J. H. Neilson1, K. Brawner1, S. Dees1, A. Chen1, J. Bibb1, C. Martin1 1University Of Alabama At Birmingham,Department Of Surgery,Birmingham, AL, USA
Introduction: Intestinal epithelial barrier function is critical for appropriate immunity and disease protection. Dysregulation of neural hormone serotonin can affect intestinal subsepratibility to inflammation, However the mechanisms regulating these findings are poorly understood. Serotonin is synthesized through the conversion of tryptophan being rate limited by the enzyme tryptophan hydroxylase (TH). Tryptophan is metabolized into a variety of different molecules, some of which are ligands to the aryl-hydrocarbon Receptor (AhR). AhR is important in intestinal immune function. We hypothesized that depleting serotonin by inhibiting TH would limit inflammation and increase the protective barrier of the gut. One mechanism of protection could be through increased AhR stimulation.
Methods: 8-week-old C57/B6 mice were treated for 5 consecutive days with a 150 mg/kg dose of the TH inhibitor 4-Chloro-L-phenylalanine (PCPA). After 5 days, drug efficacy was measured by quantifying serum serotonin levels. Immune function was measured by quantifying fecal IgA by ELISA. To assess barrier function and bacterial translocation, mesenteric lymph node (MLN) samples were homogenized and plated on Schaedler agar in aerobic conditions. Colonies were counted after 3 days of incubation. AhR ligand availability was measured using a cell-based luciferase reporter assay. HCT 116 cells were used that has been stably transfected with the DRE-driven firefly luciferase construct.
Results: Serotonin was depleted in the PCPA treated mice from an average of 7800 ng/mL (n=3) to 4300 ng/mL (n=3) to a significant degree within the intestine (P=0.004). IgA in the stool showed no difference with pretreated mice averaging 36 µg/mL and an average of 37 µg/mL after treatment (n=6, P=0.89). There was a significant decrease in bacterial translocation in treated mice. Treated mice averaged 4.5 colonies per plate (n=11) while controls averaged 1057 colonies per plate (n=6, P=0.013). The AhR luciferase assay also showed a significant increase in AhR activity in the stool showing a light intensity with an average of 2815 (n=3) for control and 5454 (n=3) for treated mice (P=0.013).
Conclusion: Serotonin depletion augments intestinal barrier function resulting in less bacterial translocation by MLN culture, but has no effect on IgA secretion. This could be due to increased AhR activity causing a variety of effects of the intestinal barrier. AhR signaling is critical for intestinal immune development. One mechanism to explain this finding could be differences in immune cell development allowing for decreased bacterial translocation in serotonin depleted mice. Targeted serotonin regulation may be a way to regulate bacterial translocation in patients at risk for infectious intestinal diseases.