合作科研工作成果与展望

本人在日本大阪国立循环器病中心工作,任生物流变研究室教授。受日本国际协力事业团派遣,于1989年4月21日～5月31日到中日友好医院临床医学研究所生物流变室与庄逢源博士进行合作研究。合作研究成果:①设置完成血液细胞变形的测定程序;②从采血中分离白血球(多核白血球、单核白血球及淋巴细胞);③确立白血球变形能的测定分析和使研究人员熟练掌握了测定技术;④测定多核白血球变形能。

Bone marrow-derived monocyte infusion improves hepatic fibrosis by decreasing osteopontin,TGF-β1,IL-13 and oxidative stress

To evaluate the therapeutic effects of bone marrow-derived CD11b+CD14+ monocytes in a murine model of chronic liver damage.METHODSChronic liver damage was induced in C57BL/6 mice by administration of carbon tetrachloride and ethanol for 6 mo. Bone marrow-derived monocytes isolated by immunomagnetic separation were used for therapy. The cell transplantation effects were evaluated by morphometry, biochemical assessment, immunohistochemistry and enzyme-linked immunosorbent assay.RESULTSCD11b+CD14+ monocyte therapy significantly reduced liver fibrosis and increased hepatic glutathione levels. Levels of pro-inflammatory cytokines, including tumor necrosis factor-α, interleukin (IL)-6 and IL-1β, in addition to pro-fibrotic factors, such as IL-13, transforming growth factor-β1 and tissue inhibitor of metalloproteinase-1 also decreased, while IL-10 and matrix metalloproteinase-9 increased in the monocyte-treated group. CD11b+CD14+ monocyte transplantation caused significant changes in the hepatic expression of α-smooth muscle actin and osteopontin.CONCLUSIONMonocyte therapy is capable of bringing about improvement of liver fibrosis by reducing oxidative stress and inflammation, as well as increasing anti-fibrogenic factors.

Macrophage colony-stimulating factor expressed in non-cancer tissues provides predictive powers for recurrence in hepatocellular carcinoma

AIM To investigate the role of macrophage colony-stimulating factor(M-CSF) in patients with hepatocellular carcinoma(HCC) after surgery. METHODS Expression of M-CSF, distribution of M2 macrophages(Mφs), and angiogenesis were assessed in the liver, including tumors and peritumoral liver tissues. The prognostic power of these factors was assessed. Mouse isolated hepatic Mφs or monocytes were cultured with media containing M-CSF. The concentration of vascular endothelial growth factor(VEGF) in media was assessed. Furthermore, the role of the M-CSF-matured hepatic Mφs on proliferation of the vascular endothelial cell(VEC) was investigated. RESULTS A strong correlation between the expressions of M-CSF and CD163 was observed in the peritumoral area. Also, groups with high density of M-CSF, CD163 or CD31 showed a significantly shorter time to recurrence(TTR) than low density groups. Multivariate analysis revealedthe expression of M-CSF or hepatic M2Mφs in the peritumoral area as the most crucial factor responsible for shorter TTR. Moreover, the expression of M-CSF and hepatic M2Mφs in the peritumoral area had better predictable power of overall survival. Values of VEGF in culture media were significantly greater in the hepatic Mφs compared with the monocytes. Proliferation of the VEC was greatest in the cells co-cultured with hepatic Mφs when M-CSF was present in media.CONCLUSION M-CSF increases hepatocarcinogenesis, most likely by enhancing an angiogenic factor derived from hepatic Mφ and could be a useful target for therapy against HCC.

Different phenotypes of monocytes in patients with newonset mild acute pancreatitis

AIM To evaluate the numbers of different subsets of monocytes and their associations with the values of clinical measures in mild acute pancreatitis(MAP) patients.METHODS The study included one group of 13 healthy controls and another group of 24 patients with new-onset MAP. The numbers of different subsets of monocytes were examined in these two groups of subjects by flow cytometry. The concentrations of plasma interleukin(IL)-10 and IL-12 were determined by cytometric bead array. The acute physiology and chronic health evaluation(APACHE) II scores of individual patients were evaluated, and the levels of plasma C-reactive protein(CRP) as well as the activities of amylase and lipase were measured. RESULTS In comparison with that in the controls, significantly increased numbers of CD14+CD163-, CD14+CD163-MAC387+ M1 monocytes, but significantly reduced numbers of CD14+CD163+IL-10+ M2 monocytes were detected in the MAP patients(P < 0.01 or P < 0.05). Furthermore, significantly higher levels of plasma IL-10 and IL-12 were observed in the MAP patients(P < 0.01 for all). More importantly, the levels of plasma CRP were positively correlated with the numbers of CD14+CD163-(R = 0.5009, P = 0.0127) and CD14+CD163-MAC387+(R = 0.5079, P = 0.0113) M1 monocytes and CD14+CD163+CD115+ M2 monocytes(R = 0.4565, P = 0.0249) in the patients. The APACHE II scores correlated with the numbers of CD14+CD163+CD115+(R = 0.4581, P = 0.0244) monocytes and the levels of plasma IL-10(R = 0.4178, P = 0.0422) in the MAP patients. However, there was no significant association among other measures tested in this population. CONCLUSION Increased numbers of CD14+CD163- and CD14+ CD163-MAC387+ monocytes may contribute to the pathogenesis of MAP, and increased numbers of CD14+CD163+CD115+ monocytes may be a biomarker for evaluating the severity of MAP.

Circulating CD14＋ monocytes in patients with aortic stenosis

BackgroundCalcific aortic stenosis （AS） is an active process sharing similarities with atherosclerosis and chronic inflammation. The pathophysiology of AS is notable for three cardinal components： inflammation, fibrosis and calcification. Monocytes play a role in each of these processes. The role of circulating monocytes in AS is not clear. The aim of the present study was to study an association between cir-culating apoptotic and non apoptotic CD14＋ monocytes and AS features.MethodsWe assessed the number of CD14＋ monocytes and apoptotic monocytes in 54 patients with significant AS （aortic valve area 0.74 ± 0.27 cm2） and compared them to 33 patients with similar risk factors and no valvular disease. The level of CD14＋ monocytes and apoptotic monocytes was assessed by flow cytometry.ResultsThere was no difference in the risk factor profile and known coronary or peripheral vascular diseases between patients with AS and controls.Pa-tients with AS exhibited increased numbers of CD14＋ monocytes as compared to controls （9.9% ± 4.9%vs. 7.7% ± 3.9%,P= 0.03）. CD14＋ monocyte number was related to age and the presence and severity of AS. In patients with AS, both CD14＋ monocytes and apoptotic mono-cytes were inversely related to aortic valve area.ConclusionsPatients with significant AS have increased number of circulating CD14＋ monocytes and there is an inverse correlation between monocyte count and aortic valve area. These findings may suggest that inflammation is operative not only in early valve injury phase, but also at later developed stages such as calcification when AS is severe.