti-GPIb monoclonal antibodies to platelets did not differ significantly between the hyperlipidemic patients and controls. However, expression of activation markers (CD62P, CD63, PAC-1, and annexin V) by platelets was higher in the hyperlipidemic patients with Type 2 diabetes. The levels of platelet-derived microparticles (PDMPs) and monocyte-derived microparticles (MDMPs) were significantly different in hyperlipidemic patients with Type 2 diabetes and controls. Soluble P-selectin (sP-selectin), soluble E-selectin (sE-selectin), and anti-Ox LDL antibody also showed higher levels in the hyperlipidemic patients with Type 2 diabetes. After treatment with eicosapentaenoic acid (EPA), the levels of CD62P, CD63, annexin V, PDMPs, and MDMPs, sE-selectin, and oxidized LDL antibody were reduced significantly. Triglyceride (TG) and total cholesterol levels were also decreased. Anti-Ox LDL antibodies and MDMPs were correlated positively with platelet CD62P (plt-CD62P) levels. These findings suggest that in hyperlipidemic patients with Type 2 diabetes, EPA may prevent complications caused by oxidized LDL, E-selectin, and activated platelets or monocytes.

Monocyte-derived microparticles may be a sign of vascular complication in patients with lung cancer

We measured and compared the levels of plasma monocyte-derived microparticles (MDMP) and platelet activation markers [plasma platelet-derived microparticles (PDMP), CD62P binding to platelets; plt-CD62P, CD63 binding to platelets; plt-CD63], to develop a better understanding of their potential contribution to vascular complications of lung cancer. The concentrations of MDMP and PDMP in lung cancer patients were significantly higher (P < 0.01) than those in normal subjects. Levels of plt-CD62P and plt-CD63 were significantly higher (P < 0.001 for each) in lung cancer patients than in controls. Levels of sE-selectin were also higher in lung cancer patients than in control subjects. MDMP correlated positively with plt-CD62P, plt-CD63, and PDMP with its relation to PDMP being particularly significant. The number of MDMPs and PDMPs are patients who are non-small cell lung cancer were significantly higher than that in small cell lung cancer patients. In addition, levels of sE-selectin were higher in non-small cell lung cancer than in small cell lung cancer patients. These findings suggest that elevated MDMPs may be a sign of vascular complication in lung cancer patients, particularly those who suffer from non-small cell lung cancer.

The effects of Helicobacter pylori eradication on chemokine production in patients with immune thrombocytopenic purpura.

To the Editor: Immune thrombocytopenic purpura (ITP) is an autoimmune disease caused by circulating antibodies that react with platelet membranes (1). Emilia et al. (2) reported a high incidence of Helicobacter pylori (HP) infection in patients with ITP as well as a significant increase in platelet count after bacterium eradication. Although the pathogenetic mechanisms of HP-induced thrombocytopenia remain unknown, HP causes an immunological response resulting in the production of high concentrations of pro-inflammatory cytokines and chemokines in non-ITP patients (3). On the contrary, we recently reported on the significance of chemokines in patients with ITP (4). Monocyte chemoattractant protein-1 (MCP-1) and regulated on activation normally T-cell expressed and secreted (RANTES) were significantly increased in ITP. This elevation was thought to be related to the CD40/CD40 ligand system. We thus investigated the concentrations of different chemokines before and after HP eradication in ITP patients. Serum samples from 42 patients (15 men and 27 women) with chronic ITP were studied. Plateletassociated immunoglobulin G (IgG) was measured by a competitive enzyme-linked immunosorbent assay. Platelet counts and platelet-associated IgG were 31 ± 6 (·10/L) and 134 ± 12 (ng/10 plt), respectively. The patients had not received any transfusions, but 21 had previously been treated for ITP (splenectomy in six patients and prednisolone therapy in 17 patients). The urea breath test was used to determine whether or not patients were infected with HP, and if found to be infected, treatment to eradicate HP included amoxicillin (1000 mg twice daily), clarithromycin (400 mg twice daily), and omeprazole (20 mg twice daily) for 1 wk. The prednisolone treatment was not changed during the eradication therapy, which was evaluated at 8 wk following treatment. We measured human 3 CC chemokines (MCP-1, RANTES, and Eotaxin) and 3 CXC chemokines [interleukin-8 (IL-8), epithelial cell-derived neutrophil attractant78; ENA-78, and stromal cell-derived factor-1; SDF-1] because HP influences both CC and CXC chemokine (5, 6) levels, which were measured with a monoclonal antibody-based ELISA kit from BioSourse International Inc. (Camarillo, CA, USA) and R & D Systems (Minneapolis, MN, USA) according to the manufacturer’s instructions. HP infection was found in 28 of 42 chronic ITP patients (66.7%). After eradication, 15 of the 28 patients (53.6%) had a significant increase in platelet count (increase of greater than 50 · 10/L vs. pretreatment platelet count), and the eradication rate was 42.9% (12/28). This eradication rate was low compared with the average rate usually obtained after a standard eradication regimen. This reason is unknown. However, one of them could be an ethnic difference. There were no significant differences in concentrations of IL-8, SDF-1 and Eotaxin between HP-positive and HP-negative ITP patients (Table 1). However, significant increases in

