重组胰岛素样生长因子-1对3C基因敲除小鼠卵巢胰岛素样生长因子-1受体相关表达和雌激素分泌的影响

Effect of recombinant insulin-like growth factor-1 (IGF-1) on ovarian IGF-1 receptor expression and estrogen secretion in MEX3C knockout mice

目的探讨重组胰岛素样生长因子-1(IGF-1)对MEX3C基因敲除小鼠卵巢发育的影响。方法采用聚合酶链式反应(PCR)法鉴定4周龄FVB(SPF级)小鼠基因型,将鉴定出的雌性小鼠随机分为4组(每组6只):野生型组、MEX3C基因敲除小鼠纯合子(简称纯合子组)、纯合子+IGF-1处理组、处理对照组。比较IGF-1注射前后各组小鼠体质量及卵巢湿重变化;酶联免疫吸附法(ELISA)测定血清中雌二醇水平;HE染色观察卵巢形态结构;免疫组织化学法和Western blotting法测定MEX3C、重组胰岛素样生长因子-1受体(IGF-1R)和p-AKT(Ser473)蛋白表达。结果IGF-1注射前,纯合子组体质量[(6.33±0.31)g]明显轻于野生型组[(15.20±0.35)g],差异有统计学意义(P<0.001)。ELISA结果显示,与处理对照组[(8.1248±0.8477)ng/L]相比,纯合子+IGF-1处理组[(17.2453±0.0731)ng/L]血清雌二醇水平显著较高,差异有统计学意义(P<0.001)。HE染色结果显示,与处理对照组相比,纯合子+IGF-1处理组中原始卵泡数(19.83±2.94)和初级卵泡数(15.50±2.69)增多,且闭锁卵泡数(7.17±1.42)减少,差异均有统计学意义(P均<0.05)。Western blotting结果证实,与处理对照组相比,纯合子+IGF-1处理组IGF-1R和p-AKT蛋白表达显著升高,差异有统计学意义(P=0.0141,P=0.0025)。结论IGF-1促进MEX3C基因敲除小鼠卵巢卵泡发育,其机制是通过上调IGF-1R和p-AKT水平,提高血清雌激素水平促进卵泡发育。

Objective To investigate the effect of recombinant insulin-like growth factor-1(IGF-1)on ovarian development in MEX3C knockout mice.Methods Mice were randomly divided into four groups(6 mice in each group):wild type group,homozygous group,homozygous+IGF-1 treatment group,and treatment control group.The weight and wet ovarian weight of mice in each group was compared before and after IGF-1 injection.The serum levels of estradiol was determined using enzyme linked immunosorbent assay(ELISA).The morphological structure of the ovary was observed by HE staining.The expression of MEX3C,IGF-1 receptor(IGF-1R),and the phosphorylation of AKT(Ser473)(p-AKT)were detected by immunohistochemistry and Western blotting.Results Before IGF-1 injection,the body weight of homozygote group[(6.33±0.31)g]was significantly lighter than that in wild type group[(15.20±0.35)g,P<0.001].Serum estradiol level was significantly higher in homozygous+IGF-1 treatment group[(17.2453±0.0731)ng/L]than in treatment control group[(8.1248±0.8477)ng/L,P=0.001].HE staining results showed that compared with the treatment control group,the primitive follicles(19.83±2.94)and primary follicles(15.50±2.69)were increased in homozygous+IGF-1 treatment group,while the atretic follicles(7.17±1.42)were decreased(all P<0.05).Western blotting results further confirmed that compared with the treatment control group,IGF-1R and p-AKT protein expressions were significantly decreased in homozygous+IGF-1 treatment group(P=0.0141,P=0.0025).Conclusion IGF-1 promotes ovarian follicular development in MEX3C knockout mice.The mechanism is related to the up-regulation of IGF-1R and P-AKT protein expression.IGF-1 can increase serum estrogen and promote follicular development.