摘要
目的探究M1巨噬细胞源外泌体(Exo)中miR-223-3p/HDAC2对肺上皮细胞(BEAS-2B)诱导的慢性阻塞性肺疾病(COPD)炎症模型的作用机制。方法电镜、NTA粒径分析和Western blot鉴定M1巨噬细胞外泌体。香烟烟雾诱导BEAS-2B建立COPD模型,将M1-Exo与COPD模型共培养,免疫荧光检测二者共定位情况,双荧光素酶验证miR-223-3p对HDAC2靶向调控关系。转染miR-223-3p mimic及inhibitor,qRT-PCR检测miR-223-3p、TNF-α、IL-1β和IL-6表达水平,Western blot检测转染后HDAC2蛋白表达。转染PcDNA及PcDNA-HDAC2,ELISA检测炎症因子、Western blot检测转染后HDAC2、CX3CL1蛋白表达。建立COPD小鼠模型,给予Exo及miR-223-3p mimic干预,评估肺组织改变。结果免疫荧光结果显示,Exo出现在BEAS-2B细胞核周围。qRT-PCR显示,miR-223-3p表达在Exo中升高,且炎症因子IL-1β、TNF-α、IL-6水平降低(P<0.05)。双荧光素酶验证miR-223-3p与HDAC2靶向结合。转染miR-223-3p mimic靶向抑制HDAC2,进而促进CX3CL1蛋白表达(P<0.05)。转染PcDNA-HDAC2可下调Exo源miR-223-3p的炎症反应及CX3CL1蛋白水平(P<0.05)。在COPD小鼠模型中,Exo及miR-223-3p mimic可减轻肺泡破坏程度(P<0.01)。结论M1巨噬细胞上调Exo中miR-223-3p表达,减少对HDAC2靶向抑制进而促进CX3CL1表达,Exo-miR-223-3p缓解COPD小鼠肺部炎症反应,减轻肺气肿。
Objective To investigate the effect of miR-223-3p/HDAC2 in M1 macrophage-derived exosomes(Exo)on chronic obstructive pulmonary disease(COPD)induced by lung epithelial cells(BEAS-2B)in a COPD mechanism of action in inflammation model.Methods Electron microscopy,NTA particle size analysis and Western blot identified M1 macrophage exosomes.Cigarette smoke induced BEAS-2B to establish a COPD model,M1-Exo was co-cultured with the COPD model,immunofluorescence was used to detect co-localisation between the two,and dual-luciferase was used to validate the target-regulatory relationship of miR-223-3p on HDAC2.Transfected with miR-223-3p mimic and inhibitor,qRT-PCR detected the expression levels of miR-223-3p,TNF-α,IL-1βand IL-6,and Western blot detected HDAC2 protein expression after transfection.PcDNA and PcDNA-HDAC2 were transfected,and inflammatory factors were detected by ELISA,and HDAC2 and CX3CL1 protein expression were detected by Western blot after transfection.A COPD mouse model was established and Exo and miR-223-3p mimic interventions were given to assess lung tissue alterations.Results Immunofluorescence results showed that Exo appeared around the nucleus of BEAS-2B cells.qRT-PCR showed that miR-223-3p expression was elevated in Exo and the levels of the inflammatory factors TNF-α,IL-1β,and IL-6 were reduced(P<0.05).Luciferase verification of miR-223-3p binding to HDAC2 target.Transfection of miR-223-3p mimic targeted inhibition of HDAC2,which in turn promoted CX3CL1 protein expression(P<0.05).Transfection of PcDNA-HDAC2 down-regulated the inflammatory response and CX3CL1 protein level of Exo-derived miR-223-3p(P<0.05).Exo and miR-223-3p mimic attenuated the degree of alveolar destruction in a COPD mouse model(P<0.01).Conclusions M1 macrophages up-regulate miR-223-3p expression in Exo,reduce inhibition of HDAC2 targeting and thus promote CX3CL1 expression,and Exo-miR-223-3p alleviates inflammatory responses in the lungs of COPD mice and reduces emphysema.
作者
赵龙
李庆虹
赵怡
李男
王敏
夏书月
ZHAO Long;LI Qinghong;ZHAO Yi;LI Nan;WANG Min;XIA Shuyue(Department of Respiratory and Critical Care Medicine,The Affiliated Central Hospital of Shenyang Medical College,Shenyang,Liaoning 110024,China;Shenyang Medical Callege,Shenyang,Liaoning 110034,China)
出处
《中国医药指南》
2025年第10期1-5,共5页
Guide of China Medicine
基金
沈阳市科技局科技计划项目(20-205-4-073)。
关键词
M1巨噬细胞
外泌体
慢性阻塞性肺疾病
炎症
M1 macrophages
Exosomes
Chronic obstructive pulmonary disease
Inflammation