Elevation of Platelet Activation Markers and Chemokines during Peripheral Blood Stem Cell Harvest with G-CSF

The kinetics of peripheral blood stem cell mobilization in response to recombinant human granulocyte colony‐stimulating factor is well established. However, there have been few investigations of platelet activation markers during peripheral blood stem cell harvest. We measured the levels of the platelet activation markers, chemokines, and soluble factors in plasma obtained from patients undergoing peripheral blood stem cell harvest. The number of leukocytes, CD34+ cells, neutrophils, monocytes, and lymphocytes peaked on day 5 after granulocyte colony‐stimulating factor treatment, but the numbers of eosinophils and basophils showed no significant change. Regulated on activation normally T‐cell expressed and secreted (RANTES) level increased through day 10, and the monocyte chemotactic peptide‐1 (MCP‐1) level peaked on day 5. Platelet counts continued to increase through day 10. The level of thrombopoietin significantly increased on day 3, peaked on day 5, and decreased slightly by day 10. The levels of soluble CD40 ligand and soluble P‐selectin increased up to day 5. The platelet‐derived microparticle level peaked on day 5, and then began to decline. CD34+ cell numbers significantly correlated with those of leucocytes, neutrophils, monocytes, and lymphocytes, as well as levels of MCP‐1, and the CD34+ cells exhibited changes similar to platelet‐derived microparticles. The patterns of change in MCP‐1, platelet‐derived microparticles, and the CD34+ cell count are similar in that each peaks on day 5 and decreases thereafter. Further study is required to determine if a cause‐and‐effect relationship in their pattern of change exists among them.

Significance of Anti-oxidized LDL Antibody and Monocyte-derived Microparticles in Anti-phospholipid Antibody Syndrome

Monocytes, platelets, endothelial cells and oxidized LDL could be very important in development of vascular complication in thrombotic diseases. We measured and compared the levels of plasma monocyte-derived microparticles (MDMPs), platelet-derived microparticles (PDMPs), and anti-oxidized LDL antibody, to develop a better understanding of their potential contribution to vascular complications in antiphospholipid antibody syndrome (APS). The concentration of MDMP in APS patients was significantly higher (p<0.01) than that in normal subjects and SLE patients. When levels of PDMPs and plt-P-selectin were compared between the control and APS patients, levels of PDMPs and plt-P-selectin were significantly higher (p<0.01 for each) in APS patients than in controls. In addition, these levels of platelet activation markers correlated with MDMP in APS patients. Twenty one of the 37 APS patients (56.8%) had elevated levels of anti-oxLDL antibody. In addition, a significant increase in MDMP was observed in anti-oxLDL antibody-positive APS patients (p<0.01). These findings suggest that elevated MDMPs may be a sign of vascular complication in APS patients, particularly those who are detected anti-oxLDL antibodies.

Significance of chemokines and soluble CD40 ligand in patients with autoimmune thrombocytopenic purpura

Abstract:  We investigated the levels of various chemokines and soluble CD40L (sCD40L) in ITP patients, in order to determine the influence of CD40–CD40L interaction on the pathogenesis of ITP. We found increases in MCP‐1 and RANTES levels in ITP patients compared with those in healthy individuals. Thirty‐eight of the 65 ITP patients (58.5%) had elevated levels of sCD40L. We found significant decreases in platelet counts in sCD40L‐positive ITP patients. Although the sCD40L level did not differ significantly between the control and nonimmune thrombocytopenia groups, but among ITP patients. sCD40L level was significantly higher in those with untreated ITP than in those with treated ITP. In addition, significant increases in RANTES, MCP‐1, sCD14, and sP‐selectin levels were observed in sCD40L‐positive ITP patients, although sE‐selectin levels were not increased in such patients. For other factors examined, however, there were no differences in level between sCD40L‐positive and ‐negative ITP patients. These findings suggests that there are two groups of ITP patients, one with elevated and one with normal of sCD40L. ITP cases in which sCD40L was increased appeared to involve changes in platelet counts and monocyte activation. The pathogenesis of ITP may in some patients include alterations of the CD40/CD40L pathway.

Gefitinib affects functions of platelets and blood vessels via changes in prostanoids balance.

