Receptor Potential Cation Channel Subfamily Research Articles

Curcumin suppresses cell proliferation and reduces cholesterol absorption in Caco-2 cells by activating the TRPA1 channel

BackgroundCurcumin (Cur) is a bioactive dietary polyphenol of turmeric with various biological activities against several cancers. Colorectal cancer (CRC) is one of the leading causes of cancer-related deaths. Intestinal cholesterol homeostasis is associated with CRC. Chemotherapy for CRC is related to varied adverse effects. Therefore, natural products with anti-cancer properties represent a potential strategy for primary prevention of CRC.MethodsThe present study used Cur as a therapeutic approach against CRC using the Caco-2 cell line. The cells were treated with different concentrations of Cur for different duration of time and then the proliferation ability of cells was assessed using Cell Counting Kit-8 and 5-Ethynyl-2′-deoxyuridine assays. Oil red O staining and cholesterol assay kit were used to evaluate cellular lipid content and cholesterol outward transportation. Finally, the protein expressions of cholesterol transport-related protein and signal transduction molecules were assessed using Western blot assay.ResultsCur inhibited cell proliferation in Caco-2 cells in a dose- and time-dependent manner by activating the transient receptor potential cation channel subfamily A member 1 (TRPA1) channel. Activation of the TRPA1 channel led to increased intracellular calcium, peroxisome proliferator-activated receptor gamma (PPARγ) upregulation, and the subsequent downregulation of the specificity protein-1 (SP-1)/sterol regulatory element-binding protein-2 (SREBP-2)/Niemann-Pick C1-like 1 (NPC1L1) signaling pathway-related proteins, and finally reduced cholesterol absorption in Caco-2 cells.ConclusionsCur inhibits cell proliferation and reduces cholesterol absorption in Caco-2 cells through the Ca2+/PPARγ/SP-1/SREBP-2/NPC1L1 signaling by activating the TRPA1 channel, suggesting that Cur can be used as a dietary supplement for the primary prevention of CRC.Graphical In Caco-2 cells, Cur first stimulates calcium influx by activating the TRPA1 channel, further upregulates PPARγ and downregulates SP-1/SREBP-2/NPC1L1 signaling pathway, and finally inhibits the absorption of cholesterol. TRPA1, transient receptor potential cation channel subfamily A member 1; NPC1L1, Niemann-Pick C1-like 1; PPARγ, peroxisome proliferator-activated receptor gamma; SP-1, specificity protein-1; SREBP-2, sterol regulatory element-binding protein-2; Cur, curcumin.

Phosphate, calcium, and vitamin D signaling, transport, and metabolism in the endometria of cyclic ewes

BackgroundRecent evidence suggests important roles for progesterone (P4) and interferon tau in the regulation of calcium, phosphate, and vitamin D signaling in the uteri of pregnant sheep. However, the effects of P4 and estradiol (E2), with respect to the expression of their receptors PGR and ESR1, respectively, in uterine epithelia on mineral signaling during the estrous cycle has not been investigated. Estrous cycles of mature Suffolk ewes were synchronized, prostaglandin F2α was administered, and ewes were observed for estrus (designated as Day 0) in the presence of vasectomized rams. On Days 1, 9, or 14 of the estrous cycle, hysterectomies were performed.Results25-hydroxyvitamin D was more abundant in plasma from ewes on Day 14 than Day 1 (P < 0.05). Expression of fibroblast growth factor receptor 2 (FGFR2), a disintegrin and metalloprotease 17 (ADAM17), and parathyroid hormone-related protein (PTHrP) mRNAs was greater in endometria on Day 9 compared to Days 1 and 14 (P < 0.01). Similarly, expression of transient receptor potential cation channel subfamily V member 6 (TRPV6) mRNA was greater in endometria on Day 9 than Day 1 (P < 0.05). ATPase plasma membrane Ca2+ transporting 4 (ATP2B4) and S100 calcium binding protein G (S100G) mRNA expression was greater in endometria on Day 14 than on Days 1 and 9 (P < 0.01). In contrast, endometrial expression of vitamin D receptor (VDR) mRNA was lower on Days 9 and 14 than Day 1 (P < 0.01). Expression of klotho (KL) (P < 0.05) and cytochrome P450 family 24 subfamily A member 1 (CYP24) (P < 0.01) mRNAs was lower on Day 14 than Days 1 and 9. ADAM17, FGF23, CYP2R1, CYP27B1, KL, and VDR proteins immunolocalized to the uterine myometrium, blood vessels, and uterine luminal (LE), superficial glandular (sGE), and glandular (GE) epithelia. S100A9 protein was weakly expressed in the uterine myometrium, LE, sGE, and GE. Immunoreactivity of CYP2R1 and KL proteins in uterine LE and sGE was less on Day 1 than on Days 9 and 14. In contrast, S100G protein was expressed exclusively by GE, and immunoreactive S100G protein was less on Day 9. S100A12 protein localized to stromal cells of the uterine stratum spongiosum and blood vessels, but not by uterine epithelial cells.ConclusionCollectively, these results implicate E2, P4, and PGR in the regulation of phosphate, calcium, and vitamin D signaling in cyclic ewes.

