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).

참고 : 모든 절차가 남부 캘리포니아 대학의 기관 동물 관리 및 사용위원회 (IACUC)의 승인 동물 프로토콜에 따라 있습니다. 수술 1. 준비 <ol><li> 필요한 수술 도구 및 소모품의 레이아웃 (1 항목 살균 - 재료 목록 18)과 해부 현미경 및 후드 또는 하강 기류 테이블에 광섬유 트랜스 조명을 정렬합니다. </li><li> 마취를 유도하기 위해 4 % 이소 플루 란과 유도 챔버 (3 개월 - - 30 G, 1 ~ 20) 성적으로 성숙한 마우스를 놓습니다. </li><li> 발 및 / 또는 꼬리 핀치 테스트 마취를 확인합니다. </li><li> 수술 중 안구 건조를 방지하기 위해 마우스의 눈에 연고를 적용합니다. </li><li> 현미경 단계로 마우스를 이동합니다. 유지 보수 호스의 코 콘에 마우스를 놓고 옆에 위치. 체온을 유지하는 데 도움 동물에 인접한 온난화 파우치를 제공합니다. 하지E :. 마우스의 위치는 개인 취향 및 / 또는 외과 의사의 손 방향에 따라 달라집니다, 왼쪽 또는 오른쪽 중 어느 쪽이 괜찮습니다. </li><li> 유지 보수를 위해 3 %, 그리고 속도를 호흡 시계 - 2 이소 플루 란을 조정합니다. </li><li> 25 G 바늘 수술 후 통증 부 프레 노르 핀 (0.05 ㎎ / ㎏) 피하 (다리의 측면 측면)를 주입한다. 다리는 정상 응답 인 트 수 있습니다. </li></ol> 2. 열기 절개 <ol><li> 촉진하여 원하는 리브를 감지하고, 면도기로 머리의 영역을 취소합니다. 참고 : 갈비뼈 1-7 폐에 근접하고 11-13 상당히 짧은 때문에 갈비 8-10 권장합니다. 갈비뼈를 계산하는 숫자 1과 가장 주동이의 리브와 함께 시작해야한다. </li><li> (3 배는 종종 동물 사용 프로토콜에 의해 권장) 포비돈 - 요오드 및 이소프로판올 (70 %) 보라고 번갈아 영역을 준비합니다. </li><li> 바로 위에, 피부를 통해 2cm 절개를 잘라 중간 크기의 미세 가위로 원하는 리브 평행. Incis기본 근육과 지방 레이어를 통해 전자. </li><li> 절개의 크기를 최소화하면서 수술 영역을 노출 트랙터에 3 층 (피부, 근육, 지방)을 놓습니다. </li></ol> 3. 리브를 절개 <ol><li> 5.0 mm 메스와 리브의 원하는 부분을 덮는 늑간 근육을 잘라. 리브와 같은 높은 곡선 아닌 경우이 같은 chondrocostal 관절 위치 5mm 근위 약이 절개입니다. 조심스럽게 좋은 팁 집게로 뼈에서 근육을 분리합니다. </li><li> 동물의 골막을 유지하면서 절제를 만들려면 5.0 mm 메스와 리브의 길이를 따라, 골막을 통해 잘라. 조심스럽게 좋은 팁 집게 옆으로 기본 뼈에서 골막을 분리합니다. 골막은 매우 섬세하고 젤라틴 일관성을 가지고 있으므로주의를 진행합니다. </li><li> 다음에 미세한 마이크로 가위 일단 뼈 통한 단면을 만든다. 필요한 경우와 절제를 측정현미경 또는 통치자 형 게이지 레티클. 