催乳激素对山羊脂肪酸合酶基因转录活性的影响

Effect of Lactogenic Hormones on the Transcriptional Activity of FASN Gene in Goats

为探讨催乳激素对体外培养的山羊乳腺上皮细胞脂肪酸合酶(fatty acid synthase,FASN)基因转录活性的调控作用,试验分别用不同浓度的胰岛素(insulin,INS)、雌激素(estradiol,E_2)、催乳素(prolactin,PRL)及不同激素组合(INS+PRL、E_2+INS+PRL)处理山羊乳腺上皮细胞24h,提取细胞总RNA,采用实时荧光定量PCR检测催乳激素对FASN基因mRNA表达水平的影响。细胞转染山羊FASN基因启动子报告基因载体,同样用不同浓度的胰岛素、雌激素、催乳素及激素组合处理24h,利用双荧光素酶报告基因系统检测催乳激素对FASN基因启动子活性的影响。结果发现,用雌激素、催乳素处理山羊乳腺上皮细胞后,FASN基因启动子活性及mRNA水平极显著或显著上调(P<0.01;P<0.05),雌激素浓度为10、100μmol/L和催乳素浓度为0.1和1μg/mL时效果最为明显,而胰岛素对FASN基因的启动子活性及mRNA水平没有显著影响(P>0.05)。用不同激素组合(INS+PRL、E_2+INS+PRL)处理细胞均能显著上调FASN基因启动子活性及mRNA表达水平(P<0.05)。结果表明,雌激素和催乳素能够调控FASN基因的转录活性,为进一步研究泌乳过程中FASN基因的分子调控机制提供理论依据。

This study was aimed to investigate the effects of lactogenic hormones on transcriptional activity of fatty acid synthase（FASN）gene in goat mammary epithelial cells cultured in vitro.The goat mammary epithelial cells were treated with different concentrations of insulin（INS）,estradiol（E_2）,prolactin（PRL）and hormone combinations（INS＋PRL,E_2＋INS＋PRL）for24 h,respectively.The relative mRNA expression of FASN gene was measured by Real-time quantitative PCR.Moreover,other cells were transfected with goat FASNgene promoter reporter vectors.Then,the cells were treated with INS,E_2,PRL and hormone combinations（INS＋PRL,E_2＋INS＋PRL）for 24 h,respectively.The promoter activity of FASN gene was detected by dual-luciferase reporter assay system.The results showed that the promoter activity and mRNA level of FASN gene were extremely significantly increased in 10 and 100μmol/L E_2 group（P〈0.01）,and also extremely significantly or significantly increased in 0.1 and 1μg/mL PRL group（P〈0.01;P〈0.05）.However,different concentrations of INS had no effect on the transcription-al activity of FASN gene（P〈0.05）.In addition,hormone combinations（INS＋PRL,E_2＋INS＋PRL）significantly increased the promoter activity and mRNA expression level of FASN gene（P〈0.05）.In conclusion,E_2 and PRL could regulate the transcriptional activity of FASN gene,which provided a theoretical basis for further study of the molecular regulation mechanism of FASN gene during lactation.