DOP66 Ileal resections from fibrotic-CD patients present microbiota dysbiosis, altered metabolomic profiles and metabolite-sensing GPCRs expression
Bauset, C.(1)*;Carda-Diéguez, M.(2);Buetas, E.(2);Lis-López, L.(1);Seco-Cervera, M.(3);Coll, S.(1);Ortiz-Masiá, D.(4);Calatayud, S.(5);Barrachina, M.D.(5);Mira, Á.(6);Cosín-Roger, J.(3);
(1)Universitat de València, Pharmacology, València, Spain;(2)FISABIO, Genomics and Health, València, Spain;(3)Universitat de València-FISABIO, Pharmacology, València, Spain;(4)Universitat de València-CIBERehd, Medicine, València, Spain;(5)Universitat de València-CIBERehd, Pharmacology, València, Spain;(6)FISABIO-CIBERESP, Genomics and Health, València, Spain;
Background
Crohn’s Disease (CD) is a subtype of IBD characterized by a chronic transmural inflammation of the gastrointestinal tract associated with several complications being intestinal fibrosis the most frequent. CD patients present microbiota dysbiosis and altered metabolomic profiles. GPCRs constitute a family of receptors which could be involved in inflammatory and fibrotic processes associated to CD. We aim to characterize microbiota composition, tissue metabolomic profile and metabolite-sensing GPCRs expression in ileal resections from fibrotic CD patients.
Methods
Ileal resections from B2-CD (n=21) and non-IBD (n=13) patients were obtained. Microbiota characterization was performed by 16S rRNA gene Illumina Miseq sequencing. Bioinformatic analysis of sequencing data was performed using constrained correspondence analysis and non-parametric Wilcoxon test to compare species proportions. Bacterial load was estimated by qPCR. Metabolomic analysis was performed by NMR. Results are expressed as μg metabolite/g tissue. Murine intestinal fibrosis was induced in C67BL/6 mice by: a) the heterotopic intestinal transplant model and b) chronic administration of 4 cycles of increasing DSS percentages. Gene expression of GPCRs was analyzed by qPCR. Data were expressed as fold induction vs control (mean±SEM) and compared by a t-test. Correlations were analyzed with the Spearman coefficient.
Results
First, microbiota analysis revealed a reduction in bacterial diversity and load in fibrotic CD patients. Then, in B2-CD samples we found at genus level Enterococcus genera significantly decreased and at species level Ruminococcus bromii and Faecalibacterium prausnitzii also reduced compared to controls.
From the metabolomic analysis, altered levels of metabolites were found in ileal resections from fibrotic CD patients as summarized in Table 1.
Next, B2-CD patients exhibited differential expression of metabolite-sensing GPCRs vs non-IBD as shown in Table 2.
Moreover, gene expression of fibrotic markers was analyzed in B2-CD patients and significantly increased levels of COL1A1 (13.22±4.38), COL3A1 (1.84±0.52), and COL4A1(7.75±2.19), were found vs controls. Of interest, GPR81, GPR84, GPR4 and GPR68 positively correlated with profibrotic markers, specifically with COL1A1 and COL4A1.
Finally, in line with human results, we also analyzed the expression of metabolite-sensing GPCRs in two different murine colitis models and results obtained are represented in Table 3.
Conclusion
Fibrotic CD patients exhibit microbial dysbiosis, joined with altered levels of metabolites and gene expression of metabolite-sensing GPCRs, which are also affected in murine colitis models. Their correlation with profibrotic markers points them as protagonists of intestinal fibrosis.