The major finding of the present study is that jump exercise during skeletal unloading showed a positive effect on the suppression of tail suspension-induced osteopenia at the femur in growing rats. Jump exercised rats retained cancellous bone mass in the distal femur, as evidenced by the increase in trabecular thickness, but did not have a change in trabecular number. These results suggest that jump exercise applied during hindlimb unloading can prevent the microarchitectural deterioration of trabecular bone by an increase in trabecular thickness while controlling the reduction of trabecular number in the distal femoral metaphysis.
Exercise has been widely accepted as being beneficial to skeletal bone. Various types of mechanical loading in rat models including treadmill running (Iwamoto et al. 1999), jumping (Umemura et al. 1997), whole body vibration (Tezval et al. 2011), swimming (Hart et al. 2001), and resistance exercise (Notomi et al. 2000) have been examined for any influence on bone loss and each has proved effective. However, the required time for daily exercise is much longer in treadmill running, vibration, swimming, and resistance exercise (for over 60 minutes) when compared to jump exercise (about one minute for 30 jumps). The effects of hindlimb unloading can be reversed after the rat is removed from the tail suspended state for an extended time. Such an exercise program requiring an extended length of time is unsuitable as a protocol during the hindlimb unloading state. Conversely, jump exercise has an advantage as an exercise program during the hindlimb unloading state, because it can be completed within approximately one minute. So far little is known about the effect of an exercise intervention during an unloading period on trabecular bone architecture. To the best of our knowledge, this study is the first to assess the effect of jump exercise on trabecular bone architecture during tail suspension.
A number of in vivo and in vitro studies have been conducted during spaceflight to clarify the mechanisms underlying the bone loss induced in a microgravity environment. However, the availability of spaceflight experiments is extremely limited. Ground-based studies have thus been conducted to investigate the mechanisms of osteopenia induced by weightlessness. The rat tail-suspension model has been well justified in previous studies as an appropriate model for the study of simulated bone changes induced by weightlessness (Martin 1990; Wronski and Morey-Holton 1987). Previous studies have reported that tail suspended rats display reduced trabecular bone mass, as evidenced by decreases in trabecular number and increases in trabecular separation (Basso et al. 2005; Ju et al. 2012; Ju et al. 2008). The present findings are consistent with these reports. In the present study, as expected, tail suspended rats demonstrated significantly decreased trabecular bone volume (−37%, p < 0.05) as a result of decreasing trabecular number (−32%, p < 0.001) and increasing trabecular separation (+19%, p < 0.001), and essentially no change in trabecular thickness in the distal femoral metaphysis. The tail suspended rats exhibited a loss of trabeculae, particularly from the central zone of the femur with a loss of trabecular number, although the remaining trabeculae did not show significant differences in thickness compared with control group (Figure 4). These results imply that loss in cancellous bone during 21 days of hindlimb unloading is predominantly due to decreases in trabecular bone number as opposed to trabecular thickness. This assumption is supported by the work of Basso et al. (Basso et al. 2005), who demonstrated that deterioration in the trabecular network after 14 days of skeletal unloading are mostly attributable to decreases in trabecular number with slight thinning of trabecular thickness. This phenomenon has also been reported in the ovariectomized rat model (Chen et al. 2001; Ito et al. 2002). Furthermore, similar findings have been confirmed in humans subjected to unloading caused by spinal cord injury (Modlesky et al. 2004) and prolonged bed rest (Armbrecht et al. 2011).
