The secondary heart field is a new site of calcineurin/Nfatc1 signaling for semilunar valve development

Division of Cardiovascular Medicine, Department of Medicine, Stanford Cardiovascular Institute, Stanford University, Stanford, California 94305, USA.
Journal of Molecular and Cellular Cardiology (Impact Factor: 4.66). 01/2012; 52(5):1096-102. DOI: 10.1016/j.yjmcc.2012.01.013
Source: PubMed


Semilunar valve malformations are common human congenital heart defects. Bicuspid aortic valves occur in 2-3% of the population, and pulmonic valve stenosis constitutes 10% of all congenital heart disease in adults (Brickner et al., 2000) [1]. Semilunar valve defects cause valve regurgitation, stenosis, or calcification, leading to endocarditis or congestive heart failure. These complications often require prolonged medical treatment or surgical intervention. Despite the medical importance of valve disease, the regulatory pathways governing semilunar valve development are not entirely clear. In this report we investigated the spatiotemporal role of calcineurin/Nfatc1 signaling in semilunar valve development. We generated conditional knockout mice with calcineurin gene disrupted in various tissues during semilunar valve development. Our studies showed that calcineurin/Nfatc1 pathway signals in the secondary heart field (SHF) but not in the outflow tract myocardium or neural crest cells to regulate semilunar valve morphogenesis. Without SHF calcineurin/Nfatc1 signaling, the conal endocardial cushions-the site of prospective semilunar valve formation--first develop and then regress due to apoptosis, resulting in a striking phenotype with complete absence of the aortic and pulmonic valves, severe valve regurgitation, and perinatal lethality. This role of calcineurin/Nfatc1 signaling in the SHF is different from the requirement of calcineurin/Nfatc1 in the endocardium for semilunar valve formation (Chang et al., 2004) [2], indicating that calcineurin/Nfatc1 signals in multiple tissues to organize semilunar valve development. Also, our studies suggest distinct mechanisms of calcineurin/Nfat signaling for semilunar and atrioventricular valve morphogenesis. Therefore, we demonstrate a novel developmental mechanism in which calcineurin signals through Nfatc1 in the secondary heart field to promote semilunar valve morphogenesis, revealing a new supportive role of the secondary heart field for semilunar valve formation.

