贞芪颗粒抑制耐甲氧西林金黄色葡萄球菌黏附及侵袭人肺上皮细胞的作用机制研究  

Mechanism of Zhenqi Granules against the adhesion and invasion of MRSA to human lung epithelial cells

作　　者：夏玉文 张璐璐 冯小玉 温博 安洪萨 陈艺幻 吕诚[1] 杨伟峰[2] 谭勇[1] XIA Yuwen;ZHANG Lulu;FENG Xiaoyu;WEN Bo;AN Hongsa;CHEN Yihuan;LV Cheng;YANG Weifeng;TAN Yong(Institute of Basic Research in Clinical Medicine,China Academy of Chinese Medical Sciences,Beijing 100700,China;Medical Experimental Center,China Academy of Chinese Medical Sciences,Beijing 100700)

机构地区：[1]中国中医科学院中医临床基础医学研究所,北京100700 [2]中国中医科学院医学实验中心,北京100700

出　　处：《北京中医药》2024年第11期1287-1292,共6页Beijing Journal of Traditional Chinese Medicine

基　　金：中国中医科学院自主选题项目(Z0735);中国中医科学院科技创新工程中医临床基础学科创新团队项目(C2021B003);中国中医科学院科技创新工程中医临床基础学科重大攻关项目子课题(CI2021A00704-2)。

摘　　要：目的探究贞芪颗粒(ZQ)对耐甲氧西林金黄色葡萄球菌(MRSA)黏附及侵袭人肺上皮细胞(BEAS-2B细胞)的抑制作用与机制。方法常规培养BEAS-2B细胞,使用不同浓度(7.5、10、12.5 mg/mL)ZQ处理细胞,同时设置对照组(Control组,只含DMEM培养液的细胞),细胞计数试剂盒8(CCK-8)法测算细胞活力,以对细胞活力无显著影响的浓度作为ZQ的实验浓度。取部分细胞随机分为Control组、ZQ组,ZQ组给予实验浓度ZQ进行预处理后建立MRSA感染BEAS-2B细胞模型,平板菌落计数法观察MRSA黏附及侵袭细胞的能力;实时荧光定量聚合酶链反应(RT-PCR)法检测ZQ对MRSA黏附基因(fnbA、clfA、clfB、icaA、cna、efb、sasX)以及BEAS-2B细胞紧密连接(TJ)蛋白编码基因(claudin-1、ZO-1、occludin)的影响;Western blotting法检测ZQ对BEAS-2B细胞TJ蛋白表达的影响。结果与Control组比较,7.5、10 mg/mL浓度的ZQ组细胞活力高(P<0.01),12.5 mg/mL组细胞活力差异无统计学意义(P>0.05)。因此选取12.5 mg/mL为ZQ的实验浓度。与Control组比较,ZQ组MRSA菌落数少(P<0.05),相对黏附率、相对侵袭率低(P<0.05),fnbA、clfA、clfB、icaA、cna基因表达低(P<0.05),efb、sasX表达差异无统计学意义(P>0.05),claudin-1、ZO-1、occludin基因与TJ蛋白表达均高(P<0.01)。结论ZQ可以抑制MRSA对BEAS-2B细胞的黏附及侵袭,该作用与降低MRSA黏附基因和提升TJ蛋白表达有关。Objective To investigate the inhibitory effect and mechanism of Zhenqi Granules(ZQ)on the adhesion and invasion of Methicillin-resistant Staphylococcus aureus(MRSA)to human lung epithelial cells(BEAS-2B).Method BEAS-2B cells were cultured routinely,and different concentrations of ZQ(7.5,10,12.5 mg/mL)were used to treat the cells.A control group(cells cultured with DMEM only)was also set up.The cell viability was measured using the cell counting kit-8(CCK-8)assay,and the concentration with no significant effect on cell viability was selected as the experimental concentration for ZQ.A portion of the cells was randomly divided into the control group and the ZQ group.The ZQ group was pretreated with the experimental concentration of ZQ before establishing an MRSA infection model in BEAS-2B cells.The adhesion and invasion abilities of MRSA were observed by plate colony counting.The effect of ZQ on MRSA adhesion genes(fnbA,clfA,clfB,icaA,cna,efb,sasX)and BEAS-2B cell tight junction(TJ)protein-coding genes(claudin-1,ZO-1,occludin)was detected by real-time fluorescence quantitative PCR(RT-PCR).Western blotting was used to assess the effect of ZQ on the expression of TJ proteins in BEAS-2B cells.Result Compared with the control group,the 7.5 and 10 mg/mL ZQ groups showed higher cell viability(P<0.01),while there was no significant difference in cell viability in the 12.5 mg/mL group(P>0.05).Therefore,12.5 mg/mL was selected as the experimental concentration for ZQ.Compared with the control group,the ZQ group showed fewer MRSA colonies(P<0.05),lower relative adhesion rate and relative invasion rate(P<0.05),and lower expression levels of MRSA adhesion-related genes fnbA,clfA,clfB,icaA,and cna(P<0.05),while the expression of efb and sasX showed no significant difference(P>0.05).The gene and protein expression levels of TJ proteins claudin-1,ZO-1,and occludin were significantly higher in the ZQ group(P<0.01).Conclusion ZQ can inhibit the adhesion and invasion of MRSA to BEAS-2B cells,which is related to the downregulation of

关 键 词：贞芪颗粒 耐甲氧西林金黄色葡萄球菌 人肺上皮细胞 黏附 侵袭 

分 类 号：R285[医药卫生—中药学]