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Blockade of TGF-β 1 Signalling Inhibits Cardiac NADPH Oxidase Overactivity in Hypertensive Rats

2011-12-31T23:00:00Z

Title: Blockade of TGF-β 1 Signalling Inhibits Cardiac NADPH Oxidase Overactivity in Hypertensive Rats Author(s) : Miguel-Carrasco, J.L. (José Luis); Baltanas, A. (Ana); Cebrian, C. (Carolina); Moreno, M.U. (María Ujué); Lopez, B. (Begoña); Hermida, N. (Nerea); Gonzalez, A. (Arantxa); Dotor, J. (Javier); Borras-Cuesta, F. (Francisco); Diez, J. (Javier); Fortuño, A. (A.); Zalba, G. (Guillermo) Abstract: NADPH oxidases constitute a major source of superoxide anion (·O(2) (-)) in hypertension. Several studies suggest an important role of NADPH oxidases in different effects mediated by TGF-β 1. In this study we show that chronic administration of P144, a peptide synthesized from type III TGF-β 1 receptor, significantly reduced the cardiac NADPH oxidase expression and activity as well as in the nitrotyrosine levels observed in control spontaneously hypertensive rats (V-SHR) to levels similar to control normotensive Wistar Kyoto rats. In addition, P144 was also able to reduce the significant increases in the expression of collagen type I protein and mRNA observed in hearts from V-SHR. In addition, positive correlations between collagen expression, NADPH oxidase activity, and nitrotyrosine levels were found in all animals. Finally, TGF-β 1-stimulated Rat-2 exhibited significant increases in NADPH oxidase activity that was inhibited in the presence of P144. It could be concluded that the blockade of TGF-β 1 with P144 inhibited cardiac NADPH oxidase in SHR, thus adding new data to elucidate the involvement of this enzyme in the profibrotic actions of TGF-β 1.

Beneficial aspects of vascular apoptosis in hypertensive heart disease

1999-12-31T23:00:00Z

Title: Beneficial aspects of vascular apoptosis in hypertensive heart disease Author(s) : Diez, J. (Javier); Fortuño, A. (A.); Gonzalez, A. (Arantxa); Ravassa, S. (Susana); Zalba, G. (Guillermo) Abstract: Arterial hypertension may be viewed as a generalized vascular disorder, with an imbalance between proliferation and apoptosis of vascular smooth muscle cells. As a consequence exaggerated accumulation of these cells may result leading to an encroachment of the tunica media into the lumen. This geometric abnormality of the vessel wall may play a critical role in the long-term maintenance of elevated blood pressure and development of hypertensive endorgan damage. In this short paper, we summarize data on alterations in the growth and death of vascular smooth muscle cells in the small coronary arteries in hypertension.

NADPH oxidase-mediated oxidative stress

2004-12-31T23:00:00Z

Title: NADPH oxidase-mediated oxidative stress Author(s) : Zalba, G. (Guillermo); San-Jose, G. (Gorka); Moreno, M.U. (María Ujué); Fortuño, A. (A.); Diez, J. (Javier) Abstract: Increased vascular production of reactive oxygen species, especially superoxide anion, significantly contributes to the oxidative stress associated with hypertension. An enhanced superoxide production causes an increased inactivation of nitric oxide that diminishes nitric oxide bioavailability, thus contributing to endothelial dysfunction and hypertrophy of vascular cells. It has been shown that NADPH oxidases play a major role as the most important sources of superoxide anion in phagocytic and vascular cells. Several experimental observations have described an enhanced superoxide generation as a result of NADPH oxidase activation in hypertension. Although these enzymes respond to stimuli such as vasoactive factors, growth factors, and cytokines, recent data suggest a significant role of the genetic background in the modulation of the expression of its different components. Several polymorphisms have been identified in the promoter and in the coding region of CYBA, the gene that encodes the essential subunit of the NADPH oxidase p22phox, some of which seem to influence significantly the activity of these enzymes in the context of cardiovascular diseases. Among CYBA polymorphisms, genetic investigations have provided a novel marker, the -930(A/G) polymorphism, which determines the genetic susceptibility of hypertensive patients to oxidative stress.

