Name: in vivo evaluation of fracture callus development during bone healing in mice using an mri-compatible osteosynthesis device for the mouse femur
Uploaded: 2017-11-14T15:00:00
Description: Watch this Scientific Journal Video about In Vivo Evaluation of Fracture Callus Development During Bone Healing in Mice Using an MRI-compatible Osteosynthesis Device for the Mouse Femur at JoVE.com

We thank Sevil Essig, Stefanie Schroth, Verena Fischer, Katja Prystaz, Yvonne H&#228;gele, and Anne Subgang for excellent technical support. We also thank the German Research Foundation (CRC1149, INST40/499-1) and the AO Trauma Foundation Germany for funding this study.

All animal experiments complied with international regulations for the care and use of laboratory animals and were approved by the regional regulatory authorities (No. 1250, Regierungspr&#228;sidium T&#252;bingen, Germany). All mice were maintained in groups of two to five animals per cage on a 14-h light, 10-h dark circadian rhythm with water and food provided ad libitum.
1. Preparation of the Surgical Material and Pre-treatment of the Mice
<ol>
	<li>Sterilize all surgical material...

<ol>
	<li>Claes, L., Recknagel, S., Ignatius, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fracture+healing+under+healthy+and+inflammatory+conditions.">Fracture healing under healthy and inflammatory conditions.</a> Nat Rev Rheumatol. 8 (3), 133-143 (2012).</li><li>Einhorn, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+cell+and+molecular+biology+of+fracture+healing.">The cell and molecular biology of fracture healing.</a> Clin Orthop Relat Res. (355), S7-S21 (1998).</li><li>Einhorn, T. A., Gerstenfeld, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fracture+healing:+mechanisms+and+interventions.">Fracture healing: mechanisms and interventions.</a> Nat Rev Rheumatol. 11 (1), 45-54 (2015).</li><li>Augat, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Local+tissue+properties+in+bone+healing:+influence+of+size+and+stability+of+the+osteotomy+gap.">Local tissue properties in bone healing: influence of size and stability of the osteotomy gap.</a> J Orthop Res. 16 (4), 475-481 (1998).</li><li>Claes, L. E., Heigele, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnitudes+of+local+stress+and+strain+along+bony+surfaces+predict+the+course+and+type+of+fracture+healing.">Magnitudes of local stress and strain along bony surfaces predict the course and type of fracture healing.</a> J Biomech. 32 (3), 255-266 (1999).</li><li>Claes, L. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+mechanical+factors+on+the+fracture+healing+process.">Effects of mechanical factors on the fracture healing process.</a> Clin Orthop Relat Res. (355), 132-147 (1998).</li><li>Hu, D. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cartilage+to+bone+transformation+during+fracture+healing+is+coordinated+by+the+invading+vasculature+and+induction+of+the+core+pluripotency+genes.">Cartilage to bone transformation during fracture healing is coordinated by the invading vasculature and induction of the core pluripotency genes.</a> Development. 144 (2), 221-234 (2017).</li><li>Hankenson, K. D., Zimmerman, G., Marcucio, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biological+perspectives+of+delayed+fracture+healing.">Biological perspectives of delayed fracture healing.</a> Injury. 45, 8-15 (2014).</li><li>Meyer, R. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Age+and+ovariectomy+impair+both+the+normalization+of+mechanical+properties+and+the+accretion+of+mineral+by+the+fracture+callus+in+rats.">Age and ovariectomy impair both the normalization of mechanical properties and the accretion of mineral by the fracture callus in rats.</a> J Orthop Res. 19 (3), 428-435 (2001).</li><li>Nikolaou, V. S., Efstathopoulos, N., Kontakis, G., Kanakaris, N. K., Giannoudis, P. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+influence+of+osteoporosis+in+femoral+fracture+healing+time.">The influence of osteoporosis in femoral fracture healing time.</a> Injury. 40 (6), 663-668 (2009).</li><li>Haffner-Luntzer, M., Kovtun, A., Rapp, A. E., Ignatius, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+Models+in+Bone+Fracture+Healing+Research.">Mouse Models in Bone Fracture Healing Research.</a> Current Molecular Biology Reports. 2 (2), 101-111 (2016).</li><li>Garcia, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rodent+animal+models+of+delayed+bone+healing+and+non-union+formation:+a+comprehensive+review.">Rodent animal models of delayed bone healing and non-union formation: a comprehensive review.</a> Eur Cell Mater. 26, 1-14 (2013).</li><li>Histing, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Small+animal+bone+healing+models:+standards,+tips,+and+pitfalls+results+of+a+consensus+meeting.">Small animal bone healing models: standards, tips, and pitfalls results of a consensus meeting.</a> Bone. 49 (4), 591-599 (2011).</li><li>Zachos, T. A., Bertone, A. L., Wassenaar, P. A., Weisbrode, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rodent+models+for+the+study+of+articular+fracture+healing.">Rodent models for the study of articular fracture healing.</a> J Invest Surg. 20 (2), 87-95 (2007).</li><li>Taha, M. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+the+efficacy+of+MRI+for+detection+of+changes+in+bone+morphology+in+a+mouse+model+of+bone+injury.">Assessment of the efficacy of MRI for detection of changes in bone morphology in a mouse model of bone injury.</a> J Magn Reson Imaging. 38 (1), 231-237 (2013).</li><li>Haffner-Luntzer, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+high-resolution+In+Vivo+MRI+for+longitudinal+analysis+of+endochondral+fracture+healing+in+mice.">Evaluation of high-resolution In Vivo MRI for longitudinal analysis of endochondral fracture healing in mice.</a> PLoS One. 12 (3), 0174283 (2017).</li><li>Beckmann, N., Falk, R., Zurbrugg, S., Dawson, J., Engelhardt, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Macrophage+infiltration+into+the+rat+knee+detected+by+MRI+in+a+model+of+antigen-induced+arthritis.">Macrophage infiltration into the rat knee detected by MRI in a model of antigen-induced arthritis.</a> Magn Reson Med. 49 (6), 1047-1055 (2003).</li><li>Al Faraj, ., Shaik A, S. u. l. t. a. n. a., Pureza, A., A, M., Alnafea, M., Halwani, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preferential+macrophage+recruitment+and+polarization+in+LPS-induced+animal+model+for+COPD:+noninvasive+tracking+using+MRI.">Preferential macrophage recruitment and polarization in LPS-induced animal model for COPD: noninvasive tracking using MRI.</a> PLoS One. 9 (3), 90829 (2014).</li><li>Rolle, A. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ImmunoPET/MR+imaging+allows+specific+detection+of+Aspergillus+fumigatus+lung+infection+in+vivo.">ImmunoPET/MR imaging allows specific detection of Aspergillus fumigatus lung infection in vivo.</a> Proc Natl Acad Sci U S A. 113 (8), 1026-1033 (2016).</li><li>Niemeyer, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Non-invasive+tracking+of+human+haemopoietic+CD34(+)+stem+cells+in+vivo+in+immunodeficient+mice+by+using+magnetic+resonance+imaging.">Non-invasive tracking of human haemopoietic CD34(+) stem cells in vivo in immunodeficient mice by using magnetic resonance imaging.</a> Eur Radiol. 20 (9), 2184-2193 (2010).</li></ol>

