Activation of TRPV1 Receptor by Eugenol in Human Odontoblasts-like Cells

Objectives: One of the dentinal hypersensitivity theories proposes that odontoblasts (ODs) could act as sensory receptor cells by the TRP receptors activity. The TRPV1 subtype in mouse ODs is activated by vanilloid agonists such as capsaicin. The structure-activity relationship analysis indicates that Eugenol, have a vanilloid structure and could also activate it. This work was aimed to evidence the TRPV1 expression and establish its possible chemical activation by Eugenol in human odontoblast-like cells (hOLC).
Methods: The dental pulp were obtained from human third molars extracted by therapeutic indication (Ethics Committee authorization B.CIEFO-122-18). The cells obtained were characterized by flow cytometry, qPCR and immunocytochemistry. The odontoblast differentiation process was performed by 7, 14 and 21 days in odontogenic induction medium with TGF-beta. TRPV1 gene expression in the hOLC was evaluated by RT-qPCR and the localization was determined by immunofluorescence. Cell viability and Eugenol EC50 were first determined by resazurin technique and the TRPV1 activation was evaluated measuring intracellular [Ca2+] using the Fluo-4AM probe. For the statistical analysis a one-way variance analysis (ANOVA) and post-hoc test was used (p<0,05).

Results: An in vitro model of hOLC was established and characterized, which identified the TRPV1 expression. A TRPV1 transcript over-expression at 7, 14 and 21 days of differentiation was evidenced (p=0.001) and positive immunostaining in the hOLC cell membrane from the seventh day of differentiation. The activation was evident by stimulation with Eugenol since calcium concentrations significantly changed (p<0,05).

Conclusions: hOLC express active TRPV1 calcium channel and could be involved as transducer of dental painful sensation. This activation could induce desensibilization and analgesia as capsaicin does. This finding provides information to understand the dental hypersensitivity and contribute to develop therapeutic alternative.