Skeletal Muscle Remodeling: Interconnections Between Stem Cells and Protein Turnover

Abstract

Nutrition and exercise are important components of a healthy lifestyle to improve rates of hypertrophic and nonhypertrophic skeletal muscle remodeling. We provide evidence to support the hypothesis that muscle stem cells and protein turnover are collaborative, not separate, mechanisms supporting muscle remodeling by facilitating protein, nuclear, and cellular turnover in response to the ingestion of protein dense foods and exercise.

In Brief

We outline evidence to show that interactions between muscle stem cells and protein turnover facilitate both hypertrophic and nonhypertrophic muscle remodeling.

Key Points

• Nutrition and exercise are key components of a healthy lifestyle to augment rates of both hypertrophic and nonhypertrophic muscle remodeling to maintain (or expand) the composition of a healthy proteome.
• Muscle stem cells and protein turnover are commonly recognized as fundamental contributors to muscle remodeling processes but often are viewed as independent regulators toward these muscle remodeling responses.
• Our hypothesis is that muscle stem cells and protein turnover are interconnected and, thus, collaborate to improve rates of muscle remodeling in response to anabolic stimuli to facilitate muscle adaptations, both hypertrophic and nonhypertrophic.

INTRODUCTION

Skeletal muscle is a highly plastic tissue that responds to a variety of physiological and pathological stimuli. The plasticity is due, at least partly, to the constant turnover or remodeling of muscle proteins, organelles, and cell populations. The nonhypertrophic remodeling of skeletal muscle fibers allows for “old” or damaged cellular components to be removed and replaced with new muscle proteins. Thus, muscle remodeling is essential for the maintenance and composition of the proteome that supports skeletal muscle health and physical performance throughout adult life. Muscle fiber remodeling is regulated on various levels including 1) the myogenic stem cells (satellite cells), 2) nonmyogenic stromal cells, 3) gene transcription, and 4) protein synthesis and breakdown. Reductionist approaches have been useful to understand the independent contribution of each of these processes on the maintenance, quality, or growth of skeletal muscle tissue in response to exercise, nutrition, or poor health. However, a more integrative approach is required to better define how these “independent” responses interact to facilitate hypertrophic and nonhypertrophic remodeling of skeletal muscle tissue in the basal state and in response to nutrition, particularly protein ingestion, and exercise stimuli applied either acutely or chronically.

The purpose of this article is to highlight that exercise and a protein dense diet are effective strategies to improve rates of muscle remodeling on a day-to-day basis and this remodeling response can be hypertrophic or nonhypertrophic in nature. We specifically provide evidence to support our hypothesis that interconnections, but temporally distinct in terms of their robustness, exist between muscle protein turnover and myogenic stem cell activity that regulate rates of muscle remodeling by facilitating protein, nuclear, and cellular turnover in response to acute and chronic nutrient intake and exercise.

Contribution of Satellite Cells to Muscle Microenvironment

Satellite cells are normally quiescent myogenic stem cells located between the basal lamina and sarcolemma that in response to various exercise-related stimuli become activated, proliferate, and differentiate to participate in muscle repair and adaptation. Satellite cells exist in a complex and dynamic niche facilitating interaction with both mature myofibers and a heterogenous population of fibro/adipogenic progenitor cells (FAP). FAP plays a key role in the remodeling of skeletal muscle tissue by producing soluble protein, cellular components (i.e., fibroblasts and adipocytes), and extracellular matrix components (i.e., collagens, fibronectin, and laminin) that comprise the interstitium (²²). Cellular components of the satellite cell niche produce signals that regulate satellite cell fate (¹¹), and that satellite cells participate in muscle remodeling by fusing to existing myofibers. However, the role of satellite cells in regulating FAP is not a traditional role applied to satellite cells but could influence muscle remodeling by altering the interstitial environment and contributing to fibrotic and adipose tissue accumulation within muscle.

Models of muscle regeneration, aging, and hypertrophy have shown that satellite cell ablation results in muscle fibrosis, intramuscular adipose tissue accumulation, and FAP expansion (^13,14,30). It was recently shown that activated satellite cells directly regulate fibroblast-mediated extracellular matrix (ECM) remodeling through miR-206-mediated cross talk (¹²). Combined, this circumstantial and direct evidence support the hypothesis that satellite cells are key regulators of FAP proliferation, differentiation, and ECM production and thus provide a new role for satellite cells in skeletal muscle remodeling. Moreover, it has been shown that satellite cells may be involved in neuromuscular junction regeneration (²⁵); a process that commonly is underappreciated in terms of exercise repair and recovery. The role of satellite cells in regulating their local niche has not been directly investigated in humans. Correlational evidence suggests that satellite cells may be serving a similar function in vivo in humans because conditions that are associated with the accumulation of fibrotic and adipose tissue in skeletal muscle such as aging, obesity, and irradiation also are characterized by decreased satellite cell content and regenerative potential (²²). Importantly, Farup et al. (¹⁰) recently demonstrated that prolonged resistance exercise training increased satellite cell content and increased quantity of proliferating FAP, suggesting that satellite cell–FAP interactions also may be occurring in humans. Together, the animal and human data support the hypothesis that satellite cells are not merely passive receivers of information, but that satellite cells may play a role in muscle remodeling through cross talk with interstitial cells and by supporting neuromuscular regeneration. Future studies will need to explore the role of satellite cells in remodeling their own microenvironment in response to both endurance and resistance exercise activities in humans.

