miR-1 as a Key Epigenetic Regulator in Early Differentiation of Cardiac Sinoatrial Region.

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.

An Update of Kaempferol Protection against Brain Damage Induced by Ischemia-Reperfusion and by 3-Nitropropionic Acid.

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.

Inhibition of RhoA and Cdc42 by miR-133a Modulates Retinoic Acid Signalling during Early Development of Posterior Cardiac Tube Segment.

It is well known that multiple microRNAs play crucial roles in cardiovascular development, including miR-133a. Additionally, retinoic acid regulates atrial marker expression. In order to analyse the role of miR-133a as a modulator of retinoic acid signalling during the posterior segment of heart tube formation, we performed functional experiments with miR-133a and retinoic acid by means of microinjections into the posterior cardiac precursors of both primitive endocardial tubes in chick embryos. Subsequently, we subjected embryos to whole mount in situ hybridisation, immunohistochemistry and qPCR analysis. Our results demonstrate that miR-133a represses RhoA and Cdc42, as well as Raldh2/Aldh1a2, and the specific atrial markers Tbx5 and AMHC1, which play a key role during differentiation. Furthermore, we observed that miR-133a upregulates p21 and downregulates cyclin A by repressing RhoA and Cdc42, respectively, thus functioning as a cell proliferation inhibitor. Additionally, retinoic acid represses miR-133a, while it increases Raldh2, Tbx5 and AMHC1. Given that RhoA and Cdc42 are involved in Raldh2 expression and that they are modulated by miR-133a, which is influenced by retinoic acid signalling, our results suggest the presence of a negative feedback mechanism between miR-133a and retinoic acid during early development of the posterior cardiac tube segment. Despite additional unexplored factors being possible contributors to this negative feedback mechanism, miR-133a might also be considered as a potential therapeutic tool for the diagnosis, therapy and prognosis of cardiac diseases.

Molecular Mechanisms of lncRNAs in the Dependent Regulation of Cancer and Their Potential Therapeutic Use.

Deep whole genome and transcriptome sequencing have highlighted the importance of an emerging class of non-coding RNA longer than 200 nucleotides (i.e., long non-coding RNAs (lncRNAs)) that are involved in multiple cellular processes such as cell differentiation, embryonic development, and tissue homeostasis. Cancer is a prime example derived from a loss of homeostasis, primarily caused by genetic alterations both in the genomic and epigenetic landscape, which results in deregulation of the gene networks. Deregulation of the expression of many lncRNAs in samples, tissues or patients has been pointed out as a molecular regulator in carcinogenesis, with them acting as oncogenes or tumor suppressor genes. Herein, we summarize the distinct molecular regulatory mechanisms described in literature in which lncRNAs modulate carcinogenesis, emphasizing epigenetic and genetic alterations in particular. Furthermore, we also reviewed the current strategies used to block lncRNA oncogenic functions and their usefulness as potential therapeutic targets in several carcinomas.

LncRNA H19 Impairs Chemo and Radiotherapy in Tumorigenesis.

Various treatments based on drug administration and radiotherapy have been devoted to preventing, palliating, and defeating cancer, showing high efficiency against the progression of this disease. Recently, in this process, malignant cells have been found which are capable of triggering specific molecular mechanisms against current treatments, with negative consequences in the prognosis of the disease. It is therefore fundamental to understand the underlying mechanisms, including the genes-and their signaling pathway regulators-involved in the process, in order to fight tumor cells. Long non-coding RNAs, H19 in particular, have been revealed as powerful protective factors in various types of cancer. However, they have also evidenced their oncogenic role in multiple carcinomas, enhancing tumor cell proliferation, migration, and invasion. In this review, we analyze the role of lncRNA H19 impairing chemo and radiotherapy in tumorigenesis, including breast cancer, lung adenocarcinoma, glioma, and colorectal carcinoma.

X-ray Characterization of Conformational Changes of Human Apo- and Holo-Transferrin.

