Coronary Vessel Development Research Articles

Coronary Vasculature in Cardiac Development and Regeneration.

Functional coronary circulation is essential for a healthy heart in warm-blooded vertebrates, and coronary diseases can have a fatal consequence. Despite the growing interest, the knowledge about the coronary vessel development and the roles of new coronary vessel formation during heart regeneration is still limited. It is demonstrated that early revascularization is required for efficient heart regeneration. In this comprehensive review, we first describe the coronary vessel formation from an evolutionary perspective. We further discuss the cell origins of coronary endothelial cells and perivascular cells and summarize the critical signaling pathways regulating coronary vessel development. Lastly, we focus on the current knowledge about the molecular mechanisms regulating heart regeneration in zebrafish, a genetically tractable vertebrate model with a regenerative adult heart and well-developed coronary system.

The Bicuspid Condition of the Aortic Valve Does Not Alter the Incidence of Accessory Coronary Artery Ostia in Syrian Hamsters (Mesocricetus auratus)

In man and Syrian hamsters (Mesocricetus auratus), the prevalence of anomalies in the origin of the coronary arteries is significantly higher in individuals with bicuspid than with normal aortic valves. In hamsters, the incidence of accessory ostia is similar in individuals with normal and anomalous coronary arteries, all of them possessing a normal (tricuspid) aortic valve. In order to evaluate whether or not the presence of bicuspid aortic valves alters the incidence of accessory ostia, 1,050 hearts from hamsters with bicuspid valves were examined. In 594 of them the coronary arteries were normal. The remaining 456 hearts showed coronary artery anomalies characterized by the absence of any artery arising from the left side of the valve. The incidence of accessory ostia was 3.9% in hamsters with normal coronary arteries and 2.2% in those with anomalous coronary patterns. Overall, 3.1% of the accessory ostia were associated with a septal artery and another 0.2% with a conal artery. These data referring to the bicuspid valves were compared with those already published on normal valves. The results of statistical analyses showed that having a bicuspid aortic valve does not alter the incidence of accessory coronary ostia. In the set of tricuspid and bicuspid valves, the incidence of accessory ostia was significantly lower on the left side than on the right side of the valve. This, together with the fact that in the present animal model the coronary anomalies were characterized by the absence of arteries on the left side of the valve, suggests that the embryonic region corresponding to the left side of the aortic valve primordium is particularly associated with preventing the normal development of coronary vessels.

CCBE1 is required for coronary vessel development and proper coronary artery stem formation in the mouse heart

Proper coronary vasculature development is essential for late-embryonic and adult heart function. The developmental regulation of coronary embryogenesis is complex and includes the coordinated activity of multiple signaling pathways. CCBE1 plays an important role during lymphangiogenesis, enhancing VEGF-C signaling, which is also required for coronary vasculature formation. However, whether CCBE1 plays a similar role during coronary vasculature development is still unknown. Here, we investigate the coronary vasculature development in Ccbe1 mutant embryos. We show that Ccbe1 is expressed in the epicardium, like Vegf-c, and also in the sinus venosus (SV) at the stages of its contribution to coronary vasculature formation. We also report that absence of CCBE1 in cardiac tissue inhibited coronary growth that sprouts from the SV endocardium at the dorsal cardiac wall. This disruption of coronary formation correlates with abnormal processing of VEGF-C propeptides, suggesting VEGF-C-dependent signaling alteration. Moreover, Ccbe1 loss-of-function leads to the development of defective dorsal and ventral intramyocardial vessels. We also demonstrate that Ccbe1 mutants display delayed and mispatterned coronary artery (CA) stem formation. CCBE1 is essential for coronary vessel formation, independent of their embryonic origin, and is also necessary for peritruncal vessel growth and proper CA stem patterning. Developmental Dynamics 247:1135-1145, 2018. © 2018 Wiley Periodicals, Inc.

