Transient Receptor Potential Vanilloid 1 mRNA 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.

Expression of the Transient Receptor Potential Vanilloid 1 ion channel in the supramammillary nucleus and the antidepressant effects of its antagonist AMG9810 in mice

The Transient Receptor Potential Vanilloid 1 (TRPV1) non-selective cation channel predominantly expressed in primary sensory neurons of the dorsal root and trigeminal ganglia mediates pain and neurogenic inflammation. TRPV1 mRNA and immunoreactivity were described in the central nervous system (CNS), but its precise expression pattern and function have not been clarified. Here we investigated Trpv1 mRNA expression in the mouse brain using ultrasensitive RNAScope in situ hybridization. The role of TRPV1 in anxiety, depression-like behaviors and memory functions was investigated by TRPV1-deficient mice and pharmacological antagonism by AMG9810. Trpv1 mRNA is selectively expressed in the supramammillary nucleus (SuM) co-localized with Vglut2 mRNA, but not with tyrosine hydroxylase immunopositivity demonstrating its presence in glutamatergic, but not dopaminergic neurons. TRPV1-deleted mice exhibited significantly reduced anxiety in the Light-Dark box and depression-like behaviors in the Forced Swim Test, but their performance in the Elevated Plus Maze as well as their spontaneous locomotor activity, memory and learning function in the Radial Arm Maze, Y-maze and Novel Object Recognition test were not different from WTs. AMG9810 (intraperitoneal injection 50 mg/kg) induced anti-depressant, but not anxiolytic effects. It is concluded that TRPV1 in the SuM might have functional relevance in mood regulation and TRPV1 antagonism could be a novel perspective for anti-depressant drugs.

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.

Ectopic endometriosis in the pelvic cavity evokes bladder hypersensitivity via transient receptor potential ankyrin 1 hyperexpression in rats.

In women with chronic pelvic pain (CPP), interstitial cystitis/bladder pain syndrome (IC/BPS) and endometriosis frequently coexist. The mechanism of these diseases coexisting is explained by cross-sensitization between endometriosis and IC/BPS. The overlapped symptoms may be related to cross-sensitization with transient receptor potential vanilloid 1 (TRPV1) and/or transient receptor potential ankyrin 1 (TRPA1) hyperexpression. This study was aimed at exploring whether bladder hypersensitivity is evoked in the surgically induced ectopic endometriosis rat and whether TRPV1 and/or TRPA1 play a vital role. A total of 63 Sprague-Dawley female rats were divided into two groups, 39 for physiological examination and 24 for molecular analysis. Surgical induction of ectopic endometriosis (ENDO, n=27), surgical sham treatment (n=18), and treatment for endometriosis by GnRH analog (ENDO-G) (n=18) were performed. Bladder function was investigated by cystometry (for TRPV1 in the sham [n=6] and ENDO [n=9] groups and for TRPA1 in the sham [n=6], ENDO [n=9], and ENDO+G [n=9] groups), and TRPV1 and TRPA1 mRNA expressions were measured using real-time qPCR in the bladder and dorsal root ganglia (DRGs). On cystometry, the relative intercontraction interval (ICI) after/before resiniferatoxin (RTx; TRPV1 activator) infusion to the bladder showed no significant difference between the two groups, whereas relative ICI after/before allyl isothiocyanate (AITC; TRPA1 activator) infusion was significantly lower in the ENDO group than in the sham group. TRPA1 mRNA expression in the bladder and L5 DRG was considerably higher in the ENDO group than in the sham group on real-time qPCR. TRPA1 mRNA hyperexpression and bladder hypersensitivity after AITC infusion were reduced in the ENDO-G group. Bladder cross-sensitization in ENDO rats occurs in association with hyperexpression of TRPA1 at both the DRG and the bladder mucosa. This can be understood by the "cross-sensitization of endometriosis to bladder" theory explaining overlapping symptoms among BPS/IC and ectopic endometriosis.

Huayu Sanjie Enema Liquid Relieves Pain in Endometriosis Model Rats by Inhibiting Inflammation, Peripheral Sensitization, and Pelvic Adhesion.

