The present data from indirect calorimetry are consistent with our previous report. The differences in RER values between lean and obese dam offspring were greater when challenged with HF diets, suggesting impaired metabolic flexibility. A recent study conducted in obese adolescents with non-alcoholic fatty liver disease reported that hepatic fat accumulation led to decreased reliance on fatty acid oxidation in the fasted state. This was accompanied by an inability to suppress fatty acid oxidation during an oral glucose tolerance test as determined by RER values. This impaired capacity to switch substrate utilization to FAO during fasting and back to carbohydrate oxidation when glucose challenged indicates metabolic inflexibility. Most importantly, impaired FAO was determined by hepatic fat content and not abdominal adiposity. Hence, it appears that there is an intricate relationship between hepatic steatosis and fatty acid oxidation. Consistent with these findings, offspring from obese dams develop increased liver weight and hepatic fat accumulation without differences in body weight or adiposity. Therefore, it is plausible that exposure to maternal obesity alters metabolic sensors leading to an impaired ability to oxidize fat. Mitochondria are typically the primary site for FAO, and since mitochondria are maternally inherited, several models of gestational programming have focused on changes in this organelle. Since our model exclusively examines the contribution of maternal obesity, mitochondrial changes may be an important conduit on how maternal obesity mediates programming of offspring metabolism. Mitochondrial dysfunction is highly associated with reduced FAO. While we found no differences in mRNA expression of mitochondrial transcription factor A between offspring of lean and obese dams, suggesting that mitochondrial numbers may not be affected, our studies identified several indications of mitochondrial dysfunction, including lower abundance of oxidative phosphorylation complexes. In addition to lower amounts of OXPHOS proteins, the function of the electron transport chain complexes and other mitochondrial proteins are highly regulated post-translationally via lysine acetylation. Recent studies have shown SIRT3, a member of the class III NAD+ dependent deacetylase family, to be located in the mitochondria and known to critically regulate OXPHOS in the liver. Sirtuins act as energy sensors and regulate metabolic processes via their deacetylation activity. The sirtuin family consists of seven isoforms that regulate distinct metabolic pathways in various cellular locations. SIRT1, 6, and 7 are located in the nucleus, SIRT2 in the cytosol, and SIRT3, 4, and 5 in the mitochondria. Our data suggest that maternal obesity affects the levels of several SIRT isoforms in the offspring liver, suggesting that the SIRT family may play a role in fetal metabolic programming. We chose to examine SIRT3 due to its mitochondrial AbMole Isoforskolin location and lack of change in mRNA expression in the other mitochondrial located isoforms. Further, Lombard et al. found that SIRT4 and SIRT5-deficient mice did not increase global lysine acetylation in contrast to SIRT3-deficient mice. Elegant studies in early mouse embryos and blastocyst also reveal that SIRT3 is maternally inherited and critical for protection from reactive oxygen species.