积雪草苷对TGF-β1诱导的肺泡上皮细胞增殖和Vimentin蛋白表达影响

Effect of asiatiside on proliferation induced by TGF-β1 and expression of Vimentin protein in alveolar epithelial cells

目的:研究积雪草苷对转化生长因子β1(TGF-β1)诱导的肺泡上皮细胞增殖和波形蛋白(Vimentin)表达的影响。方法:用5 ng/ml的TGF-β1处理A549细胞,同时分别给予40、80、160μg/ml浓度的积雪草苷处理,以正常培养的细胞作为对照,MTT测定各组细胞增殖情况,Western blot检测细胞中上皮间质转化(EMT)标志蛋白Vimentin、钙黏蛋白E(E-cadherin)的表达水平,同时检测诱导型一氧化氮合成酶(i NOS)蛋白水平。收集各组培养液上清,用ELISA法检测白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、白细胞介素-4(IL-4)、干扰素-γ(IFN-γ)、血管内皮生长因子(VEGF)含量。结果:TGF-β1培养后的A549细胞增殖活性升高,细胞间质标志物Vimentin蛋白水平升高,上皮标志物E-cadherin蛋白水平降低,同时细胞中i NOS蛋白水平也升高,细胞分泌的IL-6、TNF-α、IL-4、VEGF水平升高,IFN-γ水平降低,与正常培养的对照细胞比较,差异有统计学意义(P<0. 05)。40、80、160μg/ml的积雪草苷处理以后的细胞增殖活性降低,细胞中间质标志物Vimentin表达减少,上皮标志物E-cadherin表达增多,i NOS蛋白表达减少,细胞分泌的IL-6、TNF-α、IL-4、VEGF减少,IFN-γ增多,与未用积雪草苷处理的细胞比较,差异有统计学意义(P<0. 05)。结论:积雪草苷抑制TGF-β1诱导肺泡上皮细胞增殖活性和EMT,减少细胞分泌炎症因子,调控细胞因子的表达,具有一定的抗肺纤维化作用。

Objective:The effect of asiatiside on the proliferation induced by TGF-β1 and the expression of Vimentin protein in alveolar epithelial cells.Methods:A549 cells were treated with 5 ng/ml TGF-β1,at the same time,asiaticoside was treated with 40,80 and 160μg/ml respectively,the normal cultured cells were used as control.The proliferation of cells in each group was measured by MTT.The expression level of EMT marker protein Vimentin and E-cadherin in cells was detected by Western blot.At the same time,the level of iNOS protein was detected.Collect the supernatant of each culture medium,the contents of IL-6,TNF-α,IL-4,IFN-γand VEGF were detected by ELISA.Results:The proliferation activity of A549 cells increased after TGF-β1,the level of Vimentin protein was elevated in the cell medium marker,the level of E-cadherin protein in the epithelial marker was decreased,at the same time,the level of iNOS protein in the cells also increased,the levels of IL-6,TNF-α,IL-4 and VEGF increased,and the level of IFN-γdecreased,compared with normal cultured control cells,the difference was statistically significant(P<0.05).The cell proliferation activity was reduced after treatment with asiaticoside 40,80 and 160μg/ml,the expression of cell medium marker Vimentin was reduced,the expression of E-cadherin in the epithelial marker was increased,the expression of iNOS protein was reduced,the secretion of IL-6,TNF-α,IL-4,VEGF decreased,and IFN-γincreased,compared with the cells treated with unused Asiatic glycoside,the difference was statistically significant(P<0.05).Conclusion:Asiatiside inhibits proliferation and EMT in alveolar epithelial cells induced by TGF-β1,reduce cell secretion of inflammatory factors,regulate the expression of cytokine,it has some anti pulmonary fibrosis effect.