Name: direct mouse trauma/burn model of heterotopic ossification
Uploaded: 2015-08-06T14:00:00.000+00:00
Duration: 7 min 1 s
Description: Watch this Scientific Journal Video about Direct Mouse Trauma/Burn Model of Heterotopic Ossification at JoVE.com

We thank Amanda Fair, the CMI, and Kathy Sweet and the ORL at UM for assistance with &#181;CT imaging and analysis. Funding: BL Funded by 1K08GM109105-01 and Plastic Surgery Foundation National Endowment Award.

윤리 문 : 실험 동물 연구를위한 연구소의 여덟 번째 판 (ILAR, 2011)과 같다 : 모든 동물의 절차는 실험 동물의 사용 및 관리에 대한 가이드에 제공된 지침에 정의 된대로 좋은 동물 관행에 엄격한에 따라 수행 하였다 미시간 대학의 기관 동물 관리 및 사용위원회 (PRO0001553)에 의해 승인했다. 1. 마우스 외과 적 치료 참고 : 사용 8-10주 된 C57BL가 / 6 마​​우스. 다른 연령, 배경, 및 생쥐 균주는 또한 상이한 조건 또는 유전 적 테스트를 위해 사용될 수있다. 프로 시저의 건 절단술 부분에 대한, 얼굴 마스크, 헤어 모자, 멸균 가운, 장갑, 및 악기를 사용하여 멸균 조건을 유지한다. 외과 사이트는 무균 포비돈 - 요오드 살균 입체 재단으로 준비를해야합니다. 37 ° C의 소생 솔루션을 수의학 가열 패드를 사용하고 따뜻하게하여 체온을 피 administra 이전기. <ol><li> 2.5 % 흡입 이소 플루 란을 이용하여 마우스를 마취. 쉽게 접근 배부 및 뒷다리 - 사지를 떠나 코 콘을 사용합니다. 점막의 적절한 호흡 속도와 착색을 유지하기 위해 이소 플루 란 관리의 속도를 조절하고 근육, 발가락 핀치, 각막 반사를 확인하여 마취을 보장합니다. 절차를 수행하는 동안 건조 눈 부상을 방지하기 위해 마우스의 눈에 개성 안과 연고를 적용합니다. 수술 필드 아래 수의학 가열 패드 또는 온수 순환 장치를 사용하여 적절한 체온을 유지한다. </li><li> 무통 수술에 피하 직전에, 부 프레 노르 핀, 0.1 ㎎ / ㎏을 관리합니다. </li><li> 밀접하게 무릎 발 뒤꿈치에서 가위를 사용하여 왼쪽 뒷발에 수술 부위를 면도. </li><li> 밀접하게 척추를 통해 시작과 등쪽 정중선의 왼쪽에 면도 영역을 확장, 화상 부상을 수용 할 수있는 영역을 적어도 2cm × 3 cm의 제거, 마우스의 배부를 면도. </li><li>3 교대 포비돈 - 요오드 스크럽을 사용하여 무릎에 뒷발을 포함하여 수술 부위를 준비합니다. </li><li> 왼쪽 아킬레스 건의 안쪽면을 따라 세로 절개를 수행합니다. 아킬레스 건 쉽게 시각화 할 수 있도록 절개를 확장; 약 0.5 cm. </li><li> 날카로운 조직 가위로 중간에 힘줄의 날카로운 해부와 아킬레스 건 절단술을 수행합니다. 건 아래에있는 조직면에서 조직 가위 중 하나 블레이드를 삽입하고 블레이드 건 중간에 때까지 평면을 따라 해부. 급격히 건을 절단하기 위해 가위 블레이드를 닫습니다. 참고 : 비복근, 가자미근과 족저 건을 포함하여 모든 뒤쪽 힘줄의 포함을 확인합니다. </li><li> 멸균 거즈로 압력을 적용하여 지혈을 달성 (이 최소한의 출혈을해야한다)와 5-0 vicryl 스티치로 피부 절개를 닫습니다. </li><li> 대략 measuremen와 35g의 무게 알루미늄 블록 등의 부분 층 화상을 수행TS 2cm X 2cm X 3cm 17 초간 마우스의 등쪽에 도포 면도 수욕에서 60 ° C로 가열 하였다. 