Browsing by Author "Farías Jofré, Marcelo Enrique"
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- ItemAteroesclerosis subclínica y síndrome metabólico en niños(2013) Arnáiz Gómez, Pilar; Barja Y., Salesa; Villarroel del Pino, Luis A.; Domínguez, Angélica; Godoy J., Iván; Castillo Valenzuela, Oscar; Farías Jofré, Marcelo Enrique; Mardones, Francisco
- ItemDifferential immunophenotype of circulating monocytes from pregnant women in response to viral ligands(2023) Farías Jofré, Marcelo Enrique; Romero, Roberto; Xu, Yi; Levenson, Dustyn; Tao, Li; Kanninen, Tomi; Galaz, Jose; Arenas-Hernandez, Marcia; Liu, Zhenjie; Miller, Derek; Bhatti, Gaurav; Seyerle, Megan; Tarca, Adi L.; Gomez-Lopez, NardhyBackground Viral infections during pregnancy can have deleterious effects on mothers and their offspring. Monocytes participate in the maternal host defense against invading viruses; however, whether pregnancy alters monocyte responses is still under investigation. Herein, we undertook a comprehensive in vitro study of peripheral monocytes to characterize the differences in phenotype and interferon release driven by viral ligands between pregnant and non-pregnant women. Methods Peripheral blood was collected from third-trimester pregnant (n = 20) or non-pregnant (n = 20, controls) women. Peripheral blood mononuclear cells were isolated and exposed to R848 (TLR7/TLR8 agonist), Gardiquimod (TLR7 agonist), Poly(I:C) (HMW) VacciGrade™ (TLR3 agonist), Poly(I:C) (HMW) LyoVec™ (RIG-I/MDA-5 agonist), or ODN2216 (TLR9 agonist) for 24 h. Cells and supernatants were collected for monocyte phenotyping and immunoassays to detect specific interferons, respectively. Results The proportions of classical (CD14hiCD16−), intermediate (CD14hiCD16+), non-classical (CD14loCD16+), and CD14loCD16− monocytes were differentially affected between pregnant and non-pregnant women in response to TLR3 stimulation. The proportions of pregnancy-derived monocytes expressing adhesion molecules (Basigin and PSGL-1) or the chemokine receptors CCR5 and CCR2 were diminished in response to TLR7/TLR8 stimulation, while the proportions of CCR5− monocytes were increased. Such differences were found to be primarily driven by TLR8 signaling, rather than TLR7. Moreover, the proportions of monocytes expressing the chemokine receptor CXCR1 were increased during pregnancy in response to poly(I:C) stimulation through TLR3, but not RIG-I/MDA-5. By contrast, pregnancy-specific changes in the monocyte response to TLR9 stimulation were not observed. Notably, the soluble interferon response to viral stimulation by mononuclear cells was not diminished in pregnancy. Conclusions Our data provide insight into the differential responsiveness of pregnancy-derived monocytes to ssRNA and dsRNA, mainly driven by TLR8 and membrane-bound TLR3, which may help to explain the increased susceptibility of pregnant women to adverse outcomes resulting from viral infection as observed during recent and historic pandemics.
