Effects Of FSH On Gene Expression Related To Steroidogenesis In Bovine Follicles Cultured In Vitro

Multiple Ovulation and Embryo Transfer(MOET)is an effective technique for rapid propagation of excellent cattle herds.However,the effect of superovulation in cattle has not been significantly improved over the past 50 years,and the response of superovulation by exogenous Follicle Stimulating Hormone(FSH)in cattle is variable.Therefore,it is of great significance to study the factors affecting the effect of MOET,especially the effect of FSH on superovulation.The prerequisite of dreamful superovulation result is based on the normal development and ovulation of follicles.The normal development of follicles depends on the endocrine regulation by steroids(progesterone,androgen,estrogen,etc.)and gonadotropin hormones.Therefore,studying the effect of FSH on gene expression related to steroid hormone synthesis in follicles may provide a new way to improve the efficiency of MOET.In this study,a bovine antral follicle culture model in vitro was established,on the basis of this model,the effects of FSH on the expression of steroid synthase genes and gonadotropin receptor genes in granulosa cells and theca cells of follicles were studied by adding exogenous FSH.Aimed to provide theoretical support for improving the superovulation effect of cattle.This research mainly includes 2 aspects.The results are as follows: Experiment 1: To explore the change of expression of steroidogenesis genes in different diameter size of bovine follicles.qRT-PCR used to detect the change of gene expression that related to steroid synthesis in granulose and theca cells in different diameter size follicles of bovine ovaries(the diameter between 3-4 mm is small,5-8 mm is middle,more than 8 mm is big group).The results in granulosa cells showed that the expressions of CYP11A1,CYP19A1,3?-HSD,FSHR and LHR in granulosa cells of large follicles were significantly higher than those in small and medium follicles(P < 0.05),but there was no significant difference between small and medium follicles(P > 0.05);and the expression of 17?-HSD is no significant difference in granulosa cells between different groups.The results of theca cells showed that there was no significant difference in the expression of CYP11A1,3?-HSD,17?-HSD and FSHR between different groups(P > 0.05).The expression of CYP17A1 and LHR in theca cells of large follicles was significantly higher than that in medium and small follicles(P < 0.05).In addition,the results also confirmed that the expression of FSHR was higher in granulosa cells than that of LHR in the same group,while the expression of LHR in theca cells was higher than FSHR.Suggesting that FSHR was mainly expressed in granulosa cells and LHR was mainly expressed in theca cells.Experiment 2: To explore the influence of FSH on gene expression that related with steroidogenesis in bovine antral follicles in vitro.Follicles(9-11 mm)were isolated from bovine ovaries and cultured in vitro for 24 hours with DMEM/F12 supplemented with different concentrations of FSH.The effects of exogenous FSH treatment on the expression of steroidogenesis genes(CYP11A1,3?-HSD,CYP19A1,CYP17A1,17?-HSD)and gonadotropin receptor genes(FSHR,LHR)in bovine follicular granulosa cells and theca cells were identified by qRT-PCR.The results about granulosa cells showed that 25 ng/mL FSH treatment significantly increased the expression of CYP11A1(P < 0.05),10 ng/mL FSH treatment significantly increased the expression of 3?-HSD(P < 0.05),50 ng/mL FSH treatment significantly increased the expression of CYP19A1(P < 0.05).The results suggested that the response of steroid synthase to FSH treatment was significantly sensitive.The results about theca cells showed that 50 ng/ml FSH treatment significantly increased the expression of CYP11A1,3?-HSD,CYP17A1 and 17?-HSD(P < 0.05),but 10 and 25 ng/mL FSH treatment also significantly decreased the expression of CYP11A1,3?-HSD and CYP17A1(P < 0.05).The results about gonadotropin receptor gene expression showed that,contrast with control,in granulosa cells,25 ng/mL FSH treatment significantly increased FSHR mRNA expression(P < 0.05),10 ng/mL FSH treatment increased the expression of LHR(P < 0.05),50 ng/mL FSH treatment was no significant influence on FSHR and LHR(P > 0.05);while 50 ng/mL FSH treatments significantly increased the expression of LHR in theca cells(P < 0.05).