Contribution of Satellite Cells to Muscle Fiber Remodeling

Beyond their traditional role in muscle hypertrophy, satellite cells play an important role in muscle remodeling in response to nonhypertrophic stimuli. Specifically, recent work has characterized satellite cells at different stages of their life cycle in response to endurance exercise (^20,21,31). These studies have shown an increase in committed myogenic progenitors (Pax7⁺MyoD⁺) and an increase in differentiating myoblasts (Pax7⁻MyoD⁺) after acute endurance exercise paradigms (i.e., moderate-intensity continuous or interval cycling) (³¹) and with high-intensity interval training (²¹). Interestingly, these training-induced changes in committed and differentiating myogenic progenitors were not accompanied by an increase in muscle fiber cross-sectional area (¹⁹), thereby suggesting that satellite cell expansion and differentiation can occur without hypertrophy.

A question arises from this work: what is the physiological relevance of satellite cell differentiation without significant hypertrophy? Given that the increase in differentiating satellite cells was not accompanied by an increase in the number of myonuclei (^20,21), it could be speculated that fusion events in response to endurance exercise training are so rare that significant changes in myonuclear content cannot be detected using currently available techniques. Thus, these “rare” fusion events are not physiologically relevant. However, work from Joanisse et al. (²⁰) suggests otherwise as increased satellite cell content, including increased committed and differentiating myoblasts, was observed to be specifically associated with hybrid (i.e., expressing multiple myosin heavy chain (MHC) isoforms) muscle fibers. In addition, these hybrid muscle fibers demonstrated an increased prevalence of neonatal MHC expression and centrally located nuclei, which are both suggestive of enhanced muscle remodeling (²⁰). Interestingly, the expression of multiple MHC isoforms is indicative of neuromuscular junction remodeling (³⁵), and satellite cell–derived myonuclei were observed to be concentrated by remodeling neuromuscular junctions (²⁵). As such, satellite cell–derived myonuclear accretion in nonhypertrophic muscle adaptations may have a role in the support of neuromuscular junction remodeling.

Another important physiological role of the addition of satellite cell–derived myonuclei in the nonhypertrophic remodeling process may relate to replacement of old myonuclei with new satellite cell–derived nuclei (i.e., myonuclear turnover). Support for the hypothesis that satellite cells participate in myonuclear turnover, and that this process is important for nonhypertrophic fiber remodeling, can be observed from studies that compare myonuclear turnover in rapidly versus slowly remodeling muscles. For example, extraocular muscles (EOM) undergo continuous remodeling at a rate that exceeds limb muscles and this is facilitated by continuous satellite cell activation (²⁷). In addition, EOM exhibited an increased content of Terminal deoxynucleotidyl transferase (TdT) dUTP Nick-End Labeling (TUNEL)-positive myonuclei and caspase-3 positive myofibers compared with limb muscles (²⁷). These markers of apoptosis were extremely rare, which would be expected because these apoptosis-related assays only provide a single snapshot in time of apoptosis occurring in muscle. However, these findings provide insight that myonuclear turnover may occur at an increased frequency in actively remodeling fibers. In addition, Spalding et al. (³⁸) used ¹⁴C carbon dating to determine the age of various cell types in humans and determined that skeletal muscle cells were younger than the age of the individual from which they were derived. Because ¹⁴C incorporates into DNA during cell division, the authors concluded that skeletal muscle must undergo continuous nuclear turnover (³⁸). Together, these data provide evidence that myonuclear turnover, where old myonuclei continually are replaced by new satellite cell–derived myonuclei, actively occurs in skeletal muscle. In adults under homeostatic conditions, this process is rare resulting in slow myonuclear turnover especially in the absence of a stimulus that augments cellular remodeling (⁴). However, myonuclear turnover likely is enhanced via exercise, including nonhypertrophic modes of exercise. As such, satellite cells may be donating their nuclei to existing myofibers to provide “fresh” genetic material to promote myofiber maintenance and adaptation via protein synthesis with or without hypertrophy.

