益肾通络方调控精母细胞铁死亡治疗精索静脉曲张性不育的机制研究  

Mechanism of Yishen Tongluo Formula in Regulating Ferroptosis of Spermatocytes in Treatment of Varicocele-induced Infertility

作　　者：赵培培 陈建设[2] 孙自学[2] 陈央娣 华众 吕水林[2] ZHAO Peipei;CHEN Jianshe;SUN Zixue;CHEN Yangdi;HUA Zhong;LYU Shuilin(Henan University of Chinese Medicine,Zhengzhou 450000,China;Henan Province Hospital of Traditional Chinese Medicine,Zhengzhou 450002,China)

机构地区：[1]河南中医药大学,郑州450000 [2]河南省中医院,郑州450002

出　　处：《世界中医药》2025年第1期51-60,共10页World Chinese Medicine

基　　金：国家自然科学基金面上项目(82274536);国家中医药管理局高水平中医药重点学科建设项目(国中医药人教函〔2023〕85号);河南省重点科技攻关项目(222102310100);河南省中医药科学研究重大专项(2022ZYZD9);河南省防治生殖障碍疾病中医药重点实验室建设项目(2021ZDSYS10)。

摘　　要：目的:探讨益肾通络方含药血清调控精母细胞铁死亡治疗精索静脉曲张性不育的作用机制。方法:1)收集体外培养的精索静脉曲张(VC)患者正常和曲张的精索静脉内皮细胞进行代谢组学分析,寻找差异显著的致病代谢产物。2)培养小鼠精母细胞系,采用CCK-8法筛选出益肾通络方含药血清及致病代谢产物的最佳浓度,并随机分为正常组、模型组(代谢物组)、含药血清组(代谢物+益肾通络方组),观察各组细胞铁死亡情况;蛋白质免疫印迹法和实时荧光定量PCR检测各组铁死亡相关蛋白谷胱甘肽过氧化物酶4(GSH-Px4)、溶质载体家族7成员11(SLC7A11)、酰基辅酶A合成酶长链家族成员4(ACSL4)及铁蛋白(Fn)的蛋白及其mRNA表达水平。结果:1)铁死亡代谢通路异常与VC关系密切,最终选取氧化谷胱甘肽(GSSG)作为致病代谢产物。2)与正常组比较,模型组细胞存活率明显降低(P<0.01),GSH-Px4、SLC7A11、Fn蛋白及mRNA表达下调,ACSL4蛋白及mRNA表达上调(P<0.05,P<0.01);与模型组比较,给药组细胞存活率明显上升(P<0.01),GSH-Px4、SLC7A11、Fn蛋白及mRNA表达上调,ACSL4蛋白及mRNA表达下调(P<0.01)。结论:1)铁死亡代谢通路与VC性不育关系密切,并最终确定GSSG为主要致病代谢产物。2)益肾通络方可能通过抑制GSSG调节精母细胞铁死亡相关蛋白的表达,发挥治疗精索静脉曲张性不育的作用。Objective:To investigate the mechanism by which Yishen Tongluo Formula-containing serum regulates ferroptosis of spermatocytes to treat varicocele(VC)-induced infertility.Methods:1)Metabolomics analysis was performed on normal and VC human spermatogenic vein endothelial cells cultured in vitro to identify significantly different pathogenic metabolites.2)A mouse spermatocyte cell line was cultured,and the optimal concentration of Yishen Tongluo Formula-containing serum and pathogenic metabolites was determined using the CCK-8 assay.Cells were randomly divided into the normal group,model group(metabolite group),and the formula-containing serum group(metabolite+Yishen Tongluo Formula group).Ferroptosis in each group was observed.The expression of ferroptosis-related proteins,including glutathione peroxidase 4(GSH-Px4),solute carrier family 7 member 11(SLC7A11),acyl-CoA synthetase long-chain family member 4(ACSL4),and ferritin(Fn),and their mRNA levels were measured by Western blot and real-time quantitative PCR.Results:1)The ferroptosis metabolic pathway was closely related to VC,and oxidized glutathione(GSSG)was identified as the main pathogenic metabolite.2)Compared to the normal group,the cell viability in the model group significantly decreased(P<0.01),and the expression of GSH-Px4,SLC7A11,and Fn proteins and mRNA was downregulated,while ACSL4 protein and mRNA expression was upregulated(P<0.05,P<0.01).Compared with the model group,the drug-treated groups showed a significant increase in cell viability(P<0.01),upregulation of GSH-Px4,SLC7A11,and Fn proteins and mRNA expression,and downregulation of ACSL4 protein and mRNA expression(P<0.01).Conclusion:1)The ferroptosis metabolic pathway is closely related to VC-induced infertility,and GSSG is identified as a key pathogenic metabolite.2)Yishen Tongluo Formula may exert its therapeutic effect on VC-induced infertility by inhibiting GSSG and modulating the expression of ferroptosis-related proteins in spermatocytes.

关 键 词：益肾通络方 精索静脉曲张性不育 铁死亡 代谢组学 氧化谷胱甘肽 精索静脉内皮细胞 精母细胞 作用机制 

分 类 号：R277.5[医药卫生—中医学]