Date Published: June 10, 2019
Publisher: Public Library of Science
Author(s): Aarti Kathrani, Victor Lezcano, Edward J. Hall, Albert E. Jergens, Yeon-Jung Seo, Jonathan P. Mochel, Todd Atherly, Karin Allenspach, Simon Russell Clegg.
Dogs with protein-losing enteropathy (PLE) have decreased serum tryptophan concentrations, which may contribute to disease pathogenesis. Indoleamine-pyrrole 2,3-dioxygenase-1 (IDO-1) expression is associated with low serum tryptophan concentrations and is increased in the gastrointestinal tract of humans with inflammatory bowel disease (IBD). Therefore, the objective of our study was to determine if the mRNA expression of IDO-1 is increased in the duodenal mucosa of dogs with PLE as compared to dogs with chronic enteropathy (CE) and healthy dogs, and whether this expression is correlated with changes in serum tryptophan concentration.
Our study was a retrospective study using archived paraffin-embedded duodenal biopsy specimens from 8 healthy Beagle dogs from the Iowa State University Canine Service Colony and 18 and 6 client-owned dogs diagnosed with CE and PLE, respectively at the Bristol Veterinary School. A novel RNA in situ hybridization (ISH) technology, RNAscope, was used to identify IDO-1 mRNA mucosal expression in duodenal tissues. An IDO-1 specific probe was hybridized onto 10 duodenal biopsy sections from each dog whereby RNAscope signal (mRNA expression) was quantified by a single operator using light microscopy.
Dogs with PLE had significantly higher mRNA expression of IDO-1 in the duodenal mucosa compared to healthy dogs (mucosal percentage IDO-1 positive: P = 0.0093, (mean ± S.D) control: 19.36 ± 7.08, PLE: 34.12 ± 5.98, average fold difference: 1.76 and mucosal IDO-1 H-score: P = 0.0356, (mean ± S.D) control: 45.26 ± 19.33, PLE: 84.37 ± 19.86, average fold difference: 1.86). The duodenal mucosal mRNA expression of IDO-1 was negatively correlated with serum tryptophan concentrations in dogs with PLE (mucosal IDO-1 H-score: Spearman’s rank correlation coefficient = -0.94, P = 0.0048).
In conclusion, our study suggests that decreased serum tryptophan concentrations in dogs with PLE is associated with increased intestinal IDO-1 expression. Further studies are needed to determine potential inflammatory pathways responsible for increased expression of IDO-1 in the intestinal tract of dogs with PLE.
Indoleamine-pyrrole 2,3-dioxygenase-1 (IDO-1) is an enzyme expressed by cells of the innate immune system and acts as the interface with the adaptive immune system . IDO-1 promotes immune tolerance by influencing T-cell proliferation and clonal expansion and has also been shown to have antimicrobial properties [1, 2]. Toll-like receptor activation as well as pro-inflammatory cytokines, such as interferon-gamma and tumor necrosis factor- alpha, may induce IDO-1 expression [1, 3]. Therefore, IDO-1 expression is increased in the gastrointestinal (GI) tract of both humans with inflammatory bowel disease (IBD) and animal models of colitis, due to overexpression of pro-inflammatory cytokines and over-activation of toll-like receptors [4–6]. IDO-1 is also the initial rate-limiting step in the pathway for the oxidation of tryptophan to kynurenine . Therefore, increased IDO-1 expression is associated with lower serum tryptophan concentrations in humans with IBD due to increased tryptophan catabolism .
In this study, we show that dogs with PLE have significantly higher percentage mucosal IDO-1 positive mRNA and mucosal IDO-1 H-score in the duodenum compared to healthy Beagle control dogs. IDO-1 is the first and rate-limiting step in the catabolism of tryptophan in the kynurenine pathway [17, 18]. Through the catabolism of tryptophan and the generation of kynurenine metabolites, IDO-1 plays an important role in mucosal immune tolerance . IDO-1 activity promotes depletion of tryptophan in the microenvironment, which induces cell cycle arrest of T-cells and increases their apoptosis . However, recent evidence suggests that in a normal environment containing fatty acids, the immunosuppressive effect of IDO might not be due to decreased T cell survival and proliferation, as IDO supplied the required energy for cell survival and proliferation by increasing fatty acid oxidation . Nevertheless, the catabolism of tryptophan via IDO-1 also leads to the generation of immune-modulating kynurenine metabolites . These metabolites induce tolerogenic dendritic cells as well as T regulatory cells [23–25]. Kynurenine also selectively activates the transcription of aryl hydrocarbon, which results in the differentiation of CD4+ T cell into immunosuppressive T regulatory cells . IDO- 1 also possesses a non-enzymatic function that contributes to TGF-beta led tolerance . Therefore, IDO-1 expressing antigen presenting cells are crucial in suppressing pro-inflammatory mucosal T-cells that drive chronic intestinal inflammation. Epithelial IDO-1 activity may also function to limit microbial invasion as IL-27 has been shown to inhibit growth of intestinal bacteria and promote epithelial barrier protection via induction of IDO-1 in human and mouse intestinal epithelial cells . Therefore, intestinal IDO-1 acts as a natural brake to inflammatory responses via the metabolism of tryptophan.