Vascular oxidative stress and endothelial dysfunction

2000-12-31T23:00:00Z

Title: Vascular oxidative stress and endothelial dysfunction Author(s) : Zalba, G. (Guillermo); Gonzalez, A. (Arantxa); Beaumont, J. (Javier); San-Jose, G. (Gorka); Moreno, M.U. (María Ujué); Lopez, B. (Begoña); Ravassa, S. (Susana); Muñiz, P. (Paula); Fortuño, A. (A.); Fortuño, M.A. (María Antonia); Diez, J. (Javier)

Superoxide anion in the physiopathology of vascular diseases

2002-12-31T23:00:00Z

Title: Superoxide anion in the physiopathology of vascular diseases Author(s) : Zalba, G. (Guillermo); San-Jose, E. (Edurne); Moreno, M.U. (María Ujué); Diez, J. (Javier)

Oxidative stress in hypertension: from the SNPs to the clinical phenotype

2003-12-31T23:00:00Z

Title: Oxidative stress in hypertension: from the SNPs to the clinical phenotype Author(s) : Zalba, G. (Guillermo); Fortuño, A. (A.); San-Jose, G. (Gorka); Moreno, M.U. (María Ujué); Olivan, S. (Sara); Beloqui, O. (Óscar); Diez, J. (Javier)

Altered regulation of smooth muscle cell proliferation and apoptosis in small arteries of spontaneously hypertensive rats

1997-12-31T23:00:00Z

Title: Altered regulation of smooth muscle cell proliferation and apoptosis in small arteries of spontaneously hypertensive rats Author(s) : Diez, J. (Javier); Fortuño, M.A. (María Antonia); Zalba, G. (Guillermo); Etayo, J.C. (Juan Carlos); Fortuño, A. (A.); Ravassa, S. (Susana); Beaumont, J. (Javier) Abstract: AIM: It has been proposed that alterations of the balance between programmed cell death and cell replication might be involved in abnormalities of smooth muscle cell growth in arterial hypertension. This study was designed to analyse some regulators of apoptosis and proliferation in smooth muscle cells of small intra-myocardial arteries from the left ventricle of adult normotensive Wistar-Kyoto rats (WKY) and adult spontaneously hypertensive rats (SHR). Therefore, we assessed the expression of the cytoplasmic proteins Bax and Bcl-2, respectively a promoter and an inhibitor of apoptosis, and the expression of cyclin A, a nuclear protein that induces proliferation of smooth muscle cells. METHODS AND RESULTS: We measured the percentages of smooth muscle cells expressing these proteins using monoclonal antibodies and the avidin-biotin immunoperoxidase method. Compared with WKY, cells from SHR exhibited normal Bax expression, increased (P < 0.001) Bcl-2 expression and increased (P < 0.001) cyclin A expression. The ratio of Bax to Bcl-2, an index of cell susceptibility to apoptosis, was lower (P < 0.001) in SHR than in WKY. Systolic blood pressure was directly correlated (P < 0.01) with Bcl-2 and cyclin A in SHR. CONCLUSION: These results suggest that apoptosis and proliferation of smooth muscle cells might be inhibited and stimulated, respectively, in small arteries of adult SHR. The imbalance between these two processes may account for abnormalities of smooth muscle cell growth in the arterial wall in genetic hypertension.

Molecular mechanisms of atherosclerosis in metabolic syndrome: role of reduced IRS2-dependent signaling

2007-12-31T23:00:00Z

Title: Molecular mechanisms of atherosclerosis in metabolic syndrome: role of reduced IRS2-dependent signaling Author(s) : Gonzalez-Navarro, H. (Herminia); Vinue, A. (Angela); Vila-Caballer, M. (Marian); Fortuño, A. (A.); Beloqui, O. (Óscar); Zalba, G. (Guillermo); Burks, D. (Deborah); Diez, J. (Javier); Andres, V. (Vicente) Abstract: OBJECTIVE: The mechanisms underlying accelerated atherosclerosis in metabolic syndrome (MetS) patients remain poorly defined. In the mouse, complete disruption of insulin receptor substrate-2 (Irs2) causes insulin resistance, MetS-like manifestations, and accelerates atherosclerosis. Here, we performed human, mouse, and cell culture studies to gain insight into the contribution of defective Irs2 signaling to MetS-associated alterations. METHODS AND RESULTS: In circulating leukocytes from insulin-resistant MetS patients, Irs2 and Akt2 mRNA levels inversely correlate with plasma insulin levels and HOMA index and are reduced compared to insulin-sensitive MetS patients. Notably, a moderate reduction in Irs2 expression in fat-fed apolipoprotein E-null mice lacking one allele of Irs2 (apoE(-/-)Irs2(+/-)) accelerates atherosclerosis compared to apoE-null controls, without affecting plaque composition. Partial Irs2 inactivation also increases CD36 and SRA scavenger receptor expression and modified LDL uptake in macrophages, diminishes Akt2 and Ras expression in aorta, and enhances expression of the proatherogenic cytokine MCP1 in aorta and primary vascular smooth muscle cells (VSMCs) and macrophages. Inhibition of AKT or ERK1/2, a downstream target of RAS, upregulates Mcp1 in VSMCs. CONCLUSIONS: Enhanced levels of MCP1 resulting from reduced IRS2 expression and accompanying defects in AKT2 and Ras/ERK1/2 signaling pathways may contribute to accelerated atherosclerosis in MetS states.

