Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 19 de 19
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Mol Cell ; 76(4): 546-561.e8, 2019 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-31561952

RESUMO

Through transcriptional control of the evolutionarily conserved heat shock, or proteotoxic stress, response, heat shock factor 1 (HSF1) preserves proteomic stability. Here, we show that HSF1, a physiological substrate for AMP-activated protein kinase (AMPK), constitutively suppresses this central metabolic sensor. By physically evoking conformational switching of AMPK, HSF1 impairs AMP binding to the γ subunits and enhances the PP2A-mediated de-phosphorylation, but it impedes the LKB1-mediated phosphorylation of Thr172, and retards ATP binding to the catalytic α subunits. These immediate and manifold regulations empower HSF1 to both repress AMPK under basal conditions and restrain its activation by diverse stimuli, thereby promoting lipogenesis, cholesterol synthesis, and protein cholesteroylation. In vivo, HSF1 antagonizes AMPK to control body fat mass and drive the lipogenic phenotype and growth of melanomas independently of its intrinsic transcriptional action. Thus, the physical AMPK-HSF1 interaction epitomizes a reciprocal kinase-substrate regulation whereby lipid metabolism and proteomic stability intertwine.

2.
Bioessays ; 39(5)2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28295473

RESUMO

Beyond protein synthesis and autophagy, emerging evidence has implicated mTORC1 in regulating protein folding and proteasomal degradation as well, highlighting its prominent role in cellular proteome homeostasis or proteostasis. In addition to growth signals, mTORC1 senses and responds to a wide array of stresses, including energetic/metabolic stress, genotoxic stress, oxidative stress, osmotic stress, ER stress, proteotoxic stress, and psychological stress. Whereas growth signals unanimously stimulate mTORC1, stresses exert complex impacts on mTORC1, most of which are repressive. mTORC1 suppression, as a generic adaptive strategy, empowers cell survival under various stressful conditions. In this essay, we provide an overview of the emerging role of mTORC1 in proteostasis, the distinct molecular mechanisms through which mTORC1 reacts to diverse stresses, and the schemes exploited by cancer cells to circumvent stress-induced mTORC1 suppression. Hence, acting as a stress sensor, mTORC1 intimately couples stresses to cellular proteostasis.


Assuntos
Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Proteostase , Estresse Fisiológico , Animais , Carcinogênese , Estresse do Retículo Endoplasmático , Humanos , Modelos Biológicos , Neoplasias/metabolismo , Pressão Osmótica , Estresse Oxidativo , Estresse Psicológico
3.
Cell Cycle ; 15(23): 3155-3156, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27589382
4.
Cell Mol Life Sci ; 73(22): 4231-4248, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27289378

RESUMO

Proteome homeostasis, or proteostasis, is essential to maintain cellular fitness and its disturbance is associated with a broad range of human health conditions and diseases. Cells are constantly challenged by various extrinsic and intrinsic insults, which perturb cellular proteostasis and provoke proteotoxic stress. To counter proteomic perturbations and preserve proteostasis, cells mobilize the proteotoxic stress response (PSR), an evolutionarily conserved transcriptional program mediated by heat shock factor 1 (HSF1). The HSF1-mediated PSR guards the proteome against misfolding and aggregation. In addition to proteotoxic stress, emerging studies reveal that this proteostatic mechanism also responds to cellular energy state. This regulation is mediated by the key cellular metabolic sensor AMP-activated protein kinase (AMPK). In this review, we present an overview of the maintenance of proteostasis by HSF1, the metabolic regulation of the PSR, particularly focusing on AMPK, and their implications in the two major age-related diseases-diabetes mellitus and neurodegenerative disorders.


Assuntos
Diabetes Mellitus/metabolismo , Diabetes Mellitus/patologia , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Proteínas/toxicidade , Estresse Fisiológico/efeitos dos fármacos , Animais , Proteínas de Choque Térmico/metabolismo , Humanos , Modelos Biológicos
5.
Nat Cell Biol ; 18(5): 527-39, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-27043084

RESUMO

To cope with proteotoxic stress, cells attenuate protein synthesis. However, the precise mechanisms underlying this fundamental adaptation remain poorly defined. Here we report that mTORC1 acts as an immediate cellular sensor of proteotoxic stress. Surprisingly, the multifaceted stress-responsive kinase JNK constitutively associates with mTORC1 under normal growth conditions. On activation by proteotoxic stress, JNK phosphorylates both RAPTOR at S863 and mTOR at S567, causing partial disintegration of mTORC1 and subsequent translation inhibition. Importantly, HSF1, the central player in the proteotoxic stress response (PSR), preserves mTORC1 integrity and function by inactivating JNK, independently of its canonical transcriptional action. Thereby, HSF1 translationally augments the PSR. Beyond promoting stress resistance, this intricate HSF1-JNK-mTORC1 interplay, strikingly, regulates cell, organ and body sizes. Thus, these results illuminate a unifying mechanism that controls stress adaptation and growth.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Resposta ao Choque Térmico , Complexos Multiproteicos/metabolismo , Proteínas/toxicidade , Estresse Fisiológico/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Fatores de Transcrição/metabolismo , Animais , Tamanho Corporal/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Tamanho Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Células HEK293 , Células HeLa , Fatores de Transcrição de Choque Térmico , Resposta ao Choque Térmico/efeitos dos fármacos , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fígado/citologia , Fígado/efeitos dos fármacos , Fígado/crescimento & desenvolvimento , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Tamanho do Órgão/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Biossíntese de Proteínas/efeitos dos fármacos , Transcrição Genética/efeitos dos fármacos
6.
Mol Nutr Food Res ; 59(4): 646-57, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25581901