Modifications and Troubleshooting:
The main goal of this study was to describe a protocol for using of an MRI-compatible external fixator for femur osteotomy in the mouse with the ability to monitor callus tissue development longitudinally during the endochondral fracture-healing process. The design of a custom-made mounting device for insertion of the external fixator ensured a standardized position during repeated scans. Semi-automatic tissue segmentation allows the analysis...

Secondary fracture healing is the most common form of bone healing. It is a complex process mimicking specific aspects of ontogenic endochondral ossification1,2,3. The early fracture hematoma predominantly consists of immune cells, granulation and fibrous tissue. Low oxygen tension and high biomechanical strains hamper osteoblast differentiation at the fracture gap, but promote the differentiation of progenitor cells into chondrocytes4,5,6. These cells start to proliferate at the site of injury to form a cartilaginous matrix providing initial stability of the fractured bone. During callus maturation, chondrocytes become hypertrophic, undergo apoptosis, or trans-differentiate into osteoblasts. Neovascularization at the cartilage-to-bone transition zone provides elevated oxygen levels, allowing the formation of bony tissue7. After bony bridging of the fracture gap, biomechanical stability is increased and osteoclastic remodeling of the external fracture callus occurs to gain physiological bone contour and structure3. Therefore, the amounts of fibrous, cartilaginous, and bony tissue in the fracture callus provide important information about the bone healing process. Disturbed or delayed healing becomes visible by alterations of callus tissue development both in humans and mice8,9,10,11. Available in vivo techniques to longitudinally monitor callus tissue development in preclinical fracture healing studies using small animals include digital radiography and &#181;CT imaging12,13. However, both techniques are only able to discriminate between mineralized and non-mineralized tissue. In contrast, MRI provides excellent soft tissue contrast and might therefore be able to identify soft tissue and cartilage in the fracture callus.
Previous work showed promising results for post mortem MRI in mice with articular fractures14 and in vivo MRI in mice during intramembranous bone-defect healing15. However, both studies also stated limited spatial resolution and tissue contrast. We previously demonstrated the feasibility of high-resolution in vivo MRI for longitudinal assessment of soft callus formation during murine endochondral fracture healing16. Here, we report the protocol for using an MRI-compatible external fixator for femur osteotomy in mice in order to monitor callus tissue development longitudinally during the endochondral fracture healing process. The design of a custom-made mounting device for insertion of the external fixator ensured a standardized position during repeated scans.