그런 다음 조심스럽게 골막에서 뼈를 들어 올려 다른 쪽 끝을 잘라. 참고 :주의를 진행합니다. 이 흉막 막을 찢고 기흉 발생합니다 아래 리브를 절개하지 않고 잡아 당겨 같은 가장 섬세한 작업이다. 흉막 막이 찢어진 경우 흉강의 내부가 보이는 될 것 같은이 쉽게 알 수있을 것입니다. </li><li> 출혈을 멈추게하기 위해 5 초 - 출혈이 뼈를 절단의 결과로 발생하는 경우, 4 면봉을 잘라 끝에서 압력을 적용합니다. 참고 : 드문 경우에 출혈이 계속, 그것은 중지하고 동물 회복이 손상 될 상당한 혈액 손실을 안락사 할 필요가있다 (유체 교체없이, 안전하게 손실 될 수있는 최대 혈액량은 전체 혈액량의 10 % 7.7 -.) 200 μL 9 - 8 μL / g는 25g 마우스의 경우, 이것은 180에 해당합니다. </li><li> 즉시로 제거 리브를 배치향후 분석을 위해 4 % PFA. 주 : 뼈와 골막 모두 절제 행하는 것이라면, 골막 절개 박리 (단계 3.2)을 생략합니다. 골막은 매우 밀접하게 흉막 막에 부착되어 있기 때문에 눈물을 방지하기 위해 집게와 떨어져 흉막 막에서 리브를 괴롭 히고, 조심스럽게 진행합니다. </li></ol> 4. 절개 닫기 <ol><li> 9-0 봉합 (2 봉합은 일반적으로 충분하다)와 나머지 골막 슬리브의 맨 위에 늑간 근육을 봉합. 수술 위치에 대한 지표 역할을 직접 리브의 절단 끝 위의 봉합을 놓습니다. </li><li> 견인기를 제거합니다. (- 4 봉합 충분하다 3) 9-0 봉합과 상부의 근육과 지방을 봉합. 근육 및 / 또는 지방 특히 두꺼운 층 쥐에서 별도로 각 레이어를 봉합 (예를 들어, 근육 층의 봉합의 1 층과 지방층을위한 봉합의 1 층). </li><li> 닫기(- 5 봉합 충분하다 4) 7-0 봉합사로 피부. </li><li> 큰 집게와 함께 가장자리를 곤란, 봉합 접착제 절개를 고정합니다. </li><li> 천천히 처음 몇 분 동안 1 %로 조정하여 오프 이소 플루 란 마우스를 탈피하고 전원을 껐다 켜십시오. </li><li> 가열 램프 아래에 마우스를 놓고 의식이, 5 회복 될 때까지 남겨 - 10 분. 의식이 회복 된 후 치유 기간 동안, 마우스를 이동하고 일반적으로 ambulate과 고통의 흔적을 보여주지해야한다. </li></ol> 5. 복구 및 분석 <ol><li> 0.5 MG에서 경구 용 젤라틴 형태의 부 프레 노르 핀의 관리를 포함하여 수술 후 통증 관리를 제공 / 48 시간 동안 매 12 시간을 kg. 경구 투여는 흉부 지역에 통증이 발생할 수 있습니다 동물을 억제 할 필요가 없습니다. </li><li> 치료 기간 동안 음식과 물을 무료로 액세스 할 수있는 케이지에 마우스를 유지한다. 절개 부위가 건조 후, 여성 동물 남성 동안 공동 수용 할 수있다의 싸움을 방지하기 위해 절연되어 있어야합니다. </li><li> 치유 기간이 지나면 안락사에 대한 승인 절차를 따르십시오. 고정 및 분석을 위해 갈비뼈를 제거합니다. </li></ol>

