Transient Receptor Potential Vanilloid 1 Function Research Articles

Transient receptor potential vanilloid-1 (TRPV1) channels act as suppressors of the growth of glioma

The aim of this study was to investigate the expression and function of the transient receptor potential vanilloid 1 (TRPV1) in glioma. We found that the expression of TRPV1 mRNA and protein were upregulated in glioma compared with normal brain by qPCR and western blot analysis. In order to investigate the function of TRPV1 in glioma, short hairpin RNA (shRNA) and the inhibitor of TRPV1 were used. In vitro, the activation of TRPV1 induced cell apoptosis with decreased migration capability and inhibited proliferation, which was abolished upon TRPV1 pharmacological inhibition and silencing. Mechanistically, TRPV1 modulated glioma proliferation through the protein kinase B (Akt) signaling pathway. More importantly, in immunodeficient (NOD-SCID) mouse xenograft models, tumor size was significantly increased when TRPV1 expression was disrupted by a shRNA knockdown approach in vivo. Altogether, our findings indicate that TRPV1 negatively controls glioma cell proliferation in an Akt-dependent manner, which suggests that targeting TRPV1 may be a potential therapeutic strategy for glioma.

TRPV1 Channel in Human Eosinophils: Functional Expression and Inflammatory Modulation.

The transient receptor potential vanilloid 1 (TRPV1) is a non-selective cation channel expressed on sensory neurons and immune cells. We hypothesize that TRPV1 plays a role in human eosinophil function and is modulated by inflammatory conditions. TRPV1 expression on human eosinophils was examined by qPCR, flow cytometry, and immunohistochemistry, respectively. TRPV1 functionality was analyzed by investigating calcium flux, apoptosis, modulation by cytokines and acidic pH, and CD69 externalization using flow cytometry. Activation of TRPV1 induced calcium influx and prolonged survival. Although eosinophils were not directly activated by TRPV1 agonists, activation by IL-3 or GM-CSF was mainly restricted to TRPV1-positive eosinophils. TRPV1 surface content was increased by acidic pH, IL-3, IL-31, IL-33, TSLP, TNF-α, BDNF, and NGF-β. Interestingly, TRPV1 was also expressed by eosinophils located in proximity to peripheral nerves in atopic dermatitis (AD) skin. In conclusion, eosinophils express functional TRPV1 channels which are increased by extracellular acidification and AD-related cytokines. Since eosinophils also express TRPV1 in AD skin, our results indicate an important role of TRPV1 for neuroimmune interaction mechanisms in itchy, inflammatory skin diseases, like AD.

MiR‑3120/Hsc70 participates in forced swim stress‑induced mechanical hyperalgesia in rats in an inflammatory state.

The heat shock cognate 71kDa protein (Hsc70) is a stress‑inducible ATPase that can protect cells against harmful stimuli. Transient receptor potential vanilloid 1 (TRPV1) is a well‑documented nociceptor. Notably, Hsc70 can inhibit TRPV1 expression and function, suggesting that Hsc70 may have pain regulation potential. However, the role of Hsc70 in stress‑induced hyperalgesia remains unclear. In the present study, the participation of Hsc70 and its regulator microRNA (miR)‑3120 were investigated in forced swim (FS) stress‑induced mechanical hyperalgesia in rats in an inflammatory state. Complete Freund's adjuvant (CFA) hind paw injection was performed to induce inflammatory pain in rats (CFA rats). Furthermore, in FS+CFA rats, FS stress was performed for 3days before CFA injection. The levels of Hsc70, miR‑3120 and their downstream molecule TRPV1 were measured in the dorsal root ganglion (DRG) with western blotting, immunofluorescence, reverse transcription‑quantitative polymerase chain reaction and fluorescence insitu hybridization. The results revealed that FS stress significantly exacerbated CFA‑induced mechanical pain. Furthermore, CFA upregulated Hsc70 and TRPV1 expression, which was partially inhibited or further enhanced by FS stress, respectively. In FS+CFA rats, intrathecal injection of a lentiviral vector overexpressing Hsc70 (LV‑Hsc70) could decrease TRPV1 expression and improve the mechanical pain. Additionally, the expression levels of miR‑3120, a regulator of Hsc70, were markedly upregulated on day3 following FS stress. Finally, miR‑3120 was identified to be colocalized with Hsc70 and expressed in all sizes of DRG neurons. In CFA rats, DRG injection of miR‑3120 agomir to induce overexpression of miR‑3120 resulted in similar TRPV1 expression and behavioral changes as those caused by FS stress, which were abolished in the presence of LV‑Hsc70. These findings suggested that miR‑3120/Hsc70 may participate in FS stress‑induced mechanical hyperalgesia in rats in an inflammatory state, possibly via disinhibiting TRPV1 expression in the DRG neurons.

