Yang Lei, Chen Ting, Huang Yuanlu, Yang Yuxuan, Cheng Xiaoe, Wei Fusheng
Department of Gynecology and Obstetrics, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang 330006, Jiangxi Province, PR China.
Department of Anesthesiology and Operation, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang 330052, Jiangxi Province, PR China.
Int Immunopharmacol. 2025 Jun 17;158:114830. doi: 10.1016/j.intimp.2025.114830. Epub 2025 May 17.
Macrophage polarization plays a crucial role in acute respiratory distress syndrome (ARDS). Recently, mounting evidence has uncovered that endothelial progenitor cells (EPCs) secreted exosomes (EPCs-Exos) exert obvious therapeutic effects on the pathological inflammatory process of ARDS, but its potential mechanism is rarely reported.
The primary mouse EPCs and EPCs-Exos were isolated and identified. Absorption of EPCs-Exos by RAW264.7 cells was examined by PKH-26 staining. The polarization of RAW264.7 cells was evaluated by flow cytometry and RT-qPCR analysis. Molecular interactions were verified by dual luciferase assay, RNA pull-down and RNA immunocoprecipitation assays. ARDS mouse model was established, and pathological changes and expressions of related molecules were detected by HE staining, RT-qPCR and western blotting.
EPCs-Exos could be transferred to macrophages, and effectively reversed LPS-induced polarization of macrophages from M2 to M1 phenotype; however, these changes were diminished by activation of TLR4/NF-κB pathway. MiR-103-3p was proved to be enriched in EPC-Exos and could transfer to macrophage and inactivating TLR4/NF-κB pathway via directly binding to TLR4 3'-UTR. Moreover, miR-103-3p overexpression elevated macrophage M2 polarization and repressed M1 polarization in LPS-treated cells by inhibiting TLR4/NF-κB pathway, and knockdown of miR-103-3p in EPC-Exos abolished the regulatory roles of EPC-Exos on macrophage polarization in vitro, and lung inflammatory injury in vivo. HnRNPA2B1 was proved to interact with miR-103-3p and responsible for its exosomal secretion, which repressed pro-inflammatory macrophage polarization.
These findings suggested that hnRNPA2B1-mediated exosomal delivery of miR-103-3p from EPCs protected against macrophage inflammation in ARDS by inactivation of TLR4/NF-κB pathway.
巨噬细胞极化在急性呼吸窘迫综合征（ARDS）中起关键作用。最近，越来越多的证据表明，内皮祖细胞（EPCs）分泌的外泌体（EPCs-Exos）对ARDS的病理性炎症过程具有明显的治疗作用，但其潜在机制鲜有报道。
分离并鉴定原代小鼠EPCs和EPCs-Exos。通过PKH-26染色检测RAW264.7细胞对EPCs-Exos的摄取。通过流式细胞术和RT-qPCR分析评估RAW264.7细胞的极化情况。通过双荧光素酶测定、RNA下拉和RNA免疫沉淀测定验证分子相互作用。建立ARDS小鼠模型，通过HE染色、RT-qPCR和蛋白质免疫印迹法检测病理变化和相关分子的表达。
EPCs-Exos可转移至巨噬细胞，并有效逆转脂多糖（LPS）诱导的巨噬细胞从M2型向M1型表型的极化；然而，这些变化因TLR4/NF-κB通路的激活而减弱。已证明miR-103-3p在EPC-Exos中富集，可转移至巨噬细胞并通过直接结合TLR4的3'-UTR使TLR4/NF-κB通路失活。此外，miR-103-3p过表达通过抑制TLR4/NF-κB通路提高LPS处理细胞中巨噬细胞的M2极化并抑制M1极化，而敲低EPC-Exos中的miR-103-3p消除了EPC-Exos在体外对巨噬细胞极化以及在体内对肺炎症损伤的调节作用。已证明异质性核糖核蛋白A2B1（HnRNPA2B1）与miR-103-3p相互作用并负责其外泌体分泌，从而抑制促炎性巨噬细胞极化。
这些发现表明，hnRNPA2B1介导的EPCs来源的miR-103-3p通过使TLR4/NF-κB通路失活，在ARDS中对巨噬细胞炎症起到保护作用。
