Transient Receptor Potential Melastatin Research Articles

CD38/ADP-ribose/TRPM2-mediated nuclear Ca2+ signaling is essential for hepatic gluconeogenesis in fasting and diabetes

Hepatic glucose production by glucagon is crucial for glucose homeostasis during fasting, yet the underlying mechanisms remain incompletely delineated. Although CD38 has been detected in the nucleus, its function in this compartment is unknown. Here, we demonstrate that nuclear CD38 (nCD38) controls glucagon-induced gluconeogenesis in primary hepatocytes and liver in a manner distinct from CD38 occurring in the cytoplasm and lysosomal compartments. We found that the localization of CD38 in the nucleus is required for glucose production by glucagon and that nCD38 activation requires NAD+ supplied by PKCδ-phosphorylated connexin 43. In fasting and diabetes, nCD38 promotes sustained Ca2+ signals via transient receptor potential melastatin 2 (TRPM2) activation by ADP-ribose, which enhances the transcription of glucose-6 phosphatase and phosphoenolpyruvate carboxykinase 1. These findings shed light on the role of nCD38 in glucagon-induced gluconeogenesis and provide insight into nuclear Ca2+ signals that mediate the transcription of key genes in gluconeogenesis under physiological conditions.

Monitoring trafficking and expression of hemagglutinin-tagged transient receptor potential melastatin 4 channel in mammalian cells.

The TRPM4 gene encodes a Ca2+-activated monovalent cation channel called transient receptor potential melastatin 4 (TRPM4) that is expressed in various tissues. Dysregulation or abnormal expression of TRPM4 has been linked to a range of diseases. We introduced the hemagglutinin (HA) tag into the extracellular S6 loop of TRPM4, resulting in an HA-tagged version called TRPM4-HA. This TRPM4-HA was developed to investigate the purification, localization, and function of TRPM4 in different physiological and pathological conditions. TRPM4-HA was successfully expressed in the intact cell membrane and exhibited similar electrophysiological properties, such as the current-voltage relationship, rapid desensitization, and current size, compared to the wild-type TRPM4. The presence of the TRPM4 inhibitor 9-phenanthrol did not affect these properties. Furthermore, a wound-healing assay showed that TRPM4-HA induced cell proliferation and migration, similar to the native TRPM4. Co-expression of protein tyrosine phosphatase, non-receptor type 6 (PTPN6 or SHP-1) with TRPM4-HA led to the translocation of TRPM4-HA to the cytosol. To investigate the interaction between PTPN6 and tyrosine residues of TRPM4 in enhancing channel activity, we generated four mutants in which tyrosine (Y) residues were substituted with phenylalanine (F) at the N-terminus of TRPM4. The YF mutants displayed properties and functions similar to TRPM4-HA, except for the Y256F mutant, which showed resistance to 9-phenanthrol, suggesting that Y256 may be involved in the binding site for 9-phenanthrol. Overall, the creation of HA-tagged TRPM4 provides researchers with a valuable tool to study the role of TRPM4 in different conditions and its potential interactions with other proteins, such as PTPN6.

Metabolism of (R)-Praziquantel versus the Activation of a Parasite Transient Receptor Potential Melastatin Ion Channel.

Praziquantel (PZQ) is an essential anthelmintic drug recently established to be an activator of a Transient Receptor Potential Melastatin (TRPMPZQ ) ion channel in trematode worms. Bioinformatic, mutagenesis and drug metabolism work indicate that the cyclohexyl ring of PZQ is a key pharmacophore for activation of trematode TRPMPZQ , as well as serving as the primary site of oxidative metabolism which results in PZQ being a short-lived drug. Based on our recent findings, the hydrophobic cleft in schistosome TRPMPZQ defined by three hydrophobic residues surrounding the cyclohexyl ring has little tolerance for polarity. Here we evaluate the in vitro and in vivo activities of PZQ analogues with improved metabolic stability relative to the challenge of maintaining activity on the channel. Finally, an estimation of the respective contribution to the overall activity of both the parent and the main metabolite of PZQ in humans is reported.

