THE EFFECTS OF OVERUSE ON CELLULAR, MOLECULAR AND MORPHOMETRIC BONE HOMEOSTASIS IN A VOLUNTARY REPETITIVE STRAIN INJURY RAT MODEL

Abstract

Injuries of the hands and wrist are prevalent in many occupations requiring repetitive tasks and may be further aggravated by advancing age; these injuries are termed work related musculoskeletal disorders (WMSDs). Prior studies using an innovative operant rat model of reaching and grasping as a model of WMSDs demonstrated exposure dependent changes in forelimb bones of young adult rats performing repetitive tasks ≤ 3 months. No one has yet to examine if aging enhances forelimb bone degradative changes occurring with WMSDs, or if forelimb bones adapt or degrade further in response to moderate versus high demand repetitive tasks performed for prolonged time periods (up to 24 months). Bone remodeling is a normal biological process that allows bones to adjust to strains. Unfortunately, both aging and inflammation can deregulate the balance between bone resorption and formation. Aging mammals display increased baseline inflammatory-cytokine levels, both systemically and at the tissue level. Several inflammatory cytokines have been shown to stimulate osteoclastogenesis leading to bone resorption and reduced bone formation. We have reported increased production of inflammatory cytokines in serum and musculotendinous tissues of aged animals performing a repetitive reaching and grasping tasks for up to 12 weeks, warranting further examination of whether aged rats performing these tasks have increased bone resorptive changes, compared to young adult rats. We hypothesized that aging would enhance bone degradative changes in our model as a consequence of increased bone inflammatory responses to a moderate demand repetitive task. Therefore, our first aim was to examine forearm grip strength, trabecular and cortical bone quality, and inflammatory cytokine levels in radii of mature (14-18 mo of age) and young adult (2.5-6.5 mo of age) female Sprague Dawley rats after performance of a high repetition low force (HRLF) task for 12 weeks, compared to each other and age-matched controls. We found that mature rats performing a moderate demand repetitive task for 12 weeks had decreased bone formation and quality, particularly cortical bone quality, compared to young adult rats performing the same task, with increased inflammatory and decreased anti-inflammatory responses, and perhaps lower grip strength, as likely contributors. An adaptive bone response was observed in young adult animals performing a moderate level task of high repetition low force for 12 weeks. In contrast, a previous study showed bone degradative changes in young adult rats performing a high demand task of high repetition high force task for 12 weeks. Osteocytes are the mechanosensing cells of bones, and disruption or changes to their environment can lead to apoptosis or molecular changes. In models of forced bone loading to bone fatigue, osteocyte apoptosis increases sclerostin levels and osteoclast recruitment. Increased sclerostin also leads to increased RANKL production. In contrast, low level loading for a short period reduces sclerostin levels and encourages bone formation. We hypothesized that long-term muscle loading at high repetition low force loads would induce further bone adaptation, but that long-term high repetition high force muscle loading would result in detrimental bone loss, as well as alterations in these two bone remodeling proteins, RANKL and sclerostin. Therefore, our second aim was to determine if prolonged performance of a moderate demand upper extremity reaching and grasping task by young adult rats would continue to enhance forelimb bone formation and quality. We hypothesized that continued performance of a high repetition low force (HRLF) task for 24 weeks would lead to increased bone formation. We also hypothesized that RANKL and sclerostin, two proteins that have not been investigated in our rat model of WMSDs, would be reduced in rats performing a HRLF task for 24 weeks, as the bones reach adaptation. We found that 24 week HRLF rats showed several indices of bone formation and adaptation to the task; as well as reduced sclerostin immunoexpression, compared to controls, a reduction that likely contributed to the enhanced bone formation. To expand on this investigation, in our third aim, we investigated the impact of performance of a high repetition high force (HRHF) task for 18 weeks on young adult rat forelimb bones, and on sclerostin and RANKL levels. We observed detrimental trabecular bone remodeling in the radius, including decreased trabeculae bone volume, number and thickness, increased trabecular separation and anisotropy, and a transition to rod-shaped trabeculae in 18-week HRHF task animals, compared to food restricted control rats. In the 18-week HRHF rats, osteoclast numbers increased and osteoblast numbers decreased, concomitant with increased osteocyte apoptosis and empty lacunae, compared to control rats. Also, mRNA and protein levels of RANKL increased and sclerostin decreased in the 18-week HRHF rats, compared to to control rats. Thus, prolonged performance of a high demand task of high repetition high force induced detrimental trabecular bone changes. The increased RANKL likely contributed to these changes, and although sclerostin level decreased, a change that should contribute to enhanced osteoblast activity, bone formation was not rescued. In conclusion, prolonged performance of a HRLF task by young adult rats leads to reduced sclerostin levels and increased bone formation and bone quality. Aged rats performing the same HRLF task showed increased bone degradative changes that might increase fracture risk. In contrast, prolonged performance of a HRHF task by young adult rats leads to increased bone resorption and degradation, changes associated with RANKL expression. Sclerostin levels were reduced by the HRHF task, but failed to rescue bone formation.