<table style="border-collapse:collapse;width:823pt" width="1097">
	<colgroup>
		<col style="width:254pt" width="338" />
		<col style="width:295pt" width="393" />
		<col style="width:149pt" width="199" />
		<col style="width:125pt" width="167" />
	</colgroup>
	<tbody>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="width:254pt;height:15.75pt" width="338">Anaesthesia tube</td>
			<td class="xl65" style="width:295pt" width="393">FMI, Seeheim, Germany</td>
			<td class="xl64" style="width:149pt" width="199">ZUA-82-ANA-TUB-Mouse</td>
			<td class="xl67" style="width:125pt" width="167"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Anaesthetic machine&#160;</td>
			<td class="xl65">FMI, Seeheim, Germany</td>
			<td class="xl64" style="width:149pt" width="199">ZUA-82-GME-MA</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Artery forceps&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BH104R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Autoclave</td>
			<td class="xl65">Systec, Wettenberg, Germany</td>
			<td class="xl64" style="width:149pt" width="199">DX-150</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Autoclaving packaging</td>
			<td class="xl65">Stericlin, Feuchtwangen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">2301-04/06/10/12/16</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Avizo software</td>
			<td class="xl65">FEI, Burlington, USA</td>
			<td class="xl64" style="width:149pt" width="199">-</td>
			<td class="xl67">Version 8.0.1</td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">BioSpec 117/16 magnetic resonance imaging system</td>
			<td class="xl65">Bruker Biospin, Ettlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">117/16</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Bulldog clamp&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BH 021R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Carbon steel scalpel no. 11/15</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BA211/215</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Ceramic mounting pin 0.45 mm&#160;</td>
			<td class="xl65">RISystem, Davos, Switzerland</td>
			<td class="xl64" style="width:149pt" width="199">HS691490</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Clindamycin (300 mg / 2ml)</td>
			<td class="xl65">Ratiopharm, Ulm, Germany</td>
			<td class="xl64" style="width:149pt" width="199">-</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Dressing forceps 115 mm&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BD210R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Dressing forceps 130 mm&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BD025R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Drill bit coated 0.45 mm&#160;</td>
			<td class="xl65">RISystem, Davos, Switzerland</td>
			<td class="xl64" style="width:149pt" width="199">HS820420</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Durogrip needle holder 125 mm&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BM024R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Foliodrape&#160;</td>
			<td class="xl65">Hartmann, Heidenheim, Germany</td>
			<td class="xl64" style="width:149pt" width="199">2513026</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Frekaderm</td>
			<td class="xl65">Fresenius, Bad Homburg, Germany</td>
			<td class="xl64" style="width:149pt" width="199">4928211</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Gigli saw 0.44 mm&#160;</td>
			<td class="xl65">RISystem, Davos, Switzerland</td>
			<td class="xl64" style="width:149pt" width="199">RIS.590.110.25</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Hand drill</td>
			<td class="xl65">RISystem, Davos, Switzerland</td>
			<td class="xl64" style="width:149pt" width="199">RIS.390.130-01</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Heating plate&#160;</td>
			<td class="xl65">FMI, Seeheim, Germany</td>
			<td class="xl64" style="width:149pt" width="199">IOW-3704</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Hygonorm gloves&#160;</td>
			<td class="xl65">Hygi, Telgte, Germany</td>
			<td class="xl64" style="width:149pt" width="199">2706</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Isoflurane</td>
			<td class="xl65">Abbot, London, UK</td>
			<td class="xl64" style="width:149pt" width="199">Forene</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Micro forceps 155 mm&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BD343R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Micro scissors 120 mm&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">FD013R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Mouse FixEx L 0.7 mm&#160;</td>
			<td class="xl65">RISystem, Davos, Switzerland</td>
			<td class="xl64" style="width:149pt" width="199">RIS.611.300-10</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Needle case for drills&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BL911R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Needle holder</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BB078R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Octenisept</td>
			<td class="xl65">Sch&#252;lke, Norderstedt, Germany</td>
			<td class="xl64" style="width:149pt" width="199">121403</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Osirix software</td>
			<td class="xl65">Pixmeo SARL, Bernex, Switzerland</td>
			<td class="xl64" style="width:149pt" width="199">-</td>
			<td class="xl67">Version 4.0</td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Oxygen, medical grade</td>
			<td class="xl65">MTI, Ulm, Germany</td>
			<td class="xl64" style="width:149pt" width="199">-</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Resolon 5/0</td>
			<td class="xl65">Resorba, N&#252;rnberg, Germany</td>
			<td class="xl64" style="width:149pt" width="199">88143</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Saline 0.9%</td>
			<td class="xl65">Braun, Melsungen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">3570350</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Scalpel handle 125 mm</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BB073R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Scissors 150 mm&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BC006R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Sealer for autoclave packaging&#160;</td>
			<td class="xl65">Hawo GmbH, Obrigheim, Germany</td>
			<td class="xl64" style="width:149pt" width="199">HM500</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Sterican 27 G&#160;</td>
			<td class="xl65">Braun, Melsungen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">4657705</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Sterile surgical blades no. 11/15&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">BB511/515</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Surgical gloves&#160;</td>
			<td class="xl65">Hartmann, Heidenheim, Germany</td>
			<td class="xl64" style="width:149pt" width="199">Peha-micron 9425712</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Surgical light&#160;</td>
			<td class="xl65">Maquet SA, Ardon, France</td>
			<td class="xl64" style="width:149pt" width="199">Blue line 80</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Syringes 5 ml&#160;</td>
			<td class="xl65">Braun, Melsungen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">Injekt 4606051V</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Tissue forceps 80 mm&#160;</td>
			<td class="xl65">Aesculap, Tuttlingen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">OC091R</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Tramadol 25 mg/l</td>
			<td class="xl65">Gr&#252;nenthal, Aachen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">100mg/ml</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Vasofix Safety&#160;</td>
			<td class="xl65">Braun, Melsungen, Germany</td>
			<td class="xl64" style="width:149pt" width="199">4268113S-01</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Vicryl 5-0&#160;</td>
			<td class="xl65">Ethicon, Norderstedt, Germany</td>
			<td class="xl64" style="width:149pt" width="199">V30371</td>
			<td class="xl67"></td>
		</tr>
		<tr height="21" style="height:15.75pt">
			<td class="xl65" height="21" style="height:15.75pt">Visdisic eye ointment&#160;</td>
			<td class="xl65">Bausch &#38; Lomb, Berlin, Germany</td>
			<td class="xl64" style="width:149pt" width="199">3099559</td>
			<td class="xl67"></td>
		</tr>
	</tbody>
</table>