Targeting temperature-sensitive transient receptor potential channels in hypertension: far beyond the perception of hot and cold.

Transient receptor potential (TRP) channels are nonselective cation channels and participate in various physiological roles. Thus, changes in TRP channel function or expression have been linked to several disorders. Among the many TRP channel subtypes, the TRP ankyrin type 1 (TRPA1), TRP melastatin type 8 (TRPM8), and TRP vanilloid type 1 (TRPV1) channels are temperature-sensitive and recognized as thermo-TRPs, which are expressed in the primary afferent nerve. Thermal stimuli are converted into neuronal activity. Several studies have described the expression of TRPA1, TRPM8, and TRPV1 in the cardiovascular system, where these channels can modulate physiological and pathological conditions, including hypertension. This review provides a complete understanding of the functional role of the opposing thermo-receptors TRPA1/TRPM8/TRPV1 in hypertension and a more comprehensive appreciation of TRPA1/TRPM8/TRPV1-dependent mechanisms involved in hypertension. These channels varied activation and inactivation have revealed a signaling pathway that may lead to innovative future treatment options for hypertension and correlated vascular diseases.

Assessing the Effects of the Topical Application of L-Menthol on Pain-Related Somatosensory-Evoked Potentials Using Intra-Epidermal Stimulation.

L-menthol is known to activate transient receptor potential melastatin 8 (TRPM8) and induce analgesia to thermal stimuli. However, since thermal stimulation leads to the interaction among the other TRP channels, it was unclear whether L-menthol causes analgesia to stimuli other than thermal stimuli. Therefore, we aimed to investigate whether activating TRPM8 via topical application of 10% menthol solution attenuates pain-related somatosensory-evoked potentials (pSEPs) and affects numerical rating scale (NRS) score using intra-epidermal electrical stimulation (IES). We applied 10% L-menthol or control solution on the dorsum of the right hand of 25 healthy participants. The pSEP and NRS, elicited by IES, and sensory threshold were measured before and after each solution was applied. The results showed that the topical application of 10% L-menthol solution significantly reduced N2-P2 amplitude in pSEPs compared with the control solution. Moreover, the N2 latency was significantly prolonged upon the topical application of L-menthol solution. NRS scores were similar under both conditions. These results suggest that topical application of L-menthol does not alter subjective sensation induced using IES, although it may attenuate afferent signals at free nerve endings even with stimuli that do not directly activate TRP channels.

Antagonist of transient receptor potential melastatin 2 suppresses mechanical hypersensitivity and activation of microglia induced by infraorbital nerve ligation in male rats

Activation of microglia is known to be involved in neuropathic pain. However, the pathway that regulates the microglial activation is not completely understood. Transient receptor potential (TRP) melastatin 2 (TRPM2), which is part of the TRP superfamily, is reportedly expressed on microglia and is suggested to be involved in neuropathic pain. To explore the effect of a TRPM2 antagonist on orofacial neuropathic pain and the relationship between TRPM2 and the activation of microglia, experiments were conducted using male rats that underwent infraorbital nerve (ION) ligation as orofacial neuropathic pain models. TRPM2 expression was detected on microglia in the trigeminal spinal subnucleus caudalis (Vc). The immunoreactivity of TRPM2 in the Vc increased after ION ligation. Mechanical threshold for head-withdrawal response was measured using von Frey filament, and it decreased after ION ligation. When the TRPM2 antagonist was administered to the ION-ligated rats, the low mechanical threshold for head-withdrawal response increased, and the number of phosphorylated extracellular signal-regulated kinase (pERK)-immunoreactive cells in the Vc decreased. The number of CD68-immunoreactive cells in the Vc also decreased after the administration of the TRPM2 antagonist in the ION-ligated rats. These findings suggest that TRPM2 antagonist administration suppresses hypersensitivity to mechanical stimulation induced by ION ligation and microglial activation, and TRPM2 is also involved in microglial activation in orofacial neuropathic pain.

