Low-fat Diet Mice Research Articles

Effects of Partial Knockout of Skeletal Muscle Dynamin-Related Protein 1 on Skeletal Muscle Characteristics in Lean and Obese Mice

Dynamin-related protein 1 (Drp1) is a key regulator of mitochondrial fission and plays an essential role in maintaining skeletal muscle homeostasis. Recent studies reported muscle atrophy in mice when skeletal muscle Drp1 content was reduced. However, whether such atrophic phenotype also exists in diet-induced obese mice remains unknown. We recently reported that reducing skeletal muscle Drp1 resulted in improved whole-body glucose homeostasis and insulin sensitivity in diet-induced obese mice, but not lean healthy mice, suggesting Drp1 may exert divergent effects on physiological phenotypes under different conditions. OBJECTIVE: To determine the effects of partial knockout of skeletal muscle-specific heterozygous Drp1 (mDrp1+/−) on body composition, energy expenditure and skeletal muscle physiology in lean and obese mice. HYPOTHESIS: mDrp1+/− exhibits divergent alterations in physiological characteristics between lean healthy and obese mice. Methods: Tamoxifen-inducible mDrp1+/− mouse model was generated. Nine-week-old male and female mDrp1+/− and wildtype (WT) mice (n=6-8/group) were fed with either a high-fat diet (HFD) or low-fat diet (LFD) for four weeks, received tamoxifen injections, and remained on their respective diet for another four to six weeks. A DXA scan was conducted to measure body composition. Mice were housed in Promethion metabolic cages for 48 hours to measure energy expenditure. Isolated soleus (SOL) and extensor digitorum longus (EDL) muscles were used for assessing muscle cross sectional area (CSA) and contractility. Results: HFD-fed mice had significantly higher percentage of body fat ( p = 0.045) and lower respiratory exchange ratio ( p <0.0001) than LFD-fed mice, regardless of genotype. Regarding skeletal muscle characteristics, SOL from mDrp1+/− mice exhibited larger CSA ( p = 0.052) than WT mice, regardless of diet. However, there were no differences in muscle force or fatigue resistance in SOL between groups. In contrast, EDL muscle isolated from mDrp1+/− mice fed with HFD had greater muscle force than WT mice, but interestingly the force was lower in mDrp1+/−/LFD mice than WT/LFD mice (genotype x diet interaction effect, p = 0.001). Consistently, a significant interaction was found in both twitch and tetanic rate of force development and relaxation rates (genotype x diet interaction effect, p = 0.022, p = 0.033, p = 0.040, and p = 0.020, respectively). There was no difference in EDL muscle CSA between groups. CONCLUSION: Our data suggests that partial knockout of Drp1 in skeletal muscle induced divergent effects on CSA, force production and fatigability between diet induced obese and lean healthy mice. This study was supported by the National Institute of Health (R15DK131512). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 4154: PET imaging to characterize the tumor microenvironment in a breast cancer model of obesity

Abstract Introduction: Obesity is known to reduce efficacy for cancer treatment patients and is thought to modulate the tumor microenvironment and glucose metabolism. In this study, we aim to quantify obesity-induced differences in the breast cancer tumor microenvironment with positron emission tomography (PET) imaging of hypoxia, glucose metabolism, and granzyme B. These experiments are testing the hypothesis that tumors in obese mice have increased glucose metabolism and hypoxia, measured by [18F]F-fluorodeoxyglucose (FDG) and [18F]F-fluoromisonidazole (FMISO) uptake. We also hypothesize that immunotherapy (IMT) increases immune activation, as measured by [68Ga]Ga-Granzyme B peptide (GZP), and that obese mice will respond less to immunotherapy than lean mice. Methods: C57/Bl6 mice were placed on a high-fat diet (HFD) with 60% kcal fat (n=18) or low-fat diet (LFD) composed of 10% kcal fat (n=18) to induce obesity. Blood serum was collected for analysis of cholesterol levels prior to tumor implantation. Body weight and food intake were measured weekly. 14 weeks after initiating each diet, 5x105 E0771 cells were implanted into the 3rd mammary fat pad and allowed to grow to 50-150mm3. Tumor-bearing mice were imaged with [18F]F-FDG and [18F]F-FMISO PET to measure glucose metabolism and hypoxia, respectively prior to treatment. Following imaging, mice began treatment with either saline (n=6/diet) or checkpoint blockade IMT with 200 ug anti-PD-1 and 100 ug anti-CTLA-4 (n=12/diet). After 3 doses of IMT, mice were imaged with [68Ga]Ga-GZP PET to measure immune activation. Tumor volume measurements and treatment administration occurred every 3 days. Metrics of standardized uptake value (SUV) including mean, max, and peak were quantified from imaging data. A one-way independent ANOVA or independent t-test was used to compare differences between groups. Results: HFD-fed mice had significantly increased body weight (p<0.0001) and serum cholesterol (p=0.005) compared to LFD-fed mice. SUVmean for tumor hypoxia (p=0.003) and glucose metabolism (p=0.02) were significantly increased at baseline in HFD-fed mice compared to LFD-fed mice. IMT increases immune activation for both LFD-fed (p=0.009) and HFD-fed mice (p=0.05). IMT treatment significantly reduces tumor burden for LFD mice (p<0.0001), but not HFD mice (p=0.096). Conclusions: Obesity plays a role in the reduction of the effectiveness of immunotherapy. At baseline, tumors of obese mice have increased tumor glucose metabolism and hypoxia. Although immunotherapy increases immune activation, this is not sufficient to reduce tumor burden or improve overall survival. Understanding obesity-induced differences in tumor biology may provide imaging-directed treatment decision making. Citation Format: Shannon E. Lynch, Corinne Crawford, Addison Hunt, Luke Sligh, Benjamin M. Larimer, Suzanne E. Lapi, Anna G. Sorace. PET imaging to characterize the tumor microenvironment in a breast cancer model of obesity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4154.