in vivo evaluation of fracture callus development during bone healing in mice using an mri-compatible osteosynthesis device for the mouse femur

Endochondral fracture healing is a complex process involving the development of fibrous, cartilaginous, and osseous tissue in the fracture callus. The amount of the different tissues in the callus provides important information on the fracture healing progress. Available in vivo techniques to longitudinally monitor the callus tissue development in preclinical fracture-healing studies using small animals include digital radiography and &#181;CT imaging. However, both techniques are only able to distinguish between mineralized and non-mineralized tissue. Consequently, it is impossible to discriminate cartilage from fibrous tissue. In contrast, magnetic resonance imaging (MRI) visualizes anatomical structures based on their water content and might therefore be able to noninvasively identify soft tissue and cartilage in the fracture callus. Here, we report the use of an MRI-compatible external fixator for the mouse femur to allow MRI scans during bone regeneration in mice. The experiments demonstrated that the fixator and a custom-made mounting device allow repetitive MRI scans, thus enabling longitudinal analysis of fracture-callus tissue development.

First, the success of the surgical procedure can be confirmed by analysis of the MRI scans (see example in Figure 2). All four pins should be located in the middle of the femoral shaft. The size of the osteotomy gap should be between 0.3-0.5 mm. If the size of the osteotomy gap varies greatly from these values, the mouse should be excluded from further analysis.
Secondly, the evaluation of longitudi...

Watch this Scientific Journal Video about In Vivo Evaluation of Fracture Callus Development During Bone Healing in Mice Using an MRI-compatible Osteosynthesis Device for the Mouse Femur at JoVE.com

In Vivo Evaluation of Fracture Callus Development During Bone Healing in Mice Using an MRI-compatible Osteosynthesis Device for the Mouse Femur

The evaluation of tissue development in the fracture callus during endochondral bone healing is essential to monitor the healing process. Here, we report the use of a magnetic resonance imaging (MRI)-compatible external fixator for the mouse femur to allow MRI scans during bone regeneration in mice.

In Vivo Evaluation of Fracture Callus Development During Bone Healing in Mice Using an MRI-compatible Osteosynthesis Device for the Mouse Femur

Endochondral fracture healing is a complex process involving the development of fibrous, cartilaginous, and osseous tissue in the fracture callus. The ...