The objective of this study is to observe the effect of relieving pain of Huayu Sanjie enema liquid (HYSJ-EL) on endometriosis model rats and to explore its mechanism of action. Of 24 female Sprague Dawley rats, six were randomly selected as the sham operation group (normal control group). The remaining rats were used to establish rat models of endometriosis through autologous endometrial transplantation combined with estrogen injection. Successfully modeled rats were randomly divided into the model, indomethacin (Western medicine group), and HYSJ-EL (Chinese herbs group) treatment groups. The thermal pain threshold of rats was measured, and hematoxylin and eosin staining was used to observe pathological changes after sampling. Serum levels of prostaglandin E2 (PGE2), interleukin-6 (IL-6), macrophage inflammatory protein-2 (MIP-2), plasminogen activator inhibitor-1 (PAI-1), and transforming growth factor-β (TGF-β) were measured using an enzyme-linked immunosorbent assay (ELISA). Furthermore, the protein and mRNA expression levels of transient receptor potential vanilloid-1 (TRPV1) and tumor necrosis factor-α (TNF-α) in the endometrium and endometriotic lesions were measured using Western blotting and quantitative real-time PCR assays, respectively. Compared to the model group, the heat pain threshold of rats in the HYSJ-EL group was significantly increased (P < 0.01), and the serum levels of PGE2, IL-6, MIP-2, PAI-1, and TGF-β were significantly decreased (P < 0.01), as well as the expression of TRPV1 and TNF-α protein and mRNA in the tissue of the ectopic lesion was significantly decreased (P < 0.05). These results indicate that the Huayu Sanjie enema liquid exerts analgesic effects on endometriosis by inhibiting inflammation, peripheral nerve sensitization, and pelvic adhesion.

Optimized flow cytometric detection of transient receptor potential vanilloid-1 (TRPV1) in human hematological malignancies

The ectopic overexpression of transient receptor potential vanilloid-1 (TRPV1) has been detected in numerous solid cancers, including breast, prostate, pancreatic, and tongue epithelium cancer. However, the expression of TRPV1 in hematological malignancies remains unknown. Here we show through in silico analysis that elevated TRPV1 mRNA expression occurs in a range of hematological malignancies and presents an optimized flow cytometry method to rapidly assess TRPV1 protein expression for both cell lines and primary patient samples. Three anti-TRPV1 antibodies were evaluated for intracellular TRPV1 detection using flow cytometry resulting in an optimized protocol for the evaluation of TRPV1 in hematological malignant cell lines and patients’ peripheral blood mononuclear cells (PBMC). Overexpression of TRPV1 was observed in THP-1 (acute monocytic leukemia) and U266B1 (multiple myeloma, MM), but not U937 (histiocytic lymphoma) compared to healthy PBMC. TRPV1 was also detected in all 49 patients including B-cell non-Hodgkin’s lymphoma (B-NHL), MM, and others and 20 healthy controls. TRPV1 expression was increased in 8% of patients (MM = 2, B-NHL = 2). In conclusion, we provide an optimized flow cytometry method for routine expression analysis of clinical samples and show that TRPV1 is increased in a subset of patients with hematological malignancies.

Manual Acupuncture at ST37 Modulates TRPV1 in Rats with Acute Visceral Hyperalgesia via Phosphatidylinositol 3-Kinase/Akt Pathway.

Acupuncture can significantly ameliorate inflammatory pain in acute visceral hyperalgesia. Hyperalgesia is attenuated by inflammatory mediators that activate transient receptor potential vanilloid 1 (TRPV1), and TRPV1 is regulated by nerve growth factor (NGF)-induced phosphatidylinositol 3-kinase (PI3K)/Akt pathway. However, it is unknown whether NGF-induced PI3K/Akt pathway is associated with manual acupuncture (MA). In this study, the effect and mechanism of MA at Shangjuxu (ST37) and Quchi (LI11) were examined using an acetic acid-induced rat model with visceral hyperalgesia. We demonstrated that MA at ST37 significantly decreased abdominal withdrawal reflex (AWR) scores, proinflammatory cytokine expression (TNF-α, IL-1β, and IL-6), and TRPV1 protein and mRNA expression in rats with acute visceral hyperalgesia compared with the untreated controls, while MA at LI11 showed no effect. The effects of MA at ST37 were reversed after treatment with the PI3K agonist IGF-1 30 min before MA. In rats with visceral hyperalgesia, the upregulation of NGF, tropomyosin-receptor-kinase A (TrkA), PI3K, and phosphorylation-Akt (p-Akt) was decreased by MA at ST37, indicating that TRPV1 regulation via the NGF-induced PI3K/Akt pathway plays a vital role in the effects of MA-mediated amelioration of acute visceral hyperalgesia.

