Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 644
Filtrar
1.
Life Sci ; 244: 117306, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31953159

RESUMO

AIMS: Accumulated evidence indicates that the dysregulation of circular RNAs (circRNAs) plays pivotal roles in many human diseases including preeclampsia (PE). Circ_0063517 has been verified to be down-regulated in PE. But the role of circ_0063517 in PE is still unclear. This research aims to probe into the effect of circ_0063517 on angiogenesis in PE development. MAIN METHODS: The expression of circ_0063517, endothelin receptor type B (ETBR) and miR-31-5p was assessed by quantitative reverse transcription polymerase chain reaction (RT-qPCR). MTT assay, colony formation assay, scratch assay, transwell assay, and tube formation assay were performed to detect proliferation, migration, and angiogenesis, respectively. Dual luciferase reporter system and RNA immunoprecipitation (RIP) assay were carried out to determine the interaction between miR-31-5p and circ_0063517(or ETBR). ETBR, VEGFRA, and VEGFR2 levels were detected by western blot analysis. The effect of circ_0063517 and ETBR on angiogenesis was evaluated in N-nitro-L-arginine methyl ester hydrochloride (L-NAME)-induced PE in vivo. KEY FINDINGS: The levels of circ_0063517 and ETBR were down-regulated in the placenta tissue of PE patients. Conversely, the level of miR-31-5p was up-regulated in PE. Overexpression of circ_0063517 or knockdown of miR-31-5p facilitated growth, migration, and angiogenesis of vascular endothelial cells. Circ_0063517 knockdown-induced repression of the expression of ETBR, VEGFA, and VEGFR2 was partly counteracted by ETBR overexpression. Mechanistically, circ_0063517 sponged miR-31-5p to regulate ETBR expression. Finally, circ_0063517 promoted angiogenesis via enhancing ETBR expression in PE in vivo. SIGNIFICANCE: Our findings suggest that circ_0063517-miR-31-5p-ETBR axis regulates angiogenesis during the pathological process of PE.


Assuntos
MicroRNAs/metabolismo , Pré-Eclâmpsia/metabolismo , Receptor de Endotelina B/metabolismo , Animais , Linhagem Celular , Proliferação de Células/fisiologia , Células Endoteliais/metabolismo , Feminino , Células Endoteliais da Veia Umbilical Humana , Humanos , MicroRNAs/genética , Neovascularização Patológica/genética , Pré-Eclâmpsia/genética , Gravidez , Ratos , Ratos Sprague-Dawley
2.
Acta Derm Venereol ; 99(6): 579-586, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30809683

RESUMO

Prurigo nodularis is a highly pruritic and hyperplastic chronic dermatosis with unknown pathogenesis. Many pruritogenic mediators, including nerve growth factor, interleukin (IL)-31, thymic stromal lymphopoietin, and endothelin-1, are implicated in chronic itch and inflammation. This study investigated the mRNA levels and immunoreactivity of the nerve growth factor, IL-31, thymic stromal lymphopoietin, and endothelin axes in both lesional and perilesional skin in prurigo nodularis by using quantitative real-time PCR and immunohistochemistry studies. The nerve growth factor high-affinity receptor tyrosine kinase receptor A was upregulated while the low affinity receptor p75 neurotrophin receptor was downregulated in prurigo nodularis lesions. Downregulated expression of IL-31/IL-31 receptor A and endothelin-3/endothelin receptor B and upregulation of thymic stromal lymphopoietin receptor were found in prurigo nodularis lesions. Aberrant expression of nerve growth factor, IL-31, thymic stromal lymphopoietin and endothelin axes was found in prurigo nodularis lesions, especially in the epidermis, indicating the importance of keratinocytes in prurigo nodularis pathogenesis.


Assuntos
Epiderme/metabolismo , Queratinócitos/metabolismo , Prurigo/genética , Prurigo/metabolismo , Adulto , Citocinas/genética , Citocinas/metabolismo , Regulação para Baixo , Endotelina-3/genética , Endotelina-3/metabolismo , Feminino , Expressão Gênica , Humanos , Hipersensibilidade/complicações , Hipersensibilidade/genética , Hipersensibilidade/metabolismo , Subunidade alfa de Receptor de Interleucina-7/genética , Subunidade alfa de Receptor de Interleucina-7/metabolismo , Interleucinas/genética , Interleucinas/metabolismo , Masculino , Pessoa de Meia-Idade , Fator de Crescimento Neural/genética , Fator de Crescimento Neural/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Prurigo/complicações , RNA Mensageiro/metabolismo , Receptor de Endotelina B/genética , Receptor de Endotelina B/metabolismo , Receptor trkA/genética , Receptor trkA/metabolismo , Receptores de Citocinas/genética , Receptores de Citocinas/metabolismo , Receptores de Interleucina/genética , Receptores de Interleucina/metabolismo , Receptores de Fator de Crescimento Neural/genética , Receptores de Fator de Crescimento Neural/metabolismo , Regulação para Cima
3.
J Pharm Pharmacol ; 71(6): 988-995, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30809816

