过氧化物酶增殖体激活受体γ通过上调miR-124表达抑制急性肺损伤肺泡巨噬细胞炎症反应

Peroxisome proliferators activated receptor γ protects against acute lung injury alveolar macrophages inflammation by upregulating miR-124 expression

目的探讨PPARγ上调miR-124表达对急性肺损伤(ALI)肺泡巨噬细胞炎症反应的抑制作用及分子机制。方法分离和培养10例ALI和10例健康非吸烟者的肺泡巨噬细胞,检测PPARγ、miR-124及其靶基因TRAF6的表达变化;分别用PPARγ激动剂罗格列酮(RGZ)、PPARγ拮抗剂GW9662或miR-124抑制剂(antagomir-124)干预人肺泡巨噬细胞后,real time RT-PCR检测miR-124及其靶基因TRAF6的表达;人单核细胞THP-1细胞转染含miR-124启动子区的虫荧光素酶报告基因质粒,再经PPARγ激动剂RGZ或拮抗剂GW9662处理,检测报告基因活性;小鼠经LPS刺激后再经PPARγ激动剂RGZ处理,或经antagomir-124预处理后,再经PPARγ激动剂RGZ处理后,real time RT-PCR检测小鼠肺组织中miR-124、炎症相关因子TRAF6、IL-6、TNF-α的表达。结果 ALI患者肺泡巨噬细胞中PPARγ与miR-124的表达均降低,而miR-124靶基因TRAF6表达显著升高;激动剂活化的PPARγ能上调人肺泡巨噬细胞的miR-124,并下调miR-124靶基因TRAF6,而PPARγ拮抗剂GW9662可削弱罗格列酮上调miR-124的表达进而减弱miR-124对其靶基因TRAF6表达的抑制作用,并且miR-124抑制剂减弱PPARγ对TRAF6表达的抑制作用;活化的PPARγ显著促进miR-124启动子活性,而PPARγ拮抗剂GW9662,明显减弱PPARγ激动剂罗格列酮对miR-124启动子的转录活性的上调作用;在LPS诱导的小鼠ALI模型上,活化的PPARγ通过上调miR-124抑制小鼠肺组织IL-6、TNF-α的表达,而PPARγ拮抗剂GW9662和antagomir-124预处理均减弱PPARγ对小鼠肺组织IL-6、TNF-α的表达的抑制作用。结论PPARγ激活后可通过上调miR-124表达抑制ALI肺泡巨噬细胞的炎症反应。

Objective To explore the roles and molecular mechanisms of peroxisome proliferators activated receptor γ（ PPARγ） in suppression of acute lung injury（ ALI） alveolar macrophages inflammation via miR-124 induction. Methods Human alveolar macrophages were collected from 10 cases of ALI and health nonsmokers to detect the expression of PPARγ,miR-124 and TRAF6. Treatment of the PPARγ agonist rosiglitazone（ RGZ）, antagonist GW9662, or miR-124 inhibitor（ antagomir-124） on human alveolar macrophages,the levels of PPARγ,miR-124 and TRAF6 were determined by real time RT-PCR. THP-1 cells were transfected with the miR-124 promoter reporter,and then treated with PPARγ agonist rosiglitazone or antagonist GW9662,the report gene activities were detected. Mice were divided into two groups,one groupwere treated with LPS,then administered antagonist GW9662,another group were pretreatment of antagomir-124,after LPS intraperitoneal injection,the PPARγ agonist rosiglitazone were administered,the lung tissue were collected and the miR-124,TRAF6,IL-6,TNF-αwere detected using real time RT-PCR. Results Both PPARγ and miR-124 were decreased,but miR-124 target gent TRAF6 increased,in alveolar macrophages in ALI patients contrast to health nonsmokers. Activation of PPARγ upregulated the expression of miR-124 and downregulates its target gene TRAF6. The upregulation of miR-124 was attenuated by PPARγ antagonist GW9662,which weaken inhibition of TRAF6 targeted by miR-124. PPARγ agonist RGZ promoted the miR-124 promoter activity significantly,but the antagonist GW9662 attenuated miR- 124 promoter activity that induced by RGZ. In mice with ALI induced by LPS,activation of PPARγ can inhibit the expression of IL-6 and TNF-α in lung. However,the expression of IL-6 and TNF-α of lung tissues in ALI mice were enhanced by treatment of antagonist GW9662 and antagomir-124. Conclusions Activation of PPARγ inhinbits alveolar macrophage inflammatory response by upregulation of miR-124.