Analysis of Genome Wide Associated Variants rs2651899 (PRDM16), rs11172113 (LRP1), and rs10166942 (TRPM8) of Migraine in the Indian Episodic Migraineurs

Background: The involvement of genes in migraine pain is identified by several studies. Many genome-wide association studies (GWAS) revealed the presence of particular variants in different migraine populations worldwide. Objective: This study aims in analyzing the presence of 3 GWAS variants (rs2651899 [PRDM16], rs11172113 [LRP1], and rs10166942 Transient Receptor Potential Cation Channel Subfamily M Member 8 [TRPM8]) in the Indian episodic migraineurs. Methodology: Enrolled 200 age- and gender-matched patients and control volunteers; collected blood samples to isolate DNA to check the presence of chosen variants in them. The results obtained were statistically analyzed using SPSS version 26.0. Results: The rs10166942 (TRPM8) variant is observed in 1.5% of control, 15.88% of migraine with aura, and 5.37% of migraine without aura subjects with the highly significant P &lt; 0.0008. The other two variants are absent in the chosen sample group. Conclusion: We reported the presence of rs10166942 in the Indian episodic migraineurs and we recommend pathway analysis for confirming its association with the migraine pain progression.

TRPV4 Promotes Metastasis in Melanoma by Regulating Cell Motility through Cytoskeletal Rearrangement

The abnormal expression of Transient Receptor Potential cation channel subfamily V member 4 (TRPV4) is closely related to the progression of multiple tumors. In addition, TRPV4 is increasingly being considered a potential target for cancer therapy, especially in tumor metastasis prevention. However, the biological correlation between TRPV4 and tumor metastasis, as well as the specific role of TRPV4 in malignant melanoma metastasis, is poorly understood. In this study, we aimed to examine the role of TRPV4 in melanoma metastasis through experiments and clinical data analysis, and the underlying anticancer mechanism of Baicalin, a natural compound, and its inhibitory effect on TRPV4 with in vivo and in vitro experiments. Our findings suggested that TRPV4 promotes metastasis in melanoma by regulating cell motility via rearranging the cytoskeletal, and Baicalin can inhibit cancer metastasis, whose mechanisms reverse the recruitment of activated cofilin to leading-edge protrusion and the increasing phosphorylation level of cortactin, which is provoked by TRPV4 activation.

AIBP regulates TRPV1 activation in chemotherapy-induced peripheral neuropathy by controlling lipid raft dynamics and proximity to TLR4 in dorsal root ganglion neurons.

Nociceptive afferent signaling evoked by inflammation and nerve injury is mediated by the opening of ligand-gated and voltage-gated receptors or channels localized to cholesterol-rich lipid raft membrane domains. Dorsal root ganglion (DRG) nociceptors express high levels of toll-like receptor 4 (TLR4), which also localize to lipid rafts. Genetic deletion or pharmacologic blocking of TLR4 diminishes pain associated with chemotherapy-induced peripheral neuropathy (CIPN). In DRGs of mice with paclitaxel-induced CIPN, we analyzed DRG neuronal lipid rafts, expression of TLR4, activation of transient receptor potential cation channel subfamily V member 1 (TRPV1), and TLR4-TRPV1 interaction. Using proximity ligation assay, flow cytometry, and whole-mount DRG microscopy, we found that CIPN increased DRG neuronal lipid rafts and TLR4 expression. These effects were reversed by intrathecal injection of apolipoprotein A-I binding protein (AIBP), a protein that binds to TLR4 and specifically targets cholesterol depletion from TLR4-expressing cells. Chemotherapy-induced peripheral neuropathy increased TRPV1 phosphorylation, localization to neuronal lipid rafts, and proximity to TLR4. These effects were also reversed by AIBP treatment. Regulation of TRPV1-TLR4 interactions and their associated lipid rafts by AIBP covaried with the enduring reversal of mechanical allodynia otherwise observed in CIPN. In addition, AIBP reduced intracellular calcium in response to the TRPV1 agonist capsaicin, which was increased in DRG neurons from paclitaxel-treated mice and in the naïve mouse DRG neurons incubated in vitro with paclitaxel. Together, these results suggest that the assembly of nociceptive and inflammatory receptors in the environment of lipid rafts regulates nociceptive signaling in DRG neurons and that AIBP can control lipid raft-associated nociceptive processing.