RESUMO

OBJECTIVES: TGF-ß through hyperelongation of glycosaminoglycan (GAG) chains leads to binding of low-density lipoproteins to the proteoglycans. The vasoactive peptide, endothelin-1 (ET-1), plays a key role in the development of atherosclerosis. This study addressed the question whether ET-1 by activating the Rho kinase and cytoskeletal rearrangement can transactivate the TGF-ß receptor leading to phosphorylation of the transcription factor Smad2 and increased expression of the GAG chain synthesizing enzyme such as chondroitin synthase-1 (CHSY-1) in bovine aortic endothelial cells (BAECs). METHODS: In this study, intermediates in ET-1-induced Smad2C phosphorylation and the protein level of CHSY-1 were identified and quantified by Western blotting. KEY FINDINGS: Endothelin-1 caused time-dependent phosphorylation of Smad2C which was inhibited in the presence of the endothelin B receptor antagonist, BQ788. The response to ET-1 was inhibited by the Rho/ROCK kinase antagonist, Y27632 and by cytochalasin D, an inhibitor of actin polymerization but the ET-1-mediated pSmad2C was not inhibited by the matrix metalloproteinase (MMP) inhibitor, GM6001. ET-1 increased CHSY-1 protein level, which was inhibited in the presence of BQ788, cytochalasin D and Y27632. CONCLUSIONS: Endothelin-1 signalling via the ETB receptor utilizes cytoskeletal rearrangement and Rho kinase but not MMPs leading to TßRI transactivation signalling and phosphorylation of Smad2C and through this pathway increased the level of CHSY-1.


Assuntos
Células Endoteliais/metabolismo , Endotelina-1/metabolismo , N-Acetilgalactosaminiltransferases/genética , Receptor do Fator de Crescimento Transformador beta Tipo I/metabolismo , Amidas/farmacologia , Animais , Aorta/citologia , Western Blotting , Bovinos , Células Cultivadas , Citocalasina D/farmacologia , Oligopeptídeos/farmacologia , Fosforilação/fisiologia , Piperidinas/farmacologia , Piridinas/farmacologia , Receptor de Endotelina B/efeitos dos fármacos , Receptor de Endotelina B/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fatores de Tempo , Ativação Transcricional/fisiologia , Fator de Crescimento Transformador beta/metabolismo , Quinases Associadas a rho/metabolismo
4.
Mol Pharm ; 16(1): 305-317, 2019 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-30422662

RESUMO

ABC transporters act as efflux pumps, thereby influencing the pharmacokinetics and efficacy of many drugs. Zinc (Zn) is an essential trace element contributing to cellular growth and differentiation. It is increasingly recognized as an intracellular messenger. The present study aims at investigating the impact of Zn2+ on the function and regulation of ABC transporters at the blood-brain barrier (BBB). ABC transporter function was first studied in isolated rat brain capillaries. Zn2+ rapidly stimulated the activity of the multidrug resistance-related protein 2 (Mrp2), p-glycoprotein (P-gp), and breast cancer resistance protein (Bcrp). These short-term effects were independent of transporter de novo synthesis but based on Zn2+ triggering intracellular signaling to stimulate basal transport activity. Studies focused on Mrp2 and P-gp showed that Zn2+ induced signaling through an endothelin receptor type B (ETB)/nitric oxide synthase (NOS)/protein kinase C (PKC) pathway and caused, specifically, an activation of the isoform PKCα. Studies revealed signaling through the phosphatidylinositol 3-kinase (PI3K)/mechanistic target of rapamycin (mTOR) pathway, as well as induction of the downstream target serum- and glucocorticoid-inducible kinase 1 (SGK1). Short-term effects of Zn2+ were also demonstrated in human hCMEC/D3 cells. An initial in vivo study in rats suggested enhanced P-gp transport activity at the BBB due to elevated Zn2+ plasma levels. This work provides the first evidence for Zn2+ being a regulator of basal ABC transporter activity at the BBB, driving a rapid and nongenomic stimulation of transport function.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Encéfalo/metabolismo , Zinco/farmacologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Transportadores de Cassetes de Ligação de ATP/genética , Animais , Transporte Biológico/efeitos dos fármacos , Barreira Hematoencefálica/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Endotélio Vascular/citologia , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Humanos , Proteínas Imediatamente Precoces/metabolismo , Proteínas de Neoplasias/metabolismo , Óxido Nítrico Sintase/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Proteína Quinase C/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Ratos , Receptor de Endotelina B/metabolismo , Transdução de Sinais/efeitos dos fármacos
5.
Pain ; 160(3): 619-631, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30422869

RESUMO

Bone cancer metastasis is extremely painful and decreases the quality of life of the affected patients. Available pharmacological treatments are not able to sufficiently ameliorate the pain, and as patients with cancer are living longer, new treatments for pain management are needed. Decitabine (5-aza-2'-deoxycytidine), a DNA methyltransferases inhibitor, has analgesic properties in preclinical models of postsurgical and soft-tissue oral cancer pain by inducing an upregulation of endogenous opioids. In this study, we report that daily treatment with decitabine (2 µg/g, intraperitoneally) attenuated nociceptive behavior in the 4T1-luc2 mouse model of bone cancer pain. We hypothesized that the analgesic mechanism of decitabine involved activation of the endogenous opioid system through demethylation and reexpression of the transcriptionally silenced endothelin B receptor gene, Ednrb. Indeed, Ednrb was hypermethylated and transcriptionally silenced in the mouse model of bone cancer pain. We demonstrated that expression of Ednrb in the cancer cells lead to release of ß-endorphin in the cell supernatant, which reduced the number of responsive dorsal root ganglia neurons in an opioid-dependent manner. Our study supports a role of demethylating drugs, such as decitabine, as unique pharmacological agents targeting the pain in the cancer microenvironment.


