S3-4 H2S targets transient receptor potential receptors to relieve from inflammation and pain

Abstract

Transient Receptor Potential Vanilloid 4 (TRPV4) and Transient Receptor Potential Ankyrin 1 (TRPA1) are present both in sensory neurons and in intestinal epithelial cells, where their activation causes intracellular calcium mobilization. Activation of each receptor in sensory neurons conveys pro-nociceptive signals, while in intestinal epithelial cells, their activation confers a pro-inflammatory phenotype with chemokine release. Further, TRPV4 or TRPA1 activation in mouse intestine induced visceral hyperalgesia and colitis, confirming that these channels could participate to intestinal pain and inflammation. We hypothesized that H 2 S is able to regulate TRPV4 and TRPA1functions in intestinal epithelial cells and sensory neurons. Methods TRPV4 and TRPA1 responses were evaluated by measuring calcium flux with a Fluo-8 probe in HEK cells transfected with TRPV4 or with TRPA1, but also in normal derived Colon Mucosa (NCM460) cells, Caco-2 cells (an intestinal epithelium cell line) and mouse DRG (dorsal ganglion roots) primary neurons. Cells were pre-exposed to Sodium Hydrogen Sulfide (NaHS) a H 2 S donor, or their vehicle, 3-min before a second stimulation with the specific agonist of TRPV4, 4 α PDD (4 α -phorbol-12,13-didenoate), the agonist of TRPA1 allyl isothiocyanate (AITC) or vehicle. IL-8 expression was also evaluated after 6 h of treatment with NaHS and TRPV4 or TRPA1 stimulation. Results 4 α PDD or AITC but not NaHS increased the intracellular calcium concentration respectively in HEK TRPV4 and HEK TRPA1. TRPV4 or TRPA1 agonist induced a dose-dependent calcium flux in NCM460 cells, Caco-2 cells and neurons. Incubation of NCM460 and Caco-2 cells with NaHS (200–1000 μM) dose-dependently decreased the calcium response to a pro-inflammatory dose of TRPV4 or TRPA1 agonist (50 μM). In neurons, the calcium response to 25 μM of 4 α PDD or AITC was also significantly reduced with a NaHS pre-treatment (1 mM). Furthermore, IL-8 expression was decreased when NCM460 and Caco-2 cells were pre-treated with NaHS. Conclusions NaHS exposure decreased cellular response to TRPV4 or TRPA1 agonist. Considering the pro-inflammatory and pro-nociceptive effects of TRPV4 or TRPA1 activation in the gut, H 2 S could exert there, anti-inflammatory effects through the inhibition of TRPV4 and TRPA1. Hydrogen sulfide efficiently reduces visceral inflammation and pain as previously demonstrated, our results suggest that these protective effects of H2S donnors could be mediated by TRPV4 and TRPA1 inhibition.