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1.
Eur J Pharmacol ; 843: 96-103, 2019 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-30352200

RESUMO

It has been documented cardiac fibroblasts as the predominant cell population undergoing senescence in heart. Calcitonin gene-related peptide (CGRP) exhibits a wide range of cardiovascular protective effects. Whether CGRP protects against cardiac fibroblasts senescence in cardiac fibrosis remains unknown. Here, we detected the down-regulation of CGRP concomitant with senescence in fibrotic myocardium, both hypertension- induced left ventricular fibrosis in SHR rats and hypoxia-induced right ventricular fibrosis in pulmonary artery hypertension rats. Exogenous CGRP inhibited the cardiac fibroblasts senescence and senescence-associated secretory phenotype (SASP) induced by TGF-ß1, which was abolished by CGRP8-37, a selective CGRP receptor antagonist. Moreover, the expression of klotho, an anti-senescence protein, was down-regulated in fibrotic myocardium, and CGRP up-regulated the klotho expression in TGF-ß1-treated cardiac fibroblasts. Klotho knockdown by siRNA reversed the inhibition of CGRP on senescence and SASP induced by TGF-ß1 in cardiac fibroblasts. These results suggested that CGRP inhibited the cardiac fibroblasts senescence and SASP in cardiac fibrosis via up-regulating klotho expression.


Assuntos
Peptídeo Relacionado com Gene de Calcitonina/farmacologia , Senescência Celular/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Glucuronidase/metabolismo , Animais , Animais Recém-Nascidos , Fibroblastos/fisiologia , Fibrose , Glucuronidase/genética , Masculino , Miocárdio/citologia , Miocárdio/metabolismo , Miocárdio/patologia , RNA Interferente Pequeno , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Ratos Sprague-Dawley , Fator de Crescimento Transformador beta1/farmacologia , Regulação para Cima
2.
Pharmacol Res ; 142: 294-302, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30553824

RESUMO

Stress response refers to the systemic nonspecific response upon exposure to strong stimulation or chronic stress, such as severe trauma, shock, infection, burn, major surgery or improper environment, which disturb organisms and damage their physical and psychological health. However, the pathogenesis of stress induced disorder remains complicated and diverse under different stress exposure. Recently, studies have revealed a specific role of microRNAs (miRNAs) in regulating cellular function under different types of stress, suggesting a significant role in the treatment and prevention of stress-related diseases, such as stress ulcer, posttraumatic stress disorder, stress-induced cardiomyopathy and so on. This paper have reviewed the literature on microRNA related stress diseases in different databases including PubMed, Web of Science, and the MiRbase. It considers only peer-reviewed papers published in English between 2004 and 2018. This review summarizes new advances in principles and mechanisms of miRNAs regulating stress signalling pathway and the role of miRNAs in human stress diseases. This comprehensive review is to provide an integrated account of how different stresses affect miRNAs and how stress-miRNA pathways may, in turn, be linked with disease, which offers some potential strategies for stress disorder treatment. Furthermore, the limitation of current studies and challenges for clinical use are discussed.

3.
Cancer ; 123(14): 2634-2641, 2017 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-28301684

RESUMO

BACKGROUND: Nivolumab is a new standard of care for patients with metastatic renal cell carcinoma (mRCC) and provides an overall survival benefit of 5.40 months in comparison with everolimus. This study evaluated the cost-effectiveness of nivolumab for the second-line treatment of mRCC from the perspective of US payers and identified the range of drug costs for which the addition of nivolumab to standard therapy could be considered cost-effective from a Chinese perspective. METHODS: A partitioned survival model was constructed to estimate lifetime costs, life-years, and quality-adjusted life-years (QALYs). Costs were estimated for the US and Chinese health care systems. One-way and probabilistic sensitivity analyses were performed. RESULTS: Nivolumab provided an additional 0.29 QALYs at a cost of $151,676/QALY in the United States. The probabilistic sensitivity analysis showed that at a willingness-to-pay threshold of $100,000/QALY, at the current cost of nivolumab, the chance of nivolumab being cost-effective was 3.10%. For China, when nivolumab cost less than $7.90 or $9.70/mg, there was a nearly 90% likelihood that the incremental cost-effectiveness ratio for nivolumab would be less than $22,785 or $48,838/QALY, respectively. CONCLUSIONS: For the United States, nivolumab is unlikely to be a high-value treatment for mRCC at the current price, and a price reduction appears to be justified. In China, value-based prices for nivolumab are $7.90 and $9.70/mg for the country and Beijing City, respectively. This study could and should inform the multilateral drug-price negotiations in China that may be upcoming for nivolumab. Cancer 2017;123:2634-41. © 2017 American Cancer Society.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/uso terapêutico , Carcinoma de Células Renais/tratamento farmacológico , Neoplasias Renais/tratamento farmacológico , Anticorpos Monoclonais/economia , Antineoplásicos/economia , Carcinoma de Células Renais/patologia , China , Análise Custo-Benefício , Custos e Análise de Custo , Intervalo Livre de Doença , Custos de Medicamentos , Everolimo/economia , Everolimo/uso terapêutico , Humanos , Neoplasias Renais/patologia , Modelos Econômicos , Nivolumabe , Anos de Vida Ajustados por Qualidade de Vida , Estados Unidos
4.
Can J Physiol Pharmacol ; 94(12): 1315-1324, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27556497

