Characterizing the effects of muscle-specific GSK3α/ß reduction on murine muscle contractility and metabolism in female mice.

Dysregulation of skeletal muscle morphology and metabolism is associated with chronic diseases such as obesity and type 2 diabetes. The enzyme glycogen synthase kinase 3 (GSK3) is highly involved in skeletal muscle physiology and metabolism, acting as a negative regulator of muscle size, strength, adaptive thermogenesis, and glucose homeostasis. Correspondingly, we have shown that partial knockdown (∼40%) of GSK3 specifically in skeletal muscle increases lean mass, reduces fat mass, and activates muscle-based adaptive thermogenesis via sarco(endo)plasmic reticulum Ca2+ (SERCA) uncoupling in male mice. However, the effects of GSK3 knockdown in female mice have yet to be investigated. Here, we examined the effects of muscle-specific GSK3 knockdown on body composition, muscle size and strength, and whole body metabolism in female C57BL/6J mice. Our results show that GSK3 content is higher in the female soleus versus the male soleus; however, there were no differences in the extensor digitorum longus (EDL). Furthermore, muscle-specific GSK3 knockdown did not alter body composition in female mice, nor did it alter daily energy expenditure, glucose/insulin tolerance, mitochondrial respiration, or the expression of the SERCA uncouplers sarcolipin and neuronatin. We also did not find any differences in soleus muscle size, strength, or fatigue resistance. In the EDL, we found that an increase in absolute and specific force production, but there were no differences in fatigability. Therefore, our study highlights sex differences in the response to genetic reduction of gsk3, with most of the effects previously observed in male mice being absent in females.NEW & NOTEWORTHY Here we show that partial GSK3 knockdown has minimal effects on whole body metabolism and muscle contractility in female mice. This is partly inconsistent with previous results found in male mice, which reveal a potential influence of biological sex.

Brain atrophy, reduced cerebral perfusion, arterial stiffening, and wall thickening with aging coincide with stimulus-specific changes in fMRI-BOLD responses.

The aim of this study was to investigate how aging affects blood flow and structure of the brain. It was hypothesized older individuals would have lower gray matter volume (GMV), resting cerebral blood flow (CBF0), and depressed responses to isometabolic and neurometabolic stimuli. In addition, increased carotid-femoral pulse-wave velocity (PWV), carotid intima-media thickness (IMT), and decreased brachial flow-mediated dilation (FMD) would be associated with lower CBF0, cerebrovascular reactivity (CVR), and GMV. Brain scans (magnetic resonance imaging) and cardiovascular examinations were conducted in young (age = 24 ± 3 yr, range = 22-28 yr; n = 13) and old (age = 71 ± 4 yr; range = 67-82 yr, n = 14) participants, and CBF0, CVR [isometabolic % blood oxygen level-dependent (BOLD) in response to a breath hold (BH)], brain activation patterns during a working memory task (neurometabolic %BOLD response to N-back trial), GMV, PWV, IMT, and FMD were measured. CBF0 and to a lesser extent CVRBH were lower in the old group (P ≤ 0.050); however, the increase in the %BOLD response to the memory task was not blunted (P ≥ 0.2867). Age-related differential activation patterns during the working memory task were characterized by disinhibition of the default mode network in the old group (P < 0.0001). Linear regression analyses revealed PWV, and IMT were negatively correlated with CBF0, CVRBH, and GMV across age groups, but within the old group alone only the relationships between PWV-CVRBH and IMT-GMV remained significant (P ≤ 0.0183). These findings suggest the impacts of age on cerebral %BOLD responses are stimulus specific, brain aging involves alterations in cerebrovascular and possibly neurocognitive control, and arterial stiffening and wall thickening may serve a role in cerebrovascular aging.NEW & NOTEWORTHY Cerebral perfusion was lower in old versus young adults. %Blood oxygen level-dependent (BOLD) responses to an isometabolic stimulus and gray matter volume were decreased in old versus young adults and associated with arterial stiffening and wall thickening. The increased %BOLD response to a neurometabolic stimulus appeared unaffected by age; however, the old group displayed disinhibition of the default mode network during the stimulus. Thus, age-related alterations in cerebral %BOLD responses were stimulus specific and related to arterial remodeling.

Carnosic Acid against Lung Cancer: Induction of Autophagy and Activation of Sestrin-2/LKB1/AMPK Signalling.

