VCD-induced menopause mouse model reveals reprogramming of hepatic metabolism.

OBJECTIVE: Menopause adversely impacts systemic energy metabolism and increases the risk of metabolic disease(s) including hepatic steatosis, but the mechanisms are largely unknown. Dosing female mice with vinyl cyclohexene dioxide (VCD) selectively causes follicular atresia in ovaries, leading to a murine menopause-like phenotype. METHODS: In this study, we treated female C57BL6/J mice with VCD (160 mg/kg i.p. for 20 consecutive days followed by verification of the lack of estrous cycling) to investigate changes in body composition, energy expenditure (EE), hepatic mitochondrial function, and hepatic steatosis across different dietary conditions. RESULTS: VCD treatment induced ovarian follicular loss and increased follicle-stimulating hormone (FSH) levels in female mice, mimicking a menopause-like phenotype. VCD treatment did not affect body composition, or EE in mice on a low-fat diet (LFD) or in response to a short-term (1-week) high-fat, high sucrose diet (HFHS). However, the transition to a HFHS lowered cage activity in VCD mice. A chronic HFHS diet (16 weeks) significantly increased weight gain, fat mass, and hepatic steatosis in VCD-treated mice compared to HFHS-fed controls. In the liver, VCD mice showed suppressed hepatic mitochondrial respiration on LFD, while chronic HFHS resulted in compensatory increases in hepatic mitochondrial respiration. Also, liver RNA sequencing revealed that VCD promoted global upregulation of hepatic lipid/cholesterol synthesis pathways. CONCLUSION: Our findings suggest that the VCD-induced menopause model compromises hepatic mitochondrial function and lipid/cholesterol homeostasis that sets the stage for HFHS diet-induced steatosis while also increasing susceptibility to obesity.

Divergence in aerobic capacity and energy expenditure influence metabolic tissue mitochondrial protein synthesis rates in aged rats.

Age-associated declines in aerobic capacity promote the development of various metabolic diseases. In rats selectively bred for high/low intrinsic aerobic capacity, greater aerobic capacity reduces susceptibility to metabolic disease while increasing longevity. However, little remains known how intrinsic aerobic capacity protects against metabolic disease, particularly with aging. Here, we tested the effects of aging and intrinsic aerobic capacity on systemic energy expenditure, metabolic flexibility and mitochondrial protein synthesis rates using 24-month-old low-capacity (LCR) or high-capacity runner (HCR) rats. Rats were fed low-fat diet (LFD) or high-fat diet (HFD) for eight weeks, with energy expenditure (EE) and metabolic flexibility assessed utilizing indirect calorimetry during a 48 h fast/re-feeding metabolic challenge. Deuterium oxide (D2O) labeling was used to assess mitochondrial protein fraction synthesis rates (FSR) over a 7-day period. HCR rats possessed greater EE during the metabolic challenge. Interestingly, HFD induced changes in respiratory exchange ratio (RER) in male and female rats, while HCR female rat RER was largely unaffected by diet. In addition, analysis of protein FSR in skeletal muscle, brain, and liver mitochondria showed tissue-specific adaptations between HCR and LCR rats. While brain and liver protein FSR were altered by aerobic capacity and diet, these effects were less apparent in skeletal muscle. Overall, we provide evidence that greater aerobic capacity promotes elevated EE in an aged state, while also regulating metabolic flexibility in a sex-dependent manner. Modulation of mitochondrial protein FSR by aerobic capacity is tissue-specific with aging, likely due to differential energetic requirements by each tissue.

Divergence in aerobic capacity influences hepatic and systemic metabolic adaptations to bile acid sequestrant and short-term high-fat/sucrose feeding in rats.

