单宁酸对高糖和糖化终末产物培养条件下肾小球系膜细胞氧化应激及微炎症状态的改善作用

摘要

目的：探讨单宁酸对肾小球系膜细胞（GMC）的作用，从氧化应激及微炎症角度阐明单宁酸改善糖尿病肾病（DN）的作用机制。方法：体外培养 GMC，分别用高浓度葡萄糖（30 mmol·L-1）及糖化终末产物（AGEs）牛血清白蛋白（BSA，250 mg·L-1）处理，正常糖培养及不含糖的 BSA处理的细胞作为相应对照组，在高糖或AGEs处理的同时采用不同浓度单宁酸（10、20、40和80μmol·L-1）进行干预，培养48 h后检测细胞培养上清中丙二醛（MDA）水平及谷胱甘肽过氧化物酶（GSH-Px）、超氧化物歧化酶（SOD）和过氧化氢酶（CAT）活性， ELISA法检测细胞培养上清中8-羟基脱氧鸟苷（8-OHdG）水平，免疫组织化学法检测 GMC中细胞间黏附分子1（ICAM-1）蛋白表达， RT-PCR 法检测 GMC 中单核细胞趋化蛋白1（MCP-1）和 ICAM-1 mRNA 的表达水平。结果：与高糖组和 AGEs组比较，单宁酸处理组 MDA水平明显降低（P＜0.05），GSH-Px、SOD及CAT活性明显升高（P＜0.05或P＜0.01），GMC培养上清中8-OHdG水平明显降低（P＜0.05）。与高糖组和 AGEs组比较，40和80μmol·L-1单宁酸组 ICAM-1蛋白表达水平明显降低（P＜0.05）。与高糖组比较，单宁酸组 MCP-1和ICAM-1 mRNA表达水平明显降低（P＜0.01）。结论：单宁酸可保护 GMC免受氧化及炎症介质的损伤，从而延缓和改善DN的肾小球病变。%Objective To investigate the effect of tannic acid on glomerular mesangial cells (GMC),and to clarify the mechanism of tannic acid in improving the pathological changes of diabetic nephropathy (DN)from the aspect of oxidative stress and micro-inflammation. Methods The glomerular mesangial cells were treated with glucose (30 mmol·L-1 )or advanced glycosylation end-products (AGEs)bovine serum albumin(BSA)(250 mg·L-1 )and then different concentrations of tannic acid (10,20,40 and 80μmol·L-1 )were added into the GMC.The cells cultured by normal glucose or treated with BSA were used as control groups and then the level of malonic dialdehyde (MDA), glutathione peroxidase (GSH-Px ), superoxide Dismutase (SOD ), CAT (Catalase ) activities and 8-hydroxy-2′-deoxyguanosine(8-OHdG)levels in the culture supernatant 48 h after culture were determined by colorimetry and ELISA method. The expressions of intercellular cell adhesion molecule-1 (ICAM-1 ) protein, monocyte chemotactic protein 1 (MCP-1 ) and ICAM-1 mRNA in GMC were detected by immunohistochemical staining and RT-PCR method.Results Compared with high glucose and AGEs groups,the MDA levels in tannic acid groups were reduced significantly(P<0.05);the activities of GSH-Px,SOD and CAT were increased significantly(P<0.05 or P<0.01);the 8-OHdG levels in annic acid groups were significantly reduced (P<0.05). Compared with high glucose and AGEs groups,the expressions levels of ICAM-1 protein in 40 and 80μmol· L-1 tannic acid groups were decreased (P<0.05 ). The mRNA expressions levels of MCP-1 and ICAM-1 were significantly lower than those in high glucose group (P<0.01 ).Conclusion Tannic acid could protect GMC against the damage of oxidative and inflammatory mediators,thereby delaying and improving the glomerular lesions of DN.