Protein Metabolism in Human Obesity: A Shift in Focus From Whole‐Body to Skeletal Muscle

Insulin resistance in obesity alters carbohydrate and lipid metabolism in skeletal muscle (1). However, the effects of obesity-associated insulin resistance on skeletal muscle protein metabolism remain largely unknown. Generally, weight gain is associated with an increase in fat as well as lean mass. Also, from a cross-sectional point of view, obese individuals are generally able to produce greater absolute strength compared to lean individuals (2,3). Such observations have attenuated concerns with respect to the clinical significance of altered protein kinetics in the skeletal muscle of obese individuals, and together with inconsistent results from studies investigating protein kinetics at the whole-body level have reduced interest in the investigation of protein kinetics at the muscle level in obesity. Locomotor muscles are exposed to greater, weight bearing, mechanical loads in obese compared to lean individuals, which can have a physical training-like effect in the muscles of obese individuals and mask impaired muscle protein metabolism and function with obesity. Evidence based on nonlocomotor muscles (i.e., respiratory muscles) indicates impaired function in muscles of obese individuals (4). Further, evidence suggests that improvements in muscle strength and mass with resistance exercise are lower in obese compared to lean subjects (5,6). An overall negative correlation between resistance to the physiological effects of insulin, a common observation in human obesity, and muscle function has been reported (7,8). Within the skeletal muscle, obese insulin-resistant (IR) individuals have lower mitochondrial protein content as indicated by an up to ∼30% reduction in mitochondrial marker enzymes (9,10), and in parallel with an impaired rate of synthesis of mitochondrial proteins (11). We have recently determined that the content of individual skeletal muscle proteins involved in cytoskeletal structure, such as desmin and α-actinin-2, is ∼50% lower in muscle from obese IR subjects (12). Overall, this evidence suggests that obesity is associated with alterations in muscle protein metabolism and undesirable effects on skeletal muscle substrate metabolism and function. This review critically evaluates available evidence describing the effects of obesity on whole-body protein turnover. The main purpose of the review is to highlight recent evidence supporting an impaired rate of synthesis of skeletal muscle proteins in obese, IR individuals. Molecular and physiological mechanisms controlling protein synthesis in skeletal muscle with relevance to human obesity are also examined. Finally, clinical implications of an impaired muscle protein turnover in obesity are discussed. Higher plasma concentrations of individual amino acids in obesity have been shown in most (13,14,15,16,17,18), but not all (19,20,21,22) studies comparing obese subjects to lean controls. Further, an insulin-induced decline in the concentration of plasma amino acids, particularly branched-chain amino acid concentrations, was less evident in obese subjects (14,22). The literature is replete with studies evaluating whole-body protein metabolism in obesity with the help of amino acid tracers. Basal/postabsorptive whole-body protein turnover in obese subjects has been reported to be similar (19,23) or increased (18,24,25,26), compared to that in lean subjects. Under insulin-stimulated conditions, whole-body protein breakdown in obese subjects was either the same (14,15,19,23,27) or increased (19,28) relative to lean controls, while whole-body protein synthesis has been reported to be either increased (19), the same (19,23), or decreased (11,15,18). These discrepancies with respect to the direction of the changes in whole-body protein kinetics as a result of obesity have not been addressed. An intrinsic difficulty when evaluating whole-body protein metabolism is the means of normalizing protein kinetic data. Data can be normalized to whole-body surface area (19) or lean body mass (24), or can be expressed as a change from basal when evaluating the effects of plasma insulin (15). Basal whole-body protein kinetics are generally greater in obese compared to lean controls when expressed either as absolute whole-body protein kinetics (µmol/min) or adjusted for lean body mass (µmol/kg lean body mass/min) (15,24,26). However, these kinetics will be lower when expressed per unit of overall body weight (15,20). This can be explained by the fact that, although body fat contributes significantly to whole-body amino acid kinetics, its kinetics are less per given unit of tissue volume than those of muscle (20). At the same time, body fat contribution to whole-body protein kinetics is expected to vary greatly depending on the amount of body fat of the individual complicating the physiological interpretation of whole-body data adjusted to lean body mass. Furthermore, although the response of whole-body protein synthesis to insulin is not different between groups when expressed per unit of lean body mass, it is more than 50% lower in the obese compared to lean subjects when expressed as change from basal (15). This latter approach, which constitutes a more appropriate way to evaluate the sensitivity of whole-body protein metabolism to insulin, suggests reduced stimulation of whole-body protein turnover by insulin in obesity. Obesity has been found to be associated with lower physical activity (29), and this could also explain apparent discrepancies in the literature. In normal weight subjects, chronic aerobic exercise training does not have any effect on whole-body protein synthesis (30), but acute aerobic