Drago Sandro, El Asmar Ramzi, Di Pierro Mariarosaria, Grazia Clemente Maria, Tripathi Amit, Sapone Anna, Thakar Manjusha, Iacono Giuseppe, Carroccio Antonio, D'Agate Cinzia, Not Tarcisio, Zampini Lucia, Catassi Carlo, Fasano Alessio
Mucosal Biology Research Center, Center for Celiac Research and Division of Pediatric Gastroenterology and Nutrition, University of Maryland, School of Medicine, Baltimore, MD 21201, USA.
Scand J Gastroenterol. 2006 Apr;41(4):408-19. doi: 10.1080/00365520500235334.
Little is known about the interaction of gliadin with intestinal epithelial cells and the mechanism(s) through which gliadin crosses the intestinal epithelial barrier. We investigated whether gliadin has any immediate effect on zonulin release and signaling.
Both ex vivo human small intestines and intestinal cell monolayers were exposed to gliadin, and zonulin release and changes in paracellular permeability were monitored in the presence and absence of zonulin antagonism. Zonulin binding, cytoskeletal rearrangement, and zonula occludens-1 (ZO-1) redistribution were evaluated by immunofluorescence microscopy. Tight junction occludin and ZO-1 gene expression was evaluated by real-time polymerase chain reaction (PCR).
When exposed to gliadin, zonulin receptor-positive IEC6 and Caco2 cells released zonulin in the cell medium with subsequent zonulin binding to the cell surface, rearrangement of the cell cytoskeleton, loss of occludin-ZO1 protein-protein interaction, and increased monolayer permeability. Pretreatment with the zonulin antagonist FZI/0 blocked these changes without affecting zonulin release. When exposed to luminal gliadin, intestinal biopsies from celiac patients in remission expressed a sustained luminal zonulin release and increase in intestinal permeability that was blocked by FZI/0 pretreatment. Conversely, biopsies from non-celiac patients demonstrated a limited, transient zonulin release which was paralleled by an increase in intestinal permeability that never reached the level of permeability seen in celiac disease (CD) tissues. Chronic gliadin exposure caused down-regulation of both ZO-1 and occludin gene expression.
Based on our results, we concluded that gliadin activates zonulin signaling irrespective of the genetic expression of autoimmunity, leading to increased intestinal permeability to macromolecules.
关于麦醇溶蛋白与肠上皮细胞的相互作用以及麦醇溶蛋白穿过肠上皮屏障的机制，目前所知甚少。我们研究了麦醇溶蛋白是否对zonulin释放和信号传导有直接影响。
将离体的人小肠和肠细胞单层暴露于麦醇溶蛋白，并在存在和不存在zonulin拮抗剂的情况下监测zonulin释放和细胞旁通透性的变化。通过免疫荧光显微镜评估zonulin结合、细胞骨架重排和紧密连接蛋白-1（ZO-1）的重新分布。通过实时聚合酶链反应（PCR）评估紧密连接蛋白occludin和ZO-1的基因表达。
当暴露于麦醇溶蛋白时，zonulin受体阳性的IEC6和Caco2细胞在细胞培养基中释放zonulin，随后zonulin与细胞表面结合，细胞骨架重排，occludin-ZO1蛋白-蛋白相互作用丧失，单层通透性增加。用zonulin拮抗剂FZI/0预处理可阻断这些变化，而不影响zonulin释放。当暴露于肠腔麦醇溶蛋白时，缓解期乳糜泻患者的肠道活检标本显示肠腔zonulin持续释放且肠道通透性增加，FZI/0预处理可阻断这种增加。相反，非乳糜泻患者的活检标本显示zonulin释放有限且短暂，同时肠道通透性增加，但从未达到乳糜泻（CD）组织中的通透性水平。长期暴露于麦醇溶蛋白会导致ZO-1和occludin基因表达下调。
基于我们的结果，我们得出结论，无论自身免疫的基因表达如何，麦醇溶蛋白均可激活zonulin信号传导，导致肠道对大分子的通透性增加。