- ItemFoetoplacental epigenetic changes associated with maternal metabolic dysfunction(2018) Kerr, Bredford; Leiva Mendoza, Andrea Alejandra; Farías Jofré, Marcelo Enrique; Contreras Duarte, Susana de las Mercedes; Toledo, Fernando; Stolzenbach, Francisca; Silva, Luis; Sobrevía Luarte, Luis Alberto
- ItemHypoxia increases equilibrative nucleoside transporter 2 activity by a transcriptional independent mechanism in human umbilical vein endothelium(2006) Torres, A; San Martin, R; Farías Jofré, Marcelo Enrique; Sobrevía Luarte, Luis Alberto; Casanello Toledo, Paola CeciliaLow oxygen tension (hypoxia) reduces adenosine transport in several types of mammalian cells. Adenosine transport is mediated by human equilibrative nucleoside transporter 1 (hENT1) and hENT2 in human umbilical vein endothelium (HUVEC), a fetal cell type that grows under 5% O2 (ie. normoxia for this cell type). We studied whether hypoxia alters hENT2 expression and activity in HUVEC. Methods: Cells were cultured (0-24 h) in 5% or 2% O2 (hypoxia), and [3H]adenosine uptake (125 and 500 μM, 4 μCi/ml, 20 s, 37°C) was measured in absence or presence of 100 nM nitrobenzylthioinosine (NBMPR, hENT1 inhibitor). hENT2 mRNA was quantified by real time RT-PCR, and protein abundance was determined by Western blot. SLC29A2 (for hENT2) promoter activity was measured following transfection (electroporation, 320 V, 30 ms) with pGL3 basic plasmid (firefly/renilla luciferase reporter gene) carrying -1477 bp and -587 bp of the promoter sequence. Results: Hypoxia reduced hENT2 mRNA expression (~55%), and promoter activity (~50%), but did not alter hENT2 protein abundance. Adenosine uptake via hENT2 was increased (2-fold) in hypoxia. Conclusions: Adenosine uptake via hENT2 may be modulated by post-translational mechanisms in hypoxia in HUVEC. Supported by FONDECYT 1030781/1030607/7050030. A Torres holds a School of Medicine research fellowship, and M Farías holds a CONICYT-PhD fellowship.
- ItemInsulin requires normal expression and signaling of insulin receptor A to reverse gestational diabetes-reduced adenosine transport in human umbilical vein endothelium(2015) Westermeier, Francisco; Salomón, Carlos; Farías Jofré, Marcelo Enrique; Arroyo, Pablo; Fuenzalida Saavedra, Bárbara; Sáez, Tamara; Salsoso Rodríguez, M. Rocío; Sanhueza, Carlos; Guzmán-Gutierrez, Enrique; Pardo, Fabián; Leiva Mendoza, Andrea Alejandra; Sobrevía Luarte, Luis Alberto
- ItemPre-gestational overweight in guinea pig sows induces fetal vascular dysfunction and increased rate of large and small fetuses(2016) Krause Leyton, Bernardo; Herrera, E. A.; Díaz López, F. A.; Farías Jofré, Marcelo Enrique; Uauy, Ricardo; Casanello Toledo, Paola Cecilia
- ItemPre-pregnancy maternal obesity associates with endoplasmic reticulum stress in human umbilical vein endothelium(2018) Villalobos Labra, Roberto Esteban; Saez, P.; Subiabre Morales, Mario Enrique; Silva, L.; Toledo, F.; Westermeier, F.; Pardo, F.; Farías Jofré, Marcelo Enrique; Sobrevía Luarte, Luis Alberto
- ItemPreeclampsia associates with RECK-dependent decrease in human trophoblasts migration and invasion(2017) Gutiérrez, Jaime; Aedo, Alejandro; Mora, Jacob; Maldonado, Jorge; Salsoso Rodríguez, M. Rocío; Toledo, Fernando; Farías Jofré, Marcelo Enrique; Pardo, Fabián; Leiva Mendoza, Andrea Alejandra; Sobrevía Luarte, Luis Alberto
- ItemReduced Insulin Response in Human Umbilical Vein Endothelial Cells from Pregnancies with Maternal Obesity Is Associated with Activation of PERK Branch of Endoplasmic Reticulum Stress Pathway(2013) Farías Jofré, Marcelo Enrique; Westermeier Lafuente, Francisco David; Kusanovic Pivcevic, Juan Pedro; Poblete Lizana, José Andrés; Mardones S., Francisco; Sobrevía Luarte, Luis Alberto
- ItemTGF-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 AlbertoAims: 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.