Anatomy of the coronary artery and cardiac vein in the quail ventricle: patterns are distinct from those in mouse and human hearts.

Coronary vessel development has been investigated in avian and mouse embryonic hearts. Quail embryos are a useful tool to examine vascular development, particularly because the QH1 antibody and transgenic quail line, Tg (tie1:H2B-eYFP), are useful to trace endothelial cells. However, there are only a few descriptions of the quail coronary vessels. Using ink injection coronary angiography, we examined the course of coronary vessels in the fetal quail heart. The major coronary arteries were the right and left septal arteries, which, respectively, branched off from the right and left coronary stems. The right septal artery ran posteriorly (dorsally) and penetrated the ventricular free wall to distribute to the posterior surface of the ventricles. The left septal artery ran anteriorly (ventrally) and penetrated the ventricular free wall to distribute to the anterior surface of the ventricles. The right and left circumflex arteries were directed posteriorly along the atrioventricular sulci. The cardiac veins consisted of three major tributaries: the middle, great, and anterior cardiac veins. The middle cardiac vein ascended along the posterior interventricular sulcus and emptied into the right atrium. The great cardiac vein ran along the anterior interventricular sulcus, entered the space between the left atrium and conus arteriosus and emptied into the right atrium behind the aortic bulb. The anterior cardiac vein drained the anterior surface of the right ventricle and connected to the anterior base of the right atrium. The course of coronary vessels in the quail heart was basically the same as that observed in chick but was different from those of mouse and human.

YKL-40 levels increase with declining ankle-brachial index and are associated with long-term cardiovascular mortality in peripheral arterial disease patients

Background and aimsYKL-40 is an inflammatory marker secreted by macrophages and is expressed in atherosclerotic plaques. YKL-40 increases in coronary artery disease (CAD) with poor coronary collateral vessel development. Higher levels are linked to reduced survival in CAD patients. Studies evaluating YKL-40 in patients with peripheral arterial disease (PAD) are scarce. This study aims to elucidate a possible link between YKL-40 and PAD severity as well as cardiovascular long-term mortality. MethodsYKL-40 was measured at baseline in 365 elderly PAD patients (age 69 ± 10.4, 33.7% women, Fontaine stage I-II) by bead-based multiplex assay. Patients were followed for seven years to assess long-term cardiovascular and all-cause survival by Kaplan-Meier and Cox regression. ResultsYKL-40 levels were associated with declining ankle-brachial index (ABI) in PAD patients without Moenckeberg's mediasclerosis (R = −0.189, p=0.002). PAD patients with mediasclerosis exhibited higher YKL-40 levels (p=0.002). Baseline YKL-40 levels were significantly associated with cardiovascular mortality (HR 1.52 (1.21–1.91), p < 0.001) and all-cause mortality (HR 1.45 (1.20–1.75), p < 0.001) over a seven-year observation period. After multivariable adjustment for gender, patient age, known carotid artery disease, known coronary artery disease, smoking status, systolic blood pressure, HbA1c, low density lipoprotein cholesterol, estimated glomerular filtration rate, aspartate aminotransferase, and C-reactive protein, YKL-40 remained significantly associated with cardiovascular (HR 1.34 (1.02–1.75), p=0.033) and all-cause mortality (HR 1.25 (1.01–1.55), p=0.039). ConclusionsIncreased YKL-40 levels are independently associated with poor long-term cardiovascular survival in peripheral arterial disease patients. Furthermore, YKL-40 correlates with patients' ABI in PAD in the absence of mediasclerosis.

