热应激对猪骨骼肌卫星细胞增殖和生长的影响（英文）

Heat stress inhibits proliferation, promotes growth, and induces apoptosis in cultured Lantang swine skeletal muscle satellite cells

目的:探究热应激对蓝塘猪骨骼肌卫星细胞增殖与生长的影响,分析其作用机制。创新点:通过体外细胞模型研究了热应激对猪骨骼肌卫星细胞增殖与生长的影响,发现热应激通过Akt/m TOR/S6K途径调控细胞的生长。方法:本试验选用1日龄蓝塘猪背最长肌的骨骼肌卫星细胞,对照组为37°C正常培养,热应激组培养温度为41°C,其它培养条件相同。采用细胞计数、MTT法、流式细胞术等手段分析比较热应激对蓝塘猪骨骼肌卫星细胞增殖与生长的影响;利用实时逆转录聚合酶链式反应(RT-PCR)和蛋白质印迹法(Western blot)等方法,检测了热休克蛋白HSP70、半胱氨酸蛋白酶-3(caspase-3)及哺乳动物雷帕霉素靶蛋白(m TOR)信号通路关键基因的表达。结论:(1)41°C热处理可引起蓝塘猪骨骼肌卫星细胞产生热应激反应;(2)热应激72 h会通过改变细胞周期、抑制细胞增殖并诱导细胞凋亡等共同作用减少细胞数目;(3)热应激通过Akt/m TOR/S6K途径调控细胞的生长。

Proliferation suppression and apoptosis are the prominent characteristics induced by heat stress（HS） in cells, whereas the effects of HS on cell growth（mass accumulation） are unknown. In this study, Lantang swine（an indigenous breed of China） skeletal muscle satellite cells（SCs） were pre-cultured at 37 °C for 24 h. The HS group was subjected to HS at 41 °C, while the control group was maintained at 37 °C. Heat shock protein 70（HSP70） expression and SC size are significantly increased（P〈0.05） by HS, but cell proliferation is suppressed（P〈0.05） and apoptosis is induced（P〈0.05）. HS led to a lower percentage of SCs in the G0/G1 phase（P〈0.05） together with a higher percentage of SCs in the S phase（P〈0.05）. However, the percentage of SCs in the G2/M phase was decreased（P〈0.05） at 48 h but then increased（P〈0.05） at 72 h with HS. In addition, the phosphorylation ratios of protein kinase b（Akt）, ribosomal protein S6 kinase（S6K）, and ribosomal protein S6 were increased（P〈0.05） by HS. Nevertheless, the phosphorylation ratios of the 4E binding protein 1 and the eukaryotic initiation factor-4E were indistinguishable（P〉0.05） from those of the control group. The phosphorylation ratio of the mammalian target of rapamycin（m TOR）（Ser^2448） increased（P〈0.05） within 48 h, and apparent differences were abrogated at 72 h（P〉0.05）. Moreover, cleaved caspase-3 expression was increased at 72 h（P〈0.05）. These findings indicate that HS induces apoptosis and disrupts cell cycle distribution to decrease the number of cells. Additionally, HS can promote SC growth via an activated Akt/m TOR/S6 K signaling pathway.