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

AIM To evaluate the therapeutic effects of bone marrowderived CD11b+CD14+ monocytes in a murine model of chronic liver damage. METHODS Chronic 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. RESULTS CD11b+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 monocytetreated group. CD11b+CD14+ monocyte transplantation caused significant changes in the hepatic expression of α-smooth muscle actin and osteopontin. CONCLUSION Monocyte therapy is capable of bringing about improvement of liver fibrosis by reducing oxidative stress and inflammation,as well as increasing antifibrogenic factors.

Impaired norepinephrine regulation of monocyte inflammatory cytokine balance in heart failure

AIM To evaluate the effect of norepinephrine on inflammatory cytokine expression in ex vivo human monocytes and monocytic THP-1 cells. METHODS For human monocyte studies, cells were isolated from 12 chronic heart failure(HF)(66 ± 12 years, New York Heart Association functional class Ⅲ-Ⅳ, left ventricular ejection fraction 22% ± 9%) and 14 healthy subjects(66 ± 12 years). Monocytes(1 × 106/mL) were incubated with lipopolysaccharide(LPS) 100 ng/m L, LPS + norepinephrine(NE) 10-6 mol/L or neither(control) for 4 h. Tumor necrosis factor-alpha(TNFα) and interleukin-10(IL-10) production were determined by ELISA. Relative contribution of α- and β-adrenergic receptor subtypes on immunomodulatory activity of NE was assessed in LPSstimulated THP-1 cells incubated with NE, the α-selective agonist phenylephrine(PE), and the β-selective agonist isoproterenol(IPN). NE-pretreated THP-1 cells were also co-incubated with the β-selective antagonist propranolol(PROP), α2-selective antagonist yohimbine(YOH) or the α1-selective antagonist prazosin(PRAZ). RESULTS Basal TNFα concentrations were higher in HF vs healthy subjects(6.3 ± 3.3 pg/mL vs 2.5 ± 2.6 pg/mL, P = 0.004). Norepinephrine's effect on TNFα production was reduced in HF(-41% ± 17% HF vs -57% ± 9% healthy, P = 0.01), and proportionately with NYHA FC. Increases in IL-10 production by NE was also attenuated in HF(16% ± 18% HF vs 38% ± 23% healthy, P = 0.012). In THP-1 cells, NE and IPN, but not PE, induced a dosedependent suppression of TNFα. Co-incubation with NE and antagonists revealed a dose-dependent inhibition of the NE suppression of TNFα by PROP, but not by YOH or PRAZ. Dose-dependent increases in IL-10 production were seen with NE and IPN, but not with PE. This effect was also antagonized by PROP but not by YOH or PRAZ. Pretreatment of cells with IPN attenuated the effects of NE and IPN, but did not induce a response to PE.CONCLUSION NE regulation of monocyte inflammatory cytokine production may be reduced in moderate-severe HF, and may be mediated through β-adrenergic receptors.

Association of NFKB1 gene polymorphism (rs28362491) with levels of inflammatory biomarkers and susceptibility to diabetic nephropathy in Asian Indians

AIM To investigate the association of NFKB1 gene-94 ATTG insertion/deletion(rs28362491) polymorphism with inflammatory markers and risk of diabetic nephropathy in Asian Indians.METHODS A total of 300 subjects were recruited(100 each), normoglycemic,(NG); type 2 diabetes mellitus(T2DM) without any complications(DM) and T2 DM with diabetic nephropathy [DM-chronic renal disease(CRD)]. Analysis was carried out by polymerase chain reaction-restriction fragment length polymorphism and ELISA. Pearson's correlation, analysis of variance and logistic regression wereused for statistical analysis.RESULTS The allelic frequencies of-94 ATTG insertion/deletion were 0.655/0.345(NG), 0.62/0.38(DM) and 0.775/0.225(DM-CRD). The-94 ATTG ins allele was associated with significantly increased levels of urinary monocyte chemoattractant protein-1(u MCP-1); u MCP-1(P = 0.026) and plasma tumor necrosis factor-alpha(TNF-α); TNF-α(P = 0.030) and almost doubled the risk of diabetic nephropathy(OR = 1.91, 95%CI: 1.080-3.386, P = 0.025).CONCLUSION-94 ATTG ins/ins polymorphism might be associated with increased risk of developing nephropathy in Asian Indian subjects with diabetes mellitus.

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.

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.