RESUMO

We have found that eIF3a plays an important role in bleomycin-induced pulmonary fibrosis, and up-regulation of eIF3a induced by TGF-ß1 is mediated via the ERK1/2 pathway. Whether ERK1/2 - eIF3a signal pathway is involved in calcitonin gene-related peptide (CGRP)-mediated pathogenesis of bleomycin-induced pulmonary fibrosis remains unknown. Pulmonary fibrosis was induced by intratracheal instillation of bleomycin (5 mg/kg) in rats. Primary pulmonary fibroblasts were cultured to investigate the proliferation by BrdU incorporation method and flow cytometry. Sensory CGRP depletion by capsaicin exacerbated bleomycin-induced pulmonary fibrosis in rats, as shown by a significant disturbed alveolar structure, marked thickening of the interalveolar septa and dense interstitial infiltration by inflammatory cells and fibroblasts, accompanied with increased expression of TGF-ß1, eIF3a, phosphorylated ERK1/2, α-SMA, collagen I, and collagen III. Exogenous application of CGRP significantly inhibited TGF-ß1-induced proliferation and differentiation of pulmonary fibroblasts concomitantly with decreased expression of eIF3a, phosphorylated ERK1/2, α-SMA, collagen I, and collagen III. These effects of CGRP were abolished in the presence of CGRP8-37. These results suggest that endogenous CGRP is related to the development of pulmonary fibrosis induced by bleomycin, and the inhibitory effect of CGRP on proliferation of lung fibroblasts involves the ERK1/2 - eIF3a signaling pathway.


Assuntos
Bleomicina/toxicidade , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Peptídeo Relacionado com Gene de Calcitonina/farmacologia , Regulação para Baixo/efeitos dos fármacos , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/metabolismo , Animais , Antibióticos Antineoplásicos/toxicidade , Peptídeo Relacionado com Gene de Calcitonina/uso terapêutico , Células Cultivadas , Regulação para Baixo/fisiologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Masculino , Fibrose Pulmonar/tratamento farmacológico , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley
5.
Eur J Pharmacol ; 784: 42-8, 2016 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-27164423

RESUMO

Glutamate is a versatile molecule existing in both the central nervous system and peripheral organs. Previous studies have mainly focussed on the biological effect of glutamate in the brain. Recently, abundant evidence has demonstrated that glutamate also participates in the regulation of physiopathological functions in peripheral tissues, including the lung, kidney, liver, heart, stomach and immune system, where the glutamate/glutamate receptor/glutamate transporter system plays an important role in the pathogenesis of certain diseases, such as myocardial ischaemia/reperfusion injury and acute gastric mucosa injury. All these findings provide new insight into the biology and pharmacology of glutamate and suggest a potential therapeutic role of glutamate in non-neurological diseases.


Assuntos
Ácido Glutâmico/metabolismo , Animais , Proteínas de Transporte de Glutamato da Membrana Plasmática/metabolismo , Humanos , Especificidade de Órgãos , Receptores de Glutamato/metabolismo
6.
Naunyn Schmiedebergs Arch Pharmacol ; 389(7): 757-67, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27052575

RESUMO

Rutaecarpine has been shown to exhibit wide pharmacological effects in the cardiovascular system via stimulation of calcitonin gene-related peptide (CGRP) release. In the present study, the effect of rutaecarpine on hypoxia-induced right ventricular (RV) remodeling and the underlying mechanisms were evaluated. RV remodeling was induced by hypoxia (10 % O2, 3 weeks) in rats. Rats were treated with rutaecarpine (20 or 40 mg/kg) by intragastric administration. Proliferation of cardiac fibroblasts was induced by TGF-ß1 (5 ng/mL) and determined by MTS and EdU incorporation method. Cardiac fibroblasts were treated with exogenous CGRP (10 or 100 nM). The concentrations of CGRP and TGF-ß1 in plasma were measured by ELISA. The expression of eIF3a, p27, α-SMA, collagen-I/III, ANP, and BNP were measured by real-time PCR or western blot. Hypoxia induced an increase of right ventricle systolic pressure (RVSP), ration of RV/LV+S, and RV/tibial length in rats, while cardiac hypertrophy, apoptosis, and fibrosis were detected. The expression of ANP, BNP, α-SMA, collagen-I, collagen-III, eIF3a, and TGF-ß1 was up-regulated, and the expression of p27 was down-regulated in the right ventricle of hypoxia-treated rats. The plasma concentration of CGRP was decreased and TGF-ß1 was increased in hypoxia-treated rats. All of these effects induced by hypoxia were attenuated by rutaecarpine in a dose-dependent manner. In cultured cardiac fibroblasts, TGF-ß1 significantly promoted the proliferation and up-regulated the expression of α-SMA and collagen-I/III, while the expression of eIF3a was up-regulated and the expression of p27 was down-regulated. The effects of TGF-ß1 were attenuated by CGRP. CGRP8-37, a selective CGRP receptor antagonist, abolished the effects of CGRP. Rutaecarpine attenuates hypoxia-induced RV remodeling via stimulation of CGRP release, and the effects of rutaecarpine involve the eIF3a/p27 pathway.