주 : 그러나 즉, 블록에 추가적인 압력을 가하지 않도록, 블록의 전체 표면 영역은 마우스와 접촉하는 것을 보장하는, 마취 된 쥐의 상부에 블록을 놓아 적절한 화상 농도를 달성 할 중력 허용 장소에 블록을 들고있는 유일한 힘. 블록의 일 측면에 부착 된 얇은 플라스틱 핸들 온수욕에서 조작, 안정화, 및 검색을 위해 편리하다. 이는 약 30 %의 총 체 표면적이 8-10 주 된 C57BL / 6 마​​우스에 화상이 생성됩니다. 이 접촉 화상으로 인해 상처를 통해 화상 깊이의 균일과 동물 사이의 재현성에 다른 방법 (불꽃 또는 소독 번)를 통해 선정되었다. </li><li> 거즈로 화상 부위를 건조하고 tegaderm 드레싱을 적용합니다. </li><li> 따뜻하게 소생술 유체를 관리 : 락 테이트 링거액 1 ml에복강 내 주사하고 0.5 mL를 피하 주사. 소생이 한 시간 관리는 화상 부상과 건 절단술에서 복구에 적합합니다. 참고 : 마우스가 완전히 회복 될 때까지 모니터링에서 깨끗한 케이지에 개별적으로, 다른 마우스와 주택 케이지에 집에 마취 된 쥐를 반환하지 않습니다. 일반적인 복구는 1-6 시간 내에 발생합니다. 굽​​기 사이트는 일반적으로 2~4주 내에 치유하고 깨끗한 주거 환경에서 보관하는 경우는 거의 창상 감염에 의해 복잡하지 않습니다. </li><li> 부 프레 놀핀 0.1 ㎎ / ㎏ 피하 주사에게 절차에 따라 3 일간 매 12 시간을 관리 할 수​​ 있습니다. 적절한 간격으로 일련 μCT 검사와 HO의 성장을 모니터링합니다. 자궁외 뼈의 성장이 먼저 μCT에 의해 분명 약 삼주 수술 후입니다. 참고 : HO 개발의 대부분은 수술 후 구주로 완료됩니다. 우리는 15 주에 검사 μCT 단일 엔드 포인트에 비해 15주에 대한 격주 호 볼륨 또는 반복 μCT와 위치의 차이를 스캔 관찰되지 않았다.실험의 끝에서, 제도적 지침에 따라 CO 2 흡입으로 모든 쥐를 안락사와 10 분 후 자궁 전위와 죽음을 확인합니다. </li></ol> 2. μCT 수집 및 분석 <ol><li> 발생하기 쉬운 위치에 스캐너 침대에 마취 마우스를 고정합니다. 동 잡음 호흡 방지하기 위해 침대에 단단히 뒷다리를 테이프입니다. 이미지 보정을위한 마우스 아래에 팬텀이 포함 된 공기, 물, 및 하이드 록시 아파타이트를 포함합니다. </li><li> 뼈 분석 소프트웨어를 열고 근위 말단 뒷발의 선단에 고관절에서 양쪽 뒷다리를 포함 ROI (region of interest)를 정의한다. 다음 매개 변수를 사용하여 이미지를 얻습니다. 80 kV로, 500 μA 및 1,300 밀리 초 노출, 48 μm의 복셀 크기를 14, 15를 </li><li> 에 해당 필드에 평균 밀도를 세 팬텀 챔버의 각 투자 수익 (ROI)을 그리기 및 입력하여 하운 스 필드 단위 (HU)에 이미지를 보정소프트웨어. </li><li> 왼쪽 뒷다리의 경골 평행 동소 피질골 구조 및 HO의 묘사를위한 명확한 해부학 뷰를 허용하는 Z 축을 따라하므로 소프트웨어 &quot;그래버&quot;툴을 사용하여, 화상을 재 방향. </li><li> HO가 발견 될 때까지 무릎에서 시작하여, 이미지 조각을 원심으로 스크롤합니다. 수동 스플라인 도구를 사용하여 발을 통해 또는 HO가 초과 될 때까지 원심 계속 매 5 번째 조각에 자궁외 뼈 주변의 투자 수익 (ROI)을합니다. 확장하고 모든 호를 포함하는 하나의 투자 수익 (ROI)에 함께 로아 스티치 추정 해 도구를 사용합니다. </li><li> 3D 투자 수익 (ROI)을 확인하고 분석 메뉴를 선택합니다. 가장 뼈 윈도우 표시 하위 및 상위 임계 값을 설정하여 뼈의 양을 계산한다. 모든 검사에 대해 동일한 고정 임계 값을 사용합니다. </li></ol>