Severity of Hepatocyte Damage and Prognosis in Cirrhotic Patients Correlate with Hepatocyte Magnesium Depletion.

We aimed to evaluate the magnesium content in human cirrhotic liver and its correlation with serum AST levels, expression of hepatocellular injury, and MELDNa prognostic score. In liver biopsies obtained at liver transplantation, we measured the magnesium content in liver tissue in 27 cirrhotic patients (CIRs) and 16 deceased donors with healthy liver (CTRLs) by atomic absorption spectrometry and within hepatocytes of 15 CIRs using synchrotron-based X-ray fluorescence microscopy. In 31 CIRs and 10 CTRLs, we evaluated the immunohistochemical expression in hepatocytes of the transient receptor potential melastatin 7 (TRPM7), a magnesium influx chanzyme also involved in inflammation. CIRs showed a lower hepatic magnesium content (117.2 (IQR 110.5-132.9) vs. 162.8 (IQR 155.9-169.8) μg/g; p < 0.001) and a higher percentage of TRPM7 positive hepatocytes (53.0 (IQR 36.8-62.0) vs. 20.7 (10.7-32.8)%; p < 0.001) than CTRLs. In CIRs, MELDNa and serum AST at transplant correlated: (a) inversely with the magnesium content both in liver tissue and hepatocytes; and (b) directly with the percentage of hepatocytes stained intensely for TRPM7. The latter also directly correlated with the worsening of MELDNa at transplant compared to waitlisting. Magnesium depletion and overexpression of its influx chanzyme TRPM7 in hepatocytes are associated with severity of hepatocyte injury and prognosis in cirrhosis. These data represent the pathophysiological basis for a possible beneficial effect of magnesium supplementation in cirrhotic patients.

TRPM7 mediates endoplasmic reticulum stress and ferroptosis in sepsis-induced myocardial injury.

Transient receptor potential melastatin 7 (TRPM7), a non-selective cation channel, was significantly upregulated in the blood of patients with sepsis. This study focuses on the preliminary exploration of the probable regulatory mechanism of TRPM7 in sepsis-induced myocardial injury (SIMI).HL-1 cardiac muscle cell line was treated with lipopolysaccharide (LPS) to mimic SIMI in vitro, and TRPM7 level was assessed. The impacts of TRPM7 knockdown on cellular inflammation response, oxidative stress, apoptosis, endoplasmic reticulum (ER) stress, and ferroptosis were identified. In order to explore the mechanism, ER stress agonist tunicamycin (TM) or ferroptosis inducer erastin was applied to treat HL-1 cells. The influences of TM and erastin on the aforementioned aspects were evaluated.TRPM7 was elevated in response to LPS stimulation, and its knockdown reduced the secretion of inflammatory factors and oxidative stress degree. Moreover, TRPM7 knockdown significantly suppressed cell apoptosis, ER stress, and ferroptosis. TM and erastin reversed the functions of TRPM7 knockdown, indicating ER stress and ferroptosis mediated in the regulation of TRPM7.This research proposes the possibility of TRPM7 as a marker or target for SIMI, and provides theoretical support for follow-up research.

Vasoprotective effects of NOX4 are mediated via polymerase and transient receptor potential melastatin 2 cation channels in endothelial cells.

