We thank members of the Mariani lab for critical reading of the manuscript. Our funding sources were: the Baxter Medical Scholar Research Fellowship (to M.K.S.), USC undergraduate fellowships and the Provost, Dean Joan M. Schaeffer, and Rose Hills Fellowships, (to M.K.S.). We also acknowledge a CIRM BRIDGES fellowship through Pasadena City College (to T.T.T). and the James H. Zumberge Research and Innovation Fund, the USC Regenerative Medicine Initiative, and the NIAMS NIH under Award Number R21AR064462 (to F.V.M).

NOTA: Tutte le procedure sono in accordo con un protocollo animali approvato dalla cura e l&#39;uso degli animali Comitato Istituzionale (IACUC) presso la University of Southern California. 1. Preparazione per la chirurgia <ol><li> Lay out gli strumenti e le forniture chirurgici necessari (sterilizzato, articoli 1-18 nella lista dei materiali) e di organizzare un microscopio da dissezione e una fibra ottica trans-illuminatore in un cappuccio o su un tavolo downdraft. </li><li> Posizionare un mouse sessualmente matura (20 - 30 g, 1 - 3 mesi) di una camera di induzione con 4% isoflurano per indurre l&#39;anestesia. </li><li> Confermare anestesia con una zampa e / o test di coda pizzico. </li><li> Applicare una pomata per gli occhi del topo per evitare secchezza oculare durante l&#39;intervento chirurgico. </li><li> Spostare il mouse alla fase microscopio. Posizionare il mouse nel naso cono del tubo manutenzione e posizionare lateralmente. Fornire un sacchetto riscaldamento adiacente all&#39;animale per aiutare a mantenere la temperatura corporea. NONE:. Posizione del mouse è dipendente da preferenze personali e / o manualità del chirurgo, lato a destra oa sinistra va bene. </li><li> Regolare isoflurano al 2 - 3% per la manutenzione, e guardare la frequenza respiratoria. </li><li> Iniettare buprenorfina (0,05 mg / kg) per via sottocutanea (fianco laterale della gamba) per dolore post-operatorio con un ago 25 G. La gamba può contrarsi, che è una risposta normale. </li></ol> Incisione 2. Apertura <ol><li> Rileva la costola voluta da palpazione, e ripulire l&#39;area di capelli con un rasoio. NOTA: Ribs 8-10 sono raccomandati dal costole 1-7 sono più vicini ai polmoni e 11-13 sono notevolmente più brevi. Contando le costole dovrebbe iniziare con la costola più rostrale come numero 1. </li><li> Preparare la zona con alternanza iodopovidone e isopropanolo (70%) tampone (3x è spesso raccomandato dai protocolli di utilizzo degli animali). </li><li> Tagliare un&#39;incisione di 2 cm attraverso la pelle, direttamente sopra e parallela alla costola desiderata con le forbici microchirurgia medie dimensioni. Incise attraverso gli strati muscolari e adipose sottostanti. </li><li> Mettere tutti 3 strati (pelle, muscoli, grasso) in un riavvolgitore per esporre l&#39;area chirurgica, riducendo al minimo la dimensione della incisione. </li></ol> 3. ablazione del Rib <ol><li> Tagliare i muscoli intercostali sovrastanti la sezione desiderata della costola con un bisturi 5,0 millimetri. Posizione questa incisione attorno a 5 mm prossimale al giunto chondrocostal come questo è dove la costola non è così altamente curvo. Separare con cura il muscolo dall&#39;osso con una pinza a punta fine. </li><li> Per creare una resezione mantenendo periostio nell&#39;animale, tagliare il periostio, lungo la lunghezza della nervatura con un bisturi 5,0 millimetri. Separare con cura il periostio dall&#39;osso sottostante lateralmente con una pinza a punta fine. Procedere con cautela, come periostio è molto delicata e ha una consistenza gelatinosa. </li><li> Successivo fare una sezione