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1.
Zhen Ci Yan Jiu ; 46(1): 58-63, 2021 Jan 25.
Artigo em Chinês | MEDLINE | ID: mdl-33559427

RESUMO

OBJECTIVE: To observe the effect of electroacupuncture(EA)pretreatment on transient receptor potential vanilloid 1(TRPV1)/calcitonin gene-related peptide(CGRP)signal and nuclear factor-κB p65 (NF-κB p65) protein expression in myocardial tissue of acute myocardial ischemic injury (AMI) rats, and to investigate the possible mechanism of electroacupuncture pretreatment against AMI. METHODS: A total of 60 adult male SD rats were randomly divided into blank control, sham operation, model and EA pretreatment groups, 15 rats in each group. The acute myocardial ischemia model was established by ligating the left anterior descending (LAD)branch of the coronary artery in the model group and EA pretreatment group, while threading but no ligating at left anterior descending branch of the coronary artery was applied in the sham operation group. In the EA pretreatment group, bilateral "Neiguan"(PC6) acupoints were selected, with intensity of 2 mA and frequency of 2 Hz/100 Hz, for 20 min, once daily for 7 days before modeling. Electrocardiogram (ECG) was recorded by physiological signal acquisition system, and the ST segment potential offset values of standard Ⅱ lead were analyzed before surgery,30 min and 24 h after operation. The TTC staining was used to observe the percentage of myocardial infarction area. The HE staining was used to observe the pathological changes of myocardial tissue and the degree of inflammatory cell infiltration. And Western blot was used to detect TRPV1/CGRP signal and NF-κB p65 protein expression levels in myocardial tissue. RESULTS: Compared with the sham operation group, the ECG-J point potential in the model group was significantly increased at 30 min and decreased at 24 h after operation (P<0.05), myocardial infarction area increased significantly (P<0.05), the myocardial fibers were obviously disordered, inflammatory cell infiltration was obvious, and the expressions of TRPV1,CGRP and NF-κB p65 proteins were all increased (P<0.05). Compared with the model group, the EA pretreatment group was decreased in the ECG-J point potential at 30 min after operation(P<0.05), significantly reduced in myocardial infarction area (P<0.05), improved in the morphology of myocardial fibers, reduced ininflammatory cell infiltration, and increased in the protein expressions of TRPV1 and CGRP in myocardium (P<0.05), significantly decreased in the protein expression of NF-κB p65 (P<0.05). CONCLUSION: EA pretreatment may enhance TRPV1/CGRP signaling, down-regulate NF-κB p65 protein expression, reduce myocardial inflammatory response status, improve AMI injury, and reduce myocardial infarction area.


Assuntos
Eletroacupuntura , Isquemia Miocárdica , Pontos de Acupuntura , Animais , Calcitonina , Peptídeo Relacionado com Gene de Calcitonina/genética , Masculino , Isquemia Miocárdica/genética , Isquemia Miocárdica/terapia , NF-kappa B/genética , Ratos , Ratos Sprague-Dawley , Canais de Cátion TRPV/genética
2.
Arch Biochem Biophys ; 698: 108724, 2021 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-33309615

RESUMO

Abdominal aortic aneurysm (AAA) is a fatal vascular disease with insidious symptoms. However, the mechanism behind its development remains unclear. The transient receptor potential vanilloid (TRPV) family has crucial protective effects against cardiovascular diseases, but the role of TRPV5 in AAA has yet to be reported. In this study, ApoE-/- mice were intraperitoneally injected with AAV-GFP or AAV-TRPV5. After 30 days, mice were further administered with angiotensin II (Ang II, 1.44 mg/kg/day) by using osmotic pumps to induce the AAA model or Saline for 28 days, (i.e., Saline + AAV-GFP, Saline + AAV-TRPV5, Ang II + AAV-GFP and Ang II + AAV-TRPV5 groups were established). Compared with the control group, the incidence of AAA and the maximal diameter of the abdominal aorta markedly decreased in Ang II + AAV-TRPV5, which was detected by vascular ultrasound at 28 day. Meanwhile, less collagen and elastin degradation were observed in the Ang II + AAV-TRPV5 group by using Masson and Elastin stains. Moreover, more α-SMA and less MMP2 was observed in the abdominal aortas collected at 28 day by immunohistochemistry. In vitro, primary mouse vascular smooth muscle cells (VSMCs) were treated with Ang II (1 µM) to induce phenotype switch. Sh-TRPV5 and AdTRPV5 were used to transfect VSMCs. PCR and Western blotting were used to access the expression of contractile marker, including α-SMA and SM-22α. The results showed that the mRNA and protein level of α-SMA and SM-22α were decreased under the stimulation of Ang II, but could be attenuated by TRPV5 overexpression. The cell scratch assay demonstrated that the migration ability of VSMCs was increased in Ang II treated group and could be ameliorated by TRPV5 overexpression. Above all, VSMCs transformed from the contractile into secretory phenotype under Ang II stimuli, but could be rescued by TRPV5 overexpression. Furthermore, TRPV5 overexpression suppressed the increased expression of KLF4 induced by Ang II treatment in VSMCs. The data demonstrated that TRPV5 could inhibit AAA formation and play a critical role in the VSMC phenotype switch by downregulating KLF4, suggesting TRPV5 as a new strategy for treating AAA.


