EPA对白蛋白诱导肾小管上皮细胞损伤的保护作用的机制研究

发布时间：2018-02-16 06:51

本文关键词： 二十碳五烯酸白蛋白过氧化物酶体增殖物激活受体-γ 整合素β1 整合素连接激酶　出处：《山东大学》2014年硕士论文　论文类型：学位论文

【摘要】：研究背景与目的在各种慢性肾脏病进展中,肾脏纤维化是其共同的病理基础,也是导致终末期肾病的主要原因。。肾脏纤维化的病理改变主要包括肾小球硬化和小管间质纤维化。肾小管上皮细胞(human proximal tubular cells, HK-2)损伤并分泌大量致炎因子、促纤维形成因子和基质蛋白等,形成小管间质损伤的恶性循环,是引起肾间质纤维化的主要原因。整合素连接激酶(integrin-linked kinase, ILK)、整合素β1(integrin β1)、过氧化物酶体增殖物激活受体γ(peroxisome proliferators-activated receptor gamma, PPARy)等与肾脏纤维化的进展有着十分密切的关系。多不饱和脂肪酸(polyunsaturated Fatty Acids,PUFA)是哺乳动物体内的必需脂肪酸。但是其体内不能自身合成PUFA,只能从食物中摄取。食品来源的多不饱和脂肪酸有Ω-3(n-3PUFA和Ω-6(n-6)PUFA两种类型。从深海鱼类中提取的鱼油,含丰富的n-3PUFA,二十二碳五烯酸(eicosapentaenoic acid, EPA)和二十二碳六烯酸(docosahexaenoic acid, DHA)是其主要活性成分。研究发现鱼油除具有如抗血栓、调血脂等作用外,还对肾脏有一定的保护作用,延缓肾脏疾病的进展。本实验通过白蛋白致肾小管上皮细胞损伤模型观察EPA对HK-2ILK,整合素β1、PPARy表达的影响,探究EPA对肾脏的保护作用机制,为临床应用提供理论依据。方法取对数生长期HK-2,常规培养24h后,换无血清培养24h,使细胞同步于GO期。分为：正常对照组；白蛋白组(模型组,白蛋白20.0mg/L); EPA干预组(白蛋白20.0mg/L+EPA10μmol/L或100μmol/L)。各组分别培养24h、48h,72h后,进行指标测定。 1采用Western Blot法观察EPA对白蛋白刺激HK-2PPARγ、ILK和整合素β1表达的影响； 2采用RT-PCR法观察EPA对白蛋白刺激HK-2PPARγ、ILK和整合素p1mRNA表达的影响。结果 1.EPA对白蛋白刺激HK-2细胞PPARy表达的影响白蛋白可使HK-2细胞PPARy mRNA和蛋白表达下降；EPA可增加白蛋白诱导的HK-2细胞PPARy mRNA和蛋白的表达。 2.EPA对白蛋白刺激HK-2细胞ILK表达的影响白蛋白可增加HK-2细胞ILK mRNA和蛋白的表达；EPA可使白蛋白诱导的HK-2细胞ILK mRNA和蛋白表达下降。 3.EPA对白蛋白刺激HK-2细胞整合素β1mRNA表达的影响白蛋白可使HK-2细胞整合素β1mRNA的表达下降；EPA可增加白蛋白诱导的HK-2细胞整合素β1mRNA表达。结论 1.EPA能够促进白蛋白诱导的HK-2细胞PPARy蛋白和基因的表达。 2.EPA可以抑制白蛋白诱导的HK-2细胞ILK蛋白和基因的表达。 3.EPA可以促进白蛋白诱导的HK-2细胞整合素β1蛋白和基因的表达。
[Abstract]:Research background and purpose. Among the progress of chronic kidney disease, renal fibrosis is the common pathological basis. The pathological changes of renal fibrosis mainly include glomerulosclerosis and tubulointerstitial fibrosis. Fiber forming factors and matrix proteins form a vicious circle of tubulointerstitial injury. Integrin linked kinase (ILKN), integrin 尾 1 integrin 尾 1 (integrin 尾 1), peroxisome proliferators-activated receptor gamma (PPA Ryma) are closely related to the progression of renal fibrosis. Polyunsaturated fatty acids (PUFAs) are essential fatty acids in mammals, but they cannot synthesize PUFAs by themselves and can only be ingested from foods. There are two types of polyunsaturated fatty acids from food sources: 惟 -3n -3PUFA and 惟 -6n -6 PUFA. Fish oil extracted from deep-sea fish, Rich n-3 PUFAA, 22 carbapentaenoic acid acidand 22 carbohexaenoic acid acid. it is found that fish oil not only has some protective effects on kidney, but also has antithrombotic and lipid effects. Delay the progression of kidney disease. In this study, we observed the effects of EPA on the expression of HK-2 ILK and integrin 尾 1 PPARy in renal tubular epithelial cells induced by albumin, and explored the protective mechanism of EPA on kidney, which provided theoretical basis for clinical application. Method. The logarithmic growth phase HK-2 was taken for 24 hours after conventional culture, and the cells were cultured in go phase for 24 hours. The cells were divided into three groups: normal control group, albumin 20.0 mg / L group (model group), EPA intervention group (20.0 mg / L EPA10 渭 mol/L or 100 渭 mol / L mol/L or 100 渭 mol / L EPA) for 24 h and 48 h, respectively. The index was measured. 1the effect of EPA on the expression of HK-2PPAR 纬 -ilk and integrin 尾 1 stimulated by albumin was observed by Western Blot assay. 2the effect of EPA on the expression of HK-2PPAR 纬 -ilk and integrin p1 mRNA was observed by RT-PCR assay. Results. 1. The effect of EPA on the expression of PPARy in HK-2 cells stimulated by albumin. Albumin can decrease the expression of PPARy mRNA and protein in HK-2 cells. It can increase the expression of PPARy mRNA and protein in HK-2 cells induced by albumin. 2. The effect of EPA on the expression of ILK in HK-2 cells stimulated by Albumin. Albumin could increase the expression of ILK mRNA and protein in HK-2 cells. EPA could decrease the expression of ILK mRNA and protein in HK-2 cells induced by albumin. 3. Effect of EPA on integrin 尾 1 mRNA expression in HK-2 cells stimulated by albumin. Albumin can decrease the expression of integrin 尾 1 mRNA in HK-2 cells. EPA can increase the expression of integrin 尾 1 mRNA in HK-2 cells induced by albumin. Conclusion. 1. EPA could promote the expression of PPARy protein and gene in HK-2 cells induced by albumin. 2. EPA could inhibit the expression of ILK protein and gene in HK-2 cells induced by albumin. 3. EPA could promote the expression of integrin 尾 1 protein and gene in HK-2 cells induced by albumin.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R692

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