NOX4 activation has been implicated to have vasoprotective and blood pressure (BP)-lowering effects. Molecular mechanisms underlying this are unclear, but NOX4-induced regulation of the redox-sensitive Ca2+ channel TRPM2 and effects on endothelial nitric oxide synthase (eNOS)-nitric oxide signalling may be important. Wild-type and LinA3, renin-expressing hypertensive mice, were crossed with NOX4 knockout mice. Vascular function was measured by myography. Generation of superoxide (O2-) and hydrogen peroxide (H2O2) were assessed by lucigenin and amplex red, respectively, and Ca2+ influx by Cal-520 fluorescence in rat aortic endothelial cells (RAEC). BP was increased in NOX4KO, LinA3 and LinA3/NOX4KO mice. This was associated with endothelial dysfunction and vascular remodelling, with exaggerated effects in NOX4KO groups. The TRPM2 activator, ADPR, improved vascular relaxation in LinA3/NOX4KO mice, an effect recapitulated by H2O2. Inhibition of PARP and TRPM2 with olaparib and 2-APB, respectively, recapitulated endothelial dysfunction in NOX4KO. In endothelial cells, Ang II increased H2O2 generation and Ca2+ influx, effects reduced by TRPM2 siRNA, TRPM2 inhibitors (8-br-cADPR, 2-APB), olaparib and GKT137831 (NOX4 inhibitor). Ang II-induced eNOS activation was blocked by NOX4 and TRPM2 siRNA, GKT137831, PEG-catalase and 8-br-cADPR. Our findings indicate that NOX4-induced H2O2 production activates PARP/TRPM2, Ca2+ influx, eNOS activation and nitric oxide release in endothelial cells. NOX4 deficiency impairs Ca2+ homeostasis leading to endothelial dysfunction, an effect exacerbated in hypertension. We define a novel pathway linking endothelial NOX4/H2O2 to eNOS/nitric oxide through PARP/TRPM2/Ca2+. This vasoprotective pathway is perturbed when NOX4 is downregulated and may have significance in conditions associated with endothelial dysfunction, including hypertension.

Biochemical aspects of magnesium-enhanced bone regeneration

Current research is focused on practical implications of magnesium-based implants largely due to their biodegradability and ability to promote bone healing and formation. However, the mechanism underlying the osteogenesis regulation by magnesium is still unclear.We describe cellular and molecular mechanisms underlying the effect of magnesium ions (Mg2+) on bone growth following the device implantation. The presented data demonstrate magnesium-induced activation of canonical Wnt/β-catenin signaling pathway in human bone marrow stromal cells resulting in their differentiation into osteoblasts, osteogenic effect and recovery of bone defects. We describe the role of the molecular mechanisms responsible for osteopromotive properties of Mg2+ and associated with unique transient receptor potential melastatin 7 (TRPM7) cation channels mediating the Mg2+ influx. TRPM7-mediated Mg2+ influx is important for platelet-derived growth factor (PDGF)-induced proliferation, adhesion, and migration of human osteoblasts, as well as for promotion of Mg2+-associated bone regeneration.We discuss the effect of Mg2+ on intracellular signaling processes, expression of the vascular endothelial growth factor (VEGF), hypoxia-inducible factor-2α, and peroxisome proliferator-activated receptor-γ coactivator 1α. Mg2+ can promote bone regeneration by enhancing the production of type X collagen and VEGF by osteogenic cells in bone marrow.

TRPM7 is Involved in the Regulation of Proliferation, Migration and Osteogenic Differentiation of Human Dental Follicle Cells.

Dental follicle cells (DFCs) are promising candidates for tissue engineering. However, the molecular mechanisms that regulate the biological characteristics of DFCs are still unclear. Transient receptor potential melastatin 7 (TRPM7) is a Ca2+- and Mg2+-permeable cation channel. The aim of this study was to determine the impact of TRPM7 on the proliferation, migration and osteogenic differentiation of DFCs. PCR, Western blotting, Immunocytochemical staining and Patch clamp methods were used to identify the gene and protein expression of TRPM7 in DFCs. DFCs were infected with lentiviruses that expressed either TRPM7 specific shRNA or scrambled non-effective shRNA to investigate its functional role. Cell proliferation and migration were assessed using Cell Counting Kit-8 assays and transwell cell culture chambers separately. Cell osteogenic differentiation were determined by ALP assay kit and Alizarin Red staining. Gene and protein expression of TRPM7 were detected in DFCs, but not of TRPM6, which is a closely related channel with similar function. In the absence of Mg2+, typical whole cell TRPM7-like currents were recorded by patch clamp. These were inhibited by low concentrations of 2-APB, but activated by high concentrations of 2-APB. Functional studies demonstrated that suppression of TRPM7 expression inhibited the proliferation and migration of DFCs, and promoted their osteogenic differentiation. Furthermore, Mg2+ deficiency mimicked the effects of TRPM7 knockdown in terms of osteogenic differentiation of DFCs. These results demonstrate that TRPM7 is involved in regulating the proliferation, migration and osteogenic differentiation of DFCs.

