TRAF5通过调控IL-17A/NF-κB信号通路减轻氧-糖剥夺/再灌注引起的心肌细胞炎症和细胞凋亡
TRAF5 Attenuates Myocardial Inflammation and Apoptosis Induced by Oxy-gen-glucose Deprivation/Reperfusion by Regulating Interleukin-17A/Nuclear Factor-κB Signaling Pathway
蔡振璇 1张城炼 1许卫攀1
作者信息
- 1. 黄石市中心医院(湖北理工学院附属医院)心血管内科,湖北黄石 435000
- 折叠
摘要
目的 探讨肿瘤坏死因子受体相关因子5(tumor necrosis factor receptor-associated factor 5,TRAF5)对氧-葡萄糖剥夺/再灌注(oxygen-glucose deprivation/reperfusion,OGD/R)引起的心肌细胞炎症和细胞凋亡的影响及相关分子机制.方法 体外培养大鼠心肌细胞系H9c2细胞,构建OGD/R诱导的H9c2细胞模型,以脂质体转染法将TRAF5过表达质粒(pcDNA-TRAF5)和空载质粒(pcDNA-NC)转染至OGD/R诱导的H9c2细胞.采用实时定量聚合酶链反应(quantitative real time polymerase chain reaction,qRT-PCR)法检测细胞中TRAF5基因表达,CCK-8法测定细胞活力,试剂盒法检测细胞中乳酸脱氢酶(lactate dehydrogenase,LDH)、心肌肌钙蛋白T(cardi-ac troponin T,cTnT)和炎性因子水平,流式细胞术检测细胞凋亡,Western blot法检测细胞中相关蛋白表达.结果 与对照组比较,OGD/R组H9c2细胞中TRAF5 mRNA和蛋白表达量降低,细胞活力降低,细胞中LDH、cTnT、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-6(interleukin-6,IL-6)、白细胞介素-1β(interleukin-1β,IL-1β)、单核细胞趋化蛋白-1(monocyte chemoattractant protein-1,MCP-1)浓度升高,细胞凋亡率升高,细胞中B淋巴细胞瘤 2(B-cell lymphoma-2,Bcl-2)蛋白表达量降低,Bcl-2相关蛋白(B-cell-lympho-ma-2 related protein,Bax)、切割型半胱氨酸天冬氨酸蛋白水解酶-3(cleaved cysteine aspartate proteolytic enzyme-3,cleaved caspase-3)、白细胞介素-17A(interleukin-17A,IL-17A)蛋白表达量及磷酸化核转录因子-κB(phos-phorylated nuclear factor-κB,p-NF-κB)/NF-κB比值升高,差异均有统计学意义(P<0.05).与OGD/R组比较,OGD/R+pcDNA-TRAF5组H9c2细胞中TRAF5 mRNA和蛋白表达量升高,细胞活力升高,细胞中LDH、cTnT、TNF-α、IL-6、IL-1β、MCP-1浓度降低,细胞凋亡率降低,细胞中Bcl-2蛋白表达量升高,Bax、cleaved caspase-3、IL-17A蛋白表达量及p-NF-κB/NF-κB比值降低,差异均有统计学意义(P<0.05).结论 TRAF5可减轻OGD/R引起的心肌细胞炎症和细胞凋亡,其作用机制可能与调控IL-17A/NF-κB信号通路有关.
Abstract
Objectives To investigate the effect of tumor necrosis factor receptor-associated factor 5(TRAF5)on cardio-myocyte inflammation and apoptosis induced by oxygen-glucose deprivation/reperfusion(OGD/R)and its related molec-ular mechanism.Methods Rat cardiomyocyte line H9c2 cells were cultured in vitro,and the OGD/R induced H9c2 cell model was constructed.TRAF5 over expression plasmid(pcDNA-TRAF5)and empty plasmid(pcDNA-NC)were transfected into OGD/R induced H9c2 cells by liposome transfection.The expression of TRAF5 gene was detected by quantitative real time polymerase chain reaction(qRT-PCR).The cell viability was determined by CCK-8.The concen-trations of lactate dehydrogenase(LDH),cardiac troponin T(cTnT)and inflammatory factors were detected by kit.The apoptosis was detected by flow cytometry,and the expression of related proteins was detected by Western blot.Results Compared with control group,the expressions of TRAF5 mRNA and protein in H9c2 cells in OGD/R group decreased,the cell viability decreased,the concentrations of LDH,cTnT,tumor necrosis factor-α(TNF-α),interleukin-6(IL-6),interleukin-1β(IL-1β),monocyte chemoattractant protein-1(MCP-1)increased,the apoptosis rate increased,the expression of B-cell lymphoma-2(Bcl-2)protein decreased,the expressions of Bcl-2 related protein(Bax),cleaved cysteine aspartate proteolytic enzyme-3(cleaved caspase-3),interleukin-17A(IL-17A)protein and the ratio of phos-phorylated nuclear factor kappa B(p-NF-κB)/NF-κB increased,the differences were statistically significant(P<0.05).Compared with OGD/R group,the expressions of TRAF5 mRNA and protein in H9c2 cells in OGD/R+pcDNA-TRAF5 group increased,the cell viability increased,the concentrations of LDH,cTnT,TNF-α,IL-6,IL-1β,MCP-1 decreased,the apoptosis rate decreased,the expression of Bcl-2 protein increased,the expressions of Bax,cleaved caspase-3,IL-17A protein and the ratio of p-NF-κB/NF-κB decreased,the differences were statistically significant(P<0.05).Conclusions TRAF5 can reduce the inflammation and apoptosis of cardiomyocytes induced by OGD/R,and its mecha-nism may be related to the regulation of IL-17A/NF-κB signaling pathway.
关键词
肿瘤坏死因子受体相关因子5/氧-葡萄糖剥夺/再灌注损伤/心肌细胞/炎症反应/细胞凋亡/白细胞介素-17A/核转录因子-κB信号通路Key words
tumor necrosis factor receptor-associated factor 5/oxygen-glucose deprivation/reperfusion injury/cardio-myocytes/inflammatory response/apoptosis/interleukin-17A/nuclear factor kappa B signaling pathway引用本文复制引用
基金项目
湖北省卫生健康委青年人才项目(WJ2019Q011)
出版年
2024
NETL
NSTL
维普
万方数据