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The authors have nothing to disclose

Heterotopic ossification represents a major functional impairment faced by patients that sustain trauma, burns, and invasive musculoskeletal procedures. The most at-risk population are soldiers in modern conflicts with major blast injuries from mechanisms such as improvised explosive devices (IED).18 Improved body armor and forward positioned medical units allows for improved survival of major extremity injury. After initial stabilization and repair of their extremity injury, these patients are at high risk...

Heterotopic ossification (HO) is the formation of ectopic bone in which osteo-potent cells are aberrantly induced to form endochondral bone outside of the skeleton. While the details of the HO formation pathway are still largely unknown, an accepted paradigm includes three key factors: an inflammatory inciting incident, a permissive niche, and mesenchymal stem cells capable of forming bone.1-3 HO is a common comorbidity complicating over 60% of major burn injuries, 65% of combat-related injuries, and 10% of invasive orthopaedic surgery cases.4,5 However, it is often difficult to predict where HO will form because it can occur at sites of local injury or at distant locations that may be otherwise uninjured. This variability in location makes it difficult to intervene prophylactically to prevent reactive bone formation in a locally targeted manner. There are also congenital forms of HO such as fibrodysplasia ossificans progressiva (FOP) in which patients are prone to the development of robust HO in response to minor trauma or inflammatory insult. Powerful animal models using transgenic mice have reproduced this phenotype and provided insight to the molecular pathways that may also be important in trauma induced HO.6-9 Translational research into the pathogenesis of non-congenital HO has used a wide variety of constructs ranging from injury alone to the implantation of exogenous osteo-inductive materials and/or cells.10-13
In our prior work we have validated a simple and reliable model of HO formation in mice which does not require the administration of any exogenous material.14-17 This model created two key conditions to initiate HO: local trauma and global inflammation. This was achieved through the use of an Achilles tenotomy (local trauma) combined with a distant burn injury (global inflammation). Mice received both treatments concurrently and were found to develop a robust amount of HO that could be analyzed by histologic, radiologic, and molecular means. Interestingly, concurrent burn injury significantly increased the amount of HO that formed and accelerated its developmental time-course.14-16 HO developed at predictable sites around the calcaneus, ankle joint, and tibia/fibula of the limb that received the tenotomy. The reliability of HO development at a known location allowed for focused examination of molecular and histologic features in the early stages of ectopic ossification.14,17 To date, 100% of mice (over 50 animals) with a tenotomy and concurrent burn injury have developed HO. Additionally, longitudinal 2D and 3D imaging and spectroscopic analysis were conducted to examine the growth pattern and biochemical make-up of HO.15,16

<table><tbody><tr><td>C57BL/6 mice</td><td>Jackson Laboratory</td><td>664</td><td>8-10 weeks old</td></tr><tr><td>Isoflurane &amp;ndash; Fluriso</td><td>VET one, Boise, ID</td><td>V1 501017</td><td></td></tr><tr><td>Buprenorphine &amp;ndash; Buprenex</td><td>Reckitt Benckiser Healthcare</td><td>NDC 12496-0757-1</td><td>0.3 mg/ml solution</td></tr><tr><td>Betadine</td><td>Owens and Minor, Mechanicsville, VA</td><td>2047PVP202</td><td></td></tr><tr><td>5-0 Vicryl sutures</td><td>Ethicon, Summerville, NJ</td><td>J493</td><td></td></tr><tr><td>Tegaderm Film, 6 cm x 7 cm</td><td>3M</td><td>1624W</td><td>Cut in half to properly cover burn site</td></tr><tr><td>&amp;micro;CT - GE eXplore Locus SP</td><td>GE Healthcare Pre-Clinical Imaging, London, ON, Canada</td><td></td><td></td></tr><tr><td>Microview 2.2 Advanced Bone Analysis Application</td><td>GE Healthcare Pre-Clinical Imaging, London, ON, Canada</td><td></td><td></td></tr></tbody></table>

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.

