摘要
[目的]探讨载脂蛋白AⅠ(ApoAⅠ)和载脂蛋白AⅠ结合蛋白(AIBP)对THP-1源性巨噬细胞焦亡的影响。[方法]乳酸脱氢酶(LDH)检测试剂盒评估细胞膜完整性,Hoechst33342/PI染色观察细胞膜通透性,ELISA检测白细胞介素1β(IL-1β)和白细胞介素18(IL-18)等炎症因子水平,Western blot检测细胞焦亡相关蛋白核苷酸结合结构域富含亮氨酸重复序列和含热蛋白结构域受体3(NLRP3)、焦孔素D(GSDMD)、cleaved Caspase-1、IL-1β及IL-18的表达。[结果]氧化型低密度脂蛋白(ox-LDL)呈剂量依赖性上调NLRP3、GSDMD-N、cleaved Caspase-1、IL-1β和IL-18的表达,促进IL-1β、IL-18和LDH释放(均P<0.01),表明ox-LDL以剂量依赖的方式诱导THP-1源性巨噬细胞发生焦亡。ApoAⅠ和AIBP共同处理巨噬细胞能显著下调NLRP3、GSDMD-N、cleaved Caspase-1、IL-1β和IL-18的表达,减少IL-1β、IL-18和LDH释放,抑制ox-LDL诱导的细胞焦亡(P<0.05或P<0.01)。转染ATP结合盒转运体A1(ABCA1)siRNA后,ApoAⅠ和AIBP共同处理对细胞焦亡相关蛋白表达和炎症因子分泌无显著影响(P>0.05)。ApoAⅠ和AIBP共同处理巨噬细胞能显著降低细胞膜上嘌呤能2X7受体(P2X7R)的表达,抑制P2X7R介导的蛋白激酶R(PKR)磷酸化及NLRP3炎性小体组装(P<0.05或P<0.01)。转染P2X7R siRNA后,ApoAⅠ和AIBP共同处理对细胞焦亡相关蛋白的表达和炎症因子分泌无显著影响(P>0.05)。[结论]ApoAⅠ和AIBP通过ABCA1降低细胞膜上P2X7R的表达,抑制P2X7R/PKR/NLRP3介导的巨噬细胞焦亡。
Aim To explore the effects of apolipoprotein AⅠ(ApoAⅠ)and apolipoprotein AⅠ binding protein(AIBP)on THP-1-derived macrophage pyroptosis.Methods The lactate dehydrogenase(LDH)detection kit was used to evaluate cell membrane integrity,Hoechst33342/PI staining was used to observe cell membrane permeability,ELISA was used to detect the levels of inflammatory factors such as interleukin-1β(IL-1β)and interleukin-18(IL-18),Western blot was used to detect the expression of pyroptosis-related protein nucleotide-binding domain leucine-rich repeat and pyrin domain-containing receptor 3(NLRP3),gasdermin D(GSDMD),cleaved Caspase-1,IL-1β and IL-18.Results Oxidized low density lipoprotein(ox-LDL)upregulated the expression of NLRP3,GSDMD-N,cleaved Caspase-1,IL-1β and IL-18 in THP-1-derived macrophages in a concentration-dependent manner,and promoted the release of IL-1β,IL-18 and LDH(P<0.05 or P<0.01),indicating that ox-LDL induced pyroptosis in THP-1-derived macrophages in a concentration-dependent manner.Co-treatment of macrophages with ApoAⅠand AIBP significantly downregulated the expression of NLRP3,GSDMD-N,cleaved Caspase-1,IL-1β and IL-18,reduced the release of IL-1β,IL-18 and LDH,and inhibited ox-LDL induced pyroptosis(P<0.05 or P<0.01).After ATP-binding cassette transporter A1(ABCA1)siRNA transfection,co-treatment with ApoAⅠand AIBP had no significant effect on the expression of pyroptosis-related proteins and secretion of inflammatory factors(P>0.05).Co-treatment of macrophages with ApoAⅠand AIBP significantly reduced the expression of purinergic 2X7R receptor(P2X7R)on the cell membrane,inhibited P2X7R mediated protein kinase R(PKR)phosphorylation and NLRP3 inflammasome assembly(P<0.05 or P<0.01).After P2X7R siRNA transfection,co-treatment with ApoAⅠand AIBP had no significant effect on the expression of pyroptosis-related proteins and secretion of inflammatory factors(P>0.05).Conclusion ApoAⅠand AIBP reduce the expression of P2X7R on the cell membrane through ABCA1,inhibiting P2X7R/PKR/NLRP3 mediated macrophage pyroptosis.
作者
陈梦娇
赵真旺
王斯琦
吴剑锋
刘丹
邹瑾
张敏
CHEN Mengjiao;ZHAO Zhenwang;WANG Siqi;WU Jianfeng;LIU Dan;ZOU Jin;ZHANG Min(Institute of Cardiovascular Disease,Key Laboratory for Arteriosclerology of Hunan Province,Hunan International Scientific and Technological Cooperation Base of Arteriosclerotic Disease,Department of Bioinformatics and Medical Big Data,Hengyang Medical School,University of South China,Hengyang,Hunan 421001,China;School of Basic Medicine,Health Science Center,Hubei University of Arts and Science,Xiangyang,Hubei 441053,China;Department of Physiology,School of Basic Medicine,Qujing Medical College,Qujing,Yunan 655100,China;Department of Cardiovascular Medicine,the Second Affiliated Hospital of University of South China,Hengyang,Hunan 421001,China;Medical Education Department,Guangdong Provincial People's Hospital,Zhuhai Hospital(Jinwan Central Hospital of Zhuhai),Zhuhai,Guangdong 519041,China;Department of Cardiovascular Medicine,the First Affiliated Hospital of University of South China,Hengyang,Hunan 421001,China)
出处
《中国动脉硬化杂志》
2025年第5期402-411,共10页
Chinese Journal of Arteriosclerosis
基金
国家自然科学基金青年基金项目(82400542)
湖南省自然科学基金项目(2024JJ5347)
湖南省教育厅优秀青年项目(22B0415)
湖南省卫生健康委卫生科研项目(W20243007)
湖北省自然科学基金青年基金项目(2023AFB043)
云南省教育厅科学研究基金项目(2022J1522)。
