Hepatic Branched-chain Alpha-keto Acid Dehydrogenase Research Articles

Octanoic acid promotes branched-chain amino acid catabolisms via the inhibition of hepatic branched-chain alpha-keto acid dehydrogenase kinase in rats

ObjectiveIt has been reported that administration of octanoic acid, one of medium-chain fatty acids (MCFAs), promoted leucine oxidation in vitro and in vivo, but it remained unclear how octanoic acid stimulated leucine oxidation. Therefore, the aim of this study was to elucidate the mechanism that octanoic acid facilitates branched-chain amino acid (BCAA) catabolism. Materials/MethodsIn in vivo experiments, male rats were orally administered MCFAs as free fatty acids or triacylglycerol (trioctanoin), and then activities of hepatic branched-chain α-ketoacid dehydrogenase (BCKDH) complex (BCKDC) and BCKDH kinase (BDK) and alterations in the concentration of blood components were analyzed. In in vitro experiments, purified BCKDC associated with BDK (BCKDH–BDK complex) was reacted with various concentrations of hexanoic, octanoic, and decanoic acids. ResultsOral administration of trioctanoin in rats activated hepatic BCKDC via down-regulation of BDK activity in association with a decrease in plasma BCAA concentration and an increase in serum ketone body concentration. In vitro experiments using purified BCKDH–BDK complex showed that MCFAs (hexanoic, octanoic, and decanoic acids) inhibited BDK activity and that this inhibition was higher in hexanoic and octanoic acids than in decanoic acid. Oral administration of octanoic acid, but not decanoic acid, in rats activated hepatic BCKDC via down-regulation of BDK activity by decreasing the amount of BDK bound to the complex. The serum ketone body level was elevated by both administration of octanoic acid and decanoic acid. ConclusionThese results suggest that octanoic acid promotes BCAA catabolism in vivo by activation of BCKDC via decreasing the bound form of BDK.

Exercise Promotes BCAA Catabolism: Effects of BCAA Supplementation on Skeletal Muscle during Exercise

Branched-chain amino acids (BCAAs) are essential amino acids that can be oxidized in skeletal muscle. It is known that BCAA oxidation is promoted by exercise. The mechanism responsible for this phenomenon is attributed to activation of the branched-chain alpha-keto acid dehydrogenase (BCKDH) complex, which catalyzes the second-step reaction of the BCAA catabolic pathway and is the rate-limiting enzyme in the pathway. This enzyme complex is regulated by a phosphorylation-dephosphorylation cycle. The BCKDH kinase is responsible for inactivation of the complex by phosphorylation, and the activity of the kinase is inversely correlated with the activity state of the BCKDH complex, which suggests that the kinase is the primary regulator of the complex. We found recently that administration of ligands for peroxisome proliferator-activated receptor-alpha (PPARalpha) in rats caused activation of the hepatic BCKDH complex in association with a decrease in the kinase activity, which suggests that promotion of fatty acid oxidation upregulates the BCAA catabolism. Long-chain fatty acids are ligands for PPARalpha, and the fatty acid oxidation is promoted by several physiological conditions including exercise. These findings suggest that fatty acids may be one of the regulators of BCAA catabolism and that the BCAA requirement is increased by exercise. Furthermore, BCAA supplementation before and after exercise has beneficial effects for decreasing exercise-induced muscle damage and promoting muscle-protein synthesis; this suggests the possibility that BCAAs are a useful supplement in relation to exercise and sports.

Uteroplacental insufficiency alters liver and skeletal muscle branched-chain amino acid metabolism in intrauterine growth-restricted fetal rats.