ANGPTL4 promotes nephrotic syndrome by downregulating podocyte expression of ACTN4 and podocin

Backgroundlipopolysaccharide (LPS) can induce nephrotic syndrome-like features such as massive proteinuria, hyperlipidemia, and fusion of glomerular podocytes with foot processes (FPs) in mice. Angiopoietin-like protein 4 (ANGPTL4) neutralized the negative charge of glomerular basement membrane charge and aggravated renal injury. The mechanism of ANGPTL4 aggravating podocyte injury has not been well clarified. In this study, we aimed to investigate the potential role of ANGPTL4 on podocyte FPs fusion and podocyte signal molecules. MethodsWe built angptl4 gene knocked out in C57BL6 mice using CRISPR/Cas9 technique. Nephrotic model was built by LPS in wild type and angptl4−/− mice. Expression of ACTN4, podocin and TRPC6 in the glomerulus were determined by immunohistochemistry. ResultsIn physical condition, the wild type and angptl4−/− mice showed no significant differences in biochemical indicators and kidney pathology. But in nephrotic condition, compared with wild type mice hyperlipidemia and proteinuria with the angptl4−/− mice was significantly relieved. Moreover, the degree of FPs fusion was notably improved in the nephrotic mice knocked out angptl4 gene. Expression of ACTN4 and podocin decreased drastically in the glomerulus of wild-type nephrotic mice. Different from wild-type, the ACTN4 and podocin expression showed slight weakening in angptl4−/− nephrotic mice. As transient receptor potential cation channel subfamily member, TRPC6 expression had no visible change in glomerulus of each group. ConclusionsANGPTL4 induces hyperlipidemia and podocyte injury in nephrotic mice, thereby promoting the formation of proteinuria. Its molecular mechanism may be related to ANGPTL4 down-regulating actin cytoskeletal regulatory signals ACTN4 and podocin.

Abstract 9376: TRPM77 Knockdown Prevents Hypomagnesemia-Induced Oxidative Stress and Cardiac Diastolic Dysfunction

Introduction: Our previous study has shown that 6-week low-Mg diet-induced hypomagnesemia results in mitochondrial dysfunction, cardiac diastolic dysfunction, and seizure-related death. Transient receptor potential cation channel subfamily M 7 (TRPM7) is a Mg transporter with both channel and kinase function located in the plasma membrane. We investigated the role of TRPM7 in hypomagnesemia-associated changes. Methods: For cardiac-specific knockdown of TRPM7, pAAV[miR30]-cTnT&gt;EGFP:Scramble-shRNA as control (Con) and pAAV[miR30]-cTnT&gt;EGFP:TRPM7 shRNA as TRPM7 knockdown (T7KD) were injected into mice through the jugular vein at 10 weeks old. One week later, mice were fed with a normal diet (nlMg, 2000 mg/kg Mg) or a low-Mg diet (HypoMg, 15-30 mg/kg Mg) for 4 weeks. Results: TRPM7 was increased significantly in wild type mouse hearts under the low-Mg diet (1.45±0.18-fold of mice with normal diet, P&lt;0.01). We therefore knocked down TRPM7 in the mouse heart to see if HypoMg-induced changes could be prevented. After 4 weeks of low-Mg diet, both Con and T7KD mice had hypomagnesemia with significantly decreased serum Mg levels (0.37±0.03 and 0.45±0.10 mM, respectively, vs 1.05±0.04 mM of Con-nlMg). All T7KD mice showed significantly decreased TRPM7 protein levels in the heart tissue (&lt;30% of Con-nlMg, P&lt;0.01). Cardiac diastolic dysfunction (E/e’) was prevented in T7KD-HypoMg mice (E/e’=16.2±1.9 vs. 20.7±1.5 of Con-HypoMg). Increased total protein oxidation (measured with nitro-tyrosine immunoprecipitation) in Con-HypoMg heart tissue (1.61±0.09-fold increase, P=0.045 vs. Con-nlMg) was normalized in the T7KD-HypoMg group (1.18±0.09-fold of Con-nlMg, P=0.45 vs. Con-nlMg). Conclusions: TRPM7 was upregulated in hypomagnesemia in hearts and its knockdown prevented hypomagnesemia-induced cardiac diastolic dysfunction and increased oxidative stress.

