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HIF1-alpha expression defines metabolic compartments in the developing heart, promoting glycolytic program in the compact myocardium and mitochondrial enrichment in the trabeculae. Nonetheless, its role in cardiogenesis is debated. To assess the importance of HIF1-alpha during heart development and the influence of glycolysis in ventricular chamber formation, herein we generated conditional knockout models of Hif1a in Nkx2.5 cardiac progenitors and cardiomyocytes. Deletion of Hif1a impairs embryonic glycolysis without influencing cardiomyocyte proliferation and results in increased mitochondrial number and transient activation of amino acid catabolism together with HIF2α and ATF4 upregulation by E12.5. Hif1a mutants display normal fatty acid oxidation program and do not show cardiac dysfunction in the adulthood. Our results demonstrate that cardiac HIF1 signaling and glycolysis are dispensable for mouse heart development and reveal the metabolic flexibility of the embryonic myocardium to consume amino acids, raising the potential use of alternative metabolic substrates as therapeutic interventions during ischemic events. ; This project has been supported by Fundación Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), Spain and by grants to S.M.-P. from the European Research Council, European Union, FP7-PEOPLE-2010-RG_276891; Fundación TV3 La Marató, Spain, 201507.30.31; Comunidad de Madrid (CAM); Spain and European Union (EU), B2017/BMD-3875; Instituto de Salud Carlos III, Spain, PI17/01817; Universidad Francisco de Vitoria (UFV), Spain and LeDucq Foundation, France, 17CVD04. I.M.-M. was supported by La Caixa-CNIC and Fundacion Alfonso Martín Escudero fellowships, Spain. T.A.-G. was supported by a predoctoral award granted by CAM/EU and UFV, Spain, PEJD-2018-PRE/SAL-9529 and SM-P by a Contrato de Investigadores Miguel Servet (CPII16/00050) and UFV, Spain. ; Sí

BACKGROUND In response to pressure overload, the heart develops ventricular hypertrophy that progressively decompensates and leads to heart failure. This pathological hypertrophy is mediated, among others, by the phosphatase calcineurin and is characterized by metabolic changes that impair energy production by mitochondria. OBJECTIVES The authors aimed to determine the role of the calcineurin splicing variant CnA beta 1 in the context of cardiac hypertrophy and its mechanism of action. METHODS Transgenic mice overexpressing CnAb1 specifically in cardiomyocytes and mice lacking the unique C-terminal domain in CnA beta 1 (CnA beta 1(Delta i12) mice) were used. Pressure overload hypertrophy was induced by transaortic constriction. Cardiac function was measured by echocardiography. Mice were characterized using various molecular analyses. RESULTS In contrast to other calcineurin isoforms, the authors show here that cardiac-specific overexpression of CnA beta 1 in transgenic mice reduces cardiac hypertrophy and improves cardiac function. This effect is mediated by activation of serine and one-carbon metabolism, and the production of antioxidant mediators that prevent mitochondrial protein oxidation and preserve ATP production. The induction of enzymes involved in this metabolic pathway by CnAb1 is dependent on mTOR activity. Inhibition of serine and one-carbon metabolism blocks the beneficial effects of CnA beta 1. CnA beta 1(Delta i12) mice show increased cardiac hypertrophy and declined contractility. CONCLUSIONS The metabolic reprogramming induced by CnAb1 redefines the role of calcineurin in the heart and shows for the first time that activation of the serine and one-carbon pathway has beneficial effects on cardiac hypertrophy and function, paving the way for new therapeutic approaches. (J Am Coll Cardiol 2018; 71: 654-67) (C) 2018 The Authors. Published by Elsevier on behalf of the American College of Cardiology Foundation. This is an open access article under the CC BY-NC-ND license (http://creativecommons. org/licenses/by-nc-nd/4.0/). ; This work was supported by grants from the European Union (CardioNeT-ITN-289600 and CardioNext-608027 to Dr. Lara-Pezzi; Meet-ITN-317433 to Dr. Enriquez; UE0/MCA1108 to Dr. Acin-Perez), from the Spanish Ministry of Economy and Competitiveness (SAF2015-65722-R and SAF2012-31451 to Dr. Lara-Pezzi; SAF2015-71521-REDC, BFU2013-50448, and SAF2012-32776 to Dr. Enriquez; RyC-2011-07826 to Dr. Acin-Perez; BIO2012-37926 and BIO2015-67580-P to Dr. Vazquez), from the Spanish Carlos III Institute of Health (CPII14/00027 to Dr. Lara-Pezzi; RD12/0042/066 to Drs. Garcia-Pavia and Lara-Pezzi), from the Regional Government of Madrid (2010-BMD-2321 ``Fibroteam´´ to Dr. Lara-Pezzi; 2011-BMD-2402 ``Mitolab´´ to Dr. Enriquez) and the FIS-ISCIII (PRB2-IPT13/0001 and RD12/0042/0056-RIC-RETICS to Dr. Vazquez). This work was also supported by the Plan Estatal de IthornDthornI 2013-2016-European Regional Development Fund (FEDER) ``A way of making Europe,´´ Spain. The CNIC is supported by the Spanish Ministry of Economy and Competitiveness and by the Pro-CNIC Foundation and is a Severo Ochoa Center of Excellence (MINECO award SEV-2015-0505). Drs. Vazquez and Garcia-Pavia have served as consultants for VL39. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose. Drs. Padron-Barthe, Villalba-Orero, and Gomez-Salinero contributed equally to this work and are joint first authors. Robyn Shaw, MD, PhD, served as Guest Editor for this paper. ; Sí

