Defects in cardiac valve morphogenesis and septation of the heart chambers constitute some of the most common human congenital abnormalities. Some of these defects originate from errors in atrioventricular (AV) endocardial cushion development. Although this process is being extensively studied in mouse and chick, the zebrafish system presents several advantages over these models, including the ability to carry out forward genetic screens and study vertebrate gene function at the single cell level. In this paper, we analyze the cellular and subcellular architecture of the zebrafish heart during stages of AV cushion and valve development and gain an unprecedented level of resolution into this process. We find that endocardial cells in the AV canal differentiate morphologically before the onset of epithelial to mesenchymal transformation, thereby defining a previously unappreciated step during AV valve formation. We use a combination of novel transgenic lines and fluorescent immunohistochemistry to analyze further the role of various genetic (Notch and Calcineurin signaling) and epigenetic (heart function)pathways in this process. In addition, from a large-scale forward genetic screen we identified 55 mutants, defining 48 different genes, that exhibit defects in discrete stages of AV cushion development. This collection of mutants provides a unique set of tools to further our understanding of the genetic basis of cell behavior and differentiation during AV valve development.

Function of the heart begins long before its formation is complete. Analyses in mouse and zebrafish have shown that myocardial function is not required for early steps of organogenesis, such as formation of the heart tube or chamber specification. However, whether myocardial function is required for later steps of cardiac development, such as endocardial cushion (EC) formation, has not been established. Recent technical advances and approaches have provided novel inroads toward the study of organogenesis, allowing us to examine the effects of both genetic and pharmacological perturbations of myocardial function on EC formation in zebrafish. To address whether myocardial function is required for EC formation, we examined silent heart (sih−/−) embryos, which lack a heartbeat due to mutation of cardiac troponin T (tnnt2), and observed that atrioventricular (AV) ECs do not form. Likewise, we determined that cushion formation is blocked in cardiofunk (cfk−/−) embryos, which exhibit cardiac dilation and no early blood flow. In order to further analyze the heart defects in cfk−/− embryos, we positionally cloned cfk and show that it encodes a novel sarcomeric actin expressed in the embryonic myocardium. The Cfks11 variant exhibits a change in a universally conserved residue (R177H). We show that in yeast this mutation negatively affects actin polymerization. Because the lack of cushion formation in sih- and cfk-mutant embryos could be due to reduced myocardial function and/or lack of blood flow, we approached this question pharmacologically and provide evidence that reduction in myocardial function is primarily responsible for the defect in cushion development. Our data demonstrate that early myocardial function is required for later steps of organogenesis and suggest that myocardial function, not endothelial shear stress, is the major epigenetic factor controlling late heart development. Based on these observations, we postulate that defects in cardiac morphogenesis may be secondary to mutations affecting early myocardial function, and that, in humans, mutations affecting embryonic myocardial function may be responsible for structural congenital heart disease.

Improved safety and teamwork climate as measured by SAQ are associated with decreased patient harm and severity-adjusted mortality. Discrepancies in SAQ scores exist between different professional groups but decreased over time.

For years, biomechanical engineers have studied the physical forces involved in morphogenesis of the heart. In a parallel stream of research, molecular and developmental biologists have sought to identify the molecular pathways responsible for embryonic heart development. Recently, several studies have shown that these two avenues of research should be integrated to explain how genes expressed in the heart regulate early heart function and affect physical morphogenetic steps, as well as to conversely show how early heart function affects the expression of genes required for morphogenesis. This review combines the perspectives of biomechanical engineering and developmental biology to lay out an integrated view of the role of mechanical forces in heart development.

Integrating specific aspects of Down syndrome care into the EHR can improve adherence to guideline recommendations that span the life of a child. Future quality improvement should be focused on older children and adults with Down syndrome.