TRPV1 activation in human Langerhans cells and T cells inhibits mucosal HIV-1 infection via CGRP-dependent and independent mechanisms

Upon its mucosal transmission, HIV type 1 (HIV-1) rapidly targets genital antigen-presenting Langerhans cells (LCs), which subsequently transfer infectious virus to CD4+ Tcells. We previously described an inhibitory neuroimmune cross talk, whereby calcitonin gene-related peptide (CGRP), a neuropeptide secreted by peripheral pain-sensing nociceptor neurons innervating all mucosal epithelia and associating with LCs, strongly inhibits HIV-1 transfer. As nociceptors secret CGRP following the activation of their Ca2+ ion channel transient receptor potential vanilloid 1 (TRPV1), and as we reported that LCs secret low levels of CGRP, we investigated whether LCs express functional TRPV1. We found that human LCs expressed mRNA and protein of TRPV1, which was functional and induced Ca2+ influx following activation with TRPV1 agonists, including capsaicin (CP). The treatment of LCs with TRPV1 agonists also increased CGRP secretion, reaching its anti-HIV-1 inhibitory concentrations. Accordingly, CP pretreatment significantly inhibited LCs-mediated HIV-1 transfer to CD4+ Tcells, which was abrogated by both TRPV1 and CGRP receptor antagonists. Like CGRP, CP-induced inhibition of HIV-1 transfer was mediated via increased CCL3 secretion and HIV-1 degradation. CP also inhibited direct CD4+ Tcells HIV-1 infection, but in CGRP-independent manners. Finally, pretreatment of inner foreskin tissue explants with CP markedly increased CGRP and CCL3 secretion, and upon subsequent polarized exposure to HIV-1, inhibited an increase in LC-Tcell conjugate formation and consequently Tcell infection. Our results reveal that TRPV1 activation in human LCs and CD4+ Tcells inhibits mucosal HIV-1 infection, via CGRP-dependent/independent mechanisms. Formulations containing TRPV1 agonists, already approved for pain relief, could hence be useful against HIV-1.

Nociception and pain in humans lacking a functional TRPV1 channel.

BACKGROUNDChronic pain is a debilitating illness with currently limited therapy, in part due to difficulties in translating treatments derived from animal models to patients. The transient receptor potential vanilloid 1 (TRPV1) channel is associated with noxious heat detection and inflammatory pain, and reports of adverse effects in human trials have hindered extensive efforts in the clinical development of TRPV1 antagonists as novel pain relievers.METHODSWe examined 2 affected individuals (A1 and A2) carrying a homozygous missense mutation in TRPV1, rendering the channel nonfunctional. Biochemical and functional assays were used to analyze the mutant channel. To identify possible phenotypes of the affected individuals, we performed psychophysical and medical examinations.RESULTSWe demonstrated that diverse TRPV1 activators, acting at different sites of the channel protein, were unable to open the cloned mutant channel. This finding was not a consequence of impairment in the expression, cellular trafficking, or assembly of protein subunits. The affected individuals were insensitive to application of capsaicin to the mouth and skin and did not demonstrate aversive behavior toward capsaicin. Furthermore, quantitative sensory testing of A1 revealed an elevated heat-pain threshold but also, surprisingly, an elevated cold-pain threshold and extensive neurogenic inflammatory, flare, and pain responses following application of the TRPA1 channel activator mustard oil.CONCLUSIONOur study provides direct evidence in humans for pain-related functional changes linked to TRPV1, which is a prime target in the development of pain relievers.FUNDINGSupported by the Israel Science Foundation (368/19); Teva's National Network of Excellence in Neuroscience grant (no. 0394886) and Teva's National Network of Excellence in Neuroscience postdoctoral fellowship.

