Browsing by Author "Hernández, Ignacio"
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Item Differential expression of circulating miRNAs as a novel tool to assess BAG3-associated familial dilated cardiomyopathy.(Bioscience Reports, 2019) Zaragoza Sánchez, Carlos; Saura, Marta; Hernández, Ignacio; Ramírez Carracedo, Rafael; García García, Francisco; Zamorano, José L.; Mangas, Alipio; Toro, RocíoA new familial dilated cardiomyopathy (FDCM) was found related to mutations in BAG3 gene. MicroRNAs (miRNAs) represent new targets of FDCM, although no studies have assessed clinical association between Bcl2-associated athanogene 3 (BAG3)-related DCM and miRNAs. Here, we studied whether a clinical association between BAG3-related FDCM and circulating miRNAs may have diagnostic and prognostic value in a small cohort of familial related individuals carrying a BAG3 mutation (BAG3+) and/or diagnosed of dilated cardiomyopathy (DCM) (DCM+). The analysis of 1759 circulating miRNAs showed significant differences between BAG3+ and BAG3- individuals for miRNAs mir-3191-3p, 6769b-3p, 1249-ep, 154-5p, 6855-5p, and 182-5p, while comparisons between BAG3+/DCM+ versus BAG3+/DCM- were restricted to miRNAs mir-154-5p, 6885-5p, and 182-5p, showing significant correlation with systolic and diastolic blood pressure, A wave, left atrium length, and left atrium area. Additionally, when stratified by gender and age, miRNAs were statistically correlated with critical parameters, including left ventricle ejection fraction (LVEF) and ventricular diameter, in women and young men. Likewise, 56% of BAG3+/DCM+, significantly co-expressed mir-154-5p and mir-182-5p, and a slight 4% did not express such combination, suggesting that co-expression of mir-154-5p and mir-182-5p may potentially show diagnostic value. Further studies will require long-term follow-up, and validation in larger populations.Item Ivabradine Induces Cardiac Protection against Myocardial Infarction by Preventing Cyclophilin-A Secretion in Pigs under Coronary Ischemia/Reperfusion.(International Journal of Molecular Sciences, 2021) Hernández, Ignacio; Tesoro Santos, Laura; Ramírez Carracedo, Rafael; Díez Mata, Javier; Sánchez, Sandra; Saura, Marta; Zamorano, José Luis; Zaragoza Sánchez, Carlos; Botana, LauraIn response to cardiac ischemia/reperfusion, proteolysis mediated by extracellular matrix metalloproteinase inducer (EMMPRIN) and its secreted ligand cyclophilin-A (CyPA) significantly contributes to cardiac injury and necrosis. Here, we aimed to investigate if, in addition to the effect on the funny current (I(f)), Ivabradine may also play a role against cardiac necrosis by reducing EMMPRIN/CyPA-mediated cardiac inflammation. In a porcine model of cardiac ischemia/reperfusion (IR), we found that administration of 0.3 mg/kg Ivabradine significantly improved cardiac function and reduced cardiac necrosis by day 7 after IR, detecting a significant increase in cardiac CyPA in the necrotic compared to the risk areas, which was inversely correlated with the levels of circulating CyPA detected in plasma samples from the same subjects. In testing whether Ivabradine may regulate the levels of CyPA, no changes in tissue CyPA were found in healthy pigs treated with 0.3 mg/kg Ivabradine, but interestingly, when analyzing the complex EMMPRIN/CyPA, rather high glycosylated EMMPRIN, which is required for EMMPRIN-mediated matrix metalloproteinase (MMP) activation and increased CyPA bonding to low-glycosylated forms of EMMPRIN were detected by day 7 after IR in pigs treated with Ivabradine. To study the mechanism by which Ivabradine may prevent secretion of CyPA, we first found that Ivabradine was time-dependent in inhibiting co-localization of CyPA with the granule exocytosis marker vesicle-associated membrane protein 1 (VAMP1). However, Ivabradine had no effect on mRNA expression nor in the proteasome and lysosome degradation of CyPA. In conclusion, our results point toward CyPA, its ligand EMMPRIN, and the complex CyPA/EMMPRIN as important targets of Ivabradine in cardiac protection against IR.Item Ivabradine-Stimulated Microvesicle Release Induces Cardiac Protection against Acute Myocardial Infarction.