Human serum transferrin (Tf) is a bilobed glycoprotein whose function is to transport iron through receptor-mediated endocytosis. The mechanism for iron release is pH-dependent and involves conformational changes in the protein, thus making it an attractive system for possible biomedical applications. In this contribution, two powerful X-ray techniques, namely Macromolecular X-ray Crystallography (MX) and Small Angle X-ray Scattering (SAXS), were used to study the conformational changes of iron-free (apo) and iron-loaded (holo) transferrin in crystal and solution states, respectively, at three different pH values of physiological relevance. A crystallographic model of glycosylated apo-Tf was obtained at 3.0 Å resolution, which did not resolve further despite many efforts to improve crystal quality. In the solution, apo-Tf remained mostly globular in all the pH conditions tested; however, the co-existence of closed, partially open, and open conformations was observed for holo-Tf, which showed a more elongated and flexible shape overall.

Early Reactive A1 Astrocytes Induction by the Neurotoxin 3-Nitropropionic Acid in Rat Brain.

3-Nitropropionic acid (NPA) administration to rodents produces degeneration of the striatum, accompanied by neurological disturbances that mimic Huntington's disease (HD) motor neurological dysfunctions. It has been shown that inflammation mediates NPA-induced brain degeneration, and activated microglia secreting cytokines interleukin-1α (IL-1α) and tumor necrosis factor α (TNFα) can induce a specific type of reactive neurotoxic astrocytes, named A1, which have been detected in post-mortem brain samples of Huntington's, Alzheimer's, and Parkinson's diseases. In this work we used an experimental model based on the intraperitoneal (i.p.) administration of NPA to adult Wistar rats at doses that can elicit extensive brain degeneration, and brain samples were taken before and after extensive brain damage monitored using 2,3,5-triphenyltetrazolium chloride (TTC) staining. Western blots and immunohistochemistry of brain slices show that i.p. NPA injections elicit significant increase in the expression levels of C3α subunit, a marker of generation of neurotoxic A1 astrocytes, and of cytokines IL-1α, TNFα, and C1q within the striatum, hippocampus, and cerebellum before the appearance of the HD-related neurological dysfunctions and neuronal death induced by NPA. Noteworthy, NPA administration primarily induces the generation of A1 astrocytes in the more recent phylogenetic area of the rat cerebellum. We conclude that the activation of complement C3 protein in the brain from Wistar rats is an early event in NPA-induced brain neurodegeneration.

Negative Fgf8-Bmp2 feed-back is regulated by miR-130 during early cardiac specification.

It is known that secreted proteins from the anterior lateral endoderm, FGF8 and BMP2, are involved in mesodermal cardiac differentiation, which determines the first cardiac field, defined by the expression of the earliest specific cardiac markers Nkx-2.5 and Gata4. However, the molecular mechanisms responsible for early cardiac development still remain unclear. At present, microRNAs represent a novel layer of complexity in the regulatory networks controlling gene expression during cardiovascular development. This paper aims to study the role of miR130 during early cardiac specification. Our model is focused on developing chick at gastrula stages. In order to identify those regulatory factors which are involved in cardiac specification, we conducted gain- and loss-of-function experiments in precardiac cells by administration of Fgf8, Bmp2 and miR130, through in vitro electroporation technique and soaked beads application. Embryos were subjected to in situ hybridization, immunohistochemistry and qPCR procedures. Our results reveal that Fgf8 suppresses, while Bmp2 induces, the expression of Nkx-2.5 and Gata4. They also show that Fgf8 suppresses Bmp2, and vice versa. Additionally, we observed that Bmp2 regulates miR-130 -a putative microRNA that targets Erk1/2 (Mapk1) 3'UTR, recognizing its expression in precardiac cells which overlap with Erk1/2 pattern. Finally, we evidence that miR-130 is capable to inhibit Erk1/2 and Fgf8, resulting in an increase of Bmp2, Nkx-2.5 and Gata4. Our data present miR-130 as a necessary linkage in the control of Fgf8 signaling, mediated by Bmp2, establishing a negative feed-back loop responsible to achieve early cardiac specification.

