采用自由基损伤的方法建立大鼠睾丸间质细胞衰老模型,用流式细胞术检测各组细胞周期的变化;应用RT-qPCR、免疫荧光和Western blot技术检测睾丸间质细胞胰岛素/ IGF-1信号传导通路关键基因的表达水平.结果显示:与正常组相比,衰老组β-半乳糖苷酶阳性细胞率可达60%以上(P<0.05),G1 期所占比重明显增加,S期所占比重明显减少(P<0.05).胰岛素/ IGF-1信号传导通路关键基因表达变化: IRS1、IGF-1、IRS2、INSR在衰老组的表达水平明显低于正常组(P<0.05),IGFBP3在衰老组的mRNA表达水平明显高于正常组(P<0.05),二苯乙烯苷可逆转上述现象,上调IGF-1、INSR、IRS1、IRS2表达,下调IGFBP3的mRNA表达.结果表明二苯乙烯苷通过调控胰岛素/ IGF-1信号传导通路延缓睾丸间质细胞衰老.
A model of rat testicular leydig cell senescence was established by free radical injury. Flow cytometry was used to detect the changes of cell cycle in each group. Detection of mRNA and protein expression levels of key genes of insulin/IGF-1 signaling pathway in leydig cells by RT-qPCR, immunofluorescence and Western blot. The results show that compared with the normal group, the proportion of β-galactosidase positive cells in the aging group was more than 60%(P<0.05), and the proportion of G1 phase increased significantly, while that of S phase decreased significantly(P<0.05); Changes in the expression of key genes of the insulin/IGF-1 signaling pathway:the expression levels of IRS1,IGF-1, IRS2 and INSR in the aging group were significantly lower than those in the normal group(P<- DOI:10.3969/j.issn.1000-1565.2021.03.012二苯乙烯苷调控胰岛素/IGF-1信号通路延缓大鼠睾丸间质细胞衰老的作用机制李少华1,姜丽萍2,牛佳瑶2,王乐2,牛嗣云2,齐峰3(1.河北大学附属医院 新生儿科,河北 保定 071000;2.河北大学 基础医学院,河北 保定 071000;3.保定市第一医院 呼吸科,河北 保定 071000)摘 要:采用自由基损伤的方法建立大鼠睾丸间质细胞衰老模型,用流式细胞术检测各组细胞周期的变化;应用RT-qPCR、免疫荧光和Western blot技术检测睾丸间质细胞胰岛素/ IGF-1信号传导通路关键基因的表达水平.结果显示:与正常组相比,衰老组β-半乳糖苷酶阳性细胞率可达60%以上(P<0.05),G1 期所占比重明显增加,S期所占比重明显减少(P<0.05).胰岛素/ IGF-1信号传导通路关键基因表达变化: IRS1、IGF-1、IRS2、INSR在衰老组的表达水平明显低于正常组(P<0.05),IGFBP3在衰老组的mRNA表达水平明显高于正常组(P<0.05),二苯乙烯苷可逆转上述现象,上调IGF-1、INSR、IRS1、IRS2表达,下调IGFBP3的mRNA表达.结果表明二苯乙烯苷通过调控胰岛素/ IGF-1信号传导通路延缓睾丸间质细胞衰老.关键词:二苯乙烯苷;睾丸间质细胞;延缓衰老;胰岛素/ IGF-1信号传导通路;大鼠中图分类号:Q291 文献标志码:A 文章编号:1000-1565(2021)03-0297-07Regulation mechanism of tetrahydroxystilbene glucoside on insulin/IGF-1 signaling pathway to delay the senescence of rat leydig cells LI Shaohua1, JIANG Liping2, NIU Jiayao2, Wang Le2, NIU Siyun2, QI Feng3(1.Department of Neonatology, Affiliated Hospital of Hebei University, Baoding 071000, China; 2.Basic Medical College, Hebei University, Baoding 071000, China;3.Respiratory Department, Baoding First Hospital, Baoding 071000, China)Abstract: A model of rat testicular leydig cell senescence was established by free radical injury. Flow cytometry was used to detect the changes of cell cycle in each group. Detection of mRNA and protein expression levels of key genes of insulin/IGF-1 signaling pathway in leydig cells by RT-qPCR, immunofluorescence and Western blot. The results show that compared with the normal group, the proportion of β-galactosidase positive cells in the aging group was more than 60%(P<0.05), and the proportion of G1 phase increased significantly, while that of S phase decreased significantly(P<0.05); Changes in the expression of key genes of the insulin/IGF-1 signaling pathway:the expression levels of IRS1,IGF-1, IRS2 and INSR in the aging group were significantly lower than those in the normal group(P<- 收稿日期:2020-05-27 第一作者:李少华(1980—),女,河北定州人,河北大学附属医院主管护师,主要从事生殖衰老方向研究.E-mail:xiaolizhu2007@163.com 通信作者:牛嗣云(1967—),女,河北承德人,河北大学教授,博士,主要从事生殖衰老方向研究.E-mail:nsy1688@163.com齐峰(1966—),男,河北保定人,保定市第一医院主任医师,博士,主要从事内科相关研究.E-mail:1393270999@163.com第3期李少华等:二苯乙烯苷调控胰岛素/IGF-1信号通路延缓大鼠睾丸间质细胞衰老的作用机制0.05), and the mRNA expression levels of IGFBP3 in the aging group were significantly higher than those in the normal group(P<0.05).The above phenomena can be reversed by tetrahydroxystilbene glucoside, the expressions of IGF-1, INSR, IRS1 and IRS2 are up-regulated, and the mRNA expression of IGFBP3 is down-regulated. In this study, it was found that the senescence of testicular leydig cells was delayed by the tetrahydroxystilbene through regulating the insulin/IGF-1 signaling pathway.
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