M2型巨噬细胞外泌体对皮肤创面愈合影响的探究

The effect of M2 macrophage-derived exosomes on skin wound healing

目的:探究M2型巨噬细胞来源的外泌体(M2exos)对成纤维细胞增殖、迁移的影响。方法:极化RAW264.7细胞并收集M2型巨噬细胞上清,提取并鉴定M2exos。通过免疫荧光观察L929真皮成纤维细胞对M2exos的摄取情况,设对照组和M2exos组,通过CCK-8、细胞划痕实验分别观察在0、24、48、72 h对成纤维细胞的增殖和迁移作用。建立皮肤创口模型,随机将12只C57BL/6J小鼠分为PBS组和M2exos组。两组分别注射200μL PBS、M2exos(500μg/mL),在第1、4、7天观察、拍摄记录,并取材进行HE染色。结果:成功极化得到M2型巨噬细胞,提取和鉴定了M2exos。与对照组相比,M2exos组在48、72 h对成纤维细胞具有促进增殖(t=26.73,P<0.0001;t=6.648,P<0.01)、迁移(t=5.196,P<0.05;t=22.00,P<0.0001)作用。对C57BL/6J小鼠的研究显示,与PBS组相比,在第4、7天时M2exos组创面显著缩小(t=32.80、51.16,均P<0.0001)。结论:M2exos通过促进成纤维细胞增殖和迁移,对小鼠创面愈合有显著的促进作用。

Objective:To investigate the effect of M2 macrophage-derived exosomes(M2exos)on the proliferation and migration of fibroblasts.Methods:RAW264.7 cells were polarized and M2 macrophages supernatant was collected to extract and identify M2exos.The uptake of M2exos by L929 dermal fibroblasts was observed by immunofluorescence,and control group and M2exos group were set up,the proliferation and migration of fibroblasts were observed by CCK-8 assay and cell scratch assay at 0,24,48 and 72 h.The skin wound model was established.Twelve C57BL/6J mice were randomly divided into PBS group and M2exos group.The two groups were injected with 200μL PBS and M2exos(500μg/mL),respectively.The mice were observed and recorded on the 1st,4th and 7th day,and samples were taken for HE staining.Results:M2 macrophages were successfully polarized,extracted and identified M2exos.CCK-8 assay showed that M2exos promoted the proliferation(t=26.73,P<0.0001;t=6.648,P<0.01)and migration(t=5.196,P<0.05;t=22.00,P<0.0001)of fibroblasts at 48 h and 72 h compared with PBS group.The study of C57BL/6J mice showed that,the wound surface of M2exos group was significantly smaller than those in PBS group on the 4th and 7th day(t=32.80,51.16,all P<0.0001).Conclusion:M2exos can promote wound healing in mice by promoting the proliferation and migration of fibroblasts.