Single-nucleus multiomic landscape of congenital heart diseases reveals disease-specific genotypic profiles

Tetralogy of Fallot (TOF) is the most common cyanotic congenital heart defect (CHD), whereas hypoplastic left heart syndrome (HLHS) represents 2-3% of all CHDs. We analyzed the single-nucleus multiome profile of right ventricle samples obtained from children during routine cardiac surgery for correction of TOF or staged surgical palliation for HLHS to define cell types and characterize gene regulation states in different cell clusters. Data were integrated with pre-existing controls and analyzed using Scanpy to identify clusters and annotated using automated tool and manual curation, pycisTopic to identify cell stats and cis-regulatory topics, and SCENIC+ for enhancer-driven gene regulatory network (eRegulon). Integrated RNA-seq analysis identified 22 different cell subtypes, including five cardiomyocyte phenotypes. TOF samples showed involvement in pathway networks of the cell cycle, DNA repair, DNA replication, and RNA metabolism, whereas gene expression in HLHS samples was related to extracellular matrix organization, anatomical structure development, cell adhesion, actin-myosin filament sliding, and contractile muscle fiber pathways. In addition, the gene expression fingerprints of endothelial, fibroblast, pericyte, immune, and neuronal cell nuclei from TOF and HLHS samples exhibited nuclei-specific significant de-regulation compared to controls. We found considerable heterogeneity among the transcriptomes of TOF and HLHS, explaining the diverse clinical phenotypes. These findings can enable the development of new gene-based interventions for specific CHDs.