The discovery of (1R, 3R)-1-(3-chloro-5-fluorophenyl)-3-(hydroxymethyl)-1,2,3,4-tetrahydroisoquinoline-6-carbonitrile, a potent and selective agonist of human transient receptor potential cation channel subfamily m member 5 (TRPM5) and evaluation of as a potential gastrointestinal prokinetic agent

This publication details the discovery of a series of selective transient receptor potential cation channel subfamily M member 5 (TRPM5) agonists culminating with the identification of the lead compound (1R, 3R)-1-(3-chloro-5-fluorophenyl)-3-(hydroxymethyl)-1,2,3,4-tetrahydroisoquinoline-6-carbonitrile (39). We describe herein our biological rationale for agonism of the target, the examination of the then current literature tool molecules, and finally the process of our discovery starting with a high throughput screening hit through lead development. We also detail the selectivity of the lead compound 39 versus related family members TRPA1, TRPV1, TRPV4, TRPM4 and TRPM8, the drug metabolism and pharmacokinetics (DMPK) profile and in vivo efficacy in a mouse model of gastrointestinal motility.

The effects of transient receptor potential cation channel inhibition by BI 1358894 on cortico-limbic brain reactivity to negative emotional stimuli in major depressive disorder

Abnormal emotional processing in major depressive disorder (MDD) has been associated with increased activation to negative stimuli in cortico-limbic brain regions. The authors investigated whether treatment with BI 1358894, a small-molecule inhibitor of the transient receptor potential cation channel subfamily C leads to attenuated activity in these areas in MDD patients. 73 MDD patients were randomized to receive a single oral dose of BI 1358894 (100 mg), citalopram (20 mg), or matching placebo. Brain responses to emotional faces and scenes were investigated using functional magnetic resonance imaging. Primary endpoints were BOLD signal changes in response to negative faces in cortico-limbic brain regions, i.e. bilateral amygdala (AMY), dorsolateral prefrontal cortex, anterior insula (AI), and anterior cingulate cortex. Secondary endpoints were BOLD signal changes in response to negative scenes. For each region, separate ANOVA models were computed for the comparison of treatments (BI 1358894 or citalopram) vs. placebo. The adjusted treatment differences in the % BOLD signal changes in the faces task showed that BI 1358894 induced signal reduction in bilateral AMY and left AI. In the scenes task, BI 1358894 demonstrated significant signal reduction in bilateral AMY, AI, anterior cingulate cortex and left dorsolateral prefrontal cortex. Citalopram failed to induce any significant reductions in BOLD signal in both tasks. BI 1358894-mediated inhibition of the transient receptor potential cation channel subfamily resulted in strong signal reduction in cortico-limbic brain regions, thereby supporting development of this mechanism of action for MDD patients.

Transient Ca2+ entry by plasmalogen-mediated activation of receptor potential cation channel promotes AMPK activity.

Ethanolamine-containing alkenyl ether glycerophospholipids, plasmalogens, are major cell membrane components of mammalian cells that activate membrane protein receptors such as ion transporters and G-protein coupled receptors. However, the mechanism by which plasmalogens modulate receptor function is unknown. Here, we found that exogenously added plasmalogens activate transient receptor potential cation channel subfamily C member 4 (TRPC4) to increase Ca2+ influx, followed by calcium/calmodulin-dependent protein kinase 2-mediated phosphorylation of AMP-activated protein kinase (AMPK). Upon topical application of plasmalogens to the skin of mice, AMPK activation was observed in TRPC4-expressing hair bulbs and hair follicles. Here, TRPC4 was co-localized with the leucine-rich repeat containing G protein-coupled receptor 5, a marker of hair-follicle stem cells, leading to hair growth. Collectively, this study indicates that plasmalogens could function as gate openers for TRPC4, followed by activating AMPK, which likely accelerates hair growth in mice.

