过氧化氢对人卵巢颗粒细胞KGN增殖、自噬及凋亡的影响  

Effect of H_(2)O_(2)on proliferation,autophagy and apoptosis of human ovarian granulosa cells KGN

作　　者：冯光恩 朱晓霞 贾瑜琦 柴蔚然[2] 陈崇伟[3] 于保锋 FENG Guangen;ZHU Xiaoxia;JIA Yuqi;CHAI Weiran;CHEN Chongwei;YU Baofeng(Department of Biochemistry and Molecular Biology,School of Basic Medicine,Shanxi Medical University,Taiyuan 030001,Shanxi Province,China;不详)

机构地区：[1]山西医科大学基础医学院生物化学与分子生物学教研室,山西太原030001 [2]上海交通大学医学院附属第九人民医院辅助生殖科,上海200011 [3]山西医科大学第二医院骨科,山西太原030001

出　　处：《中国生物制品学杂志》2024年第9期1056-1061,1069,共7页Chinese Journal of Biologicals

基　　金：山西省自然科学基金(201901D111190)。

摘　　要：目的探讨氧化应激对人卵巢颗粒细胞生长发育的影响。方法体外培养人卵巢颗粒细胞KGN,用不同浓度的过氧化氢(H2O2)处理KGN细胞,采用CCK8法检测细胞增殖水平,流式细胞术检测细胞内活性氧族(reactive oxygen species,ROS)产生量及细胞凋亡情况,Western blot法检测细胞自噬和凋亡相关蛋白的表达水平。结果随着H2O2浓度的升高,颗粒细胞增殖受到明显抑制,且呈剂量依赖性(F分别为4.906、4.825、4.653、4.614和4.587,P均<0.001)。细胞内ROS水平随着H2O2浓度的增加而升高,同时细胞凋亡率不断升高;加入抗氧化剂N-乙酰-L-半胱氨酸(N-acetyl-L-cysteine,NAC)后,ROS水平恢复,细胞凋亡率下降,表明细胞凋亡与ROS的产生有关。H2O2处理后,KGN细胞中LC3-Ⅱ、cleaved-caspase3蛋白的表达水平显著升高(t分别为4.809和5.789,P均<0.05),LC3-Ⅰ、BCL-2、P62蛋白的表达水平显著降低(t分别为4.014、3.982和4.415,P均<0.05),表明H2O2能够诱导KGN细胞的自噬和凋亡。结论H2O2能够抑制人卵巢颗粒细胞增殖,诱导其自噬凋亡。氧化应激状态抑制人卵巢颗粒细胞的生长发育,降低卵母细胞质量,卵巢储备功能下降。Objective To investigate the effect of oxidative stress on the growth and development of human ovarian granulosa cells.Methods Human ovarian granulosa cells KGN were cultured in vitro and treated with different concentrations of hydrogen peroxide(H_(2)O_(2)),which were measured for the cell proliferation by CCK8 assay,detected for the production of intracellular reactive oxygen species(ROS)and the cell apoptosis by flow cytometry,and determined for the expression of proteins related to autophagy and apoptosis by Western blot.Results With the increasing concentrations of H_(2)O_(2),the proliferation of granulosa cells was significantly inhibited in a dose-dependent manner(F=4.906,4.825,4.653,4.614 and4.587,respectively,each P<0.001).The intracellular ROS level increased with the increase of H_(2)O_(2)concentration,and the apoptosis rate also increased continuously.After the treatment of N-acetyl-L-cysteine(NAC),the ROS level recovered and the apoptosis rate decreased,indicating that cell apoptosis was correlated with ROS release.After the treatment of H_(2)O_(2),the expression levels of LC3-Ⅱand cleaved-caspase 3 in KGN cells increased significantly(t=4.809 and 5.789,respectively,each P<0.05),while the expression levels of LC3-Ⅰ,BCL-2 and P62 decreased significantly(t=4.014,3.982 and 4.415,respectively,each P<0.05),which indicated the inducing effect of H_(2)O_(2)on the cell autophagy and apoptosis of KGN cells.Conclusion H_(2)O_(2)can inhibit the proliferation of human ovarian granulosa cells and induce their cell autophagy and apoptosis.Oxidative stress can inhibit the growth and development of human ovarian granulosa cells,decreased the quality of oocytes and the ovarian reserve function.

关 键 词：过氧化氢 人卵巢颗粒细胞 氧化应激 

分 类 号：R339.2[医药卫生—人体生理学]