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。

Pretransplant absolute monocyte count in peripheral blood predicts posttransplant tumor prognosis in patients undergoing liver transplantation for hepatocellular carcinoma

BACKGROUND: Preoperative absolute monocyte count in peripheral blood(AMCPB) is closely associated with prognoses in not only various malignancies but also hepatocellular carcinoma(HCC). The purpose of this study was to evaluate whether pretransplant AMCPB predicts posttransplant outcomes in patients with HCC undergoing liver transplantation(LT).METHOD: We retrospectively analyzed relationships between clinicopathologic factors involving pretransplant AMCPB and tumor recurrence or survival in 256 patients who had undergone LT for HCC between January 2005 and April 2012.RESULTS: ROC curve analysis showed that AMCPB >200/mm3was a risk factor for tumor recurrence; 43 patients showed higher AMCPB(>200/mm3), whereas 213 showed lower AMCPB(≤200/mm3) at the time of LT. On multivariate analysis,pretransplant high AMCPB, positive findings in pretransplant18F-FDG PET/CT, pathological maximal tumor size >5 cm,intrahepatic metastasis, moderately or poorly differentiated tumor and microvascular invasion were independent factors affecting recurrence-free survival. When we performed subgroup analysis based on the Milan criteria, high AMCPB was an independent factor for predicting HCC recurrence in patients with tumor beyond the Milan criteria(P=0.004), and not for patients within the criteria.CONCLUSION: This study demonstrated that pretransplant AMCPB could predict tumor recurrence after LT for HCC,especially in patients with tumor beyond the Milan criteria.

Anti-inflammatory effect of Heliotropium indicum Linn on lipopolysaccharide-induced uveitis in New Zealand white rabbits

AIM: To investigate the anti-inflammatory effect of an aqueous whole plant extract of Heliotropium indicum(HIE) on endotoxin-induced uveitis in New Zealand white rabbits.·METHODS: Clinical signs of uveitis including flares,iris hyperemia and miosis, were sought for and scored in1.0 mg/kg lipopolysaccharide(LPS)-induced uveitic rabbits treated orally with HIE(30-300 mg/kg),prednisolone(30 mg/kg), or normal saline(10 m L/kg). The number of polymorphonuclear neutrophils infiltrating, the protein concentration, as well as levels of tumor necrosis factor-α(TNF-α), prostaglandin E2(PGE2), and monocyte chemmoattrant protein-1(MCP-1) in the aqueous humor after the various treatments were also determined. A histopathological study of the anterior uveal was performed.· RESULTS: The extract and prednisolone-treatment significantly reduced(P ≤0.001) both the clinical scores of inflammation(1.0-1.8 compared to 4.40 ±0.40 in the normal saline-treated rabbits) and inflammatory cells infiltration. The level of protein, and the concentrationsof TNF-α, PGE2 and MCP-1 in the aqueous humor were also significantly reduced(P ≤0.001). Histopathological studies showed normal uveal morphology in the HIE and prednisolone-treated rabbits while normal saline-treated rabbits showed marked infiltration of inflammatory cells.· CONCLUSION: The HIE exhibits anti-inflammatory effect on LPS-induced uveitis possibly by reducing the production of pro-inflammatory mediators.

Circulating CD14＋ monocytes in patients with aortic stenosis

BackgroundCalcific 大动脉的狭窄(作为) 是与动脉粥样硬化和长期的发炎分享类似的一个活跃过程。pathophysiology 这为三个最重要的部件是著名的：发炎，纤维变性和石灰化。单核白血球在每这些过程起一个作用。传播单核白血球在的角色这不是清楚的。现在的学习的目的是学习在传播 apoptotic 之间并且非的一个协会 apoptotic CD14 + 单核白血球并且当 features.MethodsWe 在 54 个病人估计了 CD14 + 单核白血球和 apoptotic 单核白血球的数字与作为重要(大动脉的阀门区域 0.74 ±；0.27 厘米 2) 并且把他们与类似的风险因素和没有瓣膜的疾病作比较到 33 个病人。CD14 + 单核白血球和 apoptotic 单核白血球的水平被流动 cytometry.ResultsThere 估计不是在风险因素侧面的差别并且知道在病人之间的冠或外部的脉管的疾病与作为并且控制。病人与是作为与控制相比展出了 CD14 + 单核白血球的增加的数字(9.9%±；4.9% 对 7.7%±；3.9% ， P = 0.03 ) 。CD14 + 单核白血球数字与年龄和存在和严厉有关作为。在病人与作为，两个 CD14 + 单核白血球和 apoptotic 单核白血球是相反地与大动脉的阀门 area.ConclusionsPatients 与有关作为重要增加了传播 CD14 + 单核白血球的数字，在单核白血球计数和大动脉的阀门区域之间有反的关联。这些调查结果可以建议那发炎在早阀门损害阶段是起作用的不仅，而且在象石灰化那样的以后的发达阶段什么时候同样是严重的。