The overall goal of this technique is to use magnetic resonant imaging, or MRI, for the longitudinal evaluation of murine fracture callus development during bone healing. This method can help answer key questions in the bone healing field such as how fracture calluses develop over time in different mouse models. The main advantages of this technique are that fracture callus development can be monitored in vivo in real-time and that the applied external fixator provides a highly standardized fracture healing model.Demonstrating the MRI procedure will be Anne Subgang, a technician from the Small Animal Imaging Core facility. Before beginning the procedure, confirm the appropriate level of sedation by lack of response to toe pinch and inject the mouse subcutaneously with a single dose of antibiotics in a fluid depot of two times 250 microliters of 0.9%sodium chloride. Apply ointment to the animal's eyes and place the mouse on a 37-degrees-Celsius heating pad.Remove the fur from the right hind limb and scrub the surgical area with an alcohol-based disinfectant. Place a sterile drape over the animal leaving the surgical area exposed, and cover the right hind paw with a small piece from a sterile glove. Scrub the surgical area three more times with disinfectant.Then, using an autoclave-sterilized scalpel, make an approximately one-centimeter longitudinal skin incision along the interior side of the right femur and use micro scissors and forceps to bluntly separate the biceps femoris and vastus lateralis muscles. Cut the tendon insertion at the femur trochanter to allow free access to the anterolateral part of the bone taking care that the sciatic nerve is preserved, and position the external fixator parallel to the femur. Using a 0.45-millimeter drill bit, manually drill bore holes through cortex.And starting with the most proximal and distal ends, place ceramic mounting pins into the holes. When all the pins have been placed, humidify the bone with a small volume of sterile saline and use a 0.4-millimeter Gigli wire saw to create a 0.4-millimeter osteotomy through the hole bone between the two inner pins. Flush the osteotomy gap carefully with two milliliters of sterile saline to remove any bone chips from between the two fractured cortices and use a continuous resorbable suture to adapt the muscles followed by adaptation of the skin with interrupted non-resorbable sutures.Then, clean the surgical area with disinfectant and return the mouse to its cage with monitoring until full recovery. As early as three days after the surgery, insert the external fixator carefully into a custom-made mounting device and place the anesthetized mouse onto a temperature-controlled cradle for introduction into the MRI device. Rigidly attach the external fixator to the mounting device on the four-element head coil, and use a dedicated high-field small animal MRI system operating at 11.7 tesla to apply a proton-density fat-suppressed multi-sliced turbo-spin echo sequence to the fracture.When all of the images have been acquired, open the data in the appropriate image analysis software and enter the voxel size as 0.05 by 0.05 by 0.35 millimeters cubed. To segment the different tissues in the fracture callus with semi-automatic thresholding based on their intensity, first click on the edit new label field and click Add material. Rename the material to callus, and use the lasso tool to distinguish the callus area from the surrounding tissues based on the hypointense signal from the periosteum.Click Add to material, then click Add material and rename the material to cartilage. Using the threshold tool, segment the cartilage and Select only current material from the callus. Then, click cartilage and Add to material.When all of the tissues have been segmented, click Generate surface, apply None for smoothing type and click Surface View to generate 3D reconstructions of the fractured femurs based on the tissue segmentation data. A successful surgical procedure can be confirmed by MRI scan of the fracture showing all four pins in the middle of the femoral shaft with an osteotomy gap between 0.3 and 0.5 millimeters. On day 10 postsurgery, evaluation of the longitudinal scans during the fracture healing process reveals cartilaginous tissue within the middle of the fracture callus and bony tissue at the periphery of the fracture.Both the cartilaginous and bony tissues increase until day 14. The cartilaginous tissue then decreases until day 21 at which point bony bridging can be observed. After segmentation of the different tissues in the fracture callus as just demonstrated, a whole 3D femur scan can be generated with the mature cortex appearing in gray, the ceramic pins marked in yellow, the callus soft tissue in green, the cartilage tissue in red and the callus bony tissue indicated in purple.After watching this video, you should have a good understanding of how to apply an external fixator to the mouse femur to create the femur osteotomy and to insert a mouse into an MRI device to ensure a standardized scanning position. Once mastered, the external fixator application and the osteotomy technique can be completed within 20 to 30 minutes if performed properly.

in-vivo-evaluation-fracture-callus-development-during-bone-healing

Research

JoVE Journal

Medicine

Longitudinal Evaluation of Mouse Hind Limb Bone Loss After Spinal Cord Injury using Novel, in vivo, Methodology

Spinal cord injury (SCI) is often accompanied by osteoporosis in the sublesional regions of the pelvis and lower extremities, leading to a higher frequency of fractures 1. As these fractures often occur in regions that have lost normal sensory function, the patient is at a greater risk of fracture-dependent pathologies, including death. SCI-dependent loss in both bone mineral density (BMD, grams/cm2) and bone mineral content (BMC, grams) has been attributed to mechanical disuse 2, aberrant neuronal signaling 3 and hormonal changes 4. The use of rodent models of SCI-induced osteoporosis can provide invaluable information regarding the mechanisms underlying the development of osteoporosis following SCI as well as a test environment for the generation of new therapies 5-7 (and reviewed in 8). Mouse models of SCI are of great interest as they permit a reductionist approach to mechanism-based assessment through the use of null and transgenic mice. While such models have provided important data, there is still a need for minimally-invasive, reliable, reproducible, and quantifiable methods in determining the extent of bone loss following SCI, particularly over time and within the same cohort of experimental animals, to improve diagnosis, treatment methods, and/or prevention of SCI-induced osteoporosis.
An ideal method for measuring bone density in rodents would allow multiple, sequential (over time) exposures to low-levels of X-ray radiation. This study describes the use of a new whole-animal scanner, the IVIS Lumina XR (Caliper Instruments) that can be used to provide low-energy (1-3 milligray (mGy)) high-resolution, high-magnification X-ray images of mouse hind limb bones over time following SCI. Significant bone density loss was seen in the tibiae of mice by 10 days post-spinal transection when compared to uninjured, age-matched control (na&iuml;ve) mice (13% decrease, p&lt;0.0005). Loss of bone density in the distal femur was also detectable by day 10 post-SCI, while a loss of density in the proximal femur was not detectable until 40 days post injury (7% decrease, p&lt;0.05). SCI-dependent loss of mouse femur density was confirmed post-mortem through the use of Dual-energy X-ray Absorptiometry (DXA), the current "gold standard" for bone density measurements. We detect a 12% loss of BMC in the femurs of mice at 40 days post-SCI using the IVIS Lumina XR. This compares favorably with a previously reported BMC loss of 13.5% by Picard and colleagues who used DXA analysis on mouse femurs post-mortem 30 days post-SCI 9. Our results suggest that the IVIS Lumina XR provides a novel, high-resolution/high-magnification method for performing long-term, longitudinal measurements of hind limb bone density in the mouse following SCI.

