Implications of mitochondrial uncoupling in ... - Wiley Online Library

REVIEW ARTICLE

Implications of mitochondrial uncoupling in skeletal muscle in the development and treatment of obesity A. Brianne Thrush1, Robert Dent2, Ruth McPherson3 and Mary-Ellen Harper1 1 Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ontario, Canada 2 Ottawa Hospital Weight Management Clinic, Ottawa Hospital, Ontario, Canada 3 University of Ottawa Heart Institute, Ontario, Canada

Keywords diet resistance; mitochondria; obesity; oxidative phosphorylation; proton leak; uncoupling; weight loss Correspondence M.-E. Harper, BMI, Faculty of Medicine, University of Ottawa 451 Smyth Road, Ottawa, ON, K1H 8M5, Canada Fax: +1 613 562 5452 Tel: +1 613 562 5800 ex 8235 E-mail: mharper@uottawa.ca

Understanding the metabolic factors that contribute to obesity development and weight loss success are critical for combating obesity and obesity-related disorders. This review provides an overview of energy metabolism with a particular focus on mitochondrial function in health and in obesity. Mitochondrial proton leak contributes significantly to whole body energy expenditure and the potential role of energy uncoupling in weight loss success is discussed. We provide evidence to support the hypothesis that differences in energy efficiency are important regulators of body weight and weight loss success.

(Received 28 March 2013, revised 3 June 2013, accepted 17 June 2013) doi:10.1111/febs.12399

Introduction Obesity is a complex multifactorial disease and is a major risk factor for other chronic diseases, including type 2 diabetes mellitus (T2DM), cardiovascular disease, and cancer [1,2]. Obesity has reached epidemic proportions in many parts of the world; its prevalence is highest in the Americas, where it is estimated that 62% of adults are overweight or obese [3]. Understanding the myriad of biological and environmental factors that contribute to obesity is paramount for the development of personalized preventative and treatment strategies. The development of obesity is a complex issue involving the interaction of numerous environmental and biological factors. Caloric intake and energy expenditure, primarily through physical activity, are considered to be major environmental determinants in the develop-

ment of obesity. Although not discussed in detail in the present review, other environmental factors that have been associated with obesity development include (but are not limited to) socioeconomic status, education, stress and poor sleeping habits. Genetic predisposition and other biological factors such as thyroid status, which is known to alter energy efficiency, may predispose individuals to obesity and/or affect their capacity to lose weight [4–6]. An emerging area of research has also linked the gut microflora population with obesity development [7]. All of these factors, as well as others that are not listed, can contribute to energy intakes that exceed overall energy expenditure. This review discusses the biological factors that are considered to contribute to obesity susceptibility and

Abbreviations BAT, brown adipose tissue; BMR, basal metabolic rate; FA, fatty acid; IMF, intermyofibrilar; ODR, obese diet resistant; ODS, obese diet sensitive; OP1, optic atrophy 1; RMR, resting metabolic rate; ROS, reactive oxygen species; SNS, sympathetic nervous system; SS, subsarcolemmal; T2DM, type 2 diabetes mellitus; TDEE, total daily energy expenditure; UCP, uncoupling protein.

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affect the response to therapeutic strategies. In particular, the focus is on factors that affect the efficiency of energy metabolism. There is abundant evidence that the efficiency of ATP production during fuel oxidation is variable (i.e. the amount of ATP produced per unit oxygen consumed by cells and tissues varies). We examine the relevance of this physiologic fact with respect to obesity susceptibility and inter-individual variability in weight loss during hypocaloric treatment strategies with a focus on mitochondrial function in the skeletal muscle.