L-carnitine suppresses transient receptor potential vanilloid type 1 activity and myofibroblast transdifferentiation in human corneal keratocytes

Corneal stromal wound healing is a well-balanced process promoted by overlapping phases including keratocyte proliferation, inflammatory-related events, and tissue remodeling. L-carnitine as a natural antioxidant has shown potential to reduce stromal fibrosis, yet the underlying pathway is still unknown. Since transient receptor potential vanilloid 1 (TRPV1) is a potential drug target for improving the outcome of inflammatory/fibrogenic wound healing, we investigated if L-carnitine can mediate inhibition of the fibrotic response through suppression of TRPV1 activation in human corneal keratocytes (HCK). We determined TRPV1-induced intracellular calcium transients using fluorescence calcium imaging, channel currents by planar patch-clamping, and cell migration by scratch assay for wound healing. The potential L-carnitine effect on TRPV1-induced myofibroblast transdifferentiation was evaluated by immunocytochemical detection of alpha smooth muscle actin. RT-PCR analysis confirmed TRPV1 mRNA expression in HCK. L-carnitine (1 mmol/l) inhibited either capsaicin (CAP) (10 µmol/l), hypertonic stress (450 mOsmol/l), or thermal increase (>43 °C) induced Ca2+ transients and corresponding increases in TRPV1-induced inward and outward whole-cell currents. This was accompanied by suppression of injury-induced increases in myofibroblast transdifferentiation and cell migration. In conclusion, L-carnitine contributes to inhibit stromal scarring through suppressing an injury-induced intrinsic TRPV1 activity that is linked with induction of myofibroblast transdifferentiation in HCK cells.

Capsazepine decreases corneal pain syndrome in severe dry eye disease

BackgroundDry eye disease (DED) is a multifactorial disease of the ocular surface accompanied by neurosensory abnormalities. Here, we evaluated the effectiveness of transient receptor potential vanilloid-1 (TRPV1) blockade to alleviate ocular pain, neuroinflammation, and anxiety-like behavior associated with severe DED.MethodsChronic DED was induced by unilateral excision of the Harderian and extraorbital lacrimal glands of adult male mice. Investigations were conducted at 21 days after surgery. The mRNA levels of TRPV1, transient receptor potential ankyrin-1 (TRPA1), and acid-sensing ion channels 1 and 3 (ASIC1 and ASIC3) in the trigeminal ganglion (TG) were evaluated by RNAscope in situ hybridization. Multi-unit extracellular recording of ciliary nerve fiber activity was used to monitor spontaneous and stimulated (cold, heat, and acid) corneal nerve responsiveness in ex vivo eye preparations. DED mice received topical instillations of the TRPV1 antagonist (capsazepine) twice a day for 2 weeks from d7 to d21 after surgery. The expression of genes involved in neuropathic and inflammatory pain was evaluated in the TG using a global genomic approach. Chemical and mechanical corneal nociception and spontaneous ocular pain were monitored. Finally, anxiety-like behaviors were assessed by elevated plus maze and black and white box tests.ResultsFirst, in situ hybridization showed DED to trigger upregulation of TRPV1, TRPA1, ASIC1, and ASIC3 mRNA in the ophthalmic branch of the TG. DED also induced overexpression of genes involved in neuropathic and inflammatory pain in the TG. Repeated instillations of capsazepine reduced corneal polymodal responsiveness to heat, cold, and acidic stimulation in ex vivo eye preparations. Consistent with these findings, chronic capsazepine instillation inhibited the upregulation of genes involved in neuropathic and inflammatory pain in the TG of DED animals and reduced the sensation of ocular pain, as well as anxiety-like behaviors associated with severe DED.ConclusionThese data provide novel insights on the effectiveness of TRPV1 antagonist instillation in alleviating abnormal corneal neurosensory symptoms induced by severe DED, opening an avenue for the repositioning of this molecule as a potential analgesic treatment for patients suffering from chronic DED.

