C. C. Courtney1, K. M. Seiler1, E. J. Onufer1, K. G. McDonald2, R. D. Newberry2, B. W. Warner1 1Washington University,Division Of Pediatric Surgery,St. Louis, MO, USA 2Washington University,Department Of Internal Medicine,St. Louis, MO, USA
Introduction: The intestinal epithelial barrier selectively permits the absorption of nutrients while providing a defense against antigens and intraluminal toxins. It achieves this barrier function through both structural complexes and specialized delivery systems. Our lab has discovered injury to the liver associated with massive small bowel resection (SBR) independent of TPN, with associated translocation of bacteria to the liver, spleen, and lymph nodes. This suggests that there are alterations in the permeability of the intestine following SBR. The purpose of this study was to determine the most plausible mechanism by which the intestinal barrier is altered by looking at both paracellular and transcellular pathways within the intestine.
Methods: Male B6 mice underwent 50% proximal SBR or sham operation (transection with reanastomosis alone). Paracellular permeability was measured by fluorescein isothiocyanate-conguated (FITC)-dextran absorption and tight junction protein networks. FITC-dextran was injected into the distal bowel and serum uptake was quantified using a fluorimeter. Intestinal tissue was preserved in a glutaraldehyde solution and embedded for electron microscopy (EM). Single cell RNA sequencing and mass spectrometry were used to evaluate occludin and claudin-3/4 expression. The number of Goblet cell associated passages (GAPs) were quantified using intravital two-photon (2P) imaging as these structures facilitate the transcellular delivery of luminal antigens to the underlying immune cells.
Results: FITC-dextran uptake into the serum following massive SBR was increased compared to sham mice (1.83 ± 0.15 ng/µL vs. 1.34 ± 0.22 ng/µL). EM of tight junction complexes were more obscured with irregular arrangements in resected mice compared to sham operated mice (graphic 1). Single seq analysis after SBR showed a 21% decrease in occludin expression and a 30% decrease in claudin 2 and 3 expression compared to sham mice. Compared with unoperated mice, the number of GAPS appeared similarly increased after both SBR fand sham operation in both proximal and distal small intestine.
Conclusion: Massive SBR is associated with increased intestinal barrier permeability as revealed by increased FITC dextran absorption into the serum. This appears to be in part due to structural changes at the tight junction level and reduced expression of tight junction proteins occludin, claudin-2/3. The expression of GAPs was not statistically different between SBRs and shams suggesting that the alterations in permeability are primarily driven by changes at the paracellular level. These findings suggest potential targets to restoring the intestinal barrier, thus preventing intestinal resected-associated liver disease.