Cardiotrophin-1 is expressed in adipose tissue and upregulated in the metabolic syndrome

2008-12-31T23:00:00Z

Title: Cardiotrophin-1 is expressed in adipose tissue and upregulated in the metabolic syndrome Author(s) : Natal, C. (Cristina); Fortuño, M.A. (María Antonia); Restituto, P. (Patricia); Bazan, A. (Antonio); Colina, I. (Inmaculada); Diez, J. (Javier); Varo, N. (Nerea) Abstract: Adipose tissue is a target for cardiotrophin-1 (CT-1), a cytokine member of the IL-6 family of cytokines that is involved in cardiac growth and dysfunction. However, it is unknown whether adipocytes are a source of CT-1 and whether CT-1 is overexpressed in diseases characterized by increased fat depots [i.e., the metabolic syndrome (MS)]. Thus this work aimed 1) to test whether adipose tissue expresses CT-1 and whether CT-1 expression can be modulated and 2) to compare serum CT-1 levels in subjects with and without MS diagnosed by National Cholesterol Education Program Adult Treatment Panel III criteria. Gene and protein expression of CT-1 was determined by real-time RT-PCR, ELISA, and Western blotting. CT-1 expression progressively increased, along with differentiation time from preadipocyte to mature adipocyte in 3T3-L1 cells. CT-1 expression was enhanced by glucose in a dose-dependent manner in these cells. mRNA and protein CT-1 expression was also demonstrated in human adipose biopsies. Immunostaining showed positive staining in adipocytes. Finally, increased CT-1 serum levels were observed in patients with MS compared with control subjects (127 +/- 9 vs. 106 +/- 4 ng/ml, P < 0.05). Circulating levels of CT-1 were associated with glucose levels (r = 0.2, P < 0.05). Taken together, our data suggest that adipose tissue can be recognized as a source of CT-1, which could account for the high circulating levels of CT-1 in patients with MS.

G protein-coupled receptor kinase 2 plays a relevant role in insulin resistance and obesity

2009-12-31T23:00:00Z

Title: G protein-coupled receptor kinase 2 plays a relevant role in insulin resistance and obesity Author(s) : Garcia-Guerra, L. (Lucía); Nieto-Vazquez, I. (Iria); Vila-Bedmar, R. (Rocío); Jurado-Pueyo, M. (María); Zalba, G. (Guillermo); Diez, J. (Javier); Murga, C. (Cristina); Fernandez-Veledo, S. (Sonia); Mayor, F.Jr (Federico Jr.); Lorenzo, M. (Margarita) Abstract: OBJECTIVE: Insulin resistance is associated with the pathogenesis of metabolic disorders as type 2 diabetes and obesity. Given the emerging role of signal transduction in these syndromes, we set out to explore the possible role that G protein-coupled receptor kinase 2 (GRK2), first identified as a G protein-coupled receptor regulator, could have as a modulator of insulin responses. RESEARCH DESIGN AND METHODS: We analyzed the influence of GRK2 levels in insulin signaling in myoblasts and adipocytes with experimentally increased or silenced levels of GRK2, as well as in GRK2 hemizygous animals expressing 50% lower levels of this kinase in three different models of insulin resistance: tumor necrosis factor-α (TNF-α) infusion, aging, and high-fat diet (HFD). Glucose transport, whole-body glucose and insulin tolerance, the activation status of insulin pathway components, and the circulating levels of important mediators were measured. The development of obesity and adipocyte size with age and HFD was analyzed. RESULTS: Altering GRK2 levels markedly modifies insulin-mediated signaling in cultured adipocytes and myocytes. GRK2 levels are increased by ∼2-fold in muscle and adipose tissue in the animal models tested, as well as in lymphocytes from metabolic syndrome patients. In contrast, hemizygous GRK2 mice show enhanced insulin sensitivity and do not develop insulin resistance by TNF-α, aging, or HFD. Furthermore, reduced GRK2 levels induce a lean phenotype and decrease age-related adiposity. CONCLUSIONS: Overall, our data identify GRK2 as an important negative regulator of insulin effects, key to the etiopathogenesis of insulin resistance and obesity, which uncovers this protein as a potential therapeutic target in the treatment of these disorders.

