Biology subjects

Puchalska, P.

Publications and source records attributed to Puchalska, P..

7 recordsLinked to original sources

bioRxiv2023

ATP-Gated Potassium Channels Contribute to Ketogenic Diet-Mediated Analgesia in Mice.

Chronic pain is a substantial health burden and options for treating chronic pain remain minimally effective. Ketogenic diets are emerging as well-tolerated, effective therapeutic strategies in preclinical models of chronic pain, especially diabetic neuropathy. We tested whether a ketogenic diet is antinociceptive through ketone oxidation and related activation of ATP-gated potassium (KATP) channels in mice. We demonstrate that consumption of a ketogenic diet for one week reduced evoked nocifensive behaviors (licking, biting, lifting) following intraplantar injection of different noxious stimuli (methylglyoxal, cinnamaldehyde, capsaicin, or Yoda1) in mice. A ketogenic diet also decreased the expression of p-ERK, an indicator of neuronal activation in the spinal cord, following peripheral administration of these stimuli. Using a genetic mouse model with deficient ketone oxidation in peripheral sensory neurons, we demonstrate that protection against methylglyoxal-induced nociception by a ketogenic diet partially depends on ketone oxidation by peripheral neurons. Injection of tolbutamide, a KATP channel antagonist, prevented ketogenic diet-mediated antinociception following intraplantar capsaicin injection. Tolbutamide also restored the expression of spinal activation markers in ketogenic diet-fed, capsaicin-injected mice. Moreover, activation of KATP channels with the KATP channel agonist diazoxide reduced pain-like behaviors in capsaicin-injected, chow-fed mice, similar to the effects observed with a ketogenic diet. Diazoxide also reduced the number of p-ERK+ cells in capsaicin-injected mice. These data support a mechanism that includes neuronal ketone oxidation and activation of KATP channels to provide ketogenic diet-related analgesia. This study also identifies KATP channels as a new target to mimic the antinociceptive effects of a ketogenic diet.

neuroscience↗

bioRxiv2023

Ketone bodies in Right Ventricular Failure: A Unique Therapeutic Opportunity

Introductory ParagraphKetone bodies are pleotropic metabolites that play important roles in multiple biological processes ranging from bioenergetics to inflammation regulation via suppression of the NLRP3 inflammasome, and epigenetic modifications. Ketone bodies are elevated in left ventricular failure (LVF) and multiple approaches that increase ketone concentrations exert advantageous cardiac effects in rodents and humans. However, the relationships between ketone bodies and right ventricular failure (RVF) are relatively unexplored. Moreover, the cardioprotective properties of ketones in preclinical RVF are unknown. Here, we show a compensatory ketosis is absent in pulmonary arterial hypertension (PAH) patients with RVF. In the monocrotaline (MCT) rat model of PAH-mediated RVF, a dietary-induced ketosis improves RV function, suppresses NLRP3 inflammasome activation, and combats RV fibrosis. The summation of these data suggest ketogenic therapies may be particularly efficacious in RVF, and therefore future studies evaluating ketogenic interventions in human RVF are warranted.

biochemistry↗

bioRxiv2023

Metformin impairs trophoblast metabolism and differentiation in dose dependent manner

Metformin is a widely prescribed medication whose mechanism of action is not completely defined and whose role in gestational diabetes management remains controversial. In addition to increasing risks of fetal growth abnormalities and preeclampsia, gestational diabetes is associated with abnormalities in placental development including impairments in trophoblast differentiation. Given that metformin impacts cellular differentiation events in other systems, we assessed metformins impact on trophoblast metabolism and differentiation. Using established cell culture models of trophoblast differentiation, oxygen consumption rates and relative metabolite abundance were determined following 200 M (therapeutic range) and 2000 M (supra-therapeutic range) metformin treatment using Seahorse and mass-spectrometry approaches. While no differences in oxygen consumption rates or relative metabolite abundance were detected between vehicle and 200 M metformin treated cells, 2000 M metformin impaired oxidative metabolism and increased abundance of lactate and TCA cycle intermediates, -ketoglutarate, succinate, and malate. Examining differentiation, treatment with 2000 M, but not 200 M metformin, impaired HCG production and expression of multiple trophoblast differentiation markers. Overall, this work suggests that supra-therapeutic concentrations of metformin impairs trophoblast metabolism and differentiation whereas metformin concentrations in the therapeutic range do not strongly impact these processes.

cell biology↗

bioRxiv2023

Mitochondrial citrate metabolism and efflux regulates trophoblast differentiation

Cytotrophoblasts fuse to form and renew syncytiotrophoblasts necessary to maintain placental health throughout gestation. During cytotrophoblast to syncytiotrophoblast differentiation, cells undergo regulated metabolic and transcriptional reprogramming. Mitochondria play a critical role in differentiation events in cellular systems, thus we hypothesized that mitochondrial metabolism played a central role in trophoblast differentiation. In this work, we employed static and stable isotope tracing untargeted metabolomics methods along with gene expression and histone acetylation studies in an established cell culture model of trophoblast differentiation. Trophoblast differentiation was associated with increased abundance of the TCA cycle intermediates citrate and -ketoglutarate. Citrate was preferentially exported from mitochondria in the undifferentiated state but was retained to a larger extent within mitochondria upon differentiation. Correspondingly, differentiation was associated with decreased expression of the mitochondrial citrate transporter (CIC). CRISPR/Cas9 disruption of the mitochondrial citrate carrier showed that CIC is required for biochemical differentiation of trophoblasts. Loss of CIC resulted in broad alterations in gene expression and histone acetylation. These gene expression changes were partially rescued through acetate supplementation. Taken together, these results highlight a central role for mitochondrial citrate metabolism in orchestrating histone acetylation and gene expression during trophoblast differentiation.

developmental biology↗

bioRxiv2023

Ketolysis is Required for the Proper Development and Function of the Somatosensory Nervous System.