Assuntos
Antimetabólitos Antineoplásicos/uso terapêutico , Neoplasias Ósseas/complicações , Dor do Câncer/tratamento farmacológico , Dor do Câncer/etiologia , Decitabina/uso terapêutico , Animais , Densidade Óssea , Neoplasias Ósseas/diagnóstico por imagem , Neoplasias Ósseas/patologia , Dor do Câncer/diagnóstico por imagem , Linhagem Celular Tumoral , Meios de Cultivo Condicionados/farmacologia , Modelos Animais de Doenças , Endotelina-1/metabolismo , Feminino , Gânglios Espinais/citologia , Locomoção/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Naloxona/análogos & derivados , Naloxona/uso terapêutico , Neurônios/efeitos dos fármacos , Compostos de Amônio Quaternário/uso terapêutico , Receptor de Endotelina B/genética , Receptor de Endotelina B/metabolismo , Suporte de Carga/fisiologia , beta-Endorfina/metabolismo
6.
Nat Commun ; 9(1): 4711, 2018 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-30413709

RESUMO

Endothelin receptors (ETA and ETB) are class A GPCRs activated by vasoactive peptide endothelins, and are involved in blood pressure regulation. ETB-selective signalling induces vasorelaxation, and thus selective ETB agonists are expected to be utilized for improved anti-tumour drug delivery and neuroprotection. Here, we report the crystal structures of human ETB receptor in complex with ETB-selective agonist, endothelin-3 and an ETB-selective endothelin analogue IRL1620. The structure of the endothelin-3-bound receptor reveals that the disruption of water-mediated interactions between W6.48 and D2.50 is critical for receptor activation, while these hydrogen-bonding interactions are partially preserved in the IRL1620-bound structure. Consistently, functional analysis reveals the partial agonistic effect of IRL1620. The current findings clarify the detailed molecular mechanism for the coupling between the orthosteric pocket and the G-protein binding, and the partial agonistic effect of IRL1620, thus paving the way for the design of improved agonistic drugs targeting ETB.


Assuntos
Receptor de Endotelina B/química , Receptor de Endotelina B/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Cristalografia por Raios X , Endotelina-3/metabolismo , Endotelinas/química , Endotelinas/metabolismo , Endotelinas/farmacologia , Células HEK293 , Humanos , Modelos Moleculares , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Receptor de Endotelina B/agonistas , Fator de Crescimento Transformador alfa/metabolismo , beta-Arrestinas/metabolismo
7.
Nutr Metab Cardiovasc Dis ; 28(12): 1285-1295, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30392707

RESUMO

BACKGROUND AND AIMS: Intrauterine growth restriction (IUGR) is a major risk factor for perinatal morbidity and mortality, leading to long-term adverse cardiovascular outcomes. The present study aimed to investigate the potential mechanisms in IUGR-associated vascular endothelial dysfunction. METHODS AND RESULTS: Human umbilical vein endothelial cells (HUVECs) were derived from IUGR or normal newborns. We found that the proliferation of IUGR-derived HUVECs was accelerated compared to those from normal subjects. Gene profiles related to vascular function including vasomotion, oxidative stress, and angiogenesis were dysregulated in IUGR-HUVECs. Compared with HUVECs from normal newborns, nitric oxide (NO) production was reduced, with imbalance between endothelial nitric oxide synthase (eNOS) and arginase-2 (Arg-2) in IUGR. Meanwhile, intracellular asymmetric dimethylarginine (ADMA) level was elevated with diminished dimethylarginine dimethylaminohydrolase 1 (DDAH1) expression in IUGR-HUVECs. Furthermore, endothelin-1 (ET-1) and hypoxia-inducible factor 1α (HIF-1α) expression were increased, and endothelin receptor type-B (ETBR) was reduced in the IUGR group. IUGR-HUVECs exposed to hypoxia increased the ratio of ADMA to l-arginine, HIF-1α and protein arginine methyltransferase 1 (PRMT1) expression compared to controls. CONCLUSIONS: The present study demonstrated that the reduction of NO bioavailability and release results from elevated Arg-2, accumulation of intracellular ADMA, and imbalance of ET-1 and ETBR, further leading to IUGR-associated vascular endothelial dysfunction. Our study provides novel evidence on the mechanism underlying fetal programming associated with IUGR, which will serve as potential therapeutic targets in the prevention of adverse cardiovascular consequences in adulthood.


Assuntos
Arginina/metabolismo , Endotelina-1/metabolismo , Retardo do Crescimento Fetal/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Veias Umbilicais/metabolismo , Adulto , Amidoidrolases/genética , Amidoidrolases/metabolismo , Arginase/genética , Arginase/metabolismo , Arginina/análogos & derivados , Estudos de Casos e Controles , Proliferação de Células , Células Cultivadas , Feminino , Retardo do Crescimento Fetal/genética , Retardo do Crescimento Fetal/fisiopatologia , Regulação da Expressão Gênica , Humanos , Recém-Nascido , Masculino , Neovascularização Fisiológica , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , Estresse Oxidativo , Gravidez , Proteína-Arginina N-Metiltransferases/genética , Proteína-Arginina N-Metiltransferases/metabolismo , Receptor de Endotelina B/genética , Receptor de Endotelina B/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transdução de Sinais , Veias Umbilicais/fisiopatologia
8.
Sci Rep ; 8(1): 14698, 2018 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-30279475