Contributions of an Extracellular Matrix Glycoprotein to Coronary Vessel Formation

Congenital coronary artery anomalies occur in up to 5% of the human population. Such anomalies can lead to myocardial ischemia, myocardial infarction and sudden death. Coronary vessels are composed of endothelial and smooth muscle cells. The primary function of vascular endothelial cells is to provide a barrier between the vessel lumen and surrounding tissue. Vascular repair processes often recapitulate developmental mechanisms. Elucidation of signaling mechanisms that govern formation of nascent coronary endothelial cells is required to develop therapeutic strategies aimed at regrowth of congenitally malformed or damaged coronary vessels. Embryonic mice with epicardial‐specific deletion of Tbx5 (Tbx5epi−/−) exhibit delayed epicardial cell attachment to the myocardium and impaired coronary vessel formation. This leads to punctate cardiac hemorrhaging that may stem from deficits in vascular integrity. Expression of Reln mRNA, that encodes the Reelin extracellular matrix glycoprotein, is also reduced in embryonic Tbx5epi−/− mouse hearts. Reelin localizes to some, but not all, coronary vascular endothelial cells during mammalian cardiogenesis. These findings lead us to speculate that expression of Reelin in coronary vascular endothelial cells may be critical for establishing vascular integrity.Study ObjectiveThe goal of this study was to elucidate contributions of Reelin to endothelial cell function for establishing vascular integrity during coronary vessel formation.MethodsWe utilized human microvascular endothelial cells (HMECs) to assess contributions of Reelin to endothelial cell function as these cells are appropriate in vitro models of vessel formation and have endogenous Reelin expression. We achieved RELN gene silencing through a small interfering RNA‐mediated approach that led to &gt;90% reduction in both RELN mRNA and Reelin protein expression in comparison to negative control‐transfected HMECs. Next, control and RELN‐silenced endothelial cells were subjected to physiological assays to assess their ability to establish vascular integrity. First, we examined the ability of RELN‐silenced HMECs to adhere to the basement membrane through exposure of these cells to various matrices. Second, we assessed cell membrane permeability of RELN‐silenced HMECs that were plated as a monolayer on a collagen‐coated semi‐permeable barrier. This served as an indicator of in vitro vascular permeability. The cell monolayer was treated with a permeability factor and fluorescent‐labeled Dextran was added on top of the monolayer. Fluorescent Dextran passed through the cell monolayer at a rate proportional to membrane permeability. Permeability was measured by quantifying the fluorescence of the media on the basolateral side of the monolayer.ResultsOur results indicate that RELN silencing alters in vitro endothelial cell adhesion and cell membrane permeability.ConclusionsWe conclude that Reelin plays a critical role during coronary vessel development as it establishes vascular integrity through maintenance of cell adhesions and membrane permeabilitySupport or Funding InformationAHA 17AIREA3360773 and PCOM Center for Chronic Disorders of AgingThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Meet the First Authors.

Meet the First Authors.

Abstract 237: PRMT1-p53-Slug Axis Regulates Epicardial EMT and Ventricular Development

Rationale: Epicardial epithelial-to-mesenchymal trasition (EMT) is a vital process in embryonic heart development. During EMT, epicardial cells acquire migratory and invasive properties, and differentiate into new cell types, including cardiac fibroblasts and coronary smooth muscle cells. Non-histone protein methylation is an emerging modulator of cell signaling. We have recently established a role for protein arginine methyltransferase-1 (PRMT1) in TGF-β-induced EMT in cultured cells. Objective: To determine the role of PRMT1 in epicardial EMT. Methods and Results: We investigated the role of PRMT1 in epicardial EMT in mouse epicardial cells. Embryonic day 9.5 (E9.5) tamoxifen administration of WT1-Cre ERT ;PRMT1 fl/fl ;ROSA-YFP fl/fl mouse embryos was used to delete PRMT1 in the epicardium. Epicardial PRMT1 deletion led to reduced epicardial migration into the myocardium, a thinner compact myocardial layer, and dilated coronary blood vessels at E15.5. Using the epicardial cell line MEC1, we found that PRMT1 siRNA prevented the increase in mesenchymal proteins Slug and Fibronectin and the decrease in epithelial protein E-Cadherin during TGF-β treatment-induced EMT. PRMT1 siRNA also reduced the migration and invasion of MEC1 cells. We further identified that PRMT1 siRNA also increased the expression of p53, a key regulator of the Slug degradation pathway. PRMT1 siRNA increases p53 expression by decreasing p53 degradation, and shifted p53 localization to the cytoplasm. In vitro methylation assays further demonstrated that PRMT1 methylates p53. Knockdown of p53 increased Slug levels and enhanced EMT, establishing p53 as a regulator of epicardial EMT through controlling Slug expression. Furthermore, RNAseq experiments in MEC1 cells demonstrated that 40% (545/1,351) of TGF-β-induced transcriptional changes were prevented by PRMT1 siRNA. Furthermore, when p53 and PRMT1 were simultaneously knocked down, TGF-β induced transcriptional control of 37% (201/545) of these PRMT1-dependent genes was restored. Conclusions: The PRMT1-p53-Slug pathway is necessary for epicardial EMT in cultured MEC1 cells as well as in the epicardium in vivo . Epicardial PRMT1 is required for the development of compact myocardium and coronary blood vessels.

