摘要
目的观察芪风固表颗粒含药血清对肺炎克雷伯菌(Kp)感染A549细胞的保护作用。方法大鼠灌胃给予芪风固表颗粒以制备含药血清。A549细胞随机分为空白组、模型组、给药组,空白组和模型组用10%大鼠空白血清干预,给药组用10%芪风固表含药血清干预,培养48 h后,模型组和给药组每孔加入1×10^(8)cfu/mL Kp菌液1 mL,感染4 h后,HE染色观察细胞形态,流式细胞仪检测细胞周期及凋亡,平板计数法观察15、30、60、90 min细胞的细菌粘附数,RT-qPCR法检测细胞caspase5、Pannexin-1、P2RX7 mRNA表达,免疫组化法观察细胞caspase5、Pannexin-1、P2RX7蛋白定位,Western blot法检测细胞caspase5、Pannexin-1、P2RX7、IL-1β蛋白表达。结果与模型组比较,给药组细胞数量增多,S期细胞比例增加(P<0.01),G_(0)/G_(1)期细胞比例,细胞凋亡率,细菌粘附数,caspase5、Pannexin-1、P2RX7 mRNA和蛋白表达,IL-1β蛋白表达均降低(P<0.05,P<0.01)。结论芪风固表颗粒含药血清对肺炎克雷伯菌感染A549细胞引起的急性炎症有一定的预防及缓解作用,其机制可能与抑制caspase5细胞焦亡通路及加速A549细胞周期由G_(0)/G_(1)期向S期转换,减轻肺上皮细胞凋亡有关。
AIM To observe the protective effects of Qifeng Gubiao Granules-containing serum on A549 cells infected with Klebsiella pneumoniae(Kp).METHODS A549 cells were randomly divided into the blank group and the model group for administration of 10%normal rat serum,and the trial group for administration of 10% Qifeng Gubiao Granules-containing serum prepared upon the rat by gavage.After 48 h of culture,1 mL of 1×10^(8) cfu/mL Kp bacterial solution was added into each well of the model group and the trial group for 4 h of infection.Subsequently,the cells had their cell morphology observed by HE staining;their cell cycle and apoptosis detected by flow cytometry;their bacterial adhesion numbers at 15,30,60 and 90 min observed by plate counting method;their mRNA expressions of caspase5,Pannexin-1 and P2RX7 detected by RT-qPCR;their protein localization of caspase5,Pannexin-1 and P2RX7 observed by immunohistochemistry;and their protein expressions of caspase5,Pannexin-1,P2RX7 and IL-1β detected by Western blot.RESULTS Compared with the model group,the trial group displayed increases in number of cells and proportion of S-phase cells(P<0.01);and decreases in proportion of G_(0)/G_(1)-phase cells,apoptosis rate,number of bacterial adhesion,mRNA and protein expressions of caspase5,panexin-1,P2RX7,and IL-1β protein expression(P<0.05,P<0.01).CONCLUSION Qifeng Gubiao Granules-containing serum can prevent and alleviate Kp-triggered acute inflammation in A549 cells,and its mechanism may be related to the the reduction of lung epithelial cell apoptosis becaused of the inhibited pyroptosis by caspase5 pathway and the accelerated A549 cell cycle transition from G_(0)/G_(1)phase to S phase.
作者
王欣
郭雨莹
刘洋
蒙艳丽
王晓溪
王伟明
WANG Xin;GUO Yu-ying;LIU Yang;MENG Yan-li;WANG Xiao-xi;WANG Wei-ming(Heilongjiang Academy of Chinese Medicine Sciences,Harbin 150036,China)
出处
《中成药》
CAS
CSCD
北大核心
2023年第3期755-762,共8页
Chinese Traditional Patent Medicine
基金
国家自然科学基金项目(82174060)
黑龙江省自然科学基金项目(LH2019H089)
黑龙江省中医药科研项目(ZHY2021-WH024)。
