Department of Internal Medicine, College of Medicine, Brain Korea 21, Severance Biomedical Science Institute, Yonsei University, Seoul, Korea.
Lab Invest. 2012 Dec;92(12):1698-711. doi: 10.1038/labinvest.2012.132. Epub 2012 Sep 24.
Epithelial-mesenchymal transition (EMT) of peritoneal mesothelial cells (PMCs) has a role in the process of peritoneal fibrosis (PF), a serious complication in peritoneal dialysis (PD) patients. Even though monocyte chemoattractant protein-1 (MCP-1) was demonstrated to directly increase extracellular matrix (ECM) synthesis, the role of the MCP-1/CCR2 system in PD-related EMT and ECM synthesis in cultured human PMCs (HPMCs) and in an animal model of PD has never been elucidated. In vitro, HPMCs were exposed to 5.6 mM glucose (NG), NG+MCP-1 (10 ng/ml) (NG+MCP-1), or 100 mM glucose (HG) with or without CCR2 inhibitor (RS102895) (CCR2i) or a dominant-negative mutant MCP-1-expressing lentivirus (LV-mMCP-1). In vivo, PD catheters were inserted into 60 Sprague-Dawley rats, and saline (Control, C) (N=30) or 4.25% PD solution (PD) (N=30) was infused for 4 weeks. Twenty rats from each group were treated with empty LV or LV-mMCP-1 intraperitoneally. Snail, E-cadherin, α-smooth muscle actin (α-SMA), and fibronectin protein expression in HPMCs and the peritoneum was evaluated by western blot analysis. Compared with NG cells, Snail, α-SMA, and fibronectin expression was significantly increased, while E-cadherin expression was significantly decreased in HPMCs exposed to HG and NG+MCP-1, and these changes were significantly abrogated by CCR2i (P<0.05). In addition, MCP-1-induced EMT was significantly attenuated by anti-TGF-β1 antibody. In PD rats, Snail and fibronectin expression was significantly increased in the peritoneum, whereas the ratios of E-cadherin/α-SMA protein expression were significantly decreased (P<0.05). The thickness of the peritoneum and the intensity of Masson's trichrome staining in the peritoneum were also significantly higher in PD rats than in C rats (P<0.05). These changes in PD rats were significantly abrogated by LV-mMCP-1. These findings suggest that the MCP-1/CCR2 system is directly involved in PD-related EMT and ECM synthesis and that this is mediated, at least in part, via TGF-β1.
上皮-间充质转化（EMT）在腹膜间皮细胞（PMCs）中发挥作用，导致腹膜纤维化（PF），这是腹膜透析（PD）患者的严重并发症。尽管单核细胞趋化蛋白-1（MCP-1）被证明可以直接增加细胞外基质（ECM）的合成，但 MCP-1/CCR2 系统在 PD 相关 EMT 和 ECM 合成中的作用，以及在 PD 相关的动物模型中的作用，尚未得到阐明。在体外，将人 PMCs（HPMCs）暴露于 5.6mM 葡萄糖（NG）、NG+MCP-1（10ng/ml）（NG+MCP-1）或 100mM 葡萄糖（HG）中，同时或不使用 CCR2 抑制剂（RS102895）（CCR2i）或表达显性负突变 MCP-1 的慢病毒（LV-mMCP-1）。在体内，将 PD 导管插入 60 只 Sprague-Dawley 大鼠中，并用生理盐水（对照，C）（N=30）或 4.25% PD 溶液（PD）（N=30）输注 4 周。每组 20 只大鼠分别用空 LV 或 LV-mMCP-1 进行腹腔内注射。通过 Western blot 分析评估 HPMCs 和腹膜中 Snaill、E-钙黏蛋白、α-平滑肌肌动蛋白（α-SMA）和纤维连接蛋白的蛋白表达。与 NG 细胞相比，HG 和 NG+MCP-1 处理的 HPMCs 中 Snaill、α-SMA 和纤维连接蛋白的表达显著增加，而 E-钙黏蛋白的表达显著降低，而 CCR2i 可显著阻断这些变化（P<0.05）。此外，抗 TGF-β1 抗体可显著减弱 MCP-1 诱导的 EMT。在 PD 大鼠中，Snail 和纤维连接蛋白的表达在腹膜中显著增加，而 E-钙黏蛋白/α-SMA 蛋白表达的比值显著降低（P<0.05）。PD 大鼠的腹膜厚度和腹膜内 Masson 三色染色强度也明显高于 C 大鼠（P<0.05）。LV-mMCP-1 可显著阻断 PD 大鼠的这些变化。这些发现表明，MCP-1/CCR2 系统直接参与 PD 相关 EMT 和 ECM 合成，至少部分通过 TGF-β1 介导。
