Ketone Body Metabolism is Not Required for Improvement of Heart Failure by Ketogenic Diet in Mice

Abstract

Failing hearts increasingly metabolize ketone bodies, and enhancing ketosis improves heart failure (HF) remodeling. Circulating ketones are elevated by fasting/starvation, which is mimicked with a high-fat, low-carbohydrate "ketogenic diet" (KD). While speculated that KD improves HF through increased ketone oxidation, some evidence suggests KD paradoxically downregulates cardiac ketone oxidation despite increased ketone delivery. We sought to clarify the significance of cardiac ketone metabolism during KD in HF. Mice were subjected to transverse aortic constriction with apical myocardial infarction (TAC-MI) and fed either low-fat (LF) control or KD. Cardiac-specific mitochondrial pyruvate carrier 2 (csMPC2-/-) mice were used as a second model of heart failure. In both mice, feeding a KD improved HF, determined by echocardiography, heart weights, and gene expression analyses. Although KD increases plasma ketone bodies, gene expression for ketone metabolic genes is decreased in the hearts of KD-fed mice. Cardiac-specific β-hydroxybutyrate dehydrogenase 1 (csBDH1-/-), the first enzyme in ketone catabolism, mice were also studied and crossed with the csMPC2-/- mice to create double knockout (DKO) mice. These mice were aged to 16 weeks and switched to LF or KD, and KD was able to completely normalize the hearts of both csMPC2-/- and DKO mice, suggesting that ketone metabolism is unnecessary for improving heart failure with ketogenic diet. These studies were then repeated, and mice injected with U-¹³C-β-hydroxybutyrate to evaluate ketone metabolism. KD feeding significantly decreased the enrichment of the TCA cycle from ketone body carbons, as did the BDH1-deletion in DKO mice. Gene expression and respirometry suggests that KD instead increases cardiac fat oxidation. In conclusion, these results suggest that ketogenic diet decreases cardiac ketone metabolism and does not require ketone metabolism to improve heart failure.

Figure.. Ketone body metabolism is not required for ketogenic diet improvement of heart failure.

A, Echocardiography measures of end-systolic volume (ESV), end-diastolic volume (EDV), and ejection fraction (EF) of female C57BL/6J mice subjected to transverse aortic constriction with apical myocardial infarction (TAC-MI) 2 weeks post-surgery, and after 2 weeks of low-fat (LF) or ketogenic diet (KD) feeding (4-weeks post-surgery). Data also expressed as %-change from 2- to 4-weeks (n=8–9 per group). B, Left, Heart weight normalized to tibia length (HW/TL) and total plasma ketone concentrations of mice subjected to TAC-MI and fed LF or KD (n=8–9 per group). B, Right, Cardiac gene expression for heart failure and fibrosis genes, ketone metabolism genes, and fat oxidation genes (n=8–9 per group). C, Representative western blot showing cardiac deletion of BDH1 and/or MPC1/2 proteins from csBDH1−/−, csMPC2−/−, and DKO hearts. D, ESV, EDV, EF, HW/TL, and Bdh1 expression from WT (Mpc2 and Bdh1 floxed), csBDH1−/−, csMPC2−/−, and DKO mice fed LF or KD (n=5–17 per group). E, Schematic of U-¹³C-β-hydroxybutyrate (BHB) labeling, and the primary enrichment products for BHB, BHB-carnitine, acetyl-CoA, acetylcarnitine, glutamine, and succinate, as well as BHB pool size (n=4–6 per group). F, Oxygen consumption rates (OCR) of cardiac muscle fibers from LF of KD-fed mice when provided palmitoyl-CoA + carnitine, and then adenosine diphosphate (ADP) (n=9 per group). Data expressed as mean ± SD and analyzed by Student’s 2-tailed t test (A, B, and F), or 2-way ANOVA with Tukey multiple comparison’s test (D and E) with GraphPad Prism v10.3.0.