Longitudinal Evaluation of Mouse Hind Limb Bone Loss After Spinal Cord Injury using Novel, in vivo, Methodology

A longitudinal examination of bone loss in the femurs and tibiae of adult mice was performed following spinal cord injury using sequential low-dose X-ray scans. Tibia bone loss was detected throughout the study, while bone loss in the femur was not detected until 40 days post injury.

Spinal cord injury (SCI) is often accompanied by osteoporosis in the sublesional regions of the pelvis and lower extremities, leading to a higher ...

Using Quantitative Real-time PCR to Determine Donor Cell Engraftment in a Competitive Murine Bone Marrow Transplantation Model

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules that regulate HSCs. In these transplant model systems, the function of HSCs is determined by the ability of these cells to engraft and reconstitute lethally irradiated recipient mice. Commonly, the donor cell contribution/engraftment is measured by antibodies to donor- specific cell surface proteins using flow cytometry. However, this method heavily depends on the specificity and the ability of the cell surface marker to differentiate donor-derived cells from recipient-originated cells, which may not be available for all mouse strains. Considering the various backgrounds of genetically modified mouse strains in the market, this cell surface/ flow cytometry-based method has significant limitations especially in mouse strains that lack well-defined surface markers to separate donor cells from congenic recipient cells. Here, we reported a PCR-based technique to determine donor cell engraftment/contribution in transplant recipient mice. We transplanted male donor bone marrow HSCs to lethally irradiated congenic female mice. Peripheral blood samples were collected at different time points post transplantation. Bone marrow samples were obtained at the end of the experiments. Genomic DNA was isolated and the Y chromosome specific gene, Zfy1, was amplified using quantitative Real time PCR. The engraftment of male donor-derived cells in the female recipient mice was calculated against standard curve with known percentage of male vs. female DNAs. Bcl2 was used as a reference gene to normalize the total DNA amount. Our data suggested that this approach reliably determines donor cell engraftment and provides a useful, yet simple method in measuring hematopoietic cell reconstitution in murine bone marrow transplantation models. Our method can be routinely performed in most laboratories because no costly equipment such as flow cytometry is required.

Determining donor cell engraftment presents a challenge in mouse bone marrow transplant models that lack well-defined phenotypical markers. We described a methodology to quantify male donor cell engraftment in female transplant recipient mice. This method can be used in all mouse strains for the study of HSC functions.

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules ...

Development of an Audio-based Virtual Gaming Environment to Assist with Navigation Skills in the Blind

Audio-based Environment Simulator (AbES) is virtual environment software designed to improve real world navigation skills in the blind. Using only audio based cues and set within the context of a video game metaphor, users gather relevant spatial information regarding a building's layout. This allows the user to develop an accurate spatial cognitive map of a large-scale three-dimensional space that can be manipulated for the purposes of a real indoor navigation task. After game play, participants are then assessed on their ability to navigate within the target physical building represented in the game. Preliminary results suggest that early blind users were able to acquire relevant information regarding the spatial layout of a previously unfamiliar building as indexed by their performance on a series of navigation tasks. These tasks included path finding through the virtual and physical building, as well as a series of drop off tasks. We find that the immersive and highly interactive nature of the AbES software appears to greatly engage the blind user to actively explore the virtual environment. Applications of this approach may extend to larger populations of visually impaired individuals.

Audio-based Environment Simulator (AbES) is virtual environment software designed to improve real world navigation skills in the blind.

Audio-based Environment Simulator (AbES) is virtual environment software designed to improve real world navigation skills in the blind. Using only audio ...

Sequential In vivo Imaging of Osteogenic Stem/Progenitor Cells During Fracture Repair

Bone turns over continuously and is highly regenerative following injury. Osteogenic stem/progenitor cells have long been hypothesized to exist, but in vivo demonstration of such cells has only recently been attained. Here, in vivo imaging techniques to investigate the role of endogenous osteogenic stem/progenitor cells (OSPCs) and their progeny in bone repair are provided. Using osteo-lineage cell tracing models and intravital imaging of induced microfractures in calvarial bone, OSPCs can be directly observed during the first few days after injury, in which critical events in the early repair process occur. Injury sites can be sequentially imaged revealing that OSPCs relocate to the injury, increase in number and differentiate into bone forming osteoblasts. These methods offer a means of investigating the role of stem cell-intrinsic and extrinsic molecular regulators for bone regeneration and repair.

