Melissa M Thomas

The University of Calgary, Calgary, Alberta, Canada

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Publications (11)48.6 Total impact

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    ABSTRACT: AMP-activated protein kinase (AMPK) is a master regulator of metabolism. While muscle-specific AMPK β1β2 double-knockout (β1β2M-KO) mice display alterations in metabolic and mitochondrial capacity, their severe exercise intolerance suggested a secondary contributor to the observed phenotype. We find that tibialis anterior (TA), but not soleus, muscles of sedentary β1β2M-KO mice display a significant myopathy (decreased myofiber areas, increased split and necrotic myofibers, and increased centrally nucleated myofibers. A mitochondrial- and fiber-type-specific etiology to the myopathy was ruled out. However, β1β2M-KO TA muscles displayed significant (P<0.05) increases in platelet aggregation and apoptosis within myofibers and surrounding interstitium (P<0.05). These changes correlated with a 45% decrease in capillary density (P<0.05). We hypothesized that the β1β2M-KO myopathy in resting muscle resulted from impaired AMPK-nNOSμ signaling, causing increased platelet aggregation, impaired vasodilation, and, ultimately, ischemic injury. Consistent with this hypothesis, AMPK-specific phosphorylation (Ser1446) of nNOSμ was decreased in β1β2M-KO compared to wild-type (WT) mice. The AMPK-nNOSμ relationship was further demonstrated by administration of 5-aminoimidazole-4-carboxamide 1-β-d-ribofuranoside (AICAR) to β1β2-MKO muscles and C2C12 myotubes. AICAR significantly increased nNOSμ phosphorylation and nitric oxide production (P<0.05) within minutes of administration in WT muscles and C2C12 myotubes but not in β1β2M-KO muscles. These findings highlight the importance of the AMPK-nNOSμ pathway in resting skeletal muscle.-Thomas, M. M., Wang, D. C., D'Souza, D. M., Krause, M. P., Layne, A. S., Criswell, D. S., O'Neill, H. M., Connor, M. K., Anderson, J. E., Kemp, B. E., Steinberg, G. R., and Hawke, T. J. Muscle-specific AMPK β1β2-null mice display a myopathy due to loss of capillary density in nonpostural muscles.
    The FASEB Journal 02/2014; · 5.70 Impact Factor
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    ABSTRACT: While it has long been postulated that exercise training attenuates the age-related decline in heart function normally associated with increased fibrosis and collagen cross-linking, the potential benefits associated with exercise training initiated later in life are currently unclear. To address this question, Fisher 344× Brown Norway F1 rats underwent treadmill-based exercise training starting in late middle age and continued into senescence (35months) and were compared with age-matched sedentary rats. Hearts were examined for fibrosis and advanced glycation end-products in the subendocardial layer of left ventricular cross-sections. Genes for collagen synthesis and degradation were assessed by polymerase chain reaction, and matrix metalloproteinase (MMP) activity was assessed by EnzChek® Gelatinase/Collagenase Assay Kit. Exercise training of late middle-aged rats attenuated fibrosis and collagen cross-linking, while also reducing age-related mortality between late middle age and senescence. This training was also associated with an attenuated advanced glycation end-product (AGE) accumulation with aging, suggesting a decrease in collagen cross-linking. Conversely, tissue inhibitor of matrix metalloproteinase-1 (TIMP1) gene expression, TIMP and MMP1 protein expression, and MMP activity increased with age but were not significantly impacted by exercise training. While our results demonstrate that exercise training in late middle age attenuates age-related mortality and cardiac fibrosis and is accompanied by attenuated AGE accumulation indicative of less collagen cross-linking, the mechanisms explaining this attenuated replacement fibrosis did not appear to involve altered TIMP1 expression, or MMP protein and activity.