Non-small cell lung cancer (NSCLC) represents 80% of all lung cancer cases and is characterized by low survival rates due to chemotherapy and radiation resistance. Novel treatment strategies for NSCLC are urgently needed. Liver kinase B1 (LKB1), a tumor suppressor prevalently mutated in NSCLC, activates AMP-activated protein kinase (AMPK) which in turn inhibits mammalian target of rapamycin complex 1 (mTORC1) and activates unc-51 like autophagy activating kinase 1 (ULK1) to promote autophagy. Sestrin-2 is a stress-induced protein that enhances LKB1-dependent activation of AMPK, functioning as a tumor suppressor in NSCLC. In previous studies, rosemary (Rosmarinus officinalis) extract (RE) activated the AMPK pathway while inhibiting mTORC1 to suppress proliferation, survival, and migration, leading to the apoptosis of NSCLC cells. In the present study, we investigated the anticancer potential of carnosic acid (CA), a bioactive polyphenolic diterpene compound found in RE. The treatment of H1299 and H460 NSCLC cells with CA resulted in concentration and time-dependent inhibition of cell proliferation assessed with crystal violet staining and 3H-thymidine incorporation, and concentration-dependent inhibition of survival, assessed using a colony formation assay. Additionally, CA induced apoptosis of H1299 cells as indicated by decreased B-cell lymphoma 2 (Bcl-2) levels, increased cleaved caspase-3, -7, poly (ADP-ribose) polymerase (PARP), Bcl-2-associated X protein (BAX) levels, and increased nuclear condensation. These antiproliferative and proapoptotic effects coincided with the upregulation of sestrin-2 and the phosphorylation/activation of LKB1 and AMPK. Downstream of AMPK signaling, CA increased levels of autophagy marker light chain 3 (LC3), an established marker of autophagy; inhibiting autophagy with 3-methyladenine (3MA) blocked the antiproliferative effect of CA. Overall, these data indicate that CA can inhibit NSCLC cell viability and that the underlying mechanism of action of CA involves the induction of autophagy through a Sestrin-2/LKB1/AMPK signaling cascade. Future experiments will use siRNA and small molecule inhibitors to better elucidate the role of these signaling molecules in the mechanism of action of CA as well as tumor xenograft models to assess the anticancer properties of CA in vivo.

Low-dose lithium supplementation promotes adipose tissue browning and sarco(endo)plasmic reticulum Ca2+ ATPase uncoupling in muscle.

Sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) uncoupling in skeletal muscle and mitochondrial uncoupling via uncoupling protein 1 (UCP1) in brown/beige adipose tissue are two mechanisms implicated in energy expenditure. Here, we investigated the effects of glycogen synthase kinase 3 (GSK3) inhibition via lithium chloride (LiCl) treatment on SERCA uncoupling in skeletal muscle and UCP1 expression in adipose. C2C12 and 3T3-L1 cells treated with LiCl had increased SERCA uncoupling and UCP1 protein levels, respectively, ultimately raising cellular respiration; however, this was only observed when LiCl treatment occurred throughout differentiation. In vivo, LiCl treatment (10 mg/kg/day) increased food intake in chow-fed diet and high-fat diet (HFD; 60% kcal)-fed male mice without increasing body mass-a result attributed to elevated daily energy expenditure. In soleus muscle, we determined that LiCl treatment promoted SERCA uncoupling via increased expression of SERCA uncouplers, sarcolipin and/or neuronatin, under chow-fed and HFD-fed conditions. We attribute these effects to the GSK3 inhibition observed with LiCl treatment as partial muscle-specific GSK3 knockdown produced similar effects. In adipose, LiCl treatment inhibited GSK3 in inguinal white adipose tissue (iWAT) but not in brown adipose tissue under chow-fed conditions, which led to an increase in UCP1 in iWAT and a beiging-like effect with a multilocular phenotype. We did not observe this beiging-like effect and increase in UCP1 in mice fed a HFD, as LiCl could not overcome the ensuing overactivation of GSK3. Nonetheless, our study establishes novel regulatory links between GSK3 and SERCA uncoupling in muscle and GSK3 and UCP1 and beiging in iWAT.

Memory impairment in the D2.mdx mouse model of Duchenne muscular dystrophy is prevented by the adiponectin receptor agonist ALY688.

