人脐带间充质干细胞联合吡非尼酮治疗小鼠肺纤维化

Therapeutic Efficacy of Human Umbilical Cord Mesenchymal Stem Cells Combined with Pirfenidone in Pulmonary Fibrosis Mice

该文旨在研究人脐带间充质干细胞(umbilical cord-derived mesenchymal stem cells,hUC-MSCs)联合吡非尼酮(pirfenidone, PFD)对博来霉素诱导的小鼠肺纤维化的治疗作用及可能机制。C57BL/6小鼠分为正常对照组、MSCs对照组、模型组、30 mg/kg PFD组(P30)、100 mg/kg PFD组、300 mg/kg PFD组、MSCs治疗组及MSCs联合P30组。气管滴注博来霉素后第7天尾静脉注射5×105 hUC-MSCs,第7天起每日予PFD灌胃。造模后第21天收集肺组织, HE、Masson染色分别评估肺部形态变化及胶原沉积情况; Sircol法测胶原含量; PCR和Western blot检测纤维化标志物水平。超滤收集hUC-MSCs条件培养基(conditioned medium, CM),联合PFD体外培养肌成纤维细胞(myofibroblast, MFB), CCK-8与Brdu实验检测MFB生长及增殖。结果显示,与模型组相比, MSCs联合P30组显著改善小鼠生存率及肺部病理,显著降低胶原及纤维化标志物水平(P<0.001),且疗效优于单用PFD和hUC-MSCs组(P<0.05)。PFD部分抑制MFB增殖,联合CM增强了PFD对MFB增殖的抑制(P<0.001)。研究表明, hUC-MSCs联合低剂量PFD可能通过抑制MYB的增殖减轻博来霉素诱导的小鼠肺纤维化。

This study aims to investigate the therapeutic effect of human umbilical cord mesenchymal stem cells combined with pirfenidone in bleomycin-induced pulmonary fibrosis in mice and its possible mechanism. C57BL/6 mice were divided into normal control group, hUC-MSCs control group, model group, 30 mg/kg PFD group （P30）, 100 mg/kg PFD group, 300 mg/kg PFD group, hUC-MSCs treatment group and hUC-MSCs combined with P30 group. 5×10^5 hUC-MSCs were injected via the tail vein on the 7th day after intratracheal instillation of bleomycin and PFD was administered once a day orally from then on. Their lung tissues were harvested at the 2 lth day, pulmonary morphological changes and collagen deposition were assessed respectively by HE and Masson staining, the content of collagen was measured by Sircol assay and the levels of fibrosis related markers were detected by PCR and Western blot. In addition, myofibroblasts （MFB） were cultured with hUC-MSCs conditional medium （CM） collected by ultrafiltration and PFD in vitro, CCK-8 and Brdu assay were used to detect the growth and proliferation of MFB. The results showed that hUC-MSCs combined with P30 group significantly improved the survival rate and pulmonary histopathology of mice, reduced the levels of collagen and fibrosis related markers （P〈0.001）, and the efficacy of combination therapy was better than that of PFD alone and hUC-MSCs alone （P〈0.05）. PFD partially inhibited proliferation of MFB and PFD combined with CM enhanced the inhibitory effect of PFD on proliferation （P〈0.001）. This study demonstrate that hUC-MSCs combined with low-dose PFD may attenuate bleomycin-induced pulmonary fibrosis in mice by inhibiting MYB proliferation.