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    • "Furthermore, it has been demonstrated that vascular endothelial growth factor (VEGF)-mediated, calcineurinactivated NFATc-1 regulates endothelial cell fate and maintains a valvular endothelial cell (VEC) phenotype (Johnson et al., 2003). Moreover, previous reports postulated that the spatio-temporally distinct function of NFATc-1 signaling in SHF cells and endocardial cells is necessary for semilunar cushion formation as well as leaflet elongation and maturation in rodents (Chang et al., 2004; Lin et al., 2012; Wu et al., 2011). "
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    ABSTRACT: Elucidation of mechanisms in semilunar valve development may enable the development of new therapies. Here, we found differences in proliferation-associated genes and genes repressed by vascular endothelial growth factor between human semilunar valves from first and second trimester valve leaflets. The proliferation of valve interstitial cells and ventricular valve endothelial cells (VECs) and cellular density declined from the first to the second trimester. Cytoplasmic expression of nuclear factor of activated T-cells cytoplasmic 1 (NFATc-1) in VECs (4 weeks), and later cells in the leaflet/annulus junction mesenchyme expressing inactive NFATc-1 (5.5-9 weeks) were detected, indicative of EndMT in valvulogenesis. At this leaflet/annulus junction CD44+ cells clustered during elongation (11 weeks), extending toward the tip along the fibrosal layer in second trimester leaflets. Differing patterns of maturation in the fibrosa and ventricularis were detected via increased fibrosal periostin content, which tracked the presence of the CD44+ cells in the second trimester. We revealed that spatiotemporal NFATc-1 expression actively regulates EndMT during human valvulogenesis, as early as 4 weeks. Additionally, CD44+ cells play a role in leaflet maturation toward the trilaminar structure, possibly via migration of VECs undergoing EndMT, which subsequently ascend from the leaflet/annulus junction.
    Full-text · Article · Dec 2015 · Development
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    • "These results suggest that post- EMT remodeling of the valves derived from the outflow tract cushion may be more affected by the loss of Cad-11, as the valves that developed from the misaligned atrioventricular cushions were able to recover and develop properly. Additionally, there are post-EMT remodeling contributions from multiple cell types in the outflow tract that are absent from the atrioventricular tract, such as neural crest and anterior heart field mesenchyme (Lin et al., 2012), which may also be affected by loss of Cad-11. To assess the consequence of the dysmorphic aortic valve on cardiac function, Doppler in utero echocardiography was performed on the aortic valve of E17.5 embryos. "
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    ABSTRACT: Proper remodeling of the endocardial cushions into thin fibrous valves is essential for gestational progression and long-term function. This process involves dynamic interactions between resident cells and their local environment, much of which is not understood. In this study, we show that deficiency of the cell-cell adhesion protein cadherin-11 (cad-11) results in significant embryonic and perinatal lethality primarily due to valve related cardiac dysfunction. While endocardial to mesenchymal transformation is not abrogated, mesenchymal cells do not homogeneously cellularize the cushions. These cushions remain thickened with disorganized ECM, resulting in pronounced aortic valve insufficiency. Mice that survive to adulthood maintain thickened and stenotic semilunar valves, but interestingly do not develop calcification. Cad-11 -/- aortic valve leaflets contained reduced sox9 activity, β1 integrin expression, and RhoA-GTP activity, suggesting that remodeling defects are due to improper migration and/or cellular contraction. Cad-11 deletion or siRNA knockdown reduced migration, eliminated collective migration, and impaired 3D matrix compaction by aortic valve interstitial cells (VIC). Cad-11 depleted cells in culture contained few filopodia, stress fibers, or contact inhibited locomotion. Transfection of Cad-11 depleted cells with constitutively active RhoA restored cell phenotypes. Together, these results identify cadherin-11 mediated adhesive signaling for proper remodeling of the embryonic semilunar valves. Copyright © 2015. Published by Elsevier Inc.
    Full-text · Article · Jul 2015 · Developmental Biology
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    • "1 Dup 192A HLHS 18 75333697 75370277 36581 Del N NFATC1 a ( Lin et al . , 2012"
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    ABSTRACT: Background: We sought to characterize the landscape of structural variation associated with the subset of congenital cardiac defects characterized by left-sided obstruction. Methods: Cases with left-sided cardiac defects (LSCD) and pediatric controls were uniformly genotyped and assessed for copy number variant (CNV) calls. Significance testing was performed to ascertain differences in overall CNV incidence, and for CNV enrichment of specific genes and gene functions in LSCD cases relative to controls. Results: A total of 257 cases of European descent and 962 ethnically matched, disease-free pediatric controls were included. Although there was no difference in CNV rate between cases and controls, a significant enrichment in rare LSCD CNVs was detected overall (p=7.30 × 10(-3) , case/control ratio=1.26) and when restricted either to deletions (p=7.58 × 10(-3) , case/control ratio=1.20) or duplications (3.02 × 10(-3) , case/control ratio=1.43). Neither gene-based, functional nor knowledge-based analyses identified genes, loci or pathways that were significantly enriched in cases as compared to controls when appropriate corrections for multiple tests were applied. However, several genes of interest were identified by virtue of their association with cardiac development, known human conditions, or reported disruption by CNVs in other patient cohorts. Conclusion: This study examines the largest cohort to date with LSCD for structural variation. These data suggest that CNVs play a role in disease risk and identify numerous genes disrupted by CNVs of potential disease relevance. These findings further highlight the genetic heterogeneity and complexity of these disorders.
    Full-text · Article · Dec 2014 · Birth Defects Research Part A Clinical and Molecular Teratology
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