Background: During fetal development, the heart switches substrate preference from glucose to fatty acids, such that in the adult heart, 5070% of ATP is derived from fatty acid oxidation. What triggers this switch is currently unclear. In vivo, the late gestation rise in glucocorticoid levels is essential for structural and functional maturation of the fetal heart. Glucocorticoid treatment of fetal cardiomyocytes induces expression of PGC1a (a master regulator of mitochondrial phenotype), lipin1 and KLF15 (genes involved in fatty acid oxidation). We hypothesized that glucocorticoids instigate the switch to fatty acid oxidation in late gestation fetal cardiomyocytes.
Methods: Primary fetal cardiomyocytes were isolated following collagenase and pancreatin digestion of embryonic day (E)14.515.5 hearts. After 2 days in culture, cells were treated for 24 h with 1 mM dexamethasone. Oxygen consumption rate (a measure of mitochondrial respiration) and acidification rate (a measure of glycolysis) were measured using a Seahorse XF24 Analyzer using mitochondrial stress tests and glycolysis stress tests as appropriate. Respiration was measured in the presence of the fatty acid, palmitate (100 mM) and the fatty acid uptake blocker etomoxir (6 mM) or vehicle.
Results: Dexamethasone treatment did not alter glycolysis. Leak respiration was increased in dexamethasone treated cells (64.2±8.2 versus 82.02±14.25 pmol/min/protein, mean±SD, n=8). In palmitate-treated cells, dexamethasone increased basal respiration rate (517.9±48.0 versus 366.7±71 pmol/min/protein, mean±SD, n=5) and oxygen consumption (related to ATP production, 159.5±62.8 versus 297.9±35.5 pmol/min/protein, mean±SD, n=5) compared to vehicle. Etomoxir inhibited these dexamethasone-dependent increases.
Conclusion: These data support a glucocorticoid-induced switch in substrate preference towards fatty acid oxidation in fetal cardiomyocytes. The dexamethasone-induced increase in proton leak may serve to minimize DNA damage caused by mitochondrial reactive oxygen species (ROS) production, a mechanism that contributes to cardiomyocyte maturation. Future experiments will investigate whether glucocorticoids enhance mitochondrial ROS production.