Abstract Background Hypoplastic left heart syndrome (HLHS), a severe congenital heart disease, is associated with poor neurodevelopmental outcomes, microcephaly, reduced cortical brain volume, brain dysmaturation, and neurobehavioral disorders such as autism. The involvement of patient intrinsic factors was indicated, but the mechanism is largely unknown. Methods Ohia mice with HLHS causing mutations in chromatin modifier Sin3A-associated protein 130 ( Sap130 ) and cell adhesion protein ProtocadherinA9 ( Pcdha9) were investigated for brain abnormalities by histology, immunomicroscopy, and molecular profiling by RNAseq, Sap130 ChIPseq, and genome-wide methylome analysis. Additionally, adult viable Pcdha9 m/m and Emx1-cre:Sap130 f/− mice with forebrain deletion of Sap130 were examined by brain MRI and behavioral assessments. Results Ohia mice have brain abnormalities comprising forebrain hypoplasia and microcephaly in conjunction with a cortical neurogenesis defect. This is associated with loss of intermediate progenitors due to mitotic arrest and apoptosis from multipolar spindle formation, a mechanism also observed in primary microcephaly. Brain RNAseq showed perturbation of REST transcriptional regulation of neurogenesis, disruption of CREB signaling regulating synaptic plasticity and memory, and defects in neurovascular coupling indicating perturbation of brain-sparing cerebral autoregulation. Disease pathways recovered included autism, intellectual disability, and other neurobehavioral/neurological deficits. These same pathways were observed upon intersection of genes that are differentially expressed with those that are differentially methylated and also are ChIPseq targets of Sap130, suggesting the transcriptional changes are epigenetically regulated. Adult viable mice harboring either the Pcdha9 mutation or forebrain-specific Sap130 deletion showed similar learning/memory deficits and autism-like behavior, suggesting they act on convergent pathways. Conclusions Our observations indicate the intrinsic factors contributing to the adverse neurodevelopmental outcome associated with HLHS involve spindle defects causing impaired corticoneurogenesis, and brain and behavioral deficits associated with perturbed epigenetic regulation of neurodevelopmental pathways. Novelty and Significance What is known? Hypoplastic left heart syndrome (HLHS), a severe congenital heart disease, is associated with adverse neurodevelopmental outcome attributable to patient intrinsic factors. Cortical neurogenesis defect with reduced brain volume and microcephaly are observed beginning in utero, suggesting a developmental etiology. Learning impairment and autism spectrum disorder are commonly observed in HLHS. What new information does this article contribute? The Ohia HLHS mouse model exhibits neurodevelopmental deficits comprising microcephaly and cortical neurogenesis defects with loss of neural progenitors from multipolar spindle formation, as well as impaired neurovascular coupling. Molecular profiling showed disturbance of REST, transcriptional regulator of neural stem cells, and CREB signaling regulating synaptic plasticity, with neurobehavioral assessments of the mutant mice showing learning/memory and autism-like behavioral deficits. Intersection of transcriptome and DNA methylation analyses uncovered an epigenetic basis for the neurodevelopmental/neurobehavioral abnormalities, Analysis of an HLHS mouse model indicated patient intrinsic factors causing adverse neurodevelopment in HLHS are genetic and epigenetic in etiology. This may include a mitotic spindle defect that would not be rescued by in utero aortic valvuloplasty, and a defect in neurovascular coupling that is likely to reduce the efficacy of maternal hyperoxygenation. However, epigenetic therapy may provide a new avenue for treatment that should be explored.

Pathogenic mechanisms underlying adverse neurodevelopmental outcome in congenital heart disease