RESUMO

SCOPE: Epigallocatechin-3-gallate (EGCG), the most abundant catechin of green tea, has beneficial effects on physiological functions of endothelial cells (ECs), yet the detailed mechanisms are not fully understood. In this study, we investigated the role of transient receptor potential vanilloid type 1 (TRPV1), a ligand-gated nonselective calcium channel, in EGCG-mediated endothelial nitric oxide (NO) synthase (eNOS) activation and angiogenesis. METHODS AND RESULTS: In ECs, treatment with EGCG time-dependently increased the intracellular level of Ca(2+) . Removal of extracellular calcium (Ca(2+) ) by EGTA or EDTA or inhibition of TRPV1 by capsazepine or SB366791 abrogated EGCG-increased intracellular Ca(2+) level in ECs or TRPV1-transfected HEK293 cells. Additionally, EGCG increased the phsophorylation of eNOS at Ser635 and Ser1179, Akt at Ser473, calmodulin-dependent protein kinase II (CaMKII) at Thr286 and AMP-activated protein kinase (AMPK) at Thr172, all abolished by the TRPV1 antagonist capsazepine. EGCG-induced NO production was diminished by pretreatment with LY294002 (an Akt inhibitor), KN62 (a CaMKII inhibitor), and compound C (an AMPK inhibitor). Moreover, blocking TRPV1 activation prevented EGCG-induced EC proliferation, migration, and tube formation, as well as angiogenesis in Matrigel plugs in mice. CONCLUSION: EGCG may trigger activation of TRPV1-Ca(2+) signaling, which leads to phosphorylation of Akt, AMPK, and CaMKII; eNOS activation; NO production; and, ultimately, angiogenesis in ECs.


Assuntos
Catequina/análogos & derivados , Óxido Nítrico Sintase Tipo III/metabolismo , Canais de Cátion TRPV/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Proteínas Quinases Ativadas por AMP/metabolismo , Anilidas/farmacologia , Animais , Cálcio/metabolismo , Capsaicina/análogos & derivados , Capsaicina/farmacologia , Catequina/farmacologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cinamatos/farmacologia , Células Endoteliais/efeitos dos fármacos , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neovascularização Patológica/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Fosforilação , Transdução de Sinais , Canais de Cátion TRPV/antagonistas & inibidores , Canais de Cátion TRPV/genética , Chá/química
7.
Int J Biol Sci ; 10(9): 990-6, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25210497

RESUMO

14,15-epoxyeicosatrienoic acid (14,15-EET) is implicated in regulating physiological functions of endothelial cells (ECs), yet the potential molecular mechanisms underlying the beneficial effects in ECs are not fully understood. In this study, we investigated whether transient receptor potential vanilloid receptor type 1 (TRPV1) is involved in 14,15-EET-mediated Ca(2+) influx, nitric oxide (NO) production and angiogenesis. In human microvascular endothelial cells (HMECs), 14,15-EET time-dependently increased the intracellular level of Ca(2+). Removal of extracellular Ca(2+), pharmacological inhibition or genetic disruption of TRPV1 abrogated 14,15-EET-mediated increase of intracellular Ca(2+) level in HMECs or TRPV1-transfected HEK293 cells. Furthermore, removal of extracellular Ca(2+) or pharmacological inhibition of TRPV1 decreased 14,15-EET-induced NO production. 14,15-EET-mediated tube formation was abolished by TRPV1 pharmacological inhibition. In an animal experiment, 14,15-EET-induced angiogenesis was diminished by inhibition of TRPV1 and in TRPV1-deficient mice. TRPV1 may play a crucial role in 14,15-EET-induced Ca(2+) influx, NO production and angiogenesis.


