Cigarettes Make You Weak: RANKL/RANK Link Changes in Muscle and Bone.

It is increasingly appreciated that the chronic use of tobacco cigarettes leads to systemic inflammation that results in cellular and epigenetic changes throughout the whole organism. Signs of pathology can be detected in many organs even before there is evidence of lung damage. Patients with chronic obstructive pulmonary disease (COPD) often exhibit frailty owing to both a decrease in limb bone mass density (BMD) and muscle atrophy and weakness, referred to as sarcopenia (1, 2). Up to 70% of patients with COPD who exhibit osteoporosis also show signs of muscle wasting (3).

High levels of the RANKL (receptor activator of nuclear factor κ-B ligand) early on were detected in the saliva and teeth of heavy cigarette smokers and were associated with periodontal disease and tooth loss (4, 5). In bones, RANKL activates the differentiation of osteoclasts by binding and activating its receptor, RANK, thereby increasing bone resorption (6). In the extracellular space, OPG (osteoprotegerin) acts as a decoy receptor to downregulate RANKL–RANK interactions (7). Together, RANKL, RANK, and OPG expression regulate bone density. It is well known that muscle mass and force are strong indicators of bone health, and low BMD is usually detected in individuals with sarcopenia (8). More recently, it has been demonstrated that the use of tobacco cigarettes activates the RANKL/RANK/OPG pathway in osteoclasts and contributes to a low BMD and eventually to osteoporosis (9). Interestingly, postmenopausal women given a RANKL inhibitor for several years to treat their osteoporosis showed improvements in hand grip strength (10). In mice, overexpression of RANKL also leads to osteoporosis and decreases running speed, muscle mass, and oxidative myofibers, which can be restored by treating with a truncated OPG or with the RANK monoclonal antibody denosumab (10). Furthermore, in denervated muscle, RANK regulates myofiber Ca2+ storage through Stim1 (stromal interaction molecule 1), an important Ca2+ sensor that also regulates the activity of SERCA (sarco[endo]plasmic reticulum Ca2+-ATPase) and muscle force (11, 12). Thus, there is precedent for the RANKL/RANK/OPG system in regulating muscle atrophy, fiber type transition, and contractile function.

In this issue of the Journal, Xiong and colleagues (pp. 617–628) show that chronic cigarette smoke (CS) exposure in mice activates the RANKL/RANK/OPG pathway in skeletal myofibers and contributes to smoke-induced hind limb muscle dysfunction (13). The authors had previously found that patients with COPD with low BMD (14) have higher levels of plasma RANKL compared with smokers without COPD or patients with COPD with normal BMD. The present study investigates the mechanisms by which RANKL/RANK/OPG contributes to muscle dysfunction brought about by long-term smoking. Xiong and colleagues (13) found that in mice exposed to daily periods of CS for 6 months, RANKL and RANK in myofibers increased and were localized to the sarcolemma. Surprisingly, when mice were treated with a RANKL antibody, CS-induced muscle dysfunction was prevented, and muscle mass, grip strength, and exercise endurance were maintained. Importantly, treatment with RANKL antibody attenuated muscle CS-induced inflammatory signaling (TNF-α, IL-6, and NFκB-p65) and atrophy-associated genes (myostatin, atrogin-1, MuRF1) (Figure 1). Therefore, the authors concluded that RANKL/RANK/OPG activation during CS exposure triggers hind limb muscle atrophy and weakness through inflammatory and atrophy signaling pathways (13). The direct effects of smoke components were also tested in differentiated C2C12 myotubes. Myotubes incubated with the water-soluble components of CS (CS extract) showed 2- to 3-fold higher RANK and RANKL expression together with stimulated atrophy and TNF-α–related inflammatory signaling. These changes were inhibited by RANKL siRNA (13). Their data suggest that tobacco CS components enter the bloodstream of smokers and directly stimulate myofiber RANKL expression to alter the inflammatory and atrophy pathways that lead to muscle atrophy and weakness.

Figure 1.

Mechanism for muscle atrophy owing to chronic cigarette smoke exposure in mice triggered by the expression of RANKL. Three processes may be implicated in RANKL-dependent cigarette smoke–induced atrophy: 1) atrophy-associated gene activation (myostatin and Ubiquitin E3 ligases), 2) activation of proinflammatory cytokines, and 3) myofibrillar contractility controlled by intracellular Ca2+ handling and myofilament function. DHRP = dihydropyridine receptor; OPG = osteoprotegerin; MuRF1 = muscle ring-finger protein 1; RANK = receptor activator of NFκB; RANKL = RANK ligand; RYR = ryanodine receptors; SERCA = SR Ca2+-ATPase; SR = sarcoplasmic reticulum.

RANKL/RANK alters three processes known to contribute to muscle atrophy: 1) cytokine signaling, 2) upregulation of atrophy pathways, and 3) intracellular Ca2+ regulation. In the study by Xiong and colleagues (13), grip strength was used as an indicator of decreased muscle function and was accompanied by changes in cytokine signaling and atrophy pathways in CS-exposed mice. CS extract treatment in vivo has been previously shown to dysregulate myofiber Ca2+ handling through a slowing of Ca2+ pumping and SERCA function during contractions, which, in turn, reduces fatigue resistance (15). Future studies are necessary to more fully explore the role of the RANKL/RANK/OPG pathway in intracellular Ca2+ handling (e.g., Ca2+ stores and flux during contractions) and myofibrillar contractility (e.g., force development/kinetics and fatigue resistance) during smoke exposure.

Interestingly, the work of Xiong and colleagues (13) and others show that RANKL/RANK/OPG are biomarkers that can be used to monitor muscle and bone health in smokers, current and former, and patients with COPD. It should be noted that the RANKL/RANK/OPG pathway is not limited to muscle and bone but also plays a role in the immune system in bone marrow development, lymph node and thymus development, gastrointestinal tract, and central nervous system. RANKL also plays a role in the regulation of mammary glands, the thermoregulatory center, blood vessels, hair follicles, and liver (16). Interestingly, expression levels are roughly twice as high in the brain than bone (10). Thus, there is a potential for future therapies that regulate the RANKL/RANK/OPG pathway (e.g., RANK receptor antagonists, RANKL monoclonal antibodies, or OPG treatment) to treat patients with COPD. Care should be taken as it is not known whether RANKL therapies will have beneficial or harmful effects on other tissues that rely on this pathway for their normal function.

In summary, the work of Xiong and colleagues (13) provides valuable information about a critical signaling pathway involved in the activation of muscle protein degradation leading to muscle atrophy. Additional studies are needed to investigate several important unanswered questions: 1) How does smoke exposure or its components activate the expression of RANKL in myofibers? 2) Does smoking also change the extracellular concentration of OPG? 3) Are muscle calcium handling and contractile function regulated by RANKL signaling in smokers? And 4) do the effects of RANKL expression on muscle mass occur before the development of pulmonary symptoms? Their study defines a new mechanistic link between muscle and bone that is likely to be present in cigarette smokers and those that go on to develop COPD.