SOCS-3对OSM诱导的肾小管上皮细胞转分化的影响

Effects of SOCS-3 on OSM induced renal tubular epithelial-myofibroblast transdifferentiation

目的观察细胞因子信号传导抑制蛋白-3(suppressors of cytokine signaling,SOCS-3)对抑瘤素M(oncostatin M,OSM)诱导的人肾小管上皮细胞转分化的影响。方法体外培养人肾近曲小管上皮细胞株(HKC),用Lipofectamine 2000分别转染pCR3.1/SOCS-3表达质粒和pCR3.1空质粒载体,G418筛选阳性克隆,应用OSM(10 ng/ml)进行刺激。培养48 h后收集细胞及上清液,采用Western blot检测SOCS-3、CK18、α-SMA和磷酸化信号传导及转录激活因子3(phospho-Signal transducers and activators of transcription 3,p-STAT3)的表达;采用酶联免疫吸附实验测定细胞上清液中Ⅰ型胶原(collagenⅠ,ColⅠ)和纤维连接蛋白(fibronectin,FN)的分泌;采用RT-PCR检测SOCS-3 mRNA的表达。结果与对照组相比,OSM刺激组肾小管上皮细胞α-SMA和p-STAT3蛋白表达增加,细胞培养上清液ColⅠ和FN的分泌增加,而CK18蛋白表达减少。SOCS-3过表达能抑制OSM刺激引起的α-SMA和p-STAT3蛋白表达,减少ColⅠ和FN的分泌,同时能够部分恢复OSM刺激引起的CK18蛋白表达。结论 SOCS-3过表达能抑制OSM诱导的肾小管上皮细胞转分化,此过程可能与p-STAT3的磷酸化受抑有关。

Purpose To investigate the effect of suppressor of cytokine signaling-3 （ SOCS-3 ） on OSM induced renal epithelial-myofi- broblast transdifferentiation （EMT） in human renal tubular epithelial cells （HKC）. Methods Stable transfections of HKC with pCR3.1 vector and pCR3.1/SOCS-3 were performed with Lipofectamine 2000. After transfection, cells were selected with geneticin and stimulated with OSM （ 10 ng/ml）. Western blot was applied to detect the protein expressions of CK18, α-SMA and phospho-Signal transducers and activators of transcription 3 （p-STAT3）. Enzyme-linked immunoadsorbent assay （ELISA） was used to detect the pro- tein synthesis of Col [ and FN in the supernatants of the HKC. SOCS-3 mRNA was measured by reverse transcription and polymerase chain reaction （RT-PCR）. Results Compared with control group, the protein expression levels of c^-SMA and p-STAT3 were signifi- cantly increased in HKC with OSM stimulation and there was a higher concentration of Col I and FN in the supernatants. However, the expression of CK18 protein was decreased with OSM stimulation. Overexpression of SOCS-3 inhibited OSM-induced high expression of α-SMA protein and the activation of STAT3, and reversed the expression of CK18 protein. Meanwhile, overexpression of SOCS-3 re- duced the concentration of Col I and FN in the supernatants of HKC with OSM stimulation. Conclusion Overexpression of SOCS-3 inhibits OSM-induced EMT maybe partly via blocking the activation of STAT3.