紫檀芪减轻人肾小管上皮细胞高糖缺氧复氧损伤及其作用机制的研究

Pterostilbene reduces the damage of high glucose,hypoxia and reoxygenation in HK-2 cells and its mechanism

目的 探究紫檀芪(PTE)对人肾小管上皮细胞(HK-2)高糖缺氧复氧损伤的保护作用及其可能的作用机制.方法 随机将HK-2细胞分为低糖缺氧复氧组(LR)、高糖缺氧复氧组(HR)、高糖缺氧复氧+PTE 组(HR+P)、高糖缺氧复氧组+PTE+沉默信息调节因子 1(SIRT1)抑制剂组(HR+P+EX).缺氧复氧结束后采用CCK-8试剂盒和乳酸脱氢酶(LDH)检测细胞活性,超氧化物歧化酶(SOD)及丙二醛(MDA)试剂盒检测SOD活性及MDA含量;活性氧簇(ROS)染色检测细胞内ROS含量;Western blot检测HK-2细胞SIRT1、Beclin1、泛素结合蛋白(SQSTM1/p62)和微管相关蛋白轻链3B(LC3B)的表达.结果 与LR组比较,HR组LDH、MDA及ROS染色荧光强度增加,CCK-8及SOD降低,SQSTM1/p62表达上调(P<0.05),SIRT1、Beclin1、LC3B表达降低(P<0.05).与HR组比较,HR+P组CCK-8和SOD增加,LDH、MDA及ROS荧光强度下降;SIRT1、Beclin1、LC3B表达上调(P<0.05),SQSTM1/p62表达下调(P<0.05).与HR+P组比较,HR+P+EX组LDH、MDA 及 ROS 染色荧光强度增加,CCK-8 及 SOD 降低,SQSTM1/p62 表达上调(P<0.05),SIRT1、Beclin1、LC3B表达降低(P<0.05).结论 PTE通过促进HK-2细胞SIRT1表达,促进自噬的恢复,减轻高糖缺氧复氧引起的损伤.

Objective To evaluate the protective effect of pterostilbene on high glucose,hypoxia and reoxygenation injury in HK-2 cells and to explore its possible mechanism.Methods HK-2 cells were randomly divided into 4 groups:low glucose hypoxia reoxygenation group(LR),high glucose hypoxia reoxygenation group(HR),high glucose hypoxia reoxygenation+pterostilbene group(HR+P),high glucose hypoxia reoxygenation+pterostilbene+SIRT1 inhibitor group(HR+P+EX).After hypoxia and reoxygenation,cell viability was measured by CCK-8 and lactate dehydrogenase(LDH)release assay.Superoxide dismutase(SOD)activity and malondialdehyde(MDA)were detected by SOD and MDA kits.Intracellular ROS content was measured by ROS staining.The expression of SIRT1,Beclin1,SQSTM1/p62 and LC3B in HK-2 cells was measured by Western blot.Results Compared with LR group,the LDH,MDA and ROS fluorescence intensity were significantly increased,the CCK-8 and SOD were significantly decreased,the expression of SQSTM1/p62 was significantly increased(P<0.05),SIRT1,Beclin1 and LC3B were significantly decreased in HR group(P<0.05).Compared with HR group,the CCK-8 and SOD were significantly increased,LDH,MDA and ROS fluorescence intensity were decreased,the expression of SIRT1,Beclin1,LC3B were significantly increased,while the expression of SQST M1/p62 was significantly decreased in HR+P group(P<0.05).Compared with HR+P group,the fluorescence intensity of LDH,MDA and ROS were significantly increased,the CCK-8 and SOD were significantly decreased,the expression of SQSTM1/p62 was significantly increased(P<0.05),and the expression of SIRT1,Beclin1 and LC3B were significantly decreased in HR+P+EX group(P<0.05).Conclusion Pterostilbene promotes the recovery of autophagy by promoting the expression of SIRT1 protein in HK-2 cells,thereby alleviating the damage caused by hypoglycemia and hypoxia.