MEC, Spain [BFU2011-23896]; European Union [278976]; Postdok BIOGLOBE, Czech Republic [CZ.1.07/2.3.00/30.0032]; European Social Fund; state budget of the Czech Republic; Ministerio de Educacion, Cultura y Deporte, Spain; EU [238511]This research was supported by MEC, Spain, grant BFU2011-23896 (JF), European Union FP7 ANTIGONE project number 278976 (JF) and Postdok BIOGLOBE, Czech Republic CZ.1.07/2.3.00/30.0032 co-financed by the European Social Fund and the state budget of the Czech Republic (JS). N. Ayllon was funded by Ministerio de Educacion, Cultura y Deporte, Spain. S. Weisheit was supported by the POSTICK ITN (Post-graduate training network for capacity building to control ticks and tick-borne diseases) within the FP7-PEOPLE-ITN programme (EU Grant No. 238511). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.06Public library scienceSan francisco ; Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10-15% and 65-71% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in which cells convert from an intracellular reticulated, replicative form to the nondividing infectious dense-core form. The analysis of A. phagocytophilum differentially represented proteins identified stress response (GroEL, HSP70) and surface (MSP4) proteins that were over-represented in high percentage infected tick cells and salivary glands when compared to low percentage infected cells and guts, respectively. The results demonstrated that MSP4, GroEL and HSP70 interact and bind to tick cells, thus playing a role in rickettsia-tick interactions. The most important finding of these studies is the increase in the level of certain bacterial stress response and surface proteins in A. phagocytophilum-infected tick cells and salivary glands with functional implication in tick-pathogen interactions. These results gave a new dimension to the role of these stress response and surface proteins during A. phagocytophilum infection in ticks. Characterization of Anaplasma proteome contributes information on host-pathogen interactions and provides targets for development of novel control strategies for pathogen infection and transmission.

MEC, Spain [BFU2011-23896]; European Union [278976]; Postdok BIOGLOBE, Czech Republic [CZ.1.07/2.3.00/30.0032]; European Social Fund; state budget of the Czech Republic; Ministerio de Educacion, Cultura y Deporte, Spain; EU [238511]This research was supported by MEC, Spain, grant BFU2011-23896 (JF), European Union FP7 ANTIGONE project number 278976 (JF) and Postdok BIOGLOBE, Czech Republic CZ.1.07/2.3.00/30.0032 co-financed by the European Social Fund and the state budget of the Czech Republic (JS). N. Ayllon was funded by Ministerio de Educacion, Cultura y Deporte, Spain. S. Weisheit was supported by the POSTICK ITN (Post-graduate training network for capacity building to control ticks and tick-borne diseases) within the FP7-PEOPLE-ITN programme (EU Grant No. 238511). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.06Public library scienceSan francisco ; Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10-15% and 65-71% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in ...