TRPM4 contribution in mouse uterine contractions.

Inappropriate uterine contractions are a matter of concern during pregnancy or menses. We identified the transient receptor potential melastatin 4 (TRPM4) ion channel as a new actor in mouse uterine contractions highlighting this protein as a potential pharmacological target for a better control of myometrial activity. Control of uterine contractions is of interest in the context of inappropriate myometrial activity during pregnancy and at time of delivery, but it is also a matter for menstrual pain. While several molecular determinants of myometrial contractions have been described, the complete distribution of roles to the various actors is far from understood. A key phenomenon is a variation in cytoplasmic Ca2+ which leads to the activation of calmodulin in smooth muscle and also in the phosphorylation of myosin allowing contraction. The Ca2+ - TRPM4 channel which is known to modulate Ca2+- fluxes in several cell types was shown to participate in vascular as well as detrusor muscle contraction. We thus designed a study to determine whether it also participates in myometrial contraction. Uterine rings were isolated from Trpm4+/+ and Trpm4-/- non-pregnant adult mice and contractions were recorded using an isometric force transducer. In basal conditions, spontaneous contractions were similar in both groups. Application of 9-phenanthrol, a pharmacological TRPM4 inhibitor, dose-dependently reduced contraction parameters in Trpm4+/+ rings with an IC50 around 2.10-6 mol/L. The effect of 9-phenanthrol was significantly reduced in Trpm4-/- rings. The effect of oxytocin was tested and was found to be stronger in Trpm4+/+ rings compared to Trpm4-/-. Under a constant stimulation by oxytocin, 9-phenanthrol still reduced contraction parameters in Trpm4+/+ rings with a smaller effect on Trpm4-/-. Altogether it indicates that TRPM4 participates in uterine contractions in mice and may thus be evaluated as a new target to control such contractions.

The Effects of N-acetylcysteine on Transient Receptor Potential Melastatin 2 Channels Activation and Expression in Testicular Tissue of Diabetic Rats.

Diabetes mellitus (DM) is a common, chronic metabolic disease that has harmful effects on many diverse tissues, including the testis. One of the ways of tissue damage is the modification of transient receptor potential melastatin 2 (TRPM2) channels by increased reactive oxygen species (ROS). In our study for the first time, it was aimed to investigate TRPM2 channel activation in testicular tissues of diabetic rats induced by streptozotosin (STZ) and to examine the efficacy of N-acetylcysteine (NAC) treatment, which is an antioxidant. In our study, 28 Wistar albino male rats aged 8-10 weeks were used, and animals were divided into four groups: control group, NAC group, DM group, and DM + NAC group. The experimental phase was designed as eight weeks. The malondialdehyde(MDA)level, which is an indicator for lipid peroxidation due to oxidative stress, was measured by the spectrophotometric method. The Tunel assay was used to determine apoptosis on testicular tissue. TRPM2 immunoreactivity was determined by the avidin-biotin-peroxidase complex method, and quantitative polymerase chain reaction (QPCR) was used to determine TRPM2 expression levels. It was seen that MDA levels were significantly increased in the DM group and decreased after NAC treatment. Similarly, it was observed that apoptosis levels, which increased significantly in diabetic rats, decreased to the levels of the control group after treatment. It was seen that TRPM2 activation and expression levels were significantly decreased in the DM group. The results of this study show that NAC regulates TRPM2 activation in the testicular tissue of patients with diabetes and has tissue-protective properties.

