Este trabajo fue financiado por R01-AR053645 del NIAMS.

Todos los procedimientos fueron aprobados por el Cuidado de Animales de UCSF Institucional y el empleo Comisión y se ajustan a las directrices nacionales. 1. Preparación de los fijadores antes de la cirugía <ol><li> Antes de crear una fractura estabilizada, es necesario montar el dispositivo de diseño personalizado de la fijación externa. Los marcos de fijación especialmente diseñados externos consta de dos anillos de aluminio estabilizados por tres de acero inoxidable # 0/56 varillas roscadas, 8 # 2/56 tuercas hexagonales y 17 tornillos a juego (Pequeña Parte Inc., de Miami Lakes, FL 3). </li><li> Dos anillos de aluminio se necesitan por fractura. Para montar un anillo de 4 º 2/56 tuercas hexagonales (pequeña pieza Inc., de Miami Lakes, FL) y los pernos que coinciden en los agujeros correspondientes, éstos deben quedar sueltos. Este será el anillo proximal. Para montar el anillo distal 3 tuercas y tornillos, una vez más no apriete por completo. </li></ol> 2. Anestesia, fractura la Creación, y la aplicación del fijador <ol><li> Los ratones (edad, sexo, y el cuerpopeso se determinó por el usuario) se anestesiaron por inyección intraperitoneal de 50mg/ml ketamina, 0,5 mg / ml de medetomidina (0,03 ml / ratón). </li><li> Aplicar lubricante ocular para los ojos de los ratones y colocar un pedazo de cinta adhesiva en su cabeza para proteger sus ojos durante el procedimiento. </li><li> El animal se coloca sobre una almohadilla de calentamiento durante el resto del procedimiento. </li><li> Transfix las metáfisis proximal y distal de la tibia izquierda por medio de cuatro pernos 0,25 mm de los insectos estériles (Anticorro: Tool Science Fine, Foster City, CA), uniendo los pines a una herramienta de Dremel y la perforación a través del hueso. </li><li> Pasadores están orientadas perpendicularmente al eje longitudinal de la tibia, y 90 ° entre sí. Al menos 10 mm entre los pasadores proximal y distal. </li><li> Coloque el primer anillo por encima de los pernos proximales con las tuercas hacia adentro, hacia las patillas. Asegure los pasadores para el anillo de mantenimiento de la tibia posición central dentro del anillo de aluminio. </li><li> Coloque el segundo anillo por debajo de la dipines Stal y asegurar los pasadores para el anillo con las tuercas hexagonales, como antes. Añadir la cuarta tuerca de perno en combinación para la parte distal del fijador. </li><li> Crear la fractura transversal por tres puntos de flexión con un dispositivo adecuado. Utilizamos el sistema de gravedad de Bonnarens Einhorn y 4. Coloque la superficie posterolateral de la tibia a través de los bloques, aumentar el peso a una altura predeterminada a lo largo del brazo de la gota y dejar caer el peso de modo que entra en contacto con la cara anterolateral de la tibia para producir la fractura contundente. La altura del peso se deja caer desde necesita ser determinado para cada cepa de ratón para asegurar una fractura solo exclusivamente de la tibia. </li><li> Fractura se confirma por rayos-X. </li><li> Después de confirmar la fractura, tres de acero inoxidable # 0/56 varillas roscadas están unidos a los anillos. Comenzando desde el extremo distal insertar la varilla y enhebrar una tuerca en el anillo distal para bloquear la barra. A continuación, dicho de otra tuerca en la varilla, el anillo proximal se sentará en la parte superior de esta nut. Asegúrese de que los dos anillos se llevará a cabo a la misma distancia y no forzar las clavijas. Fijar el anillo proximal a las barras con una tuerca tercero. </li><li> Las inyecciones subcutáneas de buprenorfina (1,0 mg / kg) para la analgesia se administró inmediatamente después de la cirugía y los animales se vigilan para el dolor y la buprenorfina dado, según sea necesario. </li><li> La anestesia se revierte mediante la administración de atipamezol, y los animales son monitoreados hasta que son sensibles y deambular. </li></ol> 3. Osteogénesis por distracción (ver también: 12,13) Para modificar este procedimiento para dar cabida