Partitioning of energy expenditure at the whole body level Obesity usually develops gradually, and occurs when energy intake is in excess of total body energy expenditure. Energy is expended by all cells and tissues of the body to support a wide variety of fuel-consuming processes. Before focusing on the variable efficiency with which energy is used, we briefly partition out the various components of total body energy expenditure. Total daily energy expenditure (TDEE) of adults living under fairly standard conditions can be separated broadly into three major components. The largest contributor to TDEE is basal metabolic rate (BMR), which can account for 60–75% of TDEE in a sedentary individual [8]. BMR is defined as the energy expended when an individual is in a resting and postabsorptive state in a thermoneutral environment (22– 27 °C in adult humans); measurements are made in individuals resting in a supine position in the morning following sleep [9–11]. Resting metabolic rate (RMR) is much more commonly measured than BMR because it is measured in individuals at complete rest, at any time of the day, in the post-absorptive state. When measured in individuals in a supine rested state, at normal room temperature (i.e. 22–23 °C), RMR is usually within 10% of BMR [12]. BMR is dependent on many physical and biological factors. It is increased with body weight, male sex, stress, thyroid hormones, pregnancy and lactation [13–16] and is decreased with weight loss, calorie restriction, starvation and ageing [16–20]. The major determinant of BMR is fat free mass [21]. Cellular reactions that contribute to BMR include ATP-demanding processes, such as the maintenance of ion gradients across cell membranes; RNA and protein synthesis; as well as other energy demanding reactions involved in cellular turnover and repair [22]. Another important processes is mitochondrial proton leak, which, unlike the aforementioned reactions, does not consume ATP; instead, it decreases the efficiency of 5016

ATP synthesis [23]. There is evidence that mitochondrial proton leak is altered in different metabolic conditions; this is discussed below, as are the implications of proton leak to the development and treatment of obesity. Another important contributor to TDEE is the thermic effect of food, which is the increase in metabolic rate following a meal. The increase in metabolic rate is a result of the energy used for the digestion, absorption and metabolism of dietary nutrients, as well as sympathetic nervous system activation to increase blood flow to digestive organs and to peripheral tissue to promote nutrient clearance [24–26]. Dietary protein causes the largest thermogenic response, followed by carbohydrate and then fat, which has a very low thermogenic effect. The thermogenic response to eating accounts for approximately 10% of TDEE [27]. There is evidence that the thermogenic response to food is impaired in obesity [28,29]. The energy expended as a result of physical activity is also a major component of TDEE and is the most variable amongst individuals [12]. This includes both volitional exercise and energy expended as a result of non-exercising thermogenesis such as that expended during standing, talking, walking, fidgeting, daily living, spontaneous muscular contractions or shivering [8,30]. There is evidence that spontaneous movement, such as fidgeting, contributes a small but significant amount to differences in TDEE [12], and this may mitigate obesity development [31]. However, a much more significant contributor to TDEE is volitional exercise. The contributions of various tissues to overall metabolic rate have been estimated in rodents and humans [22]. The major contributors are skeletal muscle (20%), brain (20%), liver (17%), heart (11%) and gastrointestinal tract (10%). Although the metabolic rate per gram of tissue is quite low for resting skeletal muscle, it is one of the major contributors to basal metabolic rate, by virtue of its size, which is approximately 40% of adult human body mass. The brain, liver and gastrointestinal tract all account for < 2% of total body mass but are estimated to account for approximately 40% of basal metabolic rate[22,32,33].

Oxidative phosphorylation The oxidation of dietary energy substrates, mainly carbohydrate and fatty acids, results in the production of reducing equivalents in the form of NADH + H+ and FADH2. The latter are transferred through a series of shuttles and oxidation-reduction reactions in the mitochondrial electron transport chain to the final electron acceptor: molecular oxygen. The movement of FEBS Journal 280 (2013) 5015–5029 ª 2013 FEBS

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electrons through the electron transport chain is coupled to proton pumping from the mitochondrial matrix to the intermembrane space, thus generating an electrochemical gradient across the inner mitochondrial membrane, referred to as the protonmotive force. The return of protons to the matrix through the ATPsynthase complex drives ATP production. The importance of these reactions is obvious when considering the fact that most adults produce approximately 65 kg of ATP per day [34].

