Jesse Brown Veterans Affairs Medical Center, Chicago, Illinois; Division of Gastroenterology and Hepatology, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois.
Division of Gastroenterology and Hepatology, Department of Medicine, University of Illinois at Chicago, Chicago, Illinois.
Gastroenterology. 2021 Mar;160(4):1240-1255.e3. doi: 10.1053/j.gastro.2020.11.008. Epub 2020 Nov 13.
BACKGROUND & AIMS: The down-regulated in adenoma (DRA) protein, encoded by SLC26A3, a key intestinal chloride anion exchanger, has recently been identified as a novel susceptibility gene for inflammatory bowel disease (IBD). However, the mechanisms underlying the increased susceptibility to inflammation induced by the loss of DRA remain elusive. Compromised barrier is a key event in IBD pathogenesis. The current studies were undertaken to elucidate the impact of DRA deficiency on epithelial barrier integrity and to define underlying mechanisms.
Wild-type and DRA-knockout (KO) mice and crypt-derived colonoids were used as models for intestinal epithelial response. Paracellular permeability was measured by using fluorescein isothiocyanate-dextran flux. Immunoblotting, immunofluorescence, immunohistochemistry, and ribonucleoprotein immunoprecipitation assays were performed. Gut microbiome analysis was conducted to investigate the impact of DRA deficiency on gut microbial communities.
DRA-KO mice exhibited an increased colonic paracellular permeability with significantly decreased levels of tight junction/adherens junction proteins, including ZO-1, occludin, and E-cadherin. A similar expression pattern of occludin and E-cadherin was observed in colonoids derived from DRA-KO mice and short hairpin RNA-mediated DRA knockdown in Caco-2 cells. Microbial analysis showed gut dysbiosis in DRA-KO mice. However, cohousing studies showed that dysbiosis played only a partial role in maintaining tight junction protein expression. Furthermore, our results showed increased binding of RNA-binding protein CUGBP1 with occludin and E-cadherin genes in DRA-KO mouse colon, suggesting that posttranscriptional mechanisms play a key role in gut barrier dysfunction.
To our knowledge, our studies demonstrate a novel role of DRA in maintaining the intestinal epithelial barrier function and potential implications of its dysregulation in IBD pathogenesis.
SLC26A3 编码的下调腺瘤蛋白（DRA）是一种关键的肠道氯离子阴离子交换体，最近被鉴定为炎症性肠病（IBD）的新易感性基因。然而，DRA 缺失导致炎症易感性增加的机制仍不清楚。受损的屏障是 IBD 发病机制中的一个关键事件。目前的研究旨在阐明 DRA 缺乏对肠上皮屏障完整性的影响，并确定潜在的机制。
野生型和 DRA 敲除（KO）小鼠和隐窝衍生的结肠类器官被用作肠上皮反应的模型。通过荧光素异硫氰酸酯-葡聚糖通量测量细胞旁通透性。进行免疫印迹、免疫荧光、免疫组织化学和核糖核蛋白免疫沉淀测定。进行肠道微生物组分析以研究 DRA 缺乏对肠道微生物群落的影响。
DRA-KO 小鼠表现出结肠细胞旁通透性增加，紧密连接/黏附连接蛋白，包括 ZO-1、occludin 和 E-cadherin 的水平显著降低。DRA-KO 小鼠来源的结肠类器官和 Caco-2 细胞中短发夹 RNA 介导的 DRA 敲低中观察到 occludin 和 E-cadherin 的相似表达模式。微生物分析显示 DRA-KO 小鼠肠道菌群失调。然而，共栖研究表明，菌群失调仅在维持紧密连接蛋白表达方面起部分作用。此外，我们的结果表明，在 DRA-KO 小鼠结肠中，RNA 结合蛋白 CUGBP1 与 occludin 和 E-cadherin 基因的结合增加，提示转录后机制在肠道屏障功能障碍中起关键作用。
据我们所知，我们的研究表明 DRA 在维持肠上皮屏障功能方面发挥了新的作用，并提示其在 IBD 发病机制中的失调具有潜在的意义。