TRPV4 mRNA is elevated in the caudate nucleus with NPH but not in Alzheimer's disease.

Symptoms of normal pressure hydrocephalus (NPH) and Alzheimer's disease (AD) are somewhat similar, and it is common to misdiagnose these two conditions. Although there are fluid markers detectable in humans with NPH and AD, determining which biomarker is optimal in representing genetic characteristics consistent throughout species is poorly understood. Here, we hypothesize that NPH can be differentiated from AD with mRNA biomarkers of unvaried proximity to telomeres. We examined human caudate nucleus tissue samples for the expression of transient receptor potential cation channel subfamily V member 4 (TRPV4) and amyloid precursor protein (APP). Using the genome data viewer, we analyzed the mutability of TRPV4 and other genes in mice, rats, and humans through matching nucleotides of six genes of interest and one house keeping gene with two factors associated with high mutation rate: 1) proximity to telomeres or 2) high adenine and thymine (A + T) content. We found that TRPV4 and microtubule associated protein tau (MAPT) mRNA were elevated in NPH. In AD, mRNA expression of TRPV4 was unaltered unlike APP and other genes. In mice, rats, and humans, the nucleotide size of TRPV4 did not vary, while in other genes, the sizes were inconsistent. Proximity to telomeres in TRPV4 was <50Mb across species. Our analyses reveal that TRPV4 gene size and mutability are conserved across three species, suggesting that TRPV4 can be a potential link in the pathophysiology of chronic hydrocephalus in aged humans (>65years) and laboratory rodents at comparable ages.

Distribution of acid-sensing ion channel subunits in human sensory neurons contrasts with that in rodents.

Acid-sensing ion channels (ASICs) play a critical role in nociception in human sensory neurons. Four genes (ASIC1, ASIC2, ASIC3, and ASIC4) encoding multiple subunits through alternative splicing have been identified in humans. Real time-PCR experiments showed strong expression of three subunits ASIC1, ASIC2, and ASIC3 in human dorsal root ganglia; however, their detailed expression pattern in different neuronal populations has not been investigated yet. In the current study, using an in situ hybridization approach (RNAscope), we examined the presence of ASIC1, ASIC2, and ASIC3 mRNA in three subpopulations of human dorsal root ganglia neurons. Our results revealed that ASIC1 and ASIC3 were present in the vast majority of dorsal root ganglia neurons, while ASIC2 was only expressed in less than half of dorsal root ganglia neurons. The distribution pattern of the three ASIC subunits was the same across the three populations of dorsal root ganglia neurons examined, including neurons expressing the REarranged during Transfection (RET) receptor tyrosine kinase, calcitonin gene-related peptide, and a subpopulation of nociceptors expressing Transient Receptor Potential Cation Channel Subfamily V Member 1. These results strongly contrast the expression pattern of Asics in mice since our previous study demonstrated differential distribution of Asics among the various subpopulation of dorsal root ganglia neurons. Given the distinct acid-sensitivity and activity dynamics among different ASIC channels, the expression differences between human and rodents should be taken under consideration when evaluating the translational potential and efficiency of drugs targeting ASICs in rodent studies.

Integration analysis of metabolome and transcriptome reveals the effect of exogenous supplementation with mixtures of vitamins ADE, zinc, and selenium on follicular growth and granulosa cells molecular metabolism in donkeys (Equus asinus).