Prostaglandins (PGs) and thromboxane (TX) produced by cyclooxygenase (COX) have a great influence on vascular systems and platelet functions. The serum levels of epidermal growth factor (EGF) and PGs were measured in patients with lung cancer treated with gefitinib, and the influence of EGF on platelet aggregation was investigated. Twenty patients were investigated. The serum level of TXB2 increased significantly in all patients who received gefitinib for 2 weeks (before vs. after = 94.1 ± 47.3 vs. 190.9 ± 54.3, p<0.01). TXB2 also increased significantly in responders without concurrent chemotherapy (before vs. after = 79.3 ± 35.5 vs. 194.5 ± 58.1, p<0.05), but not in non-responders (before vs. after = 106. 5 ± 65.8 vs. 162. 2 ± 52.8, N.S.). PG 6-keto F1α and PGE2 did not exhibit significant changes. Furthermore, EGF showed no significant change (after vs. before = 234 ± 35 vs. 276 ± 72, N.S.). Although there was no correlation between the levels of EGF and TXB2 (N.S.), the PG 6-keto F2α/TXB2 ratio decreased significantly (before vs. after = 0.054 ± 0.018 vs 0.033 ± 0.015, p<0.05). The secondary platelet aggregation observed after high-dose adenosine diphosphate stimulation was inhibited after a 1-minute preincubation with EGF. Platelet aggregation in patients after gefitinib administration tended to accelerate and secondary aggregation was observed after low-dose adenosine diphosphate stimulation. We conclude that careful observation is needed for patients with chronic obstructive pulmonary disease, pulmonary fibrosis, and thromboembolic diseases receiving gefitinib. Furthermore, measurement of prostanoids may be a good predictor of the beneficial and adverse effects. Moreover, the combination of gefitinib with a COX inhibitor that regulates TXA2/PGI2 balance should be evaluated.

Effect of low-dose aspirin for skin rash associated with erlotinib therapy in patients with lung cancer.

Department of Respiratory Medicine, Tesseikai Neurosurgical Hospital, Osaka, Japan, Department of Neurology, Kansai Medical University, Osaka, Japan, Department of Pharmacy, Tesseikai Neurosurgikal Hospital, Osaka, Japan, Department of Thoracic Malignancy, Osaka Prefectural Medical Center for Respiratory and Allergic Disease, Osaka, Japan, Department of Pulmonary Oncology, Osaka Medical Center for Cancer and Cardiovascular Disease, Osaka, Japan, and Kishiwada City Hospital, Divison of Hematology, Osaka, Japan

Analysis of cytotoxic T lymphocytes and Fas/FasL in Japanese patients with non-small cell lung cancer associated with HLA-A2.

Abstract Purpose. In recent years, the use of immunotherapy for malignant tumors has been proposed. To explore the significance of immunotherapy for lung cancer, we examined two systems : the HLA class I and cancer-reactive CTL system, and the Fas-FasL system. Methods. HLA class I (HLA-A, -B, and -C) antigens were determined in 61 patients with lung cancer and in 30 healthy controls. The HLA class I phenotype was investigated by serological techniques. HLA-A2 alleles were investigated by polymerase chain reaction sequence-specific primer analysis. We analyzed lymphocytes isolated from 61 patients with two-color surface labeling and flow cytometry. Furthermore, we analyzed sFas and sFasL expression by enzyme-linked immunosorbent assay (ELISA). We also examined lung cancer cell line-induced apoptosis of CD8+ lymphocytes using confocal laser scanning microscopy. Results. The HLA-A2 allele was observed in 27 of 61 patients with lung cancer. There were no differences in HLA-A2 allele classifications between lung cancer patients and controls. Thirty-six of the 61 lung cancer patients (57%) had elevated levels of sFas, and 16 of the 61 patients (26.2%) had elevated levels of sFasL. The sFas level of lung cancers with HLA-A2 was significantly higher than that of cancers without HLA-A2 (P<0.01). This tendency was observed in every lung cancer tissue type, and the sFas levels of lung cancers with HLA-A2 associated significantly with the CTL levels of lung cancers with HLA-A2. Furthermore, compared to lung cancers without HLA-A2, CD8+ T-cell levels were elevated in lung cancers with HLA-A2. In contrast, sFas levels of non-small cell lung cancers without HLA-A2 were higher than those in lung cancers with HLA-A2. In an in vitro experiment using lung cancer cell lines, we observed apoptosis of CD8+ lymphocytes induced by lung cancer cells. Lung cancer-reactive CTLs are mobilized easily by restriction of HLA-A2, but this restriction is not always specific. In addition, FasL derived from lung cancer cells can induce apoptosis of CD8+ T-cells, and the frequency of this phenomenon is increased in small cell lung cancers without HLA-A2. Conclusion. Our findings suggest that the effect of immunotherapy may be insufficient for non-small cell lung cancer without HLA-A2.