Assuntos
Aneurisma da Aorta Abdominal/tratamento farmacológico , Canais de Cálcio/farmacologia , Fatores de Transcrição Kruppel-Like/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Canais de Cátion TRPV/farmacologia , Angiotensina II , Animais , Aorta/citologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Aneurisma da Aorta Abdominal/patologia , Canais de Cálcio/genética , Diferenciação Celular/efeitos dos fármacos , Dependovirus/genética , Regulação para Baixo , Técnicas de Transferência de Genes , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos , Canais de Cátion TRPV/genética , Regulação para Cima
3.
Life Sci ; 261: 118461, 2020 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-32961227

RESUMO

AIMS: Parkinson's disease (PD) is a multifactorial neurodegenerative disorder. Its molecular mechanism is still unclear. Endoplasmic reticulum (ER) stress has been highlighted in PD. Transient receptor potential vanilloid 4 (TRPV4) is a kind of nonselective calcium cation channel. A defined role for TRPV4 in PD has not been reported. The purpose of the present research was to investigate the molecular mechanisms by which TRPV4 regulates ER stress induced by the 1-methyl-4-phenylpyridinium ion (MPP+) in PC12 cells. MAIN METHODS: PC12 cells were pretreated with the TRPV4-specific antagonist HC067047 or transfected with TRPV4 siRNA followed by treatment with MPP+. Cell viability was measured by the CCK-8 Assay. The expression of TRPV4, sarco/endoplasmic reticulum Ca2+-ATPase 2 (SERCA2), glucose-regulated protein 78 (GRP78), glucose-regulated protein 94 (GRP94), C/EBP homologous protein (CHOP), procaspase-12, and tyrosine hydroxylase (TH) was detected by western blot and RT-PCR. KEY FINDINGS: The expression of TRPV4 was upregulated, while cell viability was decreased by MPP+, which was reversed by HC067047. The ER stress common molecular signature SERCA2 was depressed by MPP+. Moreover, MPP+ induced upregulation of GRP78, GRP94, CHOP, and decrease in procaspase-12 and TH. HC067047 and TRPV4 siRNA reversed MPP+-induced ER stress and restored TH production. SIGNIFICANCE: TRPV4 functions upstream of ER stress induced by MPP+ and holds promise as a prospective pharmacotherapy target for PD.


Assuntos
Apoptose , Estresse do Retículo Endoplasmático , Doença de Parkinson Secundária/patologia , Canais de Cátion TRPV/metabolismo , 1-Metil-4-fenilpiridínio , Animais , Sobrevivência Celular , Células PC12 , Doença de Parkinson Secundária/genética , Doença de Parkinson Secundária/metabolismo , Ratos , Canais de Cátion TRPV/análise , Canais de Cátion TRPV/genética , Regulação para Cima
4.
Chem Biol Interact ; 330: 109178, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32738201

RESUMO

The capsaicin (vanilloid) receptor, TRPV1, is a heat-activated cation channel modulated by inflammatory mediators and contributes to acute and chronic pain. TRPV1 channel is one of the most researched and targeted mechanisms for the development of novel analgesics. Over the years, natural products have contributed enormously to the development of important therapeutic drugs used currently in modern medicine. A literature review was conducted using Medline, Google Scholar, and PubMed. Searching the literature resulted in listing 136 natural compounds that interacted with TRPV1 channel. These compounds were phytochemicals that belong to different chemical groups including vanilloids, flavonoids, alkaloids, terpenoids, terpenyl phenols, fatty acids, cannabinoids, sulfur_containing compounds, etc. Other natural TRPV1 modulators were of animal, fungal or bacterial origin. Some natural products were small agonists or antagonists of TRPV1. Others were protein venoms. Most in vitro studies utilized electrophysiological or calcium imaging techniques to study calcium flow through the channel using primary cultures of rat dorsal root and trigeminal ganglia. Other studies used hTRPV1 or rTRPV1 expressed in HEK239, CHO cells or Xenopus oocytes. In vivo studies concentrated on different pain models conducted mainly in mice and rats. In conclusion, natural products are highly diverse in their modulatory action on TRPV1. Many gaps in natural product research are present in distinguishing modality-specific from polymodal antagonists. Species' differences in TRPV1 functionality must be taken into account in any future study. Proceeding into clinical trials needs more efforts to discover potent TRPV1 antagonists devoid of hyperthermia, the main side effect.


Assuntos
Analgésicos/farmacologia , Produtos Biológicos/farmacologia , Canais de Cátion TRPV/metabolismo , Analgésicos/efeitos adversos , Analgésicos/uso terapêutico , Animais , Produtos Biológicos/efeitos adversos , Cálcio/metabolismo , Técnicas de Cultura de Células , Febre/etiologia , Humanos , Dor/complicações , Dor/tratamento farmacológico , Canais de Cátion TRPV/agonistas , Canais de Cátion TRPV/antagonistas & inibidores , Canais de Cátion TRPV/efeitos dos fármacos , Canais de Cátion TRPV/genética
5.
Nat Commun ; 11(1): 4169, 2020 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-32820172

RESUMO

Sensing and responding to temperature is crucial in biology. The TRPV1 ion channel is a well-studied heat-sensing receptor that is also activated by vanilloid compounds, including capsaicin. Despite significant interest, the molecular underpinnings of thermosensing have remained elusive. The TRPV1 S1-S4 membrane domain couples chemical ligand binding to the pore domain during channel gating. Here we show that the S1-S4 domain also significantly contributes to thermosensing and couples to heat-activated gating. Evaluation of the isolated human TRPV1 S1-S4 domain by solution NMR, far-UV CD, and intrinsic fluorescence shows that this domain undergoes a non-denaturing temperature-dependent transition with a high thermosensitivity. Further NMR characterization of the temperature-dependent conformational changes suggests the contribution of the S1-S4 domain to thermosensing shares features with known coupling mechanisms between this domain with ligand and pH activation. Taken together, this study shows that the TRPV1 S1-S4 domain contributes to TRPV1 temperature-dependent activation.


