Su1939 Neuro-Immune Changes in IBS: A Link Between Microbiota, TLRs and Sensory-Related Markers?

Abstract

BACKGROUND: Visceral hypersensitivity is a hallmark of Irritable Bowel Syndrome (IBS). Dysbiosis of the gut microbiota and altered host-bacterial interactions appear to contribute to IBS pathology and might be involved in intestinal hypersensitivity. However, the exact mechanisms underlying these interactions are unknown. AIM: To investigate the relationship between intestinal nociceptive system and Toll-like receptors (TLRs) in colonic tissue of IBS patients and to assess if similar relationship exists in an animal model of IBS based on the hyper stimulation of TLRs in rats. METHODS: Gene expression (RT-qPCR) of sensoryrelated [cannabinoid receptor 1 (CB1) and 2 (CB2) and vanilloid receptor type 1 (TRPV1)] and host-bacterial interactions markers (TLR2, 4, 5 and 7) and antimicrobial peptides (AMPs, defensins) was assessed in sigmoidal colonic biopsies of IBS patients and healthy controls (n=13 each). SD male rats were treated intracolonically with a TLR4 agonist (LPS, Escherichia coli O5:B55, 0.2 mg/rat) or a TLR7 agonist [Imiquimod (IMQ), 0.1 mg/rat] for 5 days (n= 6 each). Colons were collected for analysis of sensory markers, TLRs, AMPs and cytokines (RT-qPCR), quantification of secretory-IgA levels (s-IgA; ELISA) and histology. RESULTS: Colonic biopsies of IBS patients showed a significant up-regulation of CB1 and CB2 (P<0.001), TRPV1 (P<0.05) and TLR2 and TLR7 (P<0.001) expression when compared to healthy individuals. A positive correlation was observed between CB1, CB2 and TRPV1 expression and TLR2 and 7 expression (P<0.001). Minor changes were detected for TLR4, TLR5 and defensin 5. Using TLR agonists in rats to mimic a dysbiotic-like state we found that selective repetitive TLR4/7 stimulation induced a mild colonic immune activation characterized by increase of pro-inflammatory cytokines and increased levels of s-IgA, without overt histological changes; which is compatible with features associated to IBS. Both TLR agonists upregulated colonic expression of host TLRs (TLR2>TLR7>TLR5>TLR4). Moreover, intracolonic LPS (TLR4) lead to an up-regulation of CB1, CB2 and TRPV1, while IMQ down-regulated the expression of TRPV1. Overall, higher changes were seen with LPS (TLR4) vs. IMQ (TLR7). Similar to IBS biopsies, minor changes were seen in the expression of AMPs (defensins). CONCLUSIONS: These observations point towards an active communication between the microbiota and intestinal neuro-immune pathways, mainly by connecting the immune TLRdependent signalling to the intestinal nociceptive system (endocannabinoid and vanilloid systems). Furthermore, the stimulation of TLRs by luminal factors appears to be enough to modulate intestinal immune and nociceptive mechanisms. Similar pathways may be part of the underlying pathophysiological mechanisms of IBS thereby linking the microbiota to intestinal neuro-immune pathways and visceral pain.