β-Klotho (β-Kl), a transmembrane protein expressed in the liver, pancreas, adipose tissues, and brain, is essential for feedback suppression of hepatic bile acid synthesis. Because bile acid is a key regulator of lipid and energy metabolism, we hypothesized potential and tissue-specific roles of β-Kl in regulating plasma lipid levels and body weight. By crossing β-kl(-/-) mice with newly developed hepatocyte-specific β-kl transgenic (Tg) mice, we generated mice expressing β-kl solely in hepatocytes (β-kl(-/-)/Tg). Gene expression, metabolomic, and in vivo flux analyses consistently revealed that plasma level of cholesterol, which is over-excreted into feces as bile acids in β-kl(-/-), is maintained in β-kl(-/-) mice by enhanced de novo cholesterogenesis. No compensatory increase in lipogenesis was observed, despite markedly decreased plasma triglyceride. Along with enhanced bile acid synthesis, these lipid dysregulations in β-kl(-/-) were completely reversed in β-kl(-/-)/Tg mice. In contrast, reduced body weight and resistance to diet-induced obesity in β-kl(-/-) mice were not reversed by hepatocyte-specific restoration of β-Kl expression. We conclude that β-Kl in hepatocytes is necessary and sufficient for lipid homeostasis, whereas nonhepatic β-Kl regulates energy metabolism. We further demonstrate that in a condition with excessive cholesterol disposal, a robust compensatory mechanism maintains cholesterol levels but not triglyceride levels in mice.

The combined activation of the cellular energy sensor AMP‐activated protein kinase (AMPK) and the nuclear transcription factor peroxisome proliferator‐activated receptor delta (PPAR δ) has been demonstrated to improve endurance and muscle function by mimicking the effects of exercise training. However, their combined pharmacological activation with exercise training has not been explored. Balb/c mice were trained on a treadmill and administered both the AMPK activator AICAR and the PPAR δ agonist GW0742 for 4 weeks. AICAR treatment potentiated endurance, but the combination of AICAR and GW0742 further potentiated endurance and increased all running parameters significantly relative to exercised and nonexercised groups (138–179% and 355% increase in running time, respectively). Despite the lack of change in basal whole‐body metabolism, a significant shift to fat as the main energy source with a decline in carbohydrate utilization was observed upon indirect calorimetry analysis at the period near exhaustion. Increased energy substrates before exercise, and elevated muscle nonesterified fatty acids (NEFA) and elevated muscle glycogen at exhaustion were observed together with increased PDK4 mRNA expression. Citrate synthase activity was elevated in AICAR‐treated groups, while PGC‐1α protein level tended to be increased in GW0742‐treated groups. At exhaustion, Pgc1a was robustly upregulated together with Pdk4, Cd36, and Lpl in the muscle. A robust upregulation of Pgc1a and a downregulation in Chrebp were observed in the liver. Our data show that combined pharmacological activation of AMPK and PPAR δ potentiates endurance in trained mice by transcriptional changes in muscle and liver, increased available energy substrates, delayed hypoglycemia through glycogen sparing accompanied by increased NEFA availability, and improved substrate shift from carbohydrate to fat.

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Although circulating fatty acids are utilized as energy substrates, they also function as ligands to the peroxisome‐proliferator activated receptors (PPARs), a family of fatty acid sensing transcription factors. Exercise training leads to various adaptations in the muscle such as elevation of glycogen content, mitochondrial number as well as upregulation of fatty acid uptake and utilization through downstream transcriptional adaptations. In line with this, CD36 has been shown to be critical in controlling fatty acid uptake and consequently, fatty acid oxidation. We show that exercise training could not ameliorate impaired endurance performance in CD36 KO mice despite intact adaptations in muscle glycogen storage and mitochondrial function. Changes in whole‐body metabolism at rest and during exercise were also suppressed in these animals. Furthermore, there was inefficient upregulation of PPAR and PPAR‐related exercise‐responsive genes with chronic training in CD36 KO mice despite normal upregulation of Pgc1a and mitochondrial genes. Our findings supplement previous observations and emphasize the importance of CD36 in endurance performance, energy production and efficient downstream transcriptional regulation by PPARs.

BackgroundDiets containing fats of different proportions and types have been demonstrated to influence metabolism. These fats differ in long chain fatty acids (LCFAs) or medium chain fatty acids (MCFAs) content. In our laboratory using swimming as the training modality, MCFAs increased endurance attributed to increased activities of oxidative enzymes. How it affects whole-body metabolism remains unexplored. The present study investigated the metabolic, biochemical and genetic adaptations with treadmill running as the training modality.MethodsC57BL/6N mice were divided into untrained and trained groups and provided with low-fat (10% kcal from soybean oil), coconut oil (10% kcal from soybean oil, 20% kcal from coconut oil) or soybean oil (30% kcal from soybean oil) diet. Training was performed on a treadmill for 30 days. After recovery, whole-body metabolism at rest and during exercise, endurance, substrate metabolism, mitochondrial enzyme activities, and gene expression of training-adaptive genes in the muscle and liver were measured.ResultsAt rest, medium-fat diets decreased respiratory exchange ratio (RER) (p < 0.05). Training increased RER in all diet groups without affecting oxygen consumption (p < 0.05). During exercise, diets had no overt effects on metabolism while training decreased oxygen consumption indicating decreased energy expenditure (p < 0.05). Coconut oil without training improved endurance based on work (p < 0.05). Training improved all endurance parameters without overt effects of diet (p < 0.05). Moreover, training increased the activities of mitochondrial enzymes likely related to the increased expression of estrogen related receptor (ERR) α and ERRβ (p < 0.05). Coconut oil inhibited peroxisome proliferator-activated receptor (PPAR) β/δ activation and glycogen accumulation in the muscle but activated PPARα in the liver in the trained state (p < 0.05). Substrate utilization data suggested that coconut oil and/or resulting ketone bodies spared glycogen utilization in the trained muscle during exercise thereby preserving endurance.ConclusionOur data demonstrated the various roles of diet and fat types in training adaptation. Diets exerted different roles in PPAR activation and substrate handling in the context of endurance exercise training. However, the role of fat types in training adaptations is limited as training overwhelms and normalizes the effects of diet in the untrained state particularly on endurance performance, mitochondrial biogenesis, and ERR expression.