p38丝裂原活化蛋白激酶-肌球蛋白轻链激酶通路对肠上皮细胞机械屏障的调节作用以及改良白头翁煎剂治疗溃疡性结肠炎的机制。

W U Tingting, Yang Xin, Zhu Huiping, Guo Jinwei, Zhu Hui, Zhang Peipei, Wang Meng, Liang Guoqiang, Sun Hongwen
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引用次数: 0

摘要

目的研究改良白头翁煎剂(MPD)在体外和体内对溃疡性结肠炎(UC)肠上皮细胞机械屏障保护作用的机制:方法:我们利用脂多糖(LPS)建立了肠上皮隐窝细胞系-6细胞屏障损伤模型。然后分别用 p38 丝裂原活化蛋白激酶-肌球蛋白轻链激酶(p38MAPK-MLCK)通路抑制剂、p38MAPK-MLCK 通路沉默基因(si-p38MAPK、si-NF-κB 和 si-MLCK)和 MPD 处理该模型。进行跨上皮电子阻力(TEER)测量和渗透性试验以评估屏障功能。对紧密连接(TJ)进行免疫荧光染色。在体内实验中,进行了葡聚糖硫酸钠诱导的大鼠结肠炎模型,以评估 MPD 和美沙拉嗪对 UC 的影响。根据大鼠的临床症状,采用疾病活动指数对其进行评分。透射电子显微镜和苏木精-伊红染色用于观察 UC 大鼠的形态学变化。Western 印迹和实时定量聚合酶链反应用于检测重要差异分子的基因和蛋白表达:结果:在体外研究中,p38MAPK-MLCK 通路抑制剂和 p38MAPK-MLCK 通路基因沉默抑制了 LPS 诱导的肠屏障功能障碍。p38MAPK-MLCK通路基因沉默会降低TJ的表达。MPD 处理可部分恢复 LPS 诱导的 TEER 减少和渗透性增加。MPD 增加了 TJ 的基因和蛋白表达,同时下调了 LPS 诱导的 p-p38MAPK 和 p-MLC 的高表达。在 UC 模型大鼠中,MPD 可改善体重下降和疾病活动指数,缓解结肠病理变化,上调 TJ 的表达,并降低 UC 大鼠结肠粘膜组织中 p-p38MAPK 和 p-MLC 的表达:结论:p38MAPK-MLCK 信号通路可影响肠上皮细胞的机械屏障功能和 TJ 表达。MPD可通过抑制p38MAPK-MLCK通路恢复TJ表达,减轻肠上皮屏障损伤。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Regulatory effects of the p38 mitogen-activated protein kinase-myosin light chain kinase pathway on the intestinal epithelial mechanical barrier and the mechanism of modified Pulsatilla decoction in the treatment of ulcerative colitis.

Objective: To investigate the mechanism of the protective effect of modified Pulsatilla decoction (, MPD) on the mechanical barrier of the ulcerative colitis (UC) intestinal epithelium in vitro and in vivo.

Methods: We established an intestinal epithelial crypt cell line-6 cell barrier injury model by using lipopolysaccharide (LPS). The model was then treated with p38 mitogen-activated protein kinase-myosin light chain kinase (p38MAPK-MLCK) pathway inhibitors, p38MAPK-MLCK pathway silencing genes (si-p38MAPK, si-NF-κB, and si-MLCK), and MPD respectively. Transepithelial electronic resistance (TEER) measurements and permeability assays were performed to assess barrier function. Immunofluorescence staining of tight junctions (TJ) was performed. In in vivo experiment, dextran sodium sulfate-induced colitis rat model was conducted to evaluate the effect of MPD and mesalazine on UC. The rats were scored using the disease activity index based on their clinical symptoms. Transmission electron microscopy and hematoxylin-eosin staining were used to examine morphological changes in UC rats. Western blotting and real-time quantitative polymerase chain reaction were performed to examine the gene and protein expression of significant differential molecules.

Results: In in vitro study, LPS-induced intestinal barrier dysfunction was inhibited by p38MAPK-MLCK pathway inhibitors and p38MAPK-MLCK pathway gene silencing. Silencing of p38MAPK-MLCK pathway genes decreased TJ expression. MPD treatment partly restored the LPS-induced decreased in TEER and increase in permeability. MPD increased the gene and protein expression of TJ, while down-regulated the LPS-induced high expression of p-p38MAPK and p-MLC. In UC model rats, MPD could ameliorate body weight loss and disease activity index, relieve colonic pathology, up-regulate TJ expression as well as decrease the expression of p-p38MAPK and p-MLC in UC rat colonic mucosal tissue.

Conclusions: The p38MAPK-MLCK signaling pathway can affect mechanical barrier function and TJ expression in the intestinal epithelium. MPD restores TJ expression and attenuates intestinal epithelial barrier damage by suppressing the p38MAPK-MLCK pathway.

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