Nonmuscle myosin IIB regulates epicardial integrity and epicardium-derived mesenchymal cell maturation.

Nonmuscle myosin IIB (NMIIB; heavy chain encoded by MYH10) is essential for cardiac myocyte cytokinesis. The role of NMIIB in other cardiac cells is not known. Here, we show that NMIIB is required in epicardial formation and functions to support myocardial proliferation and coronary vessel development. Ablation of NMIIB in epicardial cells results in disruption of epicardial integrity with a loss of E-cadherin at cell-cell junctions and a focal detachment of epicardial cells from the myocardium. NMIIB-knockout and blebbistatin-treated epicardial explants demonstrate impaired mesenchymal cell maturation during epicardial epithelial-mesenchymal transition. This is manifested by an impaired invasion of collagen gels by the epicardium-derived mesenchymal cells and the reorganization of the cytoskeletal structure. Although there is a marked decrease in the expression of mesenchymal genes, there is no change in Snail (also known as Snai1) or E-cadherin expression. Studies from epicardium-specific NMIIB-knockout mice confirm the importance of NMIIB for epicardial integrity and epicardial functions in promoting cardiac myocyte proliferation and coronary vessel formation during heart development. Our findings provide a novel mechanism linking epicardial formation and epicardial function to the activity of the cytoplasmic motor protein NMIIB.

Histological and immunocytochemical investigation of human coronary vessel development with anti-CD34 antibodies

Information about embryonic development of coronary endothelium is the main clue for the creation of new methods in tissue engineering for treatment of ischemic heart diseases. The purpose of the research was to describe human coronary vessels development on early stages of the prenatal ontogenesis. The first step in human coronary vessels development is the formation of endothelium de novo by transformation of some epicardial and, possibly, endocardial cells. The next step is the ingrowth of sinus venosus endothelium in subepicardium over ventricles and atria, which gives rise to the coronary vessels. Only after 7 days does the primitive coronary plexus of the heart communicate with aorta (third step). During this period, some subepicardial vessels invade myocardium and some intramyocardial vessels contact with the heart cavity. Such intercommunications could help in regulation of blood circulation in primitive coronary plexus before establishment of effective contacts between arterial and venous vessels—excess of blood could be discharged directly into the heart cavity. Additional population of CD34+ cells were revealed inside condensed mesenchyme of the conotruncus; it participates in the formation of vasa vasorum in the aorta. Epicardium and sinus venosus generate endothelium of coronary vessels by neovasculo- and angiogenesis, respectively. During a week after ingrowth of vessels from SV and before their ingrowth to the aorta, ventriculo-coronary communications could be found in the heart.

