Contreras Telma C, Ricciardi Elisabetta, Cremonini Eleonora, Oteiza Patricia I
Department of Nutrition, University of California, Davis, CA, USA; Department of Environmental Toxicology, University of California, Davis, CA, USA.
Department of Nutrition, University of California, Davis, CA, USA; Department of Environmental Toxicology, University of California, Davis, CA, USA.
Arch Biochem Biophys. 2015 May 1;573:84-91. doi: 10.1016/j.abb.2015.01.024. Epub 2015 Mar 17.
An increased permeability of the intestinal barrier is proposed as a major event in the pathophysiology of inflammatory bowel diseases (IBD). Tumor necrosis alpha (TNFα) plays a central role in IBD pathogenesis, in part promoting tight function (TJ) barrier dysfunction. Food extracts enriched in (-)-epicatechin (EC) prevent the development or improve the progression of IBD in animal models. This study investigated the capacity of EC to inhibit TNFα-induced permeabilization of Caco-2 cell monolayers, characterizing the underlying mechanisms. Caco-2 cells differentiated into intestinal epithelial cells were incubated in the absence/presence of TNFα, with or without the addition of 0.5-5 μM EC. TNFα triggered cell monolayer permeabilization, decreasing transepithelial electrical resistance (TEER) and increasing the paracellular transport of fluorescein sulfonic acid. The permeabilizing effects of TNFα were not due to Caco-2 cell apoptosis as evaluated by DNA fragmentation, caspase 3 and 9 activation, and cell morphology. EC prevented TNFα-triggered Caco-2 monolayer permeabilization and acted inhibiting the associated: (i) NADPH oxidase (NOX)-mediated increased oxidant production, (ii) NF-κB (IκBα phosphorylation, p50 and RelA nuclear transport, and nuclear NF-κB-DNA binding) and ERK1/2 activation, (iii) increased myosin light kinase expression, and decreased TJ protein ZO-1 levels. In summary, EC prevented TNFα-mediated Caco-2 cell barrier permeabilization in part through the inhibition of NOX/NF-κB activation and downstream TJ disruption. Diets rich in EC could contribute to ameliorate IBD-associated increased intestinal permeability.
肠屏障通透性增加被认为是炎症性肠病（IBD）病理生理学中的一个主要事件。肿瘤坏死因子α（TNFα）在IBD发病机制中起核心作用，部分促进紧密连接（TJ）屏障功能障碍。富含（-）-表儿茶素（EC）的食物提取物可预防动物模型中IBD的发生或改善其进展。本研究调查了EC抑制TNFα诱导的Caco-2细胞单层通透性的能力，并对其潜在机制进行了表征。将分化为肠上皮细胞的Caco-2细胞在有无TNFα的情况下孵育，添加或不添加0.5-5μM EC。TNFα触发细胞单层通透性增加，降低跨上皮电阻（TEER）并增加荧光素磺酸的细胞旁运输。通过DNA片段化、caspase 3和9激活以及细胞形态评估，TNFα的通透作用并非由于Caco-2细胞凋亡。EC可防止TNFα触发的Caco-2单层通透性增加，并抑制相关的：（i）NADPH氧化酶（NOX）介导的氧化剂产生增加，（ii）NF-κB（IκBα磷酸化、p50和RelA核转运以及核NF-κB-DNA结合）和ERK1/2激活，（iii）肌球蛋白轻链激酶表达增加以及TJ蛋白ZO-1水平降低。总之，EC部分通过抑制NOX/NF-κB激活和下游TJ破坏来防止TNFα介导的Caco-2细胞屏障通透性增加。富含EC的饮食可能有助于改善IBD相关的肠道通透性增加。
