| Background and aimsEpithelial cells of the small intestine migrate to the tip of the villus at which they are shed. However, under physiological conditions,97% of gaps were filled with non-permeability materials, maintaining the intestinal epithelial barrier function. But what exactly filled the gaps is unclear.The intestinal mucus is mainly secreted from goblet cells, with the main structural building block being the densely heavy O-glycosylated MUC2 mucin, which play a key role in protecting and lubricating the intestinal epithelium. This inner mucus layer is densely packed and acts as a physical barrier excluding microbes from the epithelial cells. Previous studies with ulcerative colitis (UC) patients showing bacteria penetrated into the inner mucus layer. The defects in mucus’s protective properties play a key role in the pathogenesis of UC.Confocal Laser Endomicroscopy (CLE) is an advanced endoscopic technique with the capability of a histological level of definition and magnification.CLE can clearly observe the subcellular structure, which is named as"optical biopsy". Previous studies indicate that gaps with the diameter of an individual epithelial cell can be observed in human ileal and rectal epithelium by CLE.The aims of this study were to investigate whether mucus would fill the epithelial gaps, and to assess whether gaps filled with mucus play an important role in maintaining local barrier function.Methods From June 2014 to December 2014, the terminal ileum tissues of UC patients and controls were obtained who visited the Qilu Hospital for CLE. The tissues were fixed with Carnoy’s solution overnight. In order to observe epithelial gaps, mucins were stained with Alcian Blue-Periodic Acid Schiff (AB-PAS) and Muc2. In addition, we also analyzed the CLE imagines of the terminal ileum from 39 UC patients and 34 controls, who visited the Qilu Hospital for CLE since January 2013 to December 2014. We also used the revised Watson grading system to assess the local intestinal barrier function. Watson Ⅱ is functional defect, which indicated single cell shedding with fluorescein leakage into the lumen. Watson Ⅲ is structural defect, which indicated multiple cells shedding with fluorescein leakage into the lumen. The percentage of defective perimeter of Watson Ⅱ/Ⅲ gaps were observed to evaluate the local intestinal barrier function. And the percentage of the number of goblet cells between the two groups were observed to assess whether the goblet cells were significantly reduced in patients with ulcerative colitis.ResultsMucus filled the intestinal epithelial gaps could be observed in both of the two groups under optical microscopy. The proportion of epithelial gaps without mucus in UC patients is higher than that of the control group (27.52 (25,29.68) vs 12.27 (9.78, 16.67), p<0.001). Goblet cells and the epithelial gaps could be clearly identified under CLE. Moreover, local intestinal barrier dysfunction was found in UC patients after analyzing the CLE images of terminal ileum (5.33 (3.48,6.62) vs 0.27 (0.00, 0.69), p<0.001); Comparing with control group, the proportion of goblet cells is much lower in UC patients (17.38 (16.00,19.87) vs 23.44 (20.24,26.18),p<0.001). The proportion of epithelial gaps (%) and that of goblet cells (%) were Spearman correlated, which showed a significantly negative correlation(r=-0.74,p<0.001).Conclusion1. This study firstly certified that mucus filled intestinal epithelial gaps were the direct physiological mechanism to maintain the epithelial barrier function.2. Under the observation of CLE, the proportion of epithelial gaps in UC patients was higher than the control group, while that of goblet cells was much lower, which showed a significantly negative correlation.3. The study also showed that the proportion of epithelial gaps without mucus in UC patients was higher than the control group, following with local intestinal barrier dysfunction. |
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