Vinculin b deficiency causes epicardial hyperplasia and coronary vessel disorganization in zebrafish

Coronary vessel development is a highly coordinated process during heart formation. Abnormal development and dysfunction of the coronary network are contributory factors in the majority of heart disease. Understanding the molecular mechanisms that regulate coronary vessel formation is crucial for preventing and treating the disease. We report a zebrafish gene-trap vinculin b (vclb) mutant that displays abnormal coronary vessel development among multiple cardiac defects. The mutant shows overproliferation of epicardium-derived cells and disorganization of coronary vessels, and they eventually die off at juvenile stages. Mechanistically, Vclb deficiency results in the release of another cytoskeletal protein, paxillin, from the Vclb complex and the upregulation of ERK and FAK phosphorylation in epicardium and endocardium, causing disorganization of endothelial cells and pericytes during coronary vessel development. By contrast, cardiac muscle development is relatively normal, probably owing to redundancy with Vcla, a vinculin paralog that is expressed in the myocardium but not epicardium. Together, our results reveal a previously unappreciated function of vinculin in epicardium and endocardium and reinforce the notion that well-balanced FAK activity is essential for coronary vessel development.

Transcriptional Profiling of Cultured, Embryonic Epicardial Cells Identifies Novel Genes and Signaling Pathways Regulated by TGFβR3 In Vitro.

The epicardium plays an important role in coronary vessel formation and Tgfbr3-/- mice exhibit failed coronary vessel development associated with decreased epicardial cell invasion. Immortalized Tgfbr3-/- epicardial cells display the same defects. Tgfbr3+/+ and Tgfbr3-/- cells incubated for 72 hours with VEH or ligands known to promote invasion via TGFβR3 (TGFβ1, TGFβ2, BMP2), for 72 hours were harvested for RNA-seq analysis. We selected for genes >2-fold differentially expressed between Tgfbr3+/+ and Tgfbr3-/- cells when incubated with VEH (604), TGFβ1 (515), TGFβ2 (553), or BMP2 (632). Gene Ontology (GO) analysis of these genes identified dysregulated biological processes consistent with the defects observed in Tgfbr3-/- cells, including those associated with extracellular matrix interaction. GO and Gene Regulatory Network (GRN) analysis identified distinct expression profiles between TGFβ1-TGFβ2 and VEH-BMP2 incubated cells, consistent with the differential response of epicardial cells to these ligands in vitro. Despite the differences observed between Tgfbr3+/+ and Tgfbr3-/- cells after TGFβ and BMP ligand addition, GRNs constructed from these gene lists identified NF-ĸB as a key nodal point for all ligands examined. Tgfbr3-/- cells exhibited decreased expression of genes known to be activated by NF-ĸB signaling. NF-ĸB activity was stimulated in Tgfbr3+/+ epicardial cells after TGFβ2 or BMP2 incubation, while Tgfbr3-/- cells failed to activate NF-ĸB in response to these ligands. Tgfbr3+/+ epicardial cells incubated with an inhibitor of NF-ĸB signaling no longer invaded into a collagen gel in response to TGFβ2 or BMP2. These data suggest that NF-ĸB signaling is dysregulated in Tgfbr3-/- epicardial cells and that NF-ĸB signaling is required for epicardial cell invasion in vitro. Our approach successfully identified a signaling pathway important in epicardial cell behavior downstream of TGFβR3. Overall, the genes and signaling pathways identified through our analysis yield the first comprehensive list of candidate genes whose expression is dependent on TGFβR3 signaling.

Relationship between thrombospondin-1, endostatin, angiopoietin-2, and coronary collateral development in patients with chronic total occlusion.

