Virginio García-Martínez

Cardiovascular disease displays marked sex-related heterogeneity across the life course, and the rise in cardiometabolic and vascular risk during the menopausal transition has focused attention on estrogen signaling. Healthy female hearts are not simply scaled-down male hearts; population averages differ in cardiac mass, chamber geometry, myocardial deformation, hemodynamics, and repolarization. This narrative review integrates experimental, epidemiological, genetic, and clinical evidence on estrogen action in the cardiovascular system. Estrogens signal through ERalpha and ERbeta, their membrane-associated pools, and GPER, whereas estrogen-related receptors coordinate overlapping metabolic programs without binding estrogen. These pathways regulate endothelial nitric oxide production, redox balance, inflammation, mitochondrial energetics, autophagy, calcium handling, electrophysiology, extracellular-matrix remodeling, and non-coding RNA networks. Their effects depend on cell type, age, receptor abundance, vascular substrate, comorbidities, and genetic or epigenetic variation. This context dependence helps explain why endogenous estrogen exposure cannot be equated with menopausal hormone therapy. Current evidence does not support hormone therapy for primary or secondary cardiovascular prevention; treatment of menopausal symptoms requires individualized assessment of indication, timing, formulation, route, thrombotic risk, and baseline cardiovascular health. Future studies should combine sex-aware human models, longitudinal multi-omics, and validated biomarkers to link estrogen-pathway variation with clinically meaningful cardiovascular outcomes.

Cardiogenesis begins during gastrulation with the bilateral specification of precardiac mesoderm on either side of the embryo. These fields fuse at the embryonic midline to form the linear heart tube, which subsequently undergoes rightward looping and chamber formation. Expansion and septation of the primitive atrial and ventricular chambers establish separate systemic and pulmonary circuits with correctly aligned inflow and outflow tracts. These morphogenetic events depend on the coordinated activation and repression of cell type-specific cardiac gene programmes controlled at transcriptional and post-transcriptional levels by cardiac transcription factors and paracrine/autocrine signals. Non-coding RNAs, particularly microRNAs and long non-coding RNAs, add an essential regulatory layer. MicroRNAs are endogenous ≈22-nt RNAs that modulate gene expression predominantly at the post-transcriptional level. The muscle-enriched miR-1/miR-133 cluster is essential for normal cardiogenesis, whereas dysregulation of miR-1 and miR-133 contributes to heart failure and its associated pathological processes. Here, we provide a state-of-the-art review of the genomic regulation, developmental functions, context-dependent interactions, and translational potential of the miR-1/miR-133 cluster in cardiogenesis and heart failure.

Cardiogenesis is initiated soon during gastrulation as bilateral precardiac mesoderm, progressive and symmetrically determined at both sides of the developing embryo. The precardiac mesoderm subsequently fuse at the embryonic midline constituting an embryonic linear heart tube. As development progress, the embryonic tube heart suffers an invariably rightward looping generating a prospective embryonic ventricular and atrial chambers. As cardiac development progresses, the primitive atrial and ventricular chambers enlarged and distinct left and right compartments emerge by formation of the interatrial and interventricular septa, respectively. The last steps of cardiac morphogenesis are represented by the completion of atrial and ventricular septation, resulting in the configuration of a double circuitry with distinct systemic and pulmonary chambers, each of them with distinct inlets and outlets connections. The heart development is resulted of the activation and repression of several cardiac specific gene programs in different cell types. These cardiac specific gene pathways are modulated at transcriptional and post-transcriptional levels by key cardiac transcription factors and different paracrine and autocrine signals. Over the last decade, a novel layer of complexity has emerged with the discovery of non-coding RNAs, particularly microRNAs and lncRNAs. MicroRNAs are endogenous ≈22-nt non-coding RNAs that participate in the regulation of gene expression at post-transcriptional level on most cellular processes. In this line, miR-1/miR-133 cluster is one of the muscle-specific cluster microRNAs. Proper expression of members of this cluster is essential for cardiogenesis. Furthermore, dysregulation expression of miR-1 and miR-133 expression exerts an important role in heart failure and related pathological processes. Herein, we provide a state-of-the-art review of the contribution of miR-1/miR-133 cluster in cardiogenesis and heart failure development.

