二十碳五烯酸对糖尿病肾足细胞损伤保护机制研究  

Protective mechanism of eicosapentaenoic acid on diabetic kidney podocyte injury

作　　者：张磊 于芳 曹瑞 王晓慧 徐孝娜 王枫 刘寒强 ZHANG Lei;YU Fang;CAO Rui;WANG Xiaohui;XU Xiaona;WANG Feng;LIU Hanqiang(Department of Military Health Education and Management,Ministry of Education Key Laboratory of Hazard Assessment and Control in Special Operational Environment,Shaanxi Provincial Key Laboratory of Environmental Health Hazard Assessment and Protection,Shaanxi Provincial Key Laboratory of Free Radical Biology and Medicine,School of Military Preventive Medicine,Air Force Medical University,Xi'an 710032,China)

机构地区：[1]空军军医大学军事预防医学系健康教育与管理教研室,特殊作业环境危害评估与防治教育部重点实验室,陕西省环境健康危害评估与防护重点实验室,陕西省自由基生物学与医学重点实验室,陕西西安710032

出　　处：《空军军医大学学报》2024年第8期913-919,共7页Journal of Air Force Medical University

基　　金：陕西省自然科学基金面上项目(2021JM-240)。

摘　　要：目的探究二十碳五烯酸(EPA)在模拟高糖诱导肾足细胞损伤中的作用及其潜在机制。方法体外培养小鼠MPC5肾足细胞,高糖诱导足细胞出现损伤,用不同浓度的EPA干预处理。采用CCK-8法评估MPC5细胞的活力,流式细胞术分析细胞凋亡,Western blotting检测凋亡蛋白及足细胞标志蛋白表达,活性氧(ROS)荧光探针检测法分析高糖刺激后ROS的生成,ELISA进一步检测炎性因子分泌情况。结果CCK-8细胞活力显示,EPA能维持高糖诱导的细胞活力,EPA干预后足细胞标志蛋白podocin和synaptopodin的表达明显升高(P<0.05,P<0.01);流式细胞术分析显示EPA干预后高糖刺激的足细胞凋亡率明显下降(P<0.01),促凋亡蛋白Bax和cleaved Caspase-3表达降低(P<0.05,P<0.01),抗凋亡蛋白Bcl-2的表达增加(P<0.01);相较于高糖组,EPA干预后ROS生成明显降低(P<0.01),ELISA结果显示EPA干预降低高糖诱导的TNF-α、IL-1β和IL-6炎性因子分泌(P<0.01);EPA干预后炎性信号通路蛋白TLR4和MYD88蛋白降低(P<0.05,P<0.01),EPA干预后脂质代谢蛋白SREBP1的表达下降(P<0.05,P<0.01);过表达SREBP1后,EPA对高糖诱导MPC5细胞上述检测结果被逆转。结论EPA可以抑制高糖刺激后足细胞内ROS的产生及减少足细胞凋亡,同时EPA可能通过调节SREBP1来抑制炎症通路激活,从而对糖尿病肾足细胞起到保护作用。Objective To investigate the role and potential mechanism of eicosapentaenoic acid(EPA)in simulating high glucose-induced kidney podocyte injury.Methods Mouse MPC5 kidney podocytes were cultured in vitro and induced by high glucose,and then treated with different concentrations of EPA.The viability of MPC5 cells was assessed by CCK-8 assay,apoptosis was analyzed by flow cytometry,expressions of apoptotic proteins and podocyte marker proteins were detected by Western blotting,reactive oxygen species(ROS)production after high glucose stimulation was analyzed by ROS fluorescent probe assay,and secretion of inflammatory factors was further detected by ELISA.Results CCK-8 cell viability showed that EPA maintained high glucose-induced cell viability,and the expression of podocyte marker proteins podocin and synaptopodin was significantly higher after EPA intervention(P<0.05,P<0.01).Flow cytometry analysis showed that apoptosis of high glucose-stimulated podocytes was significantly decreased after EPA intervention(P<0.01),the expressions of pro-apoptotic proteins Bax and cleaved Caspase-3 were decreased(P<0.05,P<0.01),and the expression of anti-apoptotic protein Bcl-2 was increased(P<0.01).ROS production was significantly decreased after EPA intervention compared with the high glucose group(P<0.01),and ELISA results showed that EPA intervention decreased high glucose-induced secretion of TNF-α,IL-1βand IL-6 inflammatory factors(P<0.01).After EPA intervention,the inflammatory signaling pathway proteins TLR4 and MYD88 protein were reduced(P<0.05,P<0.01),and the expression of lipid metabolism protein SREBP1 was decreased after EPA intervention(P<0.05,P<0.01).After overexpression of SREBP1,the above assay results of EPA on high glucose-induced MPC5 cells were reversed.Conclusion EPA can inhibit ROS production in podocytes stimulated by high glucose and reduce podocyte apoptosis,while EPA may inhibit the activation of inflammatory pathways by regulating SREBP1,thereby playing a protective role in diabetic kidney podocytes.

关 键 词：糖尿病肾病 二十碳五烯酸 足细胞损伤 SREBP1 

分 类 号：R587.2[医药卫生—内分泌]