TRPV1, Targeted by miR-338-3p, Induces Neuropathic Pain by Interacting with NECAB2.

A variety of studies have proposed that transient receptor potential vanilloid 1 (TRPV1) is involved in the progression of multiple diseases, including neuropathic pain. Although increased expression of TRPV1 in chronic constriction injury was described earlier, the underlying regulatory mechanisms of TRPV1 in neuropathic pain remain largely unknown. In our study, we constructed a chronic constriction injury (CCI) rat model to deeply analyze the mechanisms underlying TRPV1. RT-qPCR-indicated TRPV1 mRNA and protein expression were extremely upregulated in CCI rat dorsal spinal cord tissues. Then, TRPV1 was corroborated to interact with N-terminal EF-hand Ca2+-binding protein 2 (NECAB2). The mRNA and protein levels of NECAB2 were increased in CCI tissues. Moreover, TRPV1 and NECAB2 together regulated nociceptive procession-associated protein metabotropic glutamate receptor 5 (mGluR5), phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2), and Ca2+ in isolated microglia of CCI rats. Moreover, TRPV1 upregulation apparently increased mechanical allodynia and thermal hyperalgesia as well as the expression of inflammation-associated genes (COX-2, TNF-α, and IL-6). In addition, downregulation of NECAB2 significantly decreased mechanical allodynia and thermal hyperalgesia as well as the expression of COX-2, TNF-α, and IL-6. Furthermore, TRPV1 was confirmed to be a downstream target of miR-338-3p. TRPV1 overexpression abolished the inhibitory effect by miR-338-3p elevation on neuropathic pain development. In summary, this study proved TRPV1, targeted by miR-338-3p, induced neuropathic pain by interacting with NECAB2, which provides a potential therapeutic target for neuropathic pain treatment.

Identification and Quantification of Transient Receptor Potential Vanilloid 1 (TRPV1) in Equine Articular Tissue.

Simple summaryOsteoarthritis is affecting several species including the horse. Transient receptor potential vanilloid 1 (TRPV1)—also known as the ‘chili receptor’—is currently being investigated as a target for treating osteoarthritis in humans. To evaluate whether it could be a potential target in treating osteoarthritis in horses, we collected synovial membrane samples from healthy horse joints and horse joints with joint disease to investigate the expression of the TRPV1. Laboratory analysis showed that TRPV1 is present in horse joints, and that the levels might be elevated in diseased joints, even at a chronic state of disease. This means that TRPV1 could be used as a potential target for osteoarthritis treatment in horses. Future studies in this area will not only be beneficial to horses and their owners, but more knowledge about TRPV1, the mode of action, and possible side effects will also benefit translation into human osteoarthritis research.Joint pain and osteoarthritis (OA) are some of the most common causes of lameness in horses, and most of the available treatments focus on symptomatic relief without a disease-modifying effect. TRPV1 is a potential target for treating joint diseases, including OA, and the present study aims to investigate if the TRPV1 receptor is present in equine articular tissue and determine whether the number of receptors is upregulated in joint inflammation. Metacarpo/metatarsophalangeal (MCP/MTP) joints from 15 horses euthanised for reasons unrelated to this study were included. Based on synovial fluid analysis, macroscopic evaluation, and magnetic resonance imaging (MRI), joints were divided into two groups: healthy joints and joints with pathology. ELISA analysis was performed on synovial tissue harvested from all joints. TPRV1 was found in all joints. The mean concentration of TRPV1 compared to total protein in healthy joints (8.4 × 10−7 ng/mL) and joints with pathology (12.9 × 10−7 ng/mL) differed significantly (p = 0.01, t-test with Welch correction). Quantitative real-time reverse transcriptase PCR analysis was performed on RNA isolates from synovial tissue from all joints. TRPV1 mRNA expression ratio normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in healthy joints (0.16 (SD: 0.19)) and joints with pathology (0.24 (SD: 0.14)) did not differ significantly (p = 0.43, t-test with Welch correction). mRNA expression of interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) was very low for both groups. In conclusion, TRPV1 was detected both on mRNA and the protein level, with a higher expression of TRPV1 in samples from joints with pathology. Future studies will determine the clinical potential of equine TRPV1 as a target in the management of joint pain and inflammation.

