Department of Gastroenterology, Infectious Diseases and Rheumatology, Clinical Physiology/Nutritional Medicine, Charité - Universitätsmedizin Berlin, Berlin, Germany.
Laboratory of Mucosal Barrier Pathobiology, Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts, USA.
Ann N Y Acad Sci. 2022 Oct;1516(1):188-196. doi: 10.1111/nyas.14870. Epub 2022 Jul 26.
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.
微囊藻毒素是一种广泛存在的蓝藻细菌毒素，摄入淡水蓝绿藻后会影响肠道，导致腹泻症状。我们的研究旨在通过小肠 Caco-2 细胞模型中上皮屏障功能障碍来描述该毒素导致腹泻的机制。对微囊藻毒素处理的人 Caco-2 上皮单层进行功能和分子分析，以研究屏障功能障碍。紧密连接 (TJ) 和细胞损伤与跨上皮电阻 (TER) 变化相关进行分析。微囊藻毒素处理的 Caco-2 细胞的 TER 在 24 小时后降低到初始值的 65%；同时，荧光素的通透性增加了 2.6 倍。Western blot 分析显示 Claudin-1 表达减少，而 Claudin-3 和 Claudin-4 的表达保持不变。超分辨率受激发射损耗显微镜显示 TJ 完整性受损，上皮细胞的 TJ 域出现磨损和分裂。上皮细胞凋亡对上皮屏障功能障碍没有显著贡献，而细胞骨架肌动球蛋白收缩与 TJ 解体和屏障缺陷有关。我们的结果表明，微囊藻毒素导致肠道屏障通透性增加，这有助于解释摄入蓝藻细菌毒素后腹泻的主要发病机制为渗漏通量型。