Capsaicin receptor TRPV1 maintains quiescence of hepatic stellate cells in the liver via recruitment of SARM1

Capsaicin receptor, also known as transient receptor potential vanilloid 1 (TRPV1), is involved in pain physiology and neurogenic inflammation. Herein, we discovered the presence of TRPV1 in hepatic stellate cells (HSCs) and aimed to delineate its function in this cell type and liver fibrosis. TRPV1 expression was examined in liver biopsies from patients with liver fibrosis using quantitative real-time PCR and immunostaining. Its contribution to liver fibrosis was examined in Trpv1-/- mice, upon lentiviral delivery of the TRPV1 gene, and in human and mouse primary HSCs, using patch clamp, intracellular Ca2+ mobilization determination, FACS analyses and gain/loss of function experiments. Binding of sterile alpha and Toll/interleukin-1 receptor motif-containing protein 1 (SARM1) to TRPV1 was determined using mass spectrometry, co-immunoprecipitation, surface plasmon resonance, bioluminescence resonance energy transfer, and NanoBiT. TRPV1 mRNA levels are significantly downregulated in patients with liver fibrosis and mouse models, showing a negative correlation with F stage and α-smooth muscle actin expression, a marker of HSC activation. TRPV1 expression and function decrease during HSC activation in fibrotic livers invivo or during culture. Genetic and pharmacological inhibition of TRPV1 in quiescent HSCs leads to NF-κB activation and pro-inflammatory cytokine production. TRPV1 requires binding of its N-terminal ankyrin repeat domain to the TIR-His583 (Toll/interleukin-1 receptor) domain of SARM1 to prevent HSCs from pro-inflammatory activation. Trpv1-/- mice display increased HSC activation and more severe liver fibrosis, whereas TRPV1 overexpression is antifibrotic in various disease models. The antifibrotic properties of TRPV1 are attributed to the prevention of HSC activation via the recruitment of SARM1, which could be an attractive therapeutic strategy against liver fibrosis. We identified the neuronal channel protein TRPV1 as a gatekeeper of quiescence in hepatic stellate cells, a key driver of liver fibrogenesis and chronic liver disease. Physiologically expressed in healthy liver and consistently downregulated during liver fibrosis development, its therapeutic re-expression is expected to have few side effects, making it an attractive target diagnostic tool and drug candidate for industry and clinicians.

Morphological study of the postnatal hippocampal development in the TRPV1 knockout mice

ABSTRACT Transient receptor potential vanilloid 1 (TRPV1) is a non-selective cation channel with polymodal sensory function. TRPV1 links to fever, while, according to previous studies on TRPV1 knock-out (KO) mice, the role of the channel in the generation of febrile seizure is debated. In the hippocampal formation, functional TRPV1 channels are expressed by Cajal-Retzius cells, which have a role in guidance of migrating neurons during development. Despite the developmental aspects of febrile seizure as well as of Cajal-Retzius cells, no information is available about the hippocampal development in TRPV1 KO mouse. Therefore, in the present work postnatal development of the hippocampal formation was studied in TRPV1 KO mice. Several morphological characteristics including neuronal positioning and maturation, synaptogenesis and myelination were examined with light microscopy following immunohistochemical detection of protein markers of various neurons, synapses, and myelination. Regarding the cytoarchitectonics, neuronal migration, morphological, and neurochemical maturation, no substantial difference could be detected between TRPV1 KO and wild-type control mice. Our data indicate that synapse formation and myelination occur similarly in TRPV1 KO and in control animals. We have found slightly, but not significantly larger numbers of persisting Cajal-Retzius cells in the KO mice than in controls. Our result strengthens previous suggestion concerning the role of TRPV1 channel in the postnatal apoptotic cell death of Cajal-Retzius cells. However, the fact that the hippocampus of KO mice lacks major developmental abnormalities supports the use of TRPV1 KO in various animal models of diseases and pathological conditions.