a distracción osteogénica es sencillo. Los anillos se mantienen en posición por varillas roscadas, y girando las tuercas que sujetan las varillas en su lugar los anillos se pueden mover de separación. <ol><li> Latencia: Para la distracción osteogénica estándar de uso de este modelo de los fijadores se colocan en la tibia y se fue solo por un período de latencia de 5 días. </li><li> Distracción: osteog distracciónENESIS puede llevarse a cabo girando las tuercas de las barras estabilizadoras de giro de ¼ (0,25 mm) cada día 7 días. </li><li> Maduración: Bony puente se observa en 10 días y la consolidación completa se observa por 27 días después de la finalización de distracción. </li></ol> 4. Creación de un defecto crítico Empresas Para crear un defecto de tamaño crítico los fijadores externos se aplican como se ha descrito anteriormente con las siguientes modificaciones. <ol><li> Antes de comenzar el procedimiento de la piel se limpia con un 70% de etanol. </li><li> Los fijadores se aplican completamente antes de crear el defecto. </li><li> El ratón se coloque bajo el microscopio de disección para visualizar la tibia medial y una incisión (5-7 mm) se hace con una cuchilla # 10 en un bisturí. </li><li> El músculo se secciona suavemente y separados para exponer la mitad de diáfisis de la tibia. </li><li> Un segmento de 3 mm del hueso se retira entonces de la tijera mediados de diáfisis utilizando. Uso de tque señalar de las tijeras, el hueso se cortó lentamente lejos y fragmentos de hueso se eliminan con pinzas si es necesario. Se debe tener cuidado con las puntas de las tijeras para evitar el desplazamiento de la tibia con una gran fuerza mecánica. </li><li> Examinar el sitio quirúrgico para asegurar que no existen fragmentos de hueso, ya que podrían promover la curación. </li><li> Vuelva a colocar el músculo alrededor de los extremos romos de la tibia. </li><li> Cierre de la incisión en la piel con 2-3 suturas de monofilamento de poliaminas 6-0. </li><li> La anestesia se invierte y los animales control se elegirán para el dolor y dado analgésicos como se describió previamente. </li></ol> 5. Los resultados representativos Cuando se aplica correctamente, los fijadores externos proporcionan una mayor estabilidad rígida de la fractura cerrada de tibia con una excelente reducción (Figuras 1, 2). Sin embargo, en algunos casos la reducción inadecuada (una diferencia obvia y grande entre los extremos de los huesos o fracturas múltiples se producen (Fig. 3 </strong&gt;), Y estos ratones se excluyen de los análisis. Las fracturas estabilizado utilizando este método curar principalmente a través de osificación intramembranosa (fig. 4). En contraste, si la fractura no se estabiliza un callo grande cartílago está formado en el hueco de la fractura (fig. 5), y este es sustituido por hueso a través del proceso de osificación intramembranosa. <img alt="Figura 1" src="/files/ftp_upload/3552/3552fig1.jpg" /> Figura 1. Radiografías que ilustran un dispositivo de fijación externo que se utiliza para estabilizar una fractura tibial. Radiografía tomada después de la fractura que muestra algunos de los segmentos óseos bien alineados (punta de flecha). <img alt="Figura 2" src="/files/ftp_upload/3552/3552fig2.jpg" /> Figura 2. Imagen de un ratón después el fijador se ha aplicado. <img alt="Figura 3" src="/files/ftp_upload/3552/3552fig3.jpg" /> Figura 3. Radiografía tomada después de la fracture muestra los segmentos de hueso desalineado y fragmentado (punta de flecha). <img alt="Figura 4" src="/files/ftp_upload/3552/3552fig4.jpg" /> Figura 4. Fractura estabilizada a través de cura osificación intramembranosa. Tinción tricrómica de fractura estabilizada muestra un poco de hueso nuevo (b) en el sitio de fractura. Barras de escala = 500 micras. <img alt="Figura 5" src="/files/ftp_upload/3552/3552fig5.jpg" /> Figura 5. No estabilizado fractura se cura a través de la osificación endocondral. Tinción tricrómica de fractura no estabilizada muestra cartílago (c) y hueso (b) en lugar de la fractura. Barras de escala = 500 micras.