<ol>
	<li>Philip, S. J., Kumar, R. J., Menon, K. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Morphological+study+of+rib+regeneration+following+costectomy+in+adolescent+idiopathic+scoliosis.">Morphological study of rib regeneration following costectomy in adolescent idiopathic scoliosis.</a> Eur Spine J. 14 (8), 772-776 (2005).</li><li>Munro, I. R., Guyuron, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Split-rib+cranioplasty.">Split-rib cranioplasty.</a> Ann Plast Surg. 7 (5), 341-346 (1981).</li><li>Taggard, D. A., Menezes, A. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Successful+use+of+rib+grafts+for+cranioplasty+in+children.+Pediatric+neurosurgery.">Successful use of rib grafts for cranioplasty in children. Pediatric neurosurgery.</a> 34 (3), 149-155 (2001).</li><li>Head, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prevention+of+Regeneration+fo+the+Ribs:+A+problem+in+thoracic+surgery.">Prevention of Regeneration fo the Ribs: A problem in thoracic surgery.</a> Archives of Surgery. 14 (6), 1215-1221 (1927).</li><li>Kawanabe, Y., Nagata, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+method+of+costal+cartilage+harvest+for+total+auricular+reconstruction:+part+I.+Avoidance+and+prevention+of+intraoperative+and+postoperative+complications+and+problems.">A new method of costal cartilage harvest for total auricular reconstruction: part I. Avoidance and prevention of intraoperative and postoperative complications and problems.</a> Plastic and reconstructive surgery. 117 (6), 2011-2018 (2006).</li><li>Cheung, K. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+externally+fixed+femoral+fracture+model+for+mice.">An externally fixed femoral fracture model for mice.</a> Journal of orthopaedic research : official publication of the Orthopaedic Research Society. 21 (4), 685-690 (2003).</li><li>Matthys, R., Perren, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Internal+fixator+for+use+in+the+mouse.">Internal fixator for use in the mouse.</a> Injury. 40, S103-S109 (2009).</li><li>Cooper, G. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Testing+the+critical+size+in+calvarial+bone+defects:+revisiting+the+concept+of+a+critical-size+defect.">Testing the critical size in calvarial bone defects: revisiting the concept of a critical-size defect.</a> Plastic and reconstructive surgery. 125 (6), 1685-1692 (2010).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ask+the+Vet.">Ask the Vet.</a> JAX NOTES. 499, (2005).</li><li>Flecknell, P. A., Roughan, J. V., Stewart, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+oral+buprenorphine+('buprenorphine+jello')+for+postoperative+analgesia+in+rats--a+clinical+trial.">Use of oral buprenorphine ('buprenorphine jello') for postoperative analgesia in rats--a clinical trial.</a> Laboratory animals. 33 (2), 169-174 (1999).</li><li>Rigueur, D., Lyons, K. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Whole-mount+skeletal+staining.">Whole-mount skeletal staining.</a> Methods in molecular biology. 1130, 113-121 (2014).</li><li>Evans, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contribution+of+somitic+cells+to+the+avian+ribs.">Contribution of somitic cells to the avian ribs.</a> Developmental biology. 256 (1), 114-126 (2003).</li><li>Colnot, C., Thompson, Z., Miclau, T., Werb, Z., 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=Altered+fracture+repair+in+the+absence+of+MMP9.">Altered fracture repair in the absence of MMP9.</a> Development. 130 (17), 4123-4133 (2003).</li><li>Lu, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cellular+basis+for+age-related+changes+in+fracture+repair.">Cellular basis for age-related changes in fracture repair.</a> Journal of orthopaedic research : official publication of the Orthopaedic Research Society. 23 (6), 1300-1307 (2005).</li><li>Zilberman, Y., Gafni, Y., Pelled, G., Gazit, Z., Gazit, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioluminescent+imaging+in+bone.">Bioluminescent imaging in bone.</a> Methods in molecular biology. 455, 261-272 (2008).</li><li>Pelled, G., Gazit, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Imaging+using+osteocalcin-luciferase.">Imaging using osteocalcin-luciferase.</a> Journal of musculoskeletal. 4 (4), 362-363 (2004).</li><li>Elefteriou, F., Yang, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+mouse+models+for+bone+studies--strengths+and+limitations.">Genetic mouse models for bone studies--strengths and limitations.</a> Bone. 49 (6), 1242-1254 (2011).</li><li>Srour, M. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Natural+large-scale+regeneration+of+rib+cartilage+in+a+mouse.">Natural large-scale regeneration of rib cartilage in a mouse.</a> J. Bone Miner. , (2014).</li></ol>

