Partial gravity unloading inhibits bone healing responses in a large animal model
Introduction
It is well known that microgravity and the associated inherent reduction in mechanical loading result in substantial loss of BMD and mechanical strength. These issues are particularly important when considering fracture risk during long-duration spaceflight missions. Specifically, the mean rate of BMD loss (in lower body bones) during spaceflight has been shown to be dramatically higher than experienced on Earth, averaging losses of 1–3% per month ([1], [2]). Accordingly, simulations of alterations in mineralized tissue properties as a result of microgravity loading have predicted significantly elevated risk of fracture during long-duration (i.e. Mars) missions ([3], [4]).
What is not as well understood is how the mechanical unloading associated with spaceflight affects the fracture healing cascade. Fracture healing is a complex biological process with four distinct phases. The first step of the fracture healing cascade, occurring at the time of fracture, is the formation of a hematoma, preventing further blood accumulation at the fracture site (Kolar et al., 2011). Subsequently a soft callus forms as chondrocytes create new cartilage that bridges the two ends of the disjoined bone, providing initial mechanical competency (i.e. splinting) to the fracture (Einhorn, 1998). Finally osteoblasts replace the new cartilage with woven bone, which in turn is remodeled into a compact secondary osteonal bone structure (Schindeler et al., 2008). It has been postulated that this reparative process represents a recapitulation of development that involves complex mechanical and chemical factors ([8], [9]). Further, it has been shown that the mechanical loading environment of the healing callus has a profound effect on the resultant cell differentiation and heterogeneous matrix phenotypes (“mechanobiology”) ([10], [11], [12], [13]). In vivo experiments using fracture models have elucidated certain cellular and molecular scale events that are important in the repair process. However, alterations in the local mechanics induced via reduced gravitational loading in the repair process have not been rigorously described in bone tissues that have Haversian systems. Nonetheless it is clear that the mechanical environment, which transcends many length scales (from whole body to the subcellular), plays a key role in the subsequent fracture healing pathway and, ultimately, the nature and quality of the osseous repair. This certainly has significant implications when examining how microgravity, which impacts the relevant mechanical environment on all of these length scales, may affect bone healing.
Limited research has been performed to investigate mineralized tissue healing in microgravity environments ([14], [15], [16], [17]). Rodent studies have elucidated distinct histological and morphometric differences in animals that heal in gravitational environments versus animals that heal during spaceflight. Using a rat model, Kirchin et al. showed that bone healing was altered during spaceflight, resulting in suppression of chondrogenesis within the periosteal reaction and angiogenesis within the osteotomy gap (1995). Additionally, Durnova et al. reported decreased fracture callus size and consolidation strength resulting from inhibited osteoblast activity in rodents during a 14-day spaceflight experiment (1991). While these data demonstrate that microgravity has a deleterious effect on bone healing, the direct translation of these results to human bone healing is intractable due to the numerous differences between rodent and human bone microstructure and healing. Specifically, the basic microstructure of rodent bone can be observed as a primary lamellar structure lacking the osteonal (Haversian) systems characteristic of human bone ([18], [19]). Further, it has been shown that the rate of bone healing is known to be inversely related to the species׳ ranking on the phylogenetic scale (den Boer et al., 1999). Consequently, the healing potential of rodent bone far exceeds that of adult human tissue ([20], [21]). Due to these distinct differences between species and the limited information regarding fracture healing in Haversian systems, the objective of this study was to investigate the effects of partial gravitational unloading on long-bone fracture healing in a previously developed large animal model using parameters derived from biomechanical, histomorphometric, and radiographic assessments.
Section snippets
Methods
Skeletally mature Rambouillet Columbian ewes (age>6 years) were used in this study. Animal use approval was granted by the Colorado State University Animal Care and Use Committee (Approval #11-2938A). Hindlimb metatarsal unloading was accomplished using the technique described by Gadomski et al. (2014). Briefly, a trans-biarticular external skeletal fixation device (IMEX, Longview, TX) was implanted on the right hindlimb of 5 skeletally mature female ewes such that the metatarsal bone was
Results
All data are presented as mean (+standard deviation).
Discussion
The 3-week simulated microgravity treatment utilized in this study was performed prior to inducing the ostectomy (i.e. fracture) in order to model the expected loss of BMD encountered by the typical astronaut during a 6-month spaceflight to Mars. There is compelling evidence to indicate that the BMD loss experienced during a Mars mission will significantly alter the mechanical integrity of mineralized tissue and increase the risk of bony fracture (Keyak et al., 2009). Specifically, on average,
Conflict of interest statement
All authors have no conflicts of interest.
Acknowledgments
This work was supported by the National Aeronautics and Space Administration (NNX11AQ81G). Special thanks to Dana Ruehlman, DVM, Kimberly Menges, B.S., Desiree Shasa, M.S., and Matthew Roberts, M.S., of the Surgical Research Laboratory for animal care, Cecily Broomfield, M.S., of the Orthopaedic Research Laboratory for histological preparation, and Courtney Sadar and Alisha Eskew for experimental assistance and data analysis.
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