Microcystin is a widespread cyanobacterial toxin that affects the intestine to produce diarrheal symptoms after ingestion of freshwater blue-green algae. Our study aimed to characterize the mechanism by which the toxin leads to diarrhea via epithelial barrier dysfunction in a small intestine Caco-2 cell model. Microcystin-treated human Caco-2 epithelial monolayers were functionally and molecularly analyzed for barrier dysfunction. Tight junctions (TJs) and cell damage were analyzed in relation to transepithelial electrical resistance (TER) changes. TER of microcystin-treated Caco-2 cells was reduced by 65% of the initial value after 24 h; concomitantly, permeability for fluorescein increased 2.6-fold. Western blot analysis showed reduced claudin-1 expression, while expression of claudin-3 and -4 remained unchanged. Super-resolution stimulated emission depletion microscopy revealed that TJ integrity was compromised by fraying and splitting of the TJ domain of the epithelial cells. Epithelial apoptosis did not significantly contribute to epithelial barrier dysfunction, while cytoskeletal actomyosin constriction was associated with TJ disintegration and the barrier defect. Our results indicate that microcystin causes intestinal barrier leakiness, which helps to explain the leak flux type of diarrhea as the main pathomechanism after ingestion of cyanobacterial toxin.

中文翻译：

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解开蓝藻毒素微囊藻毒素处理的 Caco-2 单层细胞中的肠上皮屏障

微囊藻毒素是一种广泛存在的蓝藻毒素，摄入淡水蓝绿藻后会影响肠道产生腹泻症状。我们的研究旨在表征该毒素在小肠 Caco-2 细胞模型中通过上皮屏障功能障碍导致腹泻的机制。对经微囊藻毒素处理的人 Caco-2 上皮单层进行功能和分子分析，以了解屏障功能障碍。分析了紧密连接（TJ）和细胞损伤与跨上皮电阻（TER）变化的关系。微囊藻毒素处理的Caco-2细胞24小时后TER减少了初始值的65%；与此同时，荧光素的渗透性增加了 2.6 倍。Western blot分析显示claudin-1表达减少，而claudin-3和-4表达保持不变。超分辨率受激发射损耗显微镜显示，TJ 完整性因上皮细胞 TJ 结构域的磨损和分裂而受到损害。上皮细胞凋亡对上皮屏障功能障碍没有显着影响，而细胞骨架肌动球蛋白收缩与 TJ 崩解和屏障缺陷相关。我们的研究结果表明，微囊藻毒素会导致肠道屏障渗漏，这有助于解释摄入蓝藻毒素后漏流型腹泻的主要病理机制。