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目的探讨人脐带间充质干细胞(hucMSC)来源的外体(exosomes,hucMSC-Ex)对脂多糖(LPS)诱导的人肝星状细胞系LX2活化的抑制作用及其可能机制。方法体外培养LX2细胞,分别设PBS对照组、LPS组(100 ng/mL)及hucMSC-Ex组(LPS+150μg/mL hucMSC-Ex),48 h后收集细胞并提取相应的RNA及蛋白质。western blot检测LX2活化相关指标(α-SMA、ColΙα1及TGF-β1蛋白)和凋亡指标(Bcl2、Bax及caspase3蛋白)的表达情况;RT-PCR和荧光定量RT-PCR检测α-SMA mRNA表达水平;MTT法分析hucMSC-Ex对LX2细胞活化后增殖的影响。结果 LPS可刺激LX2细胞活化,活化后的LX2细胞形态由不规则形状向成纤维细胞样改变,并高表达α-SMA、ColΙα1及TGF-β1蛋白,其灰度值分别上升约2.45、2.34、2.77倍(P均<0.05);Bax及caspase3蛋白表达下降而Bcl2蛋白表达增强;LX2增殖能力增强;经hucMSC-Ex处理后,细胞形态由细长向不规则形态转变,α-SMA、ColΙα1及TGF-β1蛋白表达减弱,高表达Bax及caspase3蛋白而Bcl2蛋白表达下降,且细胞增殖减少(F=23.44,P<0.05)。结论 hucMSC-Ex能够抑制LPS诱导的LX2细胞活化,降低其增殖能力并促进细胞凋亡。
Objective To investigate the inhibitory effect of exosomes (hucMSC-Ex) derived from human umbilical cord mesenchymal stem cells (hUCCsC) on the activation of LX2 induced by lipopolysaccharide (LPS) in human hepatic stellate cell line LX2 and its possible mechanism. Methods LX2 cells were cultured in vitro. Cells in PBS control group, LPS group (100 ng / mL) and hucMSC-Ex group (LPS + 150 μg / mL hucMSC-Ex) were cultured in vitro for 48 h, and the corresponding RNA and protein were extracted. The expressions of α-SMA, ColIα1 and TGF-β1 protein and apoptosis proteins (Bcl2, Bax and caspase3) were detected by western blot. The expression of α-SMA mRNA was detected by RT-PCR and real- Level; MTT assay hucMSC-Ex LX2 cells after activation of the impact of proliferation. Results LPS stimulated the activation of LX2 cells. The activated LX2 cells changed from irregular shape to fibroblast-like shape and expressed high levels of α-SMA, ColΙα1 and TGF-β1 protein with gray values of about 2.45 and 2.34 respectively. 2.78-fold (all P <0.05). The expression of Bax and caspase3 protein decreased and the expression of Bcl2 protein increased. The proliferative ability of LX2 protein increased. After treatment with hucMSC-Ex, the cell morphology changed from slender to irregular. The expression of TGF-β1 was decreased, Bax and caspase3 were overexpressed, while the expression of Bcl2 protein was decreased, and the cell proliferation was decreased (F = 23.44, P <0.05). Conclusion hucMSC-Ex can inhibit LPS-induced LX2 cell activation, reduce its proliferation and promote apoptosis.