<ol>
	<li>Ilizarov, G. A., Lediaev, V. I., Shitin, V. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+course+of+compact+bone+reparative+regeneration+in+distraction+osteosynthesis+under+different+conditions+of+bone+fragment+fixation+(experimental+study).">The course of compact bone reparative regeneration in distraction osteosynthesis under different conditions of bone fragment fixation (experimental study).</a> Eksperimentalnaia Khirurgiia i Anesteziologiia. 14, 3-12 (1969).</li><li>Ilizarov, G. A., Deviatov, A. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surgical+elongation+of+the+leg.">Surgical elongation of the leg.</a> Ortopediia Travmatologiia i Protezirovanie. 32, (1971).</li><li>Thompson, Z., Miclau, T., Hu, D., Helms, J. 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A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Histochemical+and+molecular+analyses+of+distraction+osteogenesis+in+a+mouse+model.">Histochemical and molecular analyses of distraction osteogenesis in a mouse model.</a> Journal of Orthopaedic Research. 16, 636-642 (1998).</li><li>Choi, P., Ogilvie, C., Thompson, Z., Miclau, T., Helms, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cellular+and+molecular+characterization+of+a+murine+non-union+model.">Cellular and molecular characterization of a murine non-union model.</a> J. Orthop. Res. 22, 1100-1107 (2004).</li><li>Buckwalter, J. A., Marsh, E. T., J, L., Heckman, J. D., Bucholz, R. 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No tenemos nada que revelar.

Los huesos se curan en dos modalidades diferentes en función de la estabilidad mecánica (revisado en 14). Cuando se deja que se forme una gran inestabilidad de la plantilla del cartílago en la línea de fractura que es reemplazado por hueso para cubrir los dos extremos del hueso roto. Proximal y distal a la rotura, el hueso se forma directamente por la osificación intramembranosa en el periostio y endostio. En contraste, en la curación de fracturas estable se produce exclusivamente a través de osificaci...

<table border="1"><tbody><tr><td> Nombre del reactivo </td><td> Empresa </td><td> Número de catálogo </td><td> Comentarios </td></tr><tr><td> Pasador de insectos 0,25 mm </td><td> Herramienta de Ciencias de Bellas </td><td> 26000-25 </td><td> Acero oscuras anodizado, 0,25 mm diámetro de la varilla, la longitud de 4 cm </td></tr><tr><td> De acero inoxidable Tuerca hexagonal </td><td> Piezas pequeñas </td><td> # 2-56 </td><td> 1/8 &quot;de longitud, 56 hilos por pulgada </td></tr><tr><td> De acero inoxidable Tuerca hexagonal </td><td> Piezas pequeñas </td><td> # 0-80 </td><td> 1/8 &quot;de longitud, 80 hilos por pulgada </td></tr><tr><td> Tornillo de acero inoxidable de la máquina </td><td> Piezas pequeñas </td><td> # 0-80 </td><td> 1/8 &quot;de longitud, 80 hilos por pulgada </td></tr><tr><td> Máquina de acero inoxidable Corte de varillas roscadas </td><td> Piezas pequeñas </td><td> # 0-80 </td><td> 6 &quot;de largo, 80 hilos por pulgada </td></tr><tr><td> 18-8 de acero inoxidable Machine Head Screw </td><td> McMaster-Carr </td><td></td><td> Temas 2-56, 3/6 &quot;de largo </td></tr><tr><td> Dispositivo de fijación externa </td><td> Taller de máquinas </td><td> Diseño personalizado </td><td></td></tr></tbody></table>

creating rigidly stabilized fractures for assessing intramembranous ossification, distraction osteogenesis, or healing of critical sized defects