MiR-23b and miR-199a impair epithelial-to-mesenchymal transition during atrioventricular endocardial cushion formation.

BACKGROUND: Valve development is a multistep process involving the activation of the cardiac endothelium, epithelial-mesenchymal transition (EMT) and the progressive alignment and differentiation of distinct mesenchymal cell types. Several pathways such as Notch/delta, Tgf-beta and/or Vegf signaling have been implicated in crucial steps of valvulogenesis. We have previously demonstrated discrete changes in microRNAs expression during cardiogenesis, which are predicted to target Bmp- and Tgf-beta signaling. We now analyzed the expression profile of 20 candidate microRNAs in atrial, ventricular, and atrioventricular canal regions at four different developmental stages. RESULTS: qRT-PCR analyses of microRNAs demonstrated a highly dynamic and distinct expression profiles within the atrial, ventricular, and atrioventricular canal regions of the developing chick heart. miR-23b, miR-199a, and miR-15a displayed increased expression during early AVC development whereas others such as miR-130a and miR-200a display decreased expression levels. Functional analyses of miR-23b, miR-199a, and miR-15a overexpression led to in vitro EMT blockage. Molecular analyses demonstrate that distinct EMT signaling pathways are impaired after microRNA expression, including a large subset of EMT-related genes that are predicted to be targeted by these microRNAs. CONCLUSIONS: Our data demonstrate that miR-23b and miR-199a over-expression can impair atrioventricular EMT.

The decrease of NAD(P)H:quinone oxidoreductase 1 activity and increase of ROS production by NADPH oxidases are early biomarkers in doxorubicin cardiotoxicity.

CONTEXT: Doxorubicin cardiotoxicity displays a complex and multifactorial progression. OBJECTIVE: Identify early biochemical mechanisms leading to a sustained imbalance of cellular bioenergetics. METHODS: Measurements of the temporal evolution of selected biochemical markers after treatment of rats with doxorubicin (20 mg/kg body weight). RESULTS: Doxorubicin treatment increased lipid oxidation, catalase activity and production of H2O2 by Nox-NADPH oxidases, and down-regulated NAD(P)H: quinone oxidoreductase-1 prior eliciting changes in reduced glutathione, protein carbonyls and protein nitrotyrosines. Alterations of mitochondrial and myofibrillar bioenergetics biomarkers were detected only after this oxidative imbalance was established. NAD(P)H: quinone oxidoreductase-1 activity and increase of hydrogen peroxide production by NADPH oxidases are early biomarkers in doxorubicin cardiotoxicity.

Characterization of an extremophile bacterial acid phosphatase derived from metagenomics analysis.

Acid phosphatases are enzymes that play a crucial role in the hydrolysis of various organophosphorous molecules. A putative acid phosphatase called FS6 was identified using genetic profiles and sequences from different environments. FS6 showed high sequence similarity to type C acid phosphatases and retained more than 30% of consensus residues in its protein sequence. A histidine-tagged recombinant FS6 produced in Escherichia coli exhibited extremophile properties, functioning effectively in a broad pH range between 3.5 and 8.5. The enzyme demonstrated optimal activity at temperatures between 25 and 50°C, with a melting temperature of 51.6°C. Kinetic parameters were determined using various substrates, and the reaction catalysed by FS6 with physiological substrates was at least 100-fold more efficient than with p-nitrophenyl phosphate. Furthermore, FS6 was found to be a decamer in solution, unlike the dimeric forms of crystallized proteins in its family.

Novel Insights into the Molecular Mechanisms Governing Embryonic Epicardium Formation.

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.

Complex I and cytochrome c are molecular targets of flavonoids that inhibit hydrogen peroxide production by mitochondria.