Mitochondrial proton leak At such a high volume of ATP synthesis at the whole body level, it is easy to conceive of the potential impact of even moderate uncoupling of mitochondrial proton return from ATP synthesis; uncoupling of oxidative phosphorylation essentially means that dietary energy is not captured as ATP. Mitochondrial energy metabolism is considered as coupled or efficient when all or most of the oxygen consumed by mitochondria is associated with ATP production. Coupling efficiency approaches 90% when ADP is present, as a result of rapid ATP synthesis [35,36]. Coupling efficiency varies across different cell types and has been estimated to be 75–80% in rats [37]. Uncoupled respiration is the situation in which oxygen is consumed in the absence of ATP production. The respiration that occurs when ATP synthesis is not occurring is referred to as ‘state 4 respiration’, a metabolic state in which protons leak back across the mitochondrial inner membrane to the matrix, bypassing ATP synthase. In such uncoupled states, energy substrates [e.g. fatty acids (FA)] are oxidized, and oxygen is consumed but no ATP is synthesized. In perfused rat skeletal muscle, proton leak has been estimated to account for approximately 52% of resting oxygen consumption [33]. Considering that skeletal muscle accounts for upwards of 30% of RMR in rats [38], skeletal muscle proton leak likely contributes considerably to TDEE. Indeed, it is estimated that proton leak accounts to 20–30% of BMR in rats [22,33,37]. There are considered to be two classes of mitochondrial proton leak: basal and inducible. Basal proton leak occurs in mitochondria of all tissues and is a process that does not appear to be acutely controlled by specific factors. It is associated with adenine nucleotide translocase [39], and the novel uncoupling proteins (UCP1–3) [36]. Although the physiological purpose of basal uncoupling is unknown, it has been hypothesized that a basal level of electron movement through the electron transport chain facilitates movement to higher energy state in times of energy demand by increasing FEBS Journal 280 (2013) 5015–5029 ª 2013 FEBS


sensitivity and decreasing the response time to changes in cellular ATP utilization [22]. Basal proton leak may also function to mitigate oxidative stress [22]. Inducible proton leak is catalyzed by the uncoupling proteins UCP1, UCP2, and UCP3 and can be acutely controlled [36]; adenine nucleotide translocase may also be acutely induced to perform proton leak reactions [40,41]. UCP1 is expressed exclusively in brown adipose tissue and is considered to be the classic uncoupling protein. UCP1 is activated in response to sympathetic nervous system (SNS) activation and is necessary for brown adipose tissue (BAT) thermogenesis. In response to SNS stimulation of brown adipocytes, fatty acids are considered to relieve purine nucleotide inhibition of UCP1, thereby activating UCP1 proton leak [42]. UCP1 is an abundant mitochondrial protein (approximately 10%) and BAT is dense with mitochondria [42]. In rodent models, UCP1 activation can increase RMR up to four-fold [43] and protects against diet-induced obesity [44]. In humans, BAT is present in infants [45] and functions to maintain body temperature [46]. Recent evidence has unequivocally demonstrated that UCP1-expressing BAT is present in adults and is activated with cold exposure [47]. Indeed, BAT, identified based on histological staining of mitochondrial enzymes and multilocular tissue, had previously been detected in outdoor workers in cold climates [48]. UCP2 is expressed widely in tissues, including the stomach, thymus, spleen, macrophages, hypothalamus and pancreatic b cells. UCP3 is primarily expressed in skeletal muscle but is also found in BAT and sometimes in heart [49]. The role and mechanism of activation of UCP2 and UCP3 have been extensively debated ever since their discovery in the late 1990s. Evidence from the Brand laboratory first demonstrated that UCP1–3 are activated in the presence of reactive oxygen species (ROS). This elicited the ‘uncoupling to survive hypothesis’, which holds that proton leak acts to decrease excessive mitochondrial superoxide production to prevent oxidative damage [50]. Whether or not this is indeed the physiological functions of UCP2 and UCP3 are still under investigation. It has been noted that macrophages of UCP2 / mice have increased ROS production and mice are protected from parasitic infection compared to wild-type mice [51]. Moreover, muscle mitochondria of UCP3 / mice produce more ROS and have more evidence of oxidative damage compared to wild-type [52,53] and UCP3 overexpression lowers age-associated increases in mitochondrial ROS emission [54]. UCP3 transgenic mice are less metabolically efficient [55,56] and also are protected against diet-induced obesity [55–57] and high-fat diet5017