trasversale attraverso l&#39;osso ad una estremità con sottili micro-forbici. Se necessario, misurare la resezione conun reticolo nel microscopio o un manometro tipo righello. Quindi sollevare con attenzione l&#39;osso dal periostio e tagliare l&#39;altra estremità. NOTA: Procedere con cautela. Questo è il passo più delicato, come tirare senza asportare la costola di seguito si strappare la membrana pleurica e il risultato in un pneumotorace. Se la membrana pleurica è strappata questo sarà immediatamente evidente come l&#39;interno della cavità toracica diventa visibile. </li><li> Se l&#39;emorragia si verifica come risultato di taglio dell&#39;osso, applicare pressione alla fine del taglio con un panno di cotone per 4 - 5 sec per fermare l&#39;emorragia. NOTA: Nelle rare occasioni che il sanguinamento continua, può essere necessario per fermare e eutanasia dell&#39;animale come significativa perdita di sangue comprometterà recupero (senza sostituzione di liquidi, il volume massimo di sangue che può essere tranquillamente essere perso è del 10% del volume totale del sangue 7,7 -. 8 microlitri / g Per 25 g del mouse, questo è equivalente a circa 180-200 microlitri 9). </li><li> Porre immediatamente la nervatura rimosso in4% PFA per le analisi future. NOTA: Se la resezione sia l&#39;osso e periostio deve essere effettuata, omettere l&#39;incisione periostio e la separazione (passo 3.2). Dal momento che il periostio è molto strettamente attaccato alla membrana pleurica procedere con cautela, prendere in giro la costola di distanza dalla membrana pleurica con una pinza per evitare strappi. </li></ol> 4. Chiusura della incisione <ol><li> Suturare la muscoli intercostali sopra la parte superiore del manicotto periostio restante con 9-0 suture (2 suture sono generalmente sufficienti). Posizionare i punti di sutura direttamente sopra le estremità tagliate della costola, di agire come indicatori per posizione chirurgia. </li><li> Rimuovere il divaricatore. Suturare il muscolo sovrastante e grasso con 9-0 punti di sutura (3 - 4 punti di sutura sono generalmente sufficienti). In topi con particolarmente spessi strati muscolari e / o grasso, suturare ogni strato separatamente (ad esempio, 1 strato di suture per lo strato muscolare e 1 strato di suture per lo strato grasso). </li><li> Chiudere ilpelle con 7-0 punti di sutura (4 - 5 punti di sutura sono generalmente sufficienti). </li><li> Fissare l&#39;incisione con la colla sutura, pizzicando i bordi con le grandi pinze. </li><li> Lentamente svezzare il mouse fuori isoflurano per primo adeguamento al 1% per alcuni minuti e poi si spegne. </li><li> Posizionare il mouse sotto una lampada di calore e lasciare fino coscienza è recuperato, 5 - 10 min. Dopo la coscienza è riconquistato e per tutto il periodo di guarigione, il mouse dovrebbe muoversi e deambulare normalmente e non mostrano segni di sofferenza. </li></ol> 5. Recupero e analisi <ol><li> Fornire la gestione del dolore post-operatorio compresa la somministrazione di buprenorfina in forma di gelatina orale a 0,5 mg / kg ogni 12 ore per 48 ore. La somministrazione orale non richiede immobilizzare l&#39;animale che potrebbe causare dolore alla regione toracica. </li><li> Mantenere il mouse in una gabbia con libero accesso a cibo e acqua durante il periodo di guarigione. Dopo l&#39;incisione è asciutto, le femmine possono essere co-ospitati durante maschios deve restare isolato per evitare lotte. </li><li> Dopo il periodo di guarigione, seguire la procedura approvata per l&#39;eutanasia. Rimuovere la gabbia toracica per la fissazione e l&#39;analisi. </li></ol>