The 3-nitropropionic acid (NPA) promotes neurological alterations in the striatum, hippocampus and vicinal motor and pre-motor cortical areas, and in the cerebellum. The neurological alterations induced by systemic NPA administration resemble those found in Huntington’s disease. In previous works, we have shown that intraperitoneal (i.p.) administration of kaempferol can efficiently protect against striatum degeneration and against motor neurological dysfunctions induced by NPA. In this work, we show that i.p. administration of kaempferol also protects against the increase in pro-inflammatory cytokines that potentiate the activation of complement C3 protein (a biomarker of A1-type reactive astrocytes generation) and overproduction of neurotoxic amyloid β (Aβ) peptides in the cerebellum of rats treated with acute i.p. administration of NPA. In NPA-treated rats, large multipolar neurons of cerebellar nuclei and Purkinje neurons of the cerebellar cortex are the cells that are most intensely stained by anti-C3 and by anti-Aβ antibodies. In addition, we found that kaempferol also protects against the NPA-induced increase in phospho-tau 217 and phospho-tau 181 in the cerebellum, and our results pointed out that the NPA-induced phospho-tau 217 colocalizes with Aβ(1-42) more closely than phospho-tau 181, both in dentate nucleus and cerebellar cortex. Also, our results unveil another novel brain-protective action of i.p. kaempferol co-administration: namely, its ability to prevent microhemorrhages induced in the cerebellar nuclei area by acute NPA administration. In conclusion, the results of this work show a potent protection of kaempferol against the NPA-induced increase in degeneration biomarkers in the cerebellum.

The 3-nitropropionic acid (NPA) promotes neurological alterations in the striatum, hippocampus and vicinal motor and pre-motor cortical areas, and in the cerebellum. The neurological alterations induced by systemic NPA administration resemble those found in Huntington’s disease. In previous works, we have shown that intraperitoneal (i.p.) administration of kaempferol can efficiently protect against striatum degeneration and against motor neurological dysfunctions induced by NPA. In this work, we show that i.p. administration of kaempferol also protects against the increase of pro-inflammatory cytokines that potentiate the activation of complement C3 protein (a biomarker of A1-type reactive astrocytes generation) and overproduction of neurotoxic amyloid β (Aβ) peptides in the cerebellum of rats treated with acute i.p. administration of NPA. In NPA-treated rats, large multipolar neurons of cerebellar nuclei and Purkinje neurons of the cerebellar cortex are the cells most intensely stained by anti-C3 and by anti-Aβ antibodies. In addition, we found that kaempferol also protects against the NPA-induced increase of phospho-tau 217 and phospho-tau 181 in the cerebellum, and our results pointed out that the NPA-induced phospho-tau 217 colocalizes with Aβ(1-42) more closely than phospho-tau 181, both in dentate nucleus and cerebellar cortex. Also, our results unveil another novel brain protection action of i.p. kaempferol co-administration, namely, its ability to prevent microhemorrhages induced in the cerebellar nuclei area by acute NPA administration. In conclusion, our results lend strong support to its use in the therapeutic treatment of NPA intoxication and related β-amyloidopathies and tauopathies.

A large diversity of epigenetic factors, such as microRNAs and histones modifications, are known to be capable of regulating gene expression without altering DNA sequence itself. In particular, miR-1 is considered the first essential microRNA in cardiac development. In this study, miR-1 potential role in early cardiac chamber differentiation was analyzed through specific signaling pathways. For this, we performed in chick embryos functional experiments by means of miR-1 microinjections into the posterior cardiac precursors—of both primitive endocardial tubes—committed to sinoatrial region fates. Subsequently, embryos were subjected to whole mount in situ hybridization, immunohistochemistry and RT-qPCR analysis. As a relevant novelty, our results revealed that miR-1 increased Amhc1, Tbx5 and Gata4, while this microRNA diminished Mef2c and Cripto expressions during early differentiation of the cardiac sinoatrial region. Furthermore, we observed in this developmental context that miR-1 upregulated CrabpII and Rarß and downregulated CrabpI, which are three crucial factors in the retinoic acid signaling pathway. Interestingly, we also noticed that miR-1 directly interacted with Hdac4 and Calm1/Calmodulin, as well as with Erk2/Mapk1, which are three key factors actively involved in Mef2c regulation. Our study shows, for the first time, a key role of miR-1 as an epigenetic regulator in the early differentiation of the cardiac sinoatrial region through orchestrating opposite actions between retinoic acid and Mef2c, fundamental to properly assign cardiac cells to their respective heart chambers. A better understanding of those molecular mechanisms modulated by miR-1 will definitely help in fields applied to therapy and cardiac regeneration and repair.

