糖肾宁通过调控巨噬细胞源外泌体减轻小鼠肾小球足细胞炎症反应的机制研究

Study on the mechanism of Tangshenning alleviating the inflammatory response of mouse glomerular podocytes by regulating the exosomes derived from macrophages

目的以巨噬细胞源外泌体为切入点,观察糖肾宁对体外高糖诱导的小鼠肾小球足细胞炎症反应的影响,探讨其保护足细胞的可能机制。方法采用糖肾宁干预高糖条件培养的巨噬细胞,分为正常组、高糖组、糖肾宁组,干预48小时后,提取各组细胞上清液中外泌体,将各组外泌体分别干预高糖和非高糖条件培养的小鼠肾小球足细胞,分为正常外泌体—高糖组、高糖外泌体—高糖组、糖肾宁外泌体—高糖组、正常外泌体—非高糖组、高糖外泌体—非高糖组、糖肾宁外泌体—非高糖组、无外泌体—高糖组和无外泌体—非高糖组等8个组。48小时后,收集各组细胞并进行相关检测。CCK-8法检测各组细胞活力,酶联免疫法检测各组细胞上清液中炎症因子单核细胞趋化蛋白-1(monocyte chemotactic protein-1,MCP-1)、肿瘤坏死因子α(tumor necrosis factor-α,TNF-α)、白细胞介素6(interleukin-6,IL-6)的蛋白表达情况,实时荧光定量聚合酶链式反应检测各组细胞炎症因子(MCP-1、TNF-α、IL-6)的表达情况。结果在高糖或非高糖环境下的足细胞中,与正常外泌体干预的足细胞相比较,高糖外泌体干预的足细胞细胞活力下降(P<0.05),炎症因子(MCP-1、TNF-α、IL-6)的基因和蛋白表达均增加(P<0.05);而与高糖外泌体干预的足细胞相比,糖肾宁外泌体干预的足细胞细胞活力有所升高(P<0.05),各炎症因子的基因和蛋白表达均有所下调(P<0.05)。结论糖肾宁能通过调控巨噬细胞源外泌体减轻小鼠肾小球足细胞炎症反应。

Objective To observe the effect of Tangshenning on the inflammatory of mouse glomerular podocytes induced by high glucose in vitro,and to explore the possible mechanism of its protection against podocytes.Methods The macrophages cultured under high glucose condition were divided into normal group,high glucose group,and Tangshenning group.After 48 hours of intervention,exosomes were extracted from the supernatant of each group.The exosomes were intervened in podocytes cultured under non high glucose and high glucose conditions respectively,and were divided into blank control group,normal-normal exosomes group,normal-high glucose exosomes group,normal-Tangshenning exosomes group,model group,high glucose-normal exosomes group,high glucose-high glucose exosomes group and high glucose-Tangshenning exosomes groups.After 48 hours,cells in each group were collected and detected.CCK-8 method was used to detect the cell activity of each group.The enzyme-linked immunosorbent assay was used to detect the protein expression of inflammatory factors(MCP-1,TNF-α,IL-6)in the supernatant of each group.Real-time fluorescence quantitative polymerase chain reaction was used to detect the mRNA expression of inflammatory factors(MCP-1,TNF-α,IL-6).Results Compared with the normal-normal exosomes group,the cell viability of the normal-high glucose exosomes was decreased(P<0.05),and the genes and proteins of inflammatory factors(MCP-1,TNF-α,IL-6)were increased(P<0.05);compared with the normal-high glucose exosomes group,the cell viability of the normal-Tangshenning exosomes group was increased(P<0.05),and the genes and proteins of various inflammatory factors were decreased(P<0.05).In podocytes cultured in high glucose condition,different groups of exosomes intervention showed similar effects.Conclusion Tangshenning can reduce the inflammatory of podocytes by regulating the exosomes derived from macrophages.