Histological Examination of the Effects of Thrombopoietin Mimetic Peptide (TMP) and High-Fat Diet on Femur Fracture Healing

Background and Hypothesis: In the US, 11.3% of the population are diabetic. Impaired bone healing is a complication of diabetes that dramatically impacts quality of life. Thus, it is imperative to find effective, low-risk treatments for patients that can accelerate fracture healing. We propose treatment of femur fractures using a thrombopoietin (TPO) analogue, TMP, will expedite healing, reduce adverse side effects compared to FDA-approved BMP-2, and improve quality of life of diabetic fracture patients.
 Experimental Design or Project Methods: Tie2CreERT+ mice were bred with Mplfl/fl mice to generate mice in which the TPO receptor (Mpl) was deleted in cells of the endothelial lineage (Tie2 expressing cells) following tamoxifen induction (3 consecutive daily 10mg/kg doses). Tie2CreERT+ ; Mplfl/fl and Tie2CreERT+ ; Mpl+/+ mice served as experimental and control mice, respectively. Eight-week-old male mice of both genotypes were placed on a low-fat diet (LFD) or high-fat-diet (HFD) for 12 weeks. One week prior to surgery, mice were injected with tamoxifen to induce Cre-recombination. Mice were then subjected to femur fracture and treated with saline or TMP (33nmol/kg/day) for the first week post-surgery. Mice were euthanized at 1-, 2-, and 4-week post-surgery and injured femurs were isolated for histological evaluation of the fracture callus size and composition.
 Results: To date only Tie2CreERT+ ; Mplfl/fl specimens have been processed. As expected, untreated HFD mice exhibited impaired fracture healing compared to similarly untreated LFD mice. As would also be expected, no differences were observed in fracture healing histological parameters between saline and TMP treated Tie2 CreERT+ ; Mplfl/fl mice at similar time points post-surgery.
 Conclusion and Potential Impact: While ongoing, this study explores the efficacy of using thrombopoietic agents for fracture healing in type 2 diabetes. If promising, thrombopoietic agents could replace, BMP-2 treatment, and may improve the quality of life for individuals experiencing impaired fracture healing.

The intestinal permeability marker FITC-dextran 4kDa should be dosed according to lean body mass in obese mice

AimsTo investigate the influence of the dose in the FITC-Dextran 4kDa (FD-4) permeability test in an obese mouse model, we tested the bodyweight dose regimen and a lean body mass-based dose regimen in high fat diet (HFD) mice and low fat diet (LFD) mice. We hypothesized that the FD-4 permeation result would be dose-dependent.MethodsThe two dose regimens were compared in HFD and LFD mice. Furthermore, we conducted a dose-response study to test the effect of a low or high dose of FD-4 in weight-stratified lean mice. Gene analysis of tight junctions was also carried out.ResultsThe FD-4 intestinal permeability test was dose-dependent as we found a significant increase in plasma levels of FD-4 in obese mice with the bodyweight dose regimen. However, this difference was not detectable with the lean body mass dose regimen, even with variability-adjusted group sizes. However, the qPCR analysis revealed a decrease in tight junction gene expression in obese mice. Furthermore, we found a dose-dependent significant increase in FD-4 measured in plasma samples in lean mice. No significant difference in intestinal weight was observed between lean and obese mice.ConclusionEvaluation of the intestinal permeability by FD-4 with the typical bodyweight dose regimen in obese mice will be confounded by the significant difference in dose given when compared to a lean control group. If the test dose is based on lean body mass, no significant difference in intestinal permeability is observed, even with large group sizes. Furthermore, we showed a dose-dependent difference in plasma FD-4 levels in lean mice. Therefore, we conclude that the dose should be based on lean body mass for the FD-4 permeability test if mice with considerable obesity differences are to be compared or to use another test with fixed doses.