Assuntos
Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Células Endoteliais/fisiologia , Neovascularização Fisiológica/efeitos dos fármacos , Canais de Cátion TRPV/metabolismo , Vasodilatadores/farmacologia , Ácido 8,11,14-Eicosatrienoico/farmacologia , Animais , Cálcio/metabolismo , Regulação da Expressão Gênica/fisiologia , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico , Transdução de Sinais , Canais de Cátion TRPV/genética
8.
J Gastroenterol Hepatol ; 29(3): 494-501, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24219143

RESUMO

BACKGROUND AND AIM: Inflammatory bowel diseases (IBDs) are chronic inflammatory disorders with unclear etiology and mechanism(s). Glycine N-methyltransferase (GNMT) plays a central role in inflammatory diseases such as hepatitis and atherosclerosis. However, little is known about the impact of GNMT and the involved mechanism in the pathogenesis of IBD. In the current study, we investigated the role of GNMT in the mouse model of dextran sulfate sodium (DSS)-induced colitis. METHODS: Protein expression was determined by Western blotting or immunohistochemistry. Histopathology was examined by hematoxylin and eosin staining. Levels of pro-inflammatory cytokines were evaluated by ELISA kits. RESULTS: GNMT was expressed in the epithelium of the colon under normal conditions, and with DSS treatment, its expression was predominant in infiltrated leukocytes of lesions. Mice with genetic deletion of GNMT (GNMT(-/-) ) showed increased susceptibility to DSS induction of colitis, as revealed by the progression of colitis. Additionally, severe colonic inflammation, including increased crypt loss, leukocyte infiltration, and hemorrhage, was greater with DSS treatment in GNMT(-/-) than wild-type mice. Furthermore, the expression of adhesion molecule and inflammatory mediators in the colon was significantly higher with DSS treatment in GNMT(-/-) than wild-type mice. Moreover, loss of GNMT decreased cell apoptosis in colitis lesions with DSS treatment. CONCLUSIONS: Collectively, our findings suggest that GNMT may be a crucial molecule in the pathogenesis of DSS-induced colitis. This finding may provide new information for a potential therapeutic target in treating IBD.


Assuntos
Colite Ulcerativa/genética , Glicina N-Metiltransferase/genética , Glicina N-Metiltransferase/fisiologia , Animais , Colite Ulcerativa/induzido quimicamente , Colite Ulcerativa/patologia , Sulfato de Dextrana , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Expressão Gênica , Glicina N-Metiltransferase/metabolismo , Mediadores da Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Terapia de Alvo Molecular
9.
Am J Chin Med ; 41(5): 1079-96, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24117070

RESUMO

Paeonol, a phenolic component purified from Paeonia suffruticosa (Cortex Moutan), is used in traditional Chinese medicine to treat inflammatory diseases. However, little is known about the effect of paeonol on cholesterol metabolism. We investigated the efficacy of paeonol on cholesterol metabolism and the underlying mechanism in macrophages and apolipoprotein E deficient (apoE(-/-)) mice. Treatment with paeonol markedly attenuated cholesterol accumulation induced by oxidized LDL in macrophages, which was due to increased cholesterol efflux. Additionally, paeonol enhanced the mRNA and protein expression of ATP-binding membrane cassette transport protein A1 (ABCA1) but did not alter the protein level of ABCG1 or other scavenger receptors. Inhibition of ABCA1 activity with a pharmacological inhibitor, neutralizing antibody or small interfering RNA (siRNA), negated the effects of paeonol on cholesterol efflux and cholesterol accumulation. Furthermore, paeonol induced the nuclear translocation of liver X receptor α (LXRα) by increasing its activity. siRNA knockdown of LXRα abolished the paeonol-induced upregulation of ABCA1, promotion of cholesterol efflux and suppression of cholesterol accumulation. Moreover, atherosclerotic lesions, hyperlipidemia and systemic inflammation were reduced and the protein expression of ABCA1 was increased in aortas of paeonol-treated apoE(-/-) mice. Paeonol may alleviate the formation of foam cells by enhancing LXRα-ABCA1-dependent cholesterol efflux.


Assuntos
Transportador 1 de Cassete de Ligação de ATP/metabolismo , Acetofenonas/farmacologia , Colesterol/metabolismo , Células Espumosas/metabolismo , Expressão Gênica/efeitos dos fármacos , Receptores Nucleares Órfãos/metabolismo , Regulação para Cima/efeitos dos fármacos , Transportador 1 de Cassete de Ligação de ATP/genética , Transportador 1 de Cassete de Ligação de ATP/fisiologia , Acetofenonas/uso terapêutico , Animais , Aterosclerose/tratamento farmacológico , Aterosclerose/etiologia , Técnicas de Cultura de Células , Relação Dose-Resposta a Droga , Mediadores da Inflamação/metabolismo , Lipoproteínas LDL/metabolismo , Receptores X do Fígado , Masculino , Camundongos , Terapia de Alvo Molecular , Receptores Nucleares Órfãos/genética , Receptores Nucleares Órfãos/fisiologia , Fitoterapia , RNA Interferente Pequeno
10.
Mol Med ; 18: 805-15, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22451268

