Avaliação in silico da afinidade de drogas ao receptor TRPA1 como preditor para o manejo farmacológico da sensibilidade provocada pelo clareamento dentário / In silico assessment of drug affinity to the TRPA1 receptor as a predictor for the pharmacological management of sensitivity caused by tooth whitening

Introdução: Reduzir a sensibilidade do clareamento dental em consultório representa um desafio para os profissionais. Pesquisadores associaram o bloqueio do receptor de dor TRPA1 com a redução da sensibilidade ao clareamento. No entanto, a afinidade química dos analgésicos/anti-inflamatórios para o TRPA1 ainda precisa ser averiguada. Objetivo: Realizar uma triagem virtual de múltiplos medicamentos (analgésicos e antiinflamatórios) para verificar a afinidade química pelo receptor TRPA1. Metodologia: A estrutura cristalina das proteínas do receptor TRPA1 foi recuperada do Protein Data Bank. Os códigos SMILES dos ligantes foram extraídos do PubChem. A energia de ligação do complexo foi obtida em ∆G - kcal/mol pelo AutoDock Vina© e replicada nos servidores SwissDock©, Dockthor© e CbDock©. LigPlus© confirmou os sítios de ligação. Resultados: Apesar dos antagonistas dos receptores analisados apresentarem alta afinidade, codeína e dexametasona apresentaram regularidade em todos os servidores, mesmo apresentando valores de energia de ligação de -7,9 kcal/mol para codeína e -8,1 kcal/mol para dexametasona. Conclusão: A codeína e a dexametasona podem ser drogas potenciais para controlar a sensibilidade ao clareamento dental caso atinjam o receptor TRPA1 da polpa dentária (AU).

Introduction: Reducing in-office tooth bleaching sensitivity represents a challenge for professionals. Researchers have associated the block of the pain receptor TRPA1 with reducing bleaching sensitivity. However, the chemical affinity of analgesic/antiinflammatory drugs to the TRPA1 needs to be verified. Objective: To perform a virtual screening of multiple drugs (analgesic and anti-inflammatory drugs) to verify chemical affinity for the TRPA1 receptor. Methodology: The crystal structure of the TRPA1 receptor proteins was retrieved from the Protein Data Bank. The SMILES codes of the ligands were extracted from PubChem. The binding energy of the complex was obtained in ∆G - kcal/mol by AutoDock Vina© and replicated in the webservers SwissDock©, Dockthor©, and CbDock©. LigPlus© confirmed the binding sites. Results: Although the receptor antagonists analyzed showed high affinity, codeine and dexamethasone showed regularity among all servers, even showing binding energy values of -7.9 kcal/mol for codeine and -8.1 kcal/mol for dexamethasone. Conclusion: Codeine and dexamethasone may be potential drugs to manage tooth bleaching sensitivity if they reach the dental pulp TRPA1 receptor (AU).

Mitochondrial Reactive Oxygen Species Elicit Acute and Chronic Itch via Transient Receptor Potential Canonical 3 Activation in Mice / 神经科学通报·英文版

Mitochondrial reactive oxygen species (mROS) that are overproduced by mitochondrial dysfunction are linked to pathological conditions including sensory abnormalities. Here, we explored whether mROS overproduction induces itch through transient receptor potential canonical 3 (TRPC3), which is sensitive to ROS. Intradermal injection of antimycin A (AA), a selective inhibitor of mitochondrial electron transport chain complex III for mROS overproduction, produced robust scratching behavior in naïve mice, which was suppressed by MitoTEMPO, a mitochondria-selective ROS scavenger, and Pyr10, a TRPC3-specific blocker, but not by blockers of TRPA1 or TRPV1. AA activated subsets of trigeminal ganglion neurons and also induced inward currents, which were blocked by MitoTEMPO and Pyr10. Besides, dry skin-induced chronic scratching was relieved by MitoTEMPO and Pyr10, and also by resveratrol, an antioxidant. Taken together, our results suggest that mROS elicit itch through TRPC3, which may underlie chronic itch, representing a potential therapeutic target for chronic itch.

