Sanders S E, Madara J L, McGuirk D K, Gelman D S, Colgan S P
Department of Anesthesia, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Epithelial Cell Biol. 1995;4(1):25-34.
Cultured intestinal epithelial monolayers serve as models for mechanistic studies of intestinal inflammation. One crucial aspect of epithelial function modulated by inflammation is permeability. Indices of permeability typically obtained are transepithelial resistance or more formalized assays using Ussing chambers modified for cultured monolayers. Such Ussing chamber systems are inconvenient for screening assays and their geometry precludes evaluation of important inflammatory responses such as transepithelial migration of neutrophils. Here we use a novel flux assay to investigate the kinetics of fluorescein-labelled dextran (FD) molecules across intestinal epithelial cell lines grown on permeable membrane supports. Our results show that we could consistently demonstrate picomolar flux of FD in a range of sizes (3-70 kD) across T84 epithelial monolayers. Such FD flux was time- and dose-dependent and flux increased exponentially with decreasing transepithelial resistance. Exposure of intestinal epithelia to mucosal-derived lymphocytes or to soluble lymphocyte products (interferon-gamma, interleukin-4 or interleukin-13) increased FD flux in a dose-dependent fashion. Finally, studies of neutrophil transepithelial migration revealed qualitative and quantitative differences in FD flux depending on FD size. We conclude that in vitro transepithelial FD flux may be a useful tool to study aspects of intestinal permeability in health and in disease.
培养的肠上皮单层细胞可作为肠道炎症机制研究的模型。炎症调节上皮功能的一个关键方面是通透性。通常获得的通透性指标是跨上皮电阻，或使用为培养的单层细胞改良的尤斯灌流小室进行更规范的检测。这种尤斯灌流小室系统对于筛选检测不方便，而且其几何形状妨碍了对重要炎症反应的评估，如中性粒细胞的跨上皮迁移。在这里，我们使用一种新型通量检测方法来研究荧光素标记的葡聚糖（FD）分子穿过生长在可渗透膜支持物上的肠上皮细胞系的动力学。我们的结果表明，我们能够始终如一地证明一系列大小（3 - 70 kD）的FD在T84上皮单层细胞中的皮摩尔通量。这种FD通量是时间和剂量依赖性的，并且通量随着跨上皮电阻的降低呈指数增加。将肠上皮暴露于黏膜来源的淋巴细胞或可溶性淋巴细胞产物（干扰素 - γ、白细胞介素 - 4或白细胞介素 - 13）会以剂量依赖性方式增加FD通量。最后，对中性粒细胞跨上皮迁移的研究揭示了根据FD大小，FD通量在定性和定量上的差异。我们得出结论，体外跨上皮FD通量可能是研究健康和疾病状态下肠道通透性方面的有用工具。