RESUMO

We investigated whether AMP-activated protein kinase (AMPK), a multi-functional regulator of energy homeostasis, is involved in transient receptor potential vanilloid type 1 (TRPV1)-mediated activation of endothelial nitric oxide synthase (eNOS) in endothelial cells (ECs) and mice. In ECs, treatment with evodiamine, the activator of TRPV1, increased the phosphorylation of AMPK, acetyl-CoA carboxylase (ACC) and eNOS, as revealed by western blot analysis. Inhibition of AMPK activation by compound C or dominant-negative AMPK mutant abrogated the evodiamine-induced increase in phosphorylation of AMPK and eNOS and NO bioavailability, as well as tube formation in ECs. Immunoprecipitation and two-hybrid analysis demonstrated that AMPK mediated the evodiamine-induced increase in the formation of a TRPV1-eNOS complex. Additionally, TRPV1 activation by evodiamine increased the phosphorylation of AMPK and eNOS in aortas of wild-type mice but did not activate eNOS in aortas of TRPV1-deficient mice. In mice, inhibition of AMPK activation by compound C markedly decreased evodiamine-evoked angiogenesis in Matrigel plugs and in a hind-limb ischemia model. Moreover, evodiamine-induced phosphorylation of AMPK and eNOS in aortas of apolipoprotein E deficient (ApoE(-/-)) mice was abrogated in TRPV1-deficient ApoE(-/-) mice. In conclusion, TRPV1 activation may trigger AMPK-dependent signaling, which leads to enhanced activation of AMPK and eNOS and retarded development of atherosclerosis.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Aorta/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Canais de Cátion TRPV/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Animais , Aorta/citologia , Vasos Sanguíneos/efeitos dos fármacos , Western Blotting , Bovinos , Células Cultivadas , Colágeno , Combinação de Medicamentos , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Ativação Enzimática/efeitos dos fármacos , Membro Posterior/irrigação sanguínea , Membro Posterior/metabolismo , Injeções Intraperitoneais , Laminina , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Fosforilação/efeitos dos fármacos , Ligação Proteica , Proteoglicanas , Quinazolinas/administração & dosagem , Quinazolinas/farmacologia , Canais de Cátion TRPV/genética
11.
J Cell Physiol ; 227(8): 3053-62, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22021095

RESUMO

We investigated whether AMP-activated protein kinase (AMPK), a multi-functional regulator of energy homeostasis, participates in the regulation of erythropoietin (EPO)-mediated activation of endothelial nitric oxide synthase (eNOS) in endothelial cells (ECs) and mice. In ECs, treatment with EPO increased the phosphorylation of AMPK, acetyl-CoA carboxylase (ACC), and eNOS, as revealed by Western blot analysis. Inhibition of AMPK activation by compound C or dominant-negative AMPK mutant abrogated the EPO-induced increase in the phosphorylation of AMPK, ACC, and eNOS, as well as nitric oxide (NO) production. Additionally, suppression of AMPK activation abolished EPO-induced EC proliferation, migration and tube formation. Immunoprecipitation analysis demonstrated that AMPK mediated the EPO-induced increase in the phosphorylation of ß common receptor (ßCR) and the formation of a ßCR-AMPK-eNOS complex. In mice, inhibition of AMPK activation by compound C markedly decreased EPO-elicited angiogenesis in Matrigel plugs. Furthermore, the phosphorylation of AMPK and eNOS was significantly higher in aortas from EPO transgenic mice than wild-type mice. Moreover, treatment with EPO neutralizing antibody greatly reduced the exercise training-induced increase in phosphorylation of AMPK and eNOS in aortas of wild-type mice. Taken together, EPO may trigger AMPK-dependent signaling, which leads to enhanced phosphorylation of ßCR and eNOS, increased ßCR-AMPK-eNOS complex formation, NO production, and, ultimately, angiogenesis.


Assuntos
Proteínas Quinases Ativadas por AMP/antagonistas & inibidores , Proteínas Quinases Ativadas por AMP/metabolismo , Acetil-CoA Carboxilase/metabolismo , Eritropoetina/metabolismo , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Animais , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Endotélio/citologia , Endotélio/metabolismo , Eritropoetina/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neovascularização Fisiológica/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Pirazóis/farmacologia , Pirimidinas/farmacologia , Transdução de Sinais
12.
Cardiovasc Res ; 91(3): 492-501, 2011 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-21493704