A curcumin derivative J147 ameliorates diabetic peripheral neuropathy in streptozotocin (STZ)-induced DPN rat models through negative regulation AMPK on TRPA1

Abstract Purpose: To investigate the specific molecular mechanisms and effects of curcumin derivative J147 on diabetic peripheral neuropathy (DPN). Methods: We constructed streptozotocin (STZ)-induced DPN rat models to detected mechanical withdrawal threshold (MWT) in vivo using Von Frey filaments. In vitro, we measured cell viability and apoptosis, adenosine 5'-monophosphate-activated protein kinase (AMPK) and transient receptor potential A1 (TRPA1) expression using MTT, flow cytometry, qRT-PCR and western blot. Then, TRPA1 expression level and calcium reaction level were assessed in agonist AICAR treated RSC96cells. Results: The results showed that J147reduced MWT in vivo, increased the mRNA and protein level of AMPK, reduced TRPA1 expression and calcium reaction level in AITCR treated RSC96 cells, and had no obvious effect on cell viability and apoptosis. Besides, AMPK negative regulated TRPA1 expression in RSC96 cells. Conclusions: J147 could ameliorate DPN via negative regulation AMPK on TRPA1 in vivo and in vitro. A curcumin derivative J147might be a new therapeutic potential for the treatment of DPN.

Expressions of TRPV1 and TRPA1 in the dorsal root ganglion in the rat model of orchialgia / 中华男科学杂志

Objective@#To explore the expressions of transient receptor potential vanilloid 1 (TRPV1) and TRP ankyrin 1 (TRPA1) in the dorsal root ganglion (DRG) and their action mechanisms in the rat model of orchialgia.@*METHODS@#The models of orchialgia were established in male SD rats by injection of 2% acetic acid into the testis. Then the number of spontaneous pain responses and withdrawal latency in the model rats were recorded by behavioral tests and the expressions of TRPV1 and TRPA1 in T13－L1 DRGs determined by RT-qPCR, Western blot and immunofluorescence staining.@*RESULTS@#Compared with the normal control rats, the orchialgia models showed a significant increase in the number of spontaneous pain responses (0.13 ± 0.35 vs 22.63 ± 3.42, P<0.01) and a decrease in the withdrawal latency at 4 hours after injection (［12.75 ± 1.50］ vs ［4.85 ± 1.00］ s, P<0.05). The mRNA expressions of both TRPV1 and TRPA1 were observed in the membrane of the neurons in the DRG, the former increased by 1.77 times and the latter by 1.75 times that of the control (P<0.05).@*CONCLUSIONS@#The expressions of TRPV1 and TRPA1 were up-regulated in the DRG of the rat models of orchialgia, which may be involved in the allodynia and hyperalgesia of the rats.

TRPV1 and TRPA1 in cutaneous neurogenic and chronic inflammation: pro-inflammatory response induced by their activation and their sensitization

Cutaneous neurogenic inflammation (CNI) is inflammation that is induced (or enhanced) in the skin by the release of neuropeptides from sensory nerve endings. Clinical manifestations are mainly sensory and vascular disorders such as pruritus and erythema. Transient receptor potential vanilloid 1 and ankyrin 1 (TRPV1 and TRPA1, respectively) are non-selective cation channels known to specifically participate in pain and CNI. Both TRPV1 and TRPA1 are co-expressed in a large subset of sensory nerves, where they integrate numerous noxious stimuli. It is now clear that the expression of both channels also extends far beyond the sensory nerves in the skin, occuring also in keratinocytes, mast cells, dendritic cells, and endothelial cells. In these non-neuronal cells, TRPV1 and TRPA1 also act as nociceptive sensors and potentiate the inflammatory process. This review discusses the role of TRPV1 and TRPA1 in the modulation of inflammatory genes that leads to or maintains CNI in sensory neurons and non-neuronal skin cells. In addition, this review provides a summary of current research on the intracellular sensitization pathways of both TRP channels by other endogenous inflammatory mediators that promote the self-maintenance of CNI.