(International Journal of Molecular Sciences, 2020) Ramírez Carracedo, Rafael; Tesoro Santos, Laura; Hernández, Ignacio; Díez Mata, Javier; Botana, Laura; Saura, Marta; Sanmartin, Marcelo; Zamorano, José Luis; Zaragoza Sánchez, CarlosIvabradine can reduce heart rate through inhibition of the current I(f ) by still unexplored mechanisms. In a porcine model of ischemia reperfusion (IR), we found that treatment with 0.3 mg/kg Ivabradine increased plasma release of microvesicles (MVs) over Placebo, as detected by flow cytometry of plasma isolated from pigs 7 days after IR, in which a tenfold increase of Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) containing (both high and low-glycosylated) MVs, was detected in response to Ivabradine. The source of MVs was investigated, finding a 37% decrease of CD31+ endothelial cell derived MVs, while CD41+ platelet MVs remained unchanged. By contrast, Ivabradine induced the release of HCN4+ (mostly cardiac) MVs. While no differences respect to EMMPRIN as a cargo component were found in endothelial and platelet derived MVs, Ivabradine induced a significant release of EMMPRIN+/HCN4+ MVs by day 7 after IR. To test the role of EMMPRIN+ cardiacMVs (EMCMV), H9c2 cellmonolayers were incubated for 24 h with 107 EMCMVs, reducing apoptosis, and increasing 2 times cell proliferation and 1.5 times cell migration. The in vivo contribution of Ivabradine-induced plasma MVs was also tested, in which 108 MVs isolated from the plasma of pigs treated with Ivabradine or Placebo 7 days after IR, were injected in pigs under IR, finding a significant cardiac protection by increasing left ventricle ejection fraction and a significant reduction of the necrotic area. In conclusion ivabradine induces cardiac protection by increasing at least the release of EMMPRIN containing cardiac microvesicles.Item NIL10: A New IL10-Receptor Binding Nanoparticle That Induces Cardiac Protection in Mice and Pigs Subjected to Acute Myocardial Infarction through STAT3/NF-kB Activation.(Pharmaceutics, 2022) Tesoro Santos, Laura; Hernández, Ignacio; Ramírez Carracedo, Rafael; Díez-Mata, Javier; Alcharani, Nunzio; Jiménez-Guirado, Beatriz; Ovejero-Paredes, Karina; Filice, Marco; Zamorano, Jose Luis; Saura, Marta; Zaragoza Sánchez, Carlos; Botana, LauraBackground: Early response after acute myocardial infarction (AMI) prevents extensive cardiac necrosis, in which inflammation resolution, including expression of anti-inflammatory interleukin-10 (IL-10), may play a key role. (2) Methods: We synthesized NIL10, a micelle-based nanoparticle, to target IL-10 receptor in mice and pigs subjected to AMI. (3) Results: Administration of NIL10 induced cardiac protection of wild-type and IL-10 knockout mice and pigs subjected to AMI. Cardiac protection was not induced in IL-10-receptor null mice, as shown by a significant recovery of cardiac function, in which inflammatory foci and fibrosis were strongly reduced, together with the finding that resolving M2-like macrophage populations were increased after day 3 of reperfusion. In addition, anti-inflammatory cytokines, including IL-4, IL-7, IL-10, IL-13, IL-16, and IL-27 were also elevated. Mechanistically, NIL10 induced activation of the IL-10 receptor/STAT-3 signaling pathway, and STAT3-dependent inhibition of nuclear translocation of pro-inflammatory NF-kB transcription factor. (4) Conclusions: Taken together, we propose using NIL10 as a novel therapeutic tool against AMI-induced cardiac damageItem Niveles de quinasa ligada a la integrina endotelial y calcificación valvular: estudio molecular de la calcificación en la estenosis valvular aórtica.(Cirugía Cardiovascular, 2022) Castro-Pintoa, Mercedes; López-Menéndez, José; Sánchez-Esteban, Sandra; Hernández, Ignacio; Reventún, Paula; Rodríguez-Roda, Jorge; Zaragoza Sánchez, Carlos; Saura, MartaIntroducción y objetivos La quinasa ligada a integrina (ILK) endotelial controla la producción del óxido nítrico, modulando señales de transducción al interior celular. La disminución de ILK endotelial se ha relacionado con la calcificación valvular. Analizamos la asociación de la ILK con la calcificación de la válvula aórtica. Métodos Estudio observacional, unicéntrico y prospectivo. Recogimos tejido valvular aórtico de pacientes intervenidos de estenosis aórtica, y controles de válvulas no calcificadas. Cuantificamos los niveles de ILK endotelial empleando técnicas de análisis molecular, así como marcadores de crecimiento óseo (BMP2 y RUNX2). Para el control de factores de confusión empleamos regresión logística múltiple por pasos, introduciendo parámetros relacionados con la aterosclerosis y calcificación valvular (edad, DM, HTA, insuficiencia renal). Resultados Obtuvimos muestras de 75 pacientes y 28 controles. El 69,9% eran varones, la edad media fue de 67 años (DE 9,59 años). Los niveles de ILK fueron significativamente más bajos en válvulas patológicas