이 본 연구에서, 프로토콜은 외상의 이전에 발행 된 마우스 모델에 대한 설명 / 호. 14 ~ 17이 부상을 구울 부분 두께의 아킬레스 건 절단술 및 글로벌 염증 모욕과 동시에 지역화 근골격계 손상의 생성을 포함 유도 구울 수 있습니다. 이 시리얼 이미징 따라 할 수있는 건 절단술 사이트에서 반응 뼈의 안정적인 형성을 초래한다. 현재까지 모두 아킬레스 건 절단술을 유지하고 부상을 구울 수...

Watch this Scientific Journal Video about Direct Mouse Trauma/Burn Model of Heterotopic Ossification at JoVE.com

Direct Mouse Trauma/Burn Model of Heterotopic Ossification

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.

직접 마우스 외상 / 이소성 골화의 모델을 굽기

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

direct-mouse-traumaburn-model-of-heterotopic-ossification

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

10.0K 조회수.  University of Michigan Medical School. 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.

비디오: Direct Mouse Trauma/Burn Model of Heterotopic Ossification

Pseudofracture: An Acute Peripheral Tissue Trauma Model

Following trauma there is an early hyper-reactive inflammatory response that can lead to multiple organ dysfunction and high mortality in trauma patients; this response is often accompanied by a delayed immunosuppression that adds the clinical complications of infection and can also increase mortality.1-9 Many studies have begun to assess these changes in the reactivity of the immune system following trauma.10-15 
Immunologic studies are greatly supported through the wide variety of transgenic and knockout mice available for in vivo modeling; these strains aid in detailed investigations to assess the molecular pathways involved in the immunologic responses.16-21
The challenge in experimental murine trauma modeling is long term investigation, as fracture fixation techniques in mice, can be complex and not easily reproducible.22-30
This pseudofracture model, an easily reproduced trauma model, overcomes these difficulties by immunologically mimicking an extremity fracture environment, while allowing freedom of movement in the animals and long term survival without the continual, prolonged use of anaesthesia. The intent is to recreate the features of long bone fracture; injured muscle and soft tissue are exposed to damaged bone and bone marrow without breaking the native bone.
The pseudofracture model consists of two parts: a bilateral muscle crush injury to the hindlimbs, followed by injection of a bone solution into these injured muscles. The bone solution is prepared by harvesting the long bones from both hindlimbs of an age- and weight-matched syngeneic donor. These bones are then crushed and resuspended in phosphate buffered saline to create the bone solution.
Bilateral femur fracture is a commonly used and well-established model of extremity trauma, and was the comparative model during the development of the pseudofracture model. Among the variety of available fracture models, we chose to use a closed method of fracture with soft tissue injury as our comparison to the pseudofracture, as we wanted a sterile yet proportionally severe peripheral tissue trauma model. 31
Hemorrhagic shock is a common finding in the setting of severe trauma, and the global hypoperfusion adds a very relevant element to a trauma model. 32-36 The pseudofracture model can be easily combined with a hemorrhagic shock model for a multiple trauma model of high severity. 37

Pseudofracture, a reproducible murine model of sterile musculoskeletal trauma, allows for evaluation of late term post-traumatic immune responses. This article describes the procedural execution of the model step by step, including the potential for experimental model combinations to permit study of multiple trauma.

Pseudofracture : 급성 주변 조직 외상 모델

Following trauma there is an early hyper-reactive inflammatory response that can lead to multiple organ dysfunction and high mortality in trauma patients; ...

