The widespread use of chlorothalonil (CTL) has caused environmental residues and food contamination. Although the intestinal epithelial barrier (IEB) is directly involved in the metabolism and transportation of various exogenous compounds, there are few studies on the toxic effects of these compounds on the structure and function of IEB. The disassembly of tight junction (TJ) is a major cause of intestinal barrier dysfunction under exogenous compounds intake, but the precise mechanisms are not well understood. Here, we used Caco-2 cell monolayers as an in vitro model of human IEB to evaluate the toxicity of CTL exposure on the structure and function of IEB. Results showed that CTL exposure increased the paracellular permeability of the monolayers and downregulated mRNA levels of the TJ genes (ZO-1, OCLN, and CLDN1), polarity marker gene (SI), and anti-apoptosis gene (BCL-2) but upregulated the mRNA levels of apoptosis-related genes, including BAD, BAX, CASP3, and CASP8. Western blot analysis and immunofluorescence assay results showed the decreased levels and disrupted distribution of TJ protein network, including ZO-1 and CLDN1 in CTL-exposed IEB. In addition, the accumulation of intracellular reactive oxygen species, decreased mitochondrial membrane potential, and increased active CASP3 expression were observed in treated IEB. The result of TUNEL assay further confirmed the occurrence of cell apoptosis after CTL exposure. In addition, the phosphorylation of mitogen-activated protein kinases, including ERK, JNK and p38, was increased in CTL-exposed IEB. In summary, our results demonstrated that CTL exposure induced IEB dysfunction in Caco-2 cell monolayers by activating the mitogen-activated protein kinase pathway.

中文翻译：

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在基于 Caco-2 细胞的体外单层模型中，百菌清通过激活 MAPK 通路诱导肠上皮屏障功能障碍。

百菌清 (CTL) 的广泛使用造成了环境残留和食品污染。虽然肠上皮屏障（IEB）直接参与各种外源性化合物的代谢和运输，但关于这些化合物对IEB结构和功能的毒性作用的研究很少。紧密连接（TJ）的分解是外源性化合物摄入下肠道屏障功能障碍的主要原因，但其确切机制尚不清楚。在这里，我们使用 Caco-2 细胞单层作为人 IEB 的体外模型来评估 CTL 暴露对 IEB 结构和功能的毒性。结果表明，CTL 暴露增加了单层的细胞旁通透性，并下调了 TJ 基因（ZO-1、OCLN 和 CLDN1）、极性标记基因（SI）、和抗凋亡基因 (BCL-2)，但上调凋亡相关基因的 mRNA 水平，包括 BAD、BAX、CASP3 和 CASP8。蛋白质印迹分析和免疫荧光测定结果显示 TJ 蛋白网络的水平降低和分布中断，包括 CTL 暴露的 IEB 中的 ZO-1 和 CLDN1。此外，在处理过的 IEB 中观察到细胞内活性氧物质的积累、线粒体膜电位降低和活性 CASP3 表达增加。TUNEL检测结果进一步证实了CTL暴露后细胞凋亡的发生。此外，在 CTL 暴露的 IEB 中，丝裂原活化蛋白激酶（包括 ERK、JNK 和 p38）的磷酸化增加。总之，