Kyle L. Fulghum

Ketogenesis and ketone body metabolism are linked to brain health benefits, including delaying age-related cognitive decline and neurodegeneration. Exercise, particularly when combined with an overnight fast, stimulates ketogenesis and ketone body turnover as well as improves brain metabolism and cognition. Yet, whether ketone metabolism is obligatory for this response is unknown. Here, we use chronic exercise via voluntary wheel running plus time-restricted feeding (VWR+TRF, fasting from ZT10.5-18.5) to explore whether ketone bodies are a potential mediator of exercise-induced brain health benefits in middle-aged mice. To independently distinguish the roles of neuronal ketone body metabolism vs. hepatic ketone body production, we studied middle-age female neuronal-specific SCOT knockout mice and female hepatocyte-specific HMGCS2 knockout mice, respectively. VWR+TRF was compared to sedentary ad-libitum fed (SED+AL) mice to assess the impact on whole-body metabolism (indirect calorimetry), cognition (Barnes Maze and Y-Maze), and molecular adaptations in the hippocampus (proteomics). VWR+TRF robustly upregulated systemic lipid oxidation in all mice, regardless of genotype, during the first 6.5 hours of the dark period. In female SCOT-Neuron-KO mice, we show impaired responses to VWR+TRF in indices of short- and long-term memory. Proteomic analysis of isolated hippocampi revealed that SCOT-Neuron-KO mice failed to globally upregulate key facilitators of synaptic function, including leucine-rich repeated transmembrane proteins, neurexins, and neuroligins. In female HMGCS2-Liver-KO mice, impaired responses to VWR+TRF in indices of short-term memory were paired with an upregulation in ketogenesis machinery in the hippocampal proteome, suggesting potential in vivo evidence of cerebral ketogenesis, a mechanism mitigating an otherwise more pronounced behavioral phenotype. Together, these findings suggest that neuronal ketone body utilization is essential, and hepatic ketone production is contributory, to the full cognitive and synaptic adaptations to exercise plus time-restricted feeding, supporting ketone metabolism as a key mechanistic link between metabolic state and brain health in midlife.

This paper established a protocol to maximize ketone body respiration in isolated brain mitochondria while quantifying acetyl-CoA and energy charge in control mice compared with mice with neuron-specific deletion of succinyl-CoA-3-oxoacid-CoA transferase (SCOT) enzyme, required for ketone body oxidation. Findings are that alpha-ketoglutarate substantially increased acetoacetate (AcAc)-dependent respiration and neuron SCOT-KO had lower AcAc state 3 respiration with a decreased acetyl-CoA level.

The progression of metabolic dysfunction–associated steatotic liver disease (MASLD) to metabolic dysfunction–associated steatohepatitis (MASH) involves alterations in both liver-autonomous and systemic metabolism that influence the liver’s balance of fat accretion and disposal. Here, we quantify the contributions of hepatic oxidative pathways to liver injury in MASLD-MASH. Using NMR spectroscopy, UHPLC-MS, and GC-MS, we performed stable isotope tracing and formal flux modeling to quantify hepatic oxidative fluxes in humans across the spectrum of MASLD-MASH, and in mouse models of impaired ketogenesis. In humans with MASH, liver injury correlated positively with ketogenesis and total fat oxidation, but not with turnover of the tricarboxylic acid cycle. Loss-of-function mouse models demonstrated that disruption of mitochondrial HMG-CoA synthase (HMGCS2), the rate-limiting step of ketogenesis, impairs overall hepatic fat oxidation and induces an MASLD-MASH–like phenotype. Disruption of mitochondrial β-hydroxybutyrate dehydrogenase (BDH1), the terminal step of ketogenesis, also impaired fat oxidation, but surprisingly did not exacerbate steatotic liver injury. Taken together, these findings suggest that quantifiable variations in overall hepatic fat oxidation may not be a primary determinant of MASLD-to-MASH progression, but rather that maintenance of ketogenesis could serve a protective role through additional mechanisms that extend beyond overall rates of fat oxidation.

Ketogenesis is a dynamic metabolic conduit supporting hepatic fat oxidation particularly when carbohydrates are in short supply. Ketone bodies may be recycled into anabolic substrates, but a physiological role for this process has not been identified. Here, we use mass spectrometry–based ¹³ C-isotope tracing and shotgun lipidomics to establish a link between hepatic ketogenesis and lipid anabolism. Unexpectedly, mouse liver and primary hepatocytes consumed ketone bodies to support fatty acid biosynthesis via both de novo lipogenesis (DNL) and polyunsaturated fatty acid (PUFA) elongation. While an acetoacetate intermediate was not absolutely required for ketone bodies to source DNL, PUFA elongation required activation of acetoacetate by cytosolic acetoacetyl–coenzyme A synthetase (AACS). Moreover, AACS deficiency diminished free and esterified PUFAs in hepatocytes, while ketogenic insufficiency depleted PUFAs and increased liver triacylglycerols. These findings suggest that hepatic ketogenesis influences PUFA metabolism, representing a molecular mechanism through which ketone bodies could influence systemic physiology and chronic diseases.

