瘦素对牦牛乳腺上皮细胞中SOCS3和STAT3蛋白表达的影响及催乳素的作用

Effect of leptin on the expression of SOCS3 and STAT3 proteins in yak mammary epithelial cells and the role of prolactin

[目的]本文旨在了解瘦素(LEP)对牦牛乳腺上皮细胞(YMEC)中细胞信号转导抑制因子3(SOCS3)和信号传导及转录激活蛋白3(STAT3)蛋白表达的作用特点及催乳素(PRL)对LEP功能发挥可能产生的影响。[方法]以牦牛乳腺上皮细胞为载体,以添加和不添加催乳素(500 ng·mL^(-1))为前提,用不同剂量瘦素(0、50、100、200、400、800 ng·mL^(-1))作用YMEC 48 h以及添加JAK2/JAK3信号通路阻滞剂AG490作用48 h,采用Western blot技术检测SOCS3和STAT3蛋白的相对表达水平及STAT3蛋白磷酸化水平。[结果]无催乳素时,除50 ng·mL^(-1)瘦素外,其余各剂量均抑制SOCS3蛋白的表达,而各浓度LEP均抑制STAT3蛋白的表达,STAT3蛋白磷酸化水平高于蛋白表达水平。有催乳素时,SOCS3蛋白除了在100 ng·mL^(-1)LEP下被抑制之外,其余各剂量均有促进作用;800 ng·mL^(-1)LEP对STAT3蛋白表达有显著抑制作用,STAT3磷酸化水平除100 ng·mL^(-1)组外,均有所升高。添加AG490之后,SOCS3与STAT3都主要是通过JAK2激活诱导。[结论]高浓度瘦素会抑制SOCS3及STAT3蛋白的表达,瘦素主要通过JAK2/STAT3通路发挥生理功能,而SOCS3是JAK2/STAT3通路的负反馈调节因子;PRL能够缓解高浓度LEP导致的瘦素抵抗作用,并且能促进SOCS3和STAT3蛋白的表达。

[Objectives]The aim of this study was to understand the effect of leptin(LEP)on the expression of SOCS3 and STAT3 proteins in yak mammary epithelial cells(YMEC)and the possible effects of prolactin on leptin function.[Methods]YMEC were used as carriers,with different dosage of LEP(0,50,100,200,400 and 800 ng·mL^(-1))applied for 48 h and JAK2/JAK3 signal pathway blocker AG490 applied for 48 hours.The relative expression of SOCS3 and STAT3 at protein and STAT3 protein phosphorylation were detected by Western blot.[Results]In the absence of prolactin,all concentrations except 50 ng·mL^(-1)LEP inhibited the protein expression of SOCS3,while all concentrations of LEP inhibited the expression of STAT3 protein,and the phosphorylation level of STAT3 protein was higher than that of protein.In the presence of prolactin,SOCS3 protein was inhibited at the dosage of 100 ng·mL^(-1)LEP,and the other dosage promoted it;800 ng·mL^(-1)LEP significantly inhibited the expression of STAT3 protein,and the expression of STAT3 phosphorylation increased except 100 ng·mL^(-1)group.After adding AG490,it was found that SOCS3 and STAT3 were mainly induced by JAK2 activation,and the upstream factors of SOCS3 and STAT3 protein phosphorylation might not be just JAK2.[Conclusions]High dosage of leptin inhibits the expression of SOCS3 and STAT3 protein.Leptin exerted its physiological effects through the JAK2/STAT3 pathway,while SOCS3 is a negative feedback regulator of JAK2/STAT3 pathway;PRL can relieve leptin resistance caused by high LEP concentrations and can promote the expression of SOCS3 and STAT3 proteins.