Carvacrol enhances anti-tumor activity and mitigates cardiotoxicity of sorafenib in thioacetamide-induced hepatocellular carcinoma model through inhibiting TRPM7

AimsSorafenib (Sora) represents one of the few effective drugs for the treatment of advanced hepatocellular carcinoma (HCC), while resistance and cardiotoxicity limit its therapeutic efficacy. This study investigated the effect of transient receptor potential melastatin 7 (TRPM7) inhibitor, carvacrol (CARV), on overcoming Sora resistance and cardiotoxicity in thioacetamide (TAA) induced HCC in rats. Materials and methodsTAA (200 mg/kg/twice weekly, intraperitoneal) was administered for 16 weeks to induce HCC. Rats were treated with Sora (10 mg/Kg/day; orally) and CARV (15 mg/kg/day; orally) alone or in combination, for six weeks after HCC induction. Liver and heart functions, antioxidant capacity, and histopathology were performed. Apoptosis, proliferation, angiogenesis, metastasis, and drug resistance were assessed by quantitative real time polymerase chain reaction, enzyme-linked immunosorbent assay, and immunohistochemistry. Key findingsCARV/Sora combination significantly improved survival rate, and liver functions, reduced Alpha-Fetoprotein level, and attenuated HCC progression compared with Sora group. CARV coadministration almost obviated Sora-induced changes in cardiac and hepatic tissues. The CARV/Sora combination suppressed drug resistance and stemness by downregulating ATP-binding cassette subfamily G member 2, NOTCH1, Spalt like transcription factor 4, and CD133. CARV boosted Sora antiproliferative and apoptotic activities by decreasing cyclin D1 and B-cell leukemia/lymphoma 2 and increasing BCL2-Associated X and caspase-3. SignificanceCARV/Sora is a promising combination for tumor suppression and overcoming Sora resistance and cardiotoxicity in HCC by modulating TRPM7. To our best knowledge, this study represents the first study to investigate the efficiency of CARV/ Sora on the HCC rat model. Moreover, no previous studies have reported the effect of inhibiting TRPM7 on HCC.

TRPM4: New actor in osteogenic differentiation of human valvular interstitial cells

Transient receptor potential melastatin 4 (TRPM4) protein, a non-selective cation channel activated by calcium, is involved in cardiac fibroblasts remodeling. Its expression was reported in human valvular interstitial cells (VIC) with similar properties than fibroblasts, and its protein expression is increased on calcified aortic valves from patients suffering of aortic stenosis. VIC osteogenic differentiation is a key element in valvular calcification, which is, at term, responsible of aortic stenosis. The purpose of this study is to evaluate the implication of TRPM4 in human VIC osteogenic differentiation. VIC are isolated by enzymatic digestion from human aortic valves obtained from patients undergoing surgical aortic valve replacement. They are maintained 14 days in procalcifying media (2.7 mM calcium/2.5 mM phosphate), with or without 9-phenanthrol, a pharmacological TRPM4 inhibitor. Channel expression and functionality were followed respectively by PCR, Western Blot and electrophysiological recordings in the inside-out configuration of the patch clamp technique. Osteogenic differentiation of VIC was evaluated by measuring hydroxyapatite crystals by Alizarin red staining and by monitoring the protein expression of the osteogenic markers, BMP2 and RUNX2. A typical TRPM4 current was detected on VIC: its current-voltage relationship was linear with a conductance of 20.3 pS; the channel exhibited similar permeability for Na+ and K+; it was more activated in positive voltages and with increasing concentrations of internal Ca2+; the current was dose-dependently inhibited by 9-phenanthrol. TRPM4 mRNA and protein expression on VIC was confirmed by PCR and Western Blot. When maintained in procalcifying media, VIC developed hydroxyapatite crystals. Application of 9-phenanthrol (3.10-6 M) reduced mineralization by 30% without affecting cell viability. Moreover, it significantly reduced BMP2 and RUNX2 expression. Thereby, TRPM4 participates in osteogenic differentiation of human VIC in culture. Next studies of our unit are to evaluate whether TRPM4 repression by shRNA confirms those results.

TRPM2 regulates cell cycle through the Ca2+-CaM-CaMKII signaling pathway to promote HCC.