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The authors have nothing to disclose

La prima volta che l&#39;apprendimento di questo protocollo, determinando dove localizzare l&#39;incisione iniziale può essere impegnativo. Tuttavia, la pratica sui topi eutanasia aiuta il chirurgo imparare dove posizionare l&#39;incisione iniziale ed esporre la costola desiderato da asportato. Lavorare su cadaveri migliora anche le capacità fine-motorie necessarie per rimuovere la parte toracica, con o senza il periostio. Inoltre, qualcuno di nuovo a questa procedura potrebbe trovare manipolare le belle strumenti e s...

Debilitante lesioni scheletriche, osteoartrite cronica, ei gravi problemi connessi con la chirurgia ricostruttiva impatto produttività economica, benessere familiare, e la qualità della vita. Mentre piccole pause e le lesioni possono guarire abbastanza bene, gli esseri umani non sono in grado di riparare grandi difetti e quindi deve fare affidamento su procedure ricostruttive per ripristinare la struttura e la funzione. La ricostruzione può comportare innesti allogenici o heterogeneic, osso morcellized, ponteggi impiantati, o distrazione osteogenesi. Sfortunatamente, non solo vi sono fattori di morbilità persistenti associati a questi trattamenti, ma la forza originaria dell&#39;osso riparato è raramente raggiunto. Pertanto, sono necessari nuovi approcci clinici. Senso unico per sviluppare metodi innovativi per il trattamento di difetti segmentali è quello di studiare le situazioni in cui la riparazione su larga scala avviene naturalmente. Anfibi notoriamente in grado di rigenerare gli elementi scheletrici, mentre i mammiferi sono considerate limitate in thè la capacità. Tuttavia, dal momento che la prima parte del 20 ° secolo, alcuni rapporti di rigenerazione della costola umana sono stati pubblicati suggerisce che gli esseri umani non possono essere limitati così 1-4. Attualmente questo fenomeno è meglio conosciuto da chirurghi plastici che usano materiale costola per mascella, viso e orecchio ricostruzione, ma non è più ampiamente apprezzato 5. Per studiare questa riparazione più in dettaglio, abbiamo sviluppato un modello chirurgica utilizzando il mouse. Usando questo protocollo, i ricercatori possono individuare i fattori innati coinvolti e utilizzare queste informazioni per facilitare la guarigione scheletrico in altri luoghi. Ci sono molti vantaggi di utilizzare le costole come modello per lo studio di riparazione scheletrica. Innanzitutto, le nervature circostanti offrono un fissatore naturale (rispetto alla resezione del femore 6,7). Questo riduce il rischio di morbilità fissatori interni ed esterni e semplifica la procedura chirurgica. In secondo luogo, gli strati muscolari sottili degli wa pettoll fornire un facile accesso e una visibilità eccellente che fanno il test paragonabile alla comodità di resezioni cranica 8. In terzo luogo, in contrasto con la Calvariae che costituiscono da ossificazione intramembranosa, forma nervature da encondrale e crescere in lunghezza tramite allungamento a placche di crescita situati alle due estremità di un diafisi centrale. Pertanto, la riparazione delle costole può essere più paragonabile a riparazione delle ossa lunghe dello scheletro appendicolare. Inoltre, abbiamo trovato che rispetto al femore, il periostio della nervatura è più spessa e può essere più facilmente manipolato. Così, gli investigatori che vogliono saggiare riparazione ossea al fine di studiare il periostio o testare terapie cellulari, gli agenti farmacologici e / o impalcature tissutali potrebbero trovare questo modello chirurgica utile. In sintesi, questo modello di resezione costola fornisce un contesto all&#39;interno del quale studiare la riparazione ossea larga scala naturale nei mammiferi in quanto tale modello in uso generale attualmente esistente.

<table><tbody><tr><td>Medium sized micro-dissection scissors (Vannas-T&amp;uuml;bingen Spring Scissors 5 mm)</td><td>Fine Science Tools</td><td>15003-08</td><td></td></tr><tr><td>Fine micro-dissection scissors (Vannas Spring Scissors - 2mm Cutting Edge)</td><td>Fine Science Tools</td><td>15000-04</td><td>curved tip is beneficial</td></tr><tr><td>Micro-scalpel 5.0 mm</td><td>Fine Science Tools</td><td>10315-12</td><td>other fine scalpels can be substituted</td></tr><tr><td>Dumont 55 forceps</td><td>Fine Science Tools</td><td>11295-51</td><td></td></tr><tr><td>Retractor&amp;nbsp;</td><td>Fine Science Tools</td><td>17004-05</td><td>adjustability is convenient</td></tr><tr><td>Micro-needle holders</td><td>Fine Science Tools</td><td>12060-01</td><td></td></tr><tr><td>9.0 nylon sutures (Ethilon), taper point best</td><td>Ethicon</td><td>2819G or similar</td><td>taper point best but reverse cutting is also good</td></tr><tr><td>7.0 prolene sutures (Prolene)</td><td>Ethicon</td><td>8700H or similar</td><td>6-0 can be used too, needle point can vary</td></tr><tr><td>Large forceps (Adson Forceps)</td><td>Fine Science Tools</td><td>11006-12</td><td>other brands are fine</td></tr><tr><td>Lubricant Eye Ointment (Akwa Tears)</td><td>Akorn</td><td>17478-062-35</td><td></td></tr><tr><td>Suture glue (GLUture Topical Tissue Adhesive)</td><td>Abbot</td><td>32046-01</td><td>has excellent working time</td></tr><tr><td>Shaver</td><td>Wahl</td><td>9918-6171 or similar</td><td></td></tr><tr><td>Clamp lamp</td><td>Zoo Med</td><td>LF-5</td><td></td></tr><tr><td>Infrared Bulb, 75W</td><td>Zoo Med</td><td>RS-75</td><td></td></tr><tr><td>RC&lt;sup&gt;2&lt;/sup&gt; Rodent Anesthesia System</td><td>VetEquip&amp;nbsp;</td><td>922100</td><td></td></tr><tr><td>IsoFlo (Isoflurane)</td><td>Abbot</td><td>05260-05</td><td></td></tr><tr><td>Buprenorphine (Buprenex)</td><td>Reckitt Benckiser</td><td>12496-0757-1</td><td></td></tr><tr><td>Betadine</td><td>Purdue Frederick</td><td>67618015017</td><td></td></tr><tr><td>Flavored Gelatin, raspberry</td><td>Jell-O</td><td>B000E1FYL0</td><td>made up firm, to the consistency of &#039;jigglers&#039;</td></tr></tbody></table>