The authors have nothing to disclose

처음이 프로토콜을 학습 할 때, 어디에서 문제가 될 수있는 초기 절개를 찾을 수 결정. 그러나 안락사 마우스에 연습 외과 의사가 초기 절개를 배치하고 절제 할 원하는 리브 노출 위치를 배울 수 있습니다. 시체에서 작업도 또는 골막없이 리브 부분을 제거하는 데 필요한 미세 운동 능력을 향상시킨다. 또한,이 절차에 새로운 사람이 훌륭한 도구와 얇은 봉합을 조작하는 것은 어려운 할 수없는 경...

골격 손상, 만성 관절염을 쇠약하게하고, 재건 수술과 관련된 심각한 문제는 경제 생산성, 가족 복지, 삶의 질에 영향을. 작은 휴식과 병변이 상당히 잘 치유 할 수 있지만, 인간의 구조와 기능을 복원 재건 절차에 의존해야하므로 큰 결함을 수리하고 할 수 없습니다. 재건은 동종 또는 heterogeneic 이식, morcellized 뼈 이식 비계, 또는 사지 연장술을 포함 할 수있다. 불행하게도,뿐만 아니라 이러한 치료하지만 수리 뼈의 원래 강도가 거의 달성하지 않습니다와 관련된들이 지속적인 병적 요인이있다. 따라서, 새로운 임상 적 접근 방법이 필요하다. 분절 결함을 치료하는 혁신적인 방법을 개발 일방향 대규모 수리 자연스럽게 발생하는 상황을 연구하는 것이다. 포유 동물이 일에 제한 간주하는 동안 양서류는 유명 골격 요소를 재생성 할 수 있습니다능력이다. 그러나, 20 세기 초반 이후, 인간의 리브의 재생의보고는 적다 인간 그렇게 1-4 한정되지 않을 수 있음을 제안 발표되었다. 현재 이러한 현상은 제일 턱, 얼굴 및 귀 재건 리브 재료를 사용하여 성형 외과 의사에 의해 공지되어 있지만, 5보다 폭넓게 이해되지 않는다. 더 자세하게 수리를 연구하기 위해, 우리는 마우스를 사용하여 외과 적 모델을 개발 하였다. 이러한 프로토콜을 사용하여, 연구자들은 관련된 선천적 요인을 파악하고 다른 위치에서 골격 치유를 촉진하기 위해이 정보를 사용할 수있다. 수리 골격을 연구 모델로서 리브를 사용하는 많은 장점이있다. (6,7 대퇴골의 절제에 비해) 우선, 주변 리브 천연 고정구를 제공한다. 이것은 내부 및 외고정의 이환율의 위험을 감소하고 수술을 단순화합니다. 가슴 워싱턴의 두 번째로, 얇은 근육 층쉽게 액세스 및 두개골 절제술 (8)의 편의에 대한 분석은 비교할 수 있도록 우수한 가시성을 제공 할 것이다. 셋째, intramembranous 골화, 연골 골화에 의한 리브 형태로 구성하고 중앙 골간의 양쪽 끝에있는 성장판의 확장을 통해 길이 성장 calvariae 대조적으로. 따라서, 리브의 수리는 사지 골격의 긴 뼈의 수리에 더 비교 될 수있다. 더욱이, 우리는 대퇴골에 비해, 리브의 골막가 두껍고보다 쉽게​​ 조작 할 수 있음을 발견 하였다. 따라서, 골막을 공부하거나 세포 치료, 약리 에이전트를 테스트 및 / 또는 조직 비계의 목적을 위해 뼈 수리를 분석하고자하는 연구자들은이 수술 모델이 유용 할 수 있습니다. 요약하면,이 리브 절제 모델은 일반적으로 사용하는 이러한 모델은 현재 존재하지 않기 때문에 포유 동물에서 자연 대규모 뼈 수리를 공부하는 내에서 컨텍스트를 제공합니다.

<table border="0" cellpadding="0" cellspacing="0" width="896">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Name of Reagent/ Equipment</td>
			<td style="width:160px;">Company</td>
			<td style="width:157px;">Catalog Number</td>
			<td style="width:304px;">Comments/Description</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:275px;">Medium sized micro-dissection scissors (Vannas-T&#252;bingen Spring Scissors 5 mm)</td>
			<td style="width:160px;">Fine Science Tools</td>
			<td>15003-08</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:275px;">Fine micro-dissection scissors (Vannas Spring Scissors - 2mm Cutting Edge)</td>
			<td style="width:160px;">Fine Science Tools</td>
			<td style="width:157px;">15000-04</td>
			<td>curved tip is beneficial</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Micro-scalpel 5.0 mm</td>
			<td style="width:160px;">Fine Science Tools</td>
			<td>10315-12</td>
			<td>other fine scalpels can be substituted</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Dumont 55 forceps</td>
			<td style="width:160px;">Fine Science Tools</td>
			<td style="width:157px;">11295-51</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Retractor&#160;</td>
			<td style="width:160px;">Fine Science Tools</td>
			<td>17004-05</td>
			<td>adjustability is convenient</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Micro-needle holders</td>
			<td style="width:160px;">Fine Science Tools</td>
			<td style="width:157px;">12060-01</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:275px;">9.0 nylon sutures (Ethilon), taper point best</td>
			<td style="width:160px;">Ethicon</td>
			<td>2819G or similar</td>
			<td>taper point best but reverse cutting is also good</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">7.0 prolene sutures (Prolene)</td>
			<td style="width:160px;">Ethicon</td>
			<td>8700H or similar</td>
			<td>6-0 can be used too, needle point can vary</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Large forceps (Adson Forceps)</td>
			<td style="width:160px;">Fine Science Tools</td>
			<td style="width:157px;">11006-12</td>
			<td>other brands are fine</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Lubricant Eye Ointment (Akwa Tears)</td>
			<td style="width:160px;">Akorn</td>
			<td style="width:157px;">17478-062-35</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:275px;">Suture glue (GLUture Topical Tissue Adhesive)</td>
			<td style="width:160px;">Abbot</td>
			<td style="width:157px;">32046-01</td>
			<td>has excellent working time</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Shaver</td>
			<td style="width:160px;">Wahl</td>
			<td>9918-6171 or similar</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Clamp lamp</td>
			<td>Zoo Med</td>
			<td style="width:157px;">LF-5</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Infrared Bulb, 75W</td>
			<td style="width:160px;">Zoo Med</td>
			<td>RS-75</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:275px;">RC2 Rodent Anesthesia System</td>
			<td style="width:160px;">VetEquip&#160;</td>
			<td style="width:157px;">922100</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">IsoFlo (Isoflurane)</td>
			<td style="width:160px;">Abbot</td>
			<td style="width:157px;">05260-05</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Buprenorphine (Buprenex)</td>
			<td style="width:160px;">Reckitt Benckiser</td>
			<td style="width:157px;">12496-0757-1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Betadine</td>
			<td style="width:160px;">Purdue Frederick</td>
			<td>67618015017</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Flavored Gelatin, raspberry</td>
			<td style="width:160px;">Jell-O</td>
			<td>B000E1FYL0</td>
			<td>made up firm, to the consistency of &#39;jigglers&#39;</td>
		</tr>
	</tbody>
</table>