Characterization of sclerostin's response within white adipose tissue to an obesogenic diet at rest and in response to acute exercise in male mice.

Introduction: It is well established that sclerostin antagonizes the anabolic Wnt signalling pathway in bone, however, its physiological role in other tissues remains less clear. This study examined the effect of a high-fat diet (HFD) on sclerostin content and downstream markers of the Wnt signaling pathway (GSK3β and β-catenin) within subcutaneous inguinal white adipose tissue (iWAT), and visceral epididymal WAT (eWAT) depots at rest and in response to acute aerobic exercise. Methods: Male C57BL/6 mice (n = 40, 18weeks of age) underwent 10 weeks of either a low-fat diet (LFD) or HFD. Within each diet group, mice were assigned to either remain sedentary (SED) or perform 2h of endurance treadmill exercise at 15mmin-1 with 5° incline (EX), creating four groups: LFD + SED (N = 10), LFD + EX (N = 10), HFD + SED (N = 10), and HFD + EX (N = 10). Serum and WAT depots were collected 2h post-exercise. Results: Serum sclerostin showed a diet-by-exercise interaction, reflecting HFD + EX mice having higher concentration than HFD + SED (+31%, p = 0.03), and LFD mice being unresponsive to exercise. iWAT sclerostin content decreased post-exercise in both 28kDa (-31%, p = 0.04) and 30kDa bands (-36%, main effect for exercise, p = 0.02). iWAT β-catenin (+44%, p = 0.03) and GSK3β content were higher in HFD mice compared to LFD (+128%, main effect for diet, p = 0.005). Monomeric sclerostin content was abolished in eWAT of HFD mice (-96%, main effect for diet, p < 0.0001), was only detectable as a 30kDa band in LFD mice and was unresponsive to exercise. β-catenin and GSK3β were both unresponsive to diet and exercise within eWAT. Conclusion: These results characterized sclerostin's content to WAT depots in response to acute exercise, which appears to be specific to a reduction in iWAT and identified a differential regulation of sclerostin's form/post-translational modifications depending on diet and WAT depot.

Intermittent fasting was superior to a low-fat diet in impacting tumor growth in a pre-clinical model of obesity-driven endometrial cancer (125)

Intermittent fasting was superior to a low-fat diet in impacting tumor growth in a pre-clinical model of obesity-driven endometrial cancer (125)

Reversing the Genomic, Epigenetic, and Triple-Negative Breast Cancer-Enhancing Effects of Obesity.

Obesity is an established risk and progression factor for triple-negative breast cancer (TNBC). Given rising global rates of obesity and TNBC, strategies to reduce the burden of obesity-driven TNBC are urgently needed. We report the genomic, epigenetic, and procancer effects of obesity are reversible by various calorie restriction regimens.

Obesity and altered angiogenic-related gene expression in endometrial cancer

ObjectivesEvaluate association between obesity and angiogenic-related gene expression in endometrial cancer (EC). Evaluate interaction between diet and metformin on angiogenic-related gene expression. MethodsWe evaluated the association between 168 human angiogenic-related genes and body mass index (BMI) in the TCGA Uterine Corpus Endometrial Carcinoma cohort (endometrioid endometrial cancer (EEC) cohort n = 290, and copy number high cohort n = 55), an independent validation cohort from Gynecologic Cancer Center of Excellence (GYN-COE) (n = 62) and corresponding 185 homologous mouse genes in an LKB1fl/flp53fl/fl mouse model of EC (n = 20). Mice received 60% of calories from fat in a high-fat diet (HFD), mimicking diet-induced obesity, versus 10% of calories from fat in a low-fat diet (LFD). After tumor growth, HFD (n = 5) and LFD (n = 5) mice were treated with metformin (200 mg/kg/day) or control. Whole transcriptome analysis of mouse tumors was performed using RNA-Seq. ResultsAt a false-discovery rate of 10%, twenty-one angiogenic-related genes were differentially expressed with respect to BMI when adjusting for grade in the TCGA EEC cohort. Evaluation of these genes in the mouse model control group revealed association between increased Edil3 expression in HFD versus LFD mice (2.5-fold change (FC); unadjusted p = 0.03). An interaction was observed for expression of Edil3 between diet and metformin treatment (unadjusted p = 0.009). Association between BMI and increased expression of EDIL3 was validated in one of four EDIL3 probesets in the GYN-COE cohort (p = 0.0011, adjusted p = 0.0342). ConclusionsObesity may promote tumor progression via differential modulation of angiogenic pathways in EEC. Our exploratory findings demonstrated that EDIL3 may be a candidate gene of interest.

