Ketone-loading as A Novel Sports Nutrition Strategy: Chronic Ketone Supplementation Elicits Further Favorable Metabolic Changes in Keto-adapted Mice

Background A ketogenic diet (KD) induces nutritional ketosis (NS), benefits fatty acid oxidation (FAO), and favors moderate-intensity exercise capacity. The status that the body accommodates to produce and utilize ketone bodies (KB) and fatty acids as primary fuel sources is termed keto-adaptation. However, keto-adaptation requires time, while long-term KD also involves unfavored adverse effects. Exogenous ketone body (EKBs) administration has been introduced to elicit the advantages of NS. However, the direct use of EKBs fails to bring desired outcomes. We hypothesized that EKBs might only be effective during keto-adaptation. Methods Male C57BL/6J mice (n = 24) were divided into three groups: a control diet (Con, n = 8), a ketogenic diet (KD, n = 8), and a KD plus a ketone body (DL-β-Hydroxybutyric acid sodium salt, BHB) administration (KD+BHB, n = 8). After six weeks of KD administration, mice in the KD+BHB group receive BHB added into water bottles for another six weeks. Blood KB concentration is monitored throughout the experiment, while liver, gastrocnemius, and soleus mRNA are analyzed using RT-PCR. Results Both KD and KD+BHB induced and sustained NS and enhanced hepatic and muscular key genes regulating FAO. In addition, BHB administration upon keto-adaptation further increased circulating KB concentration and enhanced expressional levels of FAO-mediating genes ( ACO, HADH, ACADM , and MLYCD in the gastrocnemius muscle; ACO, HADH , and MLYCD in the soleus muscle), and energy-regulating genes ( PPARA and PPARG ) in the liver and skeletal muscle compared to a KD. Conclusion Compared to KD alone, chronic administration of KBs upon keto-adaptation increased the expression of key genes that favor FAO or maintain energy homeostasis in the liver and skeletal muscle. Instead of directly using EKBs in non-keto-adapted individuals, it is encouraged to use EKBs upon keto-adaptation status to elicit their energy-utilizing effects. Highlights This is the first report to evaluate the metabolic effects using exogenous ketone bodies on keto-adapted individuals. Administration of exogenous ketone body upon keto-adaptation furtherly increased circulating ketone bodies. Administration of exogenous ketone body upon keto-adaptation individuals enhanced expression of genes related to fatty acid oxidation and energy hemostasis.