Presence and upregulation of Transient Receptor Potential Vanilloid 1 (TRPV1) and Ankyrin 1 (TRPA1) in translational rat endometriosis model

Abstract The Transient Receptor Potential Vanilloid 1 (TRPV1) and Ankyrin 1 (TRPA1) are non-selective cation channels predominantly localized on capsaicin-sensitive sensory neurons; however both receptors have been described in non-neuronal tissues. It has been published that both receptors upregulated in peritoneal endometriosis in humans. Our research group demonstrated that TRPA1 and TRPV1 expression is elevated in human deep infiltrating endometriosis (DIE) lesions and the receptors have an estrogen-dependent expression pattern in rat endometrium. Here, we investigated the expression changes of TRPA1/V1 and the role of the capsaicin-sensitive sensory-nerve endings in a rat model of peritoneal endometriosis. Peritoneal endometriosis was surgically induced in 8-week-old female rats (n=7-7) for 2-weeks (acute condition) and 8-weeks (chronic condition). TRPA1/V1 mRNAs were quantified with quantitative polymerase chain reaction (qPCR). The expression levels were compared with the results obtained earlier from human DIE samples. The blockade of the TRPA1/V1 expressing capsaicin-sensitive nerve endings was induced with resiniferatoxin (RTX), followed by the measurement of the weight and size of the endometriosis lesions. We detected TRPV1 and TRPA1 mRNA in normal rat endometrium, their expression was not altered in sham-operated animals. In chronic, but not in acute endometriosis the expression was significantly elevated in the lesions, which results are consistent with our previous findings in human DIE. The elimination of capsaicin-sensitive nerve endings decreased the weight of the endometriosis lesions while the size of the ectopic tissue was not altered. Taken together, our results obtained from the rat endometriosis model are consistent with the previous human results, therefore this model is considered to have translational significance and it is suitable for functional analysis of the ion channels. The local, non-neuronal TRPA1 and TRPV1 might play a role in inflammation and sensory neuronal activation in endometriosis related pain, which is mediated by a broad range of pro-inflammatory molecules.

125I radiation downregulates TRPV1 expression through miR‑1246 in neuroblastoma cells.

Iodine‑125 (125I) seed radiation applied around the celiac ganglion can relieve the refractory pain in pancreatic cancer. In an in vitro cell radiation model of human neuroblastoma cell lines, the impact of 125I radiation on the expression of transient receptor potential vanilloid‑1 (TRPV1) was investigated. The results indicated that the radiation delivering doses <2.13 Gy did not significantly affect cell growth, whereas the doses >3.12 Gy significantly reduced cell viability. The reduced TRPV1 mRNA level was dependent on the doses, while the reduced protein level occurred at lower doses (2.63 and 4.27 Gy), then returned to normal at an intermediate dose of 5.09 Gy, and decreased again at higher doses (5.91 and 6.73 Gy). The miRNA profiling at the dose of 2.63 Gy revealed 32 and 22 miRNAs that were significantly upregulated and downregulated, respectively. In addition, the upregulated miR‑1246 target, regulated the expression of TRPV1, indicating that miR‑1246 may be a new therapeutic target for pancreatic pain.

