抑瘤素M在大鼠非酒精性脂肪肝病中表达的意义

发布时间：2018-05-20 19:30

本文选题：非酒精性脂肪肝病 + 肝星状细胞株T6　；参考：《延边大学》2017年硕士论文

【摘要】：目的:本实验通过测定抑瘤素M(oncostatinM,OSM)对大鼠肝星状细胞株T6(hepatic stellate cell line T6,HSC-T6)的活化及增殖的影响,以及抑瘤素MⅡ型受体(oncostatinM receptor β,OSMRβ)在胆碱缺乏氨基酸(cho-line-deficient-L-amino acid-defined,CDAA)饲料诱导的大鼠非酒精性脂肪肝病(non-alcoholic fatty liver disease,NAFLD)模型肝脏中的表达,探讨NAFLD发病的可能机制。方法:体外实验采用HSC-T6细胞,HSC-T6细胞培养1天、4天、7天后提取蛋白质用Western blotting测定α-平滑肌激动蛋白(α-smooth muscle actin,α-SMA)和OSMRβ的表达。在96板孔中加入3×103个/孔HSC-T6细胞适应性培养4小时后,分别加入0、5、10、20ng/ml的大鼠OSM培养24小时和48小时,再添加10μl Cell Counting Kit-8(CCK-8)试剂2个小时后在450nm波长测光密度(OD)值。体内实验选取6周龄,体重160g±20g的SD雄性大鼠30只。适应性喂养7天后,随机平均分为正常对照组和模型组。正常组给予普通饲料;模型组给予CDAA饲料,实验周期为12周。实验结束时,乙醚麻醉并处死大鼠后取其肝右叶,用于制作病理标本及目的蛋白检测。用免疫组化法检测肝组织切片中α-SMA的表达情况。用Western blotting法测定肝脏OSMRβ的表达。结果:1.HSC-T6细胞培养1天、4天和7天,随着培养时间的延长,HSC-T6细胞表达α-SMA和OSMRβ水平逐渐升高(P0.05)。2.添加OSM可促进HSC-T6细胞增殖(P0.05),其促进作用呈剂量和时间依赖性。3.CDAA组肝脏的α-SMA 水平较正常对照组升高(n=15,P0.05),同时CDAA组肝脏表达OSMRβ,且高于正常对照组(n=15,P0.05)。结论:1.HSC-T6 细胞表达 OSMRβ。2.OSM促进HSC-T6细胞活化和增殖。3.随着CDAA诱导大鼠肝纤维化,肝脏表达OSMRβ增加,提示OSM通过与HSC的OSM受体结合促进肝脏炎症反应和纤维化,在NAFLD发生发展中发挥作用。
[Abstract]:Objective: to investigate the effect of tumor suppressor Mncostatin OSMon on the activation and proliferation of rat hepatic stellate cell line T6(hepatic stellate cell line T6 (HSC-T6). And the expression of oncostatin M receptor 尾 -OSMR 尾) in the liver of non-alcoholic fatty liver disease model induced by choline deficiency (cho-line-deficiency L-amino acid-defineded-defined CDAAA) in rats with non-alcoholic fatty liver disease (NAF LDD) was studied to explore the possible mechanism of NAFLD. Methods: HSC-T6 cells were cultured in HSC-T6 cells for 1 day, 4 days and 7 days later. The expression of 伪 -smooth muscle activin (伪 -SMA) and OSMR 尾 were measured by Western blotting. After 4 hours of adaptive culture of 3 脳 103 / well HSC-T6 cells in 96 plate holes, rat OSM was cultured for 24 hours and 48 hours, respectively, and then 10 渭 l Cell Counting Kit-8 CCK-8 reagent was added for 2 hours, then the value of 450nm wavelength photometric density (450nm) was measured. The results were as follows: (1) after 4 hours of adaptive culture, 3 脳 10 3 / well HSC-T6 cells were cultured for 24 hours and 48 hours, respectively, and 10 渭 l Cell Counting Kit-8 CCK-8 reagents were added for 2 hours. In vivo, 30 SD male rats of 6 weeks old and weight of 160 g 卤20 g were selected. After 7 days of adaptive feeding, they were randomly divided into normal control group and model group. The normal group was given normal diet and the model group was given CDAA forage for 12 weeks. At the end of the experiment, the rats were anesthetized and killed. The expression of 伪 -SMA in liver sections was detected by immunohistochemical method. The expression of OSMR 尾 in liver was determined by Western blotting assay. Results 1. The expression of 伪 -SMA and OSMR 尾 in HSC-T6 cells increased gradually with the increase of culture time. Addition of OSM could promote the proliferation of HSC-T6 cells in a dose-and time-dependent manner. 3. The level of 伪 -SMA in the liver of the CDAA group was higher than that of the normal control group. Meanwhile, the expression of OSMR 尾 in the liver of the CDAA group was higher than that of the normal control group. Conclusion the expression of OSMR 尾. 2. OSM in HSC-T6 cells promotes the activation and proliferation of HSC-T6 cells. As CDAA induced liver fibrosis, the expression of OSMR 尾 in the liver increased, suggesting that OSM promoted the inflammation and fibrosis of the liver by binding to the OSM receptor of HSC, and played an important role in the pathogenesis and development of NAFLD.
【学位授予单位】：延边大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R575

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