松果菊苷对衰老大鼠生精细胞胰岛素通路IRS1、IGF-1、IGFBP3基因表达的影响

The effects of echinacoside on IRS1, IGF1 and IGFBP3 genes of insulin signaling pathways in aging spermatogenic cell model of rats

目的探讨松果菊苷对衰老大鼠生精细胞胰岛素信号通路胰岛素受体底物(IRS)1、胰岛素样生长因子(IGF)-1、IGF结合蛋白(IGFBP)3基因表达的影响。方法选用原代大鼠生精细胞为研究对象,采用自由基损伤的方法建立细胞衰老模型。运用实时荧光定量-聚合酶链反应(qRT-PCR)、Western印迹、免疫荧光技术,观察松果菊苷对生精细胞胰岛素信号传导通路关键基因IRS1、IGF-1、IGFBP3mRNA和蛋白表达变化。结果衰老组IRS1、IGF1的荧光免疫染色的平均光密度明显低于正常组(P<0.05),IGFBP3平均光密度明显高于正常组(P<0.05),Western印迹半定量分析结果与荧光免疫结果一致。衰老组IRS1、IGF1mRNA表达水平明显低于正常组(P<0.05),IGFBP3mRNA表达水平明显高于正常组(P<0.05)。松果菊苷干预后生精细胞中IRS1,IGF1mRNA和蛋白表达明显高于衰老组(P<0.05),IGFBP3表达明显低于衰老组(P<0.05)。结论松果菊苷可通过降低IGFBP3表达,提高IRS1,IGF1表达,延缓生精细胞的衰老状态。

Objective To investigate the effects of echinacoside on IRS1, IGF1 and IGFBP3 genes of insulin signaling pathways in aging spermatogenic cell model of rats. Methods The primary rat spermatogenic cells were selected as the research subjects. Free-radical oxidative damage method was used to establish cell aging model. qRT-PCR, Western blot, immunofluorescence technology were used to observe the changes of the expressions of IRS1, IGF-1 and IGFBP3 mRNA and protein in the insulin signaling pathway of spermatogenic cells. Results The average optical densities of fluorescent immunostaining of IRS1 and IGF1 in aging group were significantly lower than those in normal group. The average optical density of IGFBP3 was extremely higher than that of normal group. The results of Western blot were consistent with those of fluorescence immunoassay. The expression levels of IRS1 and IGF1 mRNA in aging group were much lower than those in normal group, and the mRNA expression level of IGFBP3 was significantly higher than that in normal group. After administration of echinacoside, the expressions of IRS1, IGF1 mRNA and protein in spermatogenic cells were much higher than those in aging group , and the expression of IGFBP3 was obviously lower than that in aging group. Conclusions Echinacoside could delay the senescence of spermatogenic cells by inhibiting the expression level of IGFBP3 and increasing the expression levels of IRS1 and IGF1.