Abstract Body: Introduction:
Congenital heart diseases (CHDs) are the most common birth defects in the U.S. There are growing numbers of infants and children living with single ventricle congenital heart disease (SVCHD), a unique congenital cardiac anomaly in which there is only one functional ventricle. The Fontan surgery is the current standard of care for SVCHD. Although operative survival has improved over the years, it is now evident that these surviving patients are facing new life-threatening challenges: severe complications resulting from the Fontan operation, including one of the most evident consequences-hepatic fibrosis, which is now recognized as Fontan-Associated Liver Disease (FALD). Unfortunately, the mechanisms underlying FALD remain little understood. Improved understanding in this knowledge gap will offer crucial insights into FALD pathophysiology that can lead to targeted therapies for SVCHD patients.

Goal:
To determine if liver metabolism changes significantly in FALD and compare these liver metabolomics changes to metabolic gene expression (our snRNA-ATAC-seq data) and reported plasma metabolomics results, and validate the key findings.

Methods:
We recently generated the first single-cell transcriptomic landscape of human FALD using snRNA-ATAC-seq, which revealed profound metabolic reprogramming in FALD. Here we utilized metabolomics to further understand this. Liver samples/biopsies were collected from healthy donors and FALD patients. The extracted metabolites were analyzed using C18 and HILIC HPLC/MS columns. Raw data files were acquired by high resolution MS on Orbitrap and processed by Compound Discoverer. Further statistical and bioinformatics analyses were performed to identify altered metabolites and potential metabolic pathways involved in FALD.

Results:
Significant changes of metabolite levels of multiple metabolic pathways were observed in Fontan livers, often consistent with the gene expression results. Overall, these data unveiled profound metabolic abnormalities in livers of patients with early-stage FALD.

Conclusions:
In conclusion, our investigation revealed alterations in metabolites involved in glycolysis/gluconeogenesis, mitochondrial electron transport chain, and amino acids in early-stage FALD. The observed correlations between the metabolomics and gene expression results reveal new insights into the mechanisms underlying FALD. FALD is not only a hepatic fibrosis disease, but also features severe dysregulation of liver metabolism.