Origin of congenital coronary arterio-ventricular fistulae from anomalous epicardial and myocardial development

Coronary Artery Fistulae (CAFs) are cardiac congenital anomalies consisting of an abnormal communication of a coronary artery with either a cardiac chamber or another cardiac vessel. In humans, these congenital anomalies can lead to complications such as myocardial hypertrophy, endocarditis, heart dilatation and failure. Unfortunately, despite their clinical relevance, the aetiology of CAFs remains unknown. In this work, we have used two different species (mouse and avian embryos) to experimentally model CAFs morphogenesis. Both conditional Itga4 (alpha 4 integrin) epicardial deletion in mice and cryocauterisation of chick embryonic hearts disrupted epicardial development and ventricular wall growth, two essential events in coronary embryogenesis. Additional transcriptomics and in vitro analyses were performed to better understand how arterio-ventricular connections are originated in the embryonic heart. Our results suggest myocardial discontinuities in the developing heart promote the formation of endocardial pouch-like structures resembling human CAF. The structure of these CAF-like anomalies was compared with histopathological data from a paediatric heart CAF, showing histomorphological and immunochemical similarities, including an accumulation of smooth muscle positive cells in the pouch-like structure wall. In vitro experiments showed the abnormal contact between the epicardium and the endocardium may promote the precocious differentiation of epicardial cells to smooth muscle. Our results suggest that myocardial discontinuities in the embryonic ventricular wall promote the early contact of the endocardium with epicardial-derived coronary progenitors at the cardiac surface, leading to ventricular endocardial extrusion, precocious differentiation of coronary smooth muscle cells, and the formation of pouch-like aberrant coronary-like structures in direct connection with the ventricular lumen. Our results may provide relevant information for the early diagnosis of these congenital anomalies and the molecular mechanisms that regulate their embryogenesis.

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Coronary smooth muscle reactivity to muscarinic stimulation after ischemia-reperfusion in porcine myocardial infarction

Journal of Applied Physiology ◽

10.1152/japplphysiol.00119.2003 ◽

2003 ◽

Vol 95 (1) ◽

pp. 81-88 ◽

Cited By ~ 7

Author(s):

Antonio Rodríguez-Sinovas ◽

Josep Bis ◽

Inocencio Anivarro ◽

Javier de la Torre ◽

Antoni Bayés-Genís ◽

...

Keyword(s):

Blood Flow ◽

Smooth Muscle ◽

Coronary Artery ◽

Coronary Blood Flow ◽

Ischemia Reperfusion ◽

Left Ventricular ◽

Severe Left Ventricular Dysfunction ◽

Coronary Smooth Muscle

This study tested whether ischemia-reperfusion alters coronary smooth muscle reactivity to vasoconstrictor stimuli such as those elicited by an adventitial stimulation with methacholine. In vitro studies were performed to assess the reactivity of endothelium-denuded infarct-related coronary arteries to methacholine ( n = 18). In addition, the vasoconstrictor effects of adventitial application of methacholine to left anterior descending (LAD) coronary artery was assessed in vivo in pigs submitted to 2 h of LAD occlusion followed by reperfusion ( n = 12), LAD deendothelization ( n = 11), or a sham operation ( n = 6). Endothelial-dependent vasodilator capacity of infarct-related LAD was assessed by intracoronary injection of bradykinin ( n = 13). In vitro, smooth muscle reactivity to methacholine was unaffected by ischemia-reperfusion. In vivo, baseline methacholine administration induced a transient and reversible drop in coronary blood flow (9.6 ± 4.6 to 1.9 ± 2.6 ml/min, P < 0.01), accompanied by severe left ventricular dysfunction. After ischemia-reperfusion, methacholine induced a prolonged and severe coronary blood flow drop (9.7 ± 7.0 to 3.4 ± 3.9 ml/min), with a significant delay in recovery ( P < 0.001). Endothelial denudation mimics in part the effects of methacholine after ischemia-reperfusion, and intracoronary bradykinin confirmed the existence of endothelial dysfunction. Infarct-related epicardial coronary artery shows a delayed recovery after vasoconstrictor stimuli, because of appropriate smooth muscle reactivity and impairment of endothelial-dependent vasodilator capacity.

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Epicardial Cells of Human Adults Can Undergo an Epithelial-to-Mesenchymal Transition and Obtain Characteristics of Smooth Muscle Cells In Vitro

Stem Cells ◽

10.1634/stemcells.2006-0366 ◽

2007 ◽

Vol 25 (2) ◽

pp. 271-278 ◽

Cited By ~ 122

Author(s):

John van Tuyn ◽

Douwe E. Atsma ◽

Elizabeth M. Winter ◽

Ietje van der Velde-van Dijke ◽

Daniel A. Pijnappels ◽

...