Transient Receptor Potential Vanilloid 1 Signaling Is Independent on Protein Kinase A Phosphorylation of Ankyrin-Rich Membrane Spanning Protein.

The sensory ion channel transient receptor potential vanilloid 1 (TRPV1) is mainly expressed in small to medium sized dorsal root ganglion neurons, which are involved in the transfer of acute noxious thermal and chemical stimuli. The Ankyrin-rich membrane spanning protein (ARMS) interaction with TRPV1 is modulated by protein kinase A (PKA) mediating sensitization. Here, we hypothesize that PKA phosphorylation sites of ARMS are crucial for the modulation of TRPV1 function, and that the phosphorylation of ARMS is facilitated by the A-kinase anchoring protein 79 (AKAP79). We used transfected HEK293 cells, immunoprecipitation, calcium flux, and patch clamp experiments to investigate potential PKA phosphorylation sites in ARMS and in ARMS-related peptides. Additionally, experiments were done to discriminate between PKA and protein kinase D (PKD) phosphorylation. We found different interaction ratios for TRPV1 and ARMS mutants lacking PKA phosphorylation sites. The degree of TRPV1 sensitization by ARMS mutants is independent on PKA phosphorylation. AKAP79 was also involved in the TRPV1/ARMS/PKA signaling complex. These data show that ARMS is a PKA substrate via AKAP79 in the TRPV1 signaling complex and that all four proteins interact physically, regulating TRPV1 sensitization in transfected HEK293 cells. To assess the physiological and/or therapeutic significance of these findings, similar investigations need to be performed in native neurons and/or in vivo.

TRPV1 as an anti-ferroptotic target in osteoarthritis.

TRPV1 as an anti-ferroptotic target in osteoarthritis.

Role of Src kinase in regulating protein kinase C mediated phosphorylation of TRPV1.

Transient receptor potential vanilloid-1 (TRPV1), activated by heat, acidic pH, endogenous vanilloids and capsaicin, is essential for thermal hyperalgesia. Under inflammatory conditions, phosphorylation of TRPV1 by protein kinase C (PKC) can sensitize the channel and decrease the activation threshold. Src kinase also phosphorylates TRPV1, promoting channel trafficking to the plasma membrane. These post-translational modifications are important for several chronic pain conditions. This study presents a previously undescribed relationship between Src and PKC phosphorylation of TRPV1, influencing the thermal hypersensitivity associated with TRPV1 activation. We assessed TRPV1 channel activity using intracellular calcium imaging and patch-clamp electrophysiology in mouse dorsal root ganglion cultures. Additionally, we used behavioural experiments to evaluate plantar thermal sensitivity following intraplantar injections of activators of known modulators of TRPV1 with and without an Src antagonist. Using calcium imaging and patch-clamp techniques, we demonstrated that pharmacological inhibition of Src kinase or mutation of the Src phosphorylation site on TRPV1 prevented PKC but not PKA-mediated sensitization of TRPV1 in vitro. We found that intraplantar injection of the PKC activator phorbol 12-myristate 13-acetate (PMA) or bradykinin produces thermal hypersensitivity that can be attenuated by pharmacological inhibition of Src. Additionally, complete Freund's Adjuvant (CFA)-induced inflammatory hypersensitivity could also be attenuated by local Src kinase inhibition. Our data demonstrate that Src phosphorylation is critical for PKC-mediated sensitization of TRPV1. Further, in a model of inflammatory pain, CFA, Src kinase inhibition could reduce thermal hypersensitivity. Targeting of Src kinase may have analgesic benefits in inflammatory pain conditions. Src kinase-mediated phosphorylation of TRPV1 is a critical regulator of the PKC-induced sensitization induced by multiple inflammatory mediators. This suggest a new regulatory mechanism governing TRPV1 function and a potential therapeutic target for inflammatory type pain, including cancer pain where Src antagonists are currently utilized.