    Experimental gerontology 11/2013; · 3.34 Impact Factor
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    ABSTRACT: Diet-induced obesity is a rising health concern which can lead to the development of glucose intolerance and muscle insulin resistance and, ultimately, type II diabetes mellitus. This research investigates the associations between glucose intolerance or muscle insulin resistance and tissue specific changes during the progression of diet-induced obesity. C57BL/6J mice were fed a normal or high-fat diet (HFD; 60% kcal fat) for 3 or 8 weeks. Disease progression was monitored by measurements of body/tissue mass changes, glucose and insulin tolerance tests, and ex vivo glucose uptake in intact muscles. Lipid metabolism was analyzed using metabolic chambers and ex vivo palmitate assays in intact muscles. Skeletal muscle, liver and adipose tissues were analyzed for changes in inflammatory gene expression. Plasma was analyzed for insulin levels and inflammatory proteins. Histological techniques were used on muscle and liver cryosections to assess metabolic and morphological changes. A rapid shift in whole body metabolism towards lipids was observed with HFD. Following 3 weeks of HFD, elevated total lipid oxidation and an oxidative fiber type shift had occurred in the skeletal muscle, which we propose was responsible for delaying intramyocellular lipid accumulation and maintaining muscle's insulin sensitivity. Glucose intolerance was present after three weeks of HFD and was associated with an enlarged adipose tissue depot, adipose tissue inflammation and excess hepatic lipids, but not hepatic inflammation. Furthermore, HFD did not significantly increase systemic or muscle inflammation after 3 or 8 weeks of HFD suggesting that early diet-induced obesity does not cause inflammation throughout the whole body. Overall these findings indicate skeletal muscle did not contribute to the development of HFD-induced impairments in whole-body glucose tolerance following 3 weeks of HFD.
    PLoS ONE 01/2013; 8(8):e71747. · 3.73 Impact Factor
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    ABSTRACT: The aging heart is characterized by a progressive decline in contractile function and diastolic relaxation. Amongst the factors implicated in these changes is a progressive replacement fibrosis secondary to cardiomyocyte death, oxidative damage, and energetic deficit, each of which may be secondary to impaired mitochondrial function. Here, we performed an in-depth examination of mitochondrial function in saponin-permeabilized cardiomyocyte bundles, a preparation where all mitochondria are represented and their structure intact, from young adult (YA) and senescent (SEN) rats (n = 8 per group). When accounting for increased fibrosis (+19%, P<0.01) and proportional decrease in citrate synthase activity in the SEN myocardium (-23%, P<0.05), mitochondrial respiration and reactive oxygen species (H(2)O(2)) emission across a range of energized states was similar between age groups. Accordingly, the abundance of electron transport chain proteins was also unchanged. Likewise, except for CuZnSOD (-37%, P<0.05), the activity of antioxidant enzymes was unaltered with aging. Although time to mitochondrial permeability transition pore (mPTP) opening was decreased (-25%, P<0.05) in the SEN heart, suggesting sensitization to apoptotic stimuli, this was not associated with a difference in apoptotic index measured by ELISA. Collectively, our results suggest that the function of existing cardiac ventricular mitochondria is relatively preserved in SEN rat heart when measured in permeabilized cells.
    PLoS ONE 01/2012; 7(8):e43003. · 3.73 Impact Factor
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    ABSTRACT: To determine whether mitochondrial dysfunction is causally related to muscle atrophy with aging, we examined respiratory capacity, H(2) O(2) emission, and function of the mitochondrial permeability transition pore (mPTP) in permeabilized myofibers prepared from four rat muscles that span a range of fiber type and degree of age-related atrophy. Muscle atrophy with aging was greatest in fast-twitch gastrocnemius (Gas) muscle (-38%), intermediate in both the fast-twitch extensor digitorum longus (EDL) and slow-twitch soleus (Sol) muscles (-21%), and non-existent in adductor longus (AL) muscle (+47%). In contrast, indices of mitochondrial dysfunction did not correspond to this differential degree of atrophy. Specifically, despite higher protein expression for oxidative phosphorylation (oxphos) system in fast Gas and EDL, state III respiratory capacity per myofiber wet weight was unchanged with aging, whereas the slow Sol showed proportional decreases in oxphos protein, citrate synthase activity, and state III respiration. Free radical leak (H(2) O(2) emission per O(2) flux) under state III respiration was higher with aging in the fast Gas, whereas state II free radical leak was higher in the slow AL. Only the fast muscles had impaired mPTP function with aging, with lower mitochondrial calcium retention capacity in EDL and shorter time to mPTP opening in Gas and EDL. Collectively, our results underscore that the age-related changes in muscle mitochondrial function depend largely upon fiber type and are unrelated to the severity of muscle atrophy, suggesting that intrinsic changes in mitochondrial function are unlikely to be causally involved in aging muscle atrophy.