Assuntos
Temperatura Alta , Ativação do Canal Iônico/fisiologia , Canais de Cátion TRPV/metabolismo , Sensação Térmica/fisiologia , Sítios de Ligação/genética , Capsaicina/química , Capsaicina/metabolismo , Dicroísmo Circular , Humanos , Ativação do Canal Iônico/genética , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Ligação Proteica , Domínios Proteicos , Canais de Cátion TRPV/química , Canais de Cátion TRPV/genética , Sensação Térmica/genética
6.
Chem Biol Interact ; 329: 109220, 2020 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-32763245

RESUMO

The sepsis is considered as serious clinic-pathological condition related with high rate of morbidity and mortality in critical care settings. In the proposed study, the hydrazides derivatives N-(benzylidene)-2-((2-hydroxynaphthalen-1-yl)diazenyl)benzohydrazides (1-2) (NCHDH and NTHDH) were investigated against the LPS-induced sepsis in rodents. The NCHDH and NTHDH markedly improved the physiological sign and symptoms associated with the sepsis such as mortality, temperature, and clinical scoring compared to negative control group, which received only LPS (i.p.). The NCHDH and NTHDH also inhibited the production of the NO and MPO compared to the negative control. Furthermore, the treatment control improved the histological changes markedly of all the vital organs. Additionally, the Masson's trichrome and PAS (Periodic Acid Schiff) staining also showed improvement in the NCHDH and NTHDH treated group in contrast to LPS-induced group. The antioxidants were enhanced by the intervention of the NCHDH and NTHDH and the level of the MDA and POD were attenuated marginally compared to the LPS-induced group. The hematology study showed marked improvement and the reversal of the LPS-induced changes in blood composition compared to the negative control. The synthetic function of the liver and kidney were preserved in the NCHDH and NTHDH treated group compared to the LPS-induced group. The NCHDH and NTHDH markedly enhanced the Nrf2, HO-1 (Heme oxygenase-1), while attenuated the Keap1 and TRPV1 expression level as compared to LPS treated group. Furthermore, the NCHDH and NTHDH treatment showed marked increased in the mRNA expression level of the HSP70/90 proteins compared to the negative control.


Assuntos
Hidrazinas/farmacologia , Insuficiência de Múltiplos Órgãos/etiologia , Sepse/etiologia , Transdução de Sinais/efeitos dos fármacos , Animais , Sobrevivência Celular/efeitos dos fármacos , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Heme Oxigenase-1/metabolismo , Hidrazinas/química , Hidrazinas/uso terapêutico , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Lipopolissacarídeos/toxicidade , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Insuficiência de Múltiplos Órgãos/tratamento farmacológico , Insuficiência de Múltiplos Órgãos/mortalidade , Fator 2 Relacionado a NF-E2/metabolismo , Óxido Nítrico/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Peroxidase/metabolismo , Sepse/tratamento farmacológico , Sepse/mortalidade , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/metabolismo
7.
Proc Natl Acad Sci U S A ; 117(28): 16626-16637, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32601236

RESUMO

Neuronal activity leads to an increase in local cerebral blood flow (CBF) to allow adequate supply of oxygen and nutrients to active neurons, a process termed neurovascular coupling (NVC). We have previously shown that capillary endothelial cell (cEC) inwardly rectifying K+ (Kir) channels can sense neuronally evoked increases in interstitial K+ and induce rapid and robust dilations of upstream parenchymal arterioles, suggesting a key role of cECs in NVC. The requirements of this signal conduction remain elusive. Here, we utilize mathematical modeling to investigate how small outward currents in stimulated cECs can elicit physiologically relevant spread of vasodilatory signals within the highly interconnected brain microvascular network to increase local CBF. Our model shows that the Kir channel can act as an "on-off" switch in cECs to hyperpolarize the cell membrane as extracellular K+ increases. A local hyperpolarization can be amplified by the voltage-dependent activation of Kir in neighboring cECs. Sufficient Kir density enables robust amplification of the hyperpolarizing stimulus and produces responses that resemble action potentials in excitable cells. This Kir-mediated excitability can remain localized in the stimulated region or regeneratively propagate over significant distances in the microvascular network, thus dramatically increasing the efficacy of K+ for eliciting local hyperemia. Modeling results show how changes in cEC transmembrane current densities and gap junctional resistances can affect K+-mediated NVC and suggest a key role for Kir as a sensor of neuronal activity and an amplifier of retrograde electrical signaling in the cerebral vasculature.