NEW FINDINGS: What is the central question of this study? Can adiponectin receptor agonism improve recognition memory in a mouse model of Duchenne muscular dystrophy? What is the main finding and its importance? Short-term treatment with the new adiponectin receptor agonist ALY688 improves recognition memory in D2.mdx mice. This finding suggests that further investigation into adiponectin receptor agonism is warranted, given that there remains an unmet need for clinical approaches to treat this cognitive dysfunction in people with Duchenne muscular dystrophy. ABSTRACT: Memory impairments have been well documented in people with Duchenne muscular dystrophy (DMD). However, the underlying mechanisms are poorly understood, and there is an unmet need to develop new therapies to treat this condition. Using a novel object recognition test, we show that recognition memory impairments in D2.mdx mice are completely prevented by daily treatment with the new adiponectin receptor agonist ALY688 from day 7 to 28 of age. In comparison to age-matched wild-type mice, untreated D2.mdx mice demonstrated lower hippocampal mitochondrial respiration (carbohydrate substrate), greater serum interleukin-6 cytokine content and greater hippocampal total tau and Raptor protein contents. Each of these measures was partly or fully preserved after treatment with ALY688. Collectively, these results indicate that adiponectin receptor agonism improves recognition memory in young D2.mdx mice.

Postexercise serum from humans influences the biological tug of war of APP processing in human neuronal cells.

Neurodegenerative diseases such as Alzheimer's disease (AD) are becoming more prevalent in our aging society. One specific neuropathological hallmark of this disease is the accumulation of amyloid-ß (Aß) peptides, which aggregate to form extraneuronal plaques. Increased Aß peptides are often observed well before symptoms of AD develop, highlighting the importance of targeting Aß-producing pathways early on in disease progression. Evidence indicates that exercise has the capacity to reduce Aß peptide production in the brain; however, the mechanisms remain unknown. Exercise-induced signaling mediators could be the driving force behind some of the beneficial effects observed in the brain with exercise. The purpose of this study was to examine if postexercise serum and the factors it contains can alter neuronal amyloid precursor protein (APP) processing. Human SH-SY5Y neuronal cells were differentiated with retinoic acid for 5 days and treated with 10% pre- or postexercise serum from humans for 30 min. Cells were collected for analysis of acute (30 min; n = 6) or adaptive (24 h posttreatment; n = 6) responses. There were no statistical differences in a disintegrin and metalloproteinase 10 (ADAM10) and ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) mRNA or protein expression with postexercise serum treatment at either time point. However, there was an increase in the ratio of soluble amyloid precursor protein α (sAPPα) to soluble amyloid precursor protein ß (sAPPß) protein content (P = 0.05) after 30 min of postexercise serum treatment. In addition, 30 min of postexercise serum treatment increased ADAM10 (P = 0.01) and BACE1 (P = 0.02) activity. These findings suggest that postexercise serum modulates important enzymes involved in APP processing, potentially pushing the cascade toward the nonamyloidogenic arm.

APP processing: a biochemical competition influenced by exercise-induced signaling mediators?

Neurodegenerative diseases, such as Alzheimer's disease (AD), are becoming more common in aging our society. One specific neuropathological hallmark of this disease is excessive accumulation of amyloid-ß (Aß) peptides, which can aggregate to form the plaques commonly associated with this disease. These plaques are often observed well before clinical diagnosis of AD. At the cellular level, both the production and aggregation of Aß peptides in the brain are detrimental to neuronal cell production, survival, and function, as well as often resulting in neuronal dysfunction and death. Exercise and physical activity have been shown to improve overall health, including brain health, and in the last several years there has been evidence to support that exercise may be able to regulate Aß peptide production in the brain. Exercise promotes the release of a wide array of signaling mediators from various metabolically active tissues and organs in the body. These exercise-induced signaling mediators could be the driving force behind some of the beneficial effects observed in brain with exercise. This review will aim to discuss potential exercise-induced signaling mediators with the capacity to influence various proteins involved in the formation of Aß peptide production in the brain.

Kynurenine metabolism is altered in mdx mice: a potential muscle to brain connection.

