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Research Papers

An In Vivo Ovine Model of Bone Tissue Alterations in Simulated Microgravity Conditions

[+] Author and Article Information
Benjamin C. Gadomski, Kirk C. McGilvray, Kevin K. Haussler

Orthopaedic Research Center,
Department of Mechanical Engineering
and School of Biomedical Engineering,
Colorado State University,
Fort Collins, CO 80523

Jeremiah T. Easley, Ross H. Palmer

Surgical Research Laboratory,
Department of Clinical Sciences,
Colorado State University,
Fort Collins, CO 80523

E. J. Ehrhart

Veterinary Diagnostic Laboratory,
Department of Clinical Sciences,
Colorado State University,
Fort Collins, CO 80523

Raymond C. Browning

Physical Activity Energetics/
Mechanics Laboratory,
Department of Health and Exercise Science,
Colorado State University,
Fort Collins, CO 80523

Brandon G. Santoni

Phillip Speigel Orthopaedic Research Laboratory,
Foundation for Orthopaedic Research
and Education,
Tampa, FL 33637

Christian M. Puttlitz

Associate Department Head for Graduate Studies
Principal Investigator,
Orthopaedic Research Center,
Department of Mechanical Engineering
and School of Biomedical Engineering,
Colorado State University,
Fort Collins, CO 80523
e-mail: puttlitz@engr.colostate.edu

1Corresponding author.

Contributed by the Bioengineering Division of ASME for publication in the Journal of Biomechanical Engineering. Manuscript received September 4, 2013; final manuscript received October 11, 2013; accepted manuscript posted October 29, 2013; published online February 5, 2014. Editor: Beth Winkelstein.

J Biomech Eng 136(2), 021020 (Feb 05, 2014) (9 pages) Paper No: BIO-13-1408; doi: 10.1115/1.4025854 History: Received September 04, 2013; Revised October 11, 2013; Accepted October 29, 2013

Microgravity and its inherent reduction in body-weight associated mechanical loading encountered during spaceflight have been shown to produce deleterious effects on important human physiological processes. Rodent hindlimb unloading is the most widely-used ground-based microgravity model. Unfortunately, results from these studies are difficult to translate to the human condition due to major anatomic and physiologic differences between the two species such as bone microarchitecture and healing rates. The use of translatable ovine models to investigate orthopedic-related conditions has become increasingly popular due to similarities in size and skeletal architecture of the two species. Thus, a new translational model of simulated microgravity was developed using common external fixation techniques to shield the metatarsal bone of the ovine hindlimb during normal daily activity over an 8 week period. Bone mineral density, quantified via dual-energy X-ray absorptiometry, decreased 29.0% (p < 0.001) in the treated metatarsi. Post-sacrifice biomechanical evaluation revealed reduced bending modulus (–25.8%, p < 0.05) and failure load (–27.8%, p < 0.001) following the microgravity treatment. Microcomputed tomography and histology revealed reduced bone volume (–35.9%, p < 0.01), trabecular thickness (–30.9%, p < 0.01), trabecular number (–22.5%, p < 0.05), bone formation rate (–57.7%, p < 0.01), and osteoblast number (–52.5%, p < 0.001), as well as increased osteoclast number (269.1%, p < 0.001) in the treated metatarsi of the microgravity group. No significant alterations occurred for any outcome parameter in the Sham Surgery Group. These data indicate that the external fixation technique utilized in this model was able to effectively unload the metatarsus and induce significant radiographic, biomechanical, and histomorphometric alterations that are known to be induced by spaceflight. Further, these findings demonstrate that the physiologic mechanisms driving bone remodeling in sheep and humans during prolonged periods of unloading (specifically increased osteoclast activity) are more similar than previously utilized models, allowing more comprehensive investigations of microgravity-related bone remodeling as it relates to human spaceflight.

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Topics: Bone , Stress
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Figures

Grahic Jump Location
Fig. 1

(a) Histological section of rodent bone demonstrating its lamellar structure and fundamental lack of secondary osteons. (b) A transverse section of mature cortical sheep bone demonstrates numerous secondary osteons. (c) Histological section of human compact bone demonstrating a similar secondary osteonal architecture as ovine bone.

Grahic Jump Location
Fig. 2

An external fixation unloading technique was utilized to differentially-unload the metatarsal bone of the right hindlimb. A total of four longitudinal connecting rods, four horseshoe rings, and six fixation pins were utilized. In this configuration, approximately 25% of the ground reaction force (GRF) is transmitted through the metatarsal bone (0.25 g) while 75% of the GRF is redirected through the external fixation device.

Grahic Jump Location
Fig. 3

Ovine hindlimbs were instrumented with orthopedic strain measurement plates and three rosette strain gages for the in vitro strain correlation experiments. (Inset) The central thickness of an orthopedic locking plate was reduced to 0.50 mm and instrumented with a rosette strain gage in order to increase its sensitivity to small strain fluctuations.

Grahic Jump Location
Fig. 4

The in vivo external fixation relative gravity was quantified by calculating the metatarsus bone strains from the acquired measurement plate strains, transforming the bone and device strains into forces using linearly elastic constitutive relationships, and taking the ratio of force transmitted through the metatarsus to total force transmitted through the system

Grahic Jump Location
Fig. 5

Normalized temporal DEXA results for the (top) microgravity and (bottom) Sham Group for the 8 week study period. The treated metatarsus displayed an acute rise in BMD after 2 weeks (*p < 0.01) followed by a rapid, linear decline in BMD due to the microgravity treatment with significant alterations in BMD versus baseline values occurring 6 weeks (+ p < 0.01) and 8 weeks (#p < 0.001) postsurgery.

Grahic Jump Location
Fig. 6

(Top) Four-point bending modulus decreased as a result of microgravity unloading. Groups denoted by like-letters are statistically different from each other: (a) p < 0.05 and (b) p < 0.05. (Bottom) Normalized diametral compression failure load decreased following the microgravity treatment while no changes were evident following the sham treatment: (c) p < 0.001, (d) p < 0.01, and (e) p < 0.01.

Grahic Jump Location
Fig. 7

(a, highlighted) DEXA, μCT, and static histomorphometric analyses were performed in the distal cancellous network of the metatarsus. Three-dimensional μCT reconstructions demonstrated decreased trabecular number, thickness, and bone volume within the cancellous microarchitecture in the (b) microgravity Group treatment metatarsi (highlighted) versus their contralateral (control) metatarsi and those of the (c) Sham Group.

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