Prolonged high force high repetition pulling induces osteocyte apoptosis and trabecular bone loss in distal radius, while low force high repetition pulling induces bone anabolism
Graphical abstract
Introduction
Most types of physical activity are considered beneficial, whether sports, planned exercise, household work or occupational tasks [1,2]. Yet, work activities are not considered exercise; the former typically lacking any cardiovascular benefits [3]. Involvement in heavy manual occupations is linked to higher incidence of musculoskeletal injuries than nonmanual and light manual occupations [4,5]. The negative effects of repetitive and high compressive forces on lumbar spine structures when performing occupational jobs are well established [[6], [7], [8]]. A small number of studies showing increased incidence of hand/wrist osteoarthritis and reduced bone mass in the hand and wrist in individuals with heavy or one-sided hand workloads [[9], [10], [11], [12]]. This is a concern because incidence of distal radial bone fractures is increasing in the U.S. and worldwide [13,14]. There is also a need for etiologic research examining mechanisms underlying tissue degradative changes occurring with high demand upper extremity occupational tasks [15].
Although not yet investigated with overuse injuries occurring as a consequence of occupational tasks, RANKL (Receptor activator of nuclear factor kappa-B ligand) is expressed by osteoblasts and osteocytes [16], and stimulates differentiation and activation of osteoclasts by binding to its receptor, RANK [17,18]. Osteocytes show metabolic responsiveness to bone loading or unloading [19,20], and RANKL is one protein released by apoptotic osteocytes following loading-induced microdamage [17,21]. Such microdamage has a catabolic effect on bones by promoting osteoclast activity in a RANKL-dependent manner [22]. Osteoprotegerin (OPG) is a RANK decoy receptor that binds RANKL, segregating the ligand and preventing activation of the RANKL/RANK pathway [23]. While osteocyte apoptosis and their subsequent release of RANKL is essential for bone remodeling, a prolonged shift in the RANKL and OPG ratio towards more RANKL enhances osteoclastogenesis and net bone resorption [17].
Sclerostin is another protein expressed by osteocytes [24] that has yet to be examined with overuse occupational injuries. It is the product of the SOST gene and a negative regulator of osteoblast differentiation and function, making it a potent inhibitor of bone formation [25,26]. Although the exact mechanisms by which sclerostin inhibits bone formation are still unclear, it is known that sclerostin selectively inhibits Wnt proteins from binding to low-density lipoprotein receptor-related protein 5/6 (LPR5/6) receptors, a change that antagonizes Wnt signaling [27]. The Wnt pathway is essential for loading-induced osteogenesis [28], and its inhibition suppresses osteoblast activity and reduces osteoblast and osteocyte viability [27]. Applied mechanical loads typically suppress sclerostin production, a change that releases the break on Wnt signaling and allows for bone formation [19]. However, loading-induced matrix microdamage in which osteocyte apoptosis occurs is associated with an enhanced release of both sclerostin and RANKL [26], changes that promote bone catabolism.
We have developed an operant and clinically relevant rat model of upper extremity WMSDs in which rats learn a reaching and lever bar pulling task for a food reward [29]. Rats reach forward using their whole forearm to pull on a lever bar located outside of the chamber (which is attached to a force transducer) at learned and defined reach rates and target forces for a food reward. Young adult rats performing the task at low force loads and high repetition rates (Low-ForceHR) for 12 weeks show adaptive bone changes, including increased trabecular bone volume in the distal metaphysis of the radius [29,30]. This is consistent with studies reporting bone anabolism in response to loading [31]. In contrast, rats performing a high force high repetition (High-ForceHR) task for 12 weeks show significant trabecular bone loss in the distal metaphysis [29,32]. Initially, an underlying inflammatory mechanism contributed to this overuse-induced trabecular bone loss, a mechanism confirmed with findings that anti-inflammatory treatments reduced this loss when provided in task weeks 4 through 12 since the inflammatory changes peaked in task week 6 [32,33]. These inflammatory responses were resolved in serum and muscles in 18 week High-ForceHR task rats [34]. However, we hypothesized that trabecular bone catabolism would persist in distal radial metaphysis of 18 week High-ForceHR rats in association with increased microdamage, osteocyte apoptosis, sclerostin and RANKL. These findings would be consistent with a Damage-Repair Theory for bone in which persistent high loading causes an accumulation of damage in bone if the loading is so high that self-repair mechanisms cannot keep pace with the level of damage or overload-induced resorption [[35], [36], [37], [38]]. That said, as stated in a recent review: “the contributions of varied, but unique, stimuli generated by muscle to bone remains to be established” [39].
To explore this hypothesis, we compared radial and ulnar bone microarchitecture changes in rats performing a High-ForceHR reaching and lever pulling task for 18 weeks to those occurring in rats performing a Low-ForceHR task for 18 weeks (extending our past 12 week studies), versus resting control (C) rats. We examined for indices of bone microdamage and changes in proteins known to modulate loading-induced bone resorption versus formation (e.g., RANKL, RANK, OPG and sclerostin), proteins not yet examined in our operant repetitive loading model or in other models of chronic muscle-contraction loading of bone.
Section snippets
Animals and overview
The Temple University Institutional Animal Care and Use Committee approved all experiments in compliance with NIH guidelines for the care and use of laboratory animals. A total of 70 young (2.5 months of age at onset) female Sprague-Dawley rats were used (one rat was excluded after onset of experiments reducing the total number of rats used to 69; Fig. 1A). All rats were housed in a central animal facility in separate cages with a 12 hour light:dark cycle, with free access to water and
Reach performance across the 18 weeks
We first investigated the consistency of performance of the operant High-ForceHR and Low-ForceHR tasks by the task rats. Each group learned to perform their respective task appropriately by task week 3 (Fig. 2A–D). The High-ForceHR rats performed the task at or near 4 reaches/min in task weeks 3 through 12, although below target levels in weeks 1 and 18 (Fig. 2A), matching past findings of symptoms of forepaw discomfort in High-ForceHR 18 week rats [34]. The Low-ForceHR rats had a tendency to
Discussion
This is the first study, to our knowledge, to show clear support of the Damage Repair Theory response in trabecular bone when loaded at high force levels, rather than just positive adaptive changes in response to prolonged muscle contraction-induced loading. Prolonged high force loading was a key factor driving radial bone microarchitecture and cellular responses. Rats performing a High-ForceHR task for 18 weeks showed several catabolic indices trabeculae in their distal radial metaphyses,
Conclusion
It is commonly assumed that bone mass increases with vigorous exercise and decreases as a result of unloading or reduced mechanical stimulation. We observed different remodeling responses when using a similar repetition rate yet two different stress magnitudes (Low-Force versus High-Force). Performance of a Low-ForceHR task for 18 weeks was matched by positive bone adaptation in both distal trabecular and mid-diaphyseal cortical bone of the radius (and somewhat in the ulna of these rats),
Acknowledgements
Research reported in this publication was supported by the National Institute of Arthritis and Musculoskeletal and Skin Diseases of the National Institutes of Health under Award Number AR056019 to MFB. This study utilized services offered by core facilities of the Center for Substance Abuse Research (Grant Number DA013429), the MicroCT and Imaging Core at Temple University School of Medicine, and Bioquant Image Analysis Corporation. We would also like to thank Vicki Kalscheur at the School of
Competing interests
None of the authors have any competing interests to declare.
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