Abstract 4121970: Early to Mid-Gestational Testosterone Excess Sex-specifically Reprograms Cardiac Transcriptome and Metabolome Profiles of Sheep Fetuses

Exposure of sheep to excess testosterone (T) from days 30-90 of gestation (term 147 days) perturbs maternal systemic lipidome in late gestation, induces fetal growth restriction and sexually dimorphic perturbation in the cardiac left ventricular (LV) transcriptome in gestational (GD) day 90 fetuses culminating in cardiometabolic dysfunction in adult female offspring. We hypothesized that the detrimental impact of prenatal T excess on fetal cardiac transcriptome is a programmed event that will persist even after cessation of excess T exposure and will be accompanied by changes in fetal metabolome. Pregnant ewes were injected with 100 mg intramuscular T propionate or vehicle (C), twice weekly from GD 30-90. On GD120±5, 30 days after cessation of T treatment fetal weights were recorded, left ventricle (LV) tissue and fetal plasma processed for transcriptomics by RNA sequencing and metabolomics analysis (Female C (FC) n=6, T (FT) n=7; Male C (MC) n=6, T (MT) n=7). LV transcriptome-based pathways enrichment analysis revealed 1) downregulation of pathways associated with oxidative phosphorylation, citric acid cycle, pyruvate metabolism; 2) upregulation of TGF-β signaling pathway in T fetuses compared to C (p<0.05, FDR<0.1); 3) selective downregulation of the glycolytic pathway in MT relative to MC as opposed to downregulation of mitochondria-related pathways in FT vs FC (p<0.05, FDR<0.1). LV lipidomics analyses found glycerophospholipids and sphingolipids as main class lipid metabolites to be enriched in FT relative to FC fetuses (p<0.05, ER>1). Fetal systemic lipidome main class metabolites enriched (p<0.05, ER>2) in T vs C fetuses included 1) sphingomyelins in both sexes 2) fatty acyls metabolites only in MT vs MC as opposed to glycerophospholipids only in FT vs FC. LV polar metabolites enriched (p<0.05, ER>2) included 1) arginine&proline metabolism pathways in T vs C in both sexes 2) nicotinate and nicotinamide metabolism only in MT vs MC (p<0.05, ER>3), and porphyrin, riboflavin metabolism and N-glycan biosynthesis pathways only in FT vs FC. Systemic polar metabolites enriched (p<0.05, ER>2) included 1) D-amino acid metabolism pathways in T vs. C, 2) valine, leucine, and isoleucine pathways only in FT vs FC and glycerophospholipid and sphingolipid metabolism only in MT vs MC. These results suggest persistence of sex-specific perturbation in fetal transcriptome and systemic and cardiac metabolome in late gestational fetuses exposed to prenatal T excess.