RESUMO

AIMS: We investigated the molecular mechanism underlying the role of transient receptor potential vanilloid type 1 (TRPV1), a Ca(2+)-permeable non-selective cation channel, in the activation of endothelial nitric oxide (NO) synthase (eNOS) in endothelial cells (ECs) and mice. METHODS AND RESULTS: In ECs, TRPV1 ligands (evodiamine or capsaicin) promoted NO production, eNOS phosphorylation, and the formation of a TRPV1-eNOS complex, which were all abrogated by the TRPV1 antagonist capsazepine. TRPV1 ligands promoted the phosphorylation of Akt, calmodulin-dependent protein kinase II (CaMKII) and TRPV1, and increased the formation of a TRPV1-Akt-CaMKII complex. Removal of extracellular Ca(2+) abolished the ligand-induced increase in the phosphorylation of Akt and CaMKII, formation of a TRPV1-eNOS complex, and eNOS activation. Inhibition of PI3K and CaMKII suppressed the ligand-induced increase in TRPV1 phosphorylation, formation of a TRPV1-eNOS complex, and eNOS activation. TRPV1 activation increased the phosphorylation of Akt, CaMKII, and eNOS in the aortas of wild-type mice but failed to activate eNOS in TRPV1-deficient aortas. Additionally, TRPV1 ligand-induced angiogenesis was diminished in eNOS- or TRPV1-deficient mice. When compared with apolipoprotein E (ApoE)-deficient mice, ApoE/TRPV1-double-knockout mice displayed reduced phosphorylation of eNOS, Akt, and CaMKII in aortas but worsened atherosclerotic lesions. CONCLUSION: TRPV1 activation in ECs may trigger Ca(2+)-dependent PI3K/Akt/CaMKII signalling, which leads to enhanced phosphorylation of TRPV1, increased TRPV1-eNOS complex formation, eNOS activation and, ultimately, NO production.


Assuntos
Células Endoteliais/enzimologia , Óxido Nítrico Sintase Tipo III/metabolismo , Óxido Nítrico/metabolismo , Canais de Cátion TRPV/metabolismo , Animais , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo , Aterosclerose/enzimologia , Aterosclerose/genética , Aterosclerose/patologia , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Capsaicina/análogos & derivados , Capsaicina/farmacologia , Bovinos , Células Cultivadas , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Células Endoteliais/efeitos dos fármacos , Ativação Enzimática , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico Sintase Tipo III/deficiência , Óxido Nítrico Sintase Tipo III/genética , Fosfatidilinositol 3-Quinase/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Quinazolinas/farmacologia , Interferência de RNA , Transdução de Sinais , Canais de Cátion TRPV/deficiência , Canais de Cátion TRPV/efeitos dos fármacos , Canais de Cátion TRPV/genética , Fatores de Tempo , Transfecção
13.
J Cell Physiol ; 226(12): 3330-9, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21321940

RESUMO

Erythropoietin (EPO), the key hormone for erythropoiesis, also increases nitric oxide (NO) bioavailability in endothelial cells (ECs), yet the definitive mechanisms are not fully understood. Increasing evidence has demonstrated that ß common receptor (ßCR) plays a crucial role in EPO-mediated non-hematopoietic effects. We investigated the role of ßCR in EPO-induced endothelial NO synthase (eNOS) activation in bovine aortic ECs (BAECs) and the molecular mechanisms involved. Results of confocal microscopy and immunoprecipitation analyses revealed that ßCR was colocalized and interacted with EPO receptor (EPOR) in ECs. Inhibition of ßCR or EPOR by neutralizing antibodies or small interfering RNA abolished the EPO-induced NO production. Additionally, blockage of ßCR abrogated the EPO-induced increase in the phosphorylation of eNOS, Akt, Src, or Janus kinase 2 (JAK2). Immunoprecipitation analysis revealed that treatment with EPO increased the interaction between ßCR and eNOS, which was suppressed by inhibition of Src, JAK2, or Akt signaling with specific pharmacological inhibitors. Furthermore, EPO-induced EC proliferation, migration, and tube formation were blocked by pretreatment with ßCR antibody and Src, JAK2, or PI3K/Akt inhibitors. Moreover, in vivo experiments showed that EPO increased the level of phosphorylated eNOS, Src, JAK2, and Akt, as well as ßCR-eNOS association in aortas and promoted the angiogenesis in Matrigel plug, which was diminished by ßCR or EPOR neutralizing antibodies. Our findings suggest that ßCR may play an integrative role in the EPO signaling-mediated activation of eNOS in ECs.


Assuntos
Subunidade beta Comum dos Receptores de Citocinas/metabolismo , Células Endoteliais/enzimologia , Eritropoetina/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Receptores de Interleucina-3/metabolismo , Transdução de Sinais , Animais , Anticorpos Neutralizantes/farmacologia , Bovinos , Movimento Celular , Proliferação de Células , Células Cultivadas , Subunidade beta Comum dos Receptores de Citocinas/genética , Subunidade beta Comum dos Receptores de Citocinas/imunologia , Células Endoteliais/efeitos dos fármacos , Ativação Enzimática , Eritropoetina/genética , Humanos , Imunoprecipitação , Janus Quinase 2/antagonistas & inibidores , Janus Quinase 2/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Microscopia Confocal , Neovascularização Fisiológica , Óxido Nítrico/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Interferência de RNA , Receptores da Eritropoetina/genética , Receptores da Eritropoetina/imunologia , Receptores da Eritropoetina/metabolismo , Receptores de Interleucina-3/genética , Receptores de Interleucina-3/imunologia , Proteínas Recombinantes , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Transfecção , Quinases da Família src/antagonistas & inibidores , Quinases da Família src/metabolismo
14.
J Nutr Biochem ; 22(11): 1015-21, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21190831

