Rab18通过JAK2/STAT3下调PLIN2并减少THP-1巨噬细胞脂质蓄积  

Rab18 down-regulates PLIN2 through JAK2/STAT3 pathway and at⁃tenuates lipid accumulation in THP-1 macrophages

作　　者：杨丽[1] 蒋思怦 张荣 郭东铭 张星星 袁中华[1] YANG Li;JIANG Si-peng;ZHANG Rong;GUO Dong-ming;Zhang Xing-xing;YUAN Zhong-hua(Institute of Cardiovascular Diseases,Key Laboratory for Arteriosclerology of Hunan Province,Hunan International Scien-tific and Technological Cooperation Base of Arteriosclerotic Diseases,Hengyang Medical School,University of South China,Hengyang 421001,China;Clinical Anatomy&Reproductive Medicine Application Institute,Hengyang Medical School,University of South China,Hengyang 421001,China;Affillated Hospital of Sichuan Nursing Vocational College,Sichuan Provincial Third People's Hospital,Chengdu 610000,China)

机构地区：[1]南华大学心血管疾病研究所,动脉硬化学湖南省重点实验室,湖南省动脉硬化性疾病国际科技创新合作基地,湖南衡阳421001 [2]南华大学衡阳医学院临床解剖与生殖医学应用研究所,湖南衡阳421001 [3]四川护理职业学院附属医院,四川省第三人民医院,四川成都610000

出　　处：《中国病理生理杂志》2022年第5期769-778,共10页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81973326)。

摘　　要：目的:研究Rab18是否通过Janus激酶2(JAK2)/信号转导及转录激活因子3(STAT3)信号通路下调脂滴包被蛋白2(PLIN2)表达,并影响人THP-1巨噬细胞脂质蓄积。方法:采用氧化低密度脂蛋白(ox-LDL)孵育人THP-1巨噬细胞不同时间(0、6、12和24 h),用Westernblot检测细胞内Rab18、JAK2、STAT3、p-JAK2、p-STAT3及PLIN2蛋白水平,使用油红O染色法观察细胞内脂质蓄积的变化。制备野生型、活化型和失活型Rab18巨噬细胞,予以ox-LDL孵育,Westernblot和RT-qPCR检测细胞内Rab18、JAK2、STAT3、p-JAK2、p-STAT3及PLIN2蛋白水平,细胞免疫荧光技术观察巨噬细胞内p-JAK2和p-STAT3的核转位情况。用JAK2抑制剂AG490预处理活化型Rab18细胞,再用ox-LDL孵育该细胞,Westernblot和RT-qPCR检测JAK2、STAT3、p-JAK2、p-STAT3及PLIN2蛋白水平,细胞免疫荧光技术观察巨噬细胞中PLIN2的表达,油红O染色和GPO-PAP酶法观察细胞内脂质蓄积情况及甘油三酯(TG)含量。结果:ox-LDL处理24 h后,Rab18、JAK2、p-JAK2、STAT3、p-STAT3及PLIN2蛋白水平显著升高(P<0.05),脂质蓄积增加。活化型和野生型Rab18上调JAK2、STAT3、p-JAK2、p-STAT3、p-JAK2/JAK2和p-STAT3/STAT3水平(P<0.05),下调PLIN2蛋白表达(P<0.05),细胞脂质蓄积及TG含量减少;免疫荧光显示Rab18与JAK2和STAT3存在共定位关系,活化型Rab18上调p-JAK2和p-STAT3水平,促进p-STAT3核移位;JAK2抑制剂AG490处理后,PLIN2表达及TG含量显著增加(P<0.05),细胞脂质蓄积增加。结论:Rab18通过JAK2/STAT3信号通路减少PLIN2表达,并抑制人THP-1巨噬细胞脂质蓄积。AIM:To investigate the effect of Rab18 on the accumulation of lipids in human THP-1 macrophages.METHODS:Human THP-1 macrophages were co-incubated with oxidized low-density lipoprotein(ox-LDL)for different time(0,6,12 and 24 h).Western blot was used to detect the protein levels of Rab18,Janus kinase 2(JAK2),signal transducer and activator of transcription 3(STAT3),p-JAK2,p-STAT3 and perilipin 2(PLIN2)in the cells.Oil red O staining was used to observe the intracellular lipid changes.Wild-type,high-activity and low-activity Rab18 cells were prepared and incubated with ox-LDL.Western blot and RT-qPCR were used to detect the protein levels of Rab18,JAK2,STAT3,p-JAK2,p-STAT3 and PLIN2,and cellular immunofluorescence was used to detect the changes of p-JAK2 and p-STAT3 in the macrophages.The high-activity Rab18 cells were pretreated with JAK2 inhibitor AG490,and then treated with ox-LDL.Western blot and RT-qPCR were used to detect the protein levels of JAK2,STAT3,p-JAK2,p-STAT3 and PLIN2,and cellular immunofluorescence was used to detect the change of PLIN2 in the macrophages.Oil red O staining and GPO-PAP enzymatic method were used to observe intracellular lipid accumulation and triglyceride(TG)content.RESULTS:The protein levels of Rab18,JAK2,p-JAK2,STAT3,p-STAT3 and PLIN2 as well as lipid accumulation were increased significantly after ox-LDL treatment for 24 h(P<0.05).High-activity and wild-type Rab18 up-regulated the protein levels of JAK2,STAT3,p-JAK2,p-STAT3,p-JAK2/JAK2 and p-STAT3/STAT3(P<0.05),down-regulated the PLIN2 expression,and reduced lipid accumulation and TG content.Immunofluorescence showed that Rab18 colocalized with JAK2 and STAT3.High-activity Rab18 up-regulated p-JAK2 and p-STAT3 levels(P<0.05),and promoted p-STAT3 nuclear translocation.The JAK2 inhibitor AG490 reversed the decreases in PLIN2 and lipid accumulation induced by high-activity Rab18.CONCLUSION:Down-regulation of PLIN2 by Rab18 through JAK2/STAT3 pathway contributes to the inhibition of lipid accumulation in human THP-1 macrophages.

关 键 词：Rab18蛋白 JANUS激酶2 信号转导及转录激活因子3 脂滴包被蛋白2 脂质蓄积 动脉粥样硬化 

分 类 号：R543[医药卫生—心血管疾病] R363.2[医药卫生—内科学]