Assuntos
Neurônios/metabolismo , Acoplamento Neurovascular , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Potássio/metabolismo , Animais , Encéfalo/irrigação sanguínea , Encéfalo/metabolismo , Circulação Cerebrovascular , Células Endoteliais/química , Células Endoteliais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Biológicos , Neurônios/química , Potássio/química , Canais de Potássio Corretores do Fluxo de Internalização/química , Canais de Potássio Corretores do Fluxo de Internalização/genética , Transdução de Sinais , Canais de Cátion TRPV/química , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/metabolismo
8.
Am J Physiol Heart Circ Physiol ; 319(2): H507-H518, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32706268

RESUMO

The lymphatic system drains and propels lymph by extrinsic and intrinsic mechanisms. Intrinsic propulsion depends upon spontaneous rhythmic contractions of lymphatic muscles in the vessel walls and is critically affected by changes in the surrounding tissue like osmolarity and temperature. Lymphatics of the diaphragm display a steep change in contraction frequency in response to changes in temperature, and this, in turn, affects lymph flow. In the present work, we demonstrated in an ex vivo diaphragmatic tissue rat model that diaphragmatic lymphatics express transient receptor potential channels of the vanilloid 4 subfamily (TRPV4) and that their blockade by both the nonselective antagonist Ruthenium Red and the selective antagonist HC-067047 abolished the response of lymphatics to temperature changes. Moreover, the selective activation of TRPV4 channels by means of GSK1016790A mirrored the behavior of vessels exposed to increasing temperatures, pointing out the critical role played by these channels in sensing the temperature of the lymphatic vessels' environment and thus inducing a change in contraction frequency and lymph flow.NEW & NOTEWORTHY The present work addresses the putative receptor system that enables diaphragmatic lymphatics to change intrinsic contraction frequency and thus lymph flow according to the changes in temperature of the surrounding environment, showing that this role can be sustained by TRPV4 channels alone.


Assuntos
Linfa/fisiologia , Vasos Linfáticos/metabolismo , Contração Muscular , Músculo Liso/metabolismo , Canais de Cátion TRPV/metabolismo , Temperatura , Animais , Diafragma , Feminino , Técnicas In Vitro , Vasos Linfáticos/efeitos dos fármacos , Masculino , Morfolinas/farmacologia , Músculo Liso/efeitos dos fármacos , Periodicidade , Pirróis/farmacologia , Ratos , Ratos Wistar , Rutênio Vermelho/farmacologia , Transdução de Sinais , Canais de Cátion TRPV/antagonistas & inibidores , Canais de Cátion TRPV/genética , Fatores de Tempo
9.
Science ; 368(6495): 1108-1113, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32499439

RESUMO

Enabling near-infrared light sensitivity in a blind human retina may supplement or restore visual function in patients with regional retinal degeneration. We induced near-infrared light sensitivity using gold nanorods bound to temperature-sensitive engineered transient receptor potential (TRP) channels. We expressed mammalian or snake TRP channels in light-insensitive retinal cones in a mouse model of retinal degeneration. Near-infrared stimulation increased activity in cones, ganglion cell layer neurons, and cortical neurons, and enabled mice to perform a learned light-driven behavior. We tuned responses to different wavelengths, by using nanorods of different lengths, and to different radiant powers, by using engineered channels with different temperature thresholds. We targeted TRP channels to human retinas, which allowed the postmortem activation of different cell types by near-infrared light.


Assuntos
Cegueira/terapia , Ouro , Raios Infravermelhos , Nanotubos , Degeneração Retiniana/terapia , Limiar Sensorial/efeitos da radiação , Canais de Cátion TRPC/fisiologia , Visão Ocular/efeitos da radiação , Animais , Cegueira/fisiopatologia , Modelos Animais de Doenças , Potenciais Evocados Visuais/fisiologia , Potenciais Evocados Visuais/efeitos da radiação , Engenharia Genética , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Estimulação Luminosa , Ratos , Células Fotorreceptoras Retinianas Cones/fisiologia , Células Fotorreceptoras Retinianas Cones/efeitos da radiação , Degeneração Retiniana/fisiopatologia , Células Ganglionares da Retina/fisiologia , Células Ganglionares da Retina/efeitos da radiação , Limiar Sensorial/fisiologia , Serpentes , Canais de Cátion TRPC/genética , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/fisiologia , Visão Ocular/fisiologia , Córtex Visual/fisiopatologia , Córtex Visual/efeitos da radiação
10.
Nat Struct Mol Biol ; 27(7): 635-644, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32572252

RESUMO

The transient receptor potential cation channel subfamily V member 3 (TRPV3) channel plays a critical role in skin physiology, and mutations in TRPV3 result in the development of a congenital skin disorder, Olmsted syndrome. Here we describe multiple cryo-electron microscopy structures of human TRPV3 reconstituted into lipid nanodiscs, representing distinct functional states during the gating cycle. The ligand-free, closed conformation reveals well-ordered lipids interacting with the channel and two physical constrictions along the ion-conduction pore involving both the extracellular selectivity filter and intracellular helix bundle crossing. Both the selectivity filter and bundle crossing expand upon activation, accompanied by substantial structural rearrangements at the cytoplasmic intersubunit interface. Transition to the inactivated state involves a secondary structure change of the pore-lining helix, which contains a π-helical segment in the closed and open conformations, but becomes entirely α-helical upon inactivation. Together with electrophysiological characterization, structures of TRPV3 in a lipid membrane environment provide unique insights into channel activation and inactivation mechanisms.