RESUMO

Wogonin, one component in Scutellaria baicalensis Georgi extracts, has several beneficial properties for cancers and inflammatory diseases. However, the efficacy of wogonin in cholesterol metabolism of macrophages remains unknown. In macrophages, cholesterol uptake is controlled by scavenger receptors (SR-A and CD36) and cholesterol efflux by SR-BI, ATP-binding cassette transporter-A1 (ABCA1) and ABCG1. In the present study, we investigated the effect and underlying molecular mechanism of wogonin on the formation of macrophage foam cells by murine J774.A1 macrophages. Wogonin attenuated oxidized low-density lipoprotein (oxLDL)-induced cholesterol accumulation in macrophages. The binding of oxLDL to macrophages and protein expression of SR-A and CD36 were not affected by wogonin. Wogonin enhanced cholesterol efflux and increased the protein level of ABCA1 without affecting the protein expression of SR-BI or ABCG1. Inhibition of ABCA1 by pharmacological inhibitor 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid disodium salt or neutralizing antibody abolished this suppressive effect of wogonin on lipid accumulation. Moreover, the up-regulation of ABCA1 protein by wogonin resulted from a decrease in degradation rate of ABCA1 protein, with no effect on ABCA1 mRNA expression. This reduction in ABCA1 degradation was due to increased protein phosphatase 2B (PP2B)-mediated ABCA1 dephosphorylation, as evidenced by increased interaction between ABCA1 and PP2B; pharmacological inhibition of PP2B would prevent wogonin-induced ABCA1 protein expression, dephosphorylation and attenuation of lipid accumulation. Collectively, wogonin increases the protein stability of ABCA1 via PP2B-mediated dephosphorylation, thus leading to reduced cholesterol accumulation in macrophage foam cells.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Calcineurina/metabolismo , Colesterol/metabolismo , Flavanonas/farmacologia , Macrófagos/metabolismo , Ácido 4,4'-Di-Isotiocianoestilbeno-2,2'-Dissulfônico/farmacologia , Transportador 1 de Cassete de Ligação de ATP , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Animais , Células Cultivadas , Células Espumosas/efeitos dos fármacos , Lipoproteínas/metabolismo , Lipoproteínas LDL/metabolismo , Macrófagos/efeitos dos fármacos , Camundongos , Receptores Depuradores Classe A/metabolismo , Receptores Depuradores Classe B/metabolismo
15.
Free Radic Biol Med ; 50(1): 47-54, 2011 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-21034810

RESUMO

α-Lipoic acid (α-LA), a key cofactor in cellular energy metabolism, has protective activities in atherosclerosis, yet the detailed mechanisms are not fully understood. In this study, we examined whether α-LA affects foam cell formation and its underlying molecular mechanisms in murine macrophages. Treatment with α-LA markedly attenuated oxidized low-density lipoprotein (oxLDL)-mediated cholesterol accumulation in macrophages, which was due to increased cholesterol efflux. Additionally, α-LA treatment dose-dependently increased protein levels of ATP-binding cassette transporter A1 (ABCA1) and ABCG1 but had no effect on the protein expression of SR-A, CD36, or SR-BI involved in cholesterol homeostasis. Furthermore, α-LA increased the mRNA expression of ABCA1 and ABCG1. The upregulation of ABCA1 and ABCG1 by α-LA depended on liver X receptor α (LXRα), as evidenced by an increase in the nuclear levels of LXRα and LXRE-mediated luciferase activity and its prevention of the expression of ABCA1 and ABCG1 after inhibition of LXRα activity by the pharmacological inhibitor geranylgeranyl pyrophosphate (GGPP) or knockdown of LXRα expression with small interfering RNA (siRNA). Consistently, α-LA-mediated suppression of oxLDL-induced lipid accumulation was abolished by GGPP or LXRα siRNA treatment. In conclusion, LXRα-dependent upregulation of ABCA1 and ABCG1 may mediate the beneficial effect of α-LA on foam cell formation.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Células Espumosas/efeitos dos fármacos , Lipoproteínas/genética , Receptores Nucleares Órfãos/fisiologia , Ácido Tióctico/farmacologia , Transportador 1 de Cassete de Ligação de ATP , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Células Cultivadas , Células Espumosas/metabolismo , Células Espumosas/patologia , Humanos , Lipoproteínas/metabolismo , Lipoproteínas LDL/metabolismo , Receptores X do Fígado , Camundongos , Receptores Nucleares Órfãos/antagonistas & inibidores , Receptores Nucleares Órfãos/metabolismo , Fosfatos de Poli-Isoprenil/farmacologia , RNA Interferente Pequeno/farmacologia , Elementos de Resposta/efeitos dos fármacos , Ativação Transcricional/efeitos dos fármacos , Ativação Transcricional/fisiologia , Transfecção , Regulação para Cima/efeitos dos fármacos
16.
Cardiovasc Res ; 88(3): 415-23, 2010 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-20615914