This study is aimed to investigate whether serum angiostatic factors (thrombospondin-1 [TSP-1] and endostatin) or angiogenic factors (angiopoietin-2 [Ang-2]) are related to coronary collateral vessel development in patients with chronic total occlusion (CTO).A total of 149 patients were enrolled in the study, and 39 patients with coronary artery disease but without significant stenosis were included in control group. In 110 patients with CTO lesion, 79 with Rentrop grades 2 to 3 collaterals were grouped as good collateral, while 31 with Rentrop grades 0 to 1 collaterals were grouped as poor collateral. Serum TSP-1, endostatin, and Ang-2 levels were studied.Serum endostatin level was significantly higher in poor collateral group compared with control group and good collateral group, respectively (96.2 ± 30.4 vs 77.8 ± 16.5 ng/mL, P = 0.007; 96.2 ± 30.4 vs 81.2 ± 30.4 ng/mL, P = 0.018). In multivariate analysis, decreased serum endostatin level was independently related to good coronary collateral development. Serum TSP-1 level was lower in patients with CTO compared with control group. However, no difference in TSP-1 level was detected between poor and good collateral group. The serum Ang-2 level did not show a significant difference among 3 groups.Circulatory endostatin may be a useful biomarker for coronary collateral development and potential target for therapeutic angiogenesis in patients with CTO.

Abstract 245: Cxcl12 Chemokine Signaling Regulates Synchronous Development of Coronary Vessels and Myocardium

Introduction: The coronary vasculature plays critical roles in heart functions. Coronary heart disease is a major cause of myocardial infarction and heart failure, which continues to be the leading cause of mortality. However, the mechanisms underlying coronary vessel development and myocardial vascularization are not fully understood. Furthermore, it is not known if myocardial developmental defects caused by lack of vascularization will lead to decreased regenerative capacity of the adult heart. Hypothesis: Synchronous development with coronary vessels is essential for normal morphogenesis and regenerative capacity of the myocardium of zebrafish heart. Methods and Results: Using transgenic reporter fish lines, we observed that during zebrafish heart development, coronary vessels form a close association with a specific subpopulation of cortical cardiomyocytes marked by the transcription factor Gata4. When gata4+ cardiomyocytes emerge onto the surface of the juvenile zebrafish hearts, they follow the tracks of coronary endothelial cells. We previously showed that cxcr4a mutant zebrafish fail to form a vascular network, whereas ectopic expression of Cxcl12b ligand induces coronary vessel formation. In cxcr4a mutant hearts, gata4+ cardiomyocytes fail to associate with coronary vessels. Importantly, cxcr4a mutant zebrafish fail to undergo heart regeneration following injury. Gene expression analysis using RNAseq was performed to dissect the molecular mechanisms underlying synchronous development of coronary vessels and gata4+ cardiomyocytes. Conclusion: Our results suggest that Cxcl12/Cxcr4 chemokine signaling has an essential role in directing cardiomyocyte migration guided by endothelial cells. Poorly developed vasculature in cxcr4a mutants likely underlies defects in myocardial development and decreased regenerative potential of zebrafish hearts.

Notch signalling in ventricular chamber development and cardiomyopathy.

The vertebrate heart is the first organ to form and function during embryogenesis. Primitive streak-derived cardiac progenitors located bilaterally move rostral to form the primitive heart tube that subsequently undergoes rightward looping, remodelling and septation to give rise to the mature four-chambered heart. Tightly regulated tissue interactions orchestrate the patterning, proliferation and differentiation processes that give rise to the adult ventricles. Studies in animal models have demonstrated the crucial function of the Notch signalling pathway in ventricular development and how alterations in human NOTCH signalling may lead to disease in the form of cardiomyopathies, such as left ventricular noncompaction (LVNC). In this review, we discuss how during trabecular formation and ventricular compaction, Dll4-Notch1 signals from chamber endocardium to regulate cardiomyocyte proliferation and differentiation in a noncell autonomous fashion and how, at later stages, myocardial Jag1 and Jag2 activate Notch1 in chamber endocardium to sustain chamber patterning and compaction with simultaneous coronary vessel development mediated by Dll4-Notch1. We suggest that alterations in these molecular mechanisms underlie MIB1-related familial LVNC and favour the hypothesis that this cardiomyopathy has a congenital nature.