Malformations of the cardiac endocardial cushions (ECs) and valves are common congenital dysmorphisms in newborn infants. Many regulators of EC development have been identified, but the process of valve maturation is less well understood. Zebrafish have been used to understand cardiogenesis through 6 days postfertilization, yet mature heart valves are not present at this stage. By analyzing valve development in larval zebrafish, we identify that valve development proceeds in two phases. Valve elongation occurs through 16 dpf independently of localized cell division. Valve maturation then ensues, resulting from deposition of extracellular matrix and thickening of the valves. Whereas elongation is consistent between larvae, maturation varies based on larval size, suggesting that maturation occurs in response to mechanical forces. Taken together, our studies indicate that zebrafish valve morphogenesis occurs in the larval period, and that zebrafish may provide a unique opportunity to study epigenetic mechanisms leading to human congenital valvular disease, when studied at the appropriate developmental stages. Developmental Dynamics 238:1796 -1802, 2009.

Safety II behavior in this unit was based on strong Safety I behaviors adapted to the Safety II environment plus innovation behaviors specific to Safety II situations. We believe these behaviors can be taught and learned. We intend to spread these concepts throughout the organization.

CULLIN 2 (CUL2) is a component of the ElonginB/C-CUL2-RBX-1-Von Hippel-Lindau (VHL) tumor suppressor complex that ubiquitinates and degrades hypoxia-inducible factor ␣ (HIF␣). HIF␣ is a transcription factor that mediates the expression of hypoxia-sensitive genes, including vascular endothelial growth factor (VEGF), which in turn regulates vasculogenesis. Whereas CUL2 participates in the degradation of HIF␣, the potential role of CUL2 in the regulation of other cellular processes is less well established. In the present study, suppression of CUL2 expression by Cul2 siRNA inhibited HIF␣ transcriptional activation of the VEGF gene in vitro, indicating that CUL2 plays a role distinct from its known function in HIF␣ degradation. Because ARNT heterodimerizes with HIF␣, we assessed whether CUL2 influenced ARNT expression. Cul2 siRNA inhibited the expression of endogenous ARNT. Ectopically expressed ARNT reversed the inhibition of HIF activity by Cul2 siRNA in the VEGF promoter, suggesting that CUL2 regulates HIF activation through ARNT. In 786-O cells lacking VHL, Cul2 siRNA suppressed the expression of both ARNT and VEGF, indicating that CUL2 regulates HIF activity independently of VHL. In transgenic zebrafish expressing GFP driven by the Flk promoter (a known HIF target), zCul2 morpholino blocked embryonic vasculogenesis in a manner similar to that caused by inhibition of VEGF-A. In the zebrafish embryos, zCul2 inhibited the expression of CUL2, VEGF, and Flk-GFP protein, indicating that CUL2 is required for expression of other vasculogenic HIF targets. Taken together, CUL2 is required for normal vasculogenesis, at least in part mediated by its regulation of HIF-mediated transcription. CULLIN 2 (CUL2)2 is a member of the CULLIN family of ubiquitin ligases (1). CUL2 associates with the von HippelLindau tumor suppressor protein (VHL), transcriptional elongation factors Elongin B/C, RING-box protein RBX1, and E3 ubiquitin-protein ligase (2-5). VHL recognizes hydroxylated hypoxia-inducible factor ␣ (HIF␣), recruits the CUL2-associating complex on HIF␣, and ubiquitinates HIF␣ (6 -11). The ubiquitinated HIF␣ is destined for degradation. In the absence of CUL2 or RBX1, the HIF-2␣ protein is increased, indicating that CUL2 and RBX1 are involved in HIF␣ protein stabilization (12). CUL2, together with ElonginB/C and RBX1, also forms a complex with MED8, a mediator subunit of the RNA polymerase II transcriptional machinery (13), though the mechanism by which CUL2 influences transcriptional regulation is unknown. CUL2 and RBX1 have been predicted to function as tumor suppressor proteins because of their protein-protein interaction with VHL. Somatic mutations in VHL have been associated with tumors including hemangioblastomas, renal cell carcinoma, and pheochromocytoma (14). However, pathogenic mutations in the Cul2 or Rbx1 gene causing carcinoma have not been identified (14 -16), indicating that CUL2 or RBX1 may not be tumor suppressor proteins. In fact, CUL2 functions as a positive cell cycle regulator in Caenorhabditis elegans ...