induced insulin resistance [58]. Accordingly, UCP1 and UCP3 protein expression is higher in the BAT and skeletal muscle mitochondria of obesity-resistant compared to obesity-prone mice [59]. UCP3 / mice are not obese, although they demonstrate higher levels of oxidative stress compared to their wild-type littermates [52,53]. After 8 and 26 weeks of high-fat feeding, UCP3 / mice demonstrate similar body weights and mitochondrial respiration but higher ROS production compared to wild-type mice [60]. Several polymorphisms in the UCP1–3 genes identified in humans have been associated with obesity phenotypes, increased weight gain, weight loss resistance and T2DM [61–64] and this has been reviewed extensively [65,66]. The most commonly studied UCP1 polymorphisms include the A1766G and A-112C at the 5′ flanking region, A3826G SNP in the promoter region and the A64T polymorphism in exon 2 [65]. Although controversial, the A3826G polymorphism is the most widely studied and has been associated with obesity, weight gain and reduced weight loss success [62–65]. In the UCP2 gene, polymorphisms at the 866 position in the promoter region have been associated with obesity and T2DM risk [66]. Carriers of the –866A allele demonstrate modest reductions in obesity rates [67] but a decreased insulin response to glucose and thus increased T2DM risk [68,69]. Carriers of the –866G allele demonstrate a higher body mass index and fat mass and an increased risk for obesity [66,67]. The UCP3 gene polymorphism –55T allele has been associated with higher rates of UCP3 mRNA expression [70] and resting energy expenditure [71], whereas heterozygosity (C/T) is associated with decreased obesity [72] and T2DM risk [73].

Physiological factors affecting mitochondrial proton leak The most widely recognized endocrine factors affecting mitochondrial proton leak are the thyroid hormones. Thyroid hormones are also the major endocrine regulators of BMR. In hypothyroidism or following thyroidectomy, BMR is decreased, whereas it is increased in hyperthyroidism [74,75]. Most of the thyroid hormone effects mitochondrial respiration are mediated by genomic mechanisms. Thyroid hormone receptors have been located in the nucleus and mitochondria in various tissues [76–78]. Major effects of thyroid hormones on proton leak and energy metabolism appear to be related to mitochondrial biogenesis. Specifically, thyroid hormones increase the expression of proteins regulating mitochondrial biogenesis, namely peroxisomeproliferator-activated receptor c coactivator a and 5018

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nuclear respiratory factor-1 [79–81]. Thyroid hormones also increase oxidative phosphorylation gene expression [81] and modulate mitochondrial respiration [82]. Mitochondrial respiration is increased and decreased in mitochondria and hepatocytes isolated from hyper- and hypothyroid rats, respectively, and approximately 50% of these differences in respiration occur as a result of changes in proton leak [83–87]. Finally, in situations of elevated thyroid hormones, resting energy expenditure, substrate oxidation and skeletal muscle mitochondrial proton leak are all increased [88]. Short- and long-term calorie restrictions have consistently been shown to decrease proton leak in rodent liver and skeletal muscle [89–92]. However, there is less consensus on the effects of high-fat feeding on mitochondrial coupling in response to a high-fat diet; the effects may depend on the tissue studied or the model used. High-fat feeding has been shown to decrease [93] or not change [94] mitochondrial coupling in isolated mitochondria from liver of high-fat fed rats. Uncoupling protein expression and mitochondrial proton leak were assessed in isolated mitochondria of BAT and hindlimb muscle in obesity-prone and obesityresistant mice [59]. In that study, UCP1 and UCP3 expression and proton leak were higher in BAT of obesity-resistant compared to obesity-prone mice on a normal chow diet. Interestingly, when challenged with a high fat diet, BAT proton leak was increased in obesity-prone mice to levels comparable with obesityresistant mice. In hindlimb muscle, UCP1 but not UCP3 protein expression is higher in obesity-resistant mice. This was previously reported to be the result of BAT tissue in the hindlimb of this strain of mouse (129 S6/SvEvTac strain) [95]. Despite this, there was no difference in skeletal muscle proton leak between the two groups when on a control or high-fat diet [59]. Mitochondrial proton leak was also reduced in the skeletal muscle of individuals with T2DM following the initiation of intensive insulin therapy, which is consistent with improved mitochondrial efficiency [96]. In situations of acute substrate oversupply, mitochondrial uncoupling was increased in saponin permeabilized fibres from hearts perfused with glucose and fatty acids [97]. Exercise training is known to increase mitochondrial oxidative capacity and mitochondrial content; however, the effect of exercise training on muscle mitochondrial coupling efficiency is not completely understood, with studies showing it to be both increased [98,99] and decreased [100] in active individuals. Recent research demonstrates that mitochondrial content and ATP flux are reduced in sedentary FEBS Journal 280 (2013) 5015–5029 ª 2013 FEBS

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compared to active individuals, despite similar rates of O2 flux, which is consistent with the conclusion that there is an increased uncoupled respiration in sedentary individuals [99]. In summary, mitochondrial proton leak contributes significantly to energy expenditure and is altered under a number of physiological conditions. It is thus feasible that alterations in mitochondrial proton leak could contribute to obesity development or weight loss success.