Transient receptor potential Vanilloid 1-based gene therapy alleviates orthodontic pain in rats

Orthodontic pain that is induced by tooth movement is an important sequela of orthodontic treatment and has a significant effect on patient quality of life. Studies have shown that the high expression of transient receptor potential vanilloid 1 (TRPV1) in trigeminal ganglions plays a vital role in the transmission and modulation of orofacial pain. However, little is known about the role of TRPV1 in orthodontic pain. In this study, male Sprague–Dawley rats were randomly assigned to six groups to study the role of TRPV1 in the modulation of tooth-movement pain. The expression levels of TRPV1 mRNA and protein were determined by real-time PCR and western blot, respectively. Moreover, pain levels were assessed using the rat grimace scale (RGS). The role of TRPV1 in modulating tooth-movement pain was examined by injecting a TRPV1 antagonist into the trigeminal ganglia of rats. A lentivirus containing a TRPV1 shRNA sequence was constructed and transduced into the rats’ trigeminal ganglia. The results showed that the expression levels of TRPV1 protein and mRNA were elevated following tooth-movement pain. Pain levels increased rapidly on the 1st day, peaked on the 3rd day and returned to baseline on the 14th day. The TRPV1 antagonist significantly reduced tooth-movement pain. The lentivirus containing a TRPV1 shRNA sequence was able to inhibit the expression of TRPV1 and relieved tooth-movement pain. In conclusion, TRPV1-based gene therapy may be a treatment strategy for the relief of orthodontic pain.

Upregulation of extraneuronal TRPV1 expression in chronic rhinosinusitis with nasal polyps.

Chronic rhinosinusitis (CRS) is a multifactorial upper airway disease with unclear etiology. Neuronal Transient Receptor Potential Vanilloid 1 (TRPV1) and Ankyrin 1 (TRPA1) channels have been implicated in the pathogenesis of CRS. We aimed to detect the expression of extraneuronal TRPV1 and TRPA1 receptors in nasal polyp (NP) tissue samples. Samples were obtained from fourty-two CRS pateints with nasal polyp and sixteen healthy controls to measure receptor gene expression by quantitative PCR, protein localization by immunohistochemistry and cytokine profile by multiplex bead immunoassay. Non-neuronal TRPV1, TRPA1 receptors were expressed in biopsy samples of NP. A population of mast cells and macrophages were immunopositive for TRPV1 and TRPA1. A fraction of plasma cells expressed TRPV1 but not TRPA1 and neither receptor was present on eosinophils. The local gene expression of extraneuronal TRPV1, TRPA1 receptors was also proven. TRPV1 mRNA levels were significantly increased in CRSwNP patients with asthma and allergic rhinitis compared to their NP counterparts. Elevated TRPV1 levels in comorbid asthma and allergy may have a function in CRSwNP. Subpopulation-specific TRPV1 presence on plasma and mast cells can indicate delicate roles in regulating activation and release of inflammatory mediators.

Expression and distribution of three transient receptor potential vanilloid(TRPV) channel proteins in human odontoblast-like cells.

Odontoblasts have been suggested to contribute to nociceptive sensation in the tooth via expression of the transient receptor potential (TRP) channels. The TRP channels as a family of nonselective cation permeable channels play an important role in sensory transduction of human. In this study, we examined the expression of transient receptor potential vanilloid-1 (TRPV1), transient receptor potential vanilloid-2 (TRPV2) and transient receptor potential vanilloid-3 (TRPV3) channels in native human odontoblasts (HODs) and long-term cultured human dental pulp cells with odontoblast phenotyoe (LHOPs) obtained from healthy wisdom teeth with the use of immunohistochemistry (IHC), immunofluorescence (IF), quantitative real-time polymerase chain reaction (qRT-PCR),western blotting (WB) and immunoelectron microscopy (IEM) assay. LHOPs samples were made into ultrathin sections, mounted on nickel grids, floated of three TRPV antibodies conjugated with 10nm colloidal gold particles and observed under IEM at 60,000 magnifications. The relative intracellular distributions of these three channels were analyzed quantitatively on IEM images using a robust sampling, stereological estimation and statistical evaluation method. The results of IHC and IF convinced that TRPV1, TRPV2 and TRPV3 channels were expressed in native HODs and (LHOPs). The result of qRT-PCR and WB confirmed that the gene and protein expression of TRPV1, TRPV2, and TRPV3 channels and TRPV1 mRNA are more abundantly expressed than TRPV2 and TRPV3 in HODs (P < 0.05). Quantitative analysis of IEM images showed that the relative intracellular distributions of these three channels are similar, and TRPV1, TRPV2 and TRPV3 proteins were preferential labeled in human odontoblast processes, mitochondria, and endoplasmic reticulum. Thus, HODs could play an important role in mediating pulp thermo-sensation due to the expression of these three TRPV channels. The difference of relative intracellular distributions of three channels suggests that special structures such as processes may have an important role to sensing of the outer stimuli first.