Assuntos
Ativação do Canal Iônico/fisiologia , Bicamadas Lipídicas/química , Canais de Cátion TRPV/química , Canais de Cátion TRPV/metabolismo , Microscopia Crioeletrônica , Bicamadas Lipídicas/metabolismo , Modelos Moleculares , Mutação , Nanoestruturas/química , Conformação Proteica , Canais de Cátion TRPV/genética
11.
Nat Commun ; 11(1): 2679, 2020 05 29.
Artigo em Inglês | MEDLINE | ID: mdl-32471994

RESUMO

The cation channel transient receptor potential vanilloid 4 (TRPV4) is one of the few identified ion channels that can directly cause inherited neurodegeneration syndromes, but the molecular mechanisms are unknown. Here, we show that in vivo expression of a neuropathy-causing TRPV4 mutant (TRPV4R269C) causes dose-dependent neuronal dysfunction and axonal degeneration, which are rescued by genetic or pharmacological blockade of TRPV4 channel activity. TRPV4R269C triggers increased intracellular Ca2+ through a Ca2+/calmodulin-dependent protein kinase II (CaMKII)-mediated mechanism, and CaMKII inhibition prevents both increased intracellular Ca2+ and neurotoxicity in Drosophila and cultured primary mouse neurons. Importantly, TRPV4 activity impairs axonal mitochondrial transport, and TRPV4-mediated neurotoxicity is modulated by the Ca2+-binding mitochondrial GTPase Miro. Our data highlight an integral role for CaMKII in neuronal TRPV4-associated Ca2+ responses, the importance of tightly regulated Ca2+ dynamics for mitochondrial axonal transport, and the therapeutic promise of TRPV4 antagonists for patients with TRPV4-related neurodegenerative diseases.


Assuntos
Sinalização do Cálcio/fisiologia , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Drosophila melanogaster/metabolismo , Doenças Neurodegenerativas/genética , Canais de Cátion TRPV/genética , Animais , Animais Geneticamente Modificados , Axônios/patologia , Cálcio/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/antagonistas & inibidores , Células Cultivadas , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Doenças Neurodegenerativas/patologia , Asas de Animais/crescimento & desenvolvimento
12.
Sci Rep ; 10(1): 8684, 2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32457384

RESUMO

Essential for calcium homeostasis, TRPV5 and TRPV6 are calcium-selective channels belonging to the transient receptor potential (TRP) gene family. In this study, we investigated the evolutionary history of these channels to add an evolutionary context to the already available physiological information. Phylogenetic analyses revealed that paralogs found in mammals, sauropsids, amphibians, and chondrichthyes, are the product of independent duplication events in the ancestor of each group. Within amniotes, we identified a traceable signature of three amino acids located at the amino-terminal intracellular region. The signature correlates with both the duplication events and the phenotype of fast inactivation observed in mammalian TRPV6 channels. Electrophysiological recordings and mutagenesis revealed that the signature sequence modulates the phenotype of fast inactivation in all clades of vertebrates but reptiles. A transcriptome analysis showed a change in tissue expression from gills, in marine vertebrates, to kidneys in terrestrial vertebrates. Our results highlight a cytoplasmatic structural triad composed by the Helix-Loop-Helix domain, the S2-S3 linker, and the TRP domain helix that is important on modulating the activity of calcium-selective TRPV channels.


Assuntos
Cálcio/metabolismo , Evolução Molecular , Canais de Cátion TRPV/metabolismo , Sequência de Aminoácidos , Anfíbios/metabolismo , Animais , Aves/metabolismo , Brânquias/metabolismo , Células HEK293 , Sequências Hélice-Alça-Hélice , Humanos , Rim/metabolismo , Mamíferos/metabolismo , Mutagênese Sítio-Dirigida , Filogenia , Alinhamento de Sequência , Canais de Cátion TRPV/química , Canais de Cátion TRPV/classificação , Canais de Cátion TRPV/genética
13.
Cancer Genomics Proteomics ; 17(3): 309-319, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32345672

RESUMO

BACKGROUND: Transient receptor potential vanilloid type 1 (TRPV1) has been studied in human malignancies, but has not been studied in epithelial ovarian cancer (EOC). We, therefore, investigated the significance of TRPV1 and correlation with phosphatase and tension homolog (PTEN) in EOC. MATERIALS AND METHODS: Immunohistochemical analyses for TRPV1 and PTEN were performed using a tissue microarray. Moreover, the role of TRPV1 in cell growth was assessed in a EOC cell line. RESULTS: High TRPV1 expression and the combination of high TRPV1 and low PTEN expression were an independent prognostic factor for overall survival and disease-free survival. In vitro results demonstrated that knockdown of TRPV1 was associated with decreased cell viability and colony formation. CONCLUSION: There is a strong association between TRPV1 and PTEN and the combination of TRPV1 and PTEN is a strong indicator of prognostic predictor in EOC.


Assuntos
Carcinoma Epitelial do Ovário/metabolismo , Neoplasias Ovarianas/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Canais de Cátion TRPV/metabolismo , Biomarcadores Tumorais/metabolismo , Carcinoma Epitelial do Ovário/genética , Carcinoma Epitelial do Ovário/patologia , Estudos de Casos e Controles , Proliferação de Células/fisiologia , Feminino , Seguimentos , Humanos , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , PTEN Fosfo-Hidrolase/biossíntese , PTEN Fosfo-Hidrolase/genética , Prognóstico , Canais de Cátion TRPV/biossíntese , Canais de Cátion TRPV/genética , Células Tumorais Cultivadas
14.
Lab Invest ; 100(8): 1057-1067, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32341518

RESUMO

Endothelial barrier disruption is a hallmark of tissue injury, edema, and inflammation. Vascular endothelial cells express the G protein-coupled receptor (GPCR) protease acctivated receptor 1 (PAR1) and the ion channel transient receptor potential vanilloid 4 (TRPV4), and these signaling proteins are known to respond to inflammatory conditions and promote edema through remodeling of cell-cell junctions and modulation of endothelial barriers. It has previously been established that signaling initiated by the related protease activated receptor 2 (PAR2) is enhanced by TRPV4 in sensory neurons and that this functional interaction plays a critical role in the development of neurogenic inflammation and nociception. Here, we investigated the PAR1-TRPV4 axis, to determine if TRPV4 plays a similar role in the control of edema mediated by thrombin-induced signaling. Using Evans Blue permeation and retention as an indication of increased vascular permeability in vivo, we showed that TRPV4 contributes to PAR1-induced vascular hyperpermeability in the airways and upper gastrointestinal tract of mice. TRPV4 contributes to sustained PAR1-induced Ca2+ signaling in recombinant cell systems and to PAR1-dependent endothelial junction remodeling in vitro. This study supports the role of GPCR-TRP channel functional interactions in inflammatory-associated changes to vascular function and indicates that TRPV4 is a signaling effector for multiple PAR family members.