La evaluación de los modos de reparación ósea es esencial para el desarrollo de terapias que se utilizan clínicamente para el tratamiento de las fracturas. La estabilidad mecánica juega un papel importante en la curación de las lesiones óseas. En el peor de los casos la inestabilidad mecánica puede llevar a los seres humanos con retraso o la no unión en el. Sin embargo, el movimiento también puede estimular el proceso de curación. En las fracturas que tienen formas de movimiento del cartílago para estabilizar los extremos de los huesos de fracturas, y este cartílago es reemplazado gradualmente por hueso a través de la recapitulación del proceso de desarrollo de la osificación endocondral. En contraste, si una fractura ósea está rígidamente estabilizado hueso se forma directamente a través de osificación intramembranosa. Clínicamente, tanto en la osificación endocondral y intramembranosa ocurren simultáneamente. Para replicar eficazmente este proceso los investigadores insertar una clavija en el canal medular del hueso fracturado como se describe por Bonnarens 4. Este método experimental proporciona una estabilidad lateral excelente permitiendo al mismo tiempo rotational de persistir la inestabilidad. Sin embargo, nuestra comprensión de los mecanismos que regulan estos dos procesos distintos también se puede mejorar de forma experimental aislar cada uno de estos procesos. Hemos desarrollado un protocolo de estabilización que proporciona la estabilización de rotación y lateral. En este modelo, la osificación intramembranosa es el único modo de sanación que se observa y los parámetros de la curación se puede comparar entre diferentes cepas de ratones genéticamente modificados 5-7, después de la aplicación de moléculas bioactivas 8,9, después de la alteración de los parámetros fisiológicos de la cicatrización 10, después de modificar la cantidad o el tiempo de estabilización 11, después de distracción osteogénica 12, después de la creación de una falta de unión 13, o después de la creación de un defecto de tamaño crítico. Aquí, se ilustra la forma de aplicar el fijador de Ilizarov modificados para el estudio de la curación de fracturas de tibia y distracción ósea en ratones.

Watch this Scientific Journal Video about Creating Rigidly Stabilized Fractures for Assessing Intramembranous Ossification, Distraction Osteogenesis, or Healing of Critical Sized Defects at JoVE.com

Creating Rigidly Stabilized Fractures for Assessing Intramembranous Ossification, Distraction Osteogenesis, or Healing of Critical Sized Defects

Este artículo describe un método para estabilizar fracturas de huesos largos que se basa en la aplicación de modificados Ilizarov fijadores externos<sup&gt; 1-3</sup&gt;. Después de la aplicación de los fijadores y la creación de la lesión ósea, la curación puede ser evaluada, la distracción osteogénica se puede realizar, o un defecto de tamaño no sindicalizados o crítica puede ser creado y utilizado para estudiar las intervenciones terapéuticas.

Creación de fracturas rígidamente estabilizados para la evaluación de osificación intramembranosa, la distracción osteogénica, o la curación de los defectos críticos de tamaño

Assessing modes of skeletal repair is essential for developing therapies to be used clinically to treat fractures. Mechanical stability plays a large role ...

creating-rigidly-stabilized-fractures-for-assessing-intramembranous

Research

JoVE Journal

Medicine

17.2K vistas.  University of California, San Francisco. Este artículo describe un método para estabilizar fracturas de huesos largos que se basa en la aplicación de modificados Ilizarov fijadores externos 1-3. Después de la aplicación de los fijadores y la creación de la lesión ósea, la curación puede ser evaluada, la distracción osteogénica se puede realizar, o un defecto de tamaño no sindicalizados o crítica puede ser creado y utilizado para estudiar las intervenciones terapéuticas.