RESUMO

LAM is a rare low-grade metastasizing lung neoplasm. Inhibitors of mTOR improve clinical outcome of LAM patients by preventing loss of lung function. Nevertheless, other cell targets may be of interest for drug development. Therefore, we explored the potential role of EDN1 (endothelin) in LAM. We report an increased endothelin blood level in LAM patients as well as EDN1 overexpression and EDN1 receptor downregulation in LAM-derived primary cells and in TSC2NEG cells mutated in TSC2. We evidenced EDN pathway dysregulation based on EDN1, EDNRA, EDNRB and ARRB1 mRNA expression in LAM-derived primary cells. We showed overexpression of EDN1 and ARRB1 mRNAs in TSC2NEG cells; these cells lost their ability to respond to stimulation by endothelin. We analyzed the effects of endothelin receptor antagonists alone or in combination with rapamycin, an mTOR inhibitor, on proliferation and migration of LAM cells. Rapamycin treatment of TSC2NEG cells significantly reduced cell proliferation or migration, while none of the tested inhibitors of EDN receptors impaired these functions. We showed that TSC2NEG cells have acquired a transformed phenotype as showed by their ability to grow as spheroids in semi-solid medium and that unlike endothelin receptors antagonists, rapamycin reduced anchorage-independent cell growth and prevented expansion of TSC2NEG spheroids.


Assuntos
Endotelina-1/metabolismo , Neoplasias Pulmonares/patologia , Linfangioleiomiomatose/patologia , Esclerose Tuberosa/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Antagonistas do Receptor de Endotelina A/farmacologia , Endotelina-1/sangue , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Pulmão/patologia , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/genética , Linfangioleiomiomatose/sangue , Linfangioleiomiomatose/genética , Cultura Primária de Células , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sirolimo/farmacologia , Esferoides Celulares , Esclerose Tuberosa/genética , Proteína 2 do Complexo Esclerose Tuberosa/genética , beta-Arrestina 1/metabolismo
9.
J Mol Biol ; 430(24): 5105-5119, 2018 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-30342934

RESUMO

Extracellular domains of G-protein-coupled receptors act as initial molecular selectivity filters for subtype specific ligands and drugs. Chimeras of the human endothelin-B receptor containing structural units from the extracellular domains of the endothelin-A receptor were analyzed after their co-translational insertion into preformed nanodiscs. A short ß-strand and a linker region in the second extracellular loop as well as parts of the extracellular N-terminal domain were identified as molecular discrimination sites for the endothelin-B receptor-selective agonists IRL1620, sarafotoxin 6c, 4Ala-ET-1 and ET-3, but not for the non-selective agonist ET-1 recognized by both endothelin receptors. A proposed second disulfide bridge in the endothelin-B receptor tethering the N-terminal domain with the third extracellular loop was not essential for ET-1 recognition and binding, but increased the receptor thermostability. We further demonstrate an experimental approach with cell-free synthesized engineered agonists to analyze the differential discrimination of peptide ligand topologies by the two endothelin receptors. The study is based on the engineering and cell-free insertion of G-protein-coupled receptors into defined membranes and may become interesting also for other targets as an alternative platform to reveal molecular details of ligand selectivity and ligand binding mechanisms.


Assuntos
Engenharia de Proteínas/métodos , Receptor de Endotelina A/química , Receptor de Endotelina B/química , Receptor de Endotelina B/metabolismo , Sítios de Ligação , Sistema Livre de Células , Dissulfetos/metabolismo , Endotelinas/farmacologia , Humanos , Ligantes , Modelos Moleculares , Fragmentos de Peptídeos/farmacologia , Domínios Proteicos , Proteínas Recombinantes/metabolismo , Termodinâmica , Venenos de Víboras/farmacologia
10.
Eur J Pharmacol ; 840: 44-49, 2018 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-30240796

RESUMO

Silent information regulator family protein 1 (Sirt1) has gained attention for protective effects against cardiovasc diseases. Vascular smooth muscle endothelin type B (ETB) receptors are related to the pathogenesis of cardiovascular diseases. Elevated oxidized low-density lipoprotein (ox-LDL) is associated with atherosclerosis. This study will investigate whether resveratrol (a Sirt1 activator, Res) is involved in oxidized low density lipoprotein (ox-LDL)-mediated- regulation of ETB receptors in rat superior mesenteric arteries (SMA). The rat SMA segments were cultured in the presence and absence of ox-LDL with or without Res and specific inhibitor (U0126) for the extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) for 24 h. After organ culture, the contractile responses to sarafotoxin 6c (S6c) were studied using a sensitive myograph, and the ETB receptor protein expression was detected using Western blotting. The results showed that Res concentration-dependently suppressed the ox-LDL -induced up-regulation of ETB receptors expression and receptor-mediated vasoconstriction. In addition, these effects could be inhibited by U0126. Furthermore, activity of ERK1/2 phosphorylation and P65 acetylation induced by ox-LDL were blocked by Res. In conclusion, Res down-regulated ETB receptors through up-regulating Sirt1 and followed by ERK1/2/NF-кB signaling pathways in the organ culture SMA.


