FSH对体外培养仔猪睾丸支持细胞cyclin D1 mRNA和cyclin E1 mRNA表达的影响

Effect of FSH on the mRNA Expression of Cyclin D1 and Cyclin E1 in the Cultured Immature Boar Sertoli Cell

【目的】确定FSH是否能调节支持细胞cyclin D1 mRNA和cyclin E1 mRNA的表达及可能的机制。【方法】以培养的仔猪睾丸支持细胞为试验材料,通过添加各种信号通路的抑制剂,应用实时荧光定量PCR检测cyclinD1 mRNA和cyclin E1 mRNA的相对表达量。【结果】不同浓度的FSH(0—100 ng.mL-1)均可促进cyclin D1 mRNA和cyclin E1 mRNA的表达(P<0.05),在FSH浓度为50 ng.mL-1时两种基因的表达量最大(P<0.05);FSH(50 ng.mL-1)也以时间依赖的方式促进了cyclin D1 mRNA和cyclin E1 mRNA的表达(P<0.05),在作用30 min时其表达量达到高峰(P<0.05)。不同浓度的Foskolin(0—20μmol.L-1)均可以促进cyclin D1 mRNA和cyclin E1 mRNA的表达(P<0.05),但均低于FSH单独作用时两种基因的表达量;Rp-cAMP(0—40μmol.L-1)、H-89(0—30μmol.L-1)和Verapamil(0—100μg.mL-1)以剂量依赖的方式抑制了FSH诱导的cyclin D1 mRNA和cyclin E1 mRNA的表达(P<0.05),而且Rp-cAMP和Verapamil联合作用对FSH的抑制效果高于单独的抑制效果(P<0.05),但低于两者之和。此外,不同浓度的U0126(0—10μmol.L-1)降低了FSH诱导的cyclin D1 mRNA和cyclin E1 mRNA的表达(P<0.05),而Rp-cAMP、H-89、Verapamil和U0126单独作用时,cyclin D1 mRNA和cyclin E1 mRNA的表达与对照相比没有显著差异(P>0.05)。【结论】FSH以剂量依赖和时间依赖的方式诱导了cyclin D1 mRNA和cyclinE1 mRNA的表达;cAMP-PKA、Ca2+和ERK1/2参与了FSH对cyclin D1 mRNA和cyclin E1 mRNA表达的调节。

【Objective】 The objective of the study is to identify whether or how FSH regulates the mRNA expression of cyclin D1 and cyclin E1 in the cultured immature boar Sertoli cell.【Method】 In the study,the cultured immature boar Sertoli cells were used as materials,and real-time PCR was used to detect the mRNA expression of cyclin D1 and cyclin E1.【Result】 Different doses of FSH（0-100 ng？mL-1） increased the mRNA expression of cyclin D1 and cyclin E1（P0.05）,which were peaking at 50 ng？mL-1（P0.05）.In addition,FSH（50 ng？mL-1） increased the mRNA expression of cyclin D1 and cyclin E1 in a time-dependent manner（P0.05） and peaked at 30 min（P0.05）.Different doses of Forskolin（0-20 μmol？L-1） increased the mRNA expression of cyclin D1 and cyclin E1（P0.05）,but lower compared with FSH alone.Rp-cAMP（0-40 μmol？L-1）,H-89（0-30 μmol？L-1） and Verapamil（0-100 μg？mL-1） inhibited FSH induced the mRNA expression of cyclin D1 and cyclin E1 in a dose-dependent manner（P0.05）,and the combination of Rp-cAMP and Verapamil had more inhibition on FSH activity than individual effect（P0.05）,but lower than addition of Rp-cAMP and Verapamil.Furthermore,different doses of U0126（0-10 μmol？L-1） reduced FSH induced the mRNA expression of cyclin D1 and cyclin E1（P0.05）,while Rp-cAMP,H-89,Verapamil or U0126 alone had no effect on the mRNA expression of cyclin D1 and cyclin E1 as compared to the control（P0.05）.【Conclusion】 FSH regulates the mRNA expression of cyclin D1 and cyclin E1 in a dose-and time-dependent manner,while cAMP-PKA,Ca2＋ and extracellular signal-regulated kinase（ERK1/2） is involved in the regulation of FSH on the mRNA expression of cyclin D1 and cyclin E1.