연구

의학

Intratibial Osteosarcoma Cell Injection to Generate Orthotopic Osteosarcoma and Lung Metastasis Mouse Models

Osteosarcoma is the most common primary bone cancer in children and adolescents, with lungs as the most common metastatic site. The five-year survival rate of osteosarcoma patients with pulmonary metastasis is less than 30%.Therefore, the utilization of mouse models mimicking the osteosarcoma development in humans is of great significance for understanding the fundamental mechanism of osteosarcoma carcinogenesis and pulmonary metastasis to develop novel therapeutics. Here, detailed procedures are reported to generate the primary osteosarcoma and pulmonary metastasis mouse models via&#160;intratibia injection of osteosarcoma cells. Combined with the bioluminescence or X-ray live imaging system, these living mouse models are utilized to monitor and quantify osteosarcoma growth and metastasis. To establish this model, a basement membrane matrix containing osteosarcoma cells was loaded in a micro-volume syringe and injected into one tibia of each athymic mouse after being anesthetized. The mice were sacrificed when the primary osteosarcoma reached the size limitation in the IACUC-approved protocol. The legs bearing osteosarcoma and the lungs with metastasis lesions were separated. These models are characterized by a short incubation period, rapid growth, severe lesions, and sensitivity in monitoring the development of primary and pulmonary metastatic lesions. Therefore, these are ideal models for exploring the functions and mechanisms of specific factors in osteosarcoma carcinogenesis and pulmonary metastasis, the tumor microenvironment, and evaluating the therapeutic efficacy in vivo.

The present protocol describes intratibia osteosarcoma cell injection to generate mouse models bearing orthotopic osteosarcoma and pulmonary metastasis lesions.

경골 내 골육종 세포 주사 동위원소 골육종 및 폐 전이 마우스 모델 생성

Osteosarcoma is the most common primary bone cancer in children and adolescents, with lungs as the most common metastatic site. The five-year survival ...

암 연구학

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.

마우스 산만 골 모델

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

발생학

Adult Mouse Digit Amputation and Regeneration: A Simple Model to Investigate Mammalian Blastema Formation and Intramembranous Ossification

Here, we present a protocol of adult mouse distal terminal phalanx (P3) amputation, a procedurally simple and reproducible mammalian model of epimorphic regeneration, which involves blastema formation and intramembranous ossification analyzed by fluorescence immunohistochemistry and sequential in-vivo microcomputed tomography (&#956;CT). Mammalian regeneration is restricted to amputations transecting the distal region of the terminal phalanx (P3); digits amputated at more proximal levels fail to regenerate and undergo fibrotic healing and scar formation. The regeneration response is mediated by the formation of a proliferative blastema, followed by bone regeneration via intramembranous ossification to restore the amputated skeletal length. P3 amputation is a preclinical model to investigate epimorphic regeneration in mammals, and is a powerful tool for the design of therapeutic strategies to replace fibrotic healing with a successful regenerative response. Our protocol uses fluorescence immunohistochemistry to 1) identify early-and-late blastema cell populations, 2) study revascularization in the context of regeneration, and 3) investigate intramembranous ossification without the need for complex bone stabilization devices. We also demonstrate the use of sequential in vivo &#956;CT to create high resolution images to examine morphological changes after amputation, as well as quantify volume and length changes in the same digit over the course of regeneration. We believe this protocol offers tremendous utility to investigate both epimorphic and tissue regenerative responses in mammals.

Here, we present a protocol of adult mouse terminal phalanx amputation to investigate mammalian blastema formation and intramembranous ossification, analyzed by fluorescent immunohistochemistry and sequential in-vivo microcomputed tomography.

성인 마우스 숫자 절단 및 재생 : 포유류 블라스터 마 형성 및 intramembranous Ossification을 조사하는 간단한 모델

Here, we present a protocol of adult mouse distal terminal phalanx (P3) amputation, a procedurally simple and reproducible mammalian model of epimorphic ...

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.

마우스의 간단한 임계 크기의 대퇴 결함 모델

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.

마우스의 분절 대퇴 임계 크기 결함 모델의 설립은 플레이트 고정술에 의해 안정화

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.

정형 외과 수술 후 인지 기능 장애 및 조직 재생 연구의 마우스 모델

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

Partial Heterotopic Hindlimb Transplantation Model in Rats

Vascularized composite allotransplantations (VCA) represent the most advanced reconstruction option for patients without autologous surgical possibilities after a complex tissue defect. Face and hand transplantations have changed disfigured patients' lives, giving them a new aesthetic and functional social organ. Despite promising outcomes, VCA is still underperformed due to life-long immunosuppression comorbidities and infectious complications. The rat is an ideal animal model for in vivo studies investigating immunological pathways and graft rejection mechanisms. Rats are also widely used in novel composite tissue graft preservation techniques, including perfusion and cryopreservation studies. Models used for VCA in rats must be reproducible, reliable, and efficient with low postoperative morbidity and mortality. Heterotopic limb transplantation procedures fulfill these criteria and are easier to perform than orthotopic limb transplants. Mastering rodent microsurgical models requires solid experience in microsurgery and animal care. Herein is reported a reliable and reproducible model of partial heterotopic osteomyocutaneous flap transplantation in rats, the postoperative outcomes, and the means of prevention of potential complications.