CXCL12 And Vascular Maturation Of The Coronary System During Heart Development

Elucidating the mechanisms involved in the patterning of coronary blood vessels in the embryo will help to improve therapies for coronary perfusion in the injured heart. In the past few years, there have been significant efforts and advances in the understanding of coronary vessel development. The major endothelial cell sources that provide coronary endothelial progenitors to the heart are the sinus venosus (the vein that delivers the venous return) and the endocardium, although the respective contribution from each source is debated. Angiogenic sprouts from the sinus venosus reach the ventricles starting at E11.5 in the mouse and form a vascular plexus in the subepicardial space. This primitive plexus undergoes a complex and incompletely understood process of vascular maturation that involves pruning, branching and reorganization into a hierarchical network of superficial veins and intramyocardial arteries. The main branches of the coronary arteries ultimately connect to the aorta through the coronary ostia by E13.5 – E14.0.CXCL12 (also known as stromal derived factor 1, SDF1) is a chemokine that acts through the G protein‐coupled receptor CXCR4. Global deficiency of Cxcl12 or Cxcr4 as well as endothelial deletion of Cxcr4 result in vascular defects in the mouse kidney and gut, but these defects do not explain the associated late gestation lethality. We hypothesized that coronary vascular defects might underlie the embryonic lethality of CXCL12 deficiency.Cxcl12 and Tie2Cre/Cxcr4 mutants form an initial coronary vascular plexus, which then fails to remodel into a hierarchical network of mature vessels, resulting in the absence of the major intramyocardial coronary arteries, hemorrhage and reduced ventricular wall perfusion. The coronary veins were mostly unaffected, and the basis of the selectivity of CXCL12 on the arterial side of the vasculature is still unclear.In vitro, CXCL12 does not induce sprouting of endothelial cells from beads embedded in a fibrin gel matrix, but instead promotes maturation of vascular sprouts preformed in response to VEGF, by increasing tube elongation, vessel branching, anastomosis and intussusception, all features of mature blood vessel networks. These features were abolished in the presence of AMD3100, a CXCR4 inhibitor. Thus, CXCL12 does not initiate vasculogenesis but rather promotes further remodeling of a plexus that is initially induced by VEGF.We conclude that failed maturation of the coronary system explains the late‐gestation lethality of Cxcl12 mutants, and highlight the role of CXCL12 as an organ‐specific arterial maturation factor of particular relevance in the coronary vasculature. Our current studies are aimed at defining the role of CXCL12 in reprogramming of venous endothelial cells from the sinus venosus into coronary arterial endothelium.Support or Funding InformationThis work was supported by NIH grant HL070123 to H.M.S. and grant 14BGIA20500059 to S.R.K. from the American Heart Association. S.C. was supported by a Postdoctoral Fellowship from the American Heart Association. H.S. was supported by a Postdoctoral Fellowship from the California Institute for Regenerative Medicine.

CpG island shore methylation of ZFPM2 is identified in tetralogy of fallot samples.

ZFPM2 gene plays an important role in heart morphogenesis and development of coronary vessels from epicardium, however, little is known regarding its epigenetic regulation in the pathogenesis of tetralogy of fallot (TOF). The methylation levels of ZFPM2 gene were measured by MassArray (Sequenom, San Diego, CA) and bisulfite sequencing polymerase chain reaction (PCR) (BSP). Real-time PCR was performed to analyze the mRNA levels for ZFPM2 gene in the myocardium of TOF. The methylation levels in the CpG island shore of ZFPM2 promoter were significantly higher in patients with TOF, with a median of 80.32% (interquartile range (IQR): 73.54-85.75%, N = 42), as compared to 59.63% in controls (IQR: 44.79-73.83%; P = 0.0186, N = 6). No significant difference was observed in the methylation status at the CpG island of ZFPM2 promoter. The ZFPM2 mRNA levels were significantly lower in patients with TOF compared to that in the controls (P < 0.05). The aberrant methylation values of ZFPM2 were negatively associated with significant changes in its mRNA level (r = -0.40, P = 0.008, N = 42). Aberrant methylation status at the promoter CpG island shore of ZFPM2 gene may be associated with its gene transcription regulation in the TOF patients.

