Molecular mechanisms underlying alcohol-induced congenital heart defects

About This Grant

PROJECT SUMMARY In the United States, up to 1 in 20 school age children have a diagnosable fetal alcohol spectrum disorder (FASD), which manifests in diverse phenotypes such as cognitive and behavioral deficits, craniofacial anomalies, and organ malformations. Importantly, up to 30% of patients with FASDs are thought to have a congenital heart defect (CHD). Despite the significant impacts of PAE on the developing cardiovascular system, the molecular and cellular mechanisms underlying EtOH-induced CHDs remain elusive along with any biomarkers that could be used to stratify individuals with FASDs by cardiovascular disease risk. In a zebrafish model of embryonic alcohol exposure (EAE), we confirmed a defect in cardiomyocyte (CM) and anterior endoderm (AE) migration that causes abnormal cardiac cone and heart tube development, resulting in ventricular structural abnormalities that persist through adulthood. Inhibition of PDGFRA and PI3K signaling through pharmacologic and genetic strategies sensitizes embryos to EtOH-induced defects in CM midline migration, suggesting a role for these pathways in failed cardiac cone fusion and downstream CHD formation. Following EAE, adults develop signs of progressive cardiomyopathy and diastolic dysfunction, which are worse in individuals with a documented CHD. RNA-sequencing of adult male and female EAE ventricles reveal consistently dysregulated gene expression, including a downregulation of gene candidates ankrd9, eya4, and slc25a33, which may contribute to the progressive cardiac dysfunction observed in EAE adults. In Specific Aim 1, we will test the hypothesis that abnormal cell migration and altered PI3K signaling in CM and AE cell populations causes failed cardiac cone fusion and CHDs. We will utilize a zebrafish model of EAE to identify changes in CMs and AE through advanced live imaging, single cell RNAseq, spatial transcriptomics, and an evaluation of PDGFRA and PI3K signaling. In Specific Aim 2, we will test the hypothesis that newly identified cardiac-expressed biomarker candidates ankrd9, eya4, and slc25a33 can modulate CHD and adult cardiomyopathy risk in individuals with FASDs. Loss of function mutants generated through CRISPR-Cas9 will be assessed for embryonic and adult cardiovascular phenotypes in the presence and absence of EAE through ISH, ICH, live imaging, and echocardiography. Taken together, the proposed studies will provide fundamental insights into the molecular mediators of CHDs and adult cardiac dysfunction following EAE.