Assuntos
Hipertrofia Ventricular Direita/prevenção & controle , Hipóxia/tratamento farmacológico , Alcaloides Indólicos/farmacologia , Quinazolinas/farmacologia , Disfunção Ventricular Direita/prevenção & controle , Função Ventricular Direita/efeitos dos fármacos , Remodelação Ventricular/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Fator de Iniciação 3 em Eucariotos/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose , Hipertrofia Ventricular Direita/etiologia , Hipertrofia Ventricular Direita/metabolismo , Hipertrofia Ventricular Direita/fisiopatologia , Hipóxia/complicações , Hipóxia/metabolismo , Hipóxia/fisiopatologia , Miocárdio/metabolismo , Miocárdio/patologia , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/metabolismo , Disfunção Ventricular Direita/etiologia , Disfunção Ventricular Direita/metabolismo , Disfunção Ventricular Direita/fisiopatologia , Pressão Ventricular/efeitos dos fármacos
7.
Life Sci ; 147: 117-24, 2016 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-26827991

RESUMO

AIMS: Activation of hypoxia inducible factor-1 (HIF) is a hallmark in hypoxia-induced pulmonary hypertension (HPH). microRNAs play a significant role in regulating proliferation of pulmonary arterial smooth muscle cells (PASMCs) in pulmonary hypertension. Previous studies have shown that HIF-1ß is a target of miR-103/107. In this present study, we aimed to investigate whether miR-103/107 regulate vascular remodeling in HPH via HIF-1ß. MAIN METHODS: The HPH model was built by hypoxia exposure in rats. Real-time PCR and Western blotting were used to determine the expression of miR-103/107 and HIF-1ß. Proliferation of PASMCs was examined by 5-bromo-2'-deoxyuridine (BrdU) incorporation method. The functions of miR-103/107 on PASMCs proliferation, HIF-1α and HIF-1ß expression were assessed by transfecting miR-103/107 mimics and inhibitors. KEY FINDINGS: Significant down-regulation of miR-103/107 was observed in remodeled intrapulmonary vascular in HPH rats and hypoxia-exposured PASMCs, whereas HIF-1α and HIF-1ß expression were up-regulated. Hypoxia exposure induced significant proliferation of PASMCs, overexpression of miR-103/107 inhibited but miR-103/107 inhibitors exacerbated PASMCs proliferation. Gain-of-function and loss-of-function experiments showed that miR-103/107 expression was inversely correlated with HIF-1ß level. No significant changes of HIF-1α expression were observed under miR-103/107 mimic treatment. SIGNIFICANCE: Loss of suppression on HIF-1ß by miR-103/107 may contribute to excess proliferation of PASMCs and vascular remodeling in hypoxic pulmonary hypertension.


Assuntos
Translocador Nuclear Receptor Aril Hidrocarboneto/metabolismo , Proliferação de Células/fisiologia , Hipertensão Pulmonar/fisiopatologia , MicroRNAs/genética , Animais , Translocador Nuclear Receptor Aril Hidrocarboneto/genética , Hipóxia Celular , Regulação para Baixo , Hipertensão Pulmonar/genética , Masculino , Miócitos de Músculo Liso/metabolismo , Artéria Pulmonar/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Transfecção , Regulação para Cima , Remodelação Vascular/fisiologia
8.
Curr Med Chem ; 23(8): 763-73, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26861004

RESUMO

Calcitonin gene-related peptide (CGRP) is extensively distributed throughout the central and peripheral nervous systems and has been shown to be a 37 amino acid multifunctional neuropeptide involved in a wide range of physiological and pathological processes. Recently, there is increasing evidence suggesting that CGRP also exists in non-nerve cells, such as epithelial cells, endothelial cells, endothelial progenitor cells (EPCs), T lymphocytes, B lymphocytes, peripheral blood mononuclear cells (PBMCs), and adipocytes. The existence of CGRP in non-neural tissue is of great importance to the regulation of multiple physiological and pathological processes via different pathways, especially through an autocrine/paracrine mode. This review integrates evidence from recent developments and aims to provide novel insights into non-neural sources of CGRP and its effects on physiological and pathological processes.