    Aging cell 09/2011; 10(6):1047-55. · 7.55 Impact Factor
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    ABSTRACT: The senescent heart has decreased systolic and diastolic functions, both of which could be related to alterations in cardiac sarcoplasmic reticulum (SR) calcium (Ca(2+)) handling. The purpose of this study was to determine if SR protein content and rates of Ca(2+) release and uptake and ATPase activity are lower in the senescent (34-36 mo) Fisher 344×Brown-Norway F1 hybrid rat heart and if a long-term exercise training program could maintain SR function. Late middle aged (29 mo) male rats underwent 5-7 mo of treadmill training. Aging resulted in a decrease in SERCA activity and modest decrease in the rate of Ca(2+) uptake but no change in Ca(2+) release rate. SERCA2a content was not decreased with age but nitrotyrosine accumulation was increased and Ser16 phosphorylated phospholamban (PLN) was decreased. Ryanodine receptor content was not decreased with age but dihydropyridine receptor content was decreased in the senescent heart. Treadmill training had no significant effect on any of the SR properties or protein contents in the senescent rat heart. These results suggest that decreases in Ca(2+) uptake and SERCA activity in the senescent F344BN rat heart are due to increased SERCA2a damage from nitrotyrosine accumulation and inhibition by PLN and that exercise training initiated at late middle age is unable to prevent these age-related changes in cardiac SR function.
    Experimental gerontology 07/2011; 46(10):803-10. · 3.34 Impact Factor
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    ABSTRACT: Mitochondria regulate critical components of cellular function via ATP production, reactive oxygen species production, Ca(2+) handling and apoptotic signaling. Two classical methods exist to study mitochondrial function of skeletal muscles: isolated mitochondria and permeabilized myofibers. Whereas mitochondrial isolation removes a portion of the mitochondria from their cellular environment, myofiber permeabilization preserves mitochondrial morphology and functional interactions with other intracellular components. Despite this, isolated mitochondria remain the most commonly used method to infer in vivo mitochondrial function. In this study, we directly compared measures of several key aspects of mitochondrial function in both isolated mitochondria and permeabilized myofibers of rat gastrocnemius muscle. Here we show that mitochondrial isolation i) induced fragmented organelle morphology; ii) dramatically sensitized the permeability transition pore sensitivity to a Ca(2+) challenge; iii) differentially altered mitochondrial respiration depending upon the respiratory conditions; and iv) dramatically increased H(2)O(2) production. These alterations are qualitatively similar to the changes in mitochondrial structure and function observed in vivo after cellular stress-induced mitochondrial fragmentation, but are generally of much greater magnitude. Furthermore, mitochondrial isolation markedly altered electron transport chain protein stoichiometry. Collectively, our results demonstrate that isolated mitochondria possess functional characteristics that differ fundamentally from those of intact mitochondria in permeabilized myofibers. Our work and that of others underscores the importance of studying mitochondrial function in tissue preparations where mitochondrial structure is preserved and all mitochondria are represented.
    PLoS ONE 01/2011; 6(3):e18317. · 3.73 Impact Factor
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    ABSTRACT: Mitochondria regulate cellular bioenergetics and apoptosis and have been implicated in aging. However, it remains unclear whether age-related loss of muscle mass, known as sarcopenia, is associated with abnormal mitochondrial function. Two technically different approaches have mainly been used to measure mitochondrial function: isolated mitochondria and permeabilized myofiber bundles, but the reliability of these measures in the context of sarcopenia has not been systematically assessed before. A key difference between these approaches is that contrary to isolated mitochondria, permeabilized bundles contain the totality of fiber mitochondria where normal mitochondrial morphology and intracellular interactions are preserved. Using the gastrocnemius muscle from young adult and senescent rats, we show marked effects of aging on three primary indices of mitochondrial function (respiration, H(2) O(2) emission, sensitivity of permeability transition pore to Ca(2+) ) when measured in isolated mitochondria, but to a much lesser degree when measured in permeabilized bundles. Our results clearly demonstrate that mitochondrial isolation procedures typically employed to study aged muscles expose functional impairments not seen in situ. We conclude that aging is associated with more modest changes in mitochondrial function in sarcopenic muscle than suggested previously from isolated organelle studies.