Uteroplacental insufficiency causes intrauterine growth restriction (IUGR) and decreases plasma levels of the branched-chain amino acids in both humans and rats. Increased fetal oxidation of these amino acids may contribute to their decline in the IUGR fetus. The rate-limiting step of branched-chain amino acid oxidation is performed by the mitochondrial enzyme branched-chain alpha-keto acid dehydrogenase (BCKAD), which is regulated by a deactivating kinase. We therefore hypothesized that uteroplacental insufficiency increases BCKAD activity through altered mRNA and protein levels of BCKAD and/or the BCKAD kinase. In IUGR fetal liver, BCKAD activity was increased 3-fold, though no difference in hepatic BCKAD protein or mRNA levels were noted. Hepatic BCKAD kinase mRNA and protein levels were significantly decreased in association with the increase in BCKAD activity. In IUGR fetal skeletal muscle, BCKAD mRNA levels were significantly increased. IUGR skeletal muscle BCKAD protein levels as well as BCKAD kinase mRNA and protein levels were unchanged. We also quantified mRNA levels of two amino acid transporters: LAT1 (system L) and rBAT (cysteine and dibasic amino acids). Both hepatic and muscle LAT1 mRNA levels were significantly increased in the IUGR fetus. We conclude that uteroplacental insufficiency significantly increases hepatic BCKAD activity in association with significantly decreased mRNA and protein levels of the deactivating kinase. We speculate that these changes contribute to the decreased serum levels of branched-chain amino acids seen in the IUGR fetus and may be an adaptation to the deprived milieu associated with uteroplacental insufficiency.

Modification by exercise training of activity and enzyme expression of hepatic branched-chain alpha-ketoacid dehydrogenase complex in streptozotocin-induced diabetic rats.

The branched-chain alpha-ketoacid dehydrogenase (BCKDH) complex is the rate-limiting enzyme in the catabolism of branched-chain amino acids. In the present study, we examined the effects of exercise training on the activity and enzyme expression of the hepatic BCKDH complex in diabetic rats. The rats were prepared by intravenous injections of streptozotocin (50 mg/kg BW), and exercise training was accomplished by treadmill running for 45 min/d for 4 wk. The total and actual activities of hepatic BCKDH complex were significantly increased to approximately 160% by 4 wk of diabetes. On the other hand, diabetic rats in the trained group had the same level of activities as those in the normal rats, indicating that exercise training inhibited the diabetes-induced increase in the enzyme activities. The activity state (% active form) of the enzyme complex was about 100% in all groups and was not affected by diabetes or training. The protein amounts of the enzyme subunits (E1alpha and E2) and the abundance of mRNA for the E2 subunit, but not for the other subunits, in the liver had the same trend as the activities. These results suggest that the capacity for branched-chain amino acid catabolism in streptozotocin-induced diabetic rats is reduced by exercise training and that this modification is associated with the suppression of diabetes-induced BCKDH complex expression in the liver.

Suppression of glycogen consumption during acute exercise by dietary branched-chain amino acids in rats.

The effects of a diet supplemented with branched-chain amino acids (BCAA; 4.8% or 6.2%) on BCAA catabolism and glycogen metabolism in rats were examined. Rats were fed a BCAA diet or control diet for 4 wk and part of the rats were subjected to exercise training during the experimental period. Feeding the BCAA diet increased serum BCAA concentrations and activity of the hepatic branched-chain alpha-keto acid dehydrogenase complex, the rate-limiting enzyme in the catabolism of BCAA, suggesting that dietary BCAA promotes BCAA catabolism. Although the serum glucose concentration and glycogen contents in the liver and gastrocnemius muscle of rested rats were not significantly affected by feeding of the BCAA diet, those in rats exhausted by acute exercise were 2-4-fold higher in rats fed the BCAA diet than in rats fed the control diet. The activity of pyruvate dehydrogenase complex in the liver and gastrocnemius muscle after acute exercise showed reverse trends; the complex activities (especially in liver) tended to be less in the BCAA diet group than in the control diet group. These results suggest that dietary BCAA spares glycogen stores in liver and skeletal muscle during exercise and that the decrease in pyruvate dehydrogenase complex activity in these tissues by dietary BCAA is involved in the mechanisms.