NEW FINDINGS: What is the central question in this study? Promoting muscle health with regular aerobic exercise can improve mental health through a kynurenine metabolic pathway: do conditions of muscle disease such as muscular dystrophy negatively influence this pathway? What is the main finding and its importance? The DBA/2J mdx model of Duchenne muscular dystrophy exhibits altered kynurenine metabolism with less kynurenic acid and peroxisome proliferator-activated receptor-γ coactivator 1-α and higher levels of tumour necrosis factor α mRNA - results associated with anxiety-like behaviour. ABSTRACT: Regular exercise can direct muscle kynurenine (KYN) metabolism toward the neuroprotective branch of the kynurenine pathway thereby limiting the accumulation of neurotoxic metabolites in the brain and contributing to mental resilience. However, the effect of muscle disease on KYN metabolism has not yet been investigated. Previous work has highlighted anxiety-like behaviours in approximately 25% of patients with Duchenne muscular dystrophy (DMD), possibly due to altered KYN metabolism. Here, we characterized KYN metabolism in mdx mouse models of DMD. Young (8-10 week old) DBA/2J (D2) mdx mice, but not age-matched C57BL/10 (C57) mdx mice, had lower levels of circulating kynurenic acid (KYNA) and lower KYNA:KYN ratio compared with their respective wild-type (WT) controls. While both C57 and D2 mdx mice displayed signs of anxiety-like behaviour, spending more time in the corners of the arena during a novel object recognition test, this effect was more prominent in D2 mdx mice. Correlational analysis detected a significant negative association between KYNA:KYN levels and time spent in corners in D2 mice, but not C57 mice. In extensor digitorum longus muscles from D2 mdx mice, but not C57 mdx mice, we found lowered protein levels of peroxisome proliferator-activated receptor-γ coactivator 1-α and kynurenine amino transferase-1 enzyme when compared with WT. Furthermore, D2 mdx quadriceps muscles had the highest level of tumour necrosis factor α expression, which is suggestive of enhanced inflammation. Thus, our pilot work shows that KYN metabolism is altered in D2 mdx mice, with a potential contribution from altered muscle health.

Exercise and estrogen: common pathways in Alzheimer's disease pathology.

Alzheimer's disease (AD) is a neurodegenerative disease that is characterized by progressive declines in cognitive function. Current epidemiological data indicate significant sex-linked disparities, where females have a higher risk of developing AD compared with male counterparts. This disparity necessitates further investigations to uncover the pathological and molecular factors influencing these sex differences. Although the underlying pathways behind this observed disparity remain elusive, recent research points to menopausal estrogen loss as a potential factor. Estrogen holds a significant role in amyloid precursor protein (APP) processing and overall neuronal health through the regulation of brain-derived neurotrophic factor (BDNF), a factor that is also reduced in postmenopausal women. BDNF is a known contributor to neuronal health and its reduced expression is typically linked to AD disorders. Exercise is known to increase BDNF and may provide an accessible activity for postmenopausal women to reduce their risk of AD. This review aims to discuss the relationship between estrogen, exercise, and BDNF in AD pathology.

Examining the effects of ovarian hormone loss and diet-induced obesity on Alzheimer's disease markers of amyloid-ß production and degradation.

After menopause, women experience declines in ovarian sex hormones, an event that has recently been associated with increased amyloid-ß peptides, a main feature of Alzheimer's disease. Diet-induced insulin resistance also increases amyloid-ß peptides; however, whether this process is exacerbated with ovarian sex hormone loss remains unknown. Female C57BL6/J mice received either bilateral ovariectomy (OVX; n = 20) or remained intact (n = 20) at 24 wk of age and were placed on either a low- or high-fat diet (LFD, n = 10 for OVX and intact; HFD, n = 10 for OVX and intact) for 10 wk. Independently, OVX led to increases in the amyloidogenic marker, soluble amyloid precursor protein ß (sAPPß). The HFD in combination with OVX led to lower insulin degrading enzyme (IDE) protein content and activity in the prefrontal cortex, indicative of decreased amyloid-ß degradation; however, no differences in amyloid-ß content were observed. Data from this study provide novel evidence of independent effects of peripheral insulin resistance and ovarian sex hormone loss in decreasing brain markers of amyloid-ß degradation. Furthermore, findings indicate how the loss of ovarian sex hormones can promote the formation of amyloidogenic APP cleavage products, independent of diet-induced insulin resistance.NEW & NOTEWORTHY This study provides novel insight into the effect of peripheral insulin resistance and ovarian hormone loss in decreasing brain markers of amyloid-ß degradation. Results demonstrate that ovarian hormone loss through ovariectomy increased the amyloidogenic marker, sAPPß, while the high-fat diet in combination with ovariectomy led to lower IDE protein content and activity in the prefrontal cortex, indicative of decreased amyloid-ß degradation. These original results provide important information for future targets in early AD pathogenesis.