1175-P: Inhibition of FBXO48 Improves Metabolic Changes Associated with NAFLD by Inhibiting Proteasomal Degradation of Phosphorylated AMPKa

Hypothesis: Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease in developed countries yet there are no FDA-approved therapies for the disease. AMPK is a cellular energy sensor that coordinates metabolic pathways to balance energy demand with production. Growing evidence suggests loss of AMPK activity contributes to NAFLD. We recently identified an orphan F-box protein, FBXO48, which mediates the ubiquitination and proteasomal degradation of phosphorylated AMPKα (pAMPK). We hypothesized that increasing pAMPK levels via inhibition of FBXO48 would improve glucose homeostasis and slow NAFLD progression. Methods: Twelve-week old male wildtype (WT) mice were fed low-fat diet (LFD) or “fast food” diet (FFD) for 24 weeks and then given vehicle (VEH) or a small molecule FBXO48 inhibitor (SMI; 5 mg/kg/d) for 8 weeks. We measured changes in body weight/composition, performed glucose tolerance tests (GTT), and collected plasma and liver for analysis. Results: SMI had no significant effect on body weight or composition, although body weight and fat mass were 2.4 and 2.1 g less in SMI FFD compared with VEH FFD mice, suggesting a modest effect on weight loss. SMI significantly improved glucose homeostasis in FFD mice and restored plasma glucose levels during GTT to those observed in LFD mice. SMI had no effect on liver triglyceride levels in LFD mice and produced a 15% non-significant reduction in FFD mice. Gene expression of inflammation markers Cd68 and Ccl2 were significantly reduced in SMI FFD compared with VEH FFD mice, as were markers of fibrosis Col1a1 and Col3a1. Conclusions: Our data suggest that increasing pAMPK levels through inhibition of FBXO48 may be a viable treatment option for improving glucose homeostasis and reducing inflammation/fibrosis associated with NAFLD. Disclosure A. Murali: None. L. R. Edmunds: None. B. Xie: None. B. Chen: Board Member; Self; Generian, Employee; Self; Generian. Y. Liu: Employee; Self; Generian. M. J. Jurczak: None. Funding National Institute of Diabetes and Digestive and Kidney Diseases (DK007052R01, DK119627)

Effects of diet, BMP-2 treatment, and femoral skeletal injury on endothelial cells derived from the ipsilateral and contralateral limbs.

Type 2 diabetes (T2D) results in physiological and structural changes in bone, contributing to poor fracture healing. T2D compromises microvascular performance, which can negatively impact bone regeneration as angiogenesis is required for new bone formation. We examined the effects of bone morphogenetic protein-2 (BMP-2) administered locally at the time of femoral segmental bone defect (SBD) surgery, and its angiogenic impacts on endothelial cells (ECs) isolated from the ipsilateral or contralateral tibia in T2D mice. Male C57BL/6 mice were fed either a low-fat diet (LFD) or high-fat diet (HFD) starting at 8 weeks. After 12 weeks, the T2D phenotype in HFD mice was confirmed via glucose and insulin tolerance testing and echoMRI, and all mice underwent SBD surgery. Mice were treated with BMP-2 (5 µg) or saline at the time of surgery. Three weeks postsurgery, bone marrow ECs were isolated from ipsilateral and contralateral tibias, and proliferation, angiogenic potential, and gene expression of the cells was analyzed. BMP-2 treatment increased EC proliferation by two fold compared with saline in LFD contralateral tibia ECs, but no changes were seen in surgical tibia EC proliferation. BMP-2 treatment enhanced vessel-like structure formation in HFD mice whereas, the opposite was observed in LFD mice. Still, in BMP-2 treated LFD mice, ipsilateral tibia ECs increased expression of CD31, FLT-1, ANGPT1, and ANGPT2. These data suggest that the modulating effects of T2D and BMP-2 on the microenvironment of bone marrow ECs may differentially influence angiogenic properties at the fractured limb versus the contralateral limb.