This paper presents a partial heterotopic osteomyocutaneous flap transplantation protocol in rats and its potential outcomes in the mid-term follow-up.

쥐의 부분 이종화 힌드래그 이식 모델

Vascularized composite allotransplantations (VCA) represent the most advanced reconstruction option for patients without autologous surgical possibilities ...

면역학 및 감염병학

A Mouse Model of Mechanotransduction-driven, Human-like Hypertrophic Scarring

Hypertrophic scarring (HTS) is an abnormal process of wound healing that results in excessive scar tissue formation. Over the past decade, we have demonstrated that mechanotransduction&#8212;the conversion of mechanical stimuli into cellular responses&#8212;drives excessive fibrotic scar healing. A mouse model to assess human-like hypertrophic scarring would be an essential tool for examining various therapeutics and their ability to reduce scarring and improve healing. Specifically, our laboratory has developed a murine wound model that increases mechanical strain to promote human-like HTS. This&#160;protocol utilizes biomechanical loading devices, made from modified 13 mm palatal expanders, whose arms are placed on either side of the incision and distracted incrementally apart in order to apply continuous tension across the wound bed during healing. Over nearly two decades of use, this model has been significantly advanced to improve efficacy and reproducibility. Using the murine HTS model, significant dermal fibrotic scars can be induced to be histologically comparable to human hypertrophic scars. This murine model provides an environment to develop biologics involved in the treatment of HTS and mechanotransduction-related conditions such as foreign body response.

This protocol will explain how to establish a hypertrophic scarring murine model that increases mechanotransduction signaling to simulate human-like scarring. This method involves increasing mechanical tension across a healing incision in a mouse and using a specialized device to create reproducible, excessive scar tissue for detailed histological and bioinformatic analyses.

Hypertrophic scarring (HTS) is an abnormal process of wound healing that results in excessive scar tissue formation. Over the past decade, we have ...

생체공학

A Murine Model of a Burn Wound Reconstructed with an Allogeneic Skin Graft

Trivial superficial wounds heal without complications by primary intention. Deep wounds, such as full thickness burns, heal by secondary intention and require surgical debridement and skin grafting. Successful integration of the donor graft into a recipient wound bed depends on timely recruitment of immune cells, robust angiogenic response and new extracellular matrix formation. The development of novel therapeutic agents, which target some key processes involved in wound healing, are hindered by the lack of reliable preclinical models with optimized objective assessment of wound closure. Here, we describe an inexpensive and reproducible model of experimental full thickness burn wound reconstructed with an allogeneic skin graft. The wound is induced on the dorsum surface of anaesthetized inbred wild type mice from the BALB/C and SKH1-Hrhr backgrounds. The burn is produced using a brass template measuring 10 mm in diameter, which is preheated to 80 &#176;C and delivered at a constant pressure for 20 s. Burn eschar is excised 24 hours after the injury and replaced with a full thickness graft harvested from the tail of a genetically similar donor mouse. No specialized equipment is required for the procedure and surgical techniques are straightforward to follow. The method may be effortlessly implemented and reproduced in most research settings. Certain limitations are associated with the model. Due to technical difficulties, the harvest of thinner split thickness skin grafts is not possible. The surgical method we describe here allows for the reconstruction of burn wounds using full thickness skin grafts. It may be used to carry out preclinical therapeutic testing.

The aim of this study was to develop a murine model of burn wound healing. A thermal burn was induced on the dorsal skin of mice using a preheated brass template. Burned tissue was debrided and overlaid with a skin graft harvested from the tail of a genetically similar donor mouse.

Trivial superficial wounds heal without complications by primary intention. Deep wounds, such as full thickness burns, heal by secondary intention and ...