Assuntos
Regulação para Baixo/efeitos dos fármacos , Músculo Liso Vascular/citologia , Receptor de Endotelina B/metabolismo , Resveratrol/farmacologia , Transdução de Sinais/efeitos dos fármacos , Animais , Lipoproteínas LDL/farmacologia , Masculino , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , NF-kappa B/metabolismo , Ratos , Ratos Sprague-Dawley , Sirtuína 1/metabolismo
11.
Cell Physiol Biochem ; 48(5): 2084-2090, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30099448

RESUMO

BACKGROUND/AIMS: Angiogenesis plays a key role during embryonic development. The vascular endothelin (ET) system is involved in the regulation of angiogenesis. Lipopolysaccharides (LPS) could induce angiogenesis. The effects of ET blockers on baseline and LPS-stimulated angiogenesis during embryonic development remain unknown so far. METHODS: The blood vessel density (BVD) of chorioallantoic membranes (CAMs), which were treated with saline (control), LPS, and/or BQ123 and the ETB blocker BQ788, were quantified and analyzed using an IPP 6.0 image analysis program. Moreover, the expressions of ET-1, ET-2, ET3, ET receptor A (ETRA), ET receptor B (ETRB) and VEGFR2 mRNA during embryogenesis were analyzed by semi-quantitative RT-PCR. RESULTS: All components of the ET system are detectable during chicken embryogenesis. LPS increased angiogenesis substantially. This process was completely blocked by the treatment of a combination of the ETA receptor blockers-BQ123 and the ETB receptor blocker BQ788. This effect was accompanied by a decrease in ETRA, ETRB, and VEGFR2 gene expression. However, the baseline angiogenesis was not affected by combined ETA/ETB receptor blockade. CONCLUSION: During chicken embryogenesis, the LPS-stimulated angiogenesis, but not baseline angiogenesis, is sensitive to combined ETA/ETB receptor blockade.


Assuntos
Antagonistas do Receptor de Endotelina B/farmacologia , Lipopolissacarídeos/farmacologia , Neovascularização Fisiológica/efeitos dos fármacos , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Animais , Galinhas , Membrana Corioalantoide/efeitos dos fármacos , Membrana Corioalantoide/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Endotelina-1/genética , Endotelina-1/metabolismo , Oligopeptídeos/farmacologia , Peptídeos Cíclicos/farmacologia , Piperidinas/farmacologia , Receptor de Endotelina A/química , Receptor de Endotelina A/genética , Receptor de Endotelina B/química , Receptor de Endotelina B/genética , Transdução de Sinais/efeitos dos fármacos , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
12.
J Mol Med (Berl) ; 96(9): 975-982, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30069745

RESUMO

Oxygen plays a central role in cardiac energy metabolism. At high altitude where the ambient oxygen level is low, we found EDNRB is associated with human hypoxia adaptation. Our subsequent study in global heterozygous knockout mice (Ednrb-/+) revealed that cardiac function was conserved in these mice when exposed to extreme hypoxia. The major goal of this study was (i) to determine the functional role of cardiomyocyte EdnrB in maintaining cardiac function under hypoxic stress and (ii) to validate the phenotypes we detected in Ednrb-/+ mice using EDNRB blockers. Unlike the global knockouts, cardiac-specific heterozygote (EdnrBflox/+) and homozygote (EdnrBflox/flox) EdnrB knockout mice were phenotypically normal. When treated with graded low levels of oxygen (10% and 5% O2), both EdnrBflox/+ and EdnrBflox/flox were hypoxia tolerant. The cardiac indexes at 10% and 5% O2 for EdnrBflox/+ were significantly higher and lactate levels were significantly lower when compared to the cre-negative controls (P < 0.05). Simultaneously, mice treated with BQ-788 (EDNRB-specific blocker) had a significantly higher cardiac index (P < 0.005) and significantly lower lactate levels (P < 0.0001) than in control mice. A similar result was obtained with mice treated with Bosentan (non-specific). These data indicate that a lower level or complete lack of EdnrB in the cardiomyocytes significantly improves cardiac performance under extreme hypoxia, a novel role of cardiomyocyte EdnrB in the regulation of cardiac function. Furthermore, this rescue under extreme hypoxia can also be achieved using EDNRB-specific pharmacological agents, e.g., BQ-788. This systematically confirms, both genetically and pharmacologically, the protective role of a lower EDNRB under extreme hypoxia stress. KEY MESSAGES: Under normal condition, cardiomyocytes-specific EdnrB knockout mice, both heterozygote and homozygote, are phenotypically normal. Under hypoxic condition, a lower level or complete deletion of cardiomyocyte EdnrB conserves cardiac function by maintaining high cardiac index. Similarly, mice treated with both specific (BQ-788) and non-specific (Bosentan) EDNRB blockers are tolerant to hypoxia by maintaining better cardiac function. The oxygen perfusion under extreme hypoxia is better in the mice with lower EDNRB, as depicted by lower lactate level at 5% oxygen. Our current study systematically confirms, both genetically and pharmacologically, the protective role of a lower EDNRB under extreme hypoxia stress. Overall, it supports our hypothesis that studies on human hypoxia adaptation provide new insight to common disease pathogenesis and treatments.