The effects of high fat diet, bone healing, and BMP-2 treatment on endothelial cell growth and function.

Angiogenesis is a vital process during the regeneration of bone tissue. The aim of this study was to investigate angiogenesis at the fracture site as well as at distal locations from obesity-induced type 2 diabetic mice that were treated with bone morphogenetic protein-2 (BMP-2, local administration at the time of surgery) to heal a femoral critical sized defect (CSD) or saline as a control. Mice were fed a high fat diet (HFD) to induce a type 2 diabetic-like phenotype while low fat diet (LFD) animals served as controls. Endothelial cells (ECs) were isolated from the lungs (LECs) and bone marrow (BMECs) 3weeks post-surgery, and the fractured femurs were also examined. Our studies demonstrate that local administration of BMP-2 at the fracture site in a CSD model results in complete bone healing within 3weeks for all HFD mice and 66.7% of LFD mice, whereas those treated with saline remain unhealed. At the fracture site, vessel parameters and adipocyte numbers were significantly increased in BMP-2 treated femurs, irrespective of diet. At distal sites, LEC and BMEC proliferation was not altered by diet or BMP-2 treatment. HFD increased the tube formation ability of both LECs and BMECs. Interestingly, BMP-2 treatment at the time of surgery reduced tube formation in LECs and humeri BMECs. However, migration of BMECs from HFD mice treated with BMP-2 was increased compared to BMECs from HFD mice treated with saline. BMP-2 treatment significantly increased the expression of CD31, FLT-1, and ANGPT2 in LECs and BMECs in LFD mice, but reduced the expression of these same genes in HFD mice. To date, this is the first study that depicts the systemic influence of fracture surgery and local BMP-2 treatment on the proliferation and angiogenic potential of ECs derived from the bone marrow and lungs.

Magnetic resonance angiography reveals increased arterial blood supply and tumorigenesis following high fat feeding in a mouse model of triple-negative breast cancer.

Breast cancer is the second most commonly diagnosed malignancy among women globally. Past MRI studies have linked a high animal fat diet (HAFD) to increased mammary cancer risk in the SV40Tag mouse model of triple-negative breast cancer. Here, serial MRI examines tumor progression and measures the arterial blood volume feeding mammary glands in low fat diet (LFD) or HAFD fed mice. Virgin female C3(1)SV40Tag mice (n = 8), weaned at 3 weeks old, were assigned to an LFD (n = 4, 3.7 kcal/g, 17.2% kcal from vegetable oil) or an HAFD (n = 4, 5.3 kcal/g, 60% kcal from lard) group. From ages 8 to 12 weeks, weekly fast spin echo MR images and time-of-flight (TOF) MR angiography of inguinal mammary glands were acquired at 9.4 T. Following in vivo MRI, mice were sacrificed. Inguinal mammary glands were excised and fixed for ex vivo MRI and histology. Tumor, blood, and mammary gland volumes for each time point were measured from manually traced regions of interest; tumors were classified as invasive by histopathology-blinded observers. Our analysis confirmed a strong correlation between total tumor volume and blood volume in the mammary gland. Tumor growth rates from weeks 8-12 were twice as high in HAFD-fed mice (0.42 ± 0.14/week) as in LFD-fed mice (0.21 ± 0.03/week), p < 0.004. Mammary gland blood volume growth rate was 2.2 times higher in HAFD mice (0.29 ± 0.11/week) compared with LFD mice (0.13 ± 0.06/week), p < 0.02. The mammary gland growth rate of HAFD-fed mice (0.071 ± 0.011/week) was 2.7 times larger than that of LFD-fed mice (0.026 ± 0.009/week), p < 0.01. This is the first non-invasive, in vivo MRI study to demonstrate a strong correlation between an HAFD and increased cancer burden and blood volume in mammary cancer without using contrast agents, strengthening the evidence supporting the adverse effects of an HAFD on mammary cancer. These results support the potential future use of TOF angiography to evaluate vasculature of suspicious lesions.