Assuntos
Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Células Endoteliais/metabolismo , Células Epiteliais/metabolismo , Linfócitos/metabolismo , Humanos
9.
Eur J Pharmacol ; 773: 42-50, 2016 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-26821114

RESUMO

Fluorofenidone is a novel derivative of l-mimosine. It has remarkable anti-fibrotic properties. In this study, we established that fluorofenidone ameliorates pulmonary fibrosis (PF) both in vivo and in vitro by specifically inhibiting the expression of eukaryotic translation initiation factor 3a (eIF3a). eIF3a plays an important role in the development and progression of PF. An animal model of PF was induced by intratracheal instillation of bleomycin (5mg/kg) in rats. Rats were orally administered with fluorofenidone (250, 500 mg/kg/d·[i.g.]) and pirfenidone (500 mg/kg/d·[i.g.]) for 28 days. Primary pulmonary fibroblasts were cultured to determine the effect of fluorofenidone on TGF-ß1-induced (5 ng/ml) proliferation and differentiation of fibroblasts. The expression/level of eIF3a, TGF-ß1, α-SMA, collagen I, and collagen III were analyzed by ELISA, real-time PCR, and western blot. The cell proliferation rate was determined by MTS assay. The results indicate that fluorofenidone significantly improves the pathological changes in lung tissues and reduces the deposition of collagen by inhibiting eIF3a in rats with bleomycin-induced PF. Moreover, in a culture of pulmonary fibroblasts, fluorofenidone decreased the up-regulation of TGF-ß1-induced eIF3a by inhibiting the proliferation of cells and reducing the expression of α-SMA, collagen I, and collagen III. These findings suggest that eIF3a is a new and special target of fluorofenidone, which could be potentially used in the development of a drug that treats PF.


Assuntos
Bleomicina/efeitos adversos , Fator de Iniciação 3 em Eucariotos/antagonistas & inibidores , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/metabolismo , Piridonas/farmacologia , Animais , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colágeno/metabolismo , Fator de Iniciação 3 em Eucariotos/genética , Fator de Iniciação 3 em Eucariotos/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/patologia , Regulação da Expressão Gênica/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Masculino , Fibrose Pulmonar/induzido quimicamente , Piridonas/uso terapêutico , Ratos , Ratos Sprague-Dawley , Fator de Crescimento Transformador beta1/sangue , Fator de Crescimento Transformador beta1/metabolismo
10.
Life Sci ; 144: 61-8, 2016 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-26612348

RESUMO

AIM: Eukaryotic translation initiation factors 3a (eIF3a) is involved in regulating cell cycle, cell division, growth and differentiation. Previous studies suggest a role of eIF3a on fibrosis disease and cellular proliferation and differentiation of fibroblasts. The present study aims to investigate the role of eIF3a on hypoxia-induced right ventricular (RV) remodeling and underlying mechanism. MAIN METHODS: RV remodeling was induced by hypoxia (10% O2, 3 weeks) in rats. Primary cardiac fibroblasts were cultured in vitro and their proliferation was investigated by MTS and EdU incorporation method. eIF3a knockdown was conducted by eIF3a siRNA. The expression/level of TGF-ß1, eIF3a, p27 and α-SMA, collagen-I, collagen-III, ANP and BNP were analyzed by ELISA, real-time PCR or Western blot. KEY FINDINGS: The expression of eIF3a was obviously increased in right ventricle of RV remodeling rats accompanied by up-regulation of α-SMA and collagens. In cultured cardiac fibroblasts, application of exogenous TGF-ß1-induced cellular proliferation and differentiation concomitantly with up-regulation of eIF3a expression and down-regulation of p27 expression. The effects of TGF-ß1-induced proliferation and up-regulation of α-SMA and collagen in cardiac fibroblasts were abolished by eIF3a siRNA. eIF3a siRNA reversed TGF-ß1 induced down-regulation of p27 expression. SIGNIFICANCE: The eIF3a plays a crucial role in hypoxia-induced RV remodeling by regulating TGF-ß1-induced proliferation and differentiation of cardiac fibroblasts, which is mediated via eIF3a/p27 pathway.