Video: Creating Rigidly Stabilized Fractures for Assessing Intramembranous Ossification, Distraction Osteogenesis, or Healing of Critical Sized Defects

Repair of a Critical-sized Calvarial Defect Model Using Adipose-derived Stromal Cells Harvested from Lipoaspirate

Craniofacial skeletal repair and regeneration offers the promise of de novo tissue formation through a cell-based approach utilizing stem cells. Adipose-derived stromal cells (ASCs) have proven to be an abundant source of multipotent stem cells capable of undergoing osteogenic, chondrogenic, adipogenic, and myogenic differentiation. Many studies have explored the osteogenic potential of these cells in vivo with the use of various scaffolding biomaterials for cellular delivery. It has been demonstrated that by utilizing an osteoconductive, hydroxyapatite-coated poly(lactic-co-glycolic acid) (HA-PLGA) scaffold seeded with ASCs, a critical-sized calvarial defect, a defect that is defined by its inability to undergo spontaneous healing over the lifetime of the animal, can be effectively show robust osseous regeneration. This in vivo model demonstrates the basis of translational approaches aimed to regenerate the bone tissue - the cellular component and biological matrix. This method serves as a model for the ultimate clinical application of a progenitor cell towards the repair of a specific tissue defect.

This protocol describes the isolation of adipose-derived stromal cells from lipoaspirate and the creation of a 4 mm critical-sized calvarial defect to evaluate skeletal regeneration.

La reparación de un modelo de defecto crítico del tamaño de calota Uso adiposo derivados de las células del estroma cosechadas desde lipoaspirado

Craniofacial skeletal repair and regeneration offers the promise of de novo tissue formation through a cell-based approach utilizing stem cells. ...

Investigación

Medicina

Murine Model of Wound Healing

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. Impaired wound healing, which occurs in diabetic patients for example, can lead to severe unfavorable outcomes such as amputation. There is, therefore, an increasing impetus to develop novel agents that promote wound repair. The testing of these has been limited to large animal models such as swine, which are often impractical. Mice represent the ideal preclinical model, as they are economical and amenable to genetic manipulation, which allows for mechanistic investigation. However, wound healing in a mouse is fundamentally different to that of humans as it primarily occurs via contraction. Our murine model overcomes this by incorporating a splint around the wound. By splinting the wound, the repair process is then dependent on epithelialization, cellular proliferation and angiogenesis, which closely mirror the biological processes of human wound healing. Whilst requiring consistency and care, this murine model does not involve complicated surgical techniques and allows for the robust testing of promising agents that may, for example, promote angiogenesis or inhibit inflammation. Furthermore, each mouse acts as its own control as two wounds are prepared, enabling the application of both the test compound and the vehicle control on the same animal. In conclusion, we demonstrate a practical, easy-to-learn, and robust model of wound healing, which is comparable to that of humans.

A murine model of cutaneous wound healing that can be used to assess therapeutic compounds in physiological and pathophysiological settings.

Modelo murino de Curación de Heridas

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. ...

Assessing Changes in Volatile General Anesthetic Sensitivity of Mice after Local or Systemic Pharmacological Intervention

One desirable endpoint of general anesthesia is the state of unconsciousness, also known as hypnosis. Defining the hypnotic state in animals is less straightforward than it is in human patients. A widely used behavioral surrogate for hypnosis in rodents is the loss of righting reflex (LORR), or the point at which the animal no longer responds to their innate instinct to avoid the vulnerability of dorsal recumbency. We have developed a system to assess LORR in 24 mice simultaneously while carefully controlling for potential confounds, including temperature fluctuations and varying gas flows. These chambers permit reliable assessment of anesthetic sensitivity as measured by latency to return of the righting reflex (RORR) following a fixed anesthetic exposure. Alternatively, using stepwise increases (or decreases) in anesthetic concentration, the chambers also enable determination of a population's sensitivity to induction (or emergence) as measured by EC50 and Hill slope. Finally, the controlled environmental chambers described here can be adapted for a variety of alternative uses, including inhaled delivery of other drugs, toxicology studies, and simultaneous real-time monitoring of vital signs.