17β-Estradiol Exacerbated Experimental Occlusal Interference-Induced Chronic Masseter Hyperalgesia by Increasing the Neuronal Excitability and TRPV1 Function of Trigeminal Ganglion in Ovariectomized Rats.

Pain symptoms in temporomandibular disorders (TMD) predominantly affect reproductive women, suggesting that estrogen regulates pain perception. However, how estrogen contributes to chronic TMD pain remains largely unclear. In the present study, we performed behavioral tests, electrophysiology, Western blot and immunofluorescence to investigate the role and underlying mechanisms of estrogen in dental experimental occlusal interference (EOI)-induced chronic masseter mechanical hyperalgesia in rats. We found that long-term 17β-estradiol (E2) replacement exacerbated EOI-induced masseter hyperalgesia in a dose-dependent manner in ovariectomized (OVX) rats. Whole-cell patch-clamp recordings demonstrated that E2 (100 nM) treatment enhanced the excitability of isolated trigeminal ganglion (TG) neurons in OVX and OVX EOI rats, and EOI increased the functional expression of transient receptor potential vanilloid-1 (TRPV1). In addition, E2 replacement upregulated the protein expression of TRPV1 in EOI-treated OVX rats. Importantly, intraganglionic administration of the TRPV1 antagonist AMG-9810 strongly attenuated the facilitatory effect of E2 on EOI-induced masseter mechanical sensitivity. These results demonstrate that E2 exacerbated EOI-induced chronic masseter mechanical hyperalgesia by increasing TG neuronal excitability and TRPV1 function. Our study helps to elucidate the E2 actions in chronic myogenic TMD pain and may provide new therapeutic targets for relieving estrogen-sensitive pain.

TRPV1 Hyperfunction Contributes to Renal Inflammation in Oxalate Nephropathy.

Inflammation worsens oxalate nephropathy by exacerbating tubular damage. The transient receptor potential vanilloid 1 (TRPV1) channel is present in kidney and has a polymodal sensing ability. Here, we tested whether TRPV1 plays a role in hyperoxaluria-induced renal inflammation. In TRPV1-expressed proximal tubular cells LLC-PK1, oxalate could induce cell damage in a time- and dose-dependent manner; this was associated with increased arachidonate 12-lipoxygenase (ALOX12) expression and synthesis of endovanilloid 12(S)-hydroxyeicosatetraenoic acid for TRPV1 activation. Inhibition of ALOX12 or TRPV1 attenuated oxalate-mediated cell damage. We further showed that increases in intracellular Ca2+ and protein kinase C α activation are downstream of TRPV1 for NADPH oxidase 4 upregulation and reactive oxygen species formation. These trigger tubular cell inflammation via increased NLR family pyrin domain-containing 3 expression, caspase-1 activation, and interleukin (IL)-1β release, and were alleviated by TRPV1 inhibition. Male hyperoxaluric rats demonstrated urinary supersaturation, tubular damage, and oxidative stress in a time-dependent manner. Chronic TRPV1 inhibition did not affect hyperoxaluria and urinary supersaturation, but markedly reduced tubular damage and calcium oxalate crystal deposition by lowering oxidative stress and inflammatory signaling. Taking all these results together, we conclude that TRPV1 hyperfunction contributes to oxalate-induced renal inflammation. Blunting TRPV1 function attenuates hyperoxaluric nephropathy.

RSV infection potentiates TRPV1-mediated calcium transport in bronchial epithelium of asthmatic children.