RESUMO

AIMS: Accumulation of foam cells in the intima is a hallmark of early-stage atherosclerotic lesions. Ginkgo biloba extract (EGb761) has been reported to exert anti-oxidative and anti-inflammatory properties in atherosclerosis, yet the significance and the molecular mechanisms of action of EGb761 in the formation of macrophage foam cells are not fully understood. METHODS AND RESULTS: Treatment with EGb761 resulted in a dose-dependent decrease in oxidized low-density lipoprotein (oxLDL)-mediated cholesterol accumulation in macrophages, a consequence that was due to a decrease in cholesterol uptake and an increase in cholesterol efflux. Additionally, EGb761 significantly down-regulated the mRNA and protein expression of class A scavenger receptor (SR-A) by decreasing expression of activator protein 1 (AP-1); however, EGb761 increased the protein stability of ATP-binding cassette transporter A1 (ABCA1) by reducing calpain activity without affecting ABCA1 mRNA expression. Small interfering RNA (siRNA) targeting haem oxygenase-1 (HO-1) abolished the EGb761-induced protective effects on the expression of AP-1, SR-A, ABCA1, and calpain activity. Accordingly, EGb761-mediated suppression of lipid accumulation in foam cells was also abrogated by HO-1 siRNA. Moreover, the lesion size of atherosclerosis was smaller in EGb761-treated, apolipoprotein E-deficient mice compared with the vehicle-treated mice, and the expression of HO-1, SR-A, and ABCA1 in aortas was modulated similar to that observed in macrophages. CONCLUSION: These findings suggest that EGb761 confers a protection from the formation of foam cells by a novel HO-1-dependent regulation of cholesterol homeostasis in macrophages.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Células Espumosas/efeitos dos fármacos , Células Espumosas/metabolismo , Heme Oxigenase-1/metabolismo , Extratos Vegetais/farmacologia , Receptores Depuradores Classe A/metabolismo , Transportador 1 de Cassete de Ligação de ATP , Animais , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo , Aterosclerose/metabolismo , Aterosclerose/patologia , Aterosclerose/prevenção & controle , Calpaína/metabolismo , Colesterol/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Células Espumosas/patologia , Homeostase/efeitos dos fármacos , Metabolismo dos Lipídeos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Knockout
17.
Circulation ; 121(16): 1828-37, 2010 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-20385932

RESUMO

BACKGROUND: In addition to the hematopoietic effect of erythropoietin, increasing evidence suggests that erythropoietin also exerts protective effects for cardiovascular diseases. However, the role of erythropoietin and its underlying mechanism in macrophage foam cell formation are poorly understood. METHODS AND RESULTS: Compared with wild-type specimens, erythropoietin was increased in atherosclerotic aortas of apolipoprotein E-deficient (apoE(-/-)) mice, mainly in the macrophage foam cells of the lesions. Erythropoietin levels in culture medium and macrophages were significantly elevated in response to oxidized low-density lipoprotein in a dose-dependent manner. Furthermore, erythropoietin markedly attenuated lipid accumulation in oxidized low-density lipoprotein-treated macrophages, a result that was due to an increase in cholesterol efflux. Erythropoietin treatment significantly increased ATP-binding cassette transporters (ABC) A1 and ABCG1 mRNA and protein levels without affecting protein expression of scavenger receptors, including scavenger receptor-A, CD36, and scavenger receptor-BI. The upregulation of ABCA1 and ABCG1 by erythropoietin resulted from liver X receptor alpha activation, which was confirmed by its prevention on expression of ABCA1 and ABCG1 after pharmacological or small interfering RNA inhibition of liver X receptor alpha. Moreover, the erythropoietin-mediated attenuation on lipid accumulation was abolished by such inhibition. Finally, reduced lipid accumulation and marked increase in ABCA1 and ABCG1 were demonstrated in erythropoietin-overexpressed macrophages. CONCLUSIONS: Our data suggest that erythropoietin suppresses foam cell formation via the liver X receptor alpha-dependent upregulation of ABCA1 and ABCG1.