Assuntos
Fatores de Iniciação em Eucariotos/genética , Hipertrofia Ventricular Direita/genética , Hipertrofia Ventricular Direita/patologia , Hipóxia/patologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Remodelação Ventricular/genética , Actinas/biossíntese , Actinas/genética , Animais , Diferenciação Celular/genética , Proliferação de Células , Células Cultivadas , Colágeno/biossíntese , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Fibroblastos , Técnicas de Silenciamento de Genes , Masculino , RNA Interferente Pequeno/farmacologia , Ratos , Ratos Sprague-Dawley , Fator de Crescimento Transformador beta1/farmacologia
11.
Eur J Pharmacol ; 765: 565-73, 2015 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-26409044

RESUMO

Mammalian Target of Rapamycin (mTOR) is involved in the proliferation and survival of pulmonary artery smooth muscle cells (PASMCs) in human pulmonary hypertension (PH) and animal PH models, and miRNAs are reported to play a key role in modulation of the proliferation of PASMCs. The purposes of this study are to determine the functions of miR-100 and mTOR in cardiovascular remodeling of the hypoxic PH rats and to clarify the correlation between them. We established a rat model of hypoxic PH, which showed an increase in right ventricle systolic pressure, right ventricular and pulmonary vascular remodeling, accompanied by an up-regulation of mTOR and a down-regulation of miR-100. Next, we established an in vitro model of hypoxia-induced proliferation of PASMCs. Consistent with the in vivo findings, hypoxia induced proliferation of PASMCs, accompanied by a down-regulation of miR-100 and an up-regulation of mTOR; these phenomena were reversed by miR-100 mimics or the antagonist of mTOR. Finally, the dual-luciferase reporter gene assay was utilized to reveal the direct interaction between miR-100 and the 3 '-UTR region of mTOR gene. Based on these observations, we conclude that miR-100 can modulate the proliferation of PASMCs in hypoxic PH rats through suppressing the mTOR expression.


Assuntos
Hipertensão Pulmonar/metabolismo , Hipóxia/metabolismo , MicroRNAs/metabolismo , Transdução de Sinais/fisiologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/biossíntese , Animais , Proliferação de Células/fisiologia , Hipertensão Pulmonar/patologia , Hipóxia/patologia , Masculino , Ratos , Ratos Sprague-Dawley
12.
Int J Cardiol ; 197: 182-91, 2015 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-26142204

RESUMO

BACKGROUND: Reactive oxygen species (ROS) is thought as a major reason of vascular injury in diabetes. Vascular peroxidase 1 (VPO1) is a newly found peroxidase playing an important role in inducing oxidative stress. In the present experiment, we tested the role of VPO1 in senescence of endothelial cells in streptozotocin (STZ)-induced diabetic rats and cultured endothelial cells. METHODS: Blood samples were collected from carotid arteries. Vasodilator responses to acetylcholine (Ach) in the isolated aortic rings were measured, serum concentration of glucose, tumor necrosis factor-α (TNF-α) and monocyte chemoattractant protein-1 (MCP-1) and the expression of VPO1 in the aorta were determined. Endothelial cells were treated with high glucose or H2O2, the concentrations of MCP-1, TNF-α and hypochlorous acid (HOCl) and the expression of VPO1 were determined. shRNA of VPO1 was used for mechanism research in cultured cells. RESULTS: Vasodilator responses to Ach were impaired markedly and the serum concentrations of glucose, TNF-α and MCP-1 were significantly increased in diabetic rats. The expression of VPO1 in the aorta was upregulated in diabetic rats. High glucose treatment significantly decreased cell viability and elevated the levels of MCP-1, TNF-α and HOCl and upregulated the expression of VPO1. H2O2 treatment significantly induced cellular senescence, inhibited eNOS expression and NO production. The effects of high glucose and H2O2 were attenuated by shRNA interference of VPO1. CONCLUSIONS: VPO1 plays an important role in senescence of endothelial cells and endothelial dysfunction by induction of oxidative stress and inflammatory reaction in type 2 diabetic rats.


Assuntos
Diabetes Mellitus Experimental/genética , Endotélio Vascular/enzimologia , Regulação da Expressão Gênica , Hemeproteínas/genética , Estresse Oxidativo , Peroxidases/genética , RNA Mensageiro/genética , Vasodilatação , Animais , Western Blotting , Diabetes Mellitus Experimental/enzimologia , Diabetes Mellitus Experimental/fisiopatologia , Diabetes Mellitus Tipo 2/enzimologia , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/fisiopatologia , Endotélio Vascular/fisiopatologia , Hemeproteínas/biossíntese , Humanos , Masculino , Peroxidases/biossíntese , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real
13.
PLoS One ; 10(6): e0130806, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26120832

RESUMO

Enhanced proliferation of pulmonary arterial vascular smooth muscle cells (PASMCs) is a key pathological component of vascular remodeling in hypoxia-induced pulmonary hypertension (HPH). Mammalian targeting of rapamycin (mTOR) signaling has been shown to play a role in protein translation and participate in the progression of pulmonary hypertension. Eukaryotic translation initiation factor-2α (eIF2α) is a key factor in regulation of cell growth and cell cycle, but its role in mTOR signaling and PASMCs proliferation remains unknown. Pulmonary hypertension (PH) rat model was established by hypoxia. Rapamycin was used to treat rats as an mTOR inhibitor. Proliferation of primarily cultured rat PASMCs was induced by hypoxia, rapamycin and siRNA of mTOR and eIF2α were used in loss-of-function studies. The expression and activation of eIF2α, mTOR and c-myc were analyzed. Results showed that mTOR/eIF2α signaling was significantly activated in pulmonary arteries from hypoxia exposed rats and PASMCs cultured under hypoxia condition. Treatment with mTOR inhibitor for 21 days attenuated vascular remodeling, suppressed mTOR and eIF2α activation, inhibited c-myc expression in HPH rats. In hypoxia-induced PASMCs, rapamycin and knockdown of mTOR and eIF2α by siRNA significantly abolished proliferation and increased c-myc expression. These results suggest a critical role of the mTOR/eIF2αpathway in hypoxic vascular remodeling and PASMCs proliferation of HPH.


