Introduction
Fiber-free diet reduces intestinal and colonic health in mice, in parallel with a reduction in GLP-1 levels. Moreover, as shown here, levels of indole-3-carboxyaldehyde (I3A) in the colonic content are reduced in mice fed a fiber-free diet. Indoles are thought to bacterial metabolites of tryptophan that increase GLP-1 levels in vitro and in vivo. We therefore set out to assess whether I3A similarly increases GLP-1 levels, and whether I3A could restore the adverse effects of fiber-free diet in mice.
Aims & Methods
We started with untargeted metabolomics using LC-MS of colonic content samples from our previous study in fiber-free diet mice. Next, we exposed in vitro GLUTag cells to different levels of I3A (1 nM to 5 mM), and cumulative GLP-1 secretion was measured after 5 min and 240 min. We then performed isolated colon perfusions in male C57BL6/JRJ mice and male Wistar rats, where I3A (500 µM - 10 mM) was administered via the luminal or arterial side. GLP-1 levels were measured in effluent collected from the portal vein. Finally, we performed an in vivo study in chow or fiber-free diet fed female C57BL6/JRJ mice given oral I3A (0.6 mg/day) or vehicle once daily. After 10 days, intestinal permeability was assessed using FITC-dextran, and animals were sacrificed and organs were removed for histology. Colonic contents were analyzed for unspecific indoles with Kovac’s method to ensure that I3A had reached the colon after oral administration.
Results
Mice fed a fiber-free diet had significantly lower I3A in their colonic content than in mice fed a control diet (7883±3375 AU, p=0.04). GLP-1 secretion was unchanged in GLUTag cells after five minutes of exposure to I3A. However, GLP-1 levels increased after 120 minutes of exposure to 1 mM (60% increase, p=0.016) and 5 mM (89% increase, p=0.0025) I3A. In isolated perfused mouse and rat colons, I3A applied into the luminal or vascular side did not affect GLP-1 secretion, regardless of concentration. Mice fed a fiber-free diet tended to weigh less compared to chow fed (mean±SEM at study end: 1.14±0.52 g, p=0.14). The small intestine and colon were smaller in fiber-free diet fed mice (reduction 0.65±0.09 %/body weight and 0.26±0.05 %/body weight, respectively; both p<0.0001); however, crypt depth, villus length, mucosal area and intestinal permeability were unaffected compared to chow fed. Supplementation of I3A to mice fed the fiber-free diet did not affect body or organ weight. Importantly, a substantial increase of indoles in colon content (1.9±0.2 mg/mL vs. 5.4±1.2, p=0.001) confirmed that I3A did reach the colonic lumen.
Conclusion
Fiber-free diet lowered colonic content of I3A in mice. I3A increases GLP-1 secretion in vitro, but not in isolated perfusion models. In prolonged in vivo intervention, the fiber-free diet decreased the weight of the small intestine and colon regardless of I3A administration. Fiber free diet did not affect body weight, crypt depth or villus length, in contrast to our previous study, perhaps due to the relatively short intervention period here. In conclusion, similar to indoles, I3A stimulates GLP-1 secretion in vitro, yet has no evident beneficial effects on intestinal health when administered in vivo. Further studies could look into possible metabolic beneficial effects.
References
Chimerel C, Emery E, Summers DK, Keyser U, Gribble FM, Reimann F. Bacterial Metabolite Indole Modulates Incretin Secretion from Intestinal Enteroendocrine L Cells. Cell Rep. 2014 Nov;9(4):1202–8.