MEC, Spain [BFU2011-23896]; European Union [278976]; Postdok BIOGLOBE, Czech Republic [CZ.1.07/2.3.00/30.0032]; European Social Fund; state budget of the Czech Republic; Ministerio de Educacion, Cultura y Deporte, Spain; EU [238511]This research was supported by MEC, Spain, grant BFU2011-23896 (JF), European Union FP7 ANTIGONE project number 278976 (JF) and Postdok BIOGLOBE, Czech Republic CZ.1.07/2.3.00/30.0032 co-financed by the European Social Fund and the state budget of the Czech Republic (JS). N. Ayllon was funded by Ministerio de Educacion, Cultura y Deporte, Spain. S. Weisheit was supported by the POSTICK ITN (Post-graduate training network for capacity building to control ticks and tick-borne diseases) within the FP7-PEOPLE-ITN programme (EU Grant No. 238511). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.06Public library scienceSan francisco ; Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10-15% and 65-71% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in which cells convert from an intracellular reticulated, replicative form to the nondividing infectious dense-core form. The analysis of A. phagocytophilum differentially represented proteins identified stress response (GroEL, HSP70) and surface (MSP4) proteins that were over-represented in high percentage infected tick cells and salivary glands when compared to low percentage infected cells and guts, respectively. The results demonstrated that MSP4, GroEL and HSP70 interact and bind to tick cells, thus playing a role in rickettsia-tick interactions. The most important finding of these studies is the increase in the level of certain bacterial stress response and surface proteins in A. phagocytophilum-infected tick cells and salivary glands with functional implication in tick-pathogen interactions. These results gave a new dimension to the role of these stress response and surface proteins during A. phagocytophilum infection in ticks. Characterization of Anaplasma proteome contributes information on host-pathogen interactions and provides targets for development of novel control strategies for pathogen infection and transmission.

MEC, Spain [BFU2011-23896]; European Union [278976]; Postdok BIOGLOBE, Czech Republic [CZ.1.07/2.3.00/30.0032]; European Social Fund; state budget of the Czech Republic; Ministerio de Educacion, Cultura y Deporte, Spain; EU [238511]This research was supported by MEC, Spain, grant BFU2011-23896 (JF), European Union FP7 ANTIGONE project number 278976 (JF) and Postdok BIOGLOBE, Czech Republic CZ.1.07/2.3.00/30.0032 co-financed by the European Social Fund and the state budget of the Czech Republic (JS). N. Ayllon was funded by Ministerio de Educacion, Cultura y Deporte, Spain. S. Weisheit was supported by the POSTICK ITN (Post-graduate training network for capacity building to control ticks and tick-borne diseases) within the FP7-PEOPLE-ITN programme (EU Grant No. 238511). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.06Public library scienceSan francisco ; Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10-15% and 65-71% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in which cells convert from an intracellular reticulated, replicative form to the nondividing infectious dense-core form. The analysis of A. phagocytophilum differentially represented proteins identified stress response (GroEL, HSP70) and surface (MSP4) proteins that were over-represented in high percentage infected tick cells and salivary glands when compared to low percentage infected cells and guts, respectively. The results demonstrated that MSP4, GroEL and HSP70 interact and bind to tick cells, thus playing a role in rickettsia-tick interactions. The most important finding of these studies is the increase in the level of certain bacterial stress response and surface proteins in A. phagocytophilum-infected tick cells and salivary glands with functional implication in tick-pathogen interactions. These results gave a new dimension to the role of these stress response and surface proteins during A. phagocytophilum infection in ticks. Characterization of Anaplasma proteome contributes information on host-pathogen interactions and provides targets for development of novel control strategies for pathogen infection and transmission.

Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10-15\% and 65-71\% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in which cells convert from an intracellular reticulated, replicative form to the nondividing infectious dense-core form. The analysis of A. phagocytophilum differentially represented proteins identified stress response (GroEL, HSP70) and surface (MSP4) proteins that were over-represented in high percentage infected tick cells and salivary glands when compared to low percentage infected cells and guts, respectively. The results demonstrated that MSP4, GroEL and HSP70 interact and bind to tick cells, thus playing a role in rickettsia-tick interactions. The most important finding of these studies is the increase in the level of certain bacterial stress response and surface proteins in A. phagocytophilum-infected tick cells and salivary glands with functional implication in tick-pathogen interactions. These results gave a new dimension to the role of these stress response and surface proteins during A. phagocytophilum infection in ticks. Characterization of Anaplasma proteome contributes information on host-pathogen interactions and provides targets for development of novel control strategies for pathogen infection and transmission. ; This research was supported by MEC, Spain, grant BFU2011-23896 (JF), European Union FP7 ANTIGONE project number 278976 (JF) and Postdok BIOGLOBE, Czech Republic CZ.1.07/2.3.00/30.0032 co-financed by the European Social Fund and the state budget of the Czech Republic (JS). N. Ayllon was funded by Ministerio de Educacion, Cultura y Deporte, Spain. S. Weisheit was supported by the POSTICK ITN (Post-graduate training network for capacity building to control ticks and tick-borne diseases) within the FP7-PEOPLE-ITN programme (EU Grant No. 238511). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. ; Sí

MEC, Spain [BFU2011-23896]; European Union [278976]; Postdok BIOGLOBE, Czech Republic [CZ.1.07/2.3.00/30.0032]; European Social Fund; state budget of the Czech Republic; Ministerio de Educacion, Cultura y Deporte, Spain; EU [238511]This research was supported by MEC, Spain, grant BFU2011-23896 (JF), European Union FP7 ANTIGONE project number 278976 (JF) and Postdok BIOGLOBE, Czech Republic CZ.1.07/2.3.00/30.0032 co-financed by the European Social Fund and the state budget of the Czech Republic (JS). N. Ayllon was funded by Ministerio de Educacion, Cultura y Deporte, Spain. S. Weisheit was supported by the POSTICK ITN (Post-graduate training network for capacity building to control ticks and tick-borne diseases) within the FP7-PEOPLE-ITN programme (EU Grant No. 238511). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.06Public library scienceSan francisco ; Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10-15% and 65-71% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in ...

Titin, as the main protein responsible for the passive stiffness of the sarcomere, plays a key role in diastolic function and is a determinant factor in the etiology of heart disease. Titin stiffness depends on unfolding and folding transitions of immunoglobulin-like (Ig) domains of the I-band, and recent studies have shown that oxidative modifications of cryptic cysteines belonging to these Ig domains modulate their mechanical properties in vitro. However, the relevance of this mode of titin mechanical modulation in vivo remains largely unknown. Here, we describe the high evolutionary conservation of titin mechanical cysteines and show that they are remarkably oxidized in murine cardiac tissue. Mass spectrometry analyses indicate a similar landscape of basal oxidation in murine and human myocardium. Monte Carlo simulations illustrate how disulfides and S-thiolations on these cysteines increase the dynamics of the protein at physiological forces, while enabling load- and isoform-dependent regulation of titin stiffness. Our results demonstrate the role of conserved cysteines in the modulation of titin mechanical properties in vivo and point to potential redox-based pathomechanisms in heart disease. ; This work was supported by the Ministerio de Ciencia e Innovación grants BIO2014-54768-P, BIO2017-83640-P, RYC-2014-16604 to JAC and PGC2018-097019-B-I00 to JV, the Regional Government of Madrid grants S2018/NMT-4443 and PEJ16/MED/TL-1593 to JAC and the Instituto de Salud Carlos III (Fondo de Investigación Sanitaria grant PRB3 (PT17/0019/0003- ISCIII-SGEFI /ERDF, ProteoRed), and "la Caixa" Banking Foundation (project code HR17-00247) to JV. We acknowledge funding from the European Research Area Network on Cardiovascular Disease through grant MINOTAUR to SS (The Austrian Science Fund – FWF, I3301) and JAC (ISCIII-AC16/00045). The CNIC is supported by ISCIII, the Ministerio de Ciencia e Innovación and the Pro CNIC Foundation, and was a Severo Ochoa Center of Excellence (SEV-2015-0505). IMM was the recipient of a CNIC-ACCIONA Masters Fellowship and holds a fellowship from "La Caixa" Foundation (ID 100010434, fellowship code LCF/BQ/DR20/11790009). CSC is the recipient of an FPI-SO predoctoral fellowship BES-2016-076638. We thank Wolfgang A. Linke and Pablo García-Pavía for critical feedback. We are also thankful for the insights of three anonymous reviewers. ; Sí