Loss of the righting reflex has long served as a standard behavioral surrogate for unconsciousness, also called hypnosis, in laboratory animals. Alterations in volatile anesthetic sensitivity caused by pharmacological interventions can be detected with a carefully controlled high-throughput assessment system, which may be adapted for delivery of any inhaled therapeutic.

La evaluación de cambios en la volátil Anestésico general La sensibilidad de los ratones después de la intervención farmacológica local o sistémica

One desirable endpoint of general anesthesia is  the state of unconsciousness, also known as hypnosis. Defining the hypnotic state in animals is  less ...

A Simple Critical-sized Femoral Defect Model in Mice

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all limb bone fractures fail to heal completely due to the extent of the trauma, disease, or age of the patient. Our ability to improve bone regenerative strategies is critically dependent on the ability to mimic serious bone trauma in test animals, but the generation and stabilization of large bone lesions is technically challenging. In most cases, serious long bone trauma is mimicked experimentally by establishing a defect that will not naturally heal. This is achieved by complete removal of a bone segment that is larger than 1.5 times the diameter of the bone cross-section. The bone is then stabilized with a metal implant to maintain proper orientation of the fracture edges and allow for mobility. 
Due to their small size and the fragility of their long bones, establishment of such lesions in mice are beyond the capabilities of most research groups. As such, long bone defect models are confined to rats and larger animals. Nevertheless, mice afford significant research advantages in that they can be genetically modified and bred as immune-compromised strains that do not reject human cells and tissue.
Herein, we demonstrate a technique that facilitates the generation of a segmental defect in mouse femora using standard laboratory and veterinary equipment. With practice, fabrication of the fixation device and surgical implantation is feasible for the majority of trained veterinarians and animal research personnel. Using example data, we also provide methodologies for the quantitative analysis of bone healing for the model.

Animal models are frequently employed to mimic serious bone injury in biomedical research. Due to their small size, establishment of stabilized bone lesions in mice are beyond the capabilities of most research groups. Herein, we describe a simple method for establishing and analyzing experimental femoral defects in mice.

A-crítico tamaño femoral Defecto sencillo modelo en ratones

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all ...

Direct Mouse Trauma/Burn Model of Heterotopic Ossification

Heterotopic ossification (HO) is the formation of bone outside of the skeleton which forms following major trauma, burn injuries, and orthopaedic surgical procedures. The majority of animal models used to study HO rely on the application of exogenous substances, such as bone morphogenetic protein (BMP), exogenous cell constructs, or genetic mutations in BMP signaling. While these models are useful they do not accurately reproduce the inflammatory states that cause the majority of cases of HO. Here we describe a burn/tenotomy model in mice that reliably produces focused HO. This protocol involves creating a 30% total body surface area partial thickness contact burn on the dorsal skin as well as division of the Achilles tendon at its midpoint. Relying solely on traumatic injury to induce HO at a predictable location allows for time-course study of endochondral heterotopic bone formation from intrinsic physiologic processes and environment only. This method could prove instrumental in understanding the inflammatory and osteogenic pathways involved in trauma-induced HO. Furthermore, because HO develops in a predictable location and time-course in this model, it allows for research to improve early imaging strategies and treatment modalities to prevent HO formation.

An Achilles tenotomy and burn injury model of heterotopic ossification allows for the reliable study of trauma induced ectopic bone formation without the application of exogenous factors.

Ratón directa Trauma / Burn modelo de osificación heterotópica

Heterotopic ossification (HO) is the formation of bone outside of the skeleton which forms following major trauma, burn injuries, and orthopaedic surgical ...