G protein-coupled receptor kinase 2 plays a relevant role in insulin resistance and obesity

2009-12-31T23:00:00Z

Association of the peroxisome proliferator-activated receptor α gene L162V polymorphism with stage C heart failure

2010-12-31T23:00:00Z

Title: Association of the peroxisome proliferator-activated receptor α gene L162V polymorphism with stage C heart failure Author(s) : Arias, T. (Teresa); Beaumont, J. (Javier); Lopez, B. (Begoña); Zalba, G. (Guillermo); Beloqui, O. (Óscar); Barba, J. (Joaquín); Valencia, F. (Félix); Gomez-Doblas, J.J. (Juan José); De-Teresa, E. (Eduardo); Diez, J. (Javier) Abstract: OBJECTIVE: To analyze whether genetic variants of PPARA are associated with the development of stage C heart failure. METHODS: We analyzed the distribution of the rs1800206, rs4253778 and rs135551 polymorphisms in genomic DNA extracted from peripheral blood cells of 534 patients in different heart failure stages and 63 healthy individuals. The mRNA expression of the peroxisome proliferator-activated receptor (PPAR)α target genes long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) and medium-chain acyl-CoA dehydrogenase (MCAD) was measured in myocardial biopsies of a subgroup of stage B and C patients. Functional studies were performed in HL-1 cardiomyocytes. RESULTS: The V162 allele of the rs1800206 polymorphism was more frequent in stage C patients than in stage A and B patients and healthy individuals. Patients with the V162 allele exhibited decreased myocardial LCHAD and MCAD mRNA expression as compared to L162 homozygote patients. In addition, stage C patients exhibited lower myocardial LCHAD and MCAD mRNA expression than stage B patients. Cardiomyocytes transfected with the V162 allele presented decreased PPARα transcriptional activity, LCHAD mRNA expression and ATP production compared to cardiomyocytes transfected with the L162 variant. CONCLUSIONS: These findings suggest that the V162 allele of the human PPARA gene can be a new risk factor in the development of stage C heart failure, likely via depressed cardiac PPARα activity

Losartan inhibits the post-transcriptional synthesis of collagen type I and reverses left ventricular fibrosis in spontaneously hypertensive rats

1998-12-31T23:00:00Z

Title: Losartan inhibits the post-transcriptional synthesis of collagen type I and reverses left ventricular fibrosis in spontaneously hypertensive rats Author(s) : Varo, N. (Nerea); Etayo, J.C. (Juan Carlos); Zalba, G. (Guillermo); Beaumont, J. (Javier); Iraburu, M.J. (María J.); Montiel-Duarte, C. (Cristina); Gil, M.J. (María José); Monreal, I. (Ignacio); Diez, J. (Javier) Abstract: 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.

Effects of loop diuretics on angiotensin II-stimulated vascular smooth muscle cell growth

2000-12-31T23:00:00Z

Title: Effects of loop diuretics on angiotensin II-stimulated vascular smooth muscle cell growth Author(s) : Muñiz, P. (Paula); Fortuño, A. (A.); Zalba, G. (Guillermo); Fortuño, M.A. (María Antonia); Diez, J. (Javier) Abstract: 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.

The loop diuretic torasemide interferes with endothelin-1 actions in the aorta of hypertensive rats

2000-12-31T23:00:00Z

Title: The loop diuretic torasemide interferes with endothelin-1 actions in the aorta of hypertensive rats Author(s) : Fortuño, A. (A.); Muñiz, P. (Paula); Zalba, G. (Guillermo); Fortuño, M.A. (María Antonia); Diez, J. (Javier) Abstract: 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.