Flavonoids can protect cells from different insults that lead to mitochondria-mediated cell death, and epidemiological data show that some of these compounds attenuate the progression of diseases associated with oxidative stress and mitochondrial dysfunction. In this work, a screening of 5 flavonoids representing major subclasses showed that they display different effects on H2O2 production by mitochondria isolated from rat brain and heart. Quercetin, kaempferol and epicatechin are potent inhibitors of H2O2 production by mitochondria from both tissues (IC50 approximately 1-2 µM), even when H2O2 production rate was stimulated by the mitochondrial inhibitors rotenone and antimycin A. Although the rate of oxygen consumption was unaffected by concentrations up to 10 µM of these flavonoids, quercetin, kaempferol and apigenin inhibited complex I activity, while up to 100 µM epicatechin produced less than 20% inhibition. The extent of this inhibition was found to be dependent on the concentration of coenzyme Q in the medium, suggesting competition between the flavonoids and ubiquinone for close binding sites in the complex. In contrast, these flavonoids did not significantly inhibit the activity of complexes II and III, and did not affect the redox state of complex IV. However, we have found that epicatechin, quercetin and kaempferol are able to stoichiometrically reduce purified cytochrome c. Our results reveal that mitochondria are a plausible main target of flavonoids mediating, at least in part, their reported preventive actions against oxidative stress and mitochondrial dysfunction-associated pathologies.

Kaempferol prevents the activation of complement C3 protein and the generation of reactive A1 astrocytes that mediate rat brain degeneration induced by 3-nitropropionic acid.

Kaempferol is a natural antioxidant present in vegetables and fruits used in human nutrition. In previous work, we showed that intraperitoneal (i.p.) kaempferol administration strongly protects against striatum neurodegeneration induced by i.p. injections of 3-nitropropionic acid (NPA), an animal model of Huntington's disease. Recently, we have shown that reactive A1 astrocytes generation is an early event in the neurodegeneration induced by NPA i.p. injections. In the present work, we have experimentally evaluated the hypothesis that kaempferol protects both against the activation of complement C3 protein and the generation of reactive A1 astrocytes in rat brain striatum and hippocampus. To this end, we have administered NPA and kaempferol i.p. injections to adult Wistar rats following the protocol described in previous work. Kaempferol administration prevents proteolytic activation of complement C3 protein and generation of reactive A1 astrocytes NPA-induced in the striatum and hippocampus. Also, it blocked the NPA-induced increase of NF-κB expression and enhanced secretion of cytokines IL-1α, TNFα, and C1q, which have been linked to the generation of reactive A1 astrocytes. In addition, kaempferol administration prevented the enhanced production of amyloid ß peptides in the striatum and hippocampus, a novel finding in NPA-induced brain degeneration found in this work.

Comparative Analysis of Non-Coding RNA Transcriptomics in Heart Failure.

Heart failure constitutes a clinical complex syndrome with different symptomatic characteristics depending on age, sex, race and ethnicity, among others, which has become a major public health issue with an increasing prevalence. One of the most interesting tools seeking to improve prevention, diagnosis, treatment and prognosis of this pathology has focused on finding new molecular biomarkers since heart failure relies on deficient cardiac homeostasis, which is regulated by a strict gene expression. Therefore, currently, analyses of non-coding RNA transcriptomics have been oriented towards human samples. The present review develops a comparative study emphasizing the relevance of microRNAs, long non-coding RNAs and circular RNAs as potential biomarkers in heart failure. Significantly, further studies in this field of research are fundamental to supporting their widespread clinical use. In this sense, the various methodologies used by the authors should be standardized, including larger cohorts, homogeneity of the samples and uniformity of the bioinformatic pipelines used to reach stratification and statistical significance of the results. These basic adjustments could provide promising steps to designing novel strategies for clinical management of patients with heart failure.