Resveratrol, Metabolic Dysregulation, and Alzheimer's Disease: Considerations for Neurogenerative Disease.

Alzheimer's disease (AD) has traditionally been discussed as a disease where serious cognitive decline is a result of Aß-plaque accumulation, tau tangle formation, and neurodegeneration. Recently, it has been shown that metabolic dysregulation observed with insulin resistance and type-2 diabetes actively contributes to the progression of AD. One of the pathologies linking metabolic disease to AD is the release of inflammatory cytokines that contribute to the development of brain neuroinflammation and mitochondrial dysfunction, ultimately resulting in amyloid-beta peptide production and accumulation. Improving these metabolic impairments has been shown to be effective at reducing AD progression and improving cognitive function. The polyphenol resveratrol (RSV) improves peripheral metabolic disorders and may provide similar benefits centrally in the brain. RSV reduces inflammatory cytokine release, improves mitochondrial energetic function, and improves Aß-peptide clearance by activating SIRT1 and AMPK. RSV has also been linked to improved cognitive function; however, the mechanisms of action are less defined. However, there is evidence to suggest that chronic RSV-driven AMPK activation may be detrimental to synaptic function and growth, which would directly impact cognition. This review will discuss the benefits and adverse effects of RSV on the brain, highlighting the major signaling pathways and some of the gaps surrounding the use of RSV as a treatment for AD.

Low-dose lithium feeding increases the SERCA2a-to-phospholamban ratio, improving SERCA function in murine left ventricles.

NEW FINDINGS: What is the central question of this study? Inhibition of glycogen synthase kinase-3 (GSK3) has been shown to improve cardiac SERCA2a function. Lithium can inhibit GSK3, but therapeutic doses used in treating bipolar disorder can have toxic effects. It has not been determined whether subtherapeutic doses of lithium can improve cardiac SERCA function. What is the main finding and its importance? Using left ventricles from wild-type mice, we found that subtherapeutic lithium feeding for 6 weeks decreased GSK3 activity and increased cardiac SERCA function compared with control-fed mice. These findings warrant the investigation of low-dose lithium feeding in preclinical models of cardiomyopathy and heart failure to determine the therapeutic benefit of GSK3 inhibition. ABSTRACT: The sarco(endo)plasmic reticulum Ca2+ -ATPase (SERCA) pump is responsible for regulating calcium (Ca2+ ) within myocytes, with SERCA2a being the dominant isoform in cardiomyocytes. Its inhibitor, phospholamban (PLN), acts by decreasing the affinity of SERCA for Ca2+ . Changes in the SERCA2a:PLN ratio can cause Ca2+ dysregulation often seen in patients with dilated cardiomyopathy and heart failure. The enzyme glycogen synthase kinase-3 (GSK3) is known to downregulate SERCA function by decreasing the SERCA2a:PLN ratio. In this study, we sought to determine whether feeding mice low-dose lithium, a natural GSK3 inhibitor, would improve left ventricular SERCA function by altering the SERCA2a:PLN ratio. To this end, male wild-type C57BL/6J mice were fed low-dose lithium via drinking water (10 mg kg-1  day-1 LiCl for 6 weeks) and left ventricles were harvested. GSK3 activity was significantly reduced in LiCl-fed versus control-fed mice. The apparent affinity of SERCA for Ca2+ was also increased (pCa50 ; control, 6.09 ± 0.03 versus LiCl, 6.26 ± 0.04, P < 0.0001) along with a 2.0-fold increase in SERCA2a:PLN ratio in LiCl-fed versus control-fed mice. These findings suggest that low-dose lithium supplementation can improve SERCA function by increasing the SERCA2a:PLN ratio. Future studies in murine preclinical models will determine whether GSK3 inhibition via low-dose lithium could be a potential therapeutic strategy for dilated cardiomyopathy and heart failure.

Acute administration of IL-6 improves indices of hepatic glucose and insulin homeostasis in lean and obese mice.

