Pirfenidone Inhibits Cell Proliferation and Collagen I Production of Primary Human Intestinal Fibroblasts
Pirfenidone Inhibits Cell Proliferation and Collagen I Production of Primary Human Intestinal Fibroblasts
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DOI：
10.3390/cells9030775
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发表时间：
2020-03-01
期刊：
影响因子：
6
通讯作者：
Faber, Klaas Nico
中科院分区：
文献类型：
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作者：
Cui, Yingying;Zhang, Mengfan;Faber, Klaas Nico
肠纤维化是炎症性肠病的常见并发症。到目前为止，还没有安全有效的治疗肠纤维化的药物。吡非尼酮是一种可用于治疗特发性肺纤维化的抗纤维化化合物。在这里，我们探索了吡非尼酮对原代人肠成纤维细胞（p-hIF）的抗增殖和抗纤维化特性。在不存在和存在吡非尼酮的情况下培养p-hIF。通过实时细胞分析仪（xCELLigence）和BrdU掺入测量细胞增殖。通过活细胞成像监测细胞运动性。通过Sytox绿色、胱天蛋白酶-3和水溶性四唑盐-1（WST-1）测定分析细胞毒性和细胞活力。通过定量逆转录PCR（RT-qPCR）测定纤维化标志物的基因表达。通过Western印迹和I型胶原蛋白表达另外通过免疫荧光显微镜分析哺乳动物雷帕霉素靶蛋白（mTOR）信号传导。吡非尼酮剂量依赖性地抑制p-HIF增殖和运动，而不诱导细胞死亡。吡非尼酮抑制有助于细胞外基质产生的基因的mRNA水平，以及基础和TGF-β 1诱导的I型胶原蛋白产生，这与抑制p-hIFs中雷帕霉素敏感性mTOR/p70 S6 K通路有关。因此，吡非尼酮通过TGF-β 1/mTOR/p70 S6 K信号通路抑制肠成纤维细胞增殖并抑制胶原蛋白I的产生，这可能是治疗肠纤维化的一种新的安全的抗纤维化策略。
Intestinal fibrosis is a common complication of inflammatory bowel disease. So far, there is no safe and effective drug for intestinal fibrosis. Pirfenidone is an anti-fibrotic compound available for the treatment of idiopathic pulmonary fibrosis. Here, we explored the anti-proliferative and anti-fibrotic properties of pirfenidone on primary human intestinal fibroblasts (p-hIFs). p-hIFs were cultured in the absence and presence of pirfenidone. Cell proliferation was measured by a real-time cell analyzer (xCELLigence) and BrdU incorporation. Cell motility was monitored by live cell imaging. Cytotoxicity and cell viability were analyzed by Sytox green, Caspase-3 and Water Soluble Tetrazolium Salt-1 (WST-1) assays. Gene expression of fibrosis markers was determined by quantitative reverse transcription PCR (RT-qPCR). The mammalian target of rapamycin (mTOR) signaling was analyzed by Western blotting and type I collagen protein expression additionally by immunofluorescence microscopy. Pirfenidone dose-dependently inhibited p-hIF proliferation and motility, without inducing cell death. Pirfenidone suppressed mRNA levels of genes that contribute to extracellular matrix production, as well as basal and TGF-beta 1-induced collagen I protein production, which was associated with inhibition of the rapamycin-sensitive mTOR/p70S6K pathway in p-hIFs. Thus, pirfenidone inhibits the proliferation of intestinal fibroblasts and suppresses collagen I production through the TGF-beta 1/mTOR/p70S6K signaling pathway, which might be a novel and safe anti-fibrotic strategy to treat intestinal fibrosis.