    Aging cell 12/2010; 9(6):1032-46. · 7.55 Impact Factor
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    ABSTRACT: Age-related loss of muscle mass and function exhibits a marked acceleration from late middle age to senescence and exercise training is one method that has been proposed to slow this process. The purpose of this study was to determine if long-term treadmill exercise training initiated at late middle age could increase endogenous antioxidant enzyme activity and attenuate the loss of skeletal muscle contractile properties in the gastrocnemius/plantaris (GAS/Plan) and soleus (SOL) muscles of senescent rats (34-36 mo) through a decrease in oxidative damage. Male Fisher 344 × Brown Norway F1-hybrid rats underwent 5-7 mo of treadmill training beginning at late middle age (29 mo). A 7 mo sedentary adult group was used to investigate age-related changes. Aging caused an increase in antioxidant enzyme activities; however, only SOD activity was further increased with exercise training. Exercise training did not attenuate the decrease in twitch or tetanic tension of the GAS/Plan or SOL. It did, however, prevent the increase in twitch half relaxation time of the SOL muscle only. Oxidative damage, as reflected in carbonyl content, was increased with age and even further with exercise training in the GAS muscle. Muscle fibre cross sectional area was decreased with age and even further with exercise training. Interestingly, small muscle fibres showed the highest accumulation of carbonyls. Overall, despite an augmentation of select antioxidant enzyme activities, exercise training from late middle age through to senescence had minimal benefits for muscle contractile properties, perhaps in part due to exacerbated oxidation.
    Experimental gerontology 11/2010; 45(11):856-67. · 3.34 Impact Factor
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    ABSTRACT: Aging skeletal muscle shows an increased time to peak force and relaxation and a decreased specific force, all of which could relate to changes in muscle Ca(2+) handling. The purpose of this study was to determine if Ca(2+)-handling protein content and function are decreased in senescent gastrocnemius muscle and if initiating a training program in late middle age (LMA, 29 mo old) could improve function in senescent (34- to 36-mo-old) muscle. LMA male Fischer 344 x Brown-Norway rats underwent 5-7 mo of treadmill training. Aging resulted in a decrease in maximal sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) activity and a decrease in Ca(2+) release rate but no change in Ca(2+) uptake rate. Efficiency of the Ca(2+) pump was increased with age, as was the content of SERCA2a. Training caused a further increase in SERCA2a content. Aging also caused an increase in protein carbonyl and reactive nitrogen species damage accumulation, and both further increased with training. Consistent with the increase in oxidative damage, heat shock protein 70 content was increased with age and further increased with training. Together, these results suggest that while initiating exercise training in LMA augments the age-related increase in expression of heat shock protein 70 and the more efficient SERCA2a isoform, it did not prevent the decrease in SERCA activity and exacerbated oxidative damage in senescent gastrocnemius muscle.
    AJP Regulatory Integrative and Comparative Physiology 03/2010; 298(5):R1269-78. · 3.28 Impact Factor
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    ABSTRACT: We previously showed that 7 wk of treadmill exercise training in late-middle-aged rats can reverse the modest reductions in skeletal muscle aerobic function and enzyme activity relative to values in young adult rats (Exp Physiol 93: 863-871, 2008). The purpose of the present study was to determine whether extending this training program into senescence would attenuate the accelerated decline in the muscle aerobic machinery normally seen at this advanced age. For this purpose, 29-mo-old Fisher 344 Brown-Norway rats underwent 5 or 7 mo of treadmill exercise training. Training resulted in greater exercise capacity during an incremental treadmill exercise test and reduced percent body fat in 34- and 36-mo-old rats and improved survival. Despite these benefits at the whole body level, in situ muscle aerobic capacity and muscle mass were not greater in the trained groups at 34 mo or 36 mo of age. Similarly, the trained groups did not have higher activities of citrate synthase (CS) or Complex IV in homogenates of either the plantaris (fast twitch) or the soleus (slow twitch) muscles at either age. Finally, protein expression of CS (a marker of mitochondrial content) and peroxisome proliferator-activated receptor-gamma coactivator-1 (relating to the drive on mitochondrial biogenesis) were not higher in the trained groups. Therefore, although treadmill training from late middle age into senescence had significant benefits on running capacity, survival, and body fat, it did not prevent the declines in muscle mass, muscle aerobic capacity, or mitochondrial enzyme activities normally seen across this age, revealing a markedly diminished plasticity of the aerobic machinery in response to endurance exercise at advanced age.
    AJP Regulatory Integrative and Comparative Physiology 08/2009; 297(3):R744-55. · 3.28 Impact Factor