Obesity can lead to impairments in hepatic glucose and insulin homeostasis, and although exercise is an effective treatment, the molecular targets remain incompletely understood. As IL-6 is an exercise-inducible cytokine, we aimed to identify whether IL-6 itself influences hepatic glucose and insulin homeostasis and whether this response differs during obesity. In vivo, male mice were fed a low-fat diet (LFD; 10% kcal) or a high-fat diet (HFD; 60% kcal) for 7 wk, which induced obesity and hepatic lipid accumulation. LFD- and HFD-fed mice were injected with IL-6 (400 ng, 75 min) or PBS and then with insulin (1 U/kg; ~15 min) or saline, at which point livers were collected. In both LFD- and HFD-fed mice, IL-6 decreased blood glucose and mRNA expression of gluconeogenic genes alongside increased phosphorylation of AKT in comparison to PBS controls, and this occurred without changes in circulating insulin. To determine whether this effect of IL-6 was directly on the liver, we completed in vitro isolated primary hepatocyte experiments from chow-fed mice and cultured with or without exposure to free fatty acid (250 µm palmitate and 250 µm oleate, 24 h) to induce lipid accumulation. In both control and free fatty acid-treated hepatocytes, IL-6 (20 ng/ml, 75 min) slightly attenuated insulin-stimulated (10 nM; ~15 min) AKT phosphorylation. Together, these data suggest that IL-6 may lead to improvements in indices of hepatic glucose and insulin homeostasis in vivo; however, this is likely due to an indirect effect on the hepatocyte. NEW & NOTEWORTHY In this study, we used lean and obese mice and found that a single injection of IL-6 improved glucose tolerance, decreased hepatic gluconeogenic gene expression, and increased hepatic phosphorylation of AKT. In primary hepatocytes cultured under control and lipid-laden conditions, IL-6 had a mild, but deleterious, effect on phosphorylation of AKT. Our results show that the beneficial effects of IL-6 on glucose and insulin homeostasis, in vivo, are maintained in obesity.

α-linolenic acid supplementation prevents exercise-induced improvements in white adipose tissue mitochondrial bioenergetics and whole-body glucose homeostasis in obese Zucker rats.

AIMS/HYPOTHESIS: While the underlying mechanisms in the development of insulin resistance remain inconclusive, metabolic dysfunction in both white adipose tissue (WAT) and skeletal muscle have been implicated in the process. Therefore, we investigated the independent and combined effects of α-linolenic acid (ALA) supplementation and exercise training on whole-body glucose homeostasis and mitochondrial bioenergetics within the WAT and skeletal muscle of obese Zucker rats. METHODS: We randomly assigned obese Zucker rats to receive a control diet alone or supplemented with ALA and to remain sedentary or undergo exercise training for 4 weeks (CON-Sed, ALA-Sed, CON-Ex and ALA-Ex groups). Whole-body glucose tolerance was determined in response to a glucose load. Mitochondrial content and bioenergetics were examined in skeletal muscle and epididymal WAT (eWAT). Insulin sensitivity and cellular stress were assessed by western blot. RESULTS: Exercise training independently improved whole-body glucose tolerance as well as insulin-induced signalling in muscle and WAT. However, the consumption of ALA during exercise training prevented exercise-mediated improvements in whole-body glucose tolerance. ALA consumption did not influence exercise-induced adaptations within skeletal muscle, insulin sensitivity and mitochondrial bioenergetics. In contrast, within eWAT, ALA supplementation attenuated insulin signalling, decreased mitochondrial respiration and increased the fraction of electron leak to reactive oxygen species (ROS). CONCLUSIONS/INTERPRETATION: These findings indicate that, in an obese rodent model, consumption of ALA attenuates the favourable adaptive changes of exercise training within eWAT, which consequently impacts whole-body glucose homeostasis. The direct translation to humans, however, remains to be determined.

Microvascular insulin resistance in skeletal muscle and brain occurs early in the development of juvenile obesity in pigs.