Establishment of a Segmental Femoral Critical-size Defect Model in Mice Stabilized by Plate Osteosynthesis

The use of tissue-engineered bone constructs is an appealing strategy to overcome drawbacks of autografts for the treatment of massive bone defects. As a model organism, the mouse has already been widely used in bone-related research. Large diaphyseal bone defect models in mice, however, are sparse and often use bone fixation which fills the bone marrow cavity and does not provide optimal mechanical stability. The objectives of the current study were to develop a critical-size, segmental, femoral defect in nude mice. A 3.5-mm mid-diaphyseal femoral ostectomy (approximately 25% of the femur length) was performed using a dedicated jig, and was stabilized with an anterior located locking plate and 4 locking screws. The bone defect was subsequently either left empty or filled with a bone substitute (syngenic bone graft or coralline scaffold). Bone healing was monitored noninvasively using radiography and in vivo micro-computed-tomography and was subsequently assessed by ex vivo micro-computed-tomography and undecalcified histology after animal sacrifice, 10 weeks postoperatively. The recovery of all mice was excellent, a full-weight-bearing was observed within one day following the surgical procedure. Furthermore, stable bone fixation and consistent fixation of the implanted materials were achieved in all animals tested throughout the study. When the bone defects were left empty, non-union was consistently obtained. In contrast, when the bone defects were filled with syngenic bone grafts, bone union was always observed. When the bone defects were filled with coralline scaffolds, newly-formed bone was observed in the interface between bone resection edges and the scaffold, as well as within a short distance within the scaffold. 
The present model describes a reproducible critical-size femoral defect stabilized by plate osteosynthesis with low morbidity in mice. The new load-bearing segmental bone defect model could be useful for studying the underlying mechanisms in bone regeneration pertinent to orthopaedic applications.

Although mouse models are invaluable tools for bone tissue engineering, models of long bone defects are sparse. This need motivated development of the present protocol which uses a locking plate with four screws and a dedicated jig to perform and stabilize a reproducible, femoral, critical-size defect with low morbidity.

Establecimiento de un defecto segmentario Modelo femoral de tamaño crítico en ratones estabilizado por osteosíntesis Plate

The use of tissue-engineered bone constructs is an appealing strategy to overcome drawbacks of autografts for the treatment of massive bone defects. As a ...

Scleral Cross-linking Using Riboflavin and Ultraviolet-A Radiation for Prevention of Axial Myopia in a Rabbit Model

Myopic individuals, especially those with severe myopia, are at higher-than-normal risk of cataract, glaucoma, retinal detachment and chorioretinal abnormalities. In addition, pathological myopia is a common irreversible cause of visual impairment and blindness1-3. Our study demonstrates the effect of scleral crosslinking using riboflavin and ultraviolet-A radiation on the development of axial myopia in a rabbit model. The axial length of the eyeball was measured by A-scan ultrasound in New Zealand white rabbits aged 13 days (male and female). The eye then underwent 360&#176; conjunctival peritomy with scleral crosslinking, followed by tarsorrhaphy. Axial elongation was induced in 13 day-old New Zealand rabbits by suturing their right eye eyelids (tarsorrhaphy). The eyes were divided into quadrants, and every quadrant had two scleral irradiation zones, each with an area of 0.2 cm&#178; and a radius of 4 mm. Crosslinking was performed by dropping 0.1% dextran-free riboflavin-5-phosphate onto the irradiation zones 20 sec before ultraviolet-A irradiation and every 20 sec during the 200 sec irradiation time. UVA radiation (370 nm) was applied perpendicular to the sclera at 57 mW/cm&#178; (total UVA light dose, 57 J/cm&#178;). Tarsorrhaphies were removed on day 55, followed by repeated axial length measurements. This study demonstrates that scleral crosslinking with riboflavin and ultraviolet-A radiation effectively prevents occlusion-induced axial elongation in a rabbit model.

We demonstrate the effect of scleral crosslinking with riboflavin and UVA on an axial elongation rabbit eye. Axial elongation was induced in 13 day-old New Zealand rabbits (male and female) by suturing their right eye eyelids (tarsorrhaphy).

