EPITHELIAL-DERIVED NADPH OXIDASE (NOX)-1 IN ENTERAL GLUCOSE-MEDIATED RESTORATION OF CRYPT ACTIVITY AND REINFORCEMENT OF BARRIER FUNCTION IN ISCHEMIC JEJUNUM

The intestinal epithelium, functioning as a partition between internal and external environments, produces physiological levels of reactive oxygen species (ROS), playing a vital role in maintaining barrier function and crypt activity. However, whether crypts retain these abilities after an ischemic challenge and the underlying mechanism remain unclear. Our previous study demonstrated that adding glucose to a ligated jejunal loop protects against superior myenteric artery (SMA) ischemia-induced barrier defects, including the mitigation of bacterial translocation (BT) into the liver and spleen. Notably, the reduction of BT occurs over the whole intestinal tract, not only restricted in the ligated glucose-containing loop. We hypothesized that local jejunal glucose-sensing might confer on the remote intestinal epithelium regeneration potential and fortify their barrier function. In this study, a 10 cm ligated jejunal loop was created and filled with a Krebs buffer containing 0 or 25 mM glucose (local jejunum) in rats. The adjacent jejunum was designated as the remote jejunum without manipulation. The rats then received a sham operation or ischemia challenge by occlusion of the SMA for 20 mins. Enteral addition of glucose in the local jejunum ameliorated ischemia-induced barrier defects, histopathological scores, and mucosal inflammation (myeloperoxidase and inflammatory cytokine production) in the remote jejunum. After ischemia, goblet cells in the remote jejunum showed depleted mucin granules and low MUC2 expression. Local addition of glucose enhanced MUC2 synthesis and stimulated mucus secretion in the remote jejunum. The activation of remote crypt stem cells after enteral glucose was evidenced by proliferating marks (Ki67) strongly colocalized with NADPH oxidase (NOX)-1 expression of the ischemic jejunum. To unveil the potential mechanism, we employed ischemic rats and mice in subsequent studies. An in vivo imaging system (IVIS) was used to visualize intestinal epithelial-derived NOX1 production by using the chemiluminescent probe L-012 (detection of NOX-derived superoxide in the gut). L-012-positive signal and NOX1 expression were diminished in ischemic rodents, which was restored by oral or enteral administration of glucose prior to ischemic challenge. Taken together, intestinal ischemia elevated phagocyte-derived inflammation while concurrently reducing epithelial-derived ROS levels, indicating the decoupling of phagocytes and epithelium in the ischemia-induced inflammatory response. Our results suggest that epithelial- derived NOX1 were involved in local glucose sensing and modulates remote epithelial function and crypt activity after ischemic challenge. Further investigation on the exact sensor of enterocyte type and mechanisms in in regulating the behavior of remote crypt cells is still needed.