- ItemThe transcriptional repression of equilibrative nucleoside transporter 1 by D-glucose rfsponses to transcription factor Sp1 and ZBP-89 in human foetal endothelium(2008) Puebla Aracena, Carlos Alberto; Farías Jofré, Marcelo Enrique; Vega Pizarro, José Luis Eduardo; Pastor-Anglada, M.; Casanello Toledo, Paola Cecilia; Sobrevía Luarte, Luis AlbertoObjectives: Adenosine is a vasodilator in most vascular beds, an effect depending on its extracellular concentration. Uptake of this nucleoside in human umbilical vein endothelial cells (HUVEC) is mainly mediated by human equilibrative nucleoside transporters 1 (hENT1). hENT1 expression and transport activity are reduced in HUVEC exposed to high D-glucose. Since specific mechanisms for these effects of D-glucose are unknown, we examined the role of Sp1 and BP-89 transcription factors on SLC29A1 (for hENT1) promoter activity in response to D-glucose. Methods: HUVEC from normal pregnancies were isolated and exposed (24 h) to 5 mM (normal) or 25 mM (high) D-glucose. Sp1 protein levels were evaluated by western blot in nuclear fractions. Reporter activity of plasmid constructs containing a wild type promoter region of SLC29A1 (-1114 bp to ATG, pGL3-hENT1-1114), or mutations (by PCR) for Sp1 (-815/-801 bp, pGL3-hENT1-1114mutSp1) or ZBP-89 (-992/-969 bp, pGL3-hENT1-1114mutZBP) or both (-1114 bp, pGL3-hENT1-1114mutSp1/ZBP) binding sites were assayed in cells over-expressing Sp1 (using pCGN-Sp1 vector). Results: Nuclear Sp1 abundance was increased, but pGL3-hENT1-1114 transcriptional activity was reduced by high D-glucose. Sp1 over-expression reduced pGL3-hENT1-1114 transcriptional activity in normal or high D-glucose. The effect of high D-glucose or Sp1 over-expression was absent in pGL3-hENT1-1114mutSp1, pGL3-hENT1-1114mutZBP and pGL3-hENT1-1114mutSp1/ZBP cells. Conclusion: A repression of the SLC29A1 promoter activity by Sp1 and ZBP-89 could explain the reduced hENT1 expression and activity exhibited by HUVEC in high extracellular D-glucose. FONDECYT 1070865/1080534/7070249 (Chile), AECI A/5484/06 (Spain). C Puebla andJL Vega hold CONICYT fellowships. M Farías holds CONICYT and PUC-School of Medicine PhD fellowships.
- ItemTranscriptional repression of equilibrative nucleoside transporter 1 by D-glucose involves nitric oxide and Sp1 in human foetal endothelium(2007) Puebla Aracena, Carlos Alberto; Farías Jofré, Marcelo Enrique; Vecchiola Cárdenas, Andrea Paola; Casanello Toledo, Paola Cecilia; Sobrevía Luarte, Luis AlbertoAdenosine uptake via the human equilibrative nucleoside transporters 1 (hENT1) is reduced in human umbilical vein endothelial cells (HUVEC) exposed to high D-glucose. Since specific mechanisms by which D-glucose reduces hENT1 expression and activity in these cells are unknown, we examined the role of nitric oxide (NO) and Sp1 as modulators of SLC29A1 (for hENT1) promoter activity in response to D-glucose. HUVEC from normal pregnancies were isolated and exposed (24 h) to 5 mM (normal) or 25 mM (high) D-glucose in absence or presence of NG-nitro-L-arginine methyl ester (LNAME, 100 mM). Sp1 protein levels were evaluated by western blot in nuclear fractions. Reporter activity of plasmid constructs containing a promoter region of SLC29A1 (-1114 bp to ATG, pGL3-hENT1-1114), Sp1 overexpression experiments (co-transfection with the expression vector pCGN-Sp1) and Sp1 binding to this region (chromatin immunoprecipitation) was assayed. pGL3-hENT1-1114 transcriptional activity was reduced by high Dglucose and in cells overexpressing Sp1. Nuclear Sp1 abundance and specific binding to its consensus sequence in SLC29A1 promoter (-815 to -801 bp) were higher in high D-glucose (1.9- and 1.7-fold, respectively) compared with normal D-glucose. All these effects were blocked by L-NAME. Thus, a NO-dependent Sp1 repression on SLC29A1 promoter could explain the reduced hENT1 expression and activity exhibited by HUVEC in high extracellular D-glucose. FONDECYT 1070865, VRAID2007-PUC (Chile). AECI A/5484/06 (Spain). C. Puebla holds a PUC-VRAID PhD fellowship. M. Farías hold CONICYT and PUC-School of Medicine PhD fellowships