a surgical procedure for resecting the mouse rib: a model for large-scale long bone repair

Questo protocollo introduce ricercatori di un nuovo modello di grande riparazione ossea utilizzando la costola del mouse. Dettagli La procedura seguente: preparazione del animale per la chirurgia, aprendo la parete del corpo toracica, esponendo la nervatura desiderato dai muscoli intercostali circostanti, asportare la sezione desiderata della costola senza indurre un pneumotorace, e chiudendo le incisioni. Rispetto alle ossa dello scheletro appendicolare, le nervature sono altamente accessibile. Inoltre, nessun fissatore interno o esterno è necessario in quanto le nervature adiacenti forniscono una fissazione naturale. L&#39;intervento utilizza commercialmente scorte disponibili, è semplice da imparare, e ben tollerato dall&#39;animale. La procedura può essere eseguita con o senza rimuovere il periostio circostante, e quindi il contributo del periostio di riparazione può essere valutata. I risultati indicano che se il periostio viene mantenuta, riparazione robusto si verifica in 1 - 2 mesi. Ci aspettiamo che l&#39;uso di questo protocollo saràstimolare la ricerca in riparazione costola e che i risultati faciliteranno lo sviluppo di nuovi modi per stimolare la riparazione ossea in altre località in tutto il corpo.

Animali tipicamente recuperare rapidamente da questa procedura, l&#39;incisione guarisce bene, e gli animali mostrano un comportamento normale. La procedura utilizza materiali di consumo disponibili sul mercato e dopo la pratica, può essere completato in meno di 1 ora. Prima di utilizzare questa procedura per generare dati sperimentali, è importante analizzare resezione al punto di tempo di giorno 0 per determinare se eventuali grandi frammenti ossei potrebbero rimanere all&#39;interno della zona di resezione. Un modo...

Watch this Scientific Journal Video about A Surgical Procedure for Resecting the Mouse Rib: A Model for Large-Scale Long Bone Repair at JoVE.com

A Surgical Procedure for Resecting the Mouse Rib: A Model for Large-Scale Long Bone Repair

The overall goal of this procedure is to successfully resect a portion of bone from the rib of a mouse. The procedure was developed as a model to study large-scale long bone repair.

Una procedura chirurgica per la resezione Rib Mouse: Un modello per le grandi ossa lunghe Repair

This protocol introduces researchers to a new model for large-scale bone repair utilizing the mouse rib. The procedure details the following: preparation ...

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Research

JoVE Journal

Medicine

13.1K Views.  Keck School of Medicine, University of Southern California. The overall goal of this procedure is to successfully resect a portion of bone from the rib of a mouse. The procedure was developed as a model to study large-scale long bone repair.

Video: A Surgical Procedure for Resecting the Mouse Rib: A Model for Large-Scale Long Bone Repair

A Mouse Distraction Osteogenesis Model

Distraction osteogenesis (DO) is a surgical procedure that involves skeletal tissue regeneration without cell transplantation. A DO model consists of the following three phases: the latency phase after osteotomy and placement of the external distractor; the distraction phase, wherein the separated bone ends are gradually and continuously distracted; and the consolidation phase. This custom-made distractor used for DO is comprised of two incomplete acrylic resin rings and an expansion screw. The process was initiated by making a mold with silicone impression material and then creating the custom-made distractor. Dental resin was poured into the formwork made of silicone impression material, and it was allowed to polymerize to create the incomplete resin rings required for the custom-made distractor. These rings were fixed with an expansion screw using transparent resin. The custom-made distractor created via this approach was attached to the tibia of mice. The tibia was fixed to the device using one pair of 25 G needles proximally, one pair of 27 G needles distally, and acrylic resin. After a latency period of 5 days, distraction was initiated at a rate of 0.2 mm/12 h. The lengthening was continued for 8 days, resulting in a total gap of 3.2 mm. The mice were sacrificed 4 weeks after distraction. Bone formation in the distraction gap was confirmed using both radiography and histology.

