RESUMEN
BACKGROUND AND PURPOSE: Pulpitis is associated with tooth hypersensitivity and results in pulpal damage. Thermosensitive transient receptor potential (TRP) ion channels expressed in the dental pulp may be key transducers of inflammation and nociception. We aimed at investigating the expression and role of thermo-TRPs in primary human dental pulp cells (hDPCs) in normal and inflammatory conditions. EXPERIMENTAL APPROACH: Inflammatory conditions were induced in hDPC cultures by applying polyinosinic:polycytidylic acid (poly(I:C)). Gene expression and pro-inflammatory cytokine release were measured by RT-qPCR and ELISA. Functions of TRPA1 channels were investigated by monitoring changes in intracellular Ca2+ concentration. Mitochondrial superoxide production was measured using a fluorescent substrate. Cellular viability was assessed by measuring the activity of mitochondrial dehydrogenases and cytoplasmic esterases. TRPA1 activity was modified by agonists, antagonists, and gene silencing. KEY RESULTS: Transcripts of TRPV1, TRPV2, TRPV4, TRPC5, and TRPA1 were highly expressed in control hDPCs, whereas TRPV3, TRPM2, and TRPM3 expressions were much lower, and TRPM8 was not detected. Poly(I:C) markedly up-regulated TRPA1 but not other thermo-TRPs. TRPA1 agonist-induced Ca2+ signals were highly potentiated in inflammatory conditions. Poly(I:C)-treated cells displayed increased Ca2+ responses to H2O2, which was abolished by TRPA1 antagonists. Inflammatory conditions induced oxidative stress, stimulated mitochondrial superoxide production, resulted in mitochondrial damage, and decreased cellular viability of hDPCs. This inflammatory cellular damage was partly prevented by the co-application of TRPA1 antagonist or TRPA1 silencing. CONCLUSION AND IMPLICATIONS: Pharmacological blockade of TRPA1 channels may be a promising therapeutic approach to alleviate pulpitis and inflammation-associated pulpal damage.
Asunto(s)
Pulpa Dental , Estrés Oxidativo , Pulpitis , Canal Catiónico TRPA1 , Regulación hacia Arriba , Humanos , Estrés Oxidativo/efectos de los fármacos , Canal Catiónico TRPA1/metabolismo , Canal Catiónico TRPA1/antagonistas & inhibidores , Pulpitis/metabolismo , Pulpitis/patología , Pulpa Dental/citología , Pulpa Dental/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Células Cultivadas , Canales de Potencial de Receptor Transitorio/metabolismo , Canales de Potencial de Receptor Transitorio/antagonistas & inhibidores , Poli I-C/farmacología , Supervivencia Celular/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/efectos de los fármacos , Calcio/metabolismo , Superóxidos/metabolismoRESUMEN
BACKGROUND: Our previous studies demonstrated that sour cherry anthocyanins (AC) reduce the salivary count of Streptococcus mutans and inhibit salivary amylase activity within 30 minutes after chewing AC gum. AC gum and changing toothbrushes after scaling reduced the Gram-negative species in the unstimulated salivary microbiota. The present study examined the effect of AC gums on salivary factors, including changes in microbiome. METHODS: The study was conducted over three weeks with two groups; young adults (18-30) and adults (30-45). Ten participants changed their toothbrushes, while the other 10 participants did not change after the control period. After scaling, all participants received three doses of AC gum daily. The salivary mRNA and protein levels of cytokines, mucins, melatonin, and the microbiota of unstimulated and stimulated saliva were determined by polymerase chain reaction, enzyme-linked immunosorbent assay, and 16S rRNA gene sequencing. RESULTS: Significantly higher levels of tumor necrosis factor α (TNFα), interleukin-1ß (IL-1ß), mucin5B (MUC5B), mucin7 (MUC7), and melatonin were detected in stimulated saliva. Correlation analysis of these factors with the microbiota showed positive correlations with the genera Lachnospiraceae, Eikenella, Saccharibacteria_(TM7), Streptococcus, Prevotella, and Haemophilus. CONCLUSIONS: AC chewing gum has a beneficial effect on the composition of the oral microbiome, and toothbrush replacement leads to changes in the levels of salivary pro-inflammatory cytokines.