Effects of Bushen Tiaochong Recipe (补肾调冲方) containing serum on ovarian granulosa cell proliferation, steroidogenesis and associated gene expression in rats

Abstract

To observe the effect of bushen tiaochong recipe (BSTCR) on rats' ovarian granulosa cell (GC) proliferation, steroidogenesis and follicle-stimulating hormone receptor (FSHR), and insulin-like growth factor-1 (IGF-1) mRNA expression using serum pharmacological method. Rats' GCs were incubated with 10% blank serum (as negative control group), follicle-stimulating hormone (FSH)-containing serum (S-FSH, as positive control group), or BSTCR (in different dosages) containing serum (S-BSTCR, as the BSTCR groups) for 48 h. 3H-TdR incorporation was then performed; DNA was measured to analyze the distribution of GCs in the cell cycle and their proliferation index (PI) using a flow cytometer; estradiol (E2) and progesterone (P) content in the culture fluid were examined by radioimmunoassay; and levels of FSHR and IGF-1 mRNA expression in GCs were measured by real-time RT-PCR. A dose-dependent increase of 3H-TdR incorporation in GC was shown in the BSTCR groups. Cells in G0/G1 phase had markedly less, while those in S phase had a significantly higher increase in the BSTCR groups compared with the negative control group. A high value of PI was also shown in the BSTCR groups, especially in the high dose group where the influence of cell proliferation was stronger than that in the positive control group. The levels of E2 and P in the BSTCR groups of all dosages were significantly higher than those in the negative control group, and did not show any significant difference compared with those in the positive control group. Levels of FSHR and IGF-1 mRNA expression in the BSTCR groups increased in a dose-dependent manner at levels higher than those in the negative control group. S-BSTCR can obviously stimulate the proliferation and steroidogenesis of ovarian GCs. It is speculated that BSTCR could play a regulatory action on ovarian function through two different pathways of endocrine and autocrine by promoting FSHR and IGF-1 mRNA expression.