目的:克罗恩病(CD)是胃肠道的慢性炎症性疾病。纤维化,CD的严重并发症,当活化的肠成纤维细胞在受影响的区域沉积过量的细胞外基质(ECM)时发生。ECM的主要成分是高分子量透明质酸(HA),当解聚成低分子量片段时,变得促炎和促纤维化。HA降解的机制尚未完全了解,但是最近发现的新型蛋白质KIAA1199可以降解HA。我们假设KIAA1199蛋白在CD结肠成纤维细胞中增加,并产生促进炎症和纤维化的HA片段。
方法:从CD和非炎症性肠病对照(ND)患者手术切除的结肠组织的外植体中分离成纤维细胞。通过免疫印迹和免疫染色评估KIAA1199的蛋白水平和组织分布。并对功能性HA降解进行生化测定。
结果:与对照相比,培养的CD成纤维细胞产生并沉积在ECM中的KIAA1199蛋白水平升高。用促炎细胞因子白细胞介素(IL)6处理成纤维细胞增加了KIAA1199在ECM中的沉积。与对照组相比,CD成纤维细胞也产生显著更高水平的IL6,CD结肠成纤维细胞中IL6受体的抗体阻断降低了ECM中KIAA1199蛋白的水平。结肠成纤维细胞降解HA,然而,KIAA1199的小干扰RNA沉默消除了该能力。
结论:CD成纤维细胞主要通过IL6驱动的自分泌机制产生升高水平的KIAA1199。这导致HA的过度降解和促炎HA片段的产生,这有助于维持肠道炎症和纤维化。
OBJECTIVE: Crohn\'s Disease (CD) is a chronic inflammatory disease of the gastrointestinal tract. Fibrosis, a serious complication of CD, occurs when activated intestinal fibroblasts deposit excessive amounts of extracellular matrix (ECM) in affected areas. A major component of the ECM is high-molecular-weight hyaluronan (HA) that, when depolymerized to low-molecular-weight fragments, becomes proinflammatory and profibrotic. Mechanisms for HA degradation are incompletely understood, but the novel protein KIAA1199 recently was discovered to degrade HA. We hypothesized that KIAA1199 protein is increased in CD colon fibroblasts and generates HA fragments that foster inflammation and fibrosis.
METHODS: Fibroblasts were isolated from explants of surgically resected colon tissue from CD and non-inflammatory bowel disease control (ND) patients. Protein levels and tissue distribution of KIAA1199 were assessed by immunoblot and immunostaining, and functional HA degradation was measured biochemically.
RESULTS: Increased levels of KIAA1199 protein were produced and deposited in the ECM by cultured CD fibroblasts compared with controls. Treatment of fibroblasts with the proinflammatory cytokine interleukin (IL) 6 increased deposition of KIAA1199 in the ECM. CD fibroblasts also produce significantly higher levels of IL6 compared with controls, and antibody blockade of IL6 receptors in CD colon fibroblasts decreased the level of KIAA1199 protein in the ECM. Colon fibroblasts degrade HA, however, small interfering RNA silencing of KIAA1199 abrogated that ability.
CONCLUSIONS: CD fibroblasts produce increased levels of KIAA1199 primarily through an IL6-driven autocrine mechanism. This leads to excessive degradation of HA and the generation of proinflammatory HA fragments, which contributes to maintenance of gut inflammation and fibrosis.