Testing Epithelial Permeability in Fetal Tissue-Derived Enteroids.
Testing Epithelial Permeability in Fetal Tissue-Derived Enteroids.
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DOI：
10.3791/64108
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发表时间：
2022-06-16
期刊：
影响因子：
--
通讯作者：
Ho TT
中科院分区：
文献类型：
--
作者：
Llerena A;Urmi S;Amin J;Cha B;Ho TT
人类胎儿组织来源的肠样蛋白正在成为研究早产儿肠道损伤的一种有前途的体外模型。肠类表现出极性，由具有顶端边界的管腔、紧密连接和暴露于生长介质的基底外侧外层组成。肠道损伤的后果包括粘膜炎症和通透性增加。在脆弱的早产人类受试者中测试肠道通透性通常是不可行的。因此，需要体外胎儿组织来源的肠道模型来研究早产儿的肠道损伤。肠类可用于测试紧密连接蛋白调节的上皮通透性的变化。在肠类中，肠干细胞分化为所有上皮细胞类型，并在小鼠肉瘤细胞分泌的基底膜基质上形成三维结构。在本文中，我们描述了用于从胎儿肠道组织建立肠样、用免疫荧光成像表征肠样紧密连接蛋白以及测试上皮通透性的方法。由于革兰氏阴性优势细菌生态失调是肠道损伤的已知危险因素，我们使用脂多糖（LPS）（一种由革兰氏阴性菌产生的内毒素）来诱导肠类的通透性。将荧光素标记的葡聚糖显微注射到肠腔中，并测量渗漏到培养基中的连续葡聚糖浓度，以量化细胞旁通透性的变化。实验表明，顶端暴露于 LPS 会以浓度依赖性方式诱导上皮细胞通透性。这些发现支持革兰氏阴性显性菌群失调导致早产儿肠道损伤机制的假设。
Human fetal tissue-derived enteroids are emerging as a promising in vitro model to study intestinal injuries in preterm infants. Enteroids exhibit polarity, consisting of a lumen with an apical border, tight junctions, and a basolateral outer layer exposed to growth media. The consequences of intestinal injuries include mucosal inflammation and increased permeability. Testing intestinal permeability in vulnerable preterm human subjects is often not feasible. Thus, an in vitro fetal tissue-derived intestinal model is needed to study intestinal injuries in preterm infants. Enteroids can be used to test changes in epithelial permeability regulated by tight junction proteins. In enteroids, intestinal stem cells differentiate into all epithelial cell types and form a three-dimensional structure on a basement membrane matrix secreted by mouse sarcoma cells. In this article, we describe the methods used for establishing enteroids from fetal intestinal tissue, characterizing the enteroid tight junction proteins with immunofluorescent imaging, and testing epithelial permeability. As gram-negative dominant bacterial dysbiosis is a known risk factor for intestinal injury, we used lipopolysaccharide (LPS), an endotoxin produced by gram-negative bacteria, to induce permeability in the enteroids. Fluorescein-labeled dextran was microinjected into the enteroid lumen, and serial dextran concentrations leaked into the culture media were measured to quantify the changes in paracellular permeability. The experiment showed that apical exposure to LPS induces epithelial permeability in a concentration-dependent manner. These findings support the hypothesis that gram-negative dominant dysbiosis contributes to the mechanism of intestinal injury in preterm infants.
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影响因子：
3.8
作者：
Walker RW;Clemente JC;Peter I;Loos RJF
通讯作者：
Loos RJF
影响因子：
9
作者：
Humberg A;Fortmann I;Siller B;Kopp MV;Herting E;Göpel W;Härtel C;German Neonatal Network, German Center for Lung Research and Priming Immunity at the beginning of life (PRIMAL) Consortium
通讯作者：
German Neonatal Network, German Center for Lung Research and Priming Immunity at the beginning of life (PRIMAL) Consortium
影响因子：
2.5
作者：
Dheer R;Young VB
通讯作者：
Young VB
影响因子：
--
作者：
Alganabi, Mashriq;Lee, Carol;Pierro, Agostino
通讯作者：
Pierro, Agostino
DOI：
10.1016/j.jcmgh.2018.04.008
发表时间：
2018
影响因子：
7.2
作者：
Tsai YH;Czerwinski M;Wu A;Dame MK;Attili D;Hill E;Colacino JA;Nowacki LM;Shroyer NF;Higgins PDR;Kao JY;Spence JR
通讯作者：
Spence JR