Impaired microvascular insulin signaling may develop before overt indices of microvascular endothelial dysfunction and represent an early pathological feature of adolescent obesity. Using a translational porcine model of juvenile obesity, we tested the hypotheses that in the early stages of obesity development, impaired insulin signaling manifests in skeletal muscle (triceps), brain (prefrontal cortex), and corresponding vasculatures, and that depressed insulin-induced vasodilation is reversible with acute inhibition of protein kinase Cß (PKCß). Juvenile Ossabaw miniature swine (3.5 mo of age) were divided into two groups: lean control ( n = 6) and obese ( n = 6). Obesity was induced by feeding the animals a high-fat/high-fructose corn syrup/high-cholesterol diet for 10 wk. Juvenile obesity was characterized by excess body mass, hyperglycemia, physical inactivity (accelerometer), and marked lipid accumulation in the skeletal muscle, with no evidence of overt atherosclerotic lesions in athero-prone regions, such as the abdominal aorta. Endothelium-dependent (bradykinin) and -independent (sodium nitroprusside) vasomotor responses in the brachial and carotid arteries (wire myography), as well as in the skeletal muscle resistance and 2A pial arterioles (pressure myography) were unaltered, but insulin-induced microvascular vasodilation was impaired in the obese group. Blunted insulin-stimulated vasodilation, which was reversed with acute PKCß inhibition (LY333-531), occurred alongside decreased tissue perfusion, as well as reduced insulin-stimulated Akt signaling in the prefrontal cortex, but not the triceps. In the early stages of juvenile obesity development, the microvasculature and prefrontal cortex exhibit impaired insulin signaling. Such adaptations may underscore vascular and neurological derangements associated with juvenile obesity.

Carnosol Increases Skeletal Muscle Cell Glucose Uptake via AMPK-Dependent GLUT4 Glucose Transporter Translocation.

Skeletal muscle is a major insulin-target tissue and plays an important role in glucose homeostasis. Insulin action in muscle activates the phosphatidylinositol-3 kinase (PI3K)/Akt signaling pathway causing the translocation of intracellularly stored GLUT4 glucose transporters to the plasma membrane and increased glucose uptake. Impaired insulin action in muscle results in insulin resistance and type 2 diabetes mellitus (T2DM). Activation of the energy sensor AMP-activated kinase (AMPK) increases muscle glucose uptake and the use of AMPK activators is viewed as an effective strategy to combat insulin resistance. Rosemary extract (RE) has been shown to stimulate muscle AMPK and glucose uptake, but the exact components responsible for these effects are unknown. In the current study, we investigated the effect of carnosol, a RE polyphenol, in L6 rat muscle cells. Carnosol stimulated glucose uptake in L6 myotubes in a dose- and time-dependent manner, did not affect Akt, increased AMPK phosphorylation and plasma membrane GLUT4 levels. The carnosol-stimulated glucose uptake and GLUT4 translocation was significantly reduced by the AMPK inhibitor compound C (CC). Our study is the first to show an AMPK-dependent increase in muscle glucose uptake by carnosol. Carnosol has potential as a glucose homeostasis regulating agent and deserves further study.

Filling the void: a role for exercise-induced BDNF and brain amyloid precursor protein processing.

Inactivity, obesity, and insulin resistance are significant risk factors for the development of Alzheimer's disease (AD). Several studies have demonstrated that diet-induced obesity, inactivity, and insulin resistance exacerbate the neuropathological hallmarks of AD. The aggregation of ß-amyloid peptides is one of these hallmarks. ß-Site amyloid precursor protein-cleaving enzyme 1 (BACE1) is the rate-limiting enzyme in amyloid precursor protein (APP) processing, leading to ß-amyloid peptide formation. Understanding how BACE1 content and activity are regulated is essential for establishing therapies aimed at reducing and/or slowing the progression of AD. Exercise training has been proven to reduce the risk of AD as well as decrease ß-amyloid production and BACE1 content and/or activity. However, these long-term interventions also result in improvements in adiposity, circulating metabolites, glucose tolerance, and insulin sensitivity making it difficult to determine the direct effects of exercise on brain APP processing. This review highlights this large void in our knowledge and discusses our current understanding of the direct of effect of exercise on ß-amyloid production. We have concentrated on the central role that brain-derived neurotrophic factor (BDNF) may play in mediating the direct effects of exercise on reducing brain BACE1 content and activity as well as ß-amyloid production. Future studies should aim to generate a greater understanding of how obesity and exercise can directly alter APP processing and AD-related pathologies. This knowledge could provide evidence-based hypotheses for designing therapies to reduce the risk of AD and dementia.