Protein Turnover and its Role in Muscle Remodeling

Protein turnover at a constant rate is important for the maintenance and composition of the proteome of a tissue. Exercise and protein ingestion, separately or together, have been shown to improve rates of muscle protein remodeling by their ability to modulate changes in protein synthesis and breakdown rates. For example, after eating a protein dense meal, it has been shown that approximately 10% of the ingested protein-derived amino acids being released into circulation directly are used for postprandial muscle protein synthesis and nonhypertrophic remodeling (¹⁶). Exercise before protein ingestion results in more of the dietary protein-derived amino acids being used for the stimulation of postprandial muscle protein synthesis rates in healthy adults (³²), and this enhanced amino acid sensitivity of muscle to food can persist up to 2 d into recovery from a single bout of exercise (⁶). In fact, Boirie et al. (³) has proposed that “new” dietary amino acids ingested in a meal preferentially are used to synthesize proteins and “old” amino acids are shifted toward oxidative pathways. As such, the regular performance of exercise and maintaining a healthy eating pattern that includes protein ingestion is important for facilitating the removal of aged damaged proteins and their replacement with newly synthesized proteins, thereby promoting the skeletal muscle adaptive response.

The type and duration of exercise that is regularly performed dictate whether hypertrophic or nonhypertrophic muscle remodeling will be most dominant after training. Resistance exercise is well documented to enhance muscle protein synthesis within the mitochondrial, sarcoplasmic, and myofibrillar protein fractions (^5,6). The latter usually is measured as a total value of contractile proteins such as myosin, actin, troponin, and tropomyosin. Hasten et al. (¹⁹) demonstrated that resistance exercise is especially potent at hypertrophic remodeling of the MHC component of myofibrillar proteins by enhancing its synthesis rates leading to increases in muscle mass. Lesser studied is the muscle protein remodeling response during recovery from endurance-based exercise. However, endurance exercise also is capable of enhancing the muscle protein synthesis rates within the mitochondrial and myofibrillar protein fractions (⁹) and generally results in nonhypertrophic remodeling by augmenting MHC I isoforms with decreases in hybrid fibers and increased muscle oxidative capacity (¹⁸). It remains to be identified, however, the exact mechanism that underpins how sarcomeres undergo remodeling without disrupting their function. It is believed that newly synthesized myofibrillar proteins, such as myosin isoforms, may be held in a prepool in the sarcoplasm before protein exchange with the outer surface of the myofibril (¹⁵).

Tracer dilution methods for directly measuring muscle protein breakdown rates are challenging to apply during nonsteady conditions (e.g., bolus feeding and exercise), which has limited its assessment in human investigations. As such, the role of catabolic processes in the regulation of muscle protein remodeling to feeding and exercise is less well understood. It seems likely that the processes involved in protein remodeling (e.g., synthesis and breakdown) are coordinated with the amino acids derived from protein breakdown (³⁴) or increases in amino acid oxidation rates (²⁹) also serving as signals for the regulation of changes in protein synthesis. Certainly, postprandial/postexercise muscle protein synthesis rates seem to be more responsive to exercise and protein/amino acid interactions versus proteolytic responses in healthy adults (¹). However, measurements of turnover at the whole body level likely provide the most ideal opportunity to obtain concurrent protein synthesis and breakdown assessments to improve our understanding of the nutritional, using more real-world feeding patterns, and exercise interactions on the “total” protein remodeling response in vivo in humans.

Overall, nutrition and exercise interactions induced hypertrophic or nonhypertrophic remodeling of the proteome through changes in protein synthesis and breakdown to promote the skeletal muscle adaptive response. A program of resistance exercise training is most effective at enhancing hypertrophic remodeling rates to augment muscle strength, and to lesser extent muscle oxidative capacity. Endurance exercise training generally induces nonhypertrophic remodeling rates aimed at improving muscle oxidative capacity with less influence on hypertrophic remodeling to improve muscle strength.

Interconnections Between Stem Cells and Protein Turnover for Muscle Remodeling

From a systems perspective, it seems that the activation of stem cells and protein turnover is interconnected to facilitate muscle remodeling but regulated in a temporally distinct manner despite being exposed to the same nutrient, metabolic, and hormonal milieu after eating or exercise stimuli in vivo in humans. Protein synthesis requires the steady supply of nuclear-derived genetic information to code the formation of new proteins. The accumulation of DNA damage and subsequent deregulation of gene expression results in a decrease in tissue remodeling (³⁷). As such, exercise-induced myonuclear turnover facilitated by the addition of new satellite cell–derived nuclei likely contributes to the maintenance of myonuclear integrity and ensures the continued contribution of genetic information for protein synthesis (Figure). Direct evidence for this hypothesis in vivo in humans is lacking because of the traditional approach of evaluating the role of protein synthesis and satellite cells as separate mechanisms for muscle remodeling.