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

이 프로토콜은 마우스 리브를 이용하여 대규모 뼈 수리에 대한 새로운 모델 연구원을 소개합니다. 주변 늑간근에서 원하는 리브 노출 흉추 원통 벽을 개방 기흉 유도없이 리브의 원하는 부분을 절개하고, 절개 폐쇄 수술에 대한 동물의 제조 : 절차는 다음과 같은 세부. 팔다리 뼈대의 뼈에 비해 매우 리브 접근 가능하다. 인접한 리브 자연 고정을 제공하기 때문에 또한, 더 내부 또는 외부 고정 장치는 필요하지 않습니다. 수술은 시판 공급 장치를 사용 배우기 간단하고, 동물에 의해 잘 용납입니다. 절차 또는 주변 골막을 제거하지 않고 수행 할 수 있기 때문에 보수 골막에의 기여가 평가 될 수있다. 2 개월 - 결과는 골막이 유지되는 경우, 강력한 수리 (1)에서 발생하는 것을 나타냅니다. 우리는 것이 프로토콜의 사용을 기대리브 수리에 대한 연구를 자극하고 결과는 몸 주위에 다른 위치에서 뼈 수리를 자극 할 수있는 새로운 방법의 개발을 촉진됩니다.

동물은 일반적으로 절개가 잘 치유,이 과정에서 신속하게 복구하고, 동물은 정상적인 동작을 나타낸다. 절차는 시판 공급 장치를 사용하고 실시 후, 1 시간에 따라 완성 될 수있다. 실험 데이터를 생성하는 절차를 사용하기 전에, 어떤 큰 뼈 조각이 절제 영역 내에 남아있을 수 있는지 확인 일 0 시점 절제술을 분석하는 것이 중요하다. 이것은 전형적으로 특별한 방사선 안전 승인을 필요로하더라?...

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.

마우스 리브를 절제하는 수술 : 대규모 긴 뼈 수리에 대한 모델

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

a-surgical-procedure-for-resecting-mouse-rib-model-for-large-scale

Research

JoVE Journal

Medicine

13.0K 조회수.  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.

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

A Novel Surgical Approach for Intratracheal Administration of Bioactive Agents in a Fetal Mouse Model

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and acquired diseases. Apart from congenital or inherited abnormalities with the requirement for long-term expression of the delivered gene, several non-inherited perinatal conditions, where short-term gene expression or pharmacological intervention is sufficient to achieve therapeutic effects, are considered as potential future indications for this kind of approach. Candidate diseases for the application of short-term prenatal therapy could be the transient neonatal deficiency of surfactant protein B causing neonatal respiratory distress syndrome1,2 or hyperoxic injuries of the neonatal lung3. Candidate diseases for permanent therapeutic correction are Cystic Fibrosis (CF)4, genetic variants of surfactant deficiencies5 and &alpha;1-antitrypsin deficiency6.
Generally, an important advantage of prenatal gene therapy is the ability to start therapeutic intervention early in development, at or even prior to clinical manifestations in the patient, thus preventing irreparable damage to the individual. In addition, fetal organs have an increased cell proliferation rate as compared to adult organs, which could allow a more efficient gene or stem cell transfer into the fetus. Furthermore, in utero gene delivery is performed when the individual's immune system is not completely mature. Therefore, transplantation of heterologous cells or supplementation of a non-functional or absent protein with a correct version should not cause immune sensitization to the cell, vector or transgene product, which has recently been proven to be the case with both cellular and genetic therapies7.
In the present study, we investigated the potential to directly target the fetal trachea in a mouse model. This procedure is in use in larger animal models such as rabbits and sheep8, and even in a clinical setting9, but has to date not been performed before in a mouse model. When studying the potential of fetal gene therapy for genetic diseases such as CF, the mouse model is very useful as a first proof-of-concept because of the wide availability of different transgenic mouse strains, the well documented embryogenesis and fetal development, less stringent ethical regulations, short gestation and the large litter size.
Different access routes have been described to target the fetal rodent lung, including intra-amniotic injection10-12, (ultrasound-guided) intrapulmonary injection13,14 and intravenous administration into the yolk sac vessels15,16 or umbilical vein17. Our novel surgical procedure enables researchers to inject the agent of choice directly into the fetal mouse trachea which allows for a more efficient delivery to the airways than existing techniques18.

