下调PDCD4抑制MAP2K3和p38 MAPK的表达减轻LPS诱导的肾小管上皮细胞炎症和凋亡  

Downregulating PDCD4 alleviated the LPS induced inflammation and apoptosis of renal tubular epithelial cells by suppressing the expression of MAR2K3 and p38 MAPK

作　　者：蒋伟[1] 张益楠 张健锋[3] 黄中伟[1] Jiang Wei;Zhang Yinan;Zhang Jianfeng;Huang Zhongwei(Department of Emergency Medicine,the Affiliated Hospital of Nantong University,Nantong 226001,China)

机构地区：[1]南通大学附属医院急诊内科,江苏南通226001 [2]南通大学附属医院皮肤科,江苏南通226001 [3]南通大学附属医院消化内科,江苏南通226001

出　　处：《中国急救医学》2024年第4期305-313,共9页Chinese Journal of Critical Care Medicine

基　　金：南通市科技计划项目(JCZ2022020)。

摘　　要：目的研究程序性细胞死亡蛋白4(programmed cell death protein 4,PDCD4)在脓毒症诱导的急性肾损伤(acute kidney injury,AKI)中的作用机制,以及调控PDCD4表达通过丝裂原活化蛋白激酶3(mitogen-activated protein kinase 3,MAP2K3)和p38蛋白激酶(p38 mitogen-activated protein kinase,p38 MAPK)对脓毒症AKI起到潜在治疗作用。方法用脂多糖(lipopolysaccharide,LPS)刺激人肾小管上皮细胞(HK-2)构建脓毒症AKI细胞模型。进一步用腺病毒介导siRNA和过表达载体抑制和上调AKI细胞模型中PDCD4的表达;CCK-8法检测细胞增殖;用DCFH-DA及激光共聚焦显微镜检测细胞中ROS水平,用总SOD活性检测试剂和MDA检测试剂盒检测细胞中SOD和MDA水平;免疫共沉淀验证PDCD4和MAP2K3之间的蛋白相互作用;TUNEL染色法检测细胞凋亡;RT-qPCR和Western blot检测PDCD4及相关基因的mRNA和蛋白表达水平;ELISA法检测患者血清中炎症相关因子水平。结果LPS诱导可以促进HK-2细胞中PDCD4表达,下调PDCD4可抑制LPS诱导的HK-2细胞的炎症、氧化应激及细胞凋亡。数据库预测及免疫共沉淀证实PDCD4可以与MAP2K3相互作用,且在LPS诱导的HK-2细胞中,MAP2K3表达水平显著增强。MAP2K3过表达和p38 MAPK激动剂可以减轻PDCD4下调对LPS诱导的细胞炎症和氧化应激的影响并抑制细胞凋亡。结论下调PDCD4可以通过抑制MAP2K3和p38 MAPK从而抑制LPS诱导的肾小管上皮细胞的炎症和凋亡。Objective To investigate the mechanism of programmed cell death protein 4(PDCD4)in sepsis-associated acute kidney injury(AKI)and its potential therapeutic effects on sepsis-associated AKI by regulating the expression of mitogen-activated protein kinase 3(MAP2K3)and p38 mitogen-activated protein kinase(p38 MAPK).Methods Lipopolysaccharide(LPS)was used to stimulate human renal tubular epithelial cells(HK-2)to induce sepsis-associated AKI.Next,the expression of PDCD4 was knockdown by using RNA interfering in LPS induced HK-2 cells.Commercial kits and TUNEL staining were used for the detection of cellular inflammation,oxidative stress and apoptosis.Then,co-immunoprecipitation assay was performed for the examination of the association between PDCD4 and MAP2K3.The MAP2K3-overexpressing HK-2 cells were constructed,following which their inflammation and apoptosis were determined by using the same aforementioned methods.Results Knockdown of PDCD4 relieved the LPS-induced inflammation,apoptosis and the higher levels of reactive oxygen species in HK-2.The co-immunoprecipitation assays also showed that PDCD4 can interact directly with MAP2K3.The overexpression of MAP2K3 and the application of a p38 MAPK signal agonist abolished the effect of PDCD4 knockdown on the LPS-induced inflammation,apoptosis and oxidative stress of these cells.Conclusions Inhibition of PDCD4 alleviates the LPS-induced inflammation and apoptosis of renal tubular epithelial cells by suppressing the expression of MAP2K3 and p38 MAPK.

关 键 词：急性肾损伤 脓毒症 程序性细胞死亡蛋白4 丝裂原活化蛋白激酶3 P38蛋白激酶 活性氧 超氧化物歧化酶 丙二醛 

分 类 号：R459.7[医药卫生—急诊医学] R692[医药卫生—治疗学]