The transient receptor potential vanilloid 1 (TRPV1) channel is expressed in human bronchial epithelium (HBE), where it transduces Ca2+ in response to airborne irritants. TRPV1 activation results in bronchoconstriction, cough, and mucus production, and may therefore contribute to the pathophysiology of obstructive airway disease. Since children with asthma face the greatest risk of developing virus-induced airway obstruction, we hypothesized that changes in TRPV1 expression, localization, and function in the airway epithelium may play a role in bronchiolitis and asthma in childhood. We sought to measure TRPV1 protein expression, localization, and function in HBE cells from children with versus without asthma, both at baseline and after RSV infection. We determined changes in TRPV1 protein expression, subcellular localization, and function both at baseline and after RSV infection in primary HBE cells from normal children and children with asthma. Basal TRPV1 protein expression was higher in HBE from children with versus without asthma and primarily localized to plasma membranes (PMs). During RSV infection, TRPV1 protein increased more in the PM of asthmatic HBE as compared with nonasthmatic cells. TRPV1-mediated increase in intracellular Ca2+ was greater in RSV-infected asthmatic cells, but this increase was attenuated when extracellular Ca2+ was removed. Nerve growth factor (NGF) recapitulated the effect of RSV on TRPV1 activation in HBE cells. Our data suggest that children with asthma have intrinsically hyperreactive airways due in part to higher TRPV1-mediated Ca2+ influx across epithelial membranes, and this abnormality is further exacerbated by NGF overexpression during RSV infection driving additional Ca2+ from intracellular stores.

Type I diabetes suppresses intracellular calcium ion increase normally evoked by heat stress in rat skeletal muscle.

Heat stress, via its effects on muscle intracellular Ca2+ concentrations ([Ca2+]i), has been invoked as a putative therapeutic countermeasure to type 1 diabetes-induced muscle atrophy. Using a circulation- and neurally intact in vivo muscle preparation, we tested the hypothesis that impaired muscle Ca2+ homeostasis in type 1 diabetic rats is due to attenuated heat stress tolerance mediated via transient receptor potential vanilloid 1 (TRPV1). Male Wistar rats were randomly assigned to one of the following four groups: 1) healthy control 30°C (CONT 30°C); 2) CONT 40°C; 3) diabetes 30°C (DIA 30°C); and 4) DIA 40°C. The temperature of 40°C was selected because it exceeds the TRPV1 activation threshold. Spinotrapezius muscles of Wistar rats were exteriorized in vivo and loaded with the fluorescent Ca2+ probe Fura-2 AM. [Ca2+]i was estimated over 20 min using fluorescence microscopy (340/380 nm ratio) in quiescent muscle held at the required temperature, using a calibrated heat source applied to the ventral muscle surface. Western blotting was performed to determine the protein expression levels of TRPV1 in spinotrapezius muscle. After 20 min of heat stress, the CONT 40°C condition induced a 12.3 ± 5% [Ca2+]i (P < 0.05) elevation that was markedly absent in the DIA 40°C or other conditions. Thus, no significant differences were found among DIA 40°C, DIA 30°C, and CONT 30°C. TRPV1 protein expression was decreased by 42.0 ± 9% in DIA compared with CONT (P < 0.05) and, unlike CONT, heat stress did not increase TRPV1 phosphorylation. In conclusion, diabetes suppresses TRPV1 protein expression and function and inhibits the elevated myocyte [Ca2+]i evoked normally by heat stress. These results suggest that capsaicin or other therapeutic strategies to increase Ca2+ accumulation via TRPV1 might be more effective than hyperthermic therapy for type 1 diabetic patients.

The interaction between P2X3 and TRPV1 in the dorsal root ganglia of adult rats with different pathological pains

Peripheral inflammatory and neuropathic pain are closely related to the activation of purinergic receptor P2X ligand-gated ion channel 3 (P2X3) and transient receptor potential vanilloid 1 (TRPV1), but the interaction between P2X3 and TRPV1 in different types of pathological pain has rarely been reported. In this study, complete Freund’s adjuvant (CFA)-induced inflammatory pain and spared nerve injury (SNI)-induced neuropathic pain models were established in adult rats. The interactions between P2X3 and TRPV1 in the dorsal root ganglion were observed by pharmacological, co-immunoprecipitation, immunofluorescence and whole-cell patch-clamp recording assays. TRPV1 was shown to promote the induction of spontaneous pain caused by P2X3 in the SNI model, but the induction of spontaneous pain behaviour by TRPV1 was not completely dependent on P2X3 in vivo. In both the CFA and SNI models, the activation of peripheral P2X3 enhanced the effect of TRPV1 on spontaneous pain, while the inhibition of peripheral TRPV1 reduced the induction of spontaneous pain by P2X3 in the CFA model. TRPV1 and P2X3 had inhibitory effects on each other in the inflammatory pain model. During neuropathic pain, P2X3 facilitated the function of TRPV1, while TRPV1 had an inhibitory effect on P2X3. These results suggest that the mutual effects of P2X3 and TRPV1 differ in cases of inflammatory and neuropathic pain in rats.

