Increased ketone body oxidation provides additional energy for the failing heart without improving cardiac efficiency

Abstract

Aims: The failing heart is energy-starved and inefficient due to perturbations in energy metabolism. Although ketone oxidation has been shown recently to increase in the failing heart, it remains unknown whether this improves cardiac energy production or efficiency. We therefore assessed cardiac metabolism in failing hearts and determined whether increasing ketone oxidation improves cardiac energy production and efficiency.

Methods and results: C57BL/6J mice underwent sham or transverse aortic constriction (TAC) surgery to induce pressure overload hypertrophy over 4-weeks. Isolated working hearts from these mice were perfused with radiolabelled β-hydroxybutyrate (βOHB), glucose, or palmitate to assess cardiac metabolism. Ejection fraction decreased by 45% in TAC mice. Failing hearts had decreased glucose oxidation while palmitate oxidation remained unchanged, resulting in a 35% decrease in energy production. Increasing βOHB levels from 0.2 to 0.6 mM increased ketone oxidation rates from 251 ± 24 to 834 ± 116 nmol·g dry wt-1 · min-1 in TAC hearts, rates which were significantly increased compared to sham hearts and occurred without decreasing glycolysis, glucose, or palmitate oxidation rates. Therefore, the contribution of ketones to energy production in TAC hearts increased to 18% and total energy production increased by 23%. Interestingly, glucose oxidation, in parallel with total ATP production, was also significantly upregulated in hearts upon increasing βOHB levels. However, while overall energy production increased, cardiac efficiency was not improved.

Conclusions: Increasing ketone oxidation rates in failing hearts increases overall energy production without compromising glucose or fatty acid metabolism, albeit without increasing cardiac efficiency.

Keywords: Beta-hydroxybutyrate; Cardiac Energy metabolism; Heart failure; Hypertrophy; Ketone body oxidation.

Figure 1

Assessment of in vivo and ex vivo cardiac function and morphology in TAC and sham C57Blk/6J mice. (A) Echocardiographic assessment of ejection fraction (unpaired two-tailed t-test, n = 20 for sham, 26 for TAC). (B) Echocardiographic assessment of diastolic dysfunction as measured by the ‘e’ to ‘e prime’ ratio (unpaired two-tailed t-test, n = 18 for sham, 23 for TAC). (C) Echocardiographic assessment of systolic function by a measure of fractional shortening (unpaired two-tailed t-test, n = 20 for sham, 26 for TAC). (D) Ex vivo assessment of cardiac function in the isolated perfused heart by a measure of cardiac work in which the concentration of βOHB was increased from 200 µM to 600 µM at 30 min (multiple t-tests with correction for multiple comparisons using the Bonferroni–Dunn method, n = 19 for sham, 22 for TAC). (A–C) Sham groups are represented by white circles while TAC groups are represented by red squares. Data are expressed as mean ± SEM. *P < 0.05 compared to the sham group.

Figure 2

Absolute metabolic rates of the failing heart in the presence of either 200 µM βOHB or 600 µM βOHB. (A) βOHB (ketone body) oxidation (n = 6 for sham, n = 6 for TAC). (B) Palmitate (fatty acid) oxidation (n = 8 for sham, n = 9 for TAC). (C) Glucose oxidation (n = 13 for sham, n = 16 for TAC). (D) Glycolysis (n = 10 for sham, n = 13 for TAC). Sham groups are represented by white circles while TAC groups are represented by red squares. A two-way ANOVA with Bonferroni correction for multiple comparisons was carried out for each panel in this figure. Data are expressed as mean ± SEM. *P < 0.05 compared to the sham group. ^†P < 0.05 compared to the 200 μM βOHB group.

Figure 3

Normalized metabolic rates of the failing heart in the presence of either 200 µM βOHB or 600 µM βOHB. (A) βOHB (ketone body) oxidation normalized to cardiac work (n = 6 for sham, n = 6 for TAC). (B) Palmitate (fatty acid) oxidation normalized to cardiac work (n = 8 for sham, n = 8 for TAC). (C) Glucose oxidation normalized to cardiac work (n = 13 for sham, n = 13 for TAC). (D) Glycolysis normalized to cardiac work (n = 10 for sham, n = 11 for TAC). Sham groups are represented by white circles while TAC groups are represented by red squares. A two-way ANOVA with Bonferroni correction for multiple comparisons was carried out for each panel in this figure. Data are expressed as mean ± SEM. *P < 0.05 compared to the sham group. ^†P < 0.05 compared to the 200 μM βOHB group.

Figure 4

TCA acetyl CoA production, ATP production, and cardiac efficiency in the failing heart in the presence of either 200 µM βOHB or 600 µM βOHB. (A) TCA acetyl CoA production from glucose oxidation, palmitate oxidation or βOHB oxidation. (B) ATP production from glucose oxidation, glycolysis, palmitate oxidation, or βOHB oxidation. (C) The percent contribution of glucose oxidation, glycolysis, palmitate oxidation, or βOHB oxidation to the total ATP production. (D) Cardiac efficiency as determined by normalizing cardiac work for total acetyl CoA production (n = 6 for all groups). In (D), sham groups are represented by white circles while TAC groups are represented by red squares. A two-way ANOVA with Bonferroni correction for multiple comparisons was carried out for each panel in this figure. Data are expressed as mean ± SEM. *P < 0.05 compared to the sham group. ^†P < 0.05 compared to the 200 μM βOHB group.

Figure 5

Protein expression levels of fatty acid and ketone body oxidative enzymes in the murine pressure overload failing heart. (A) Protein expression of all metabolic enzymes assessed at 600 µM βOHB (top to bottom): long chain acyl CoA dehydrogenase (LCAD), β-hydroxy-acyl-CoA-dehydrogenase (βHAD), succinyl-CoA-3-oxaloacid CoA transferase (SCOT), β-hydroxybutyrate dehydrogenase 1 (BDH1), and voltage-dependent anion channel (VDAC) (multiple t-tests with correction for multiple comparisons using the Bonferroni–Dunn method, n = 7 for all groups). All proteins were normalized to the loading control, VDAC. (B) Overall protein lysine acetylation in sham vs. TAC hearts at 600 µM βOHB (unpaired two-tailed t-test, n = 7 for both groups) whereby the loading control used was VDAC. (C) Assessment of LCAD acetylation status in sham vs. TAC hearts at 600 µM βOHB in which normalization was to the heavy chain of the antibody used to pull down all of the lysine-acetylated proteins (unpaired two-tailed t-test, n = 4 for sham and 7 for TAC). Densitometric analysis is shown on the right for panels (A–C). Sham groups are represented by white circles while TAC groups are represented by red squares. Data are expressed as mean ± SEM. *p < 0.05 compared to the sham group.

Figure 6

Summary diagram of the failing heart’s increased reliance on ketone oxidation for energy production. The percent contribution of βOHB to total TCA acetyl CoA production in the sham heart (A) and TAC heart (B) at 200 µM βOHB, and the TAC heart at 600 µM βOHB (C). The total energy production decreases in TAC hearts (B) compared to sham hearts (A) but increasing βOHB availability to the failing heart (C) increases its contribution to energy production by more than two-fold making it a significant substrate contributing to increased overall energy production for the energy starved heart.