白藜芦醇通过调节SIRT1改善环磷酰胺诱导的人颗粒细胞氧化应激损伤  

Resveratrol ameliorates oxidative stress damage of human granulosa cells induced by cyclophosphamide by regulating SIRT1

作　　者：吴海峰[1] 贾建雷[1] 李倩[1] 甄秀丽[2] 魏雅楠[2] 乜照燕[2] WU Hai-feng;JIA Jian-lei;LI Qian;ZHEN Xiu-li;WEI Ya-nan;NIE Zhao-yan(Department of Laboratory Medicine, Hebei Chest Hospital, Shijiazhuang 050041, China;Department of Reproductive Medicine, the Fourth Hospital of Hebei Medical University, Shijiazhuang 050011, China)

机构地区：[1]河北省胸科医院检验科,河北石家庄050041 [2]河北医科大学第四医院生殖医学科,河北石家庄050011

出　　处：《河北医科大学学报》2022年第7期832-838,共7页Journal of Hebei Medical University

基　　金：河北省民生科技专项(192777106D);河北省医学科学研究重点课题计划(20210332)。

摘　　要：目的探讨白藜芦醇(resveratrol,RES)对4-过氧化氢环磷酰胺(4-hydroperoxy cyclophosphamide,4-HC)诱导人颗粒细胞(granulosa cells,GCs)氧化应激损伤改善作用。方法收集体外受精-胚胎移植患者卵泡液GCs进行体外培养。苏木精-伊红染色和免疫组织化学染色对GCs鉴定,Cell Counting Kit-8(CCK-8)检测4-HC诱导GCs毒性和RES对4-HC诱导GCs毒性改善作用。将GCs分为3组:对照组、4-HC组、RES+4-HC组。CCK-8测定GCs存活率,活性氧荧光探针DCFH-DA检测GCs内活性氧(reactive oxygen species,ROS),水溶性四氮唑1测定超氧化物歧化酶(superoxide dismutase,SOD),钼酸铵法检测过氧化氢酶(catalase,CAT),Western Blot测定沉默信息调节因子2相关酶1(silent information regulator 1,SIRT1)蛋白表达。结果4-HC(0,1.0,2.5,5.0,7.5μmol/L)处理GCs,CCK-8结果显示随4-HC浓度增加,GCs存活率降低,与对照组比较,差异有统计学意义(P<0.05),选择2.5μmol/L 4-HC为后续4-HC模型浓度。RES(0、5、10、25、50、75μmol/L)处理GCs 2 h后加入2.5μmol/L 4-HC,结果显示,RES可提高GCs存活率,与4-HC组比较,差异有统计学意义(P<0.05)。选择50μmol/L RES为后续改善浓度。对3组GCs处理后各项检测结果显示,与CON组比较,4-HC组GCs存活率、SOD、CAT降低,ROS增加(P<0.05)。与4-HC组比较,RES+4-HC组GCs细胞活率、SOD、CAT增加,ROS降低。Western Blot显示RES+4-HC组SIRT1升高,与其他2组比较,差异有统计学意义(P<0.05)。结论RES可能通过增加SIRT1表达改善4-HC诱导GCs损伤。Objective To investigate the role of resveratrol(RES)in ameliorating 4-hydroperoxy cyclophosphamide(4-HC)-induced oxidative stress damage in human granulosa cells(GCs).Methods The GCs were collected from patients undergoing in vitro fertilization and embryo transfer(IVF-ET)for in vitro culture.Hematoxylin-eosin staining(HE)and immunochemical staining were performed to identify GCs.Cell Counting Kit-8(CCK-8)was used to detect the toxicity of 4-HC-induced GCs and the improvement effect of RES on the toxicity of 4-HC-induced GCs.The GCs were divided into three groups:the control group,4-HC group and RES+4-HC group.The CCK-8 was used to measure survival rate of GCs,and DCFH-DA fluorescent probe was used to detect reactive oxygen species(ROS)in GCs.Water-soluble tetrazolium-1(WST-1)was used to detect superoxide dismutase(SOD),and ammonium molybdate method was used to detect catalase(CAT).The protein expression of Silent Information Regulator 2-related enzyme 1(SIRT1)was determined by Western Blot.Results GCs were treated with 4-HC(0,1.0,2.5,5.0,7.5μmol/L).CCK-8 results showed the survival rate of GCs decreased with the increase of 4-HC,which showed significant differences,as compared with the control group(P<0.05).2.5μmol/L 4-HC was selected as the subsequent concentration of 4-HC model.RES(0,5,10,25,50,75μmol/L)was used to treat GCs for 2 h and then added with 2.5μmol/L 4-HC.The results showed that RES improved the survival rate of GCs,suggesting significant differences compared with the 4-HC group(P<0.05).50.0μmol/L RES was selected as the subsequent improvement concentration.The detection results of GCs after treatment in the three groups showed that survival rate of GCs,SOD and CAT decreased,and ROS increased in 4-HC group,compared with CON group(P<0.05).Compared with the 4-HC group,the survival rate of GCs,SOD and CAT in the RES+4-HC group significantly increased,and the ROS significantly decreased.Western blot showed that SIRT1 in the RES+4-HC group significantly increased,suggesting significant difference,

关 键 词：白藜芦醇 环磷酰胺 体外受精 

分 类 号：R711.5[医药卫生—妇产科学]