The Generation of Closed Femoral Fractures in Mice: A Model to Study Bone Healing

Summary

The murine closed femoral fracture model is a powerful platform to study fracture healing and novel therapeutic strategies to accelerate bone regeneration. The goal of this surgical protocol is to generate unilateral closed femoral fractures in mice using an intramedullary steel rod to stabilize the femur.

Abstract

Bone fractures impose a tremendous socio-economic burden on patients, in addition to significantly affecting their quality of life. Therapeutic strategies that promote efficient bone healing are non-existent and in high demand. Effective and reproducible animal models of fractures healing are needed to understand the complex biological processes associated with bone regeneration. Many animal models of fracture healing have been generated over the years; however, murine fracture models have recently emerged as powerful tools to study bone healing. A variety of open and closed models have been developed, but the closed femoral fracture model stands out as a simple method for generating rapid and reproducible results in a physiologically relevant manner. The goal of this surgical protocol is to generate unilateral closed femoral fractures in mice and facilitate a post-fracture stabilization of the femur by inserting an intramedullary steel rod. Although devices such as a nail or a screw offer greater axial and rotational stability, the use of an intramedullary rod provides a sufficient stabilization for consistent healing outcomes without producing new defects in the bone tissue or damaging nearby soft tissue. Radiographic imaging is used to monitor the progression of callus formation, bony union, and subsequent remodeling of the bony callus. Bone healing outcomes are typically associated with the strength of the healed bone and measured with torsional testing. Still, understanding the early cellular and molecular events associated with fracture repair is critical in the study of bone tissue regeneration. The closed femoral fracture model in mice with intramedullary fixation serves as an attractive platform to study bone fracture healing and evaluate therapeutic strategies to accelerate healing.

Introduction

Fractures are among the most common injuries occurring to the musculoskeletal system and are associated with a tremendous socioeconomic burden, including treatment costs that are projected to surpass $25 billion annually in the United States^1,2. Although the majority of fractures heal without incident, healing is associated with substantial downtime and loss of productivity. Approximately 5 - 10% of all fractures result in a delayed healing or non-union, due to age or other underlying chronic health conditions, such as osteoporosis and diabetes mellitus^3,

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Protocol

The following procedure was performed with approval from the Indiana University School of Medicine Institutional Animal Care and Use Committee (IACUC). All survival surgeries were performed under sterile conditions as outlined by the NIH guidelines. Pain and risk of infections were managed with proper analgesics and antibiotics to ensure a successful outcome.

1. Anesthesia and Preparation

1. Weigh the mouse and anesthetize it with a mixture of ketamine (100 mg/kg) and xylazine (10 mg/kg) administered via the intraperitoneal (I.P.) route. Place the mouse in an empty cage and monitor it until it is fully sedated.
2. E....

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Results

The successful implementation of the surgical procedure was monitored with radiographic imaging. Key steps include the insertion of an intramedullary needle, the placement of a guide wire, the induction of a transverse fracture at the femoral midshaft, and the proper stabilization with an intramedullary rod (Figure 2Ai - 2Aiv). The healing progression of the fracture callus was monitored with weekly radiographic images up to .......

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Discussion

The goal of this surgical procedure is to generate standardized closed femoral fractures in mice. A key advantage of this model is that the internal fixation takes place after the generation of the fracture, thereby avoiding an angulation of the intramedullary rod. Perhaps the most critical aspect of this protocol is the generation of a standardized transverse fracture at the femoral midshaft, as the fracture geometry is dependent on the applied bending force and the positioning of the hind limb. Improper positioning of .......

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Materials

Name	Company	Catalog Number	Comments
Oster Minimax Trimmer	Animal World Network	78049-100
POVIDONE-IODINE	Thermo Fisher Scientific	395516
OPHTHALMIC OINTMENT	Thermo Fisher Scientific	NC0490117
Styker T/Pump Warm Water Recirculator	Kent Scientific Corporation	TP-700
1ml Sub-Q Syringe	Thermo Fisher Scientific	309597
ENCORE Sensi-Touch PF	Moore Medical LLC	30347	Latex, powder-free surgical glove
PrecisionGlide 25G Hypodermic Needles	Thermo Fisher Scientific	14-826-49
Ultra-High-Temperature Tungsten Wire,	McMaster-Carr	3775K37	0.005" Diameter, 1/16 lb. Spool, 380' Long
304 stainless steel, 24G thin walled tubing	Microgroup Inc	304h24tw-5ft
#15 Scalpel Blades	Fine Science Tools	10015-00
#10 Scalpel Blades	Fine Science Tools	10010-00
Narrow Pattern Forceps	Fine Science Tools	11002-12	Serrated/Straight/12cm
Iris Forceps	Fine Science Tools	11066-07	1x2 Teeth/Straight/7cm
Dissector Scissors	Fine Science Tools	14081-09	Slim Blades/Angled to Side/Sharp-Sharp/10cm
Fine Scissors	Fine Science Tools	14058-11	ToughCut/Straight/Sharp-Sharp/11.5cm
Olsen-Hegar Needle Holder with Suture Cutter	Fine Science Tools	12002-12	Straight/Serrated/12cm/with Lock
Crile Hemostat	Fine Science Tools	13004-14	Serrated/Straight/14cm
Tungsten Wire Cutter	ACE Surgical Supply Co., Inc.	08-051-90	ACE #150 Wire Cutter, tungsten carbide tips
3-0 VICRYL Suture	Ethicon Suture	J423H	3-0 VICRYL UNDYED 27" FS-2 CUTTING
piXarray 100 Digital Specimen Radiography System	Bioptics, Inc		Cabinet x-ray system
Einhorn 3-Point Bending Device	N/A	N/A	Custom Built