Keyword(s):

Smooth Muscle ◽

Smooth Muscle Cells ◽

Epithelial To Mesenchymal Transition ◽

Muscle Cells ◽

Mesenchymal Transition ◽

Epicardial Cells ◽

Human Adults

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Effect of Paclitaxel+Hirudin on the TLR4-MyD88 Signaling Pathway During Inflammatory Activation of Human Coronary Artery Smooth Muscle Cells and Mechanistic Analysis

Cellular Physiology and Biochemistry ◽

10.1159/000494588 ◽

2018 ◽

Vol 50 (4) ◽

pp. 1301-1317 ◽

Cited By ~ 1

Author(s):

Hongmei Li ◽

Xian Wang ◽

Anlong Xu

Keyword(s):

Smooth Muscle ◽

Coronary Artery ◽

Smooth Muscle Cells ◽

Muscle Cells ◽

Dependent Manner ◽

Human Coronary Artery ◽

Post Intervention ◽

Inflammatory Activation ◽

Myd88 Pathway

Background/Aims: Approximately 10%-20% of patients with acute cardiovascular disease who have received coronary intervention suffer restenosis and high inflammation. The stent compound paclitaxel+hirudin was prepared for the treatment of post-intervention restenosis. This study aimed to explore the anti-inflammatory and anti-restenosis mechanisms of paclitaxel+hirudin with regard to the TLR4/MyD88/NF-κB pathway. Methods: Human coronary artery smooth muscle cells (HCASMCs) at 4-6 generations after in vitro culture were used as a model. Lipopolysaccharide (LPS) was used as an inducer to maximally activate the TLR4/MyD88/NF-κB inflammation pathway. After MyD88 knockdown and selective blocking of MyD88 degradation with epoxomicin, the effects of paclitaxel+hirudin stenting on key sites of the TLR4/MyD88/NF-κB pathway were detected using ELISA, Q-PCR, and western blot analysis. Results: LPS at 1 μg/mL for 48 h was the optimal modeling condition for inflammatory activation of HCASMCs. Paclitaxel+hirudin inhibited the levels of key proteins and the gene expression, except for that of the MyD88 gene, of the TLR4-MyD88 pathway. The trend of the effect of paclitaxel+hirudin on the pathway proteins was similar to that of MyD88 knockdown. After epoxomicin intervention, the inhibitory effects of paclitaxel+hirudin on the key genes and proteins of the TLR4-MyD88 pathway were significantly weakened, which even reached pre-intervention levels. Paclitaxel+hirudin affected the MyD88 protein in a dosage-dependent manner. Conclusion: The paclitaxel+hirudin compound promotes MyD88 degradation in the TLR4/MyD88/NF-κB pathway to reduce the activity of TLR4 and NF-κB p65 and to weaken the LPS-initiated inflammatory reactions of IL-1β, IL-6, and TNF-α.

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Extracellular calcium-dependent and -independent effects of adenosine in bovine coronary artery

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y90-088 ◽

1990 ◽

Vol 68 (5) ◽

pp. 608-613 ◽

Cited By ~ 5

Author(s):

Mudumbi V. Ramagopal ◽

S. Jamal Mustafa

Keyword(s):

Smooth Muscle ◽

Coronary Artery ◽

Coronary Arteries ◽

Extracellular Calcium ◽

Free Medium ◽

Response Curves ◽

Coronary Smooth Muscle ◽

Calcium Dependent ◽

Relaxation Responses ◽

The Right

Adenosine relaxes the coronary arteries of various species through A2 receptors. The aim of the present investigation was to evaluate the relaxing effects of adenosine in relation to the role of calcium in bovine coronary arteries by studying the vasodilatory effect of adenosine in normal and calcium-free medium and on calcium-45 efflux into calcium-free medium. Acetylcholine (ACh) and norepinephrine (NE) were used to induce tone in coronary artery rings. Adenosine, 5′-(N-ethylcarboxamido)adenosine (NECA), and N6-(L-phenylisopropyl)adenosine (L-PIA) produced concentration-dependent relaxations of the coronary artery rings. Both in normal and calcium-free medium, the order of potency for adenosine analogs (NECA > L-PIA > adenosine) was similar and 8-phenyltheophylline antagonized the relaxation responses to adenosine and its analogs. Removal of extracellular calcium shifted the concentration–response curves to the right in a parallel fashion, slowed the rate of relaxation, and in NE contracted rings reduced the maximum responses for adenosine and its analogs. In calcium-free medium, adenosine was without an effect on calcium-45 efflux in the presence of ACh. However, adenosine inhibited the stimulated calcium-45 efflux induced by NE. The data suggest that the vasodilatory action of adenosine in bovine coronary smooth muscle has both extracellular calcium-dependent and -independent components.Key words: adenosine receptors, calcium, coronary circulation, vascular smooth muscle, acetylcholine, norepinephrine.

