双氢睾酮对小鼠卵泡颗粒细胞增殖与抗苗勒管激素表达的影响

doi:10.16431/j.cnki.1671-7236.2022.08.019

Abstract

Abstract: 【Objective】 The aim of this study was to investigate the mechanism of dihydrotestosterone (DHT) involved in the regulation proliferation and the expression of anti-Müllerian hormone (AMH) of mouse granulosa cells.【Method】 Kunming mice(3 weeks)were injected with pregnant horse serum gonadotropin (PMSG,10 IU/mouse) to obtain granulosa cells.After granulosa cells was passaged for 48 h,the morphology of granulosa cells was identified by HE staining,the growth curve of granulosa cells was drawn,and the expression of follicle stimulating hormone receptor (FSHR) was identified by immunofluorescence.When the confluence of the second generation granulosa cells reached 50%,they were starved with serum-free DMEM/F12 medium for 12 h,and different concentrations of DHT (0,10^-9,10^-8,10^-7,10^-6,10^-5 mol/L) were added to the medium.After 48 h of culture,the proliferation of granulosa cells was detected.The expression of AMH gene and AMH protein were measured by Real-time quantitative PCR and ELISA,respectively. 10^-6 mol/L flutamide (AR specific inhibitor),10^-7 mol/L DHT,10^-7 mol/L DHT and 10^-6 mol/L flutamide,10^-5 mol/L DHT,10^-5 mol/L DHT and 10^-8 mol/L 11 ketodihydroprosterone (AR specific agonist) were added to the medium of granulosa cells,which were recorded as F,D7,DF,D5 and DK groups respectively,and the control group was not added with drugs.After 48 h of culture,the proliferation of granulosa cells and the content of AMH protein were detected.【Result】 Mouse granulosa cells cultured in vitro were spindle or paving stone shaped,and the growth curve was S-shaped.The cells were generally expressed FSHR.Compared with 0 mol/L DHT group,the cell proliferation of 10^-8 and 10^-7 mol/L DHT group were significantly and extremely significantly increased(P<0.05;P<0.01),and that of 10^-5 mol/L DHT group was decreased significantly (P<0.05). The relative expression of AMH gene and the content of AMH protein in 10^-7 mol/L DHT group were extremely significantly increased (P<0.01).Compared with D7 group,the proliferation of granulosa cells and the content of AMH protein in DF group were extremely significantly decreased (P<0.01).Compared with D5 group,the proliferation of granulosa cells and AMH protein in DK group were extremely significantly increased (P<0.01).【Conclusion】 10^-7mol/L DHT could significantly promote the proliferation and AMH expression of granulosa cells,while 10^-5 mol/L DHT significantly reduced the proliferation and AMH expression.Moreover,AR could mediate DHT to regulate proliferation and AMH expression of granulosa cells.

Key words: mice; granulosa cells; dihydrotestosterone(DHT); androgen receptor(AR); anti-Müllerian hormone(AMH); cell proliferation

CLC Number:

YU Kai, ZHAO Yufen, WENG Yu, WANG Qin, HAO Shaoyu, YU Boyang, DU Chenguang, SUBUDENG Gerile, LI Haijun. Effects of Dihydrotestosterone on Proliferation and Expression of Anti-Müllerian Hormone of Mouse Granulosa Cells[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(8): 3036-3043.

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Effects of Dihydrotestosterone on Proliferation and Expression of Anti-Müllerian Hormone of Mouse Granulosa Cells

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