Deficiency of Inositol Monophosphatase Activity Decreases Phosphoinositide Lipids and Enhances TRPV1 Function In Vivo.

Membrane remodeling by inflammatory mediators influences the function of sensory ion channels. The capsaicin- and heat-activated transient receptor potential vanilloid 1 (TRPV1) channel contributes to neurogenic inflammation and pain hypersensitivity, in part because of its potentiation downstream of phospholipase C-coupled receptors that regulate phosphoinositide lipid content. Here, we determined the effect of phosphoinositide lipids on TRPV1 function by combining genetic dissection, diet supplementation, and behavioral, biochemical, and functional analyses in Caenorhabditis elegans As capsaicin elicits heat and pain sensations in mammals, transgenic TRPV1 worms exhibit an aversive response to capsaicin. TRPV1 worms with low levels of phosphoinositide lipids display an enhanced response to capsaicin, whereas phosphoinositide lipid supplementation reduces TRPV1-mediated responses. A worm carrying a TRPV1 construct lacking the distal C-terminal domain features an enhanced response to capsaicin, independent of the phosphoinositide lipid content. Our results demonstrate that TRPV1 activity is enhanced when the phosphoinositide lipid content is reduced, and the C-terminal domain is key to determining agonist response in vivo.

Shaoyao-Gancao Decoction Relieves Visceral Hyperalgesia in TNBS-Induced Postinflammatory Irritable Bowel Syndrome via Inactivating Transient Receptor Potential Vanilloid Type 1 and Reducing Serotonin Synthesis.

Postinflammatory irritable bowel syndrome (PI-IBS) is a common functional gastrointestinal disorder, which is characterized by abdominal pain, low-grade inflammation, and visceral hypersensitivity. Shaoyao-Gancao decoction (SGD) has been used to improve the clinical symptoms of abdominal spasmodic pain accompanying acute gastroenteritis, but the underlying therapeutic mechanism has not been fully elucidated. In the present study, a rat model of PI-IBS was established via rectal administration of TNBS. Rats were scored daily for 28 days using disease activity index (DAI). Abdominal withdrawal reflex (AWR) was used to measure the pain threshold. After SGD (6.25, 12.5, and 25 g/kg/d) treatment for 14 days, rat colonic tissue was collected for histopathological grading, enterochromaffin (EC) cell count, and 5-HT content measurement. RT-qPCR and western blot analyses were employed to detect the gene and protein level of tryptophan hydroxylase (TPH), serotonin reuptake transporter (SERT), and transient receptor potential vanilloid 1 (TRPV1). To further validate the effect of SGD on TRPV1, another experiment was performed in cells. The results revealed that visceral hyperalgesia, reflected by increased DAI, AWR, pathological injury score, 5-HT content, and EC cell count in PI-IBS rats, was significantly ameliorated by SGD. In cells, SGD markedly inhibited the expression and function of TRPV1. Moreover, the expression levels of TPH were also repressed by SGD. The findings of the present study indicated that the therapeutic effect of SGD on visceral hyperalgesia may be closely associated with the regulatory role of TRPV1 and 5-HT signaling pathways.

Inhibition of TRPV1 by SHP-1 in nociceptive primary sensory neurons is critical in PD-L1 analgesia.

