摘要
通过建立A549肺泡上皮细胞氧糖剥夺/恢复(oxygen-glucose deprivation/recovery,OGD/R)模型,模拟临床体外循环中肺泡上皮细胞缺血–再灌注过程,观察OGD/R是否引发A549细胞焦亡,为进一步探讨CPB相关肺损伤发生机制提供参考。采用RT-qPCR法和Western blot法检测焦亡相关基因Caspase1、ASC、NLRP3、GSDMD的mRNA转录和蛋白质表达水平,并进一步观察Caspase1特异性拮抗剂VX-765对Caspase1、焦亡效应蛋白GSDMD、培养上清液中Caspase1焦亡效应产物IL-1β、IL-18表达水平以及细胞存活率的影响。结果显示与对照组相比,OGD/R后A549细胞Caspase1活性增高(P<0.05),焦亡相关基因Caspase1、ASC、NLRP3、GSDMD的mRNA和蛋白质表达均上调(P<0.05);采用拮抗剂VX-765预处理后,细胞焦亡相关蛋白Caspase1、GSDMD表达量明显减少(P<0.05),细胞培养上清液中焦亡标志产物细胞因子IL-1β、IL-18水平显著下降(P<0.05),细胞存活率得到恢复。该研究表明,氧糖剥夺/恢复可诱导A549肺泡上皮细胞发生焦亡,抑制细胞焦亡的发生对氧糖剥夺/恢复细胞模型具有保护作用。
To further explore the mechanisms of cardiopulmonary bypass-related acute lung injury,an OGD/R(oxygen-glucose deprivation/recovery)model using A549 alveolar epithelial cell has been established to simulate the ischemia-reperfusion process of alveolar epithelial cell during clinical cardiopulmonary bypass.The pyroptosis of A549 cell caused by OGD/R was observed.The mRNA and protein expression levels of pyroptosis related gene Caspase1,ASC,NLRP3,GSDMD were detected by RT-qPCR and Western blot.Then the effects of Caspase1 inhibitor VX-765 on Caspase1 and GSDMD expression,the A549 cell survival rate,as well as the concentrations of IL-1βand IL-18 in cell culture supernatant were measured,respectively.The results showed that OGD/R induced an significant increase in Caspase1 activity(P<0.05),and the mRNA and protein expression levels of pyroptosis-related gene Caspase1,ASC,NLRP3,GSDMD were up-regulated significantly(P<0.05).When pretreatment with the Caspase1 inhibitor VX-765,the Caspase1 and GSDMD expression levels,the concentration of IL-1βand IL-18 in cell culture supernatant were decreased,and the A549 cell survival rate after OGD/R was restored.This study suggests that OGD/R induces A549 alveolar epithelial cell pyroptosis,and inhibition of the occurrence of pyroptosis has a protective effect on OGD/R cell model,which may be involved in the mechanisms of cardiopulmonary bypass-related lung injury.
作者
王小燕
肖宗懿
易寒
安雨轩
宋娟
陈菲
王寿勇
WANG Xiaoyan;XIAO Zongyi;YI Han;AN Yuxuan;SONG Juan;CHEN Fei;WANG Shouyong(Department of Anesthesiology,Children’s Hospital of Chongqing Medical University,National Clinical Research Center for Child Health and Disorders,Ministry of Education Key Laboratory of Child Development and Disorders,Chongqing Key Laboratory of Pediatrics,Chongqing 400014,China)
出处
《中国细胞生物学学报》
CAS
CSCD
2020年第11期1969-1974,共6页
Chinese Journal of Cell Biology
基金
重庆市卫生计生委医学科研项目(批准号:2015MSXMO40)
国家自然科学基金(批准号:30700785)资助的课题。