Vitamins and microelements play essential roles in mammalian ovarian physiology, including follicle development, ovulation, and synthesis and secretion of hormones and growth factors. However, it is nevertheless elusive to what extent exogenous supplementation with mixtures of vitamins ADE, zinc (Zn), and selenium (Se) affects follicular growth and granulosa cells (GCs) molecular function. We herein investigated their effect on follicular growth and GCs physiological function. We showed that follicular growth and ovulation time was accelerated and shortened with the increases of vitamins ADE, Zn, and Se doses by continually monitoring and recording (one estrus cycle of about 21 days) with an ultrasound scanner. Integrated omics analysis showed that there was a sophisticated network relationship, correlation expression, and enrichment pathways of the genes and metabolites highly related to organic acids and their derivatives and lipid-like molecules. Quantitative real-time PCR (qPCR) results showed that vitamin D receptor (VDR), transient receptor potential cation channel subfamily m member 6 (TRPM6), transient receptor potential cation channel subfamily v member 6 (TRPV6), solute carrier family 5 member 1 (SLC5A1), arachidonate 5-lipoxygenase (ALOX5), steroidogenic acute regulatory protein (STAR), prostaglandin-endoperoxide synthase 2 (PTGS2), and insulin like growth factor 1 (IGF-1) had a strong correlation between the transcriptome data. Combined multi-omics analysis revealed that the protein digestion and absorption, ABC transporters, biosynthesis of amino acids, aminoacyl-tRNA biosynthesis, mineral absorption, alanine, aspartate and glutamate metabolism, glycine, serine and threonine metabolism, arginine biosynthesis, and ovarian steroidogenesis were significantly enriched. We focused on the gene-metabolite interactions in ovarian steroidogenesis, founding that insulin receptor (INSR), phospholipase a2 group IVA (PLA2G4A), adenylate cyclase 6 (ADCY6), cytochrome p450 family 1 subfamily b member 1 (CYP1B1), protein kinase camp-activated catalytic subunit beta (PRKACB), cytochrome p450 family 17 subfamily a member 1 (CYP17A1), and phospholipase a2 group IVF (PLA2G4F) were negatively correlated with β-estradiol (E2), progesterone (P4), and testosterone (T) (P < 0.05). while ALOX5 was a positive correlation with E2, P4, and T (P < 0.05); cytochrome p450 family 19 subfamily a member 1 (CYP19A1) was a negative correlation with cholesterol (P < 0.01). In mineral absorption, our findings further demonstrated that there was a positive correlation between solute carrier family 26 member 6 (SLC26A6), SLC5A1, and solute carrier family 6 member 19 (SLC6A19) with Glycine and L-methionine. Solute carrier family 40 member 1 (SLC40A1) was a negative correlation with Glycine and L-methionine (P < 0.01). TRPV6 and ATPase Na+/K+ transporting subunit alpha 1 (ATP1A1) were positively associated with Glycine (P < 0.05); while ATPase Na+/K+ transporting subunit beta 3 (ATP1B3) and cytochrome b reductase 1 (CYBRD1) were negatively related to L-methionine (P < 0.05). These outcomes suggested that the vitamins ADE, Zn, and Se of mixtures play an important role in the synthesis and secretion of steroid hormones and mineral absorption metabolism pathway through effects on the expression of the key genes and metabolites in GCs. Meanwhile, these also are required for physiological function and metabolism of GCs. Collectively, our outcomes shed new light on the underlying mechanisms of their effect on follicular growth and GCs molecular physiological function, helping explore valuable biomarkers.

N-Oleoyldopamine promotes the differentiation of mouse trophoblast stem cells into parietal trophoblast giant cells

The placenta plays various roles in a healthy pregnancy, and abnormalities in the placenta result in adverse outcomes. Adequate differentiation of trophoblast subtypes is necessary for placental function, but the molecular mechanisms that determine trophoblast cell fate remain unclear. Here, we screened small molecular compound (SMC) libraries (1904 SMCs) to identify particular SMCs which regulate trophoblast differentiation in mouse trophoblast stem cells (mTSCs) to understand the molecular mechanisms underlying cell fate decision in trophoblast cells. The two-step screening revealed a novel effect of N-oleoyldopamine (OLDA), an endogenic vanilloid, to promote differentiation into parietal trophoblast giant cells (P-TGCs) and repress them into spongiotrophoblast cells in mTSCs. Analyses by gene deletion and inhibitor treatments indicated that transient receptor potential cation channel subfamily V member 3 (Trpv3), one of the candidates for targeting by OLDA, was involved in maintaining stem status and P-TGC differentiation in mTSCs. Finally, transcriptome analysis revealed that Fosl1, a key regulatory factor in differentiation into P-TGCs, was upregulated by OLDA treatment, suggesting that OLDA promoted the differentiation of mTSCs into P-TGCs via regulation of Fosl1 expression.

Piperazine Derivative Stabilizes Actin Filaments in Primary Fibroblasts and Binds G-Actin In Silico.