HCC is one of the most common causes of cancer-related deaths. Transient receptor potential melastatin 2 (TRPM2), a Ca2+-permeable cation channel, was reported to be involved in carcinogenesis and tumor growth recently. However, whether TRPM2 is involved in the pathogenesis and progression of HCC remains unclear. Herein, we systematically elucidated the functional role of TRPM2 in HCC cell cycle regulation and proliferation. We determine TRPM2 expression to be strongly upregulated in the tumor tissues of HCC patients and associated with a negative prognosis. TRPM2 is highly expressed in HCC cell lines Huh-7 and HepG2 cells, rather than in normal hepatocytes. Inhibition or silencing of TRPM2, or inhibition of the downstream Ca2+-CaM-CaMKII signaling pathway, significantly suppressed the proliferation of Huh-7 and HepG2 cells by arresting the cell cycle at the G1/S phase, accompanied with reduced expression of G1/S checkpoint proteins. Importantly, inhibition or depletion of TRPM2 remarkably slowed down the growth of patient-derived xenografts and Huh-7 xenografts in mice. Our results indicate that TRPM2 promotes HCC cell proliferation via activating the Ca2+-CaM-CaMKII signaling pathway to induce the expression of the key G1/S regulatory proteins and accelerate the cell cycle. This study provides compelling evidence of TRPM2 involvement in a previously unrecognized mechanism that drives HCC progression and demonstrates that TRPM2 is a potential target for HCC treatment.

Curcumin attenuates hydroxychloroquine-mediated apoptosis and oxidative stress via the inhibition of TRPM2 channel signalling pathways in a retinal pigment epithelium cell line.

Hydroxychloroquine (HCQ) is used in the treatment of several diseases, such as malaria, Sjögren's disease, Covid-19, and rheumatoid arthritis. However, HCQ induces retinal pigment epithelium death via the excessive increase of cytosolic (cROS) and mitochondrial (mROS) free oxygen radical production. The transient receptor potential melastatin 2 (TRPM2) cation channel is stimulated by ADP-ribose (ADPR), cROS, and mROS, although it is inhibited by curcumin (CRC). We aimed to investigate the modulating action of CRC on HCQ-induced TRPM2 stimulation, cROS, mROS, apoptosis, and death in an adult retinal pigment epithelial 19 (ARPE19) cell line model. ARPE19 cells were divided into four groups: control (CNT), CRC (5µM for 24h), HCQ (60µM for 48h), and CRC + HCQ groups. The levels of cell death (propidium iodide positive cell numbers), apoptosis markers (caspases -3, -8, and -9), oxidative stress (cROS and mROS), mitochondria membrane depolarization, TRPM2 current density, and intracellular free Ca2+ and Zn2+ fluorescence intensity were upregulated in the HCQ group after stimulation with hydrogen peroxide and ADPR, but their levels were downregulated by treatments with CRC and TRPM2 blockers (ACA and carvacrol). The HCQ-induced decrease in retinal live cell count and cell viability was counteracted by treatment with CRC. HCQ-mediated overload Ca2+ influx and retinal oxidative toxicity were induced in an ARPE19 cell line through the stimulation of TRPM2, although they were attenuated by treatment with CRC. Hence, CRC may be a potential therapeutic antioxidant for TRPM2 activation and HCQ treatment-induced retinal oxidative injury and apoptosis.

Abstract 5277: Activation of TRPM8 channel suppresses prostate cancer growth and progression

Abstract Prostate cancer (PC) is one of the most prevalent male malignancies and a leading cause of cancer-related deaths in men. While androgen-deprivation therapy is successful in the early stages of PC, tumor cells eventually become resistant to its effects. With a median survival of 18 months, the shift to androgen-independent prostate cancer (AIPC) has a poor prognosis. Mechanisms causing this change have yet to be explained, however mounting evidence implicates the loss of transient receptor potential melastatin 8 (TRPM8) as a significant contributor. The prostate epithelium normally expresses the ionotropic receptor TRPM8. Our team has recently shown that testosterone-induced activation of TRPM8 enhances Ca2+ absorption and causes apoptosis. This prompted us to propose that increased TRPM8 activity on the plasma membrane is cytotoxic to PC cells and that TRPM8 internalization is a crucial step in the pathogenesis of PC. We examined the amounts of TRPM8 mRNA in benign tumor and metastatic PC patient datasets. TRPM8 mRNA is initially elevated in the early stages of PC but is increasingly lost during the progression to AIPC. In addition, male and female TRPM8−/− mice exhibited increased serum testosterone levels, heightened AR activity, and activation of cell cycle, invasion, and adhesion-related effectors. In both AR+ and AR− xenograft models, our research reveals that TRPM8 possesses potent antitumor properties. Given this, investigating the in vivo role of TRPM8 in PC has the potential to significantly enhance patient outcomes by preventing progression to the androgen-independent state. Citation Format: Swapna Asuthkar. Activation of TRPM8 channel suppresses prostate cancer growth and progression. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5277.