Reduced ATGL-mediated lipolysis attenuates ß-adrenergic-induced AMPK signaling, but not the induction of PKA-targeted genes, in adipocytes and adipose tissue.

5'-AMP-activated protein kinase (AMPK) is activated as a consequence of lipolysis and has been shown to play a role in regulation of adipose tissue mitochondrial content. Conversely, the inhibition of lipolysis has been reported to potentiate the induction of protein kinase A (PKA)-targeted genes involved in the regulation of oxidative metabolism. The purpose of the current study was to address these apparent discrepancies and to more fully examine the relationship between lipolysis, AMPK, and the ß-adrenergic-mediated regulation of gene expression. In 3T3-L1 adipocytes, the adipose tissue triglyceride lipase (ATGL) inhibitor ATGListatin attenuated the Thr(172) phosphorylation of AMPK by a ß3-adrenergic agonist (CL 316,243) independent of changes in PKA signaling. Similarly, CL 316,243-induced increases in the Thr(172) phosphorylation of AMPK were reduced in adipose tissue from whole body ATGL-deficient mice. Despite reductions in the activation of AMPK, the induction of PKA-targeted genes was intact or, in some cases, increased. Similarly, markers of mitochondrial content and respiration were increased in adipose tissue from ATGL knockout mice independent of changes in the Thr(172) phosphorylation of AMPK. Taken together, our data provide evidence that AMPK is not required for the regulation of adipose tissue oxidative capacity in conditions of reduced fatty acid release.

Prior exercise training blunts short-term high-fat diet-induced weight gain.

High-fat diets rapidly cause weight gain and glucose intolerance. We sought to determine whether these changes could be mitigated with prior exercise training. Male C57BL/6J mice were exercise-trained by treadmill running (1 h/day, 5 days/wk) for 4 wk. Twenty-four hours after the final bout of exercise, mice were provided with a high-fat diet (HFD; 60% kcal from lard) for 4 days, with no further exercise. In mice fed the HFD prior to exercise training, the results were blunted weight gain, reduced fat mass, and a slight attenuation in glucose intolerance that was mirrored by greater insulin-induced Akt phosphorylation in skeletal muscle compared with sedentary mice fed the HFD. When ad libitum-fed sedentary mice were compared with sedentary high-fat fed mice that were calorie restricted (-30%) to match the weight gain of the previously trained high-fat fed mice, the same attenuated impairments in glucose tolerance were found. Blunted weight gain was associated with a greater capacity to increase energy expenditure in trained compared with sedentary mice when challenged with a HFD. Although mitochondrial enzymes in white adipose tissue and UCP-1 protein content in brown adipose tissue were increased in previously exercised compared with sedentary mice fed a HFD, ex vivo mitochondrial respiration was not increased in either tissue. Our data suggest that prior exercise training attenuates high-fat diet-induced weight gain and glucose intolerance and is associated with a greater ability to increase energy expenditure in response to a high-fat diet.

Evidence for fatty acids mediating CL 316,243-induced reductions in blood glucose in mice.

CL 316,243, a ß3-adrenergic agonist, was developed as an antiobesity and diabetes drug and causes rapid decreases in blood glucose levels in mice. The mechanisms mediating this effect have not been fully elucidated; thus, the purpose of the current study was to examine the role of fatty acids and interleukin-6, reputed mediators of insulin secretion, in this process. To address this question, we used physiological and pharmacological approaches in combination with knockout mouse models. CL 316,243 treatment in male C57BL6 mice increased plasma fatty acids, glycerol, interleukin-6, and insulin and reduced blood glucose concentrations 2 h following injections. The ability of CL 316,243 to increase insulin and fatty acids and reduce glucose was preserved in interleukin-6-deficient mice. CL 316,243-induced drops in blood glucose occurred in parallel with increases in circulating fatty acids but prior to increases in plasma interleukin-6. CL 316,243-mediated increases in plasma insulin levels and reductions in blood glucose were attenuated when mice were pretreated with the lipase inhibitor nicotinic acid or in whole body adipose tissue triglyceride lipase knockout mice. Collectively, our findings demonstrate an important role for fatty acids in mediating the effects of CL 316,243 in mice. Not only do our results provide new insight into the mechanisms of action of CL 316,243, but they also hint at an unappreciated aspect of adipose tissue -pancreas cross-talk.