que en controles (0,96 vs. 0,76; p < 0,001). Existió correlación lineal negativa significativa entre la ILK y la calcificación valvular (coeficiente de Pearson = −0,35; p = 0,002). En el modelo multivariante, ILK presentó una asociación significativa con la calcificación valvular (OR 0,068; p < 0,001), así como la DM (OR 11,80; p = 0,03). Observamos un aumento de marcadores de crecimiento óseo según avanzaba la calcificación, con correlación inversa significativa de ILK y BMP2 (R = −0,417; p < 0,001) y RUNX2 (R = −0,587; p < 0,001). Conclusiones La disminución de la expresión endotelial de ILK puede influir en la calcificación de la válvula aórtica, marcando el inicio del proceso de transdiferenciación del endotelio valvular a tejido con capacidad osteogénica.Item Non-Invasive Detection of Extracellular Matrix Metalloproteinase Inducer EMMPRIN, a New Therapeutic Target against Atherosclerosis, Inhibited by Endothelial Nitric Oxide.(International Journal of Molecular Sciences, 2018) Ramírez Carracedo, Rafael; Tesoro Santos, Laura; Hernández, Ignacio; Díez Mata, Javier; Filice, Marco; Toro, Rocío; Rodríguez Piñero, Manuel; Zamorano, José Luis; Saura, Marta; Zaragoza Sánchez, CarlosLack of endothelial nitric oxide causes endothelial dysfunction and circulating monocyte infiltration, contributing to systemic atheroma plaque formation in arterial territories. Among the different inflammatory products, macrophage-derived foam cells and smooth muscle cells synthesize matrix metalloproteinases (MMPs), playing a pivotal role in early plaque formation and enlargement. We found increased levels of MMP-9 and MMP-13 in human endarterectomies with advanced atherosclerosis, together with significant amounts of extracellular matrix (ECM) metalloproteinase inducer EMMPRIN. To test whether the absence of NO may aggravate atherosclerosis through EMMPRIN activation, double NOS3/apoE knockout (KO) mice expressed high levels of EMMPRIN in carotid plaques, suggesting that targeting extracellular matrix degradation may represent a new mechanism by which endothelial NO prevents atherosclerosis. Based on our previous experience, by using gadolinium-enriched paramagnetic fluorescence micellar nanoparticles conjugated with AP9 (NAP9), an EMMPRIN-specific binding peptide, magnetic resonance sequences allowed non-invasive visualization of carotid EMMPRIN in NOS3/apoE over apoE control mice, in which atheroma plaques were significantly reduced. Taken together, these results point to EMMPRIN as a new therapeutic target of NO-mediated protection against atherosclerosis, and NAP9 as a non-invasive molecular tool to target atherosclerosis.Item Targeting TLR4 with ApTOLL Improves Heart Function in Response to Coronary Ischemia Reperfusion in Pigs Undergoing Acute Myocardial Infarction.(Biomolecules, 2020) Ramírez Carracedo, Rafael; Tesoro Santos, Laura; Hernández, Ignacio; Díez Mata, Javier; Piñeiro, David; Hernández Jiménez, Macarena; Zamorano, José Luis; Zaragoza Sánchez, CarlosToll-like receptor 4 (TLR4) contributes to the pathogenesis of coronary ischemia/reperfusion (IR). To test whether the new TLR4 antagonist, ApTOLL, may prevent coronary IR damage, we administered 0.078 mg/kg ApTOLL or Placebo in pigs subjected to IR, analyzing the levels of cardiac troponins, matrix metalloproteinases, pro-, and anti-inflammatory cytokines, heart function, and tissue integrity over a period of 7 days after IR. Our results show that ApTOLL reduced cardiac troponin-1 24 h after administration, improving heart function, as detected by a significant recovery of the left ventricle ejection fraction (LVEF) and the shortening fraction (FS) cardiac parameters. The extension of necrotic and fibrotic areas was also reduced, as detected by Evans blue/2,3,5-triphenyltetrazolium chloride (TTC) staining, Hematoxylin/Eosine, and Masson Trichrome staining of heart sections, together with a significant reduction in the expression of the extracellular matrix-degrading, matrix metalloproteinase 9. Finally, the expression of the following cytokines, CCL1, CCL2, MIP1-A-B, CCL5, CD40L, C5/C5A, CXCL1, CXCL10, CXCL11, CXCL12, G-CSF, GM-CSF, ICAM-1, INF-g, IL1-a, ILI-b, IL-1Ra, IL2, IL4, IL5, IL6, IL8, IL10, IL12, IL13, IL16, IL17-A, IL17- E, IL18, IL21, IL27, IL32, MIF, SERPIN-E1, TNF-a, and TREM-1, were also assayed, detecting a pronounced decrease of pro-inflammatory cytokines after 7 days of treatment with ApTOLL. Altogether, our results show that ApTOLL is a promising new tool for the treatment of acute myocardial infarction (AMI).