Local upregulation of transient receptor potential ankyrin 1 and transient receptor potential vanilloid 1 ion channels in rectosigmoid deep infiltrating endometriosis.

Transient Receptor Potential Vanilloid 1 (TRPV1) and Transient Receptor Potential Ankyrin 1 (TRPA1) expressed mainly by primary sensory neurons function as major nociceptive integrators. They are also present on the rat endometrium in an oestrogen-regulated manner. TRPV1 is upregulated in peritoneal and ovarian endometriosis patients, but there is no information about TRPA1 and their pathophysiological significances. In this study, patients undergoing laparoscopic surgery were investigated: severe dysmenorrhoea due to rectosigmoid deep infiltrating endometriosis (n = 15), uterine fibroid-induced moderate dysmenorrhoea (n = 7) and tubal infertility with no pain (n = 6). TRPA1 and TRPV1 mRNA and protein expressions were determined by quantitative polymerase chain reaction and semi-quantitative immunohistochemistry from the endometrium samples taken by curettage. Results were correlated with the clinical characteristics including pain intensity. TRPA1 and TRPV1 receptors were expressed in the healthy human endometrium at mRNA and protein levels. Sparse, scattered cytoplasmic TRPA1 and TRPV1 immunopositivities were found in the stroma and epithelial layers. We detected upregulated mRNA levels in deep infiltrating endometriosis lesions, and TRPV1 gene expression was also elevated in autocontrol endometrium of deep infiltrating endometriosis patients. Histological scoring revealed significant TRPA1 and TRPV1 difference between deep infiltrating endometriosis stroma and epithelium, and in deep infiltrating endometriosis epithelium compared to control samples. Besides, we measured elevated stromal TRPV1 immunopositivity in deep infiltrating endometriosis. Stromal TRPA1 and TRPV1 immunoreactivities strongly correlated with dysmenorrhoea severity, as well TRPV1 expression on ectopic epithelial cells and macrophages with dyspareunia. Epithelial TRPA1 and stromal TRPV1 immunopositivity also positively correlated with dyschezia severity. We provide the first evidence for the presence of non-neuronal TRPA1 receptor in the healthy human endometrium and confirm the expression of TRPV1 channels. Their upregulations in rectosigmoid deep infiltrating endometriosis lesions and correlations with pain intensity suggest potential roles in pathophysiological mechanisms of the disease.

Effects of a TRPV1 agonist capsaicin on respiratory rhythm generation in brainstem-spinal cord preparation from newborn rats.

The heat-sensitive transient receptor potential vanilloid 1 (TRPV1) channels are expressed in the peripheral and central nervous systems. However, there is no report on how the activation of TRPV1 causes the modulation of neuronal activity in the medullary respiratory center. We examined effects of capsaicin, a specific agonist of TRPV1 channels, on respiratory rhythm generation in brainstem-spinal cord preparation from newborn rats. Capsaicin induced a biphasic response in the respiratory rhythm (a transient decrease followed by an increase in the C4 rate). The second-phase excitatory effect (but not the initial inhibitory effect) in the biphasic response was partly blocked by capsazepine or AMG9810 (TRPV1 antagonists). Capsaicin caused strong desensitization. After its washout, the strength of C4 burst inspiratory activity was augmented once per four to five respiratory cycles. The preinspiratory and inspiratory neurons showed tonic firings due to membrane depolarization during the initial inhibitory phase. In the presence of TTX, capsaicin increased the fluctuation of the membrane potential of the CO2-sensitive preinspiratory neurons in the parafacial respiratory group (pFRG), accompanied by slight depolarization. The C4 inspiratory activity did not stop, even 60-90min after the application of 50/100μM capsaicin. Voltage-sensitive dye imaging demonstrated that the spatiotemporal pattern of the respiratory rhythm generating networks after application of capsaicin (50μM, 70-90min) was highly similar to the control. A histochemical analysis using TRPV1 channel protein antibodies and mRNA demonstrated that the TRPV1 channel-positive cells were widely distributed in the reticular formation of the medulla, including the pFRG. Our results showed that the application of capsaicin in the medulla has various influences on the respiratory center: transient inhibitory and subsequent excitatory effects on the respiratory rhythm and periodical augmentation of the inspiratory burst pattern. The effects of capsaicin were partially blocked by TRPV1 antagonists but could be also induced at least partially via the non-specific action. Our results also suggested a minor contribution of the TRPV1 channels to central chemoreception.