Assuntos
Inflamação/genética , Receptor PAR-1/genética , Receptor PAR-2/genética , Transdução de Sinais/genética , Canais de Cátion TRPV/genética , Animais , Cálcio/metabolismo , Permeabilidade Capilar/genética , Edema/genética , Edema/metabolismo , Células HEK293 , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Inflamação/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor PAR-1/metabolismo , Receptor PAR-2/metabolismo , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo , Canais de Cátion TRPV/metabolismo
15.
BMC Med Genet ; 21(1): 64, 2020 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-32228492

RESUMO

BACKGROUND: The calcium-selective channel TRPV6 (transient receptor potential cation channel subfamily V member 6) is crucial for maternal-fetal calcium transport across the placenta. TRPV6 mutations have recently been associated with an antenatally severe under-mineralising skeletal dysplasia accompanied by postnatal biochemical abnormalities. This is the first post-mortem report in a patient with TRPV6 skeletal dysplasia. CASE PRESENTATION: The female infant had severe antenatal and postnatal skeletal abnormalities by 20 weeks gestation and was ventilator-dependent from birth. These skeletal abnormalities were apparent at an earlier gestational age than in previous reported cases and a more severe clinical course ensued. Biochemical and skeletal abnormalities, including bone density, improved postnatally but cardiac arrest at 4 months of age led to withdrawal of intensive care. Compound heterozygous TRPV6 variants (c.1978G > C p.(Gly660Arg) and c.1528C > T p.(Arg510Ter)) were identified on exome sequencing. Post-mortem identified skeletal abnormalities but no specific abnormalities in other organ systems. No placental pathology was found, multi-organ histological features reflected prolonged intensive care only. Post-mortem macroscopic examination indicated reduced thoracic size and short, pale and pliable ribs. Histological examination identified reduced number of trabeculae in the diaphyses (away from the growth plates), whereas metaphyses showed adequate mineralisation and normal number of trabeculae, but with slightly enlarged reactive chondrocytes, indicating post-natal skeletal growth recovery. Post-mortem radiological findings demonstrated improved bone density, improved rib width, healed fractures, although ribs were still shorter than normal. Long bones (especially humerus and femur) had improved from initial poorly defined metaphyses and reduced bone density to sharply defined metaphyses, prominent growth restart lines in distal diaphyses and bone-in-bone appearance along diaphyses. CONCLUSIONS: This case provide bone histological confirmation that human skeletal development is compromised in the presence of TRPV6 pathogenic variants. Post-mortem findings were consistent with abnormal in utero skeletal mineralisation due to severe calcium deficit from compromised placental calcium transfer, followed by subsequent phenotypic improvement with adequate postnatal calcium availability. Significant skeletal recovery occurs in the early weeks of postnatal life in TRPV6 skeletal dysplasia.


Assuntos
Desenvolvimento Ósseo , Osso e Ossos/patologia , Canais de Cálcio/genética , Desenvolvimento Infantil/fisiologia , Osteocondrodisplasias/genética , Osteocondrodisplasias/patologia , Canais de Cátion TRPV/genética , Autopsia , Desenvolvimento Ósseo/genética , Osso e Ossos/anormalidades , Calcificação Fisiológica/genética , Cálcio/metabolismo , Canais de Cálcio/análise , Análise Mutacional de DNA , Feminino , Humanos , Lactente , Osteocondrodisplasias/reabilitação , Parto/fisiologia , Canais de Cátion TRPV/análise
16.
Am J Physiol Cell Physiol ; 318(5): C969-C980, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32293931

RESUMO

The porcine lens response to a hyperosmotic stimulus involves an increase in the activity of an ion cotransporter sodium-potassium/two-chloride cotransporter 1 (NKCC1). Recent studies with agonists and antagonists pointed to a mechanism that appears to depend on activation of transient receptor potential vanilloid 1 (TRPV1) ion channels. Here, we compare responses in lenses and cultured lens epithelium obtained from TRPV1-/- and wild type (WT) mice. Hydrostatic pressure (HP) in lens surface cells was determined using a manometer-coupled microelectrode approach. The TRPV1 agonist capsaicin (100 nM) caused a transient HP increase in WT lenses that peaked after ∼30 min and then returned toward baseline. Capsaicin did not cause a detectable change of HP in TRPV1-/- lenses. The NKCC inhibitor bumetanide prevented the HP response to capsaicin in WT lenses. Potassium transport was examined by measuring Rb+ uptake. Capsaicin increased Rb+ uptake in cultured WT lens epithelial cells but not in TRPV1-/- cells. Bumetanide, A889425, and the Akt inhibitor Akti prevented the Rb+ uptake response to capsaicin. The bumetanide-sensitive (NKCC-dependent) component of Rb+ uptake more than doubled in response to capsaicin. Capsaicin also elicited rapid (<2 min) NKCC1 phosphorylation in WT but not TRPV1-/- cells. HP recovery was shown to be absent in TRPV1-/- lenses exposed to hyperosmotic solution. Bumetanide and Akti prevented HP recovery in WT lenses exposed to hyperosmotic solution. Taken together, responses to capsaicin and hyperosmotic solution point to a functional role for TRPV1 channels in mouse lens. Lack of NKCC1 phosphorylation and Rb+ uptake responses in TRPV1-/- mouse epithelium reinforces the notion that a hyperosmotic challenge causes TRPV1-dependent NKCC1 activation. The results are consistent with a role for the TRPV1-activated signaling pathway leading to NKCC1 stimulation in lens osmotic homeostasis.