We developed a novel surgical approach for intratracheal administration of bioactive agents into the mouse fetus. The delivery route is more efficient in targeting the fetal mouse lungs than the commonly used intra-amniotic injection. This procedure has to date not been described in a mouse model.

태아 마우스 모델에서 Bioactive 에이전트의 Intratracheal 관리를위한 소설 외과 접근

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and ...

연구

의학

Surgical Technique for Spinal Cord Delivery of Therapies: Demonstration of Procedure in Gottingen Minipigs

This is a compact visual description of a combination of surgical technique and device for the delivery of (gene and cell) therapies into the spinal cord. While the technique is demonstrated in the animal, the procedure is FDA-approved and currently being used for stem cell transplantation into the spinal cords of patients with ALS. While the FDA has recognized proof-of-principle data on therapeutic efficacy in highly characterized rodent models, the use of large animals is considered critical for validating the combination of a surgical procedure, a device, and the safety of a final therapy for human use. The size, anatomy, and general vulnerability of the spine and spinal cord of the swine are recognized to better model the human. Moreover, the surgical process of exposing and manipulating the spinal cord as well as closing the wound in the pig is virtually indistinguishable from the human. We believe that the healthy pig model represents a critical first step in the study of procedural safety.

Short visual description of the surgical technique and device used for the delivery of (gene and cell) therapies into the spinal cord. The technique is demonstrated in the animal but is entirely translatable and currently being used for human application.

치료의 척추 배달을위한 수술 기법 : 괴팅겐 Minipigs의 절차의 데모

This is a  compact visual description of a combination of surgical technique and device  for the  delivery of (gene and cell) therapies into the spinal ...

A Mouse Fetal Skin Model of Scarless Wound Repair

Early in utero, but not in postnatal life, cutaneous wounds undergo regeneration and heal without formation of a scar. Scarless fetal wound healing occurs across species but is age dependent. The transition from a scarless to scarring phenotype occurs in the third trimester of pregnancy in humans and around embryonic day 18 (E18) in mice. However, this varies with the size of the wound with larger defects generating a scar at an earlier gestational age. The emergence of lineage tracing and other genetic tools in the mouse has opened promising new avenues for investigation of fetal scarless wound healing. However, given the inherently high rates of morbidity and premature uterine contraction associated with fetal surgery, investigations of fetal scarless wound healing in vivo require a precise and reproducible surgical model. Here we detail a reliable model of fetal scarless wound healing in the dorsum of E16.5 (scarless) and E18.5 (scarring) mouse embryos.

During mammalian development, early gestational skin wounds heal without a scar. Here we detail a reliable and reproducible model of fetal scarless wound healing in the cutaneous dorsum of E16.5 (scarless) and E18.5 (scarring) mouse embryos.

흉터 상처 수리의 마우스 태아의 피부 모델

Early in utero, but not in postnatal life, cutaneous wounds undergo regeneration and heal without formation of a scar. Scarless fetal wound healing occurs ...

The Mouse Round-window Approach for Ototoxic Agent Delivery: A Rapid and Reliable Technique for Inducing Cochlear Cell Degeneration

Investigators have utilized a wide array of animal models and investigative techniques to study the mammalian auditory system. Much of the basic research involving the cochlea and its associated neural pathways entails exposure of model cochleae to a variety of ototoxic agents. This allows investigators to study the effects of targeted damage to cochlear structures, and in some cases, the self-repair or regeneration of those structures. Various techniques exist for delivery of ototoxic agents to the cochlea. When selecting a particular technique, investigators must consider a number of factors, including the induction of inadvertent systemic toxicity, the amount of cochlear damage produced by the surgical procedure itself, the type of lesion desired, animal survivability, and reproducibility/reliability of results. Currently established techniques include parenteral injection, intra-peritoneal injection, trans-tympanic injection, endolymphatic sac injection, and cochleostomy with perilymphatic perfusion. Each of these methods has been successfully utilized and is well described in the literature; yet, each has various shortcomings. Here, we present a technique for topical application of ototoxic agents directly to the round window niche. This technique is non-invasive to inner ear structures, produces rapid onset of reliably targeted lesions, avoids systemic toxicity, and allows for an intra-animal control (the contra-lateral ear). Results stemming from this approach have helped deeper understanding of auditory pathophysiology, cochlear cell degeneration, and regenerative capacity in response to an acute injury. Future investigations may use this method to conduct interventional studies involving gene therapy and stem cell transplantation to combat hearing loss.

Various methods exist for introducing ototoxic agents to the cochleae of animal models. Presented is a surgical protocol for delivery of ototoxic agents to the round window niche. The procedure is reliable, creates targeted intra-cochlear lesions, and avoids mechanical damage to the microarchitecture. Examination of cochlear self-repair/regeneration is possible.

