Tu1398 Direct and Specific Inhibition of Constitutive Nitric Oxide Mediated Regulation of Brush Border Membrane NHE3 and SGLT1 in Intestinal Epithelial Cells

Abstract

Background and aim:diarrhea is a common disease in gastroenterology which is caused by various affects such as intestinal infection, non-infectious inflammation, tumors and so on. The imbalance of fluid/sodium absorption and secretion is considered playing a crucial role in diarrhea. Sodium, as an important electrolyte, involves three mechanisms to transport. Of note, sodium/hydrogen exchanger (NHE) may occupy a crucial position in modulating intestinal water and sodium absorption. Previous study had shown that somatostatin (SST) could stimulate NHE8 expression in physiological intestine. To determine the benefical effect of SST on NHE8 protein expression in colitis mice and its mechanisms, experimental colitis was induced in mice utilizing dextran sulfate sodium (DSS), models of Caco-2 cells intervened by TNF-α were also established to further explore the mechanism of somatostatin modulating NHE8 expression. Methods: To induce diarrhea via intestinal inflammation, mice were fed with 3% Dextran sulfate sodium (DSS) water for seven days. On the eighth day, treatment groups were administrated with octreotide at dose of 50 μg/ kg body weight three times a day for three days. On the eleventh day, mice were euthanized and colonic tissues were collected. Diarrheal symptoms were assessed every other day. Diarrheal score was recorded based on fecal shape, color and hardness. For the TNF-α study, Caco-2 cells were incubated with TNF-α for 18 hours before adding somatostatin. Cells were exposed to somatostatin for 1 hour before harvest. To further explore the mechanism of somatostatin modulating NHE8 expression, Caco-2 cells were incubated with TNF-α for 18 hours. Subsequently, Caco-2 cells pretreated with MAPKK inhibitor (PD98059) were administratedwith SST for 1 hour before harvest.Results: For DSS colitis mice, the expression of somatostatin in colon were decreased in DSS colitis mice compared with the control mice. Moreover, SST could not only ameliorate diarrhea in inflammatory colitis (diarrheal score: 1.7 ± 0.78 vs. 3.6 ± 0.16, n = 3, P<0.05) but also stimulate NHE8 expression in proximal and distal colon compared with DSS colitis mice (proximal colon: 0.61±0.08 vs. 0.31±0.04, n=3, P<0.05; distal colon: 0.74±0.1 vs. 0.36±0.06, n=3, P<0.05). For TNF-α intervened Caco-2 cells, SST could stimulate NHE8 expression compared with TNF-α intervened cells (0.5 ± 0.04 vs. 0.25 ± 0.01; n=3, P<0.05). Furthermore, compared with TNF-α intervened cells, SST could decline the phosphorylation of ERK1/2 (P<0.05). conclusions :In conclusion, the present study suggested that somatostatin could up-regulate the expression of NHE8 protein in both DSS colitis mice and TNF-α intervened Caco-2 cells by blocking the activation of the MAPK-ERK1/2 pathway.