Assuntos
Cristalino/metabolismo , Membro 2 da Família 12 de Carreador de Soluto/genética , Canais de Cátion TRPV/genética , Animais , Bumetanida/farmacologia , Capsaicina/farmacologia , Linhagem Celular , Epitélio/efeitos dos fármacos , Epitélio/metabolismo , Humanos , Pressão Hidrostática/efeitos adversos , Cristalino/efeitos dos fármacos , Camundongos , Camundongos Knockout , Fosforilação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Suínos
17.
Am J Physiol Cell Physiol ; 318(5): C1018-C1029, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32293932

RESUMO

Autophagy is a highly conserved self-protection mechanism that plays a crucial role in cardiovascular diseases. Cardiomyocyte hypoxic injury promotes oxidative stress and pathological alterations in the heart, although the interplay between these effects remains elusive. The transient receptor potential vanilloid 1 (TRPV1) ion channel is a nonselective cation channel that is activated in response to a variety of exogenous and endogenous physical and chemical stimuli. Here, we investigated the effects and mechanisms of action of TRPV1 on autophagy in hypoxic cardiomyocytes. In this study, primary cardiomyocytes isolated from C57 mice were subjected to hypoxic stress, and their expression of TRPV1 and adenosine 5'-monophosphate-activated protein kinase (AMPK) was regulated. The autophagy flux was assessed by Western blotting and immunofluorescence staining, and the cell viability was determined through Cell counting kit-8 assay and Lactate dehydrogenase assays. In addition, the calcium influx after the upregulation of TRPV1 expression in cardiomyocytes was examined. The results showed that the number of autophagosomes in cardiomyocytes was higher under hypoxic stress and that the blockade of autophagy flux aggravated hypoxic damage to cardiomyocytes. Moreover, the expression of TRPV1 was induced under hypoxic stress, and its upregulation by capsaicin improved the autophagy flux and protected cardiomyocytes from hypoxic damage, whereas the silencing of TRPV1 significantly attenuated autophagy. Our observations also revealed that AMPK signaling was activated and involved in TRPV1-induced autophagy in cardiomyocytes under hypoxic stress. Overall, this study demonstrates that TRPV1 activation mitigates hypoxic injury in cardiomyocytes by improving autophagy flux through the AMPK signaling pathway and highlights TRPV1 as a novel therapeutic target for the treatment of hypoxic cardiac disease.


Assuntos
Autofagia/genética , Traumatismos Cardíacos/genética , Proteínas Quinases/genética , Canais de Cátion TRPV/genética , Animais , Cálcio/metabolismo , Capsaicina/farmacologia , Sobrevivência Celular/genética , Modelos Animais de Doenças , Regulação da Expressão Gênica/genética , Traumatismos Cardíacos/patologia , Humanos , Hipóxia/genética , Hipóxia/metabolismo , Hipóxia/patologia , Camundongos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Transdução de Sinais/genética
18.
Food Funct ; 11(4): 3516-3526, 2020 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-32253400

RESUMO

Chronic obstructive pulmonary disease (COPD) is a chronic, progressive lung disease with few successful treatments, and is strongly associated with cigarette smoking (CS). Since the novel coronavirus has spread worldwide seriously, there is growing concern that patients who have chronic respiratory conditions like COPD can easily be infected and are more prone to having severe illness and even mortality because of lung dysfunction. Loquat leaves have long been used as an important material for both pharmaceutical and functional applications in the treatment of lung disease in Asia, especially in China and Japan. Total flavonoids (TF), the main active components derived from loquat leaves, showed remarkable anti-inflammatory and antioxidant activities. However, their protective activity against CS-induced COPD airway inflammation and oxidative stress and its underlying mechanism still remain not well-understood. The present study uses a CS-induced mouse model to estimate the morphological changes in lung tissue. The results demonstrated that TF suppressed the histological changes in the lungs of CS-challenged mice, as evidenced by reduced generation of pro-inflammatory cytokines including interleukin 6 (IL-6), IL-1ß, tumor necrosis factor α (TNF-α), nitric oxide (NO), and inducible nitric oxide synthase (iNOS) and diminished the protein expression of transient receptor potential vanilloid 1 (TRPV1). Moreover, TF also inhibited phosphorylation of IKK, IκB and NFκB and increased p-Akt. Interestingly, TF could inhibit CS-induced oxidative stress in the lungs of COPD mice. TF treatment significantly inhibited the level of malondialdehyde (MDA) and increased the activity of superoxide dismutase (SOD). In addition, TF markedly downregulated TRPV1 and cytochrome P450 2E1 (CYP2E1) and upregulated the expression of SOD-2, while the p-JNK level was observed to be inhibited in COPD mice. Taken together, our findings showed that the protective effect and putative mechanism of the action of TF resulted in the inhibition of inflammation and oxidative stress through the regulation of TRPV1 and the related signal pathway in lung tissues. It suggested that TF derived from loquat leaves could be considered to be an alternative or a new functional material and used for the treatment of CS-induced COPD.