New Insights into the Roles of lncRNAs as Modulators of Cytoskeleton Architecture and Their Implications in Cellular Homeostasis and in Tumorigenesis.

The importance of the cytoskeleton not only in cell architecture but also as a pivotal element in the transduction of signals that mediate multiple biological processes has recently been highlighted. Broadly, the cytoskeleton consists of three types of structural proteins: (1) actin filaments, involved in establishing and maintaining cell shape and movement; (2) microtubules, necessary to support the different organelles and distribution of chromosomes during cell cycle; and (3) intermediate filaments, which have a mainly structural function showing specificity for the cell type where they are expressed. Interaction between these protein structures is essential for the cytoskeletal mesh to be functional. Furthermore, the cytoskeleton is subject to intense spatio-temporal regulation mediated by the assembly and disassembly of its components. Loss of cytoskeleton homeostasis and integrity of cell focal adhesion are hallmarks of several cancer types. Recently, many reports have pointed out that lncRNAs could be critical mediators in cellular homeostasis controlling dynamic structure and stability of the network formed by cytoskeletal structures, specifically in different types of carcinomas. In this review, we summarize current information available about the roles of lncRNAs as modulators of actin dependent cytoskeleton and their impact on cancer pathogenesis. Finally, we explore other examples of cytoskeletal lncRNAs currently unrelated to tumorigenesis, to illustrate knowledge about them.

Prostatic artery occlusion: initial findings on pathophysiological response in a canine prostate model.

Background: Prostatic artery embolization (PAE) is an alternative treatment for symptomatic benign prostatic hyperplasia (BPH) in men. A technical modification of conventional PAE has been developed in a canine prostate model consisting of prostatic artery occlusion (PAO) using Onyx® whose therapeutic effect is prostate shrinkage. However, the underlying mechanisms are not well clarified. The purpose was to evaluate the biological mechanisms responsible for therapeutic effects of PAO in the canine prostate. Methods: Ten adult male beagles (5.0±0.82 years) underwent PAO with Onyx-18 (n=7) and prostatic artery angiography as control (n=3). Blood samples were taken at different time points of follow-up (baseline, 1 week, 2 weeks, 1 month, 3 months and 6 months) to measure the serum canine prostate specific esterase (CPSE). MRI examinations were also performed to document the prostate volume (PV) before and after interventions at different time points of follow-up. Prostates were harvested at 2 weeks (n=2) in the PAO-group, and the remaining ones (n=8) at 6 months for the determinations of intraprostatic testosterone and dihydrotestosterone (DHT) by ELISA, apoptosis by TUNEL assay and histopathological study. Results: The mean serum CPSE concentration started to decrease significantly from 2 weeks to 6 months after PAO along with PV compared with baseline data. In addition, a moderate but significant correlation was observed between CPSE and PV (r=0.655, P=0.000). Regarding intraprostatic androgens, testosterone was significantly higher after PAO than control (19.70 vs. 4.87 ng/mL, P=0.002), whereas DHT was lower but no significant (112.52 vs. 138.35 pg/mL, P=0.144). In histological study, PAO induced a severe hemorrhagic necrosis in the whole prostates along with inflammatory cell infiltration at early 2 weeks, and then diffuse interstitial fibrosis with atrophy of the glandular epithelium and intraprostatic cavity formation at 6 months. Apoptosis was detected in all specimens with higher apoptotic index after PAO at 2 weeks (7.35%) and at 6 months (4.38%) compared with control (2.64%), without statistically significant difference between groups. Conclusions: PAO induces hemorrhagic ischemia predominantly resulting in necrosis rather than apoptosis with prostate shrinkage. CPSE is a potential biomarker to assess the response to PAO in the canine prostate.

Transarterial prostatic embolization: initial experience in a canine model.

