PI3K/PKB信号通路在TGF-β_1诱导人肝星状细胞表达骨桥蛋白中的作用

Effects of PI3K/PKB signaling pathway on expression of osteopontin in human hepatic stellate cells induced by transforming growth factor-β_1

目的:研究磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/PKB)信号通路在转化生长因子β1(TGF-β1)诱导人肝星状细胞表达骨桥蛋白(OPN)的调控作用。方法:体外培养LX-2人肝星状细胞株,予TGF-β1(终浓度2.5、5、10、20μg/L)刺激24 h或予TGF-β1(终浓度10μg/L)刺激12 h、24 h、48 h;先经PI3K/PKB信号通路特异性抑制剂wortmannin(0.1μmol/L)预处理1 h,再予10μg/L TGF-β1刺激24 h,收集细胞,采用real-time PCR及Western blotting法检测OPN表达情况。结果:TGF-β1能够促进LX-2细胞表达OPN,在一定浓度和时间范围内,其表达量随着TGF-β1浓度和时间的增加而增加,呈剂量和时间依赖性关系;经wortmannin预处理再予TGF-β1刺激的LX-2细胞,与对照组相比,OPN表达受到明显抑制(P<0.01)。结论:TGF-β1对LX-2人肝星状细胞OPN表达具有诱导作用,此作用可能受PI3K/PKB信号通路的调控。

AIM：To investigate the regulatory effects of phosphatylinositol 3-kinase/protein kinase B （PI3K/PKB） signaling pathway on the expression of osteopontin （ OPN） in transforming growth factor-β1 （ TGF-β1 ）-induced hu-man hepatic stellate cells .METHODS：Human hepatic stellate cell line LX-2 was cultured in DMEM and stimulated by TGF-β1 at the final concentration of 2.5, 5, 10 and 20 μg/L for 24 h or at final concentration of 10 μg/L for 12 h, 24 h and 48 h.LX-2 cells were pretreated with wortmannin , a specific inhibitor of PI3K/PKB signaling pathway , at final con-centration of 0.1 μmol/L for 1 h, followed by incubation with TGF-β1 at final concentration of 10μg/L for 24 h.The cells were collected.The expression of OPN was detected by real-time PCR and Western blotting .RESULTS： In LX-2 cells, the expression of OPN was apparently elevated when incubated with TGF-β1 .With the increase in TGF-β1 concentration or the extension of incubation hours , the expression of OPN was increased gradually in a dose-and time-dependent manner with certain limits.LX-2 cells pretreated with wortmannin and incubated with TGF-β1 had a significant decrease in the OPN expression as compared with control group （P〈0.01）.CONCLUSION：The expression of OPN in TGF-β1-induced LX-2 cells is regulated by the PI3K/PKB signaling pathway.