Assuntos
Fumar Cigarros/efeitos adversos , Medicamentos de Ervas Chinesas/administração & dosagem , Eriobotrya/química , Flavonoides/administração & dosagem , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Canais de Cátion TRPV/imunologia , Animais , Citocromo P-450 CYP2E1/genética , Citocromo P-450 CYP2E1/imunologia , Humanos , Interleucina-6/genética , Interleucina-6/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estresse Oxidativo/efeitos dos fármacos , Folhas de Planta/química , Doença Pulmonar Obstrutiva Crônica/etiologia , Doença Pulmonar Obstrutiva Crônica/imunologia , Transdução de Sinais/efeitos dos fármacos , Fumaça/efeitos adversos , Superóxido Dismutase/genética , Superóxido Dismutase/imunologia , Canais de Cátion TRPV/genética
19.
Invest Ophthalmol Vis Sci ; 61(4): 2, 2020 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-32271891

RESUMO

Purpose: Contact lenses, osmotic stressors, and chemical burns may trigger severe discomfort and vision loss by damaging the cornea, but the signaling mechanisms used by corneal epithelial cells (CECs) to sense extrinsic stressors are not well understood. We therefore investigated the mechanisms of swelling, temperature, strain, and chemical transduction in mouse CECs. Methods: Intracellular calcium imaging in conjunction with electrophysiology, pharmacology, transcript analysis, immunohistochemistry, and bioluminescence assays of adenosine triphosphate (ATP) release were used to track mechanotransduction in dissociated CECs and epithelial sheets isolated from the mouse cornea. Results: The transient receptor potential vanilloid (TRPV) transcriptome in the mouse corneal epithelium is dominated by Trpv4, followed by Trpv2, Trpv3, and low levels of Trpv1 mRNAs. TRPV4 protein was localized to basal and intermediate epithelial strata, keratocytes, and the endothelium in contrast to the cognate TRPV1, which was confined to intraepithelial afferents and a sparse subset of CECs. The TRPV4 agonist GSK1016790A induced cation influx and calcium elevations, which were abolished by the selective blocker HC067047. Hypotonic solutions, membrane strain, and moderate heat elevated [Ca2+]CEC with swelling- and temperature-, but not strain-evoked signals, sensitive to HC067047. GSK1016790A and swelling evoked calcium-dependent ATP release, which was suppressed by HC067027 and the hemichannel blocker probenecid. Conclusions: These results demonstrate that cation influx via TRPV4 transduces osmotic and thermal but not strain inputs to CECs and promotes hemichannel-dependent ATP release. The TRPV4-hemichannel-ATP signaling axis might modulate corneal pain induced by excessive mechanical, osmotic, and chemical stimulation.


Assuntos
Epitélio Anterior/metabolismo , Mecanotransdução Celular/fisiologia , Trifosfato de Adenosina/metabolismo , Animais , Cálcio/metabolismo , Sinalização do Cálcio/fisiologia , Células Cultivadas , Eletrofisiologia , Feminino , Regulação da Expressão Gênica/fisiologia , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pressão Osmótica , Técnicas de Patch-Clamp , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Canais de Cátion TRPV/genética
20.
Pharm Biol ; 58(1): 208-218, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32114881

RESUMO

Context: It is common sense that chewing a mint leaf can cause a cooling feeling, while chewing ginger root will produce a burning feeling. In Traditional Chinese Medicine (TCM), this phenomenon is referred to as 'cold/hot' properties of herbs. Herein, it is reported that TCM with different "cold/hot" properties have different effects on the variation of cells.Objective: To explore the intrinsic 'cold/hot' properties of TCM from the perspective of cellular and molecular biology.Materials and methods: A375 cells were selected using Cancer Cell Line Encyclopaedia (CCLE) analysis and western blots. Hypaconitine and baicalin were selected by structural similarity analysis from 56 and 140 compounds, respectively. A wireless thermometry system was used to measure cellular temperature change induced by different compounds. Alteration of intracellular calcium influx was investigated by means of calcium imaging.Results: The IC50 values of GSK1016790A, HC067047, hypaconitine, and baicalin for A375 cells are 8.363 nM, 816.4 µM, 286.4 µM and 29.84 µM, respectively. And, 8 µM hypaconitine induced obvious calcium influx while 8 µM baicalin inhibited calcium influx induced by TRPV4 activation. Cellular temperature elevated significantly when treated with GSK1016790A or hypaconitine, while the results were reversed when cells were treated with HC067047 or baicalin.Discussion and conclusions: The changes in cellular temperature are speculated to be caused by the alteration of intracellular calcium influx mediated by TRPV4. In addition, the 'cold/hot' properties of compounds in TCM can be classified by using cellular temperature detection.


Assuntos
Proliferação de Células/efeitos dos fármacos , Medicamentos de Ervas Chinesas/farmacologia , Queratinócitos/efeitos dos fármacos , Termogênese/efeitos dos fármacos , Aconitina/análogos & derivados , Aconitina/farmacologia , Cálcio/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Temperatura Baixa , Flavonoides/farmacologia , Expressão Gênica/efeitos dos fármacos , Temperatura Alta , Humanos , Leucina/análogos & derivados , Leucina/farmacologia , Medicina Tradicional Chinesa/métodos , Morfolinas/farmacologia , Pirróis/farmacologia , Sulfonamidas/farmacologia , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/metabolismo
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