Assuntos
Fator de Iniciação 2 em Eucariotos/metabolismo , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/metabolismo , Hipóxia/complicações , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Hemodinâmica/efeitos dos fármacos , Hipertensão Pulmonar/patologia , Hipertensão Pulmonar/fisiopatologia , Hipóxia/patologia , Hipóxia/fisiopatologia , Masculino , Miócitos de Músculo Liso/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Artéria Pulmonar/patologia , RNA Interferente Pequeno/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Regulação para Cima/efeitos dos fármacos , Remodelação Vascular/efeitos dos fármacos
14.
Basic Res Cardiol ; 110(4): 38, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25982880

RESUMO

Recent studies demonstrated that NADPH oxidase 2 (NOX2) expression in myocardium after ischemia-reperfusion (IR) is significantly upregulated. However, the underlying mechanisms remain unknown. This study aims to determine if nuclear cardiac myosin light chain 2 (MYL2), a well-known regulatory subunit of myosin, functions as a transcription factor to promote NOX2 expression following myocardial IR in a phosphorylation-dependent manner. We examined the phosphorylation status of nuclear MYL2 (p-MYL2) in a rat model of myocardial IR (left main coronary artery subjected to 1 h ligation and 3 h reperfusion) injury, which showed IR injury and upregulated NOX2 expression as expected, accompanied by elevated H2O2 and nuclear p-MYL2 levels; these effects were attenuated by inhibition of myosin light chain kinase (MLCK). Next, we explored the functional relationship of nuclear p-MYL2 with NOX2 expression in H9c2 cell model of hypoxia-reoxygenation (HR) injury. In agreement with our in vivo findings, HR treatment increased apoptosis, NOX2 expression, nuclear p-MYL2 and H2O2 levels, and the increases were ameliorated by inhibition of MLCK or knockdown of MYL2. Finally, molecular biology techniques including co-immunoprecipitation (Co-IP), chromatin immunoprecipitation (ChIP), DNA pull-down and luciferase reporter gene assay were utilized to decipher the molecular mechanisms. We found that nuclear p-MYL2 binds to the consensus sequence AGCTCC in NOX2 gene promoter, interacts with RNA polymerase II and transcription factor IIB to form a transcription preinitiation complex, and thus activates NOX2 gene transcription. Our results demonstrate that nuclear MYL2 plays an important role in IR injury by transcriptionally upregulating NOX2 expression to enhance oxidative stress in a phosphorylation-dependent manner.


Assuntos
Miosinas Cardíacas/fisiologia , Glicoproteínas de Membrana/genética , Miocárdio/metabolismo , Cadeias Leves de Miosina/fisiologia , NADPH Oxidases/genética , Animais , Miosinas Cardíacas/análise , Núcleo Celular/química , Células Cultivadas , Masculino , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Cadeias Leves de Miosina/análise , Quinase de Cadeia Leve de Miosina/antagonistas & inibidores , NADPH Oxidase 2 , Estresse Oxidativo , Fosforilação , Ratos , Ratos Sprague-Dawley
15.
Int Immunopharmacol ; 27(1): 53-64, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25957199

RESUMO

It has also been shown that the decreased expression of eukaryotic translation initiation factor 3a (eIF3a) by L-mimosine caused cell cycle arrest. Our previous study has found that eIF3a is involved in bleomycin-induced pulmonary fibrosis. Whether the eIF3a/p27 signal pathway is involved in the inhibitory effect of L-mimosine on bleomycin-induced pulmonary fibrosis remains unknown. Pulmonary fibrosis was induced by intratracheal instillation of bleomycin (5 mg/kg) in rats. Primary pulmonary fibroblasts were cultured to investigate the proliferation by BrdU incorporation method and flow cytometry. The expression of eIF3a, p27, α-SMA, collagen I and collagen III was analyzed by qPCR and Western blot. In vivo, L-mimosine treatment significantly ameliorated the bleomycin-mediated histological fibrosis alterations and blocked collagen deposition concomitantly with reversing bleomycin-induced expression up-regulation of eIF3a, α-SMA, collagen I and collagen III (both mRNA and protein) and expression down- regulation of p27. In vitro, L-mimosine remarkably attenuated proliferation of pulmonary fibroblasts and expression of α-SMA, collagen I and collagen III induced by TGF-ß1, and this inhibitory effect of L-mimosine was accompanied by inhibiting eIF3a expression and increasing p27 expression. Knockdown of eIF3a gene expression reversed TGF-ß1-induced proliferation of fibroblasts, down-regulation of p27 expression and up-regulation of α-SMA, collagen I, and collagen III expression. These results suggest that L-mimosine inhibited the progression of bleomycin-induced pulmonary fibrosis in rats via the eIF3a/p27 pathway.