Assuntos
Antagonistas do Receptor de Endotelina B/farmacologia , Hipóxia/genética , Hipóxia/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Receptor de Endotelina B/genética , Receptor de Endotelina B/metabolismo , Adaptação Fisiológica , Animais , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Marcação de Genes , Loci Gênicos , Camundongos , Camundongos Knockout , Oxigênio/metabolismo , Fenótipo , Sensibilidade e Especificidade
13.
Pflugers Arch ; 470(12): 1815-1827, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30094478

RESUMO

Complex regional pain syndrome (CRPS) is a disorder that involves abnormal inflammation and nerve dysfunction frequently resistant to a broad range of treatments. Peripheral nerve stimulation with electroacupuncture (EA) has been widely used in different clinical conditions to control pain and inflammation; however, the use of EA in the treatment of CRPS is under investigation. In this study, we explore the effects of EA on hyperalgesia and edema induced in an animal model of chronic post-ischemia pain (CPIP model) and the possible involvement of endothelin receptor type B (ETB) in this effect. Female Swiss mice were subjected to 3 h hind paw ischemia/reperfusion CPIP model. EA treatment produced time-dependent inhibition of mechanical and cold hyperalgesia, as well as edema in CPIP mice. Peripheral administration (i.pl.) of BQ-788 (10 nmol), an ETB antagonist, prevented EA-induced antihyperalgesia while intrathecal administration prolonged EA's effect. Additionally, peripheral pre-treatment with sarafotoxin (SRTX S6c, 30 pmol, ETB agonist) increased EA anti-hyperalgesic effect. Furthermore, the expression of peripheral ETB receptors was increased after EA treatments, as measured by western blot. These results may suggest that EA's analgesic effect is synergic with ETB receptor activation in the periphery, as well as central (spinal cord) ETB receptor blockade. These data support the use of EA as a nonpharmacological approach for the management of CRPS-I, in an adjuvant manner to ETB receptor targeting drugs.


Assuntos
Síndromes da Dor Regional Complexa/terapia , Eletroacupuntura/métodos , Hiperalgesia/terapia , Receptor de Endotelina B/metabolismo , Animais , Síndromes da Dor Regional Complexa/metabolismo , Antagonistas do Receptor de Endotelina B/administração & dosagem , Antagonistas do Receptor de Endotelina B/farmacologia , Feminino , Hiperalgesia/metabolismo , Camundongos , Oligopeptídeos/administração & dosagem , Oligopeptídeos/farmacologia , Nervos Periféricos/efeitos dos fármacos , Piperidinas/administração & dosagem , Piperidinas/farmacologia , Receptor de Endotelina B/agonistas , Medula Espinal/efeitos dos fármacos , Venenos de Víboras/administração & dosagem , Venenos de Víboras/farmacologia
14.
Exp Eye Res ; 176: 207-209, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30025919

RESUMO

PURPOSE: Whilst the pathogenesis of retinal vein occlusion (RVO) is still unclear, systemic hypertension and increased level of endothelin-1 (ET-1) are known risk factors. Therefore, we studied the influence of ET-1 on the retinal veins in hypertensive rats. METHODS: We focused on the behavior of retinal veins in spontaneous hypertensive rats (SHR). To determine whether ET-1 was associated with the blood flow in eyes of SHRs, the chorioretinal blood flow in the rats was assessed using laser speckle flowgraphy (LSFG-Micro, Softcare, Fukuoka, Japan) before and after an intravenous injection of ET-1 under general anesthesia. In addition, retinas from SHRs and age-matched normotensive Wistar-Kyoto rats (WKYs) were removed, and retinal sections were immunostained for the ET-A and ET-B receptors. The protein levels of both ET-1 receptors and hypoxia-inducible factor 1 (HIF-1) in the retinal tissues were also determined by western blot analysis. RESULTS: One of the retinal veins became exceptionally constricted and was nearly occluded, and the chorioretinal blood flow significantly decreased in the retinas of SHRs following the injection of ET-1. Immunoreactivity to ET-A receptor was higher in SHR retinas than in WKY retinas. The protein levels of ET-A receptor and HIF-1 were also significantly higher in SHR retinas than in WKY retinas. CONCLUSIONS: An increase of ET-1 in circulating blood leads to the local constriction of retinal veins and this effect is accentuated in hypertensive rats by an upregulation of ET-A receptor. It is plausible that such a constriction of retinal veins increases retinal venous pressure, and may even contribute to the pathogenesis of RVO.


Assuntos
Endotelina-1/administração & dosagem , Oclusão da Veia Retiniana/etiologia , Oclusão da Veia Retiniana/fisiopatologia , Veia Retiniana/fisiopatologia , Animais , Corioide/irrigação sanguínea , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Injeções Intravenosas , Fluxometria por Laser-Doppler , Masculino , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Fluxo Sanguíneo Regional/fisiologia , Oclusão da Veia Retiniana/metabolismo , Vasos Retinianos/fisiologia , Vasoconstrição
15.
Physiol Res ; 67(Suppl 1): S83-S94, 2018 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-29947530

RESUMO

The global epidemic of diabetes is of significant concern. Diabetes associated vascular disease signifies the principal cause of morbidity and mortality in diabetic patients. It is also the most rapidly increasing risk factor for cognitive impairment, a silent disease that causes loss of creativity, productivity, and quality of life. Small vessel disease in the cerebral vasculature plays a major role in the pathogenesis of cognitive impairment in diabetes. Endothelin system, including endothelin-1 (ET-1) and the receptors (ET(A) and ET(B)), is a likely candidate that may be involved in many aspects of the diabetes cerebrovascular disease. In this review, we took a brain-centric approach and discussed the role of the ET system in cerebrovascular and cognitive dysfunction in diabetes.