Estrogens Exacerbate Nociceptive Pain via Up-Regulation of TRPV1 and ANO1 in Trigeminal Primary Neurons of Female Rats.

Several trigeminal pain disorders show sex differences, and high levels of estrogens may underlie these differences. The interaction between transient receptor potential vanilloid 1 (TRPV1) and anoctamin 1 (ANO1) plays an important role in peripheral nociception. However, whether TRPV1 and ANO1 are involved in estrogen-modulated trigeminal pain sensitivity is unclear. In this study, we examined estradiol (E2) modulation of nociception through behavioral and immunohistological experiments after application of capsaicin (Cap), a selective TRPV1 agonist, onto the ocular surface in ovariectomized rats treated with high-dose E2 (HE) or low-dose E2 (LE) for 2 days. In addition, we used real-time PCR to study the effects of E2 on the expression levels of TRPV1 and ANO1 mRNA in trigeminal ganglia. In the behavioral experiment, the HE group showed significant potentiation of Cap-evoked nocifensive behavior compared with the LE group. Immunohistochemistry showed that Cap evoked a significantly greater number of cells that were immunoreactive for c-Fos, a marker of nociceptive activation, in the trigeminal subnucleus caudalis/upper cervical cord in the HE group than in the LE group. The number of c-Fos-immunoreactive cells in the ventral trigeminal interpolaris/caudalis were similar in the 2 groups. Real-time PCR showed that the levels of TRPV1 and ANO1 mRNA in the HE group were significantly higher than levels in the LE group. Thus, high levels of estrogens may be a risk factor for Cap-evoked nociceptive pain, and estrogen-dependent increases in TRPV1 and ANO1 are likely involved in modulating the nociceptive response in the trigeminal area.

Analysis of transient receptor potential vanilloid 1 and substance P gene expression in the mouse tongue following oral ginger administration

Ginger rhizome (Zingiber officinale) exhibits multiple pharmacological actions. For example, its pungent components target the transient receptor potential vanilloid 1 (TRPV1) ion channel and thus contribute to swallowing reflex recovery by elevating the neuropeptide substance P. However, the precise mechanism underlying this action remains unclear. To examine TRPV1 and substance P gene expression in the mouse tongue in response to stimulation by orally administered ginger, quantitative real-time polymerase chain reaction and immunohistochemistry were performed to evaluate mRNA and protein expression. TRPV1 mRNA expression in the mouse tongue was upregulated 30 min after oral ginger stimulation. In the gingerstimulated mouse, TRPV1 protein expression was increased and concentrated in the plasma membranes of the mucous glandular cells of the tongue epithelium. No significant differences in substance P mRNA expression relative to the control were observed after ginger stimulation. However, immunohistochemistry revealed that the amount of substance P protein expression increased in the mucous glandular cells of the tongue epithelium in ginger-stimulated mice, and this expression appeared to concentrate in the secretory granules of these cells. Activation of TRPV1 promotes the secretion of substance P in saliva, and clinically, saliva levels of substance P can be measured noninvasively and can provide a useful biomarker of the swallowing function. An increased level of substance P in the saliva could indicate improved dysphagia. Our data suggest that ginger activates TRPV1 and promotes the secretion of substance P in saliva. Ginger is therefore expected to serve as a functional agent for improving dysphagia.