Several previous histomorphometric analyses have found that the increase in trabecular bone mass that occurs with resistance exercise is primarily due to increased trabecular thickness, rather than noticeable changes in numbers of trabeculae (Holy and Zerath 2000; Notomi et al. 2000; Notomi et al. 2001). Moreover, we recently found that restoration of trabecular bone architecture induced by jump exercise during remobilization is predominantly attributable to increased trabecular thickness (Ju et al. 2012). The present findings are consistent with previous studies where jump exercise applied during a skeletal unloading period produced a significant increase in distal femur trabecular bone mass, primarily by increased trabecular thickness without significant change in trabecular number. These results imply that the cancellous bone gain induced by jump exercise during hindlimb unloading is predominantly attributable to increases in trabecular thickness while controlling the reduction in trabecular number. In contrast, Swift et al. (2011) employing simulated resistance training during periods of disuse, found that muscle contractions once every three days produced significantly larger increases in proximal tibial cancellous bone mass associated both with greater trabecular thickness and number than untreated hindlimb unloaded rats. The reason for the apparent discrepancy between these studies is twofold. First, the difference in the mode of mechanical stimulation applied during hindlimb unloading might produce the difference in trabecular architecture. Since Swift et al. (2011) used the low-magnitude, high-frequency mechanical stimulation, the distribution and duration of mechanical stress in bone by the stimulation could be substantially different from the stress induced by jump exercise. In humans, high-impact activity such as jump exercise elicits greater bone gain than repetitive high-frequency activities such as running or walking (Fuchs et al. 2001; Kohrt et al. 2004). Thus, different mechanical loading activities may have different mechanisms of action on the trabecular structural characteristics. Further study is needed to elucidate the mechanism of differential effects among different types of mechanical loading on trabecular bone architecture. It is also possible that different findings are related to differing assessment methods (i.e., analyses of the 3D trabecular bone microarchitecture by micro-CT versus 2D trabecular bone microarchitecture by histomorphometry). The volume of interest provided by 2D sections may not accurately reflect the change across the entire volume of the metaphyseal region. However, the micro-CT may allow for the assessment of true 3D quantification of trabecular bone microarchitecture.
Changes in body weight and muscle mass may play important roles in the regulation of bone mass. Both body weight and hindlimb muscle mass have decreased in rats subjected to hindlimb unloading with tail suspension. A greater weight loss during hindlimb unloading was observed in slow-twitch soleus muscle (47%) than in the fast-twitch gastrocnemius muscle (18%), which is consistent with previous findings (Hurst and Fitts 2003). It has been well documented that the soleus muscle plays a major role in anti-gravity function and is highly dependent on gravity for the normal expression of protein mass and slow phenotype; whereas the gastrocnemius muscle is physically active and does not have the same type of anti-gravity function. During hindlimb unloading, anti-gravity muscles such as the soleus are therefore more vulnerable to atrophy than the gastrocnemius muscle. In the present study, hindlimb muscle atrophy was not fully prevented with jump exercise. Nevertheless, the final bone volume was greater in the JUM group than in the CON and SUS groups. A similar effect was described by Notomi et al. (2003), who reported that the climbing exercise increased bone mass and strength without significantly increasing hindlimb muscle weight after 8 weeks of exercise. In the jump exercise protocol that was used, the bones of the lower limb were loaded only by the ground-reaction force and muscle contraction force at the point of jump take-off. There was no ground-reaction force at landing because the rats were gently placed with their forelimbs on the floor by a technician. These data suggest that the suppression of bone loss and architectural deterioration observed in jump-exercised rats is derived primarily from the mechanical stress generated by the ground-reaction or muscle contraction occurring at take-off.
The present study displays several limitations. First, the study was conducted on the rat hindlimb unloading model combined with the jump exercise training. Experimental animals such as rats are useful for defining biological mechanisms but findings are not necessarily directly applicable to the human situation, since rat bone-modeling patterns are different from human patterns. Furthermore, jump exercise is actually difficult to apply to astronauts during missions or patients with disuse osteoporosis. In the present study, jump exercise was performed in a gravity environment after removal from the tail suspension apparatus. Our results might differ if the same exercise was performed in space. Hence, specially designed workout equipment has to be developed to provide the stimulus to bone, in order to apply this methodology to astronauts or patients with disuse osteoporosis. Nonetheless, the results of this study will improve our understanding of the architectural adaptation of trabecular bone with mechanical stimulation applied during skeletal unloading such as weightlessness and immobility. Bone loss and architectural deterioration caused by inactivity and immobility is one of the major factors for osteoporosis. Recently, several anabolic agents for the treatment of osteoporosis are being developed, which could partly share mechanisms of action with the exercise stimulation. Thus, our data may provide valuable insights into the development of therapeutic agents with effect similar to exercise stress. Secondly, we did not perform the histomorphometric analysis and cannot confirm tissue-level mechanisms for the functional adaptation of the trabecular architecture to mechanical loading observed in the present study. Further study will be needed to clarify the mechanism of mechanical stimulation on trabecular bone architecture.