Assuntos
Encéfalo/metabolismo , Complicações do Diabetes/metabolismo , Endotelinas/metabolismo , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Animais , Encéfalo/efeitos dos fármacos , Circulação Cerebrovascular/efeitos dos fármacos , Circulação Cerebrovascular/fisiologia , Complicações do Diabetes/tratamento farmacológico , Antagonistas do Receptor de Endotelina A/administração & dosagem , Antagonistas do Receptor de Endotelina A/metabolismo , Antagonistas do Receptor de Endotelina B/administração & dosagem , Antagonistas do Receptor de Endotelina B/metabolismo , Endotelinas/agonistas , Endotelinas/antagonistas & inibidores , Humanos
16.
Physiol Res ; 67(Suppl 1): S95-S113, 2018 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-29947531

RESUMO

Endothelin B (ET(B)) receptors present in abundance the central nervous system (CNS) have been shown to have significant implications in its development and neurogenesis. We have targeted ET(B) receptors stimulation using a highly specific agonist, IRL-1620, to treat CNS disorders. In a rat model of cerebral ischemia intravenous administration IRL-1620 significantly reduced infarct volume and improved neurological and motor functions compared to control. This improvement, in part, is due to an increase in neuroregeneration. We also investigated the role of IRL-1620 in animal models of Alzheimer's disease (AD). IRL-1620 improved learning and memory, reduced oxidative stress and increased VEGF and NGF in Abeta treated rats. IRL-1620 also improved learning and memory in an aged APP/PS1 transgenic mouse model of AD. These promising findings prompted us to initiate human studies. Successful chemistry, manufacturing and control along with mice, rat and dog toxicological studies led to completion of a human Phase I study in healthy volunteers. We found that a dose of 0.6 microg/kg of IRL-1620 can be safely administered, three times every four hours, without any adverse effect. A Phase II clinical study with IRL-1620 has been initiated in patients with cerebral ischemia and mild to moderate AD.


Assuntos
Endotelinas/uso terapêutico , Regeneração Nervosa/efeitos dos fármacos , Doenças Neurodegenerativas/tratamento farmacológico , Fragmentos de Peptídeos/uso terapêutico , Receptor de Endotelina B/agonistas , Animais , Doenças do Sistema Nervoso Central/tratamento farmacológico , Doenças do Sistema Nervoso Central/metabolismo , Ensaios Clínicos como Assunto/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Endotelinas/farmacologia , Humanos , Regeneração Nervosa/fisiologia , Doenças Neurodegenerativas/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Fragmentos de Peptídeos/farmacologia , Receptor de Endotelina B/metabolismo
17.
Physiol Res ; 67(Suppl 1): S149-S154, 2018 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-29947535

RESUMO

Renal medullary endothelin B receptors (ET(B)) mediate sodium excretion and blood pressure (BP) control. Several animal models of hypertension have impaired renal medullary ET(B) function. We found that 4-week high-caloric diet elevated systolic BP in Dahl salt-sensitive (Dahl S) rats (126+/-2 vs. 143+/-3 mm Hg, p<0.05). We hypothesized that renal medullary ET(B) function is dysfunctional in DS rats fed a high-caloric diet. We compared the diuretic and natriuretic response to intramedullary infusion of ET(B) agonist sarafotoxin 6c (S6c) in DS rats fed either a normal or high-caloric diet for 4 weeks. Urine was collected during intramedullary infusion of saline for baseline collection followed by intramedullary infusion of either saline or S6c. We first examined the ET(B) function in DS rats fed a normal diet. S6c increased urine flow (2.7+/-0.3 microl/min during baseline vs. 5.1+/-0.6 microl/min after S6c; p<0.05; n=5) and sodium excretion (0.28+/-0.05 vs. 0.81+/-0.17 micromol/min; p<0.05), suggesting that DS rats have renal medullary ET(B) function. However, DS rats fed a high-caloric diet displayed a significant increase in urine flow (2.7+/-0.4 vs. 4.2+/-0.4 microl/min, baseline vs. S6c infusion, respectively; p<0.05, n=6), but no significant change in sodium excretion in response to S6c (0.32+/-0.06 vs. 0.45+/-0.10 micromol/min). These data demonstrate that renal medullary ET(B) function is impaired in DS rats fed a high-caloric diet, which may be contributed to the elevation of blood pressure during high-caloric feeding in this model.


Assuntos
Dieta Hiperlipídica/efeitos adversos , Ingestão de Alimentos/fisiologia , Hipertensão/metabolismo , Medula Renal/metabolismo , Receptor de Endotelina B/agonistas , Receptor de Endotelina B/metabolismo , Animais , Pressão Sanguínea/efeitos dos fármacos , Pressão Sanguínea/fisiologia , Ingestão de Alimentos/efeitos dos fármacos , Hipertensão/etiologia , Medula Renal/efeitos dos fármacos , Masculino , Peptídeos/farmacologia , Ratos , Ratos Endogâmicos Dahl , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Venenos de Víboras
18.
Physiol Res ; 67(Suppl 1): S257-S264, 2018 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-29947545

RESUMO

The endothelin axis (endothelins and their receptors) is strongly involved in physiological and pathological processes. ET-1 plays a crucial role in particular in tumor diseases. Endothelin-1 receptors (ET(A) and ET(B)) are deregulated and overexpressed in several tumors such as melanoma and glioma. We studied the binding of 24 monoclonal antibodies directed against human ET(B) receptors (hET(B)) to different melanoma cell lines. Few of these mAbs bound to all the melanoma cell lines. One of them, rendomab B49, bound to ET(B) receptors expressed at the surface of human glioma stem cells. More recently, we produced new antibodies directed against human ET(A) receptor (hET(A)). Several antibodies have been isolated and have been screened on different tumoral cells lines. As for the mAbs directed against the hET(B) receptor only some of new antibodies directed against ET(A) receptor are capable to bind the human tumoral cell lines. Rendomab A63 directed against hET(A) is one of them. We report the specificity and binding properties of these mAbs and consider their potential use in diagnosis by an in vivo imaging approach.