인공 와우 세포 변성을 유도하기위한 신속하고 신뢰할 수있는 기술 : Ototoxic 에이전트 배달을 위해 마우스 라운드 창 접근

Investigators have utilized a wide array of animal models and investigative techniques to study the mammalian auditory system. Much of the basic research ...

A Novel Murine Model of Arteriovenous Fistula Failure: The Surgical Procedure in Detail

The arteriovenous fistula (AVF) still suffers from a high number of failures caused by insufficient remodeling and intimal hyperplasia from which the exact pathophysiology remains unknown. In order to unravel the pathophysiology a murine model of AVF-failure was developed in which the configuration of the anastomosis resembles the preferred situation in the clinical setting. A model was described in which an AVF is created by connecting the venous end of the branch of the external jugular vein to the side of the common carotid artery using interrupted sutures. At a histological level, we observed progressive stenotic intimal lesions in the venous outflow tract that is also seen in failed human AVFs. Although this procedure can be technically challenging due to the small dimensions of the animal, we were able to achieve a surgical success rate of 97% after sufficient training. The key advantage of a murine model is the availability of transgenic animals. In view of the different proposed mechanisms that are responsible for AVF failure, disabling genes that might play a role in vascular remodeling can help us to unravel the complex pathophysiology of AVF failure.

Here we present a murine model of arteriovenous fistula (AVF) failure in which a clinically relevant anastomotic configuration is incorporated. This model can be used to study the pathophysiology and to test possible therapeutic interventions.

동정맥루 실패의 소설 쥐 모델 : 세부 수술

The arteriovenous fistula (AVF) still suffers from a high number of failures caused by insufficient remodeling and intimal hyperplasia from which the ...

Surgical Angiogenesis in Porcine Tibial Allotransplantation: A New Large Animal Bone Vascularized Composite Allotransplantation Model

Segmental bone loss resulting from trauma, infection malignancy and congenital anomaly remains a major reconstructive challenge. Current therapeutic options have significant risk of failure and substantial morbidity.
Use of bone vascularized composite allotransplantation (VCA) would offer both a close match of resected bone size and shape and the healing and remodeling potential of living bone. At present, life-long drug immunosuppression (IS) is required. Organ toxicity, opportunistic infection and neoplasm risks are of concern to treat such non-lethal indications.
We have previously demonstrated that bone and joint VCA viability may be maintained in rats and rabbits without the need of long-term-immunosuppression by implantation of recipient derived vessels within the VCA. It generates an autogenous, neoangiogenic circulation with measurable flow and active bone remodeling, requiring only 2 weeks of IS. As small animals differ from man substantially in anatomy, bone physiology and immunology, we have developed a porcine bone VCA model to evaluate this technique before clinical application is undertaken. Miniature swine are currently widely used for allotransplantation research, given their immunologic, anatomic, physiologic and size similarities to man. Here, we describe a new porcine orthotopic tibial bone VCA model to test the role of autogenous surgical angiogenesis to maintain VCA viability.
The model reconstructs segmental tibial bone defects using size- and shape-matched allogeneic tibial bone segments, transplanted across a major swine leukocyte antigen (SLA) mismatch in Yucatan miniature swine. Nutrient vessel repair and implantation of recipient derived autogenous vessels into the medullary canal of allogeneic tibial bone segments is performed in combination with simultaneous short-term IS. This permits a neoangiogenic autogenous circulation to develop from the implanted tissue, maintaining flow through the allogeneic nutrient vessels for a short time. Once established, the new autogenous circulation maintains bone viability following cessation of drug therapy and subsequent nutrient vessel thrombosis.

Currently any kind of vascularized composite allotransplantation depends on long-term-immunosuppression, difficult to support for non-life-critical indications. We present a new porcine tibial VCA model that can be used to study bone VCA and demonstrate the use of surgical angiogenesis to maintain bone viability without the need of long-term immune-modulation.

돼지 Tibial Allotransplantation 외과 신생: 새로운 큰 동물 뼈 나가도록 복합 Allotransplantation 모델

Segmental bone loss resulting from trauma, infection malignancy and congenital anomaly remains a major reconstructive challenge. Current therapeutic ...