A large diversity of epigenetic factors, such as microRNAs and histones modifications, are known to be capable of regulating gene expression without altering DNA sequence itself. In particular, miR-1 is considered the first essential microRNA in cardiac development. In this study, miR-1 potential role in early cardiac chamber differentiation is analyzed through specific signaling pathways. For this, we performed in chick embryos functional experiments by means of miR-1 microinjections into the posterior cardiac precursors -of both primitive endocardial tubes- committed to sinoatrial region fates. Subsequently, embryos were subjected to whole mount in situ hybridization, immunohistochemistry and RT-qPCR analysis. As a relevant nov-elty, our results revealed that miR-1 increases Tbx5, Gata4 and AMHC1, while this microRNA diminishes Mef2c expression, during early differentiation of cardiac sinoatrial region. Further-more, we observed in this developmental context that miR-1 upregulates CRABPII and RARß, and downregulates CRABPI, which are three crucial factors in retinoic acid signaling pathway. Interestingly, we also noticed that miR-1 directly interacts with HDAC4 and Calmodulin, as well as with Erk2/MAPK1, which are three key factors actively involved in Mef2c regulation. Our study shows, for the first time, a key role of miR-1 as an epigenetic regulator in early differenti-ation of cardiac sinoatrial region through orchestrating opposite actions between retinoic acid and Mef2c, fundamental to properly assign cardiac cells to their respective heart chambers. A better understanding of those molecular mechanisms modulated by miR-1 will definitely help in fields applied to therapy and cardiac regeneration and repair.

Kaempferol, a flavonoid present in many food products, has chemical and cellular antioxidant properties that are beneficial for protection against the oxidative stress caused by reactive oxygen and nitrogen species. Kaempferol administration to model experimental animals can provide extensive protection against brain damage of the striatum and proximal cortical areas induced by transient brain cerebral ischemic stroke and by 3-nitropropionic acid. This article is an updated review of the molecular and cellular mechanisms of protection by kaempferol administration against brain damage induced by these insults, integrated with an overview of the contributions of the work performed in our laboratories during the past years. Kaempferol administration at doses that prevent neurological dysfunctions inhibit the critical molecular events that underlie the initial and delayed brain damage induced by ischemic stroke and by 3-nitropropionic acid. It is highlighted that the protection afforded by kaempferol against the initial mitochondrial dysfunction can largely account for its protection against the reported delayed spreading of brain damage, which can develop from many hours to several days. This allows us to conclude that kaempferol administration can be beneficial not only in preventive treatments, but also in post-insult therapeutic treatments.

The epicardium is a very dynamic cardiac layer with pivotal contributions during cardiogenesis, acting in the postnatal period as an apparently dormant single-cell layer. In mammalian embryos, the epicardium, which originates form the proepicardium, translocates into the pericardial cavity and subsequently rests on the surface of the myocardium. Later, it gives rise to the epicardium-derived cells, which migrate into subepicardial space, invade the developing myocardium, promoting its growth, and contribute to different cell types. Anomalies in the process of epicardial development, the generation of epicardium-derived cells and their signaling mechanisms in different experimental models lead to defective cardiac development, reminiscent of human congenital heart diseases. Furthermore, recent studies have reported that epicardial derivates in adults, i.e., epicardial adipose tissue, are associated with electrophysiological cardiovascular anomalies. Herein, we provide a state-of-the-art review focusing on both congenital and adult heart diseases associated with epicardial development.

The embryonic epicardium originates from the proepicardium, an extracardiac primordium constituted by a cluster of mesothelial cells. In early embryos, the embryonic epicardium is characterized by a squamous cell epithelium resting on the myocardium surface. Subsequently, it invades the subepicardial space and thereafter the embryonic myocardium by means of an epithelial–mesenchymal transition. Within the myocardium, epicardial-derived cells present multilineage potential, later differentiating into smooth muscle cells and contributing both to coronary vasculature and cardiac fibroblasts in the mature heart. Over the last decades, we have progressively increased our understanding of those cellular and molecular mechanisms driving proepicardial/embryonic epicardium formation. This study provides a state-of-the-art review of the transcriptional and emerging post-transcriptional mechanisms involved in the formation and differentiation of the embryonic epicardium.