Escleral Entrecruzamiento El uso de riboflavina y Ultravioleta-A de la radiación para la Prevención de la miopía axial en un modelo de conejo

Myopic individuals, especially those with severe myopia, are at higher-than-normal risk of cataract, glaucoma, retinal detachment and chorioretinal ...

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

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.

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.

La generación de fracturas femorales cerradas en ratones: un modelo para el estudio del hueso curativo

Bone fractures impose a tremendous socio-economic burden on patients, in addition to significantly affecting their quality of life. Therapeutic strategies ...

Fracture Apparatus Design and Protocol Optimization for Closed-stabilized Fractures in Rodents

The reliable generation of consistent stabilized fractures in animal models is essential for understanding the biology of bone regeneration and developing therapeutics and devices. However, available injury models are plagued by inconsistency resulting in wasted animals and resources and imperfect data. To address this problem of fracture heterogeneity, the purpose of the method described herein is to optimize fracture generation parameters specific to each animal and yield a consistent fracture location and pattern. This protocol accounts for variations in bone size and morphology that may exist between mouse strains and can be adapted to generate consistent fractures in other species, such as rat. Additionally, a cost-effective, adjustable fracture apparatus is described. Compared to current stabilized fracture techniques, the optimization protocol and new fracture apparatus demonstrate increased consistency in stabilized fracture patterns and locations. Using optimized parameters specific to the sample type, the described protocol increases the precision of induced traumas, minimizing the fracture heterogeneity typically observed in closed-fracture generation procedures.

The goal of the protocol is to optimize the fracture generation parameters to yield consistent fractures. This protocol accounts for the variations in bone size and morphology that may exist between animals. Additionally, a cost-effective, adjustable fracture apparatus is described.

Fractura de diseño de aparatos y optimización de protocolo para las fracturas cerradas-estabilizado en roedores

The reliable generation of consistent stabilized fractures in animal models is essential for understanding the biology of bone regeneration and developing ...

3D Planning and Printing of Patient Specific Implants for Reconstruction of Bony Defects

We are in the midst of the 3D era in most aspects of life, and especially in medicine. The surgical discipline is one of the major players in the medical field using the constantly developing 3D planning and printing capabilities. Computer-assisted design (CAD) and computer assisted manufacturing (CAM) are used to describe the 3D planning and manufacturing of the product. The planning and manufacturing of 3D surgical guides and reconstruction implants is performed almost exclusively by engineers. As technology advances and software interfaces become more user-friendly, it raises a question regarding the possibility of transferring the planning and manufacturing to the clinician. The reasons for such a shift are clear: the surgeon has the idea of what he wants to design, and he also knows what is feasible and could be used in the operating room. It allows him to be prepared for any scenario/unexpected results during the operation and allows the surgeon to be creative and express his new ideas using the CAD software. The purpose of this method is to provide clinicians with the ability to create their own surgical guides and reconstruction implants. In this manuscript, a detailed protocol will provide a simple method for segmentation using segmentation software and implant planning using a 3D design software. Following the segmentation and stl file production using segmentation software, the clinician could create a simple patient specific reconstruction plate or a more complex plate with a cradle for bone graft positioning. Surgical guides can be created for accurate resection, hole preparation for proper reconstruction plate positioning or for bone graft harvesting and re-contouring. A case of lower jaw reconstruction following plate fracture and nonunion healing of a trauma sustained injury is detailed.

This protocol describes the use of 3D planning and printing for reconstruction of bony defects. We use segmentation tools to create 3D models followed by 3D design software to create patient specific implants for reconstruction purposes concomitant to ablative surgery or as a second stage.

Planificación e impresión 3D de implantes específicos para el paciente para la reconstrucción de defectos óseos

We are in the midst of the 3D era in most aspects of life, and especially in medicine. The surgical discipline is one of the major players in the medical ...