摘要
目的探讨纤溶酶原激活物抑制物-1(PAI-1)对棕榈酸刺激后小鼠原代心肌细胞凋亡的作用及其可能机制。方法分离培养小鼠原代心肌细胞,分为对照组与棕榈酸组,Western blotting检测PAI-1及凋亡蛋白剪切型的半胱天冬酶3(cleaved-caspase 3)、BCL2关联X蛋白(Bax)表达。建立PAI-1低表达及过表达的心肌细胞,荧光定量PCR检测PAI-1基因的mRNA表达,Western blotting检测PAI-1、cleaved-caspase 3及Bax蛋白的表达。确定PAI-1小干扰RNA条件后给予棕榈酸刺激,分为对照组、PAI-1低表达组、棕榈酸组、PAI-1低表达+棕榈酸组,Western blotting检测cleaved-caspase 3、Bax及磷酸化核因子κB(p-NF-κB)的表达。最后,将棕榈酸组细胞给予NF-κB抑制剂Bay11-7082处理,分为棕榈酸组、PAI-1低表达+棕榈酸组、Bay11-7082+棕榈酸组、PAI-1低表达+Bay11-7082+棕榈酸组,Western blotting检测cleaved-caspase 3及Bax蛋白的表达。结果棕榈酸组的cleaved-caspase 3、Bax及PAI-1蛋白表达水平明显高于对照组;PAI-1低表达+棕榈酸组的cleaved-caspase 3、Bax及p-NF-κB蛋白表达水平明显低于棕榈酸组,差异均有统计学意义(P<0.01)。此外,Bay11-7082+棕榈酸组的cleaved-caspase 3及Bax蛋白表达水平明显低于棕榈酸组,PAI-1低表达+Bay11-7082+棕榈酸组的cleaved-caspase 3及Bax蛋白表达水平明显低于Bay11-7082+棕榈酸组,差异均有统计学意义(P<0.01)。结论 PAI-1通过影响NF-κB通路参与对棕榈酸刺激后心肌细胞凋亡的调控。
Objective To investigate the effect and mechanism of plasminogen activator inhibitor-1(PAI-1) on cardiomyocytes apoptosis stimulated by palmitate.Methods Mouse primary cardiomyocytes were isolated and cultured,and then divided into control group and palmitic acid (PA) group,the expressions of PAI-1,apoptotic protein cleaved-caspase 3 and BCL2-associated X protein (Bax) were examined using Western blotting.PAI-1 low expression cells and over expression cells were established,and the expressions of PAI-1,cleaved-caspase 3 and Bax were examined by real-time PCR and Western blotting.After PAI-1 siRNA conditions were determined,cells were stimulated with palmitate,and then divided into four groups including control,PAI-1 low expression group,palmitate group and PAI-1 low expression+palmitate group.Western blotting was performed to examine the expressions of cleaved-caspase 3,Bax and phosphorylated-nuclear factor kappa-B (p-NF-κB).At last,the cells in palmitate group were given with NF-κB inhibitor Bay11-7082,and then divided into palmitate group,PAI-1 low expression + palmitate group,Bay11-7082 + palmitate group,and PAI-1 low expression + Bay11-7082 + palmitate group,then the expressions of cleaved-caspase 3 and Bax were detected by Western blotting.Results The expressive levels of cleaved-caspase 3,Bax and PAI-1 were significantly higher in palmitate group than those in control group (P<0.01).The expressions of cleaved-caspase 3,Bax and p-NF-κB were obviously lower in PAI-1 low expression+palmitate group than those in palmitate group (P<0.01).Besides,the expressions of cleaved-caspase 3 and Bax were obviously lower in Bay11-7082+palmitate group than those in palmitate group (P<0.01),and the expressions of cleaved-caspase 3 and Bax were markedly lower in PAI-1 low expression+Bay11-7082+palmitate group than those in Bay11-7082+palmitate group (P<0.01).Conclusion By affecting the NF-κB pathway,PAI-1 could involve in the apoptosis regulation of palmitate stimulated cardiomyocytes.
作者
裴培
成小丽
邢瑞楠
田孝祥
刘丹
Pei Pei;Cheng Xiao-Li;Xing Rui-Nan;Tian Xiao-Xiang;Liu Dan(Department of Cardiology,General Hospital of Northern Theater Command,Shenyang 110016,China)
出处
《解放军医学杂志》
CAS
CSCD
北大核心
2020年第5期513-517,共5页
Medical Journal of Chinese People's Liberation Army
基金
沈阳市科技计划项目(19-112-4-056)
辽宁省自然科学基金(20180550368)
军事科技领域青年人才托举工程项目(17-JCJQ-QT-028)。
作者简介
裴培,医学硕士,主要从事冠心病防治的基础研究。E-mail:18900960781@163.cm;通信作者:刘丹,E-mail:ljmuer@sina.com。
