Dietary branched-chain amino acid restriction alters fuel selection and reduces triglyceride stores in hearts of Zucker fatty rats

Abstract

Elevations in circulating levels of branched-chain amino acids (BCAAs) are associated with a variety of cardiometabolic diseases and conditions. Restriction of dietary BCAAs in rodent models of obesity lowers circulating BCAA levels and improves whole-animal and skeletal-muscle insulin sensitivity and lipid homeostasis, but the impact of BCAA supply on heart metabolism has not been studied. Here, we report that feeding a BCAA-restricted chow diet to Zucker fatty rats (ZFRs) causes a shift in cardiac fuel metabolism that favors fatty acid relative to glucose catabolism. This is illustrated by an increase in labeling of acetyl-CoA from [1-¹³C]palmitate and a decrease in labeling of acetyl-CoA and malonyl-CoA from [U-¹³C]glucose, accompanied by a decrease in cardiac hexokinase II and glucose transporter 4 protein levels. Metabolomic profiling of heart tissue supports these findings by demonstrating an increase in levels of a host of fatty-acid-derived metabolites in hearts from ZFRs and Zucker lean rats (ZLRs) fed the BCAA-restricted diet. In addition, the twofold increase in cardiac triglyceride stores in ZFRs compared with ZLRs fed on chow diet is eliminated in ZFRs fed on the BCAA-restricted diet. Finally, the enzymatic activity of branched-chain ketoacid dehydrogenase (BCKDH) is not influenced by BCAA restriction, and levels of BCAA in the heart instead reflect their levels in circulation. In summary, reducing BCAA supply in obesity improves cardiac metabolic health by a mechanism independent of alterations in BCKDH activity.

Keywords: Zucker fatty rat; branched-chain amino acids; cardio metabolic diseases; heart metabolism; obesity.

Fig. 1.

Dietary branched-chain amino acid restriction alters fuel selection in isolated Zucker fatty rat (ZFR) hearts. Male ZFR were fed chow diet (Obese control) or a chow diet in which BCAA were restricted by 45% (Obese restr) for 15 wk. The hearts were then isolated and perfused with [1-¹³C]palmitate and [U-¹³C]glucose. A: labeling strategy and ¹³C enrichment in the acetyl-CoA pool. B: 2-deoxyglucose-6-phosphate concentrations. C: ¹³C enrichment in the malonyl-CoA pool. n = 7–8 per group. Data represent mean ± SE. *P < 0.05, **P < 0.005. 2DG6P, 2 deoxyglucose-6-phosphate; PDH, pyruvate dehydrogenase.

Fig. 2.

Effect of dietary branched-chain amino acid (BCAA) restriction on fatty-acid oxidation-related metabolites. Zucker lean rats and Zucker fatty rats (ZFRs) were fed a chow diet (lean control and obese control, respectively) or BCAA-restricted diet (lean restr, and obese restr, respectively). Even-chain acylcarnitine (AC) concentrations (A), even-chain hydroxylated-acylcarnitine (OH-AC) concentrations (B), and acyl-CoA concentrations (C) in snap-frozen hearts from a previously described animal cohort (19). n = 9–15 per group. Acyl-CoA concentrations (D) in hearts from ZFRs fed a chow diet (obese control) or BCAA-restricted diet (obese restr) snap frozen at the conclusion of the perfusion study described in Fig. 1. n = 7–8 per group. Data represent mean ± SE. Statistical differences indicated by: *P < 0.05, **P < 0.005 for obesity effect and #P < 0.05, ##P < 0.005 for diet effect.

Fig. 3.

Effect of dietary branched-chain amino acid (BCAA) restriction on cardiac triglyceride (TG) concentrations, gene expression, and protein abundance. A: Zucker lean rats and Zucker fatty rats (ZFRs) were fed a chow diet (lean control and obese control, respectively) or BCAA-restricted diet (lean restr and obese restr, respectively) and cardiac TG concentrations were measured. n = 9–15 per group. *P < 0.05 vs. lean control. B: mRNA levels in perfused hearts from ZFR fed a chow diet (obese control) or BCAA-restricted diet (obese restr). C: representative immunoblots of glucose transporter (GLUT) 1, GLUT4, hexokinase (HK) II, phosphorylated AKT (p-AKT), and AKT with indicated loading controls and corresponding densitometric measurements. n = 7–8 per group. Data represent mean ± SE. *P < 0.05 for diet effect. CPT, carnitine palmitoyltransferase.

Fig. 4.

Effect of dietary branched-chain amino acid (BCAA) restriction on cardiac BCAA metabolism. Zucker lean rats and Zucker fatty rats were fed a chow diet (lean control and obese control, respectively) or a BCAA-restricted diet (lean restr and obese restr, respectively). A: BCAA concentrations. B: concentrations of BCAA catabolic products C3, C5, and C5-OH/C3-DC acylcarnitines. C: activity of the rate-limiting enzyme in BCAA catabolism, branched-chain ketoacid dehydrogenase (BCKDH). Data represent mean ± SE. Statistical differences indicated by: *P < 0.05, **P < 0.005 for obesity effect and #P < 0.05, ##P < 0.005 for diet effect. KIV, alpha-keto-isovalerate; Leu/Ile, leucine/isoleucine; Val, valine.