Energy balance in response to weight gain or loss Resting metabolic rate is higher in obesity, primarily as a result of an increase in body mass. When weight loss occurs, resting energy expenditure is decreased [19,101–103] and this persists beyond the weight reduction phase [19]. Equations have been developed to estimate energy expenditure based on that measured in healthy individuals. These equations typically use sex, age, height and weight to estimate energy expenditure [104–106]. Although controversial, resting energy expenditure [18,102,103,107,108] and exercise energy expenditure [104] in response to weight loss have been shown to be less than predicted. The reduction in resting energy expenditure following weight loss has often been attributed to a loss in lean mass. However, a recent study showed that RMR and TDEE were still significantly reduced to values that were lower than predicted in individuals who had predominantly maintained lean mass after drastic weight reduction following a demanding combination of diet and exercise [103]. It has been suggested that reductions in RMR and TDEE in response to weight loss may be a result of adaptive mechanisms to protect body mass [109]. Such adaptive mechanisms are complex and poorly understood but may include hormonal, SNS signalling and behavioural changes. Increased appetite and food intake [110] have been observed in response to decreased energy intake. Increased food cravings [111], calories to reach satiation and meal size [112, 113] have been reported following weight loss. Following weight reduction, increased neural activity in reward centres of the brain and in areas responsible for processing of food-related stimuli and decreased activity in areas of the brain responsible for food restrain have been observed in humans [114]. Interestingly, postweight loss leptin administration largely reverses the weight loss-induced changes in neural activity [114,115], and increases post-meal satiation [112]. Nonexercise activity thermogenesis is also decreased and FEBS Journal 280 (2013) 5015–5029 ª 2013 FEBS


increased in response to energy restriction and overfeeding, respectively [31,107], and may also be a mechanism to protect body mass. Circulating hormones are also altered in situations of energy imbalance and may play an important role in weight regulation. For example, circulating thyroid hormones and 24-h urine catecholamine output are increased and decreased following weight gain and loss, respectively, and are associated with changes in resting energy expenditure [116]. Decreases in circulating leptin have been associated with weight lossinduced decreases in energy expenditure and whole body fat oxidation [117], and may contribute to weight regain post-weight loss [118]. Restoring circulating leptin levels to pre-weight loss values with leptin administration prevents weight loss-induced reductions in energy expenditure and circulating thyroid hormones, and also restores sympathetic tone [112,114,118]. Hormonal and metabolic changes that occur in obesity and weight loss can also affect mitochondrial function and efficiency, which would contribute to changes in energy expenditure. Indeed, diet-induced weight loss has been shown to reduce mitochondrial oxidative capacity as measured in permeabilized muscle fibres, despite no changes in mitochondrial content [119]. In summary, it appears that there are numerous physiological, metabolic and behavioural adaptations that occur, in an attempt to protect body weight, during energy imbalance that would ultimately affect weight loss success.

The biological basis of weight loss variability There is considerable evidence demonstrating variability in response to positive or negative energy balance. For example, the biological basis for weight gain variability has been demonstrated in highly controlled, overfeeding studies in monozygotic twins. This research demonstrated a greater between-pair variability than within-pairs with respect to weight gain and changes in RMR and fat mass gain [4]. Research conducted by our group has investigated the metabolic and genetic basis for weight loss variability in humans. The weight loss programme at the Ottawa Hospital Weight Management Clinic is a highly regimented meal replacement weight loss programme. Individuals enrolled in the programme consume 900 kcalday 1 in the form of a meal replacement for 6 or 12 weeks. Meal replacement is accompanied by weekly visits with physicians and information sessions. Food is slowly re-introduced at 12 weeks. In highly-adherent subjects, we have 5019

