[目的]研究微小RNA let-7a(microRNA let-7a,miR-let-7a)对脂磷壁酸(Lipoteichoic acid,LTA)诱导的炎性奶牛乳腺上皮细胞(MAC-T)的增殖和凋亡作用.[方法]通过实时荧光定量PCR(Quantitative real time polymerase chain reaction,qRT-PCR)、实时细胞分析仪和流式细胞术检测过表达和抑制表达miR-let-7a在炎性MAC-T中的影响.[结果]qRT-PCR检测结果显示,转染miR-let-7a模拟物会使炎性MAC-T细胞中miR-let-7a的表达量极显著上调(P＜0.001),转染miR-let-7a抑制剂会使细胞中miR-let-7a的表达量极显著下降(P＜0.001);RTCA结果显示,转染miR-let-7a模拟物能明显促进MAC-T细胞增殖,转染miR-let-7 a抑制物会使MAC-T细胞的增殖受到抑制;流式细胞术结果显示,转染miR-let-7a模拟物的MAC-T细胞的凋亡率显著降低(P＜0.01);而转染miR-let-7a抑制剂则与之相反,MAC-T细胞的凋亡率显著升高(P＜0.01).即过表达miR-let-7a可以促进MAC-T细胞增殖,并降低细胞的凋亡率.而抑制表达miR-let-7a的作用与之相反,抑制表达miR-let-7a使MAC-T细胞的增殖下降,细胞的凋亡率增大.[结论]miR-let-7a是一种抑制炎症调节因子,其过表达可以缓解奶牛乳腺炎,为阐明奶牛乳腺炎的分子调控机制奠定基础.
Effect of miR-let-7a on proliferation and apoptosis of inflammatory bovine mammary epithelial cells induced by lipoteichoic acid
[Objective]The study aimed to investigate the effect of microRNA let-7a(miR-let-7a)on the proliferation and apoptosis of inflam-matory bovine mammary epithelial cells(MAC-T)induced by lipoteichoic acid(LTA).[Method]miR-let-7a was over-expressed and inhibi-ted on inflammatory MAC-T,and its function was detected by Quantitative real time polymerase chain reaction(qRT-PCR),ANNEXIN V-FITC/PI,and real time cell analysis.[Result]The qRT-PCR results showed that transfection of miR-let-7a mimetic significantly upregulated the expression of miR-let-7a in inflammatory MAC-T cells(P＜0.001),while transfection of miR-let-7a inhibitor significantly decreased the expression of miR-let-7a in MAC-T cells(P＜0.001).The RTCA results showed that transfection of miR-let-7a mimetic significantly pro-moted the proliferation of MAC-T cells,while transfection of miR-let-7a inhibitors inhibited the proliferation of MAC-T cells.And the results of flow cytometry showed that the apoptosis rate of MAC-T cells transfected with miR-let-7a mimetic was significantly reduced(P＜0.01);On the contrary,transfection with miR-let-7a inhibitor significantly increased the apoptosis rate of MAC-T cells(P＜0.01).In summary,the results showed that over-expressed of miR-let-7a could promote the proliferation of MAC-T,and reduce the apoptosis rate of MAC-T cells.The effect of inhibiting the expression of miR-let-7a was opposite.Inhibiting the expression of miR-let-7a leaded to a decrease in prolifer-ation of MAC-T cells and an increase in cell apoptosis rate.[Conclusion]miR-let-7a is an anti-inflammatory regulator factor,and its over-ex-pression can alleviate bovine mastitis.These findings lay a foundation for elucidating the molecular regulation mechanism of bovine mastitis.
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