: The progression of metabolic-dysfunction-associated steatotic liver disease (MASLD) to metabolic-dysfunction-associated steatohepatitis (MASH) involves complex alterations in both liver-autonomous and systemic metabolism that influence the liver’s balance of fat accretion and disposal. Here, we quantify the relative contribution of hepatic oxidative pathways to liver injury in MASLD-MASH. Using NMR spectroscopy, UHPLC-MS, and GC-MS, we performed stable-isotope tracing and formal flux modeling to quantify hepatic oxidative fluxes in humans across the spectrum of MASLD-MASH, and in mouse models of impaired ketogenesis. We found in humans with MASH, that liver injury correlated positively with ketogenesis and total fat oxidation, but not with turnover of the tricarboxylic acid cycle. The use of loss-of-function mouse models demonstrated that disruption of mitochondrial HMG-CoA synthase (HMGCS2), the rate-limiting step of ketogenesis, impairs overall hepatic fat oxidation and induces a MASLD-MASH-like phenotype. Disruption of mitochondrial β-hydroxybutyrate dehydrogenase (BDH1), the terminal step of ketogenesis, also impaired fat oxidation, but surprisingly did not exacerbate steatotic liver injury. Taken together, these findings suggest that quantifiable variations in overall hepatic fat oxidation may not be a primary determinant of MASLD-to-MASH progression, but rather, that maintenance of hepatic ketogenesis could serve a protective role through additional mechanisms that extend beyond quantified overall rates of fat oxidation.

Summary: Therapeutic interventions targeting hepatic lipid metabolism in metabolic dysfunction-associated steatotic liver disease (MASLD) and steatohepatitis (MASH) remain elusive. Using mass spectrometry-based stable isotope tracing and shotgun lipidomics, we established a novel link between ketogenesis and MASLD pathophysiology. Our findings show that mouse liver and primary hepatocytes consume ketone bodies to support fatty acid (FA) biosynthesis via both de novo lipogenesis (DNL) and FA elongation. Analysis of ¹³ C-labeled FAs in hepatocytes lacking mitochondrial D-β-hydroxybutyrate dehydrogenase (BDH1) revealed a partial reliance on mitochondrial conversion of D-βOHB to acetoacetate (AcAc) for cytoplasmic DNL contribution, whereas FA elongation from ketone bodies was fully dependent on cytosolic acetoacetyl-CoA synthetase (AACS). Ketone bodies were essential for polyunsaturated FA (PUFA) homeostasis in hepatocytes, as loss of AACS diminished both free and esterified PUFAs. Ketogenic insufficiency depleted liver PUFAs and increased triacylglycerols, mimicking human MASLD, suggesting that ketogenesis supports PUFA homeostasis, and may mitigate MASLD-MASH progression in humans.

Few studies have detailed the physiological adaptations that occur in the maternal heart. We and others have shown that pregnancy-induced cardiac growth is associated with significant changes in cardiac metabolism. Here, we examined mitochondrial respiration and substrate preference in isolated mitochondria from the maternal heart. We show that following birth, cardiac mitochondria are “primed” to respire on carbohydrate, amino acid, and ketone bodies. However, heightened respiratory efficiency is observed only with carbohydrate and amino acid sources. These results suggest that significant changes in mitochondrial respiration occur in the maternal heart in the postpartum period.

Purpose of Review: Pregnancy and exercise are systemic stressors that promote physiological growth of the heart in response to repetitive volume overload and maintenance of cardiac output. This type of remodeling is distinct from pathological hypertrophy and involves different metabolic mechanisms that facilitate growth; however, it remains unclear how metabolic changes in the heart facilitate growth and if these processes are similar in both pregnancy- and exercise-induced cardiac growth. Recent Findings: The ability of the heart to metabolize a myriad of substrates balances cardiac demands for energy provision and anabolism. During pregnancy, coordination of hormonal status with cardiac reductions in glucose oxidation appears important for physiological growth. During exercise, a reduction in cardiac glucose oxidation also appears important for physiological growth, which could facilitate shuttling of glucose-derived carbons into biosynthetic pathways for growth. Understanding the metabolic underpinnings of physiological cardiac growth could provide insight to optimize cardiovascular health and prevent deleterious remodeling, such as that which occurs from postpartum cardiomyopathy and heart failure. Summary: This short review highlights the metabolic mechanisms known to facilitate pregnancy-induced and exercise-induced cardiac growth, both of which require changes in cardiac glucose metabolism for the promotion of growth. In addition, we mention important similarities and differences of physiological cardiac growth in these models as well as discuss current limitations in our understanding of metabolic changes that facilitate growth.

E-cigarette use has surged, but the long-term health effects remain unknown. E-cigarette aerosols containing nicotine and acrolein, a combustion and e-cigarette byproduct, may impair cardiac electrophysiology through autonomic imbalance. Here we show in mouse electrocardiograms that acute inhalation of e-cigarette aerosols disturbs cardiac conduction, in part through parasympathetic modulation. We demonstrate that, similar to acrolein or combustible cigarette smoke, aerosols from e-cigarette solvents (vegetable glycerin and propylene glycol) induce bradycardia, bradyarrhythmias, and elevations in heart rate variability during inhalation exposure, with inverse post-exposure effects. These effects are slighter with tobacco- or menthol-flavored aerosols containing nicotine, and in female mice. Yet, menthol-flavored and PG aerosols also increase ventricular arrhythmias and augment early ventricular repolarization (J amplitude), while menthol uniquely alters atrial and atrioventricular conduction. Exposure to e-cigarette aerosols from vegetable glycerin and its byproduct, acrolein, diminish heart rate and early repolarization. The pro-arrhythmic effects of solvent aerosols on ventricular repolarization and heart rate variability depend partly on parasympathetic modulation, whereas ventricular arrhythmias positively associate with early repolarization dependent on the presence of nicotine. Our study indicates that chemical constituents of e-cigarettes could contribute to cardiac risk by provoking pro-arrhythmic changes and stimulating autonomic reflexes.