COLONIC CRYPT PERMEABILITY BARRIER IN CROHNS DISEASE

胃肠道上皮的屏障功能赋予了胃肠道高的过电选择性。几项研究表明，肠道对大分子的渗透性增加可能与克罗恩病的发病机制有关。胃肠道通透性的决定因素是复杂的，不完全了解。我们已经在兔结肠隐窝细胞的顶端边界建立了一个不寻常的氨（弱碱）渗透屏障。根据临床研究者奖K08 DKO2410，我们最近确定存在渗透性屏障以及弱酸CO2（一种气体，如NH3）和丁酸，所有这些都是结肠腔中发酵的丰富产物。这些分子中的每一个都具有中性亲脂性形式，可渗透大多数细胞膜，影响细胞内ph。事实上，使用最近开发的制剂，我们发现极化结肠表面细胞，不像它们位于隐窝的祖细胞，对NH3、丁酸和CO2具有高的根尖渗透性。结肠腺对气体/亲脂分子的屏障可能对胃肠道上皮的整体屏障功能很重要。因此，了解改变的顶端“微分子”屏障如何影响上皮细胞健康，以及改变的微分子屏障是否在与克罗恩病相关的“大分子”通透性增加之前和/或易感性是很重要的。由于结肠隐窝不寻常的屏障功能直到最近才被认识到，目前还没有关于其与人类疾病关系的数据。目前炎症性肠病的动物模型不能很好地代表最终导致人类克罗恩病的病理生理过程。幸运的是，我们可以在灌注的人隐窝中研究隐窝细胞内pH值。在最近的研究中，我们在人类隐窝中观察到一个对NH3扩散但不是对NH4+运输的根尖屏障。此外，我们已经能够在包含人类隐窝的细胞之间动力学表征酸碱运输。这些技术进步应用于乙状窦镜活检获得的隐窝，允许直接研究人类微分子屏障，并能够评估改变的微分子屏障在克罗恩病发病机制中的作用。虽然根尖酸碱屏障现象的基础生物学研究在膜生物学中有广泛的应用，但在这一应用中概述的研究主要集中在与克罗恩病有关的屏障生物学和结肠菌pH。

The barrier function of gastrointestinal epithelium confers high permselectivity to the GI tract. Several studies have demonstrated increased intestinal permeability to macromolecules that may be linked to the pathogenesis of Crohn's Disease. The determinants of gastrointestinal permeability are complex and incompletely understood. We have established the presence of an unusual barrier to the permeation of ammonia (a weak base) at the apical borders of rabbit colonic-crypt cells. Under Clinical Investigator Award K08 DKO2410, we recently determined that a permeability barrier exists as well to the weak acids CO2 (a gas, like NH3) and butyric acid, all of which are abundant products of fermentation in the colonic lumen. Each of these molecules possesses neutral lipophilic forms that permeate most cell membranes, affecting intracellular pH. Indeed, using a recently- developed preparation, we found that polarized colonic surface-cells, unlike their progenitors located in the crypt, possess high apical permeability to NH3, butyric acid, and CO2. The barrier to gases/lipophilic molecules in colonic glands is likely important to the overall barrier function of gastrointestinal epithelium. As such, it is important to know how altered apical "micromolecular" barriers affect epithelial cell health, and whether an altered micromolecular barrier precedes and/or predisposes to the increased "macromolecular" permeability associated with Crohn's Disease: As the unusual barrier function of colonic crypts has only recently been appreciated, no data as yet exist on its relationship to human disease. Present animal models of inflammatory bowel disease do not represent optimally the pathophysiologic processes that culminate in human Crohn's disease. Fortunately, we can study crypt cell intracellular pH in perfused human crypts. In recent work, we observed in human crypts an apical barrier to NH3 diffusion but not to NH4+ transport in all patients. In addition, we have been able to kinetically characterize acid-base transport among cells that comprise human crypts. These technical advances, applied to crypts obtained by sigmoidoscopic biopsy, permits direct study of micromolecular barriers in humans, and enables evaluation of the role of altered micromolecular barriers in the pathogenesis of Crohn's Disease. Although studies of the fundamental biology of apical acid-base barrier phenomenon have broad application to membrane biology, studies outlined in this application are focused long-term on the biology of the barrier and colonocyte pH as it relates to Crohn's Disease.