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observed a 10-fold difference in weight loss during the first 6 weeks of weight loss. When factors known to effect metabolic rate are taken into account (age, weight, sex, thyroid hormone status), a two-fold difference in weight loss remains unexplained [120]. Metabolic and genomic studies have been conducted in patients in the upper quintile for rate of weight loss (obese diet sensitive; ODS) and lower quintile for rate of weight loss (obese diet resistant; ODR) (Fig. 1). To address the impact of possible differences in physical activity between groups, planned physical activity and work-related activity were compared and found not to be different between the ODS and ODR groups [121]. This suggests that there may be biological factors contributing to the observed differences in rate of weight loss. Indeed, in isolated mitochondria of rectus femoris, proton leak was 51% higher in ODS compared to ODR. Considering that muscle energy expenditure accounts for approximately 30% of RMR in humans [22], the observed differences in muscle mitochondrial proton leak could reflect a 5% difference in RMR between ODS and ODR individuals [6]. Further investigations revealed distinct differences in the skeletal muscle of ODS and ODR individuals. Skeletal muscle (vastus lateralis) from ODS has more type I oxidative muscle fibers and exhibited fiber hypertrophy compared to ODR individuals, whereas type IIa (oxidative-glycolytic) fibres were higher in ODR than ODS individuals and IIx fibers were higher in ODR compared to lean individuals [121]. Type IIx fiber expression has also been shown to correlate with insulin resistance in obesity and T2DM [122]. Overfeeding

studies in monozygotic twins have demonstrated that those individuals with higher proportions of type I and lower type IIa fibers gain less fat mass in response to overfeeding [123], which follows a pattern that is similar to that observed in the impaired weight loss response in our studies [121]. Skeletal muscle (rectus femoris) of ODS subjects also demonstrated higher mRNA expression of UCP3 and the genes involved in oxidative metabolism compared to muscle of ODR subjects [121]. Interestingly, this difference in gene expression pattern (e.g. ox phos gene transcripts) has also been observed in blood samples of ODS and ODR individuals before the weight loss programme; hence, blood-based predictor tests may be available in the future [124]. Taken together, this research suggests that the skeletal muscle of ODS individuals has a greater oxidative capacity, although oxidative phosphorylation is less efficient and this likely contributes to the observed differences in weight loss success (Fig. 2).

ODR

ODS

Fig. 1. Rate of weight loss in ODS and ODR women. Weight loss data are shown for the time of entry into the weight management programme (week 0) and over the subsequent 32 weeks. The dashed line indicates the point at which rate of weight loss is calculated (6 weeks) (n = 12 per group). Reproduced with permission [6].

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Fig. 2. Schematic representation of proposed mechanism of weight loss variability. ODS individuals demonstrate a higher expression of oxidative phosphorylation genes, type I oxidative muscle fiber and increased levels of proton leak compared to ODR individuals. Collectively, this results in higher rates of weight loss success [6,121]. ANT, adenine nucleotide translocase.


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Mitochondrial dysfunction in obesity and T2DM Numerous studies have demonstrated that skeletal muscle metabolism and mitochondrial function are impaired in obesity. Specifically, skeletal muscle from obese humans is characterized by increased FA uptake, lipid accumulation and oxidative stress [125– 128]. The accumulation of reactive lipid species, namely diacyglycerol and ceramide, are associated with impaired insulin sensitivity in this tissue [129–132]. Many studies have also demonstrated that FA oxidation is reduced in obesity and T2DM [133–136], which may be a contributing factor in lipid accumulation and oxidative stress. Other studies, however, have demonstrated that FA oxidation is either moderately increased or not different compared to lean controls in rodents and humans [137–139]. The observed differences in mitochondrial oxidation may be a result of differences in mitochondrial content [137,140]. Mitochondrial content has been shown to be reduced in the skeletal muscle of obese individuals and individuals with T2DM compared to lean controls [141]. This suggests that mitochondrial dysfunction may in part be the result of a reduced mitochondrial mass. In accordance with this, the levels of mitochondrial proteins and their (predominantly nuclear) genes are reduced in the skeletal muscle in obesity and T2DM [142,143] and also in individuals with an impaired weight loss response [121,124]. Some studies have demonstrated that FA oxidation is incomplete in the skeletal muscle of obese rodents and humans and following a high-fat diet [144–146]. Incomplete FA oxidation has been observed in obesity [144] and can lead to the accumulation of by products of metabolism, namely acylcarnitines. Acylcarnitines have been proposed to cause mitochondrial dysfunction and insulin resistance [145,146]; however, the metabolic effects and mechanisms of action of these metabolites remain unknown. In skeletal muscle, mitochondria form a reticulum and are categorized into two distinct compartments: subsarcolemmal (SS) and intermyofibrilar (IMF). SS mitochondria are predominantly responsible for energy demands at the cell surface, such as substrate transport, ion exchange and cellular signalling, whereas IMF are important for muscle contraction [147,148]. Studies have demonstrated that SS and IMF mitochondria respond differently to various metabolic perturbations and there may be a different pattern of response in rodents compared to humans [141,149–151]. In obese Zucker rats, FA oxidation and mitochondrial content (number, width, density) are increased in SS but not IMF mitochondria compared FEBS Journal 280 (2013) 5015–5029 ª 2013 FEBS


