Regulation Of AMH,AMHR-?,and BMPs(2,6)Genes In Bovine(Bos Taurus)Granulosa Cells

Anti-Mullerian hormone(AMH)is an important marker of ovarian reserve,which is produced by granulosa cells(GCs)from pre-antral and early-antral ovarian follicles in several species,including cattle.This hormone plays a vital role during the recruitment of primordial follicles and follicle stimulating hormone(FSH)-dependent follicular growth.However,the regulatory mechanism of AMH expression in follicles is still unclear.In this study,we compared the expressions of AMH,AMHR-?,BMP2,BMP6,FSHR,LHCGR,PGR,ERa,and ER? genes during follicular development.In-vitro expressions of AMH,AMHR-?,BMP2,and BMP6 genes were investigated in bovine GCs treated by FSH,and the expressions of AMH,AMHR-?,BMP2,BMP6,PGR,ERa,and ER? genes were also observed in bovine GCs after luteinization in vitro.The relationships among their mRNA expressions and hormone level were estimated.The GCs were collected from small(3-8mm).medium(9-12mm)and large size(13-24mm)follicles which were at before,onset and after deviation.respectively.Furthermore,mRNA expressions,hormones(AMH,FSH,LH,P4 and E2),apoptosis of GCs,and cell viability were detected by qRT-PCR,ELISA,flow cytometry,and spectrophotometry,respectively.AMH,AMHR-?,BMP2,and FSHR genes were highly expressed in small and medium follicles compared with large ones.In addition,the highest level of AMH protein(84.14 ± 5.41 ng/ml)was found in medium-size follicles.During follicle development,mRNA expressions of PGR,ER? and ER? in in bovine GCs were higher in medium and large follicles than small ones.However,AMH and BMP2 transcript abundance were the highest in medium-size follicles while AMHR-? was the highest in small size follicIes.Lower doses of FSH(1,5,10 and 25 ng/ml)treatment in vitro increased the viability of bovine GCs while higher doses(50 ng/inl)damaged it,and FSH treatment significantly increased ex pressions of AMH,AMHR-?,and BMP2 at the lower doses,while their expressions were repressed at the higher doses.The optimum concentration of FSH(25 ng/ml)indcuced higher AMH and BMP2 expresions.During in-vitro luteinization process,mRNA expressions of PGR,ERa and ER? were increased while AMH and AMHR-? were declined.Similarly,the concentration of AMH was decreased in luteinized culture media.In summary,AMH,AMHR-II,and BMP2 genes showed higher expressions in GCs from the follicles which developments were depended on FSH;and lower doses of FSH demonstrated a stimulatory effect on AMH and BMP2 expression.The AMH and AMHR-? expressions were significantly decreased during in-vitro luteinization of bovine GCs.Our results provided a better understanding the regulations of AMH,AMHR ?,and BMPs(2,6)in bovine GCs during folliculogenesis,which will be helpful to improve the outcomes of conventional assisted reproductive technologies,such as superovulation and oestrus synchronization in bovines.