High versus low aerobic capacity significantly impacts the risk for metabolic diseases. Rats selectively bred for high or low intrinsic aerobic capacity differently modify hepatic bile acid metabolism in response to high-fat diets (HFDs). Here we tested if a bile acid sequestrant would alter hepatic and whole body metabolism differently in rats with high and low aerobic capacity fed a 1-wk HFD. Male rats (8 mo of age) that were artificially selected to be high (HCR) and low-capacity runners (LCR) with divergent intrinsic aerobic capacities were transitioned from a low-fat diet (LFD, 10% fat) to an HFD (45% fat) with or without a bile acid sequestrant (BA-Seq, 2% cholestyramine resin) for 7 days while maintained in an indirect calorimetry system. HFD + BA-Seq increased fecal excretion of lipids and bile acids and prevented weight and fat mass gain in both strains. Interestingly, HCR rats had increased adaptability to enhance fecal bile acid and lipid loss, resulting in more significant energy loss than their LCR counterpart. In addition, BA-Seq induced a greater expression of hepatic CYP7A1 gene expression, the rate-limiting enzyme of bile acid synthesis in HCR rats both on HFD and HFD + BA-Seq diets. HCR displayed a more significant reduction of RQ in response to HFD than LCR, but HFD + BA-Seq lowered RQ in both groups compared with HFD alone, demonstrating a pronounced impact on metabolic flexibility. In conclusion, BA-Seq provides uniform metabolic benefits for metabolic flexibility and adiposity, but rats with higher aerobic capacity display adaptability for hepatic bile acid metabolism.NEW & NOTEWORTHY The administration of bile acid sequestrant (BA-Seq) has uniform metabolic benefits in terms of metabolic flexibility and adiposity in rats with high and low aerobic capacity. However, rats with higher aerobic capacity demonstrate greater adaptability in hepatic bile acid metabolism, resulting in increased fecal bile acid and lipid loss, as well as enhanced fecal energy loss.

VCD-induced menopause mouse model reveals reprogramming of hepatic metabolism.

Menopause adversely impacts systemic energy metabolism and increases the risk of metabolic disease(s) including hepatic steatosis, but the mechanisms are largely unknown. Dosing female mice with vinyl cyclohexene dioxide (VCD) selectively causes follicular atresia in ovaries, leading to a murine menopause-like phenotype. In this study, we treated female C57BL6/J mice with VCD (160mg/kg i.p. for 20 consecutive days followed by verification of the lack of estrous cycling) to investigate changes in body composition, energy expenditure (EE), hepatic mitochondrial function, and hepatic steatosis across different dietary conditions. VCD treatment induced ovarian follicular loss and increased follicle-stimulating hormone (FSH) levels in female mice, mimicking a menopause-like phenotype. VCD treatment did not affect body composition, or EE in mice on a low-fat diet or in response to a short-term (1-week) high-fat, high sucrose diet (HFHS). However, the transition to a HFHS lowered cage activity in VCD mice. A chronic HFHS diet (16 weeks) significantly increased weight gain, fat mass, and hepatic steatosis in VCD-treated mice compared to HFHS-fed controls. In the liver, VCD mice showed suppressed hepatic mitochondrial respiration on LFD, while chronic HFHS diet resulted in compensatory increases in hepatic mitochondrial respiration. Also, liver RNA sequencing revealed that VCD promoted global upregulation of hepatic lipid/cholesterol synthesis pathways. Our findings suggest that the VCD- induced menopause model compromises hepatic mitochondrial function and lipid/cholesterol homeostasis that sets the stage for HFHS diet-induced steatosis while also increasing susceptibility to obesity.

Acute exercise dynamically modulates the hepatic mitochondrial proteome.

Exercise powerfully increases energy metabolism and substrate flux in tissues, a process reliant on dramatic changes in mitochondrial energetics. Liver mitochondria play a multi-factorial role during exercise to fuel hepatic glucose output. We previously showed acute exercise activates hepatic mitophagy, a pathway to recycle low-functioning/damaged mitochondria, however little is known how individual bouts of exercise alters the hepatic mitochondrial proteome. Here we leveraged proteomics to examine changes in isolated hepatic mitochondria both immediately after and 2 hours post an acute, 1 hour bout of treadmill exercise in female mice. Further, we utilized leupeptin, a lysosomal inhibitor, to capture and measure exercise-induced changes in mitochondrial proteins that would have been unmeasured due to their targeting for lysosomal degradation. Proteomic analysis of enriched hepatic mitochondria identified 3241 total proteins. Functional enrichment analysis revealed robust enrichment for proteins critical to the mitochondria including metabolic pathways, tricarboxylic acid cycle, and electron transport system. Compared to the sedentary condition, exercise elevated processes regulating lipid localization, Il-5 signaling, and protein phosphorylation in isolated mitochondria. t-SNE analysis identified 4 unique expressional clusters driven by time-dependent changes in protein expression. Isolation of proteins significantly altered with exercise from each cluster revealed influences of leupeptin and exercise both independently and cooperatively modulating mitochondrial protein expressional profiles. Overall, we provide evidence that acute exercise rapidly modulates changes in the proteins/pathways of isolated hepatic mitochondria that include fatty acid metabolism/storage, post-translational protein modification, inflammation, and oxidative stress. In conclusion, the hepatic mitochondrial proteome undergoes extensive remodeling with a bout of exercise.