to lean controls [151]. By contrast to rodent models, IMF but not SS mitochondrial content is decreased in obese insulin-resistant and obese T2DM individuals compared to lean insulin-sensitive individuals. IMF mitochondrial content was also positively associated with insulin sensitivity [150]. The electron transport chain activity is also reduced in SS and IMF mitochondria of obese individuals with T2DM compared to lean controls [141]. It is worth noting that physical activity levels were reportedly higher in the lean population, which may impact upon the observed findings [141]. In summary, the role of subcellular locations of mitochondria in mitochondrial dysfunction in humans under different metabolic states (obesity, diabetes, insulin resistance, weight loss) is not completely understood and warrants further investigation. As discussed above, fiber type may play an important role in skeletal muscle function and weight loss success in obesity. Type IIx glycolytic fibre expression is higher in the skeletal muscle of diabetic individuals [122] and type IIa fibre expression is positively correlated with the weight gain response to overfeeding [123]. Type II fibres have a reduced capacity to oxidize fat [152] and may also have a a reduced capacity to deal with oxidative stress [153], thus potentially contributing to the increases oxidative stress observed in obesity and T2DM. Changes in mitochondrial morphology have also been observed in obesity and T2DM. Higher rates of mitochondrial fission have been implicated with diabetic neuropathy [154,155]. Shorter, rounder mitochondria and increases in mitochondrial fission proteins dynamin-related protein 1 and fission protein 1 have been observed in the skeletal muscle of ob/ob and high-fat fed mice, and palmitate-treated C2C12 cells [156]. The fusion protein, mitofusin-1 is reduced in the skeletal muscle of obese rodents and humans, and this is associated with smaller mitochondria and a fragmented mitochondrial network [157]. Lower levels of the fusion proteins mitofusin 1 and optic atrophy 1 have also been observed in the skeletal muscle of individuals with T2DM [158].

Exercise, weight loss maintenance and mitochondrial function Exercise training has been shown to improve mitochondrial function and content in obesity and T2DM [159–163]. Exercise training stimulates mitochondrial biogenesis resulting in increased mitochondrial size and content [159] and increases the expression of proteins involved in oxidative phosphorylation and FA utilization [164–166]. In addition, exercise training 5021

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increases FA oxidation, promotes more complete FA oxidation and increases oxidative phosphorylation [100,145,167]. Diet-induced weight loss decreases skeletal muscle oxidative metabolism [119] and exercise may prevent this [160,167,168]. In addition, weight loss from diet and exercise demonstrates increased mitochondrial density and capacity (NADH-oxidase activity) compared to diet alone [169]. In addition to the beneficial effects of exercise-induced weight loss on mitochondrial function and insulin sensitivity, there is evidence to suggest that regular exercise is important for weight loss maintenance [170,171]. In summary, the mechanism of the development of muscle mitochondrial dysfunction in obesity and T2DM is as yet poorly understood. Mitochondrial dysfunction combined with decreases in mitochondrial content apparently increases the risk for the development of obesity and obesity-related disease (e.g. muscle insulin resistance). However, the latter is the subject of continuing debate [172,173].

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Concluding remarks Energy intake and expenditure, and thus body weight, are regulated by a complex interplay between of physiological and behavioural factors, which are only beginning to be understood. Research suggests that mitochondrial function and/or content is altered in obesity, and likely contributes to the development of the disease. It appears that mitochondrial efficiency is an important factor in human energy expenditure and body weight regulation, with higher rates of muscle mitochondrial proton leak in those individuals who lose weight much more quickly than others. However, more research is needed to determine the contribution of mitochondrial efficiency to body weight regulation and energy expenditure. Understanding mitochondrial dysfunction associated with obesity and weight loss success is important for developing appropriate prevention and treatment strategies.

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