OBJECTIVE: The purpose of this study was to prospectively evaluate pathologic responses to transarterial prostatic embolization and its technical safety in a canine model. MATERIALS AND METHODS: Ten adult male beagle dogs were surgically castrated and given hormonal therapy for 4 months to induce prostatic hyperplasia. After three months of hormonal therapy, the dogs were randomly assigned to a transarterial prostatic embolization group (n = 7) or a control group (n = 3). Dogs in the transarterial prostatic embolization group were subjected to embolization with microspheres 300-500 µm in diameter. Four months after the study was begun, all dogs were sacrificed for pathologic study. Transrectal ultrasound and MRI were performed to evaluate pathologic responses. The data on prostate size acquired with transrectal ultrasound were processed for statistical analysis by paired Student t test. RESULTS: The canine prostatic hyperplasia model was successfully established in 10 dogs. The increase in mean prostate size being as great as 572% after 3 months of hormonal therapy. An intraprostatic cavity was detected 1 month after transarterial prostatic embolization in all seven dogs. Four dogs had significant shrinkage of the prostate, and the other three had an increase in prostate size. Imaging examinations and necropsy revealed a huge cavity occupying almost the entire prostate in the three dogs with increased prostate size. No complications associated with transarterial prostatic embolization were encountered. CONCLUSION: Transarterial prostatic embolization is a safe procedure that can induce prostatic infarction and ablate the prostate. The findings suggest the procedure has potential clinical applications in the care of patients with benign prostatic hyperplasia.

Non-Coding RNAs in Retinoic Acid as Differentiation and Disease Drivers.

: All-trans retinoic acid (RA) is the most active metabolite of vitamin A. Several studies have described a pivotal role for RA signalling in different biological processes such as cell growth and differentiation, embryonic development and organogenesis. Since RA signalling is highly dose-dependent, a fine-tuning regulatory mechanism is required. Thus, RA signalling deregulation has a major impact, both in development and disease, related in many cases to oncogenic processes. In this review, we focus on the impact of ncRNA post-transcriptional regulatory mechanisms, especially those of microRNAs and lncRNAs, in RA signalling pathways during differentiation and disease.

Dynamic MicroRNA Expression Profiles During Embryonic Development Provide Novel Insights Into Cardiac Sinus Venosus/Inflow Tract Differentiation.

MicroRNAs have been explored in different organisms and are involved as molecular switches modulating cellular specification and differentiation during the embryonic development, including the cardiovascular system. In this study, we analyze the expression profiles of different microRNAs during early cardiac development. By using whole mount in situ hybridization in developing chick embryos, with microRNA-specific LNA probes, we carried out a detailed study of miR-23b, miR-130a, miR-106a, and miR-100 expression during early stages of embryogenesis (HH3 to HH17). We also correlated those findings with putative microRNA target genes by means of mirWalk and TargetScan analyses. Our results demonstrate a dynamic expression pattern in cardiac precursor cells from the primitive streak to the cardiac looping stages for miR-23b, miR-130a, and miR-106a. Additionally, miR-100 is later detectable during cardiac looping stages (HH15-17). Interestingly, the sinus venosus/inflow tract was shown to be the most representative cardiac area for the convergent expression of the four microRNAs. Through in silico analysis we revealed that distinct Hox family members are predicted to be targeted by the above microRNAs. We also identified expression of several Hox genes in the sinus venosus at stages HH11 and HH15. In addition, by means of gain-of-function experiments both in cardiomyoblasts and sinus venosus explants, we demonstrated the modulation of the different Hox clusters, Hoxa, Hoxb, Hoxc, and Hoxd genes, by these microRNAs. Furthermore, we correlated the negative modulation of several Hox genes, such as Hoxa3, Hoxa4, Hoxa5, Hoxc6, or Hoxd4. Finally, we demonstrated through a dual luciferase assay that Hoxa1 is targeted by miR-130a and Hoxa4 is targeted by both miR-23b and miR-106a, supporting a possible role of these microRNAs in Hox gene modulation during differentiation and compartmentalization of the posterior structures of the developing venous pole of the heart.