We present a mouse tibial distraction osteogenesis model developed using a custom-made distractor. The use of a mouse as an analysis target is advantageous for advancing research.

Un modello di osteogenesi di distrazione del topo

Distraction osteogenesis (DO) is a surgical procedure that involves skeletal tissue regeneration without cell transplantation. A DO model consists of the ...

Ricerca

Biologia dello sviluppo

Mouse Pneumonectomy Model of Compensatory Lung Growth

In humans, disrupted repair and remodeling of injured lung contributes to a host of acute and chronic lung disorders which may ultimately lead to disability or death. Injury-based animal models of lung repair and regeneration are limited by injury-specific responses making it difficult to differentiate changes related to the injury response and injury resolution from changes related to lung repair and lung regeneration. However, use of animal models to identify these repair and regeneration signaling pathways is critical to the development of new therapies aimed at improving pulmonary function following lung injury. The mouse pneumonectomy model utilizes compensatory lung growth to isolate those repair and regeneration signals in order to more clearly define mechanisms of alveolar re-septation. Here, we describe our technique for performing mouse pneumonectomy and sham pneumonectomy. This technique may be utilized in conjunction with lineage tracing or other transgenic mouse models to define molecular and cellular mechanism of lung repair and regeneration.

Mouse pneumonectomy is a commonly employed model of compensatory lung growth. This procedure can be used in conjunction with lineage tracing or transgenic mouse models to elucidate underlying mechanisms.

Topo Pneumonectomia Modello di crescita compensatoria Lung

In humans, disrupted repair and remodeling of injured lung contributes to a host of acute and chronic lung disorders which may ultimately lead to ...

Medicina

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.

Un semplice Critical dimensioni femorale Difetto Modello 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 ...

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.

Istituzione di un segmentale femorale critico-size difetto del modello in topi stabilizzati da osteosintesi Piastra

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 ...

A Mouse Model of Orthopedic Surgery to Study Postoperative Cognitive Dysfunction and Tissue Regeneration

Surgery is commonly used to improve and maintain quality of life. Unfortunately, in vulnerable patients such as the elderly, complications may occur and significantly diminish the outcome. Indeed, after routine orthopedic surgery to repair a fracture, as many as 50% of elderly patients suffer from neurologic complications like delirium. Also, the capacity to heal and regenerate tissue after surgery decreases with age, and can impact the quality of fracture repair and even osseous integration of implants. Thus, a better understanding of mechanisms that drive these age-dependent changes could provide strategic targets to minimize risk for such complications and optimize outcomes. Here, we introduce a clinically relevant mouse model of tibial fracture. The postoperative changes in these mice mimic some of the cognitive impairments commonly observed after routine orthopedic surgery in humans. Briefly, an incision is performed in the right hind limb under strictly aseptic conditions. Muscles are disassociated, and a 0.38-mm stainless steel pin is inserted into the upper crest of the tibia, inside the intramedullary canal. Osteotomy is then performed, and the wound is stapled. We have used this model to investigate the effects of surgical trauma on postoperative neuroinflammation and behavioral changes. By applying this fracture model in combination with parabiosis, a surgical model in which 2 mice are anastomosed, we have studied cells and secreted factors that systemically rejuvenate organ function and tissue regeneration after injury. By following our step-by-step protocol, these models can be reproduced with high fidelity, and can be adapted to interrogate many biologic pathways that are altered by surgical trauma.

This protocol describes a mouse model of orthopedic surgery that has been used to study mechanisms of postoperative neuroinflammation and behavioral changes, and when combined with parabiosis, to study tissue regeneration during aging.

Un modello murino di chirurgia ortopedica per studiare la disfunzione cognitiva postoperatoria e rigenerazione tissutale

Surgery is commonly used to improve and maintain quality of life. Unfortunately, in vulnerable patients such as the elderly, complications may occur and ...

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.

