美沙拉秦介导TGF-β1/Smad信号通路减轻脂多糖诱导的结肠上皮细胞炎症及凋亡

Mesalazine mediates TGF-β1/Smad signaling pathway to alleviate lipopolysaccharide-induced colonic epithelial cell inflammation and apoptosis

目的:探讨美沙拉秦(MS)对脂多糖(LPS)诱导的结肠炎(UC)模型细胞增殖、凋亡和炎症损伤的作用,研究转化生长因子-β1(TGF-β1)/Smad信号通路在其中发挥的功能。方法:体外培养人结肠上皮细胞NCM-460,LPS处理诱导UC模型,分为Con组(不做处理)、LPS组(1 mg/L LPS处理)、MS组(0.1、0.2、0.4 mg/L MS+1 mg/L LPS共处理)及inhibitor组(10μmol/L TGF-β1/Smad信号通路抑制剂LY2109761+0.2 mg/L MS+1 mg/L LPS共处理)。分别用倒置显微镜、EdU法、Hoechst 33258染色法、ELISA和Western blot检测不同处理组NCM-460细胞形态、增殖、凋亡、炎症因子分泌水平及TGF-β1/Smad通路关键蛋白表达。结果:LPS处理导致NCM-460细胞增殖率及Smad7蛋白水平较Con组显著下降,而凋亡细胞数、炎症因子TNF-α、IL-6、可溶性白细胞介素-2受体(sIL-2R)释放量以及TGF-β1、p-Smad2、p-Smad3蛋白表达显著增加(P<0.05);MS显著扭转LPS诱导的上述作用,且呈一定剂量依赖性(P<0.05)。与0.2 mg/L MS组相比,inhibitor组NCM-460细胞增殖率和Smad7表达显著增加,而凋亡细胞数、TNF-α、IL-6、sIL-2R分泌水平以及TGF-β1、p-Smad2、p-Smad3蛋白水平显著降低(P<0.05)。结论:MS可缓解LPS诱导的NCM-460细胞凋亡和炎症反应,该保护作用可能与TGF-β1/Smad信号通路抑制有关。

Objective:To investigate effects of mesalazine(MS)on proliferation,apoptosis and inflammatory injury of cell model of ulcerative colitis(UC)induced by lipopolysaccharide(LPS),as well as transforming growth factor-β1(TGF-β1)/Smad signaling pathway effect in this study.Methods:Human colonic epithelial cells NCM-460 cultured in vitro were induced UC model by LPS,and divided into Con group(no treatment),LPS group(1 mg/L LPS),MS group(0.1,0.2,0.4 mg/L MS+1 mg/L LPS)and inhibitor group(10μmol/L TGF-β1/Smad signaling pathway inhibitor LY2109761+0.2 mg/L MS+1 mg/L LPS).Cell morphology,proliferation,apoptosis and levels of inflammatory factors and TGF-β1/Smad pathway-related markers were examined by inverted microscope,EdU assay,Hoechst 33258 staining,ELISA and Western blot.Results:LPS treatment highly induced cell proliferation rate and Smad7 pro-tein level compared with Con group,while apoptotic cells,inflammatory factors TNF-αand IL-6,soluble interleukin-2 receptor(sIL-2R)release,as well as TGF-β1,p-Smad2,p-Smad3 protein expressions were increased;the above effects induced by LPS was reversed by MS in a dose-dependent manner(P<0.05).Compared with 0.2 mg/L MS group,NCM-460 cells proliferation rate and Smad7 expression were increased,while apoptotic cells,TNF-αand IL-6,sIL-2R releases,and TGF-β1,p-Smad2,p-Smad3 protein expressions were decreased(P<0.05).Conclusion:MS can attenuate LPS-induced apoptosis and inflammatory injury in NCM-460 cells,and this protection was possibly through suppressing TGF-β1/Smad signaling pathway.