基于铁死亡途径探讨“秩边透水道”针法改善弱精子症小鼠生殖功能机制研究  

Mechanism research of"Zhibian"(BL54)-toward-"Shuidao"(ST28)acupuncture technique for improving reproductive function in mice with asthenospermia based on the ferroptosis pathway

作　　者：郝健亨 常博雅 靳清凯 任佳 王海军 冀来喜 HAO Jianheng;CHANG Boyd;JIN Qingkai;REN Jia;WANG Haijun;JI Laixi(College of Acupuncture-Moxibustion and Tuina,Chengdu University of TCM,Chengdu 610075,Sichuan Province,China;Second Clinical College,Shanxi University of CM,Jinzhong 030619)

机构地区：[1]成都中医药大学针灸推拿学院,四川成都610075 [2]山西中医药大学第二临床学院,晋中030619

出　　处：《中国针灸》2025年第3期351-360,共10页Chinese Acupuncture & Moxibustion

基　　金：国家自然科学基金资助项目:82074549;山西省中医药管理局基金项目:2023ZYYB048;师怀堂新九针学术流派传承工作室项目:2020PY-LP-03。

摘　　要：目的:探讨“秩边透水道”针法调控铁死亡途径改善弱精子症小鼠生殖功能的可能机制。方法:将60只雄性C57BL/6小鼠随机分成空白组、模型组、针刺组和Fer-1组,每组15只。除空白组外,其余3组小鼠给予环磷酰胺(50 mg·kg^(-1)·d^(-1))腹腔注射诱导弱精子症模型,连续5 d。针刺组小鼠给予“秩边透水道”针法干预,每次20 min;Fer-1组给予铁死亡抑制剂Fer-1溶液(1 mg/kg)腹腔注射。两组均每日1次,连续干预2周。称量各组小鼠睾丸湿重并计算睾丸系数,采用精子质量检测系统检测小鼠精子质量;ELISA法检测小鼠血清睾酮(T)、卵泡刺激素(FSH)、黄体生成激素(LH)含量;HE染色法观察小鼠睾丸和附睾形态;免疫荧光法检测小鼠睾丸活性氧(ROS)表达水平;生化法检测小鼠睾丸组织丙二醛(MDA)、还原型谷胱甘肽(GSH)和总铁离子(TFe)含量;透射电镜观察小鼠睾丸组织线粒体结构;JC-1染色检测小鼠睾丸组织线粒体膜电位水平;实时荧光定量PCR法和Western blot法检测小鼠睾丸组织溶质转运蛋白家族7成员11(SLC7A11)、谷胱甘肽过氧化物酶4(GPX4)、铁蛋白重链1(FTH1)、长链脂酰-Co A合成酶4(ACSL4)mRNA和蛋白表达。结果:与空白组比较,模型组小鼠睾丸湿重和睾丸系数降低(P<0.01);精子浓度和精子活力降低(P<0.01);血清T、FSH和LH含量降低(P<0.01);睾丸内生精小管结构疏松、管腔变形,精子细胞排列紊乱、精子数量减少;附睾小管管壁变薄,管腔内精子数量较少;睾丸组织ROS表达水平及MDA、TFe含量升高(P<0.01),GSH含量降低(P<0.01);线粒体数量减少,嵴结构严重破坏;线粒体膜电位水平降低(P<0.01);睾丸组织SLC7A11、GPX4、FTH1mRNA和蛋白表达降低(P<0.01),ACSL4 mRNA和蛋白表达升高(P<0.01)。与模型组比较,针刺组和Fer-1组小鼠睾丸湿重和睾丸系数升高(P<0.01);精子浓度和精子活力升高(P<0.01);血清T、FSH和LH含量升高(P<0.01);睾丸和附睾组织病理结Objective To investigate the underlying mechanism of"Zhibian"(BL54)-toward-"Shuidao"(ST28)acupuncture technique for improving reproductive function in mice with asthenospermia by regulating ferroptosis pathway.Methods Sixty male C57BL/6 mice were randomly divided into a blank group,a model group,an acupuncture group and a Fer-1 group,15 mice in each one.Except the blank group,the intraperitoneal injection with cyclophosphamide(50·kg^(-1)·d^(-1))was administered to establish the asthenospermia model in the mice of the rest 3 groups for 5 consecutive days.In the acupuncture group,"Zhibian"(BL54)-toward-"Shuidao"(ST28)acupuncture technique was operated in the mice,for 20 min each time;and in the Fer-1 group,Fer-1 solution(1 mg/kg)was injected intraperitoneally.The interventions of these two groups were delivered once daily and for 2 consecutive weeks.The testicular wet weight was measured and the testicular coefficient was calculated.Using sperm quality detection system,the sperm quality was detected.With ELISA used,the contents of testosterone(T),follicle-stimulating hormone(FSH)and luteinizing hormone(LH)in the serum were detected.With HE staining,testicular and epididymal morphology was observed.Immunofluorescence was used to detect the expression of reactive oxygen species(ROS)in the testes.Biochemical assay was conducted to determine the contents of malondialdehyde(MDA),reduced glutathione(GSH),and total iron ion(TFe)in the testicular tissue.Transmission electron microscopy was used to examine mitochondrial structure of the testis,while JC-1 staining was used to assess mitochondrial membrane potential in the testicular tissue.Fluorescence quantitative PCR and Western blot analyses were employed to measure the mRNA and protein expression of solute carrier family 7 member 11(SLC7A11),glutathione peroxidase 4(GPX4),ferritin heavy chain 1(FTH1),and acyl-CoA synthetase long-chain family member 4(ACSL4)in the testicular tissue.Results Compared with the blank group,in the model group,the testicular wet weight and te

关 键 词：弱精子症 “秩边透水道”针法 铁死亡 氧化应激 线粒体 

分 类 号：R245[医药卫生—针灸推拿学]