exercise increases skeletal muscle protein synthesis (31). When short-term physical activity is controlled by having subjects either avoid exercise during the days leading-up to the study (25) or by admitting them to the research unit prior to the study day (24,32), a 24% increase in the absolute rate of basal whole-body protein synthesis in obesity has been evident. These higher rates of protein turnover in the basal state can also be reduced. Acute exercise is associated with a decrease in whole-body protein turnover in obese subjects (33), but considerable absolute decreases (∼15%) in whole-body protein turnover in obese subjects occur only after weight loss (33). Therefore, increased whole-body protein kinetics in obesity appear to relate less to effects of physical activity and more to the effects of body composition, namely increased percent body fat. Positive correlations between whole-body protein turnover and several indexes of body fatness have been reported (32,34), even after accounting for the greater degree of lean body mass in obesity (34). Women have higher rates of basal whole-body protein turnover for a given amount of fat-free body mass when compared to males (35). Further, whole-body protein turnover is increased in women with higher waist-to-hip circumference (24). Upper body obesity is associated with an impaired ability to suppress whole-body protein breakdown during hyperinsulinemia (28). Although the exact mechanism(s) are not known, increased plasma free fatty acid concentrations may be implicated. At the muscle level, we have recently reported that a prolonged increase in plasma free fatty acid concentrations in healthy humans decreases protein breakdown (36) by 27%, which could have explained the past observation of a lower rate of muscle protein breakdown in obesity (20). These data also suggest a disassociation in protein kinetics between muscle and whole-body, with whole-body protein kinetics being more closely affected (i.e., increased) by whole-body fat accumulation (34). Further, women with increased body fat accumulation in the abdominal area have decreased muscle protein breakdown compared to lean women (20). This suggests that body fat distribution, rather than gender per se, may be more important in altering protein metabolism in obesity. Whole-body protein kinetics in obesity often have been studied in parallel with whole-body glucose kinetics (19,23). Besides intrinsic differences in the metabolism of these two substrates across human tissues, protein metabolism is more sensitive than glucose metabolism to the plasma insulin concentration both at the whole-body (37), as well as the skeletal muscle (38) levels. This is in line with the impaired suppression of whole-body protein breakdown in the obese compared to lean subjects being evident at relatively low (i.e., 10 mU/m2/min) (19,28) but not higher (i.e., 40 mU/m2/min) plasma insulin concentrations (19,27). Whether stimulation of protein synthesis by insulin can be assessed appropriately in the presence of reduced plasma amino acid concentrations has been argued (39,40), because plasma amino acids represent important stimuli for both whole-body (41,42), and skeletal muscle (43,44) protein synthesis. Changes in both proteins synthesis and degradation will affect plasma amino acid concentrations. Insulin infusion acutely reduces the concentration of plasma amino acids (14,19,23), with greater effect in lean compared to obese subjects (23). Unequal background plasma amino acid concentrations between groups of lean and obese subjects can become a confounding factor when assessing the role of plasma insulin in protein synthesis. Further, the role of plasma insulin in regulating protein synthesis cannot be assessed properly in the presence of large experimental increases in the concentration of plasma amino acids (19). Increases in plasma amino acid concentrations can mediate sufficient delivery of substrate to tissues for stimulation of protein synthesis in both lean and obese subjects, therefore masking any impaired response of protein synthesis in obese subjects that may be linked directly to plasma insulin. During investigation of the role of insulin on protein metabolism, clamping the plasma amino acids at similar physiological levels between lean and obese subjects can control for any bias due to the specific effects of plasma amino acids themselves on both whole-body as well as tissue-specific protein synthesis. When the plasma amino acid concentration is clamped at postabsorptive levels (15) or concentrations simulating the fed state (11), an impaired response of whole-body protein synthesis to insulin is documented in obesity. Evidence supporting a negative effect of body fat on muscle protein metabolism and function can be found in the literature of aging. Obese older subjects have poorer muscle quality than that of nonobese older subjects (45), suggesting that poor muscle quality in obese older subjects may be linked to body fat accumulation. In humans, body fat accumulates with age despite maintaining a stable body weight across ages (46). At the same time, a decline in muscle mass is observed around the fourth decade in life (47,48). When eventually both effects do occur, the increased body fat observed simultaneously with decreased muscle mass (known as sarcopenia) in elderly has been termed “sarcopenic obesity”. This condition is characterized by the presence of a vicious cycle where obesity and sarcopenia reinforce each other (49): adverse metabolic changes due to obesity can have negative impact on muscle protein turnover leading to loss of muscle mass, whereas at the same time sarcopenia-related decreases in resting metabolic rate and physical