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Effects of zotarolimus, sirolimus, dexamethasone, and paclitaxel on the in vitro migration of human coronary artery smooth muscle and endothelial cells

Cardiovascular Revascularization Medicine ◽

10.1016/j.carrev.2006.03.083 ◽

2006 ◽

Vol 7 (2) ◽

pp. 119

Author(s):

M.M. Mack ◽

Y. Xiao ◽

S.E. Burke

Keyword(s):

Endothelial Cells ◽

Smooth Muscle ◽

Coronary Artery ◽

Human Coronary Artery

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GW26-e1243 Inhibitory Effects of Hirudin on the Growth and Proliferation of Human Coronary Artery Smooth Muscle Cells and Endothelial Cells in Vitro

Journal of the American College of Cardiology ◽

10.1016/j.jacc.2015.06.408 ◽

2015 ◽

Vol 66 (16) ◽

pp. C105

Author(s):

Hongmei Li ◽

Mengqiong Sun ◽

Ang Gao ◽

Zhen Wang ◽

Xueqing Yang ◽

...

Keyword(s):

Endothelial Cells ◽

Smooth Muscle ◽

Coronary Artery ◽

Smooth Muscle Cells ◽

Muscle Cells ◽

Inhibitory Effects ◽

Human Coronary Artery

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Ca2+ antagonist-insensitive coronary smooth muscle contraction involves activation of ϵ-protein kinase C-dependent pathway

AJP Cell Physiology ◽

10.1152/ajpcell.00066.2003 ◽

2003 ◽

Vol 285 (6) ◽

pp. C1454-C1463 ◽

Cited By ~ 19

Author(s):

Andrea Dallas ◽

Raouf A. Khalil

Keyword(s):

Smooth Muscle ◽

Coronary Artery ◽

Muscle Contraction ◽

Smooth Muscle Contraction ◽

Channel Blockers ◽

Cell Contraction ◽

Coronary Smooth Muscle ◽

Calphostin C ◽

Gf 109203X ◽

Dependent Pathway

Certain angina and coronary artery disease forms do not respond to Ca2+ channel blockers, and a role for vasoactive eicosanoids such as PGF2α in Ca2+ antagonist-insensitive coronary vasospasm is suggested; however, the signaling mechanisms are unclear. We investigated whether PGF2α-induced coronary smooth muscle contraction is Ca2+ antagonist insensitive and involves activation of a PKC-dependent pathway. We measured contraction in single porcine coronary artery smooth muscle cells and intracellular free Ca2+ concentration ([Ca2+]i) in fura 2-loaded cells and examined cytosolic and particulate fractions for PKC activity and reactivity with isoform-specific PKC antibodies. In Hanks' solution (1 mM Ca2+), PGF2α (10-5 M) caused transient [Ca2+]i increase followed by maintained [Ca2+]i increase and 34% cell contraction. Ca2+ channel blockers verapamil and diltiazem (10-6 M) abolished maintained PGF2α-induced [Ca2+]i increase but only partially inhibited PGF2α-induced cell contraction to 17%. Verapamil-insensitive PGF2α contraction was inhibited by PKC inhibitors GF-109203X, calphostin C, and ϵ-PKC V1-2. PGF2α caused Ca2+-dependent α-PKC and Ca2+-independent ϵ-PKC translocation from cytosolic to particulate fractions that was inhibited by calphostin C. Verapamil abolished PGF2α-induced α-but not ϵ-PKC translocation. PMA (10-6 M), a direct activator of PKC, caused 21% contraction with no significant [Ca2+]i increase and ϵ-PKC translocation that were inhibited by calphostin C but not verapamil. Membrane depolarization by 51 mM KCl, which stimulates Ca2+ influx, caused 36% cell contraction and [Ca2+]i increase that were inhibited by verapamil but not GF-109203X or calphostin C and did not cause α- or ϵ-PKC translocation. Thus a significant component of PGF2α-induced contraction of coronary smooth muscle is Ca2+ antagonist insensitive, involves Ca2+-independent ϵ-PKC activation and translocation, and may represent a signaling mechanism of Ca2+ antagonist-resistant coronary vasospasm.

