Zhu Liangying, Fu Xiao, Chen Xiang, Han Xiaodong, Dong Ping
Immunology and Reproductive Biology Laboratory, Medical School, Nanjing University, Hankou Road 22, Nanjing, 210093, China.
The Affiliated Drum Tower Hospital of Nanjing University Medical School, Nanjing, Jiangsu Province, China.
Cell Biol Int. 2017 Sep;41(9):960-968. doi: 10.1002/cbin.10788. Epub 2017 Jul 9.
IPF is characterized by fibroblast accumulation, collagen deposition, and ECM remodeling, with myofibroblasts believed to be the effector cell type. Myofibroblasts develop due to EMT of lung alveolar epithelial cells, which can be induced by TGF-β. M2 macrophages, a macrophage subpopulation, secrete large amounts of TGF-β. To clarify the relationship between IPF, EMT, TGF-β, and M2 macrophages, a bleomycin-induced pulmonary fibrosis mouse model was used. Seventeen days after mice were treated with bleomycin, the successful establishment of a pulmonary fibrosis model was confirmed by HE stain and Masson's trichrome stain. We found evidence in support of EMT, such as elevated protein levels of α-SMA in lung tissue and decreased levels of E-cadherin and CK-18. Additionally, increased TGF-β levels and TGF-β/Smad2 signaling activation was observed. Macrophages were recruited to pulmonary alveoli. Alveolar macrophages were phenotyped and identified as M2 macrophages, with up-regulated CD206 on the cell surfaces. For in vitro studies, we treated RAW 264.7 cells with IL-4 for 24 h, and the cells were then utilized as M2 macrophages. TGF-β levels increased significantly in the culture supernatant. Forty-eight hours after lung epithelial cells (MLE-12) were co-cultured with the M2 macrophages, the expression of α-SMA increased, and E-cadherin and CK-18 decreased. When a TGF-β receptor inhibitor, LY2109761 was used, the EMT induced by M2 macrophages was blocked. In conclusion, we demonstrated that M2 macrophages induce EMT through the TGF-β/Smad2 signaling pathway.
特发性肺纤维化（IPF）的特征是成纤维细胞积聚、胶原蛋白沉积和细胞外基质重塑，其中肌成纤维细胞被认为是效应细胞类型。肌成纤维细胞由肺泡上皮细胞的上皮-间质转化（EMT）产生，而转化生长因子-β（TGF-β）可诱导这种转化。M2巨噬细胞是巨噬细胞的一个亚群，可分泌大量TGF-β。为了阐明IPF、EMT、TGF-β和M2巨噬细胞之间的关系，我们使用了博来霉素诱导的肺纤维化小鼠模型。在用博来霉素处理小鼠17天后，通过苏木精-伊红（HE）染色和马松三色染色证实成功建立了肺纤维化模型。我们发现了支持EMT的证据，例如肺组织中α-平滑肌肌动蛋白（α-SMA）蛋白水平升高，而E-钙黏蛋白和细胞角蛋白18（CK-18）水平降低。此外，还观察到TGF-β水平升高以及TGF-β/ Smad2信号通路激活。巨噬细胞被募集到肺泡。对肺泡巨噬细胞进行表型分析并鉴定为M2巨噬细胞，其细胞表面的CD206上调。对于体外研究，我们用白细胞介素-4（IL-4）处理RAW 264.7细胞24小时，然后将这些细胞用作M2巨噬细胞。培养上清液中的TGF-β水平显著升高。肺上皮细胞（MLE-12）与M2巨噬细胞共培养48小时后，α-SMA的表达增加，而E-钙黏蛋白和CK-18减少。当使用TGF-β受体抑制剂LY2109761时，M2巨噬细胞诱导的EMT被阻断。总之，我们证明M2巨噬细胞通过TGF-β/ Smad2信号通路诱导EMT。