Cannabinoids increase mechanosensitivity of trigeminal ganglion neurons innervating the inner walls of rat anterior chambers via activation of TRPA1 / 华中科技大学学报（医学）（英德文版）

Our previous study found that some trigeminal ganglion (TG) nerve endings in the inner walls of rat anterior chambers were mechanosensitive, and transient receptor potential ankyrin 1 (TRPA1) was an essential mechanosensitive channel in the membrane. To address the effect of cannabinoids on the mechanosensitive TG nerve endings in the inner walls of anterior chambers of rat eye, we investigated the effect of the (R)-(+)-WIN55, 212-2 mesylate salt (WIN), a synthetic cannabinoid on their cell bodies in vitro. Rat TG neurons innervating the inner walls of the anterior chambers were labeled by 1,1'-dilinoleyl-3,3,3',3'-tetramethylindocarbocyanine, 4-chlorobenzenesulfona (FAST DiI). Whole cell patch clamp was performed to record the currents induced by drugs and mechanical stimulation. Mechanical stimulation was applied to the neurons by buffer ejection. WIN evoked inward currents via TRPA1 activation in FAST DiI-labeled TG neurons. WIN enhanced mechanosensitive currents via TRPA1 activation in FAST DiI-labeled TG neurons. Our results indicate that cannabinoids can enhance the mechanosensitivity of TG endings in the inner walls of anterior chambers of rat eye via TRPA1 activation.

Gingerol activates noxious cold ion channel TRPA1 in gastrointestinal tract / 中国天然药物

TRPA1 channels are non-selective cation channels that could be activated by plant-derived pungent products, including gingerol, a main active constituent of ginger. Ginger could improve the digestive function; however whether ginger improves the digestive function through activating TRPA1 receptor in gastrointestinal tract has not been investigated. In the present study, gingerol was used to stimulate cell lines (RIN14B or STC-1) while depletion of extracellular calcium. TRPA1 inhibitor (rethenium red) and TRPA1 gene silencing via TRPA1-specific siRNA were also used for mechanistic studies. The intracellular calcium and secretion of serotonin or cholecystokinin were measured by fura-2/AM and ELISA. Stimulation of those cells with gingerol increased intracellular calcium levels and the serotonin or cholecystokinin secretion. The gingerol-induced intracellular calcium increase and secretion (serotonin or cholecystokinin) release were completely blocked by ruthenium red, EGTA, and TRPA1-specific siRNA. In summary, our results suggested that gingerol derived from ginger might improve the digestive function through secretion releasing from endocrine cells of the gut by inducing TRPA1-mediated calcium influx.

Expressions of transient receptor potential A1 and related inflammatory factors in the rat model of prostatic inflammation / 中华男科学杂志

<p><b>OBJECTIVE</b>To explore the molecular mechanism of pain associated with chronic prostatitis and chronic pelvic pain syndrome (CP/CPPS) in the rat model of prostatic inflammation.</p><p><b>METHODS</b>Thirty-six male SD rats were equally randomized to an experimental and a control group, the former injected with 50 μl of 3% λ-carrageenan into the ventral prostate to make the model of non-bacterial prostatic inflammation, while the latter with the same volume of sterile saline solution. At 1, 2 and 4 weeks after modeling, the prostate, L6-S1 dorsal root ganglion (DRG) and spinal cord were harvested for examination of the expressions of the nerve growth factor (NGF), transient receptor potential ankyrin 1 (TRPA1), and calcitonin-gene-related peptide (CGRP) by immunohistochemistry and Western blot.</p><p><b>RESULTS</b>The expressions of NGF, TRPA1 and CGRP in the prostatic tissue were all significantly increased in the experimental group as compared with the control (P <0.05), with a gradual decrease with the prolonging of time (P <0.05). In the L6-S1 DRG and spinal cord, the expressions of NGF, TRPA1 and CGRP exhibited no significant differences between the experimental and control groups at 1 week after modeling (P >0.05) and kept at high levels in the experimental group at 2 and 4 weeks, though not significantly different from those at 1 week (P >0.05). Statistically significant differences were observed in the expressions of the three proteins in the experimental rats among different time points (P <0.05), but not between the two groups at any time point (P >0.05).</p><p><b>CONCLUSION</b>The molecular mechanism of CP/CPPS can be evaluated in the rat model of prostatic inflammation established by injecting λ-carrageenan into the prostate. TRPA1 may play an important role in connecting the upstream and down-stream pathways of CP/CPPS-associated pain.</p>

Transient receptor potential A1 is involved in cold-induced contraction in the isolated rat colon smooth muscle / 生理学报