Sequential In vivo Imaging of Osteogenic Stem/Progenitor Cells During Fracture Repair

Quantitative measurement of bone progenitor function in fracture healing requires high resolution serial imaging technology. Here, protocols are provided for using intravital microscopy and osteo-lineage tracking to sequentially image and quantify the migration, proliferation and differentiation of endogenous osteogenic stem/progenitor cells in the process of repairing bone fracture.

Bone turns over continuously and is highly regenerative following injury. Osteogenic stem/progenitor cells have long been hypothesized to exist, but in ...

An Improved Method for Rapid Intubation of the Trachea in Mice

Despite some anatomical and physiological differences, mouse models continue to be an essential tool for studying human lung disease. Bleomycin toxicity is a commonly used model to study both acute lung injury and fibrosis, and multiple methods have been developed for administering bleomycin (and other toxic agents) into the lungs. However, many of these approaches, such as transtracheal instillation, have inherent drawbacks, including the need for strong anesthetics and survival surgery. This paper reports a quick, reproducible method of intratracheal intubation that involves mild inhaled anesthesia, visualization of the trachea, and the use of a surrogate spirometer to confirm exposure. As a proof of concept, 8-12 week old C57BL/6 mice were administered either 2.0 U/kg of bleomycin or an equivalent volume of PBS, and both damage and fibrotic endpoints were measured post-exposure. This procedure allows researchers to treat a large cohort of mice in a relatively short period with little expense and minimal post-procedure care.

This article presents a rapid and simple method for administering bleomycin directly into the mouse trachea via intubation. Key advantages of this method are that it is highly reproducible, easy to master, and does not require specialized equipment or lengthy recovery times.

Despite some anatomical and physiological differences, mouse models continue to be an essential tool for studying human lung disease. Bleomycin toxicity ...

Depletion of Mouse Cells from Human Tumor Xenografts Significantly Improves Downstream Analysis of Target Cells

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic patient tumors in different assays1. Human tumor xenografts represent the gold standard with respect to tumor biology, drug discovery, and metastasis research 2-4. Tumor xenografts can be derived from different types of material like tumor cell lines, tumor tissue from primary patient tumors4 or serially transplanted tumors. When propagated in vivo, xenografted tissue is infiltrated and vascularized by cells of mouse origin. Multiple factors such as the tumor entity, the origin of xenografted material, growth rate and region of transplantation influence the composition and the amount of mouse cells present in tumor xenografts. However, even when these factors are kept constant, the degree of mouse cell contamination is highly variable.
Contaminating mouse cells significantly impair downstream analyses of human tumor xenografts. As mouse fibroblasts show high plating efficacies and proliferation rates, they tend to overgrow cultures of human tumor cells, especially slowly proliferating subpopulations. Mouse cell derived DNA, mRNA, and protein components can bias downstream gene expression analysis, next-generation sequencing, as well as proteome analysis 5. To overcome these limitations, we have developed a fast and easy method to isolate untouched human tumor cells from xenografted tumor tissue. This procedure is based on the comprehensive depletion of cells of mouse origin by combining automated tissue dissociation with the benchtop tissue dissociator and magnetic cell sorting. Here, we demonstrate that human target cells can be can be obtained with purities higher than 96% within less than 20 min independent of the tumor type.

Human tumor xenografts are vascularized and infiltrated by cells of mouse origin during the growth phase in vivo. To circumvent the bias caused by these contaminating cells during downstream analysis, we developed a method allowing for comprehensive depletion of all mouse cells by magnetic cell sorting.

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic ...

An Intramedullary Locking Nail for Standardized Fixation of Femur Osteotomies to Analyze Normal and Defective Bone Healing in Mice

Bone healing models are essential to the development of new therapeutic strategies for clinical fracture treatment. Furthermore, mouse models are becoming more commonly used in trauma research. They offer a large number of mutant strains and antibodies for the analysis of the molecular mechanisms behind the highly differentiated process of bone healing. To control the biomechanical environment, standardized and well-characterized osteosynthesis techniques are mandatory in mice. Here, we report on the design and use of an intramedullary nail to stabilize open femur osteotomies in mice. The nail, made of medical-grade stainless steel, provides high axial and rotational stiffness. The implant further allows the creation of defined, constant osteotomy gap sizes from 0.00 mm to 2.00 mm. Intramedullary locking nail stabilization of femur osteotomies with gap sizes of 0.00 mm and 0.25 mm result in adequate bone healing through endochondral and intramembranous ossification. Stabilization of femur osteotomies with a gap size of 2.00 mm results in atrophic non-union. Thus, the intramedullary locking nail can be used in healing and non-healing models. A further advantage of the use of the nail compared to other open bone healing models is the possibility to adequately fix bone substitutes and scaffolds in order to study the process of osseous integration. A disadvantage of the use of the intramedullary nail is the more invasive surgical procedure, inherent to all open procedures compared to closed models. A further disadvantage may be the induction of some damage to the intramedullary cavity, inherent to all intramedullary stabilization techniques compared to extramedullary stabilization procedures.