Regulation of Rat Liver Branched-Chain α-Keto Acid Dehydrogenase Activity by Meal Frequency and Dietary Protein

Effects of feeding frequency on liver branched-chain alpha-keto acid dehydrogenase (BCKAD) activity are unknown. In the present study, rats were trained to consume their daily allotment of food in 6 h (meal-feeding). Rats were fed diets containing 0, 9, 25 or 50% casein and after 10 d were killed before or 3 h after the meal. The enzyme in rats fed diets containing 0, 9 and 25% casein was activated three- to sixfold after meal consumption. Previous studies showed that the liver enzyme is essentially fully activated in post-absorptive rats fed an adequate protein diet ad libitum. Meal-feeding an adequate protein (25% casein) diet resulted in a marked decrease in the postabsorptive percentage of active complex compared to ad libitum feeding of the same diet (29 +/- 6% vs. 93 +/- 6% active). Administration of alpha-ketoisocaproate (200 mumol/100 g body weight, an inhibitor of BCKAD kinase) reversed the meal-feeding-induced inactivation of the complex within 10 min. We conclude that the frequency of food intake, in addition to the level of dietary protein, influences the proportion of liver BCKAD in the active state. Inactivation of hepatic BCKAD in rats trained to feed once a day may be an adaptive mechanism that results in increased efficiency of branched-chain amino acid utilization between meals.

Nutritional and hormonal regulation of the activity state of hepatic branched-chain alpha-keto acid dehydrogenase complex.

The hepatic branched-chain alpha-keto acid dehydrogenase complex plays an important role in regulating branched-chain amino acid levels. These compounds are essential for protein synthesis but are toxic if present in excess. When dietary protein is deficient, the hepatic enzyme is present in the inactive, phosphorylated state to allow conservation of branched-chain amino acids for protein synthesis. When dietary protein is excessive, the enzyme is in the active, dephosphorylated state to commit the excess branched-chain amino acids to degradation. Inhibition of protein synthesis by cycloheximide, even when the animal is starving for protein, results in activation of the hepatic branched-chain alpha-keto acid dehydrogenase complex to prevent accumulation of branched-chain amino acids. Likewise, the increase in branched-chain amino acids caused by body wasting during starvation and uncontrolled diabetes is blunted by activation of the hepatic branched-chain alpha-keto acid dehydrogenase complex. The activity state of the hepatic branched-chain alpha-keto acid dehydrogenase complex is regulated in the short term by the concentration of branched-chain alpha-keto acids (inhibitors of branched-chain alpha-keto acid dehydrogenase kinase) and in the long term by alteration in the total branched chain alpha-keto acid dehydrogenase kinase activity.

Mechanism of activation of hepatic branched-chain alpha-ketoacid dehydrogenase by a muscle factor.

We recently reported that a skeletal muscle factor specifically activates the branched-chain alpha-ketoacid (BCKA) dehydrogenase in liver mitochondria (Paul, H. S., and Adibi, S. A. (1982) J. Biol. Chem. 257, 12581-12588). The evidence suggested that the muscle factor may be a phosphatase or may stimulate the phosphatase in liver mitochondria, thus converting the inactive BCKA dehydrogenase into an active form. In the present study we have investigated these possibilities. The muscle factor did not increase the activity of BCKA dehydrogenase which was previously activated in vitro by preincubation of mitochondria or by the addition of Triton or Ca2+ to the incubation medium. The muscle factor had little or no stimulatory effect on BCKA dehydrogenase when this enzyme was activated in vivo by treatment of rats with streptozotocin or clofibrate. The addition of NaF (an inhibitor of phosphatase) to the incubation medium entirely abolished the stimulatory effect of the muscle factor on BCKA dehydrogenase. Elimination of phosphatase by partial purification of BCKA dehydrogenase resulted in loss of stimulation of the dehydrogenase by the muscle factor. These results suggest that the muscle factor activates the BCKA dehydrogenase in liver mitochondria by stimulating its associated phosphatase.