Transient receptor potential (TRP) A1, a member of TRP channel family, is activated by noxious cold. The aims of this study were to determine if TRPA1 contributed to cold-induced contractions in the isolated rat colon preparations and explore the potential mechanisms. The colon smooth muscle layers were surgically isolated from the male Wistar rats and changes in isotonic tension of longitudinal muscle under various treatments were recorded as colonic motilities. Cold stimuli were obtained by the reperfusion with Krebs-Henseleit solution at given temperature using Constant Flow Pump. The mRNA expressions of TRPA1, TRPV1 and TRPM8 in rat colon smooth muscle layer were examined by using reverse transcription-polymerase chain reaction (RT-PCR) techniques. The results showed that the contractions induced by cold stimuli (from 37 degrees C to 12 degrees C stepwise) were inversely proportional to the temperature with a maximum contraction at 17 degrees C in both proximal and distal colons (P<0.01). RT-PCR analysis revealed the expression of TRPA1, but not TRPM8 and TRPV1, in the rat proximal and distal colon smooth muscle layers. Cold-induced colonic contractions were specially inhibited by TRPA1 blocker, ruthenium red (30 μmol/L), in the proximal and distal colon (P<0.05). The cold-induced contractions of proximal (P<0.01, P<0.05) and distal colons (both P<0.001) were almost abolished or inhibited by the pretreatments of TRPA1 agonists, Allyl isothiocyanate (AITC, 300 μmol/L) and cinnamaldehyde (CA, 1 mmol/L). Extracellular calcium removal (EGTA, 1 mmol/L), PLC blocker (U73122, 10 μmol/L) and IP(3) receptor blocker (2-aminoethoxydiphenyl borate, 2-APB, 30 μmol/L) all decreased the contractions evoked by the cooling at 17 degrees C in the proximal and distal colon (P<0.001, P<0.05, P<0.001). Atropine (1 μmol/L) had no effects on these contractions. L-type Ca(2+) channels blocker nifedipine (1 μmol/L) and neurotoxin tetrodotoxin (TTX, 2 μmol/L) decreased the contractile response in the distal colon (P<0.01, P<0.05), but not in the proximal colon. In conclusion, TRPA1 contributes to cold-induced contractions of the rat colon smooth muscle, and the mechanism of TRPA1 activation involves PLC/IP(3)/Ca(2+) pathway. L-type Ca(2+) channel and neurogenic mechanism other than muscarinic receptor might be partially involved in cold-induced contraction of the distal colon, which probably resulted in higher contraction of distal colon compared with that of proximal colon.

Bradykinin modulates ion channel in inflammatory pain / 药学学报

Injury or inflammation induces release of a range of inflammatory mediators. Bradykinin is one of the most important inflammatory mediators and plays a crucial role in mediating inflammatory pain. It is well known that multiple ion channels located in the nociceptors participate in pain sensation. Recent studies demonstrate an important role of bradykinin in regulating the function and expression of pain-related ion channels. This paper summarizes the recent advances in the understanding of the role of bradykinin in modulation of the channels and discusses future possibilities in the treatment of inflammatory pain.

Effects of the neurotrophic factor artemin on sensory afferent development and sensitivity / 生理学报

Artemin is a neuronal survival and differentiation factor in the glial cell line-derived neurotrophic factor family. Its receptor GFRalpha3 is expressed by a subpopulation of nociceptor type sensory neurons in the dorsal root and trigeminal ganglia (DRG and TG). These neurons co-express the heat, capsaicin and proton-sensitive channel TRPV1 and the cold and chemical-sensitive channel TRPA1. To further investigate the effects of artemin on sensory neurons, we isolated transgenic mice (ARTN-OE mice) that overexpress artemin in keratinocytes of the skin and tongue. Enhanced levels of artemin led to a 20% increase in the total number of DRG neurons and increases in the level of mRNA encoding TRPV1 and TRPA1. Calcium imaging showed that isolated sensory neurons from ARTN-OE mice were hypersensitive to the TRPV1 agonist capsaicin and the TRPA1 agonist mustard oil. Behavioral testing of ARTN-OE mice also showed an increased sensitivity to heat, cold, capsaicin and mustard oil stimuli applied either to the skin or in the drinking water. Sensory neurons from wildtype mice also exhibited potentiated capsaicin responses following artemin addition to the media. In addition, injection of artemin into hindpaw skin produced transient thermal hyperalgesia. These findings indicate that artemin can modulate sensory function and that this regulation may occur through changes in channel gene expression. Because artemin mRNA expression is up-regulated in inflamed tissue and following nerve injury, it may have a significant role in cellular changes that underlie pain associated with pathological conditions. Manipulation of artemin expression may therefore offer a new pain treatment strategy.