Un modello come minimo dilagante per analizzare Endochondral frattura la guarigione nei topi in condizioni standardizzate di biomeccaniche

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

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 generazione delle fratture chiuse femorale in topi: un modello per studiare la guarigione ossea

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

Transverse Fracture of the Mouse Femur with Stabilizing Pin

Fracture repair is an essential function of the skeleton that cannot be reliably modeled in vitro. A mouse injury model is an efficient approach to test whether a gene, gene product or drug influences bone repair because murine bones recapitulate the stages observed during human fracture healing. When a mouse or human breaks a bone, an inflammatory response is initiated, and the periosteum, a stem cell niche surrounding the bone itself, is activated and expands. Cells residing in the periosteum then differentiate to form a vascularized soft callus. The transition from the soft callus to a hard callus occurs as the recruited skeletal progenitor cells differentiate into mineralizing cells, and the bridging of the fractured ends results in the bone union. The mineralized callus then undergoes remodeling to restore the original shape and structure of the healed bone. Fracture healing has been studied in mice using various injury models. Still, the best way to recapitulate this entire biological process is to break through the cross-section of a long bone that encompasses both cortices. This protocol describes how a stabilized, transverse femur fracture can be safely performed to assess healing in adult mice. A surgical protocol including detailed harvesting and imaging techniques to characterize the different stages of fracture healing is also provided.

This protocol describes a method to perform fractures on adult mice and monitor the healing process.

Frattura trasversale del femore del topo con perno stabilizzante

Fracture repair is an essential function of the skeleton that cannot be reliably modeled in vitro. A mouse injury model is an efficient approach to test ...

Biologia

Establishing a Diaphyseal Femur Fracture Model in Mice

Bones have a significant regenerative capacity. However, fracture healing is a complex process, and depending on the severity of the lesions and the age and overall health status of the patient, failures can occur, leading to delayed union or nonunion. Due to the increasing number of fractures resulting from high-energy trauma and aging, the development of innovative therapeutic strategies to improve bone repair based on the combination of skeletal/mesenchymal stem/stromal cells and biomimetic biomaterials is urgently needed. To this end, the use of reliable animal models is fundamental to better understanding the key cellular and molecular mechanisms that determine the healing outcomes. Of all the models, the mouse is the preferred research model because it offers a wide variety of transgenic strains and reagents for experimental analysis. However, the establishment of fractures in mice may be technically challenging due to their small size. Therefore, this article aims to demonstrate the procedures for the surgical establishment of a diaphyseal femur fracture in mice, which is stabilized with an intramedullary wire and resembles the most common bone repair process, through cartilaginous callus formation.

This protocol describes a surgical procedure for the establishment of a diaphyseal fracture in the femur of mice, which is stabilized with an intramedullary wire, for fracture healing studies.

Stabilire un modello di frattura del femore diafisario nei topi

Bones have a significant regenerative capacity. However, fracture healing is a complex process, and depending on the severity of the lesions and the age ...

Visualizzazione 3D del rimodellamento osseo sotto carico di trazione in topi ad espansione di sutura

A 3-D Visualization Technique for Bone Remodeling in a Suture Expansion Mouse Model

Craniofacial sutures play a crucial role beyond being fibrous joints connecting craniofacial bones; they also serve as the primary niche for calvarial and facial bone growth, housing mesenchymal stem cells and osteoprogenitors. As most craniofacial bones develop through intramembranous ossification, the sutures' marginal regions act as initiation points. Due to this significance, these sutures have become intriguing targets in orthopedic therapies like spring-assisted cranial vault expansion, rapid maxillary expansion, and maxillary protraction. Under orthopedic tracing force, suture stem cells are rapidly activated, becoming a dynamic source for bone remodeling during expansion. Despite their importance, the physiological changes during bone remodeling periods remain poorly understood. Traditional sectioning methods, primarily in the sagittal direction, do not capture the comprehensive changes occurring throughout the entire suture. This study established a standard mouse model for sagittal suture expansion. To fully visualize bone remodeling changes post-suture expansion, the PEGASOS tissue clearing method was combined with whole-mount EdU staining and calcium chelating double labeling. This allowed the visualization of highly proliferating cells and new bone formation across the entire calvarial bones following expansion. This protocol offers a standardized suture expansion mouse model and a 3-D visualization method, shedding light on the mechanobiological changes in sutures and bone remodeling under tensile force loading.

Autore in primo piano: Tecnica di pulizia dei tessuti PEGASOS per visualizzare il rimodellamento osseo 

This protocol presents a standardized suture expansion mouse model and a 3-D visualization method to study the mechanobiological changes of the suture and bone remodeling under tensile force loading.

Una tecnica di visualizzazione 3D per il rimodellamento osseo in un modello murino di espansione della sutura

Craniofacial sutures play a crucial role beyond being fibrous joints connecting craniofacial bones; they also serve as the primary niche for calvarial and ...