Department of Pathology, The University of Chicago, Chicago, Illinois, USA.
Gastroenterology. 2011 Apr;140(4):1208-1218.e1-2. doi: 10.1053/j.gastro.2011.01.004. Epub 2011 Jan 13.
BACKGROUND & AIMS: Tumor necrosis factor (TNF) increases intestinal epithelial cell shedding and apoptosis, potentially challenging the barrier between the gastrointestinal lumen and internal tissues. We investigated the mechanism of tight junction remodeling and barrier maintenance as well as the roles of cytoskeletal regulatory molecules during TNF-induced shedding.
We studied wild-type and transgenic mice that express the fluorescent-tagged proteins enhanced green fluorescent protein-occludin or monomeric red fluorescent protein 1-ZO-1. After injection of high doses of TNF (7.5 μg intraperitoneally), laparotomies were performed and segments of small intestine were opened to visualize the mucosa by video confocal microscopy. Pharmacologic inhibitors and knockout mice were used to determine the roles of caspase activation, actomyosin, and microtubule remodeling and membrane trafficking in epithelial shedding.
Changes detected included redistribution of the tight junction proteins ZO-1 and occludin to lateral membranes of shedding cells. These proteins ultimately formed a funnel around the shedding cell that defined the site of barrier preservation. Claudins, E-cadherin, F-actin, myosin II, Rho-associated kinase (ROCK), and myosin light chain kinase (MLCK) were also recruited to lateral membranes. Caspase activity, myosin motor activity, and microtubules were required to initiate shedding, whereas completion of the process required microfilament remodeling and ROCK, MLCK, and dynamin II activities.
Maintenance of the epithelial barrier during TNF-induced cell shedding is a complex process that involves integration of microtubules, microfilaments, and membrane traffic to remove apoptotic cells. This process is accompanied by redistribution of apical junctional complex proteins to form intercellular barriers between lateral membranes and maintain mucosal function.
肿瘤坏死因子（TNF）可增加肠上皮细胞脱落和凋亡，从而可能破坏胃肠道腔与内部组织之间的屏障。我们研究了紧密连接重塑和屏障维持的机制，以及 TNF 诱导脱落过程中细胞骨架调节分子的作用。
我们研究了表达荧光标记蛋白增强型绿色荧光蛋白-occludin 或单体红色荧光蛋白 1-ZO-1 的野生型和转基因小鼠。经腹腔注射大剂量 TNF（7.5μg）后，进行剖腹术，并切开小肠段，通过视频共聚焦显微镜观察黏膜。使用药理学抑制剂和基因敲除小鼠来确定半胱天冬酶激活、肌动球蛋白、微管重塑和膜转运在上皮细胞脱落中的作用。
检测到的变化包括紧密连接蛋白 ZO-1 和 occludin 向脱落细胞的侧膜重新分布。这些蛋白最终在脱落细胞周围形成一个漏斗，定义了屏障保留的部位。Claudins、E-cadherin、F-actin、肌球蛋白 II、Rho 相关激酶（ROCK）和肌球蛋白轻链激酶（MLCK）也被募集到侧膜。半胱天冬酶活性、肌球蛋白马达活性和微管对于起始脱落是必需的，而完成这个过程需要微丝重排以及 ROCK、MLCK 和 dynamin II 的活性。
在 TNF 诱导的细胞脱落过程中，上皮屏障的维持是一个复杂的过程，涉及微管、微丝和膜转运的整合，以去除凋亡细胞。这个过程伴随着顶端连接复合体蛋白的重新分布，在侧膜之间形成细胞间屏障，维持黏膜功能。