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Sprouty1 Controls Genitourinary Development via its N-Terminal Tyrosine

Journal of the American Society of Nephrology ◽

10.1681/asn.2018111085 ◽

2019 ◽

Vol 30 (8) ◽

pp. 1398-1411

Author(s):

Marta Vaquero ◽

Sara Cuesta ◽

Carlos Anerillas ◽

Gisela Altés ◽

Joan Ribera ◽

...

Keyword(s):

Urinary Tract ◽

Knockout Mice ◽

Congenital Anomalies ◽

Molecular Mechanisms ◽

Structural Domains ◽

Reporter Mice ◽

Ectopic Overexpression ◽

Whole Mount ◽

Ureteric Buds

BackgroundStudies in mice suggest that perturbations of the GDNF-Ret signaling pathway are a major genetic cause of congenital anomalies of the kidney and urinary tract (CAKUT). Mutations in Sprouty1, an intracellular Ret inhibitor, results in supernumerary kidneys, megaureters, and hydronephrosis in mice. But the underlying molecular mechanisms involved and which structural domains are essential for Sprouty1 function are a matter of controversy, partly because studies have so far relied on ectopic overexpression of the gene in cell lines. A conserved N-terminal tyrosine has been frequently, but not always, identified as critical for the function of Sprouty1 in vitro.MethodsWe generated Sprouty1 knockin mice bearing a tyrosine-to-alanine substitution in position 53, corresponding to the conserved N-terminal tyrosine of Sprouty1. We characterized the development of the genitourinary systems in these mice via different methods, including the use of reporter mice expressing EGFP from the Ret locus, and whole-mount cytokeratin staining.ResultsMice lacking this tyrosine grow ectopic ureteric buds that will ultimately form supernumerary kidneys, a phenotype indistinguishable to that of Sprouty1 knockout mice. Sprouty1 knockin mice also present megaureters and vesicoureteral reflux, caused by failure of ureters to separate from Wolffian ducts and migrate to their definitive position.ConclusionsTyrosine 53 is absolutely necessary for Sprouty1 function during genitourinary development in mice.

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Glucocorticoid and TNF signaling converge at A20 (TNFAIP3) to repress airway smooth muscle cytokine expression

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00179.2016 ◽

2016 ◽

Vol 311 (2) ◽

pp. L421-L432 ◽

Cited By ~ 17

Author(s):

Sarah K. Sasse ◽

Mohammed O. Altonsy ◽

Vineela Kadiyala ◽

Gaoyuan Cao ◽

Reynold A. Panettieri ◽

...

Keyword(s):

Smooth Muscle ◽

Airway Smooth Muscle ◽

Molecular Mechanisms ◽

Nuclear Factor Κb ◽

Airway Epithelial Cells ◽

Therapeutic Effects ◽

Target Tissue ◽

Airway Epithelial ◽

Fatal Asthma

Airway smooth muscle is a major target tissue for glucocorticoid (GC)-based asthma therapies, however, molecular mechanisms through which the GC receptor (GR) exerts therapeutic effects in this key airway cell type have not been fully elucidated. We previously identified the nuclear factor-κB (NF-κB) inhibitor, A20 (TNFAIP3), as a mediator of cytokine repression by glucocorticoids (GCs) in airway epithelial cells and defined cooperative regulation of anti-inflammatory genes by GR and NF-κB as a key mechanistic underpinning of airway epithelial GR function. Here, we expand on these findings to determine whether a similar mechanism is operational in human airway smooth muscle (HASM). Using HASM cells derived from normal and fatal asthma samples as an in vitro model, we demonstrate that GCs spare or augment TNF-mediated induction of A20 ( TNFAIP3), TNIP1, and NFKBIA, all implicated in negative feedback control of NF-κB-driven inflammatory processes. We applied chromatin immunoprecipitation and reporter analysis to show that GR and NF-κB directly regulate A20 expression in HASM through cooperative induction of an intronic enhancer. Using overexpression, we show for the first time that A20 and its interacting partner, TNIP1, repress TNF signaling in HASM cells. Moreover, we applied small interfering RNA-based gene knockdown to demonstrate that A20 is required for maximal cytokine repression by GCs in HASM. Taken together, our data suggest that inductive regulation of A20 by GR and NF-κB contributes to cytokine repression in HASM.

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