BACKGROUND: Arrhythmogenic cardiomyopathy/arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited cardiac disease characterized by fibrofatty replacement of the myocardium, resulting in heart failure and sudden cardiac death. The most aggressive arrhythmogenic cardiomyopathy/ARVC subtype is ARVC type 5 (ARVC5), caused by a p.S358L mutation in TMEM43 (transmembrane protein 43). The function and localization of TMEM43 are unknown, as is the mechanism by which the p.S358L mutation causes the disease. Here, we report the characterization of the first transgenic mouse model of ARVC5. METHODS: We generated transgenic mice overexpressing TMEM43 in either its wild-type or p.S358L mutant (TMEM43-S358L) form in postnatal cardiomyocytes under the control of the α-myosin heavy chain promoter. RESULTS: We found that mice expressing TMEM43-S358L recapitulate the human disease and die at a young age. Mutant TMEM43 causes cardiomyocyte death and severe fibrofatty replacement. We also demonstrate that TMEM43 localizes at the nuclear membrane and interacts with emerin and β-actin. TMEM43-S358L shows partial delocalization to the cytoplasm, reduced interaction with emerin and β-actin, and activation of glycogen synthase kinase-3β (GSK3β). Furthermore, we show that targeting cardiac fibrosis has no beneficial effect, whereas overexpression of the calcineurin splice variant calcineurin Aβ1 results in GSK3β inhibition and improved cardiac function and survival. Similarly, treatment of TMEM43 mutant mice with a GSK3β inhibitor improves cardiac function. Finally, human induced pluripotent stem cells bearing the p.S358L mutation also showed contractile dysfunction that was partially restored after GSK3β inhibition. CONCLUSIONS: Our data provide evidence that TMEM43-S358L leads to sustained cardiomyocyte death and fibrofatty replacement. Overexpression of calcineurin Aβ1 in TMEM43 mutant mice or chemical GSK3β inhibition improves cardiac function and increases mice life span. Our results pave the way toward new therapeutic approaches for ARVC5. ; This work was supported by grants from the European Union (CardioNeTITN-289600 and CardioNext-608027 to Dr Lara-Pezzi), the Spanish Ministry of Economy and Competitiveness (RTI2018-096961-B-I00, SAF2015-65722-R, and SAF2012-31451 to Dr Lara-Pezzi; SAF2015-71863-REDT to Dr Garcia-Pavia), the Spanish Carlos III Institute of Health (PI14/0967 to Dr Garcia-Pavia, CPII14/00027 to Dr Lara-Pezzi; RD012/0042/0066 to Drs Garcia-Pavia and Lara-Pezzi), the Regional Government of Madrid (2010-BMD-2321 "Fibroteam" to Dr Lara-Pezzi), the Isabel Gemio Foundation (Todos somos Raros grant to Dr Garcia-Pavia), and the Spanish Society of Cardiology (2014 Basic Research Grant to Dr Garcia-Pavia). This work was also supported by the Plan Estatal de I+D+I 2013-2016–European Regional Development Fund (FEDER) "A way of making Europe," Spain. The Centro Nacional de Investigaciones Cardiovasculares Carlos III is supported by the Ministerio de Ciencia, Innovación y Universidades (MCNU), and Pro Centro Nacional de Investigaciones Cardiovasculares Carlos III Foundation and is a Severo Ochoa Center of Excellence (SEV-2015-0505). ; Sí