补肾调冲方对大鼠卵巢颗粒细胞增殖与分泌及其IGF-1mRNA表达的影响

目的:观察补肾调冲方对大鼠卵巢颗粒细胞增殖与分泌功能及其IGF-1 mRNA表达的影响。方法:制备原代培养的大鼠卵巢颗粒细胞,预培养24h后,分别加入FSH(50ng/mL)和不同浓度的补肾调冲方(100、10、1、0.1μg/mL),继续培养48h后,放免法测定颗粒细胞培养液中E2、P水平及细胞内cAMP含量,用Real-tim e RT-PCR方法实时定量测定颗粒细胞IGF-1 mRNA的表达量。结果:①补肾调冲方可明显增加3H-TdR掺入颗粒细胞DNA中的数量,并促进颗粒细胞分泌E2、P,增加颗粒细胞中cAMP的含量;②补肾调冲方可促进基础状态下颗粒细胞IGF-1 mRNA的表达,在一定范围内呈剂量依赖性。结论:补肾调冲方在无FSH作用的条件下,可直接促进颗粒细胞内IGF-1 mRNA的表达,使细胞内cAMP的含量增加,从而调节颗粒细胞增殖与分泌功能。

Effects of Bushen Tiaochong Recipe on Rats Ovarian Granulosa Cell Proliferation,Steroidogenesis and IGF - 1 mRNA Expression

Objective:To observe the effect of Bushen Tiaochong Recipe(BSTCR) on rats ovarian granulosa cell proliferation,steroidogenesis and IGF-1 mRNA expression.Methods:Granulosa cells isolated from rats ovaries and cultured in medium containing 20% calf serum for 24 hours preincubation,followed by culture in medium containing FSH(50ng/mL) or varying dosage of BSTCR(100,10,1,0.1 μg/mL) for a further 48 hours.~(3)H-Thymidine incorporation assay were performed using a scintillation counter.Quantification of Estrodial,progestogen and cAMP were examined by radio-immunity.IGF-1 mRNA expression were measured by semiquantitative and real-time RT-PCR(SYBR GreenI).Results:①BSTCR increased thymidine incorporation into DNA of granulosa cells,promoted granulosa cell steroidogenesis and improved the content of cAMP significantly;②BSTCR stimulated the expression of IGF-1 mRNA with a certain dose-dependent manner.Conclusion:In culture of absence of FSH,BSTCR regulates granulosa cells function directly through a increase in IGF-1 mRNA expression,which leads to increase of cAMP.