摘要
目的:探讨积雪草苷对人增生性瘢痕成纤维细胞(HSFB)中miR-564及其介导的TGF-β_(1)/Smad信号通路中相关蛋白表达的影响。方法:HSFB分为对照组、0.05mg/mL、0.1mg/mL、0.5mg/mL和1mg/mL组,其中对照组加入100μL的DMEM培养基进行培养,0.05mg/mL、0.1mg/mL、0.5mg/mL和1mg/mL积雪草苷组分别加入含相应浓度AS的DMEM培养基继续培养。分别采用MTT法检测细胞增殖能力,流式细胞术检测细胞凋亡水平,酶联免疫吸附试验检测细胞MMP-2、COLⅠ和COLⅢ的表达水平,免疫荧光检测细胞α-SMA蛋白的表达,RT-PCR法检测细胞中miR-564的表达,Westernblot检测TGF-β_(1)/Smad信号通路中相关蛋白的表达。结果:与对照组相比,积雪草苷干预后HSFB增殖水平显著降低,凋亡水平显著增加,细胞中MMP-2、COL-Ⅰ、COL-Ⅲ、α-SMA、TGF-β_(1)、p-Smad2,p-Smad3蛋白的表达和miR-564的表达均明显降低,差异均具有统计学意义(P<0.05或P<0.01),其中以1mg/mL组对HSFB作用最佳。结论:积雪草苷能够通过抑制HSFB中miR-564的表达,进而抑制TGF-β_(1)/Smad信号通路的激活,达到促进细胞凋亡、抑制细胞增殖和胶原分泌的作用。
Objective:To investigate the effect of asiaticoside on the expressions of miR-564 and its related proteins of TGF-β_(1)/Smad signaling pathway in human hypertrophic scar fibroblasts(HSFB).Methods:HSFB were divided into the control group,as well as the groups of 0.05 mg/mL,0.1 mg/mL,0.5 mg/mL and 1 mg/mL.HSFB in the control group was added into 100μL DMEM medium,and those in the groups of 0.05 mg/mL,0.1 mg/mL,0.5 mg/mL and 1 mg/mL were added into DMEM medium containing corresponding concentrations of asiaticoside.MTT assay was used to detect cell proliferation,flow cytometry was used to detect apoptosis,enzyme-linked immunosorbent assay(ELISA)was used to detect the expressions of MMP-2,COL-I and COL-III,immunofluorescence was used to detect the expression ofα-SMA protein,RT-PCR was used to detect the expression of miR-564,and Western blot was used to detect the expression of related proteins of TGF-β_(1)/Smad signaling pathway.Results:Compared to those in the control group,the proliferation level of HSFB cells were significantly decreased,whereas the apoptosis level of HSFB cells were significantly increased,the expressions of MMP-2,COL-I,COL-III,α-SMA,TGF-β_(1),p-Smad2,p-smad3 and miR-564 were significantly decreased after asiaticoside intervention(P<0.05,P<0.01);of which the group of 1 mg/mL had the best effect on HSFB.Conclusion:Asiaticoside can promote cell apoptosis,inhibit cell proliferation and collagen secretion by inhibiting the expression of miR-564 in HSFB,and thus to inhibit the activation of TGF-β_(1)/Smad signaling pathway.
作者
王瑶
袁锐
刘国良
袁星星
WANG Yao;YUAN Rui;LIU Guoliang;YUAN Xingxing(The First Affiliated Hospital of Heilongjiang University of Chinese Medicine,Harbin 150040,China;Jiamusi College of Heilongjiang University of Chinese Medicine,Jiamusi 154007,China;Heilongjiang University of Chinese Medicine,Harbin 150040,China;Nangang Branch of Heilongjiang Academy of Chinese Medical Sciences,Harbin 150006,China)
出处
《中医药学报》
CAS
2021年第8期32-36,共5页
Acta Chinese Medicine and Pharmacology
基金
黑龙江中医药大学校科研基金项目(201726,201516)
哈尔滨市科技局青年后备人才(2017RAQXJ184)。
