补肾活血方对人胚肺成纤维细胞IL-17A、LC3-Ⅱ影响的实验研究  

Experimental Study of the Bushen Huoxue Prescription on the Regulation Role of IL-17A and LC3-Ⅱ in Lung Fibroblasts

作　　者：刘晓明[1] 李芮[1] 高善语[1] Liu Xiaoming;Li Rui;Gao Shanyu(The Affiliated Hospital of Shandong University of Traditional Chinese Medicine,Jinan 250014,China)

机构地区：[1]山东中医药大学附属医院,济南250014

出　　处：《世界科学技术-中医药现代化》2022年第5期2043-2052,共10页Modernization of Traditional Chinese Medicine and Materia Medica-World Science and Technology

基　　金：山东省中医药管理局山东中医药科技发展计划项目(2019-0088):补肾活血方调控成纤维细胞自噬干预肺纤维化的实验研究,负责人:刘晓明;济南市科学技术局临床医学科技创新计划项目(202019159):补肾活血方调控IL-17A/PI3K/GSK3β信号通路介导细胞自噬研究治疗肺纤维化的作用机制,负责人:刘晓明。

摘　　要：目的 探讨补肾活血方对人胚肺成纤维细胞的作用机制。方法 首先,观察不同浓度的含药血清对人胚肺成纤维细胞的抑制情况及细胞内的胶原蛋白的含量,确定出实验用含药血清的最佳血药浓度;其次,应用TGF-β1诱导人胚肺成纤维细胞,将实验细胞分为空白对照组,TGF-β1处理组,TGF-β1处理+最佳浓度含药血清干预1 h组(含药血清组),TGF-β1处理+基因敲除阴性对照组(sh-NC组),TGF-β1处理+IL-7A基因敲减组(sh-IL-17A组),TGF-β1处理+最佳浓度含药血清干预1 h+过表达质粒阴性对照组(Vector组),TGF-β1处理+最佳浓度含药血清干预1 h+IL-17A过表达质粒组(IL-17AOE组),进行相应药物培养后,检测各组细胞的增值率、胶原蛋白及IL-17A表达量的变化。结果 TGF-β1处理组中人胚肺成纤维细胞的增值率、胶原蛋白、IL-17A蛋白含量及IL-17A mRNA表达量较空白对照组明显增多(P<0.05);含药血清组细胞的增值率、胶原蛋白、IL-17A蛋白含量及IL-17A mRNA表达量较TGF-β1处理组明显较少,而自噬小体的数量较TGF-β1处理组明显增多;sh-IL-17A组细胞的增值率、胶原蛋白、IL-17A蛋白含量及IL-17A mRNA表达量较sh-NC组明显较少,而自噬小体的数量较sh-NC组明显增多;IL-17AOE组细胞的增值率、胶原蛋白及IL-17A mRNA等各项指标较Vector组明显增多,而自噬小体的数量较Vector组明显减少。结论 补肾活血方可以对MRC-5细胞中的胶原显著降解。这一作用与抑制IL-17A介导的通路,激活细胞自噬,进一步降解胶原蛋白有关。Objective To investigate the mechanism of Bushen Huoxue prescription on human embryonic lung fibroblasts.Methods Firstly, the inhibition and the collagen production of different concentrations of drug-containing serum on human embryonic lung fibroblasts were performed to determine the optimal concentration of drug-containing serum for experimental use. Secondly, human embryonic lung fibroblasts were induced by TGF-β1 to establish cell model. The experimental cells were divided into blank control group, TGF-β1 treatment group, TGF-β1 treatment+optimal concentration of drug-containing serum intervention group for 1 hour(drug-containing serum group), TGF-β1treatment+gene knockout negative control group(sh-NC group), TGF-β1 treatment+IL-7A gene knockdown group(shIL-17A group), TGF-β1 treatment+optimal concentration of drug-containing serum for 1h + negative control group(Vector group), TGF-β1 treatment+optimal concentration of drug-containing serum for 1h+IL-17A overexpression plasmid group(IL-17AOE group), TGF-β1 treatment+optimal concentration of drug-containing serum for 1h+negative control group(IL-17AOE group). Further, after the drug treatment, the proliferation rate, expression levels of collagen and IL-17A in each group were detected.Results The growth rate and the expression level of collagen protein, IL-17A mRNA and protein in human embryonic lung fibroblasts with TGF-β1 treatment were significantly higher than the blank control group(P<0.05). Moreover, the growth rate and the expression level of collagen protein, IL-17A mRNA and protein in human embryonic lung fibroblasts with the drug-containing serum treatment group were significantly lower than those in the TGF-β1 treatment group, while the number of autophagosome was significantly higher than that in the TGF-β1 treatment group. The growth rate and the expression level of collagen protein, IL-17A mRNA and protein in human embryonic lung fibroblasts with sh-IL-17A treatment group were significantly lower than those in sh-NC group,while the num

关 键 词：补肾活血方 人胚肺成纤维细胞 COL-Ⅰ IL-17A 自噬小体LC3-Ⅱ 

分 类 号：R285.5[医药卫生—中药学]