Assuntos
Fator de Iniciação 3 em Eucariotos/metabolismo , Fibroblastos/efeitos dos fármacos , Pulmão/patologia , Mimosina/administração & dosagem , Fibrose Pulmonar/tratamento farmacológico , Actinas/genética , Actinas/metabolismo , Animais , Bleomicina/administração & dosagem , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Colágeno/metabolismo , Inibidor de Quinase Dependente de Ciclina p27/genética , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Fator de Iniciação 3 em Eucariotos/genética , Fibroblastos/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Mimosina/efeitos adversos , Fibrose Pulmonar/induzido quimicamente , RNA Interferente Pequeno/genética , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta/metabolismo
16.
Biochem Biophys Res Commun ; 458(4): 869-76, 2015 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-25701782

RESUMO

The risk of cardiovascular complications in diabetic patients is mainly associated with endothelial dysfunction. Reduced number of EPCs and impaired function of EPCs in diabetes result in imbalance of endothelial homeostasis and dysfunction of vessels. In patients with diabetes mellitus, plasma levels of asymmetric dimethylarginine (ADMA) were elevated, while the expression and activity of dimethylarginine dimethylaminohydrolase (DDAH) were reduced. In the present study, we investigated the role of the DDAH2/ADMA pathway in the senescence of EPCs in type 2 diabetic patients and cultured EPCs treated with high glucose. The results showed that the percentage of senescent EPCs increased while the expression of DDAH2 decreased concomitantly with an increase in the plasma levels of ADMA in patients with type 2 diabetes mellitus (T2DM). Similar results were seen in cultured EPCs treated with high glucose. Exogenous application of ADMA accelerated the senescence of EPCs in a dose-dependent manner, and overexpression of DDAH2 inhibited high glucose-induced EPCs senescence. In addition, it has also been reported that DDAH/ADMA pathway is regulated by silent information regulator 1 (SIRT1) in endothelial cell. In the present study, we found decreased expression of SIRT1 both in T2DM patients and EPCs pretreated with high glucose. And resveratrol (activating SIRT1) inhibited high glucose-induced EPCs senescence by upregulating the expression of DDAH2 and decreasing the levels of ADMA. Taken together, we concluded that DDAH2/ADMA is involved in the accelerated senescence of EPCs in diabetes, which is associated with the activation of SIRT1.


Assuntos
Amidoidrolases/metabolismo , Arginina/análogos & derivados , Diabetes Mellitus Tipo 2/enzimologia , Diabetes Mellitus Tipo 2/patologia , Células Progenitoras Endoteliais/patologia , Arginina/sangue , Arginina/metabolismo , Senescência Celular , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/metabolismo , Células Progenitoras Endoteliais/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Sirtuína 1/metabolismo
17.
Free Radic Biol Med ; 83: 115-28, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25701432

RESUMO

Nuclear myosin regulates gene transcription and this novel function might be modulated through phosphorylation of the myosin regulatory light chain (p-MLC20). Nonmuscle MLC20 (nmMLC20) is also present in the nuclei of cardiomyocytes and a potential nmMLC20 binding sequence has been identified in the promoter of the xanthine oxidase (XO) gene. Thus, we investigated its function in the regulation of XO transcription after myocardial ischemia/reperfusion (IR). In a rat model of myocardial IR and a cardiomyocyte model of hypoxia/reoxygenation (HR) injury, the cardiac or cell injury, myosin light chain kinase (MLCK) content, XO expression and activity, XO-derived products, and level of nuclear p-nmMLC20 were detected. Coimmunoprecipitation (co-IP), chromatin immunoprecipitation, DNA pull-down, and luciferase reporter gene assays were used to decipher the molecular mechanisms through which nmMLC20 promotes XO expression. IR or HR treatment dramatically elevated nuclear p-nmMLC20 level, accompanied by increased XO expression, activity, and products (H2O2 and uric acid), as well as the IR or HR injury; these effects were ameliorated by inhibition of MLCK or knockdown of nmMLC20. Our findings from these experiments demonstrated that nuclear p-nmMLC20 binds to the consensus sequence GTCGCC in the XO gene promoter, interacts with RNA polymerase II and transcription factor IIB to form a transcription preinitiation complex, and hence activates XO gene transcription. These results suggest that nuclear p-nmMLC20 plays an important role in IR/HR injury by transcriptionally upregulating XO gene expression to increase oxidative stress in myocardium. Our findings demonstrate nuclear nmMLC20 as a potential new therapeutic target to combat cardiac IR injury.


