Browsing by Author "Vásquez, Rodrigo"

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    d-glucose increased l-arginine transport and nitric oxide synthesis through an autocrine mechanism involving TGF-β1 and TGF-β receptor II (TβRII) in human umbilical vein endothelium
    (2006) Vásquez, Rodrigo; Farías Jofré, Marcelo Enrique; San Martín, Rody; Casanello Toledo, Paola Cecilia; Sobrevía Luarte, Luis Alberto
    High D-glucose increases L-arginine transport, nitric oxide (NO) synthesis, and release of Transforming Growth Factor β1 (TGF-β1) in human umbilical vein endothelium (HUVEC). Changes in cell proliferation in response to D-glucose have been explained by a TGF-β1 autocrine mechanism in this cell type. However, the involvement of TGF-β1 and TGF-β1 receptor II (TβRII) on D-glucose effect on endothelial L-arginine/NO pathway has not been reported. L-[3H]Arginine transport (100 μM, 2 μCi/ml) and endothelial NO synthase (eNOS) activity [L-[3H]citrulline formation from L-[3H]arginine, 4 μCi/ml, 30 min, ± NG-nitro-L-arginine methyl ester (L-NAME), 100 μM] were determined in primary cultures of HUVEC exposed (6 h) to 5 mM (normal) or 25 mM (high) D-glucose, in absence or presence of TGF-β1 (2 ng/ml). Supernatant TGF-β1 level was measured by ELISA. HUVEC were infected (2% sera for 12 h) with an adenovirus expressing a negative dominant truncated TβRII (AdTβRIIt). Expression of TβRIIt was determined by Western blot. High D-glucose and TGF-β1 [half maximal effect (Ks): 0.28 ng/ml] increased L-arginine transport, effects that were significantly (P< 0.005) reduced in AdTβRIIt infected cells. L-Arginine transport was not further increased in non-infected infected cells co-incubated with high D-glucose and TGF-β1. However, L-arginine transport in AdTβRIIt infected cells co-incubated with these molecules was reduced. High D-glucose and TGF-β1 increased L-citrulline formation only in non-infected cells. High D-glucose also increased supernatant TGF-β1 level. We propose that stimulation of endothelial L-arginine/NO pathway by high D-glucose could result from a mechanism involving activation of TβRII by TGF-β1 as a consequence of increased release of this growth factor in HUVEC. Supported by FONDECYT 1030781 & 1030607 (Chile). R.V. holds a DIPUC-School of Medicine PhD (Chile) fellowship. M.F. holds CONICYT and School of Medicine–PhD (Chile) fellowships
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    Oxidative Damage of Lysozyme and Human Serum Albumin and Their Mixtures. A Comparison of Photosensitized and Peroxyl Radical Promoted Processes
    (2011) Arenas Lissi, Andrea Beatriz; Vásquez, Rodrigo; López Alarcón, Camilo Ignacio; Lissi, Eduardo; Silva Stevens, Eduardo Andrés
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    TGF-beta1 inhibits expression and activity of hENT1 in a nitric oxide-dependent manner in human umbilical vein endothelium
    (2009) Vega Pizarro, José Luis Eduardo; Puebla Aracena, Carlos Alberto; Vásquez, Rodrigo; Farías Jofré, Marcelo Enrique; Alarcón, Julio; Pastor-Anglada, Marçal; Krause Leyton, Bernardo; Casanello Toledo, Paola Cecilia; Sobrevía Luarte, Luis Alberto
    Aims: We studied whether transforming growth factor β1 (TGF-β1) modulates human equilibrative nucleoside transporters 1 (hENT1) expression and activity in human umbilical vein endothelial cells (HUVECs). hENT1-mediated adenosine transport and expression are reduced in gestational diabetes and hyperglycaemia, conditions associated with increased synthesis and release of nitric oxide (NO) and TGF-β1 in this cell type. TGF-β1 increases NO synthesis via activation of TGF-β receptor type II (TβRII), and NO inhibits hENT1 expression and activity in HUVECs. Methods and results: HUVECs (passage 2) were used for experiments. Total and hENT1-mediated adenosine transport was measured in the absence or presence of TGF-β1, NG-nitro-L-arginine methyl ester (L-NAME, NO synthase inhibitor), S-nitroso-N-acetyl-L,D-penicillamine (SNAP, NO donor), and/or KT-5823 (protein kinase G inhibitor) in control cells and cells expressing a truncated form of TGF-β1 receptor type II (TTβRII). Western blot and real-time PCR were used to determine hENT1 protein abundance and mRNA expression. SLC29A1 gene promoter and specific protein 1 (Sp1) transcription factor activity was assayed. Vascular reactivity was assayed in endothelium-intact or -denuded umbilical vein rings. TGF-β1 reduced hENT1-mediated adenosine transport, hENT1 protein abundance, hENT1 mRNA expression, and SLC29A1 gene promoter activity, but increased Sp1 binding to DNA. TGF-β1 effect was blocked by L-NAME and KT-5823 and mimicked by SNAP in control cells. However, TGF-β1 was ineffective in cells expressing TTβRII or a mutated Sp1 consensus sequence. Vasodilatation in response to TGF-β1 and S-(4-nitrobenzyl)-6-thio-inosine (an ENT inhibitor) was endothelium-dependent and blocked by KT-5823 and ZM-241385. Conclusion: hENT1 is down-regulated by activation of TβRII by TGF-β1 in HUVECs, a phenomenon where NO and Sp1 play key roles. These findings comprise physiological mechanisms that could be important in diseases where TGF-β1 plasma level is increased as in gestational diabetic mothers or patients with diabetes mellitus.