Mitochondrial function and Aß in Alzheimer's disease postmortem brain.

INTRODUCTION: Mitochondrial dysfunction is observed in Alzheimer's disease (AD). However, the relationship between functional mitochondrial deficits and AD pathologies is not well established in human subjects. METHODS: Post-mortem human brain tissue from 11 non-demented (ND) and 12 AD subjects was used to examine mitochondrial electron transport chain (ETC) function. Data were analyzed by neuropathology diagnosis and Apolipoprotein E (APOE) genotype. Relationships between AD pathology and mitochondrial function were determined. RESULTS: AD subjects had reductions in brain cytochrome oxidase (COX) function and complex II Vmax. APOE ε4 carriers had COX, complex II and III deficits. AD subjects had reduced expression of Complex I-III ETC proteins, no changes were observed in APOE ε4 carriers. No correlation between p-Tau Thr 181 and mitochondrial outcomes was observed, although brains from non-demented subjects demonstrated positive correlations between Aß concentration and COX Vmax. DISCUSSION: These data support a dysregulated relationship between brain mitochondrial function and Aß pathology in AD.

Acute exercise rapidly activates hepatic mitophagic flux.

Exercise is critical for improving metabolic health and putatively maintains or enhances mitochondrial quality control in metabolic tissues. Although previous work has shown that exercise elicits hepatic mitochondrial biogenesis, it is unknown if acute exercise activates hepatic mitophagy, the selective degradation of damaged or low-functioning mitochondria. We tested if an acute bout of treadmill running increased hepatic mitophagic flux both right after and 2-h postexercise in 15- to 24-wk-old C57BL/6J female mice. Acute exercise did not significantly increase markers of autophagic flux, however, mitophagic flux was activated 2-h post-treadmill running as measured by accumulation of both LC3-II and p62 in isolated mitochondria in the presence of leupeptin, an inhibitor of autophagosome degradation. Furthermore, mitochondrial-associated ubiquitin, which recruits the autophagy receptor protein p62, was also significantly increased at 2 h. Further examination via Western blot and proteomics analysis revealed that acute exercise elicits a time-dependent, dynamic activation of mitophagy pathways. Moreover, the results suggest that exercise-induced hepatic mitophagy is likely mediated by both polyubiquitination and receptor-mediated signaling pathways. Overall, we provide evidence that acute exercise activates hepatic mitophagic flux while also revealing specific receptor-mediated proteins by which exercise maintains mitochondrial quality control in the liver.NEW & NOTEWORTHY This study provides evidence that acute exercise activates hepatic mitophagic flux and mitochondrial polyubiquitination while additionally revealing specific receptor-mediated proteins by which exercise maintains mitochondrial quality control in the liver.

Heat Treatment Improves Hepatic Mitochondrial Respiratory Efficiency via Mitochondrial Remodeling.

Nonacholic fatty liver disease, or hepatic steatosis, is the most common liver disorder affecting the western world and currently has no pharmacologic cure. Thus, many investigations have focused on alternative strategies to treat or prevent hepatic steatosis. Our laboratory has shown that chronic heat treatment (HT) mitigates glucose intolerance, insulin resistance, and hepatic steatosis in rodent models of obesity. Here, we investigate the direct bioenergetic mechanism(s) surrounding the metabolic effects of HT on hepatic mitochondria. Utilizing mitochondrial proteomics and respiratory function assays, we show that one bout of acute HT (42°C for 20 min) in male C57Bl/6J mice (n = 6/group) triggers a hepatic mitochondrial heat shock response resulting in acute reductions in respiratory capacity, degradation of key mitochondrial enzymes, and induction of mitophagy via mitochondrial ubiquitination. We also show that chronic bouts of HT and recurrent activation of the heat shock response enhances mitochondrial quality and respiratory function via compensatory adaptations in mitochondrial organization, gene expression, and transport even during 4 weeks of high-fat feeding (n = 6/group). Finally, utilizing a liver-specific heat shock protein 72 (HSP72) knockout model, we are the first to show that HSP72, a protein putatively driving the HT metabolic response, does not play a significant role in the hepatic mitochondrial adaptation to acute or chronic HT. However, HSP72 is required for the reductions in blood glucose observed with chronic HT. Our data are the first to suggest that chronic HT (1) improves hepatic mitochondrial respiratory efficiency via mitochondrial remodeling and (2) reduces blood glucose in a hepatic HSP72-dependent manner.