Alzheimer's disease (AD) is characterized by synaptic dysfunction, which is expressed through the loss of dendritic spines and changes in their morphology. Pharmacological compounds that are able to protect spines in the AD brain are suggested to be novel drugs that would be able to slow down the disease progression. We have recently shown that a positive modulator of transient receptor potential cation channel subfamily C member 6 (TRPC6), the compound N-(2-chlorophenyl)-2-(4-phenylpiperazine-1-yl) acetamide (51164), causes the upregulation of postsynaptic neuronal store-operated calcium entry, maintains mushroom spine percentage, and recovers synaptic plasticity in amyloidogenic mouse models of Alzheimer's disease. Here, using confocal microscopy and calcium imaging methods, we present the experimental data indicating that 51164 possesses an alternative mechanism of action. We demonstrated that 51164 can increase the mushroom spine percentage in neurons with the downregulated activity of TRPC6-dependent neuronal store-operated calcium entry. Moreover, we report the binding of 51164 to G-actin in silico. We observed that 51164 interacts with Lys 336, Asp157, and Ser14 of G-actin, amino acids involved in the stabilization/polymerization of the G-actin structure. We showed that interactions of 51164 with G-actin are much stronger in comparison to the well-characterized F-actin stabilizing and polymerizing drug, jasplakinolide. The obtained results suggest an alternative protective mechanism of 51164 that is related to the preservation of actin filaments in vitro.

The distinctive role of menthol in pain and analgesia: Mechanisms, practices, and advances.

Menthol is an important flavoring additive that triggers a cooling sensation. Under physiological condition, low to moderate concentrations of menthol activate transient receptor potential cation channel subfamily M member 8 (TRPM8) in the primary nociceptors, such as dorsal root ganglion (DRG) and trigeminal ganglion, generating a cooling sensation, whereas menthol at higher concentration could induce cold allodynia, and cold hyperalgesia mediated by TRPM8 sensitization. In addition, the paradoxical irritating properties of high concentrations of menthol is associated with its activation of transient receptor potential cation channel subfamily A member 1 (TRPA1). Under pathological situation, menthol activates TRPM8 to attenuate mechanical allodynia and thermal hyperalgesia following nerve injury or chemical stimuli. Recent reports have recapitulated the requirement of central group II/III metabotropic glutamate receptors (mGluR) with endogenous κ-opioid signaling pathways for menthol analgesia. Additionally, blockage of sodium channels and calcium influx is a determinant step after menthol exposure, suggesting the possibility of menthol for pain management. In this review, we will also discuss and summarize the advances in menthol-related drugs for pathological pain treatment in clinical trials, especially in neuropathic pain, musculoskeletal pain, cancer pain and postoperative pain, with the aim to find the promising therapeutic candidates for the resolution of pain to better manage patients with pain in clinics.

VP.59 A single-arm, open-label, multicenter study of tranilast for advanced heart failure in patients with muscular dystrophy

The transient receptor potential cation channel subfamily V member 2 (TRPV2) is a stretch-sensitive calcium channel. Myocytes' damage induces TRPV2 expression on the sarcolemma, which causes calcium influx into the cytoplasm, and triggers degeneration. TRPV2 inhibition was effective in animal models of cardiomyopathy and muscular dystrophy (MD). Our pilot study showed that tranilast, a TRPV2 inhibitor, reduced brain natriuretic peptide (BNP) levels in two MD patients with advanced heart failure. Then, we planned a study to evaluate the safety and efficacy of tranilast for heart failure of MD patients. Subjects were MD patients whose serum BNP levels exceeded 100 pg/mL despite receiving standard therapy. Tranilast was administered orally at 100 mg thrice daily. The primary endpoint was the change in log (BNP) (⊿log [BNP]) at 6 months from baseline. The null hypothesis was determined based on a previous carvedilol study that resulted in a mean population ⊿log [BNP] of 0.18. TRPV2 expression on the mononuclear cell (MNC) surface, cardiac events, left ventricular fractional shortening (FS), human atrial natriuretic peptide (hANP), creatine kinase, and pinch strength were also assessed. Because of the poor general condition of many patients, among 18 patients included, 13 patients could be treated according to the protocol throughout the 6-month period. There were no serious adverse events related to tranilast except diarrhea, a known adverse effect. TRPV2 expression on the MNC surface was elevated at baseline and reduced after treatment. BNP, hANP, and FS remained stable. In the per-protocol set group, ⊿log [BNP] was -0.2 and significantly lower than that in the null hypothesis. In conclusion, tranilast is safe and effectively inhibits TRPV2 expression, even in MD patients with advanced heart failure. To evaluate the efficacy of tranilast in preventing heart failure, motor impairment, and respiratory failure, we are planning a study for mild MD patients.