Assuntos
Núcleo Celular/metabolismo , Regulação Enzimológica da Expressão Gênica , Isquemia Miocárdica/patologia , Traumatismo por Reperfusão Miocárdica/patologia , Cadeias Leves de Miosina/metabolismo , Xantina Oxidase/genética , Animais , Apoptose , Western Blotting , Núcleo Celular/genética , Proliferação de Células , Células Cultivadas , Imunoprecipitação da Cromatina , Peróxido de Hidrogênio/metabolismo , Imunoprecipitação , Masculino , Isquemia Miocárdica/genética , Isquemia Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/metabolismo , Cadeias Leves de Miosina/genética , Estresse Oxidativo , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Xantina Oxidase/metabolismo
18.
Eur J Pharmacol ; 749: 89-97, 2015 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-25592322

RESUMO

Eukaryotic translation initiation factor 3a (eIF3a) is a multifunctional protein and plays an important role in regulation of cellular function including proliferation and differentiation. In the present study, we tested the function of eIF3a in pulmonary fibrosis. Pulmonary fibrosis was induced by intratracheal instillation of bleomycin (5mg/kg) in rats. Primary pulmonary fibroblasts were cultured for proliferation investigation by BrdU incorporation method and flow cytometry. The expression/level of eIF3a, TGF-ß1, ERK1/2 and α-SMA were analyzed by ELISA, real-time PCR or western blot. Results showed that the expression of eIF3a was obviously increased in lungs of pulmonary fibrosis rats accompanied by up-regulation of α-SMA and collagens. In cultured pulmonary fibroblasts, application of exogenous TGF-ß1 induced cell proliferation and differentiation concomitantly with up-regulation of eIF3a expression and ERK1/2 phosphorylation. The effects of TGF-ß1-induced proliferation of fibroblasts and up-regulation of α-SMA were abolished by eIF3a siRNA. TGF-ß1-induced eIF3a expression was reversed in the presence of PD98059, an inhibitor of ERK1/2. These findings suggest that eIF3a plays an important role in bleomycin-induced pulmonary fibrosis by regulating pulmonary fibroblasts׳ function, and up-regulation of eIF3a induced by TGF-ß1 is mediated via the ERK1/2 pathway.


Assuntos
Antibióticos Antineoplásicos/efeitos adversos , Bleomicina/efeitos adversos , Fator de Iniciação 3 em Eucariotos/metabolismo , Fibrose Pulmonar/metabolismo , Animais , Colágeno/metabolismo , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/patologia , Ratos Sprague-Dawley , Fator de Crescimento Transformador beta1/farmacologia
19.
J Cardiovasc Pharmacol ; 64(5): 460-4, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25098345

RESUMO

The aim of this study was to investigate whether N-arachidonic acid ethanolamine (anandamide, AEA) transporter contributed to calcitonin gene-related peptide (CGRP) expression mediated by nitroglycerin (GTN) in peripheral blood mononuclear cells (PBMCs) of healthy volunteers and its association with the mitochondrial aldehyde dehydrogenase-2 (ALDH2) Glu504Lys (ALDH2*2) polymorphism. In 10 ALDH2*2-genotyped Chinese volunteers, we assessed the activity of AEA transporter and expression of CGRP messenger ribonucleic acid (mRNA) in cultured PBMCs treated with different concentration of GTN with or without pretreatment with AM404 (the AEA transporter blocker). In this study, the activity of AEA transporter and expression of CGRP mRNA elevated with the increase in the concentration of GTN. Pretreatment of the cells with AM404 (1 µM) 2 hours before GTN reduced the GTN-induced increase in both AEA transporter activity and CGRP mRNA expression significantly, and cells with the ALDH2*1/*1 homozygote genotype showed significantly higher activity of AEA transporter and CGRP mRNA expression than carriers of the ALDH2*2 allele. Therefore, we strongly suggested that GTN can stimulate CGRP expression by elevating the AEA transporter activity, which is affected by ALDH2 Glu504Lys polymorphism.


Assuntos
Aldeído Desidrogenase/genética , Ácidos Araquidônicos/metabolismo , Peptídeo Relacionado com Gene de Calcitonina/genética , Endocanabinoides/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Alcamidas Poli-Insaturadas/metabolismo , Adulto , Aldeído-Desidrogenase Mitocondrial , Ácidos Araquidônicos/farmacologia , Grupo com Ancestrais do Continente Asiático/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Genótipo , Humanos , Leucócitos Mononucleares/metabolismo , Masculino , Nitroglicerina/farmacologia , Polimorfismo Genético , RNA Mensageiro/metabolismo
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