Figure: Exercise and nutrition signals are sensed at the sarcolemma to augment rates of hypertrophic and nonhypertrophic muscle fiber remodeling. Activated satellite cells engage in cross talk with fibro/adipogenic progenitor cells (FAP) in the interstitium that regulates extracellular matrix deposition and satellite cell differentiation. Differentiation and fusion of satellite cells contribute myonuclei that support both hypertrophic and nonhypertrophic muscle fiber remodeling by providing additional genetic material that provides a substrate for the protein synthetic machinery. The remodeling of the muscle protein fractions, such as myofibrillar, sarcoplasmic, and mitochondrial proteins, occurs through modulations in simultaneously occurring processes of synthesis and breakdown rates. Dietary amino acids are transported into muscle via specific muscle amino acid transporters whereby they can be used to stimulate postexercise muscle protein synthesis and support the muscle adaptive response. Amino acids also can have the amino group removed and converted into a source of metabolic fuel.

Indeed, based on the current methods, these two processes are disconnected temporally because alterations to protein synthesis most commonly are measured on the time course of hours, whereas satellite cell responses are measured in days. In the acute setting, for example, increases in protein synthesis occur first, and being the most robust within hours of an anabolic stimuli and wane over time, whereas maximal satellite cell responses occur days later. What is noteworthy is that the stimulation of muscle protein synthesis rates on a regular basis could lead to hypertrophy independent of a satellite cell response as shown in mouse models. Specifically, it has been demonstrated that significant hypertrophy occurred over the short term without myonuclear accretion via genetic manipulation of pathways known to regular muscle hypertrophy (^2,24) or with overload in satellite cell–depleted mice resulting in an increased myonuclear domain (²⁶). Thus, in the short term, a new set point for the myonuclear domain that facilities protein synthesis seems possible. However, this capacity to expand the myonuclear domain may be determined by the quantity of existing myonuclei (¹⁷), and the ability to maintain this temporary expanded domain eventually requires the addition of new satellite cell–derived myonuclei. Given all this, the data suggest that the initial hypertrophic response mediated by an increased stimulation of muscle protein synthesis rates may be regulated by the number of existing satellite cell–derived myonuclei, and the continued maintenance of hypertrophy eventually requires the addition of new, satellite cell–derived myonuclei to potentially maintain the elevated basal muscle protein synthetic responses commonly observed in the trained versus untrained state (²³).

Human studies have not been conducted to evaluate the temporal relation between satellite cell–derived myonuclear accretion and protein synthesis; however, it is likely that the two processes collaborate to support hypertrophic and nonhypertrophic muscle remodeling. The use of nonsubstrate-specific tracer methods, such as deuterium oxide (D₂O), will provide better insight into the temporal relations between stem cells and protein synthesis that contribute to the various muscle remodeling responses by capturing a greater window under the protein synthesis × time curve into exercise-induced adaptive processes. Recent evidence has shown by the use of D₂O methods to monitor integrated muscle protein synthetic responses that the early remodeling response during a program of resistance training is aimed at nonhypertrophic repair and the late (10th week of training) muscle protein synthetic responses supporting hypertrophic remodeling (⁸).

CONCLUSIONS

Most exercise studies have assessed the regulation of stem cells and protein turnover to the remodeling process during recovery from resistance exercise or overload models, and, as such, their contribution to the remodeling process often is viewed through the lens of “hypertrophic potential” or myofiber repair. This article suggests that stem cell activation and increased protein turnover in response to protein ingestion and exercise have important roles in the remodeling of skeletal muscle tissue beyond hypertrophy. For example, in response to an exercise stimulus, satellite cells regulate their own external microenvironment and contribute new nuclei to myofibers that could assist with the muscle remodeling for nonhypertrophic processes. These new nuclei can replace or compensate for old existing nuclei and thus refresh the cellular transcriptome to provide “optimum” genetic information to the protein synthetic machinery that results in the nonhypertrophic remodeling of the muscle proteome to maintain tissue health.

A healthy lifestyle aimed at optimizing hypertrophic and nonhypertrophic muscle remodeling rates should consist of a nutrient dense eating pattern that includes ingestion of adequate protein (e.g., 0.4 g protein·kg⁻¹ per meal × five daily (³³)) combined with sufficient levels of daily physical activity (³⁶), and depending on the preferred phenotypic goal, should incorporate either resistance- or endurance-based activities into the lifestyle routine. Certainly, resistance exercise–based activities and the acute assessment of muscle remodeling markers can provide qualitative predictive insight into training-induced gains in muscle mass (^5,7,28). However, eating alone or endurance-based activities provide greater insight into nonhypertrophic remodeling potential with predictive chronic adaptations being more related to mitochondrial, sarcoplasmic, MHC remodeling, and protein renewal leading to improved metabolic quality/health, but not overt hypertrophy.