activity contribute to positive energy balance leading to obesity. In older individuals we have observed that the rate of postabsorptive muscle protein synthesis is lower in subjects with increased percent body fat (Figure 1). Similar relationship has been observed in obese young individuals (11), a circumstance devoid of the intrinsic physiological effects of aging itself on muscle protein synthesis. Given that an increase in body fat accumulation in the general population is evident as early as in the third decade of life (50) preceding the decline in muscle mass observed in middle-aged adults (47,48), increased body fat leading to insulin resistance early in life may be a precursor to not only diabetes but also the age-associated loss of muscle protein. Correlation (Pearson's product-moment, r) between percent body fat and postabsorptive skeletal muscle protein fractional synthesis rate in older (60–79 years old) subjects (unpublished data—see acknowledgements note). While there is speculation that fat mass gains observed in middle-aged adults may predispose these individuals to muscle loss as they age (51), evidence describing protein turnover in human obesity, per se, at the skeletal muscle level is limited. Considerable evidence over the past 30 years from animal studies, however, consistently shows that obese rats have less protein content in skeletal muscle compared to their lean controls (52,53,54,55). This evidence has been collected in parallel with evidence showing lower rates of amino acid uptake (53) and protein synthesis (52,54,56,57,58,59,60) in the muscles from obese compared to lean animals. Recently, an impaired response of muscle protein synthesis to nutrient ingestion has been reported in obese animals (61). In a study using the arterio-venous balance approach to describe protein kinetics in obesity, muscle protein breakdown in the postabsorptive state was ∼60% lower in the obese compared to lean subjects (20). When the rate of synthesis of mixed-muscle proteins was measured in the postabsorptive state in obese IR subjects it was found to be ∼46% lower in the obese compared to that of lean controls (11). On the other hand, in the same study, combined hyperaminoacidemia-hyperinsulinemia resulted in a similar stimulation of muscle protein synthesis in both lean and obese IR subjects (11). This latter finding is different from that reported with combined hyperaminoacidemia-hyperinsulinemia in the elderly, who demonstrated lower muscle protein synthesis compared to that in young subjects (62). Therefore, extrapolating any findings from studies in elderly to obese, IR humans can lead to discrepant interpretations about the role of obesity in altering muscle protein kinetics. Although aging and obesity may both be referred to as states of “sarcopenic obesity” (49), the possibility exists that these conditions impair muscle protein synthesis through different physiological mechanisms with overlapping features. Although combined hyperaminoacidemia-hyperinsulinemia increases the average rate of synthesis of total mixed-muscle proteins in the obese to the same extent as in lean subjects, this same stimulus did not increase the rate of synthesis of mitochondrial proteins in the muscle of the obese IR subjects (11). Differences in the physical activity levels between obese and lean subjects may have played a role in the impaired response of protein synthesis in muscle mitochondria in the obese subjects. However, the two groups were well-matched for maximal oxygen uptake (11), a function that is largely supported by the mitochondria. Impaired response in the synthesis of muscle mitochondrial proteins, but not total muscle proteins under the same circumstances, suggests that physiological mechanisms regulating the turnover rate of functionally related groups of proteins are uniquely impaired in the skeletal muscle of obese IR individuals. Increasing plasma amino acid concentrations stimulates overall protein synthesis in muscle in parallel with activation of the rapamycin-sensitive mammalian target of rapamycin (mTOR) pathway (63). On the other hand, decreasing plasma amino acid concentrations decreases the rate of overall protein synthesis in muscle, and in parallel with decreased activity of molecular markers implicated in protein synthesis (64). Activation of the mTOR pathway in skeletal muscle, leading to the subsequent activation of p70 ribosomal protein S6 kinase 1, is currently the best described molecular mechanism linking increased plasma amino acid and insulin concentrations to muscle protein synthesis (63,65) (Figure 2). Although care needs to be exercised when extrapolating data from rodents to humans (66), in the absence of any relevant evidence in humans, impaired responses with respect to the activation of the mTOR pathway by insulin (67) and exercise (68) have recently been reported in obese compared to lean rats. A simplified depiction of positive and negative factors that regulate protein synthesis in skeletal muscle with relevance to the obesity/insulin-resistant (IR) state. IRS, insulin receptor substrate; Akt, protein kinase B; TSC2, tuberous sclerosis complex-2; AMPK, AMP-activated protein kinase; mTOR, mammalian target of rapamycin; S6K1, p70 ribosomal protein S6 kinase 1; 4E-BP1, eukaryotic translation initiation factor 4E-binding protein 1. In normal human muscle, insulin alone can increase muscle protein synthesis up to ∼150% (69) and this is observed in conjunction with increased activity of the Akt/mTOR pathway (65). Activation of Akt in muscle can increase the ability of Akt to phosphorylate and inactivate tuberous sclerosis complex-2 (TSC2), a suppressor of mTOR (Figure 2). On the other hand, decreased