Ketogenic diets are emerging as protective interventions in preclinical and clinical models of somatosensory nervous system disorders. Additionally, dysregulation of succinyl-CoA 3-oxoacid CoA-transferase 1 (SCOT, encoded by Oxct1), the fate-committing enzyme in mitochondrial ketolysis, has recently been described in Friedreichs ataxia and amyotrophic lateral sclerosis. However, the contribution of ketone metabolism in the normal development and function of the somatosensory nervous system remains poorly characterized. We generated sensory neuron-specific, Advillin-Cre knockout of SCOT (Adv-KO-SCOT) mice and characterized the structure and function of their somatosensory system. We used histological techniques to assess sensory neuronal populations, myelination, and skin and spinal dorsal horn innervation. We also examined cutaneous and proprioceptive sensory behaviors with the von Frey test, radiant heat assay, rotarod, and grid-walk tests. Adv-KO-SCOT mice exhibited myelination deficits, altered morphology of putative A{delta} soma from the dorsal root ganglion, reduced cutaneous innervation, and abnormal innervation of the spinal dorsal horn compared to wildtype mice. Synapsin 1-Cre-driven knockout of Oxct1 confirmed deficits in epidermal innervation following a loss of ketone oxidation. Loss of peripheral axonal ketolysis was further associated with proprioceptive deficits, yet Adv-KO-SCOT mice did not exhibit drastically altered cutaneous mechanical and thermal thresholds. Knockout of Oxct1 in peripheral sensory neurons resulted in histological abnormalities and severe proprioceptive deficits in mice. We conclude that ketone metabolism is essential for the development of the somatosensory nervous system. These findings also suggest that decreased ketone oxidation in the somatosensory nervous system may explain the neurological symptoms of Friedreichs ataxia.

neuroscience↗

bioRxiv2022

Loss of hepatic phosphoenolpyruvate carboxykinase 1 dysregulates metabolic responses to acute exercise but enhances adaptations to exercise training in mice

Acute exercise increases liver gluconeogenesis to supply glucose to working muscle. Concurrently, elevated liver lipid breakdown fuels the high energetic cost of gluconeogenesis. This functional coupling between liver gluconeogenesis and lipid oxidation has been proposed to underlie the ability of regular exercise to enhance liver mitochondrial oxidative metabolism and decrease liver steatosis in individuals with non-alcoholic fatty liver disease. Herein we tested whether repeated bouts of increased hepatic gluconeogenesis are necessary for exercise training to lower liver lipids. Experiments used diet-induced obese mice lacking hepatic phosphoenolpyruvate carboxykinase 1 (KO) to inhibit gluconeogenesis and wild type (WT) littermates. 2H/13C metabolic flux analysis quantified glucose and mitochondrial oxidative fluxes in untrained mice at rest and during acute exercise. Circulating and tissue metabolite levels were determined during sedentary conditions, acute exercise, and refeeding post-exercise. Mice also underwent six weeks of treadmill running protocols to define hepatic and extrahepatic adaptations to exercise training. Untrained KO mice were unable to maintain euglycemia during acute exercise resulting from an inability to increase gluconeogenesis. Liver triacylglycerides were elevated following acute exercise and circulating {beta}-hydroxybutyrate was higher during post-exercise refeeding in untrained KO mice. In contrast, exercise training prevented liver triacylglyceride accumulation in KO mice. This was accompanied by pronounced increases in indices of skeletal muscle mitochondrial oxidative metabolism in KO mice. Together, these results show that hepatic gluconeogenesis is dispensable for exercise training to reduce liver lipids. This may be due to responses in ketone body metabolism and/or metabolic adaptations in skeletal muscle to exercise.

physiology↗

bioRxiv2022

Ketolysis is a metabolic driver of CD8+ T cell effector function through histone acetylation

Environmental nutrient availability influences T cell metabolism, impacting T cell function and shaping immune outcomes. However, the metabolic pathways critical for optimal T cell responses remain poorly understood. Here, we identify ketone bodies (KBs) - including {beta}-hydroxybutyrate ({beta}OHB) and acetoacetate (AcAc) - as essential fuels supporting CD8+ T cell metabolism and effector function. Ketolysis is an intrinsic feature of highly functional CD8+ T effector (Teff) cells and {beta}OHB directly increases CD8+ Teff cell IFN-{gamma} production and cytolytic activity. Using metabolic tracers, we establish that CD8+ Teff cells preferentially use KBs over glucose to fuel the tricarboxylic acid (TCA) cycle in vitro and in vivo. KBs directly boost the respiratory capacity of CD8+ T cells and TCA cycle-dependent metabolic pathways that fuel T cell growth. Mechanistically, we find that {beta}OHB is a major substrate for acetyl-CoA production in CD8+ T cells and regulates effector responses through effects on histone acetylation. Together, our results identify cell-intrinsic ketolysis as a metabolic and epigenetic driver of optimal CD8+ T cell effector responses. One Sentence summaryKetone bodies promote CD8+ T cell metabolism and effector function through regulation of epigenetic programming

immunology↗