Inflammation associated weight cycling contributes to an obesogenic memory phenotype in adipose tissue

Abstract Energy expenditure through physical activity and dieting promotes weight loss. However, sustaining a healthy weight can be challenging and often a pattern known as weight cycling emerges. We evaluated the inflammatory status of mice undergoing weight cycling. Male C57BL/6 mice were randomized into four groups (n=10/group): low fat diet (LFD) for 32 weeks (LFD), high fat diet (HFD) for 32 weeks (HFD), LFD for 28 weeks then changed to HFD for 4 weeks (LFD-&amp;gt;H), HFD for 21 weeks then changed to LFD for 7 weeks and then changed to HFD for 4 weeks (HFD-&amp;gt;L-&amp;gt;H). LFD-&amp;gt;H and HFD-&amp;gt;L-&amp;gt;H mice did not significantly differ in body weight, body fat weight, or body fat percentage, but were significantly higher than LFD mice while remaining lower than HFD mice (p&amp;lt;0.05). In addition, HFD-&amp;gt;L-&amp;gt;H mice had smaller adipocytes of the epididymal fat pad compared to HFD and LFD-&amp;gt;H mice which both had larger adipocytes (p&amp;lt;0.05) compared to LFD. TNFα and IFNγ were higher (p&amp;lt;0.05) in the epididymal fat pad of HFD-&amp;gt;L-&amp;gt;H mice compared to LFD and LFD-&amp;gt;H mice. This was consistent with more M1-type macrophages (p&amp;lt;0.05) and cytotoxic T-cells (p&amp;lt;0.05) as indicated by ITGAX and CD8a gene expression, respectively. However, MCP1 was significantly lower (p&amp;lt;0.05) in HFD-&amp;gt;L-&amp;gt;H mice compared to LFD-&amp;gt;H. Liver histopathological and protein analysis revealed HFD-&amp;gt;L-&amp;gt;H mice had greater NASH and NAS (p&amp;lt;0.05) scores, increased total STAT3 and NF-kB (p&amp;lt;0.05) compared to LFD controls. Taken together, weight cycling can lead to aberrations in the storage of triglycerides when challenged with an increased caloric intake. This is further supported by an increase in adipose tissue inflammation in weight cycling mice which is likely contributing to the observed obesogenic memory phenotype.

Cardiac complex II activity is enhanced by fat and mediates greater mitochondrial oxygen consumption following hypoxic re-oxygenation.

Recent evidence suggests that mitochondrial complex II is an essential mediator of myocardial ischemia-reperfusion injury. The present study aimed to investigate the effects of fatty acid supplementation or high-fat diet (HFD) on cardiac mitochondrial activity. The changes of complex I and complex II activities and mitochondrial oxygen consumption rate (OCR) following hypoxia and re-oxygenation under these conditions were studied. Our results have shown that OCR (mitochondrial activity) was significantly increased with palmitoylcarnitine supplementation in mitochondria-enriched fraction from C57BL/6 mice hearts. Mitochondrial complex I activity was unaffected by palmitoylcarnitine but complex II activity was enhanced. Re-oxygenation following 30-min hypoxia transiently increased OCR but such an effect on OCR was abolished by complex II inhibitor, malonate, but not by complex I inhibitor, rotenone, despite that complex I activity was significantly increased with re-oxygenation following hypoxia in the presence of palmitoylcarnitine. Furthermore, OCR and complex II activity were significantly increased in the mitochondria from high-fat diet mice heart compared with those of normal or low-fat diet mice. Re-oxygenation to mitochondria following 30-min hypoxia increased OCR in all three groups but significantly more in HFD. Malonate abolished re-oxygenation-induced OCR increment in all groups. Our results indicate that complex II activity and OCR are enhanced with palmitoylcarnitine or in HFD mice heart. Although re-oxygenation following hypoxia enhanced complex II and complex I activities, complex II plays an important role in increasing mitochondrial activity, which may be instrumental in myocardial injury following ischemic reperfusion.