Lianmei Qiwu Decoction relieves diabetic cardiac autonomic neuropathy by regulating AMPK/TrkA/TRPM7 signaling pathway

This study investigated the effect of Lianmei Qiwu Decoction(LMQWD) on the improvement of cardiac autonomic nerve remodeling in the diabetic rat model induced by the high-fat diet and explored the underlying mechanism of LMQWD through the AMP-activated protein kinase(AMPK)/tropomyosin receptor kinase A(TrkA)/transient receptor potential melastatin 7(TRPM7) signaling pathway. The diabetic rats were randomly divided into a model group, an LMQWD group, an AMPK agonist group, an unloaded TRPM7 adenovirus group(TRPM7-N), an overexpressed TRPM7 adenovirus group(TRPM7), an LMQWD + unloaded TRPM7 adenovirus group(LMQWD+TRPM7-N), an LMQWD + overexpressed TRPM7 adenovirus group(LMQWD+TRPM7), and a TRPM7 channel inhibitor group(TRPM7 inhibitor). After four weeks of treatment, programmed electrical stimulation(PES) was employed to detect the arrhythmia susceptibility of rats. The myocardial cell structure and myocardial tissue fibrosis of myocardial and ganglion samples in diabetic rats were observed by hematoxylin-eosin(HE) staining and Masson staining. The immunohistochemistry, immunofluorescence, real-time quantitative polymerase chain reaction(RT-PCR), and Western blot were adopted to detect the distribution and expression of TRPM7, tyrosine hydroxylase(TH), choline acetyltransferase(ChAT), growth associated protein-43(GAP-43), nerve growth factor(NGF), p-AMPK/AMPK, and other genes and related neural markers. The results showed that LMQWD could significantly reduce the arrhythmia susceptibility and the degree of fibrosis in myocardial tissues, decrease the levels of TH, ChAT, and GAP-43 in the myocardium and ganglion, increase NGF, inhibit the expression of TRPM7, and up-regulate p-AMPK/AMPK and p-TrkA/TrkA levels. This study indicated that LMQWD could attenuate cardiac autonomic nerve remodeling in the diabetic state, and its mechanism was associated with the activation of AMPK, further phosphorylation of TrkA, and inhibition of TRPM7 expression.

MITOCHONDRIA: the dual function of the transient receptor potential melastatin 2 channels from cytomembrane to mitochondria.

Mitochondria are closely related to oxidative stress and play an important role in maintaining cell functional homeostasis and meeting cell energy demand. The transient receptor potential melastatin 2 (TRPM2) channel affects the occurrence and progression of diseases by regulating mitochondrial function. TRPM2 channel promotes Ca2+ influx to affect 18kDa translocator protein (TSPO), mitochondrial membrane potential (MMP), reactive oxygen species (ROS), adenosine triphosphate (ATP) production, and mitochondrial autophagy. The mechanism of Ca2+ influx into the mitochondria by TRPM2 is abundant. Interestingly, the TRPM2 channel inhibits the production of mitochondrial ROS in cancer cells and promotes the production of mitochondrial ROS in normal cells, which induces cell death in normal cells but proliferation in cancer cells. TRPM2 can be a potential target for the treatment of various diseases due to its role as a molecular link between mitochondria and Ca2+ signals.