Evaluating the Procedure for Performing Awake Cystometry in a Mouse Model

Awake filling cystometry has been used for a long time to evaluate bladder function in freely moving mice, however, the specific methods used, vary among laboratories. The goal of this study was to describe the microsurgical procedure used to implant an intravesical tube and the experimental technique for recording urinary bladder pressure in an awake, freely moving mouse. In addition, experimental data is presented to show how surgery, as well as tubing type and size, affect lower urinary tract function and recording sensitivity. The effect of tube diameter on pressure recording was assessed in both polyethylene and polyurethane tubing with different internal diameters. Subsequently, the best performing tube from both materials was surgically implanted into the dome of the urinary bladder of male C57BL/6 mice. Twelve-hour, overnight micturition frequency was recorded in healthy, intact animals and animals 2, 3, 5, and 7 days post-surgery. At harvest, bladders were assessed for signs of swelling using gross observation and were subsequently processed for pathological analysis. The greatest extent of bladder swelling was observed on day 2 and 3, which correlated with behavioral voiding data showing significantly impaired bladder function. By day 5, bladder histology and voiding frequency had normalized. Based on the literature and evidence provided by our studies, we propose the following steps for in vivo recording of intravesical pressure and voided volume in an awake mouse: 1) Perform the surgery using an operating microscope and microsurgical tools, 2) Use polyethylene-10 tubing to minimize movement artifacts, and 3) Perform cystometry on post-operative day 5, when bladder swelling resolves.

This study describes the surgical procedures and experimental techniques for performing awake cystometry in a freely moving mouse. In addition, it provides experimental evidence to support its optimization and standardization.

마우스 모델에서 깨어나는 방광 측정을 수행하기위한 절차 평가

Awake filling cystometry has been used for a long time to evaluate bladder function in freely moving mice, however, the specific methods used, vary among ...

A Surgical Procedure for the Administration of Drugs to the Inner Ear in a Non-Human Primate Common Marmoset (Callithrix jacchus)

Hearing research has long been facilitated by rodent models, although in some diseases, human symptoms cannot be recapitulated. The common marmoset (Callithrix jacchus) is a small, easy-to-handle New World monkey which has a similar anatomy of the temporal bone, including the middle ear ossicular chains and inner ear to humans, than in comparison with that of rodents. Here, we report a reproducible, safe, and rational surgical approach to the cochlear round window niche for the drug delivery to the inner ear of the common marmoset. We adopted posterior tympanotomy, a procedure used clinically in human surgery, to avoid manipulation of the tympanic membrane that may cause conductive hearing loss. This surgical procedure did not lead to any significant hearing loss. This approach was possible due to the large bulla structure of the common marmoset, although the lateral semicircular canal and vertical portion of the facial nerve should be carefully considered. This surgical method allows us to perform the safe and accurate administration of drugs without hearing loss, which is of great importance in obtaining pre-clinical proof of concept for translational research.

A Surgical Procedure for the Administration of Drugs to the Inner Ear in a Non-Human Primate Common Marmoset Callithrix jacchus

We report a surgical method to administrate drugs to the inner ear of a non-human primate, the common marmoset (Callithrix jacchus), via the round window membrane.

비 인간 영장류 일반 비단 (Callithrix jacchus)에서 내가 의약품의 관리에 대 한 수술

Hearing research has long been facilitated by rodent models, although in some diseases, human symptoms cannot be recapitulated. The common marmoset ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

Myography 와이어를 사용 하 여 마우스 Mesenteric 동맥의 isometric 수축 측정

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Standardized Histomorphometric Evaluation of Osteoarthritis in a Surgical Mouse Model

One of the most prevalent joint disorders in the United States, osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage, primarily in the hip and knee joints, which results in significant impacts on patient mobility and quality of life. To date, there are no existing curative therapies for OA able to slow down or inhibit cartilage degeneration. Presently, there is an extensive body of ongoing research to understand OA pathology and discover novel therapeutic approaches or agents that can efficiently slow down, stop, or even reverse OA. Thus, it is crucial to have a quantitative and reproducible approach to accurately evaluate OA-associated pathological changes in the joint cartilage, synovium, and subchondral bone. Currently, OA severity and progression are primarily assessed using the Osteoarthritis Research Society International (OARSI) or Mankin scoring systems. In spite of the importance of these scoring systems, they are semiquantitative and can be influenced by user subjectivity. More importantly, they fail to accurately evaluate subtle, yet important, changes in the cartilage during the early disease states or early treatment phases. The protocol we describe here uses a computerized and semiautomated histomorphometric software system to establish a standardized, rigorous, and reproducible quantitative methodology for the evaluation of joint changes in OA. This protocol presents a powerful addition to the existing systems and allows for more efficient detection of pathological changes in the joint.

The current protocol establishes a rigorous and reproducible method for quantification of morphological joint changes that accompany osteoarthritis. Application of this protocol can be valuable in monitoring disease progression and evaluating therapeutic interventions in osteoarthritis.

외과 마우스 모형에 있는 골관절염의 표준화한 조직 형측정 평가

One of the most prevalent joint disorders in the United States, osteoarthritis (OA) is characterized by progressive degeneration of articular cartilage, ...