activation of Akt in muscle secondary to reduced plasma insulin concentration has been linked to decreased activation of the mTOR pathway and reduced rate of muscle protein synthesis (70).When insulin-stimulated Akt phosphorylation is reduced in IR muscle (71), an impaired Akt activation can maintain the suppressive effects of TSC2 on mTOR, resulting in a reduced rate of muscle protein synthesis. The mTOR pathway also has been implicated in the stimulation of muscle protein synthesis by mechanical stimuli (reviewed in (72)). Studies from obese rats suggest impaired muscle mTOR activation in response to exercise, concurrent with impaired Akt phosphorylation (68). Akt phosphorylation tends to increase after exercise in humans, accompanied by increased muscle protein synthesis (73). Further, whereas the phosphorylation of TSC2 by Akt in selected amino acid residues (Ser939, Thr1462) results in increased mTOR activity, phosphorylation of TSC2 on alternative residues (Thr1271, Ser1387) increases the activity of TSC2 and results in decreased mTOR activity (74). Phosphorylation of the latter amino acid residues has been linked to the activation of muscle AMP-activated protein kinase, that can also inhibit mTOR by directly phosphorylating the mTOR-binding partner raptor (74). This latter mechanism has been implicated in recent findings showing that mechanical overload-induced hypertrophy was impaired as much as 30% in muscles from obese compared to lean rats (75). Increased plasma insulin concentrations following mixed meal ingestion are observed simultaneously with plasma amino acid concentrations maintained at normal or high levels. In an experimental circumstance of insulin infusion, local/intra-arterial infusion of insulin can result in maintenance of plasma amino acid concentrations at basal levels (76). Under conditions where plasma amino acid concentrations are maintained, insulin can stimulate muscle protein synthesis by increasing the delivery of plasma amino acids into the muscle, where plasma amino acids serve both as signaling molecules as well as substrate for protein synthesis (Figure 2). Increasing plasma insulin concentration stimulates muscle blood flow and enhances muscle protein synthesis (76). On the other hand, a blunted blood flow response to insulin, as seen with aging, has been associated with an impaired stimulation of muscle protein synthesis (77). In obese, IR individuals, impaired insulin-mediated vasodilation (78) and muscle capillary recruitment (79,80,81), have been documented, which can impair plasma amino acid delivery into the skeletal muscle and reduce protein synthesis. Recent evidence showing that in healthy humans protein synthesis is not stimulated by insulin when the insulin-induced vasodilation and capillary recruitment are experimentally inhibited (e.g., via inhibition of the nitric oxide pathway) (82), provides support for the above proposed mechanism. Plasma insulin has also been implicated in the stimulation of transmembrane amino acid transport in muscle (83). In vitro evidence, with transportable amino acid analogues, suggests impaired plasma membrane amino acid transport both in the basal as well as in the insulin-stimulated state in obesity (84). The protein content in muscle is determined by the balance between the processes of protein synthesis and breakdown and maintaining a given rate of muscle protein synthesis is central to the maintenance of adequate protein turnover and the renewal of proteins in skeletal muscle. As described above, excess body fat and IR are associated with a reduced rate of protein synthesis in skeletal muscle (11), a condition that can have significant clinical implications given the physiological importance of skeletal muscle in health and disease (85). Reduced muscle mitochondrial content in obesity, as a consequence of an impaired rate of synthesis of mitochondrial proteins (11), can (i) contribute to abnormal metabolic responses related to substrate oxidation and energy production and be a factor to insulin in skeletal muscle leading eventually to the of An impaired muscle protein synthesis in obesity may also have important clinical implications when in the of energy which is to reduce excess body fat in obese individuals. Increased plasma amino acid concentrations due to a physiological state that can muscle protein synthesis provides a stimulus for increased protein synthesis and the maintenance of muscle mass over time in healthy humans may result in less than of amino During energy in obese individuals, lean mass is in to tissue mass This loss of lean body mass may be to a decreased rate of muscle protein a recent observation (i.e., decrease in protein reported in healthy adults following acute energy In both obesity-associated and free fatty (36) insulin increased plasma amino acid concentrations stimulate protein synthesis in exercise combined with acid could therefore be in lean body mass with weight loss in a similar to that described in elderly a decrease in muscle protein synthesis with energy was observed despite an of the for protein (i.e., in healthy adults Such a response may result in decrease in fat-free mass in to the expected decrease in overall body weight as a result of the energy However, when resistance training was during the of energy in obese older individuals fat-free mass was in conjunction with apparent changes in muscle protein synthesis the metabolic of obesity and insulin resistance on muscle protein together with the of both physical activity and protein during energy in muscle protein synthesis in obesity, will be important in the to that loss of body fat and of lean body mass. 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