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

doi:10.3969/j.issn.1007-3205.2022.07.018

河北医科大学学报 ›› 2022, Vol. 43 ›› Issue (7): 832-838.doi: 10.3969/j.issn.1007-3205.2022.07.018

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

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

出版日期:2022-07-25 发布日期:2022-07-26
作者简介:吴海峰(1977-)，男，河北磁县人，河北省胸科医院副主任检验师，医学硕士，从事临床检验诊断研究。
基金资助:
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）

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

1.Department of Laboratory Medicine, Hebei Chest Hospital, Shijiazhuang 050041, China;
2. Department of Reproductive Medicine, the Fourth Hospital of Hebei
Medical University, Shijiazhuang 050011, China

Online:2022-07-25 Published:2022-07-26

摘要/Abstract

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(0，1.0，2.5，5.0，7.5 μmol/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2.5 μmol/L 4-HC为后续4-HC模型浓度。RES（0、5、10、25、50、75 μmol /L）处理GCs 2 h后加入2.5 μmol/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50 μmol/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。

关键词: 白藜芦醇, 环磷酰胺, 体外受精

Abstract: 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, 7.5 μmol/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). 2.5 μmol/L 4-HC was selected as the subsequent concentration of 4-HC model. RES(0, 5, 10, 25, 50, 75 μmol/L) was used to treat GCs for 2 h and then added with 2.5 μmol/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). 50.0 μmol/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, compared with the other two groups(P＜0.05).
Conclusion RES possibly ameliorates 4-HC-induced GCs damage by increasing SIRT1.

Key words: resveratrol, cyclophosphamide, fertilization in vitro

吴海峰, 贾建雷, 李倩, 甄秀丽, 魏雅楠, 乜照燕. 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 [J]. 河北医科大学学报, 2022, 43(7): 832-838.

WU Hai-feng, JIA Jian-lei, LI Qian, ZHEN Xiu-li, WEI Ya-nan, NIE Zhao-yan. Resveratrol ameliorates oxidative stress damage of human granulosa cells induced by cyclophosphamide by regulating SIRT1[J]. Journal of Hebei Medical University, 2022, 43(7): 832-838.

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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

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