Cadmium and High-Fat Diet Disrupt Renal, Cardiac and Hepatic Essential Metals

Exposure to the environmental toxicant cadmium (Cd) contributes to the development of obesity-associated diseases. Obesity is a risk factor for a spectrum of unhealthy conditions including systemic metabolic dyshomeostasis. In the present study, the effects of whole-life exposure to environmentally-relevant concentrations of Cd on systemic essential metal distribution in adult mice fed a high-fat diet (HFD) were examined. For these studies, male and female mice were exposed to Cd-containing drinking water for >2 weeks before breeding. Pregnant mice and dams with offspring were exposed to Cd-containing drinking water. After weaning, offspring were continuously exposed to the same Cd concentration as their parents, and divided into HFD and normal (low) fat diet (LFD) groups. At 10 and 24 weeks, mice were sacrificed and blood, liver, kidney and heart harvested for metal analyses. There were significant concentration dependent increases in Cd levels in offspring with kidney > liver > heart. Sex significantly affected Cd levels in kidney and liver, with female animals accumulating more metal than males. Mice fed the HFD showed > 2-fold increase in Cd levels in the three organs compared to similarly treated LFD mice. Cadmium significantly affected essential metals levels in blood, kidney and liver. Additionally, HFD affected essential metal levels in these three organs. These findings suggest that Cd interacts with HFD to affect essential metal homeostasis, a phenomenon that may contribute to the underlying mechanism responsible for the development of obesity-associated pathologies.

Diet and Exercise Training Influence Skeletal Muscle Long-Chain acyl-CoA Synthetases

Long-chain acyl-CoA synthetases (ACSL) are implicated as regulators of oxidation and storage of fatty acids within skeletal muscle; however, to what extent diet and exercise alter skeletal muscle ACSL remains poorly understood. This study aimed to determine the effects of diet and exercise training on skeletal muscle ACSL and to examine relationships between ACSL1 and ACSL6 and fat oxidation and fat storage, respectively. Male C57BL/6J mice consumed a 60% high-fat diet (HFD) for 12 wk to induce obesity compared with low-fat diet (LFD). At week 4, mice began aerobic exercise (EX-Tr) or remained sedentary (SED) for 8 wk. At week 12, the protein abundance of five known ACSL isoforms and mRNA expression for ACSL1 and ACSL6 were measured in gastrocnemius muscle, as was skeletal muscle lipid content. Fat oxidation was measured using metabolic cage indirect calorimetry at week 10. Of the five known ACSL isoforms, four were detected at the protein level. HFD resulted in greater, yet nonsignificant, ACSL1 protein abundance (+18%, P = 0.13 vs LFD), greater ACSL6 (+107%, P < 0.01 vs LFD), and no difference in ACSL4 or ACSL5. Exercise training resulted in greater ACSL6 protein abundance in LFD mice (P = 0.05 LFD EX-Tr vs SED), whereas ACSL4 was lower after exercise training compared with sedentary, regardless of diet. Under fasted conditions, skeletal muscle ACSL1 protein abundance was not related to measures of whole-body fat oxidation. Conversely, skeletal muscle ACSL6 protein abundance was positively correlated with intramyocellular lipid content (P < 0.01, r = 0.22). We present evidence that ACSL isoforms 1, 4, and 6 may undergo regulation by HFD and/or exercise training. We further conclude that increased skeletal muscle ACSL6 may facilitate increased intramyocellular fat storage during HFD-induced obesity.

Maternal exercise before and during pregnancy alleviates metabolic dysfunction associated with high-fat diet in pregnant mice, without significant changes in gut microbiota

Although maternal exercise before and during pregnancy is beneficial, the effects of exercise on microbiota changes during pregnancy are unknown. Here we tested the hypothesis that maternal exercise before and during pregnancy would positively affect glucose homeostasis, pancreatic cell function, and gut microbiota dysbiosis in high-fat diet (HFD) fed dams. Female C57BL/6 mice were fed either a HFD or a low-fat diet (LFD) for 12 weeks. The HFD mice were split into two groups for 4 weeks prior to pregnancy initiation and throughout the pregnancy: sedentary (HFD) or exercised (HFD + Ex). Food intake, body weight, body composition, and glucose and insulin tolerance were measured. At gestation day 19, blood, pancreas, gonadal visceral and subcutaneous fat, plantaris muscle, and cecum were collected for analysis. Both HFD and HFD + Ex mice had impaired glucose clearance compared to LFD mice at 15 days of gestation. No changes were found in pancreatic α- or β-cell health. HFD + Ex mice had significantly reduced visceral fat mass, serum insulin, and leptin levels and increased high-density lipoprotein levels, compared to HFD-fed mice. In contrast to our hypothesis, microbiota diversity and composition were not different among groups. The relative abundance of five bacterial phyla, such as Firmicutes, Bacteroidetes, Verrucomicrobia, Deferribacteres, and Actinobacteria, were not significantly altered with diet or exercise during pregnancy. Our findings suggest that maternal exercise prevents excess visceral fat accumulation, hyperinsulinemia, and hyperleptinemia associated with a HFD, but not through the alterations of gut microbiota composition or diversity during pregnancy.