Endocannabinoids Modulate Production of Osteoclastogenic Factors by Stem Cells of the Apical Papilla In Vitro

IntroductionMany mediators are produced during pulp inflammation and necrosis, including endocannabinoids (ECbs), which might affect the function of stem cells of the apical papilla (SCAP), cells of paramount importance for root formation, and regenerative endodontic treatment. The aim of this study was to evaluate the production of osteoclastogenesis-related mediators by SCAP modulated by ECbs and lipopolysaccharide (LPS) in vitro. MethodsSCAP were cultured and treated with ECb anandamide (AEA), 2-arachidonoylglycerol, or N-arachidonoylaminophenol. All groups were incubated in the presence of a vehicle or LPS and the antagonist of transient receptor potential cation channel subfamily V member 1, capsazepine. After 24 hours, the culture medium supernatants were collected for further quantification of tumor necrosis factor alpha, CCL2, macrophage colony-stimulating factor, osteoprotegerin, and receptor activator of nuclear factor kappa B ligand. ResultsSmall amounts of tumor necrosis factor alpha and receptor activator of nuclear factor kappa B ligand were detected in SCAP supernatants, and none of the experimental conditions altered their production. A down-regulation in constitutive CCL2 production was observed in the AEA group compared with that in the LPS group. The production of macrophage colony-stimulating factor was significantly increased in all groups treated with AEA compared with the control and LPS-treated groups. Osteoprotegerin was significantly increased by AEA alone and by 2-arachidonoylglycerol and N-arachidonoylaminophenol in the presence of LPS and capsazepine. ConclusionsAEA modulates some of the osteoclastogenic factors produced by SCAP in a bone resorption protective fashion.

Novel Cinnamaldehyde Derivatives Inhibit Peripheral Nerve Degeneration by Targeting Schwann Cells.

Peripheral nerve degeneration (PND) is a preparative process for peripheral nerve regeneration and is regulated by Schwann cells, a unique glial cell in the peripheral nervous system. Dysregulated PND induces irreversible peripheral neurodegenerative diseases (e.g., diabetic peripheral neuropathy). To develop novel synthetic drugs for these diseases, we synthesized a set of new cinnamaldehyde (CAH) derivatives and evaluated their activities in vitro, ex vivo, and in vivo. The 12 CAH derivatives had phenyl or naphthyl groups with different substitution patterns on either side of the α,β-unsaturated ketone. Among them, 3f, which had a naphthaldehyde group, was the most potent at inhibiting PND in vitro, ex vivo, and in vivo. To assess their interactions with transient receptor potential cation channel subfamily A member 1 (TRPA1) as a target of CAH, molecular docking studies were performed. Hydrophobic interactions had the highest binding affinity. To evaluate the underlying pharmacological mechanism, we performed bioinformatics analysis of the effect of 3f on PND based on coding genes and miRNAs regulated by CAH, suggesting that 3f affects oxidative stress in Schwann cells. The results show 3f to be a potential lead compound for the development of novel synthetic drugs for the treatment of peripheral neurodegenerative diseases.

Ehrlich Tumor Induces TRPV1-Dependent Evoked and Non-Evoked Pain-like Behavior in Mice.

We standardized a model by injecting Ehrlich tumor cells into the paw to evaluate cancer pain mechanisms and pharmacological treatments. Opioid treatment, but not cyclooxygenase inhibitor or tricyclic antidepressant treatments reduces Ehrlich tumor pain. To best use this model for drug screening it is essential to understand its pathophysiological mechanisms. Herein, we investigated the contribution of the transient receptor potential cation channel subfamily V member 1 (TRPV1) in the Ehrlich tumor-induced pain model. Dorsal root ganglia (DRG) neurons from the Ehrlich tumor mice presented higher activity (calcium levels using fluo-4 fluorescent probe) and an increased response to capsaicin (TRPV1 agonist) than the saline-injected animals (p < 0.05). We also observed diminished mechanical (electronic von Frey) and thermal (hot plate) hyperalgesia, paw flinching, and normalization of weight distribution imbalance in TRPV1 deficient mice (p < 0.05). On the other hand, TRPV1 deficiency did not alter paw volume or weight, indicating no significant alteration in tumor growth. Intrathecal injection of AMG9810 (TRPV1 antagonist) reduced ongoing Ehrlich tumor-triggered mechanical and thermal hyperalgesia (p < 0.05). Therefore, the contribution of TRPV1 to Ehrlich tumor pain behavior was revealed by genetic and pharmacological approaches, thus, supporting the use of this model to investigate TRPV1-targeting therapies for the treatment of cancer pain.