Congenital coronary artery anomalies: a bridge from embryology to anatomy and pathophysiology—a position statement of the development, anatomy, and pathology ESC Working Group

Congenital coronary artery anomalies are of major significance in clinical cardiology and cardiac surgery due to their association with myocardial ischaemia and sudden death. Such anomalies are detectable by imaging modalities and, according to various definitions, their prevalence ranges from 0.21 to 5.79%. This consensus document from the Development, Anatomy and Pathology Working Group of the European Society of Cardiology aims to provide: (i) a definition of normality that refers to essential anatomical and embryological features of coronary vessels, based on the integrated analysis of studies of normal and abnormal coronary embryogenesis and pathophysiology; (ii) an animal model-based systematic survey of the molecular and cellular mechanisms that regulate coronary blood vessel development; (iii) an organization of the wide spectrum of coronary artery anomalies, according to a comprehensive anatomical and embryological classification scheme; (iv) current knowledge of the pathophysiological mechanisms underlying symptoms and signs of coronary artery anomalies, with diagnostic and therapeutic implications. This document identifies the mosaic-like embryonic development of the coronary vascular system, as coronary cell types differentiate from multiple cell sources through an intricate network of molecular signals and haemodynamic cues, as the necessary framework for understanding the complex spectrum of coronary artery anomalies observed in human patients.

Vinculin b deficiency causes epicardial hyperplasia and coronary vessel disorganization in zebrafish.

Coronary vessel development is a highly coordinated process during heart formation. Abnormal development and dysfunction of the coronary network are contributory factors in the majority of heart disease. Understanding the molecular mechanisms that regulate coronary vessel formation is crucial for preventing and treating the disease. We report a zebrafish gene-trap vinculin b (vclb) mutant that displays abnormal coronary vessel development among multiple cardiac defects. The mutant shows overproliferation of epicardium-derived cells and disorganization of coronary vessels, and they eventually die off at juvenile stages. Mechanistically, Vclb deficiency results in the release of another cytoskeletal protein, paxillin, from the Vclb complex and the upregulation of ERK and FAK phosphorylation in epicardium and endocardium, causing disorganization of endothelial cells and pericytes during coronary vessel development. By contrast, cardiac muscle development is relatively normal, probably owing to redundancy with Vcla, a vinculin paralog that is expressed in the myocardium but not epicardium. Together, our results reveal a previously unappreciated function of vinculin in epicardium and endocardium and reinforce the notion that well-balanced FAK activity is essential for coronary vessel development.

Sequential Notch activation regulates ventricular chamber development.

Ventricular chambers are essential for the rhythmic contraction and relaxation occurring in every heartbeat throughout life. Congenital abnormalities in ventricular chamber formation cause severe human heart defects. How the early trabecular meshwork of myocardial fibres forms and subsequently develops into mature chambers is poorly understood. We show that Notch signalling first connects chamber endocardium and myocardium to sustain trabeculation, and later coordinates ventricular patterning and compaction with coronary vessel development to generate the mature chamber, through a temporal sequence of ligand signalling determined by the glycosyltransferase manic fringe (MFng). Early endocardial expression of MFng promotes Dll4-Notch1 signalling, which induces trabeculation in the developing ventricle. Ventricular maturation and compaction require MFng and Dll4 downregulation in the endocardium, which allows myocardial Jag1 and Jag2 signalling to Notch1 in this tissue. Perturbation of this signalling equilibrium severely disrupts heart chamber formation. Our results open a new research avenue into the pathogenesis of cardiomyopathies.