Poster abstracts
Poster number 54 submitted by Adrianna P. Matos-Nieves
Understanding the role of Netrin signaling in the developing heart and congenital heart disease
Adrianna P. Matos-Nieves (Molecular, Cellular and Developmental Biology ), Uddalak Majumdar, Sathiyanarayanan Manivannan, Emily M. Cameron (Center for Cardiovascular Research and The Heart Center, Nationwide Childrens Hospital, Columbus, OH), Stephanie LaHaye (Institute for Genomic Medicine, Nationwide Childrens Hospital, Columbus, OH), Vidu Garg (Center for Cardiovascular Research and The Heart Center, Nationwide Childrens Hospital, Columbus, OH, Departments of Pediatrics and Molecular Genetics, The Ohio State University, Columbus, OH )
Abstract:
Malformations involving the cardiac outflow tract (OFT) and semilunar valves account for >50% of congenital heart disease (CHD) when including bicuspid aortic valve (BAV). We previously reported that endothelial cell-derived nitric oxide activates Notch1 signaling and Notch1+/-;Nos3-/- compound mutant mice are a model of cardiac OFT malformations as they display a spectrum of CHD including BAV, thickened semilunar valves, and tetralogy of Fallot-like phenotypes. In order to identify underlying genetic programs that govern the development of the OFT, transcriptomic profiling of E13.5 Notch1+/-;Nos3-/- OFT was performed and compared to wildtype littermate controls. We analyzed this data using Gene set enrichment analysis pipeline (SeqGSEA), with canonical signaling pathway gene sets. SeqGSEA analysis revealed the top disrupted pathways including known CHD-associated pathways such as Notch, TGF-beta, and Wnt signaling. In addition, axon guidance was among the top enriched pathways. While, Slit/Robo signaling has already been demonstrated to be critical for cardiac OFT development, the role of Netrin signaling has not been studied. Using qPCR, we validated the downregulation of Netrin-1 in the E13.5 Notch1+/-;Nos3-/- OFT and demonstrated its expression from E9.5-E13.5 in the developing mouse OFT along with its receptors Dcc, Neogenin and Unc5b. Single-cell RNA sequencing data derived from E6.5 – E14.5 embryonic mouse hearts suggest that Netrin-1, Neogenin and Unc5b co-localize with Tnnt2+ cells which serves as a marker for myocardial cells. Using a Netrin-1LacZ mutant mouse line, we have demonstrated that Netrin-1 is expressed in the E9.5 – E12.5 trabecular myocardium and the developing OFT cushions. Netrin-1-/- embryos in a C57BL/6J background display embryonic lethality by E10.5 however, the cause of lethality is yet unknown. Molecular analysis of Notch1+/-;Nos3-/- embryos has implicated Netrin signaling in aortic valve development and supports genetic investigations in humans with CHD.
Keywords: development, genetics, heart