Whole blueberry protects pancreatic beta-cells in diet-induced obese mouse

BackgroundBlueberry is rich in bioactive substances and possesses powerful antioxidant potential, which can protect against oxidant-induced and inflammatory cell damage and cytotoxicity. The aim of this study was to determine how blueberry affects glucose metabolism and pancreatic β-cell proliferation in high fat diet (HFD)-induced obese mice.MethodsWild type male mice at age of 4 weeks received two different kinds of diets: high-fat diet (HFD) containing 60% fat or modified HFD supplemented with 4% (wt:wt) freeze-dried whole blueberry powder (HFD + B) for 14 weeks. A separate experiment was performed in mice fed with low-fat diet (LFD) containing 10% fat or modified LFD + B supplemented with 4% (wt:wt) freeze-dried whole blueberry powder. The metabolic parameters including blood glucose and insulin levels, glucose and insulin tolerances were measured.ResultsBlueberry-supplemented diet significantly increased insulin sensitivity and glucose tolerance in HFD + B mice compared to HFD mice. However, no difference was observed in blood glucose and insulin sensitivity between LFD + B and LFD mice. In addition, blueberry increased β-cell survival and prevented HFD-induced β-cell expansion. The most important finding was the observation of presence of small scattered islets in blueberry treated obese mice, which may reflect a potential role of blueberry in regenerating pancreatic β-cells.ConclusionsBlueberry-supplemented diet can prevent obesity-induced insulin resistance by improving insulin sensitivity and protecting pancreatic β-cells. Blueberry supplementation has the potential to protect and improve health conditions for both type 1 and type 2 diabetes patients.

Let-7e-5p Regulates GLP-1 Content and Basal Release From Enteroendocrine L Cells From DIO Male Mice.

Characterization of enteroendocrine L cells in diabetes is critical for better understanding of the role of glucagon-like peptide-1 (GLP-1) in physiology and diabetes. We studied L-cell transcriptome changes including microRNA (miRNA) dysregulation in obesity and diabetes. We evaluated the regulation of miRNAs through microarray analyses on sorted enteroendocrine L cells from control and obese glucose-intolerant (I-HFD) and hyperglycemic (H-HFD) mice after 16 weeks of respectively low-fat diet (LFD) or high-fat diet (HFD) feeding. The identified altered miRNAs were studied in vitro using the mouse GLUTag cell line to investigate their regulation and potential biological functions. We identified that let-7e-5p, miR-126a-3p, and miR-125a-5p were differentially regulated in L cells of obese HFD mice compared with control LFD mice. While downregulation of let-7e-5p expression was observed in both I-HFD and H-HFD mice, levels of miR-126a-3p increased and of miR-125a-5p decreased significantly only in I-HFD mice compared with controls. Using miRNA inhibitors and mimics we observed that modulation of let-7e-5p expression affected specifically GLP-1 cellular content and basal release, whereas Gcg gene expression and acute GLP-1 secretion and cell proliferation were not affected. In addition, palmitate treatment resulted in a decrease of let-7e-5p expression along with an increase in GLP-1 content and release, suggesting that palmitate acts on GLP-1 through let-7e-5p. By contrast, modulation of miR-125a-5p and miR-126a-3p in the same conditions did not affect content or secretion of GLP-1. We conclude that decrease of let-7e-5p expression in response to palmitate may constitute a compensatory mechanism contributing to maintaining constant glycemia in obese mice.

Effects of High-Fat Diet and Body Mass on Bone Morphology and Mechanical Properties in 1100 Advanced Intercross Mice.

Obesity is generally protective against osteoporosis and bone fracture. However, recent studies indicate that the influence of obesity on the skeleton is complex and can be detrimental. We evaluated the effects of a high-fat, obesogenic diet on the femur and radius of 1100 mice (males and females) from the Large-by-Small advanced intercross line (F34 generation). At age 5 months, bone morphology was assessed by microCT and mechanical properties by three-point bending. Mice raised on a high-fat diet had modestly greater cortical area, bending stiffness, and strength. Size-independent material properties were unaffected by a high-fat diet, indicating that diet influenced bone quantity but not quality. Bone size and mechanical properties were strongly correlated with body mass. However, the increases in many bone traits per unit increase in body mass were less in high-fat diet mice than low-fat diet mice. Thus, although mice raised on a high-fat diet have, on average, bigger and stronger bones than low-fat-fed mice, a high-fat diet diminished the positive relationship between body mass and bone size and whole-bone strength. The findings support the concept that there are diminishing benefits to skeletal health with increasing obesity. © 2019 American Society for Bone and Mineral Research.