Recently programmed death-ligand 1 (PD-L1) receptor PD-1 was found in dorsal root ganglion (DRG) neurons, and PD-L1 activates PD-1 to inhibit inflammatory and neuropathic pain by modulating neuronal excitability. However, the downstream signaling of PD-1 in sensory neurons remains unclear. Here, we show that PD-L1 activated Src homology 2 domain-containing tyrosine phosphatase-1 (SHP-1) to downregulate transient receptor potential vanilloid 1 (TRPV1) in DRG neurons and inhibit bone cancer pain in mice. Local injection of PD-L1 produced analgesia. PD-1 in DRG neurons colocalized with TRPV1 and SHP-1. PD-L1 induced the phosphorylation of SHP-1 in DRG TRPV1 neurons and inhibited TRPV1 currents. Loss of TRPV1 in mice abolished bone cancer–induced thermal hyperalgesia and PD-L1 analgesia. Conditioned deletion of SHP-1 in NaV1.8+ neurons aggravated bone cancer pain and diminished the inhibition of PD-L1 on TRPV1 currents and pain. Together, our findings suggest that PD-L1/PD-1 signaling suppresses bone cancer pain via inhibition of TRPV1 activity. Our results also suggest that SHP-1 in sensory neurons is an endogenous pain inhibitor and delays the development of bone cancer pain via suppressing TRPV1 function.

Gain of Function of Ion Channel TRPV1 Exacerbates Experimental Colitis by Promoting Dendritic Cell Activation

Dysregulated mucosal immunity plays an essential role in the pathophysiology of inflammatory bowel disease (IBD). Transient receptor potential vanilloid 1 (TRPV1) is a Ca2+-permeable ion channel that is implicated in modulating immune responses. However, its role in the pathogenesis of intestinal inflammation remains elusive. Here, we found that TRPV1 gain of function significantly increased the susceptibility of mice to experimental colitis, and that was associated with excessive recruitment of dendritic cells and enhanced Th17 immune responses in the lamina propria of colon. TRPV1 gain of function promoted dendritic cell activation and cytokine production upon inflammatory stimuli, and consequently enhanced dendritic cell-mediated Th17 cell differentiation. Further mechanistic studies showed that TRPV1 gain of function in dendritic cells enhanced activation of calcineurin/nuclear factor of activated T cells (NFATc2) signaling induced by inflammatory stimuli. Moreover, in patients with IBD, TRPV1 expression was increased in lamina propria cells of inflamed colon compared with healthy controls. Our findings identify an important role for TRPV1 in modulating dendritic cell activation and sustaining Th17 responses to inflammatory stimuli, which suggest that TRPV1 might be a potential therapeutic target in controlling mucosal immunity and IBD.

Inhibition of Inositol Monophosphatase Enhances TRPV1 Function in vivo

Plasma membrane lipid changes play an essential role in regulating various cellular processes including ion channel function. Membrane remodeling by inflammatory mediators exerts an extra level of modulation in these proteins. The transient receptor potential vanilloid 1 (TRPV1) is an excitatory cation channel involved in neurogenic inflammation and pain hypersensitivity, and its function is regulated by phosphoinositides content. Here, we determined the effect of phosphoinositides on TRPV1 activity in Caenorhabditis elegans using genetic dissection, diet supplementation, and functional analyses. As in mammalian cells, transgenic TRPV1 worms’ aversive response can be positively or negatively modulated with arachidonic acid, a tarantula toxin, and capsazepine. Chemical inhibition of the inositol monophosphatase enzyme (TTX-7) with lithium chloride enhances TRPV1 function in vivo. Moreover, TRPV1 worms lacking the function of TTX-7 have low level of phosphoinositide lipids and display enhanced aversive behavior to capsaicin. On the contrary, TRPV1 worms supplemented with phosphoinositides displayed reduced aversive behavior. Calcium imaging in TRPV1-expressing neurons of live worms shows that capsaicin elicited robust Ca2+ transients in TRPV1 worms fed with control diet, but significantly less changes in worms grown in phosphoinositide supplemented media. Behavior analysis of a TRPV1 worm mutant lacking the C terminus phosphoinositide-interaction site demonstrates that this region modulates channel agonist sensitivity. Our results support that TRPV1 function is negatively regulated by phosphoinositide lipids in vivo.