This protocol describes an osteosynthesis technique using an intramedullary locking nail for standardized fixation of femur osteotomies, which can be used to analyze normal and defective bone healing in mice.

Bone healing models are essential to the development of new therapeutic strategies for clinical fracture treatment. Furthermore, mouse models are becoming ...

Development of a Direct Pulp-capping Model for the Evaluation of Pulpal Wound Healing and Reparative Dentin Formation in Mice

Dental pulp is a vital organ of a tooth fully protected by enamel and dentin. When the pulp is exposed due to cariogenic or iatrogenic injuries, it is often capped with biocompatible materials in order to expedite pulpal wound healing. The ultimate goal is to regenerate reparative dentin, a physical barrier that functions as a "biological seal" and protects the underlying pulp tissue. Although this direct pulp-capping procedure has long been used in dentistry, the underlying molecular mechanism of pulpal wound healing and reparative dentin formation is still poorly understood. To induce reparative dentin, pulp capping has been performed experimentally in large animals, but less so in mice, presumably due to their small sizes and the ensuing technical difficulties. Here, we present a detailed, step-by-step method of performing a pulp-capping procedure in mice, including the preparation of a Class-I-like cavity, the placement of pulp-capping materials, and the restoration procedure using dental composite. Our pulp-capping mouse model will be instrumental in investigating the fundamental molecular mechanisms of pulpal wound healing in the context of reparative dentin in vivo by enabling the use of transgenic or knockout mice that are widely available in the research community.

We describe a step-by-step method of performing direct pulp capping on mice teeth for the evaluation of pulpal wound healing and reparative dentin formation in vivo.

Dental pulp is a vital organ of a tooth fully protected by enamel and dentin. When the pulp is exposed due to cariogenic or iatrogenic injuries, it is ...

An Anesthesia, Surgery, and Harvest Method for the Evaluation of Transpedicular Screws Using an In Vivo Porcine Lumbar Spine Model

Pedicle screw fixation is the gold standard for the treatment of spinal diseases. However, many studies have reported the issue of loosening pedicle screws after spinal surgery, which is a serious concern. To address this problem, diverse types of pedicle screws have been examined to identify those with good fixation strength and osseointegration in spine bone. The porcine spine is a good alternative for the human spine in the evaluation of pedicle screws due to the anatomical size, mechanical characteristics, and cost. Although several studies have reported that pedicle screws are efficient in the porcine model, no study has described detailed protocols for the evaluation of a pedicle screw using the porcine model. Here, we describe a detailed method for evaluating transpedicular screws using an in vivo porcine lumbar spine model. The technical details for anesthesia, spine surgery, and harvest provided here will facilitate with the evaluation of the transpedicular screw fixation model.

An Anesthesia, Surgery, and Harvest Method for the Evaluation of Transpedicular Screws Using an In Vivo Porcine Lumbar Spine Model

Here, we introduce a method to evaluate transpedicular screws by using an in vivo porcine lumbar spine model.

Pedicle screw fixation is the gold standard for the treatment of spinal diseases. However, many studies have reported the issue of loosening pedicle ...

A Minimally Invasive Model to Analyze Endochondral Fracture Healing in Mice Under Standardized Biomechanical Conditions

Bone healing models are necessary to analyze the complex mechanisms of fracture healing to improve clinical fracture treatment. During the last decade, an increased use of mouse models in orthopedic research was noted, most probably because mouse models offer a large number of genetically-modified strains and special antibodies for the analysis of molecular mechanisms of fracture healing. To control the biomechanical conditions, well-characterized osteosynthesis techniques are mandatory, also in mice. Here, we report on the design and use of a closed bone healing model to stabilize femur fractures in mice. The intramedullary screw, made of medical-grade stainless steel, provides through fracture compression an axial and rotational stability compared to the mostly used simple intramedullary pins, which show a complete lack of axial and rotational stability. The stability achieved by the intramedullary screw allows the analysis of endochondral healing. A large amount of callus tissue, received after stabilization with the screw, offers ideal conditions to harvest tissue for biochemical and molecular analyses. A further advantage of the use of the screw is the fact that the screw can be inserted into the femur with a minimally invasive technique without inducing damage to the soft tissue. In conclusion, the screw is a unique implant that can ideally be used in closed fracture healing models offering standardized biomechanical conditions.

This protocol describes a minimally invasive osteosynthesis technique using an intramedullary screw for standardized stabilization of femur fractures, which can be used to analyze endochondral bone healing in mice.

Bone healing models are necessary to analyze the complex mechanisms of fracture healing to improve clinical fracture treatment. During the last decade, an ...