Assuntos
Anticorpos Monoclonais/metabolismo , Antagonistas do Receptor de Endotelina A/metabolismo , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/administração & dosagem , Células CHO , Linhagem Celular Tumoral , Cricetinae , Cricetulus , Antagonistas do Receptor de Endotelina A/administração & dosagem , Endotelina-1/genética , Endotelina-1/metabolismo , Feminino , Humanos , Camundongos , Camundongos Nus , Ligação Proteica/fisiologia , Receptor de Endotelina A/genética , Receptor de Endotelina B/genética , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
19.
Biomed Pharmacother ; 99: 704-714, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29710468

RESUMO

The aim of this study was to investigate the role of the ß-adrenergic blocker bucindolol on endothelial dysfunction and pulmonary vascular remodeling in rats with pulmonary arterial hypertension (PAH). Male Wistar rats were divided into four groups: control, monocrotaline (MCT), control?+?bucindolol and monocrotaline?+?bucindolol (MCT?+?BCD). PAH was induced by an injection of monocrotaline (60?mg/kg i.p.). After two weeks, the animals were treated for seven days with bucindolol (2?mg/kg/day i.p.) or vehicle. Echocardiography was performed upon treatment completion to analyze pulmonary vascular resistance (PVR) and right ventricle (RV) myocardial performance index. Lungs were collected for oxidative stress and western blot analysis, and the pulmonary artery was analyzed for histological and immunohistochemical parameters. The MCT?+?BCD group showed a decrease (32%) in the protein expression of endothelin-1 type A receptor (ETAR) and in the ratio of ETA/endothelin-1 type B receptor (ETBR) (62%) as compared to the MCT group. Bucindolol treatment did not alter oxidative stress, as determined by lipid peroxidation analysis and antioxidant enzyme activities and expression, endothelial nitric oxide synthase immunocontent and decreased nitrotyrosine levels. Moreover, bucindolol improved vascular remodeling of the pulmonary artery in the MCT?+?BCD group by decreasing (21%) PVR and increasing RV workload in relation to MCT.


Assuntos
Antagonistas Adrenérgicos beta/farmacologia , Hipertensão Pulmonar/tratamento farmacológico , Propanolaminas/farmacologia , Artéria Pulmonar/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Ecocardiografia , Hipertensão Pulmonar/fisiopatologia , Masculino , Monocrotalina/toxicidade , Óxido Nítrico Sintase Tipo III , Estresse Oxidativo/efeitos dos fármacos , Artéria Pulmonar/metabolismo , Ratos , Ratos Wistar , Receptor de Endotelina A/efeitos dos fármacos , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/efeitos dos fármacos , Receptor de Endotelina B/metabolismo , Remodelação Vascular/efeitos dos fármacos
20.
Ann Vasc Surg ; 52: 207-215, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29758325

RESUMO

Autophagy plays an important role in cardiovascular diseases. High glucose (HG) upregulates endothelin subtype B (ETB) receptors in vascular smooth muscle cells (VSMCs). However, it is unclear as to whether autophagy is involved in HG-induced upregulation of ETB receptors in VSMCs. The present study was designed to examine the hypothesis that HG upregulates ETB receptors by inhibiting autophagy in VSMCs. We studied HG-treated rat superior mesenteric artery (SMA) without endothelium in the presence and absence of 5-aminoimidazole-4-carboxamide 1-ß-D-ribofuranoside (AICAR), rapamycin, or MHY1485 for 24 hr. We measured contractile responses to sarafotoxin 6c (S6c) (an ETB receptor agonist) using a sensitive myograph. Levels of protein expression were determined using Western blotting. HG impaired autophagy and increased the levels of ETB receptor protein expression and ETB receptor-mediated contractile responses to S6c in SMA. However, these effects were reversed by AICAR (an agonist of adenosine monophosphate [AMP]-activated protein kinase [AMPK]) and rapamycin (an inhibitor of mammalian target of rapamycin [mTOR]). However, MHY1485 (an agonist of mTOR) did not upregulate the AICAR-inhibited ETB receptor-mediated contractile responses or ETB receptor protein expression in the presence of HG. These data suggest that HG upregulated ETB receptors by inhibiting autophagy in VSMCs via AMPK and mTOR signaling pathways.


Assuntos
Autofagia/efeitos dos fármacos , Glucose/toxicidade , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Receptor de Endotelina B/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Masculino , Artéria Mesentérica Superior/efeitos dos fármacos , Artéria Mesentérica Superior/metabolismo , Artéria Mesentérica Superior/patologia , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Técnicas de Cultura de Órgãos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Regulação para Cima , Vasoconstrição/efeitos dos fármacos
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA