RESUMO
BACKGROUND: Telomeres are nucleoprotein structures that protect the ends of eukaryote chromosomes. Shorter telomere length (TL) is associated with some age-related human disorders, but its relationship with obesity or adiposity parameters remains unclear. OBJECTIVE: The aim of this study was to assess the relationship between TL and changes in adiposity indices after a 5-year nutritional intervention. DESIGN AND SUBJECTS: TL was measured by quantitative real-time PCR in 521 subjects (55-80 years, 55% women). Participants were randomly selected from the PREDIMED-NAVARRA centre after they completed a 5-year intervention programme. Anthropometric parameters were directly measured by trained personnel at baseline and on a yearly basis thereafter. TL at baseline and changes in TL after a 5-year intervention were assessed. RESULTS: Higher baseline TL significantly predicted a greater decrease in body weight (B=-1.09 kg, 95% confidence interval (CI): -2.01 to -0.16), body mass index (BMI) (B=-0.47 kg m(-2), 95% CI: -0.83 to -0.11), waist circumference (B=-1.15 cm, 95% CI: -2.28 to -0.01) and waist to height ratio (B=-0.008, 95% CI: -0.010 to -0.001) in multiple-adjusted models. In addition, changes in TL during the 5-year intervention were inversely associated with changes in the four anthropometric variables. The reduction in adiposity indices during the intervention, associated with increasing TL, was even higher among subjects with the longest telomeres at baseline. Logistic regression analysis showed that the risk of remaining obese after 5 years was lower in those participants who initially had the longest telomeres and increased their TL after intervention (odds ratio=0.27, 95% CI: 0.03-2.03). CONCLUSIONS: Our research suggests that TL is inversely associated with changes in obesity parameters. The assessment of TL can provide further insights for biological pathways leading to adiposity. We show for the first time an improvement of obesity indices when an increase in TL is observed after a 5-year Mediterranean diet intervention.
Assuntos
Adiposidade/genética , Envelhecimento/metabolismo , Dieta Mediterrânea , Obesidade/genética , Encurtamento do Telômero , Telômero/genética , Idoso , Idoso de 80 Anos ou mais , Envelhecimento/genética , Biomarcadores/metabolismo , Índice de Massa Corporal , Dieta com Restrição de Gorduras , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Obesidade/epidemiologia , Obesidade/metabolismo , Fenótipo , Valor Preditivo dos Testes , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Risco , Homeostase do Telômero/genética , Encurtamento do Telômero/genética , Aumento de Peso/genéticaRESUMO
It has been shown that NADPH oxidase plays a role in oxidative stress which has been involved in the development of metabolic syndrome. The -930A/G polymorphism of the CYBA gene (that codes p22phox, a major component of the NADPH oxidase) has been associated with human hypertension and with a reduction in NADPH oxidase activity. In this work, we have examined the influence of the -930A/G polymorphism on obesity risk and insulin resistance in a case-control study of Spanish subjects (n=313). In the obese group (n=159), there was a statistically significant association between the GG genotype of the -930A/G polymorphism of the CYBA gene and fasting insulin levels and HOMA index. This outcome agrees with previous findings concerning functional analyses of this polymorphism and reinforces the hypothesis that insulin resistance is associated with oxidative stress. In conclusion, a protective effect in carriers of the -930A/G polymorphism of the p22phox gene against insulin resistance in a population of Spanish obese adults has been found.
Assuntos
Resistência à Insulina/genética , NADPH Oxidases/genética , Obesidade/genética , Polimorfismo Genético , Adulto , Alelos , Composição Corporal , Predisposição Genética para Doença , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , NADPH Oxidases/metabolismo , Análise de Regressão , Espanha , Adulto JovemRESUMO
BACKGROUND: Shorter telomeres have been associated with elevated risk for age-related diseases. However, little is known about the biomarker role of telomere length (TL) for predicting inflammation and glucose alterations. OBJECTIVE: The objective of this research is to evaluate the association between TL, inflammatory markers and glucose levels after a 2-month weight-loss programme in obese adolescents. METHODS: Telomere length was measured using a quantitative polymerase chain reaction in 66 obese adolescents aged 12-17 years (51% men) from the EVASYON programme. The adolescents were genotyped for the polymorphism -174G/C (rs1800795) in the IL-6gene, and anthropometric and biochemical markers as well as inflammatory cytokines were analysed. RESULTS: Multiple-adjusted models showed that longer telomeres at baseline were associated with a higher reduction in glucose (B = -4.08, 95% confidence interval: -6.66 to -1.50) and IL-6 (B = -1.03, 95% confidence interval: -2.01 to -0.05) serum levels after 2 months of the weight-loss treatment. The -174G/C polymorphism modulated the association between basal TL and changes in IL-6 (P interaction = 0.029). Thus, subjects with the GG + GC genotype and with longer telomeres showed a higher decrease in IL-6 levels than CC homozygotes. CONCLUSION: Longer telomeres are associated with an improvement in glucose tolerance and inflammation after a weight-loss programme in obese adolescents. Moreover, the -174G/C polymorphism may influence the relationship between TL and IL-6 changes.
Assuntos
Interleucina-6/genética , Obesidade Infantil/genética , Telômero/genética , Programas de Redução de Peso/métodos , Adolescente , Antropometria , Biomarcadores , Glicemia/genética , Criança , Feminino , Genótipo , Humanos , Inflamação/genética , Masculino , Obesidade Infantil/fisiopatologia , Obesidade Infantil/terapia , Polimorfismo Genético , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Introducción y objetivos: Los telómeros son regiones no codificantes localizadas al final de los cromosomas de células eucariotas, y su acortamiento se ha visto relacionado con la enfermedad cardiovascular y sus factores de riesgo. El objetivo de este estudio es evaluar la asociación entre el índice de salud cardiovascular ideal y el riesgo de telómero corto en una población de sujetos de edad avanzada de la cohorte Seguimiento Universidad de Navarra (SUN). Métodos: Se valoró a 886 adultos mayores de 55 años (645 varones y 241 mujeres). La longitud telomérica se midió utilizando qPCR (quantitative protein chain reaction) en tiempo real y reacción única. El índice de salud cardiovascular «Life's simple 7» se definió según la American Heart Association mediante la puntuación de 7 ítems con valores de 0 a 2 para cada uno: tabaquismo, actividad física, dieta, índice de masa corporal, presión arterial, colesterol total y glucosa en sangre. La máxima puntuación del índice corresponde a 14 puntos. Se categorizó en terciles: pobre (0-9 puntos), intermedio (10-11 puntos) e ideal (12-14 puntos). El riesgo de telómero corto se definió como una longitud telomérica por debajo del percentil 20. Resultados: Sujetos con altos valores en el índice de salud cardiovascular ideal tenían menos riesgo de telómero corto (OR ajustada=0,60; IC95%, 0,34-1,05; p de tendencia lineal=0,052). Esta asociación fue significativa en varones (OR ajustada=0,37; IC95%, 0,17-0,83; p de tendencia lineal=0,025), pero no en mujeres. Conclusiones: En varones mayores de 55 años, existe una asociación inversa entre el índice de salud cardiovascular y el riesgo de tener telómeros cortos (AU)
Introduction and objectives: Telomeres are noncoding regions located at the end of chromosomes and their shortening has been associated with risk factors and cardiovascular disease. The aim of this study was to evaluate the association between ideal cardiovascular health (Life's simple 7) and the odds of having short telomeres in a subsample of participants older than 55 years from the Seguimiento Universidad de Navarra (SUN) study. Methods: We included 886 participants older than 55 years (645 men and 241 women). Telomere length was measured using a real-time quantitative polymerase chain reaction. Cardiovascular health score was defined by the American Heart Association as a composite score of 7 key risk factors (smoking status, physical activity, diet, body mass index, blood pressure, total cholesterol, and fasting blood glucose) with 0 to 2 points for each factor. We categorized this score in tertiles as poor (0-9 points), intermediate (10-11 points) and ideal (12-14 points). The odds of having short telomeres was defined as telomere length below the 20th percentile. Results: Individuals with higher ideal cardiovascular health had a lower prevalence of having short telomeres (adjusted OR, 0.60; 95%CI, 0.34-1.05; P trend=.052). This association was statistically significant in men (adjusted OR, 0.37; 95%CI, 0.17-0.83; P trend=.025) but not in women. Conclusions: An inverse association between cardiovascular health score and short telomeres was found especially for men older than 55 years in the SUN population (AU)
Assuntos
Humanos , Masculino , Feminino , Pessoa de Meia-Idade , Idoso , Doenças Cardiovasculares/genética , Exercício Físico , Telômero/genética , Estilo de Vida , Índice de Massa Corporal , Fatores de Risco , Reação em Cadeia da Polimerase , Estudos ProspectivosRESUMO
In a previous study, we found that the p22(phox) subunit of the NADH/NADPH oxidase is overexpressed in vascular smooth muscle cells (VSMCs) from spontaneously hypertensive rats (SHRs) with enhanced vascular production of superoxide anion ((.)O(2)(-)). Thus, we have investigated whether changes in the sequence or activity of the promoter region of p22(phox) gene are present in SHRs. To carry out this analysis, first of all, we characterized the rat gene structure and promoter region for the p22(phox) subunit. The p22(phox) gene spans approximately 10 kb and contains 6 exons and 5 introns. Primer extension analysis indicated the transcriptional start site 100 bp upstream from the translational start site. The immediate promoter region of the p22(phox) gene does not contain a TATA box, but there are a CCAC box and putative recognition sites for nuclear factors, such as SP1, gamma-interferon, and nuclear factor-kappaB. Using reporter-gene transfection analysis, we found that this promoter was functional in VSMCs. Furthermore, we observed that p22(phox) promoter activity was significantly higher in VSMCs from SHRs than from normotensive Wistar-Kyoto rats. In addition, we found that there were 5 polymorphisms in the sequence of p22(phox) promoter between Wistar-Kyoto rats and SHRs and that they were functional. The results obtained in this study provide a tool to explore the mechanisms that regulate the expression of p22(phox) gene in rat VSMCs. Furthermore, our findings show that changes in the sequence of p22(phox) gene promoter and in the degree of activation of VSMCs are responsible for upregulated expression of p22(phox) in SHRs.
Assuntos
Proteínas de Membrana Transportadoras , Músculo Liso Vascular/metabolismo , NADPH Desidrogenase/genética , NADPH Desidrogenase/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Polimorfismo Genético/genética , Regiões Promotoras Genéticas/genética , Regiões 5' não Traduzidas/genética , Animais , Sequência de Bases , Sítios de Ligação/genética , Células Cultivadas , Clonagem Molecular , Éxons/genética , Genes Reporter , Biblioteca Genômica , Íntrons/genética , Dados de Sequência Molecular , Músculo Liso Vascular/citologia , Mutagênese Sítio-Dirigida , NADPH Oxidases , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Análise de Sequência de DNA , Especificidade da Espécie , Superóxidos/metabolismo , Transfecção , Regulação para Cima/genéticaRESUMO
Torasemide is a loop diuretic that is effective at low once-daily doses in the treatment of arterial hypertension. Because its antihypertensive mechanism of action may not be based entirely on the elimination of salt and water from the body, a vasodilator effect of this drug can be considered. In the present study, the ability of different concentrations of torasemide to modify angiotensin II (Ang II)-induced vascular responses was examined, with the use of an organ bath system, in endothelium-denuded aortic rings from spontaneously hypertensive rats. Ang II-induced increases of intracellular free calcium concentration ([Ca(2+)](i)) were also examined by image analysis in cultured vascular smooth muscle cells (VSMCs) from spontaneously hypertensive rats. A dose-response curve to Ang II was plotted for cumulative concentrations (from 10(-9) to 10(-6) mol/L) in endothelium-denuded aortic rings (pD(2)=7.5+/-0.3). Isometric contraction induced by a submaximal concentration of Ang II (10(-7) mol/L) was reduced in a dose-dependent way by torasemide (IC(50)=0.5+/-0.04 micromol/L). Incubation of VSMCs with different concentrations of Ang II (from 10(-10) to 10(-6) mol/L) resulted in a dose-dependent rise of [Ca(2+)](i) (pD(2)=7.5+/-0.3). The stimulatory effect of [Ca(2+)](i) induced by a submaximal concentration of Ang II (10(-7) mol/L) was blocked by torasemide (IC(50)=0.5+/-0.3 nmol/L). Our findings suggest that torasemide blocks the vasoconstrictor action of Ang II in vitro. This action can be related to the ability of torasemide to block the increase of [Ca(2+)](i) induced by Ang II in VSMCs. It is proposed that these actions might be involved in the antihypertensive effect of torasemide observed in vivo.
Assuntos
Anti-Hipertensivos/farmacologia , Aorta/metabolismo , Cálcio/metabolismo , Hipertensão/fisiopatologia , Membranas Intracelulares/metabolismo , Sulfonamidas/farmacologia , Vasoconstrição/efeitos dos fármacos , Angiotensina II/farmacologia , Animais , Aorta/efeitos dos fármacos , Compostos de Bifenilo/farmacologia , Células Cultivadas , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/fisiologia , Furosemida/farmacologia , Hipertensão/metabolismo , Irbesartana , Masculino , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Ratos , Ratos Endogâmicos SHR , Tetrazóis/farmacologia , TorasemidaRESUMO
An association of increased apoptosis with overexpression of the proapoptotic protein Bax-alpha has been reported in the left ventricle of adult spontaneously hypertensive rats (SHR). Both alterations were corrected in SHR that received long-term treatment with the AT1 antagonist losartan. To gain insight into the regulation of cardiac Bax-alpha protein in genetic hypertension, we investigated the expression of the protein p53 (a BAX gene transcription factor) and BAX mRNA in the left ventricle of 30-week-old Wistar-Kyoto rats (WKY), SHR, and SHR treated with losartan (20 mg. kg-1. d-1) during 14 weeks before death. The expression of p53 and Bax proteins was assessed by Western blot analysis. The expression of BAX mRNA was assessed by Northern blot analysis. The density of apoptotic cells was assessed by direct immunoperoxidase detection of biotin-labeled deoxyuridine nucleotides. Compared with WKY, untreated SHR exhibited increased apoptosis (P<0.05), increased Bax-alpha protein (P<0.05), and similar levels of p53 protein and BAX mRNA. Losartan given long term was associated with the normalization of apoptosis and Bax-alpha protein expression. The expression of BAX mRNA was decreased (P<0. 05) in treated SHR compared with untreated SHR. No changes in the expression of p53 protein were observed in losartan-treated SHR. These results suggest that overexpression of the Bax-alpha protein seen in the left ventricle of adult SHR with increased apoptosis is not related to a p53-mediated upregulation of BAX gene transcription. Our data also suggest that normalization of Bax-alpha protein observed in SHR after long-term blockade of angiotensin II type 1 receptors may be due to the inhibition of BAX gene transcription.
Assuntos
Regulação da Expressão Gênica , Hipertensão/genética , Miocárdio/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2 , Proteínas Proto-Oncogênicas/genética , Transcrição Gênica , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose , Técnica Direta de Fluorescência para Anticorpo , Regulação da Expressão Gênica/efeitos dos fármacos , Ventrículos do Coração , Hipertensão/metabolismo , Hipertensão/patologia , Losartan/farmacologia , Miocárdio/citologia , Miocárdio/patologia , Proteínas Proto-Oncogênicas/análise , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Valores de Referência , Transcrição Gênica/efeitos dos fármacos , Proteína X Associada a bcl-2RESUMO
OBJECTIVE: Previous studies have shown that as well as left ventricular hypertrophy, myocardial fibrosis develops early in rats with spontaneous hypertension (SHR). The present study was designed to investigate whether chronic treatment with the angiotensin II type 1 (AT1) receptor antagonist losartan modifies collagen type I metabolism and reverses left ventricular fibrosis in young SHR with left ventricular hypertrophy. DESIGN: The study was performed in 30-week-old normotensive Wistar-Kyoto (WKY) rats, untreated SHR and SHR treated with losartan (20 mg/mg per day, orally) for 14 weeks before they were killed. METHODS: Ventricular pro-alpha 1 (I) collagen messenger RNA was analyzed by Northern blot. Serum levels of the carboxy-terminal propeptide of procollagen type I (PIP) and the pyridoline cross-linked telopeptide domain of collagen type I (CITP) were determined by specific radioimmunoassays as markers of collagen type I synthesis and degradation, respectively. Collagen volume fraction was determined in the left ventricle by quantitative morphometry. RESULTS: Compared with WKY rats, SHR exhibited increased (P < 0.05) mean arterial pressure, pro-alpha 1 (I) collagen messenger RNA, PIP and left ventricular collagen volume fraction, and similar CITP values. After the treatment period, mean arterial pressure was higher (P < 0.05) in losartan-treated SHR than in WKY rats. Compared with untreated SHR, treated SHR showed no left ventricular hypertrophy and diminished (P < 0.05) values of mean arterial pressure, PIP and left ventricular collagen volume fraction. No changes in pro-alpha 1 (I) collagen messenger RNA and CITP values were observed with treatment in SHR. No significant differences in the left ventricular collagen volume fraction were observed between treated SHR with normal blood pressure and treated SHR with abnormally high blood pressure at the end of the treatment period. CONCLUSIONS: These results suggest that chronic AT1 blockade with losartan decreases the post-transcriptional synthesis of fibril-forming collagen type I molecules in young SHR. This effect may be involved in the ability of this drug to reverse left ventricular fibrosis in young rats with genetic hypertension. Apart from its antihypertensive action, other mechanisms may mediate the antifibrotic effect of losartan in this animal model.
Assuntos
Anti-Hipertensivos/farmacologia , Cardiomiopatias/tratamento farmacológico , Colágeno/metabolismo , Hipertensão/tratamento farmacológico , Losartan/farmacologia , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Transcrição Gênica , Animais , Biomarcadores/sangue , Pressão Sanguínea/efeitos dos fármacos , Northern Blotting , Cardiomiopatias/metabolismo , Cardiomiopatias/patologia , Colágeno/sangue , Colágeno/genética , Colágeno Tipo I , Fibrose/tratamento farmacológico , Fibrose/metabolismo , Fibrose/patologia , Ventrículos do Coração/metabolismo , Ventrículos do Coração/patologia , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Hipertrofia Ventricular Esquerda/tratamento farmacológico , Hipertrofia Ventricular Esquerda/metabolismo , Hipertrofia Ventricular Esquerda/patologia , Masculino , Peptídeos/sangue , Pró-Colágeno/sangue , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKYRESUMO
The term oxidative stress refers to a situation in which cells are exposed to excessive levels of either molecular oxygen or chemical derivatives of oxygen (ie, reactive oxygen species). Three enzyme systems produce reactive oxygen species in the vascular wall: NADH/NADPH oxidase, xanthine oxidoreductase, and endothelial nitric oxide synthase. Among vascular reactive oxygen species superoxide anion plays a critical role in vascular biology because it is the source for many other reactive oxygen species and various vascular cell functions. It is currently thought that increases in oxidant stress, namely excessive production of superoxide anion, are involved in the pathophysiology of endothelial dysfunction that accompanies a number of cardiovascular risk factors including hypercholesterolemia, hypertension and cigarette smoking. On the other hand, vascular oxidant stress plays a pivotal role in the evolution of clinical conditions such as atherosclerosis, diabetes and heart failure.
Assuntos
Vasos Sanguíneos/enzimologia , Estresse Oxidativo/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Animais , Vasos Sanguíneos/metabolismo , Doenças Cardiovasculares/etiologia , Humanos , Complexos Multienzimáticos/metabolismo , NADH NADPH Oxirredutases/metabolismo , Óxido Nítrico/metabolismo , Peróxidos , Superóxidos/metabolismoRESUMO
1. o-Iodosobenzoic acid (IOB) caused the formation of a disulfide bridge between SH1 and SH2 groups of myosin SF1 rendering inactive its ATPase activity. 2. IOB at high concentrations provoked fragmentation of SF1 at its tryptophan residues. 3. The main fragmentation point was located at 15 K from the amino terminus of the myosin heavy chain. 4. Actin was not fragmented by IOB. It protected SF1 tryptophans from IOB attack. 5. These results suggest a possible use of IOB as a reagent to study protein tryptophan under nondenaturing conditions.
Assuntos
Iodobenzoatos/farmacologia , Miosinas/metabolismo , Reagentes de Sulfidrila/farmacologia , Actinas/metabolismo , Animais , Galinhas , Dissulfetos/análise , Ditiotreitol/farmacologia , Eletroforese , Cinética , Subfragmentos de Miosina/metabolismo , Oxirredução , TripsinaRESUMO
Hypertensive heart disease is a progressive condition in which the compensatory left ventricular hypertrophy that maintains cardiac output leads to myocardial remodeling, characterized by fibrosis, insufficient vascularization, and alterations in cardiomyocytes, including contractile disturbances, changes in gene expression, and decrease in the number of cells. Structural abnormalities in the myocardial wall accelerate the development of diastolic and systolic dysfunction, resulting in heart failure. Many observations point to the apoptotic cell death of cardiomyocytes as a relevant factor in the transition from compensatory hypertrophy to pump failure in experimental and human hypertension. Potential inducers of cardiomyocyte apoptosis in overloaded hearts include extrinsic factors, such as mechanical forces, neurohormonal activation, oxidative stress, hypoxia, and cytokines. Some lines of evidence indicate that angiotensin II and the overstretching of cardiomyocytes are originally involved in the triggering of apoptosis in hypertension, whereas other factors are being investigated. Furthermore, intracellular changes, such as downregulation of survival proteins or activation of death proteins, seem to play an important role. The assumption that the apoptosis of cardiomyocytes worsens hypertensive heart disease prognosis brings forth new approaches to avoid or slow the transition to pump failure. In this respect, experimental data indicate that currently used antihypertensive drugs interfere with cardiomyocyte apoptosis. Moreover, the knowledge of intracellular apoptotic processes in cardiomyocytes provides novel therapeutic strategies to be added to the multimodal approach in the prevention of heart failure.
Assuntos
Apoptose , Hipertensão/fisiopatologia , Angiotensina II/metabolismo , Animais , Anti-Hipertensivos/farmacologia , Apoptose/efeitos dos fármacos , Humanos , Hipertensão/tratamento farmacológico , Miocárdio/metabolismo , Estresse Mecânico , Remodelação VentricularRESUMO
BACKGROUND: Torasemide and furosemide are diuretics that inhibit the Na(+), K(+), 2Cl(-) co-transporter localized in cells from the ascending limb of the loop of Henle. The effects of torasemide and furosemide on cell growth induced by angiotensin II (Ang II) were investigated in cultured vascular smooth muscle cells (VSMCs) obtained from the aorta of adult spontaneously hypertensive rats (SHR). METHODS: Cell growth was determined by DNA and protein synthesis as measured by [3H]thymidine and [3H]leucine incorporation, respectively. Proliferation of VSMCs was measured using a non-radioactive colorimetric cell proliferation assay. RESULTS: Ang II (10(-7) M) signficantly increased DNA and protein synthesis and cell proliferation in VSMCS: These effects were completely abolished by the Ang II type 1 receptor antagonist irbesartan (10(-6) M). Ang II-induced [3H]leucine incorporation was reduced in a dose-dependent way by torasemide (IC(50) value: 7.7+/-0.8x10(-7) M) but not by furosemide. Neither torasemide nor furosemide modified Ang II-stimulated [3H]thymidine incorporation or proliferation in VSMCs. CONCLUSIONS: These results indicate that torasemide, but not furosemide, inhibits Ang II-induced protein synthesis in VSMCs from SHR. Thus, it is suggested that the capacity of torasemide to block this trophic action of Ang II in rat VSMCs is not mediated by inhibition of the Na(+), K(+), 2Cl(-) co-transport mechanism.
Assuntos
Angiotensina II/farmacologia , Divisão Celular/efeitos dos fármacos , Diuréticos/farmacologia , Furosemida/farmacologia , Músculo Liso Vascular/citologia , Sulfonamidas/farmacologia , Antagonistas de Receptores de Angiotensina , Animais , Aorta Torácica/citologia , Aorta Torácica/efeitos dos fármacos , Compostos de Bifenilo/farmacologia , Proteínas de Transporte/antagonistas & inibidores , Células Cultivadas , Cloretos/metabolismo , Irbesartana , Leucina/metabolismo , Alça do Néfron/fisiologia , Músculo Liso Vascular/efeitos dos fármacos , Potássio/metabolismo , Ratos , Ratos Endogâmicos SHR , Receptor Tipo 1 de Angiotensina , Receptor Tipo 2 de Angiotensina , Sódio/metabolismo , Simportadores de Cloreto de Sódio-Potássio , Tetrazóis/farmacologia , Timidina/metabolismo , TorasemidaRESUMO
BACKGROUND: The direct effects of torasemide and furosemide on vasoconstriction and increases in intracellular free calcium concentration ([Ca(2+)]i) induced by endothelin-1 (ET-1) were investigated in the aorta of spontaneously hypertensive rats (SHR). METHODS: Vascular responses were assessed in endothelium-denuded aortic rings using an organ bath system. Changes of [Ca(2+)]i in cultured vascular smooth muscle cells (VSMCs) were assessed using fura-2 methodology. RESULTS: ET-1-induced vasoconstriction was reduced in a dose-dependent way by torasemide and furosemide (IC(50) values: 4.3+/-1.4x10(-5) and 9.8+/-5.6 x10(-5) M, respectively). The ET-1-induced biphasic [Ca(2+)]i increase was blocked by torasemide (IC(50)=2.0+/-0.2x10(-8) and 2.7+/-0.6x10(-6) M, respectively). Furosemide inhibited the initial rise in [Ca(2+)]i induced by ET-1, with no effect on the second rise. The specific chloride (Cl(-)) channel blocker diphenylamine-2-carboxylate inhibits both ET-1-induced responses in VSMCs (IC(50)=8.0+/-0.3x10(-9) and 2.5+/-0.7x10(-7) M, respectively). CONCLUSIONS: These results suggest that the ability of loop diuretics to interfere with the vascular actions of ET-1 may involve different molecular mechanisms. The ability of torasemide to block the vasoconstrictive action of ET-1 could include an inhibitory action on Cl(-) channels.
Assuntos
Aorta Torácica/efeitos dos fármacos , Diuréticos/farmacologia , Endotelina-1/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Sulfonamidas/farmacologia , Vasoconstrição/efeitos dos fármacos , Animais , Anti-Hipertensivos/farmacologia , Aorta Torácica/fisiologia , Aorta Torácica/fisiopatologia , Bosentana , Cálcio/metabolismo , Canais de Cloreto/antagonistas & inibidores , Endotélio Vascular/fisiologia , Furosemida/farmacologia , Técnicas In Vitro , Cinética , Músculo Liso Vascular/fisiologia , Músculo Liso Vascular/fisiopatologia , Ratos , Ratos Endogâmicos SHR , Torasemida , Vasoconstrição/fisiologia , ortoaminobenzoatos/farmacologiaRESUMO
Using a combination of screening, RACE, and RT-PCR, we have isolated a new rat brain cDNA, we refer to as rMNK2, that showed strong homology to known MAP-kinases. The deduced amino acid sequence of rMNK2 indicated that it is the rat homolog of human p63(mapk), showing 94.5% identity. rMNK2 showed 77% homology with rat ERK3 and its human homolog p97(mapk), and 43% homology with both rat genes rMNK1(ERK1) and ERK2, within the kinase domain. This suggest that rMNK2 and ERK3 belong to a separate subfamily within the rat MAP-kinase multigene family. The most interesting difference lies in subdomain VIII, where this new subfamily contain a SEG/SPR motif instead of the TEY/APE found in the ERK subfamily, the TPY/APE found in the JNK/SAPK subfamily or the TGY/APE found in the p38/RK subfamily. The human homologs of ERK3 and rMNK2 (p97(mapk) and p63(mapk)) also show this significant change. Expression of rMNK2 has been detected in brain and to a lesser extent in lung by reverse transcription/PCR (RT-PCR). In situ hybridization of rat brain slices demonstrated a restricted expression of rMNK2 in the choroid plexus and hippocampus. This is interesting because the human homolog p63(mapk) maps to 18q12-21, a region that might be implicated in manic-depressive illness.
Assuntos
Proteínas Quinases Dependentes de Cálcio-Calmodulina/biossíntese , Proteínas Quinases Dependentes de Cálcio-Calmodulina/química , MAP Quinases Reguladas por Sinal Extracelular , Proteínas Quinases Ativadas por Mitógeno/biossíntese , Proteínas Quinases Ativadas por Mitógeno/química , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Sequência Conservada , DNA Complementar/isolamento & purificação , Humanos , Hibridização In Situ , Proteína Quinase 6 Ativada por Mitógeno , Proteínas Quinases Ativadas por Mitógeno/genética , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Ratos , Homologia de Sequência de AminoácidosRESUMO
The complete genomic structure of the human p44(mapk) gene (HMGW-approved symbol PRKM3) has been determined. The gene covers 9 kb and is composed of nine exons and eight introns. This structure is identical to the previously reported mouse p44(mapk) gene, indicating a high degree of evolutionary conservation. A sequence differing by one nucleotide from the consensus TATA box is present 132 positions upstream of the main transcription initiation point. This point has been located 415 nucleotides upstream of the translation initiation codon ATG and perfectly meets the consensus criteria for an initiator element (Inr). Multiple consensus sequences for factors that regulate either basal transcription or gene expression during cell differentiation and proliferation can be found in the putative promoter region. Some of them, such as several G/C boxes located downstream from the transcription initiation point, are also present in the homologous mouse gene, where they were shown to be functional.
Assuntos
Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Proteínas Quinases Ativadas por Mitógeno , Regiões 5' não Traduzidas/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas Quinases Dependentes de Cálcio-Calmodulina/química , Clonagem Molecular , Códon de Iniciação/genética , Sequência Consenso , Éxons , Humanos , Íntrons , Camundongos , Proteína Quinase 3 Ativada por Mitógeno , Dados de Sequência Molecular , Iniciação Traducional da Cadeia Peptídica/genética , Regiões Promotoras Genéticas/genética , Sequências Reguladoras de Ácido NucleicoRESUMO
Previous findings have shown that hypotensive doses of losartan prevent the excess of apoptosis present in the hypertrophied left ventricle of adult spontaneously hypertensive rats (SHR). This study was designed to determine whether angiotensin II facilitates apoptosis in cardiomyocytes of adult SHR. Primary cultures of ventricular cardiomyocytes from 30-week-old normotensive Wistar-Kyoto rats (WKY) and SHR with left ventricular hypertrophy were exposed to 10(-)(9) mol/L angiotensin II for 24 hours. Apoptotic cells were assessed by terminal deoxynucleotidyl transferase assay and confirmed by Annexin V detection. The expression of Bax-alpha, Bcl-2, p53, and caspase-3 proteins was assessed by Western blot assays. The expression of BAX gene was assessed by Northern blot. Angiotensin II increased (P<0.01) cardiomyocyte apoptosis, and this effect was higher (P<0.001) in SHR cells than in WKY cells. Whereas losartan (10(-7) mol/L) blocked the apoptotic effect of the octapeptide in cells from the two strains of rats, PD123319 (10(-7) mol/L) inhibited angiotensin II-mediated apoptosis only in SHR cells. Angiotensin II stimulated (P<0.01) Bax-alpha protein, and this effect was higher (P<0.01) in SHR cells than in WKY cells. Angiotensin II did not modify Bcl-2, p53, and BAX mRNA in cells from the two strains of rats. Angiotensin II induced a similar increase (P<0.05) in the ratio caspase-3/procaspase-3 (an index of caspase-3 activation) in cardiomyocytes from the two strains of rats. The present in vitro results indicate that SHR cardiomyocytes exhibit enhanced susceptibility to angiotensin II-induced apoptosis. Ligand binding to angiotensin II type 1 and type 2 receptors leading to changes in posttranscriptional processing of Bax-alpha and accumulation of this proapoptotic protein may be involved in the abnormal response of SHR cardiomyocytes. These data support a role for angiotensin II in apoptosis observed in the left ventricle of these rats.
Assuntos
Angiotensina II/farmacologia , Apoptose , Coração/efeitos dos fármacos , Hipertensão/patologia , Miocárdio/patologia , Animais , Pressão Sanguínea , Caspase 3 , Caspases/metabolismo , Células Cultivadas , Ativação Enzimática , Ventrículos do Coração/efeitos dos fármacos , Ventrículos do Coração/patologia , Hipertensão/complicações , Hipertrofia Ventricular Esquerda/etiologia , Hipertrofia Ventricular Esquerda/patologia , Masculino , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Proteína Supressora de Tumor p53/metabolismo , Proteína X Associada a bcl-2RESUMO
Increased vascular reactive oxygen species production, especially superoxide anion, contributes significantly in the functional and structural alterations present in hypertension. An enhanced superoxide production causes a diminished NO bioavailability by an oxidative reaction that inactivates NO. Exaggerated superoxide levels and a low NO bioavailability lead to endothelial dysfunction and hypertrophy of vascular cells. It has been shown that the enzyme NAD(P)H oxidase plays a major role as the most important source of superoxide anion in vascular cells. Several experimental observations have shown an enhanced superoxide generation as a result of the activation of vascular NAD(P)H oxidase in hypertension. Although this enzyme responds to stimuli such as vasoactive factors, growth factors, and cytokines, some recent data suggest the existence of a genetic background modulating the expression of its different components. New polymorphisms have been identified in the promoter of the p22(phox) gene, an essential subunit of NAD(P)H oxidase, influencing the activity of this enzyme. Genetic investigations of these polymorphisms will provide novel markers for determination of genetic susceptibility to oxidative stress in hypertension.