Assuntos
Aterosclerose/tratamento farmacológico , Cardiotônicos/farmacologia , Eritropoetina/farmacologia , Células Espumosas/efeitos dos fármacos , Receptores Nucleares Órfãos/metabolismo , Transportador 1 de Cassete de Ligação de ATP , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Doenças da Aorta/tratamento farmacológico , Doenças da Aorta/imunologia , Doenças da Aorta/patologia , Apolipoproteínas E/genética , Aterosclerose/imunologia , Aterosclerose/patologia , Antígenos CD36/genética , Células Cultivadas , Células Espumosas/metabolismo , Células Espumosas/patologia , Lipídeos/biossíntese , Lipoproteínas/genética , Lipoproteínas/metabolismo , Lipoproteínas LDL/farmacologia , Receptores X do Fígado , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , RNA Mensageiro/metabolismo , Receptores Depuradores Classe A/genética , Receptores Depuradores Classe B/genética
18.
Cardiovasc Res ; 82(3): 468-75, 2009 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-19307231

RESUMO

AIMS: Valsartan, a selective angiotensin II type 1 receptor (AT1R) blocker, has beneficial effects in the cardiovascular system in part by its increase of nitric oxide (NO) bioavailability, yet the mechanisms are unclear. We investigated the molecular mechanisms underlying this effect in endothelial cells (ECs). METHODS AND RESULTS: NO production was examined by Griess reagent assay, DAF-2 DA fluorescence staining and cGMP ELISA kits. Protein interaction was determined by western blotting and immunoprecipitation. Treating bovine or human aortic ECs with valsartan increased NO production, as evidenced by elevated level of stable NO metabolites and intracellular cGMP. Valsartan increased the phosphorylation but not the protein level of endothelial NO synthase (eNOS). Inhibition of phosphoinositide-3 kinase (PI3K)/Akt and Src pathways by specific inhibitors suppressed valsartan-induced NO release. In addition, valsartan increased the tyrosine residue phosphorylation of AT1R, which was attenuated by inhibition of Src but not PI3K activities. Valsartan also suppressed the interaction of eNOS and AT1R, which was blocked by Src or PI3K inhibition. CONCLUSION: Valsartan-induced NO production in ECs is mediated through Src/PI3K/Akt-dependent phosphorylation of eNOS. Valsartan-induced AT1R phosphorylation depends on Src but not PI3K, whereas valsartan-induced suppression of AT1R-eNOS interaction depends on Src/PI3K/Akt signalling. These results indicate a novel vasoprotective mechanism of valsartan in upregulating NO production in ECs.


Assuntos
Anti-Hipertensivos/farmacologia , Células Endoteliais/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Receptor Tipo 1 de Angiotensina/metabolismo , Tetrazóis/farmacologia , Valina/análogos & derivados , Animais , Aorta/citologia , Bovinos , Células Cultivadas , Humanos , Óxido Nítrico/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Valina/farmacologia , Valsartana , Quinases da Família src/metabolismo
19.
Life Sci ; 84(3-4): 97-104, 2009 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-19041881

RESUMO

AIMS: Resistin promotes macrophage-foam cell formation, but the mechanisms are unclear. In macrophages, lipid uptake is regulated by scavenger receptors (SR-A and CD36), while the cholesterol efflux is regulated by SR-BI, ATP-binding cassette transporter-A1 (ABCA1) and ABCG1. We investigated the mechanisms underlying the dysregulation by resistin of these regulators leading to promotion of lipid accumulation in bone marrow-derived macrophages. MAIN METHODS: Western blotting, real-time PCR and oil red O staining were performed. KEY FINDINGS: Resistin exacerbated lipid accumulation in oxLDL-treated macrophages. Resistin treatment of oxLDL-untreated macrophages showed increased SR-A and CD36 mRNA and protein levels, and decreased ABCA1 protein level, while having no effect on SR-BI or ABCG1 expression. Up-regulation of SR-A and CD36 by resistin resulted from activation of AP-1 and PPARgamma, respectively, and this was confirmed by the lack of activation of either after AP-1 inhibition using curcumin or SP600125, or PPARgamma inhibition using GW9662, respectively. The down-regulation of ABCA1 by resistin was not accompanied by a reduced mRNA level or an activation of LXRalpha/RXR, but resulted from enhanced protein degradation as revealed by the abolition of the down-regulation after inhibition of the proteasome pathway using ALLN or MG-132. A combined inhibition by SP600125, GW9662 and ALLN prevented resistin-induced exacerbation of lipid accumulation in oxLDL-treated macrophages. SIGNIFICANCE: Resistin promotes foam cell formation via dysregulation of SR-A, CD36 and ABCA1. SR-A and CD36 are transcriptionally up-regulated by resistin through AP-1 and PPARgamma, respectively, whereas ABCA1 is down-regulated by resistin through proteasome-mediated enhancement of protein degradation.


Assuntos
Transportadores de Cassetes de Ligação de ATP/fisiologia , Antígenos CD36/fisiologia , Metabolismo dos Lipídeos , Macrófagos/metabolismo , Resistina/fisiologia , Receptores Depuradores Classe A/fisiologia , Transportador 1 de Cassete de Ligação de ATP , Animais , Colesterol/metabolismo , Lipoproteínas LDL/fisiologia , PPAR gama/fisiologia , Complexo de Endopeptidases do Proteassoma/fisiologia , Ratos , Ratos Sprague-Dawley , Fator de Transcrição AP-1/fisiologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA