이 작품은 (HPL에) NIH 보조금 R01 GM087609, (MTL에) (HPL에) 안토니 슈, NIH 보조금 U01의 HL099776의 명예에 잉그리드 라이 빌 슈에서 선물, Hagey 연구소에 대한로부터 연구비 일부 지원 소아 재생 의학 및 (MTL과 HPL에) 오크 재단. GGW 의학의 스탠포드 대학, 스탠포드 의료 과학자 훈련 프로그램에 의해 지원되며, NIGMS 훈련 보조금 GM07365했다. MSH는 CIRM 임상 연구원 교육 그랜트 TG2-01159에 의해 지원되었다. WXH가 노프 심장 혈관 재단 기금에 의해 지원되었다.

참고 :이 문서에 설명 된 모든 절차는 실험 동물 관리에 스탠포드 행정 패널 (APLAC)에 의해 설립 지침에 따라 수행된다. 1. 시간 제한 임신 - 과배란 기술 (그림 1) 참고 : 정확하게 E16.5 및 E18.5에서 태아 수술을 위해 마우스 배아의 임신 기간 타이밍은 매우 중요합니다. 과배란을 유도하기 위해 임신 한 암컷 혈청 (PMS) 및 인간 융모 성 성선 자극 호르몬 (HCG) 주사를 사용하여 마우스 임신 타이밍에 대한 우리의 프로토콜이 절 우리의 세부 사항. <ol><li> 1:00 하루 1 3:00 PM 사이에 100 μl의 PBS의 볼륨 PMS의 3.0-5.0 국제 단위 (IU)와 여성 마우스 (케이지 당 &lt;5) 복강 내 (IP)를 주입한다. </li><li> 12:00 3 일 (사십 오 PMS 주입 후 시간 47) 2:00시 사이에, 100 μl의 PBS의 볼륨 HCG의 3.0-5.0 IU와 여성 쥐 IP를 주입. 참고 : HCG 주입 유도한다<html> 배란 약 12​​ 시간 후 분사. </li><li> 즉시 HCG 주사 다음 8-16 주령의 수컷과 암컷을 짝짓기. 참고 : 우리는 일반적으로 개별 남성의 케이지에 두 여성을 배치합니다. </li><li> 태아 연령 E0.5으로 - (10:00 AM 6:00) 기록 4 일의 아침에 남성에서 분리 된 여성. 참고 : 질 플러그가이 시간에 확인할 수 있습니다; 그러나, 질 플러그의 관찰은 임신을 보장하지 않으며 어떠한 플러그가 관찰되지 않을 때 여성은 임신이 될 수 있습니다. 임신 4 일의 아침에 질 플러그 검사, 임신 나이 E16.5 및 E18.5에서 육안 검사 및 / 또는 촉진에 의해 일반적으로 관찰 할 수 있음을 감안할 때 꼭 필요한 것은. 경험상, 상기 균주에 따라 슈퍼 배란 여성의 약 30 % ~ 50 %가 여기에 설명 된 기술을 사용하여 임신. </li></ol>PG &quot;/&gt; 그림 과배란 기법에 대한 도식. 마우스에서 과배란 초과 임신에 대한 회로도를 보여주는 절차. <a href="https://www.jove.com/files/ftp_upload/52297/52297fig1large.jpg" target="_blank">이 그림의 더 큰 버전을 보려면 여기를 클릭하십시오.</a> 2. 쥐과 태아 수술 E16.5 및 E18.5 배아에 (등쪽 부상) (그림 2) <ol><li> 절차 전에, 70 % 이소 프로필 알코올 수술실 및 장비의 모든 표면을 청소합니다. 또한,이를 오토 클레이브에 의해 절차에서 사용될 모든 수술 용품 및기구 소독. 일부 기관들은 뜨거운 비드 살균 이후의 사용을 허용 할 수 있습니다. 작업 당, 거즈 및 수술 도구를 포함 무균 팩을 사용합니다. </li><li> 분당 1 L에 유지 보수 마취 다음에 분당 2 L에서 2.5 % 이소 플루 란 / 산소 혼합물에서 임신 한 어머니의 마취 (태아 연령 E16.5 또는 E18.5)를 유도한다. </li><li> 사기하려면회사 적절한 마취, 마우스의 깊은 페달 반사가 억제되어 있는지 확인하고 발생하기 쉬운 위치에 마우스를 놓습니다. </li><li> Puralube이 절차를 수행하는 동안 눈에 자극이나 건조 함을 방지하기 위해 같은 수의사 안과 연고를 적용합니다. </li><li> 더 이상 30 초 (그림 2A)을 위해 제모 크림의 빛 응용 프로그램을 투여하여 복부를 준비합니다. </li><li> 포비돈 - 요오드 알코올 (그림 2B 및 2C) 무균 수술을 위해 복부를 준비합니다. </li><li> 미세 수술 가위 (그림 2D 및 2E)를 사용하여 현미경으로 중간 선 개복술을 수행합니다. </li><li> 조심스럽게 수술 (그림 2 층)으로 선택 자궁과 태아를 노출합니다. </li><li> 뭉툭한 팁 바늘을 사용하여 따뜻한 (38 ° C) 인산염 완충 식염수 (PBS)와 수술 필드를 관개 참고 : 하나는 조심스럽게 큰 구멍 바늘의 끝 부분을 구부려서 할 수있다. (그림 2G 및 2H). </li><li> 허용하는 방식으로 태아의 위치를배부에 대한 전체 액세스 권한을에요. </li><li> 의도 지느러미 상처 (그림 2I)의 사이트를 덮는 자궁을 통해 7-0 나일론 봉합사를 사용하여 지갑 문자열 스티치를 전달합니다. 위치 지갑 왼쪽 또는 척수의 오른쪽에 배부의 영역을 통해 문자열, 큰 혈관이없는 자궁 벽의 영역에서. </li><li> 지갑 문자열 (그림 2J)의 중심에 자궁 벽과 양막을 통해 3mm의 절개를합니다. </li><li> 따뜻한 (38 ° C) PBS와 절개 사이트를 관개. </li><li> 미세 수술 가위를 사용하여 태아의 등쪽에서, 하나의 전층 상처 절제 길이 약 1mm 잘라. </li><li> 부드럽게 얼룩 절개 사이트는면 팁 어플리케이터를 건조. </li><li> 상처의 위치 (그림 2K)를 표시하기 위해 상처 부위에 3 μL 볼륨 인도 잉크 피하 주입한다. </li><li> 잉크가 상처 사이트 내에서 유지 된 보장하기 위해 따뜻한 (38 ° C) PBS로 세척하십시오. </li><li> 수술 보조 난 되세요nject 지갑 문자열로 양막에 무딘 팁 10 G 주사기를 통해 따뜻한 (38 ° C) PBS (그림 2L)를 폐쇄한다. 지갑 문자열 폐쇄 완료 (그림 2M)에 가까워지면서 주사기를 집어 넣으십시오. </li><li> 조심스럽게 복강 (그림 2N)로 자궁을 반환합니다. </li><li> 에버트 피부와 복막. </li><li> 따뜻한 (38 ° C) PBS로 복강 수술 보조 관개했다. </li><li> 피부와 복막을 스테이플 링으로 빠르게 닫기 복부 (그림 2O)을 마감했다. 표준 클로저는 두 층에서 수행된다; 복막과 제 2 층에서의 하나의 층, 피하 조직 및 피부 복근. 즉시 희생과 태아의 수확을 위해, 우리의 시위는 한 층에 폐쇄를 보여줍니다. </li><li> 30 분 동안이나 동물이 흉골 드러 누움을 유지하기에 충분한 의식을 회복 할 때까지 37 ° C로 설정 따뜻한 인큐베이터에서 관찰에서 동물을 놓습니다. </li><li> 마지막 ret하지 마십시오완전히 절차에서 회복 될 때까지 다른 동물의 회사에 동물을 항아리. </li><li> 마취에서 그 이후 48 시간 동안 각성시, 부 프레 노르 핀의 피하 주사 (0.05 ㎎ / ㎏) 통증 평가에 따라 필요에 따라 진통제마다 12 시간을 관리 할 수​​ 있습니다. 필요에 따라 추가 수술 후 통증 완화를 위해 피하 주사를 통해 카프로 펜 (5 ㎎ / ㎏)을 관리합니다. </li><li> 케이지에 동물을 반환하고 물과식이로 제공합니다. </li><li> 통증의 양상 면밀히 모니터링합니다. </li><li> 48 시간의 수술 후는, 이소 플루 란과 수확 부상 태아의 과다 복용으로 임신 한 어머니를 희생. 이를 위해, 5 % 이상으로 이소 플루 란 농도를 조정하고 호흡의 중단 후 1 분 동안 노출을 유지한다. 자궁 경부 전위와 안락사를 확인합니다. 연령대 제어를위한 unwounded 배아를 수확. 늦은 배아는 같은 IACUC의 권고와 일치 안락사의 별도의 방법이 있어야 드인두세, 자궁 전위, 또는 화학 물질 주입. </li></ol><img alt="그림 2" src="/files/ftp_upload/52297/52297fig2highres.jpg" /> 그림 2. 회로도 쥐과 태아 외과. E16.5 및 E18.5 마우스 배아에 상처를 입는 등의 일반 단계를 반복합니다. (A) 마우스 복부의 탈모. (B 및 C) 마우스 복부의 준비. (D) 현미경 수술에 사용 자궁을 통해 지갑 문자열 봉합의 과정. (E) 중간 선 개복술. (F) 자궁의 노출. 뭉툭한 팁 바늘 (G) 작성. 따뜻한 식염수와 자궁의 (H) 관개. (I) 작성. (J) 절개 벽과 1mm 전체 두께 절제 상처 발생. (K) 인도 잉크의 피하 주사. (L과 M) 지갑 문자열 봉합의 폐쇄. (N과 O)복부의 폐쇄. <a href="https://www.jove.com/files/ftp_upload/52297/52297fig2large.jpg" target="_blank">이 그림의 더 큰 버전을 보려면 여기를 클릭하십시오.</a>

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The authors have nothing to disclose.

여기에 제시된 수술 프로토콜은 먼저 우리의 실험실 (10)에 의해 2006 년에 출판 된 태아 쥐의 흉터 치유 절제 모델에 대해 설명합니다. 절제 상처 (11)의 다른 모델의 확립 외에, 뮤린 태아 흉터, 창상 치유의 모델 (12, 13)뿐만 아니라 존재한다. 원숭이, 양, 토끼, 쥐, 쥐 태아 흉터 상처 치유의 조사는 14 ~ 17보고되었다. 그러나 마우스는 그들의 비교적 낮은 당 일일 케이...

태아 피부 상처 임신 1 늦게까지 신속하고 scarlessly 치유. 태아 흉터 상처 수리는 정상 조직의 구조와 기능의 재생이 특징입니다. 흉터 표현형에 흉터에서의 전환은 인간 및 마우스 2,3에 배아 일 18 (E18)의 주위에 임신의 세 번째 임신에서 발생합니다. 성인에 비해, 상처 복구 태아 급속한 상피화, 결합 조직의 증착, 및 섬유 아세포 이동을 특징으로한다. 많은 연구 초기 태아의 개발 기간 동안 흉터 상처 치유의 현상에 대한 가능한 설명을 제안했다. 염증은 성인 상처 수리의 기본 구성 요소입니다; 그러나, 태아의 상처는 급성 염증 (4)의 부족을 특징으로한다. 이 단계 동안 여부 태아 면역계의 작용 미성숙 결과는 불분명하다. 최근 연구 제안이 풍부, 매트의 차이urity 및 E18 태아의 피부 대 E15에서 비만 세포의 기능 이상 마우스 3 흉터 표현형으로의 전환에 대한 책임이 있습니다. 다른 연구는 속성과 태아와 성인 상처 대 식세포의 풍요 로움의 차이가 태아의 상처 수리 (5) 동안 정상 세포 외 기질 (ECM)의 개혁에 대한 책임이 있음을 가정하는. 태아 및 성인 개발시 환경 요인의 차이도 상처 복구에 영향을 미칠 수 있습니다. Longaker와 동료들은 태아의 상처 액이 6 유체 상처 성인 없음에 비해 히알루 론산 자극 활동의 높은 수준을 보유하고 있음을 보여 주었다. 따라서, 태아 상처 환경에서 히알루 론산, 세포 운동성 및 확산에 도움이 미세 환경을 촉진하는 글리코 사 미노 글리 칸, 높은 수준의 초기 태아의 개발 과정에서 볼 수있는 흉터의 표현형에 대한 책임을 할 수있다. 태아는 사실에 근거 포인트의 다른 라인L 상처 환경은 상대적으로 저산소증과 성장 인자 7 풍부한 무균 양수에 잠겨있다. 그러나, 확실한 대답은 섬유 성 수리에 흉터 재생으로의 전환을 트리거 배아 중 중요한 사건이나 요인에 제공 없습니다. 태아의 흉터 치유 메커니즘 책임을 이해하는 것은 정확하고 재현성 모델을 필요로한다. 여기 세부 E16.5 (흉터) 및 E18.5 (흉터) 마우스 배아의 배부 태아 흉터 상처 치유의 재현 모델. 또한,이 모델의 경미한 변화는 이러한 태아 피부 및 상처 8,9의 유전자 발현 분석과 같은 추가의 연구들을 수행하는데 이용 될 수있다. 정확하게 시간이 초과 임신이 태아의 흉터 상처 치유 모델의 성공적인 반복 설에 중요한 점을 감안, 우리 과배란 초과 임신에 대해서도 상세하게 우리의 프로토콜입니다.

<table><tbody><tr><td>7-O MONOSOF Suture</td><td>eSuture</td><td>SN-1647G</td><td></td></tr><tr><td>Surgical Forceps</td><td>Kent Scientific</td><td>INS650916</td><td></td></tr><tr><td>Micro-scissors</td><td>Kent Scientific</td><td>INS600127</td><td></td></tr><tr><td>Autoclip 9 mm</td><td>Texas Scientific Instruments</td><td>205060</td><td></td></tr><tr><td>Insulin Syringe</td><td>Thermo Fisher Scientific</td><td>22-272-382</td><td></td></tr><tr><td>Black Pigment</td><td>AIMS</td><td>242</td><td></td></tr><tr><td>BD Safety-Lok 3 ml Syringe</td><td>BD Biosciences</td><td>309596</td><td></td></tr><tr><td>Phosphate Buffered Saline</td><td>Life Technologies</td><td>10010-049</td><td></td></tr><tr><td>OPMI-MD Surgical Microscope</td><td>Carl Zeiss Surgical Inc</td><td></td><td></td></tr><tr><td>Pregnant Mares Serum (PMS)</td><td>Millipore</td><td>367222</td><td></td></tr><tr><td>Human Chorionic Gonadotropin (HCG)</td><td>Sigma-Aldrich</td><td>CG10</td><td></td></tr><tr><td>Povidone Iodine Prep Solution</td><td>Dynarex</td><td>1415</td><td></td></tr><tr><td>Nair (depilatory cream)</td><td>Church and Dwight Co.</td><td>22600267058</td><td></td></tr></tbody></table>

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.

조직 학적 분석을 위해, E16.5 및 E18.5 마우스 배아의 등 피부에 피부의 상처는 48 시간의 PFA 4 %에 고정 후 상처, 및 파라핀을 수확해야합니다. 형광 형질 전환 모델에서, 빛 간섭 단층 냉동 보존이 적합 할 수 있습니다. 세포 및 결합 조직 구조를 시각화하는데 이용 될 수있는 몇 가지 얼룩이있다. 헤 마톡 실린 및 에오신 파란색 핵 및 산성 구조 (즉., 세포질과 세포 외 콜라겐) 레드, 핑크, 오렌?...

Watch this Scientific Journal Video about A Mouse Fetal Skin Model of Scarless Wound Repair at JoVE.com

A Mouse Fetal Skin Model of Scarless Wound Repair

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 ...

a-mouse-fetal-skin-model-of-scarless-wound-repair

Research

JoVE Journal

Medicine

12.2K 조회수.  Stanford University School of Medicine. 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.

비디오: A Mouse Fetal Skin Model of Scarless Wound Repair

The Three-Dimensional Human Skin Reconstruct Model: a Tool to Study Normal  Skin and Melanoma Progression

Most in vitro studies in experimental skin biology have been done in 2-dimensional (2D) monocultures, while accumulating evidence suggests that cells behave differently when they are grown within a 3D extra-cellular matrix and also interact with other cells (1-5). Mouse models have been broadly utilized to study tissue morphogenesis in vivo. However mouse and human skin have significant differences in cellular architecture and physiology, which makes it difficult to extrapolate mouse studies to humans. Since melanocytes in mouse skin are mostly localized in hair follicles, they have distinct biological properties from those of humans, which locate primarily at the basal layer of the epidermis. The recent development of 3D human skin reconstruct models has enabled the field to investigate cell-matrix and cell-cell interactions between different cell types. The reconstructs consist of a "dermis" with fibroblasts embedded in a collagen I matrix, an "epidermis", which is comprised of stratified, differentiated keratinocytes and a functional basement membrane, which separates epidermis from dermis. Collagen provides scaffolding, nutrient delivery, and potential for cell-to-cell interaction. The 3D skin models incorporating melanocytic cells recapitulate natural features of melanocyte homeostasis and melanoma progression in human skin. As in vivo, melanocytes in reconstructed skin are localized at the basement membrane interspersed with basal layer keratinocytes. Melanoma cells exhibit the same characteristics reflecting the original tumor stage (RGP, VGP and metastatic melanoma cells) in vivo. Recently, dermal stem cells have been identified in the human dermis (6). These multi-potent stem cells can migrate to the epidermis and differentiate to melanocytes.

In this report, we describe the three-dimensional skin reconstruct model which mimics human skin in architecture and composition. Melanocyte physiology, melanoma progression and the fate of dermal stem cells have been investigated using the skin reconstruct model. The model is also useful as a preclinical tool for drug assessment.

3 차원의 인간 피부 모델을 재구성 : 도구는 일반 피부와 흑색증의 진행을 공부하기

Most in vitro studies in experimental skin biology have been done in 2-dimensional (2D) monocultures, while accumulating evidence suggests that cells ...

연구

생체공학

A Full Skin Defect Model to Evaluate Vascularization of Biomaterials In Vivo

Insufficient vascularization is considered to be one of the main factors limiting the clinical success of tissue-engineered constructs. In order to evaluate new strategies that aim at improving vascularization, reliable methods are required to make the in-growth of new blood vessels into bio-artificial scaffolds visible and quantify the results. Over the past couple of years, our group has introduced a full skin defect model that enables the direct visualization of blood vessels by transillumination and provides the possibility of quantification through digital segmentation. In this model, one surgically creates full skin defects in the back of mice and replaces them with the material tested. Molecules or cells of interest can also be incorporated in such materials to study their potential effect. After an observation time of one&#8217;s own choice, materials are explanted for evaluation. Bilateral wounds provide the possibility of making internal comparisons that minimize artifacts among individuals as well as of decreasing the number of animals needed for the study. In comparison to other approaches, our method offers a simple, reliable and cost effective analysis. We have implemented this model as a routine tool to perform high-resolution screening when testing vascularization of different biomaterials and bio-activation approaches.

A Full Skin Defect Model to Evaluate Vascularization of Biomaterials In Vivo

Vascularization is key to approaches in successful tissue engineering. Therefore, reliable technologies are required to evaluate the development of vascular networks in tissue-constructs. Here we present a simple and cost-effective method to visualize and quantify vascularization in vivo.

전체 피부 결함 모델은 생체의 혈관 신생을 평가하기<em&gt; 생체</em

Insufficient vascularization is considered to be one of the main factors limiting the clinical success of tissue-engineered constructs. In order to ...

Murine Model of Wound Healing

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

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

상처 치유의 생쥐 모델

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

의학

Generation of a Three-dimensional Full Thickness Skin Equivalent and Automated Wounding

In vitro models are a cost effective and ethical alternative to study cutaneous wound healing processes. Moreover, by using human cells, these models reflect the human wound situation better than animal models. Although two-dimensional models are widely used to investigate processes such as cellular migration and proliferation, models that are more complex are required to gain a deeper knowledge about wound healing. Besides a suitable model system, the generation of precise and reproducible wounds is crucial to ensure comparable results between different test runs. In this study, the generation of a three-dimensional full thickness skin equivalent to study wound healing is shown. The dermal part of the models is comprised of human dermal fibroblast embedded in a rat-tail collagen type I hydrogel. Following the inoculation with human epidermal keratinocytes and consequent culture at the air-liquid interface, a multilayered epidermis is formed on top of the models. To study the wound healing process, we additionally developed an automated wounding device, which generates standardized wounds in a sterile atmosphere.

The goal of this protocol is to build up a three-dimensional full thickness skin equivalent, which resembles natural skin. With a specifically constructed automated wounding device, precise and reproducible wounds can be generated under maintenance of sterility.

3 차원 전체 두께 피부의 세대 동등한 및 자동화 부상

In vitro models are a cost effective and ethical alternative to study cutaneous wound healing processes. Moreover, by using human cells, these models ...

Ischemic Tissue Injury in the Dorsal Skinfold Chamber of the Mouse: A Skin Flap Model to Investigate Acute Persistent Ischemia

Despite profound expertise and advanced surgical techniques, ischemia-induced complications ranging from wound breakdown to extensive tissue necrosis are still occurring, particularly in reconstructive flap surgery. Multiple experimental flap models have been developed to analyze underlying causes and mechanisms and to investigate treatment strategies to prevent ischemic complications. The limiting factor of most models is the lacking possibility to directly and repetitively visualize microvascular architecture and hemodynamics. The goal of the protocol was to present a well-established mouse model affiliating these before mentioned lacking elements. Harder et al. have developed a model of a musculocutaneous flap with a random perfusion pattern that undergoes acute persistent ischemia and results in ~50% necrosis after 10 days if kept untreated. With the aid of intravital epi-fluorescence microscopy, this chamber model allows repetitive visualization of morphology and hemodynamics in different regions of interest over time. Associated processes such as apoptosis, inflammation, microvascular leakage and angiogenesis can be investigated and correlated to immunohistochemical and molecular protein assays. To date, the model has proven feasibility and reproducibility in several published experimental studies investigating the effect of pre-, peri- and postconditioning of ischemically challenged tissue.

The window of the murine dorsal skinfold chamber presented visualizes a zone of acute persistent ischemia of a musculocutaneous flap. Intravital epi-fluorescence microscopy permits for direct and repetitive assessment of the microvasculature and quantification of hemodynamics. Morphologic and hemodynamic results can further be correlated with histological and molecular analyses.

마우스의 등쪽 피하 지방 상공 회의소에서 허혈성 조직 손상 : 피부 플랩 모델은 급성 영구 허혈을 조사하기

Despite profound expertise and advanced surgical techniques, ischemia-induced complications ranging from wound breakdown to extensive tissue necrosis are ...

Assessment of Acute Wound Healing using the Dorsal Subcutaneous Polyvinyl Alcohol Sponge Implantation and Excisional Tail Skin Wound Models.

Wound healing is a complex process that requires the orderly progression of inflammation, granulation tissue formation, fibrosis, and resolution. Murine models provide valuable mechanistic insight into these processes; however, no single model fully addresses all aspects of the wound healing response. Instead, it is ideal to use multiple models to address the different aspects of wound healing. Here, two different methods that address diverse aspects of the wound healing response are described. In the first model, polyvinyl alcohol sponges are subcutaneously implanted along the mouse dorsum. Following sponge retrieval, cells can be isolated by mechanical disruption, and fluids can be extracted by centrifugation, thus allowing for a detailed characterization of cellular and cytokine responses in the acute wound environment. A limitation of this model is the inability to assess the rate of wound closure. For this, a tail skin excision model is utilized. In this model, a 10 mm x 3 mm rectangular piece of tail skin is excised along the dorsal surface, near the base of the tail. This model can be easily photographed for planimetric analysis to determine healing rates and can be excised for histological analysis. Both described methods can be utilized in genetically altered mouse strains, or in conjunction with models of comorbid conditions, such as diabetes, aging, or secondary infection, in order to elucidate wound healing mechanisms.

Here, two murine wound healing models are described, one designed to assess cellular and cytokine wound healing responses and the other to quantify the rate of wound closure. These methods can be used with complex disease models such as diabetes to determine mechanisms of various aspects of poor wound healing.

등지성 폴리비닐 알코올 스폰지 이식 및 절제 꼬리 피부 상처 모델을 사용하여 급성 상처 치유의 평가.

Wound healing is a complex process that requires the orderly progression of inflammation, granulation tissue formation, fibrosis, and resolution. Murine ...

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.

Mechanotransduction-driven, human-like hypertrophic scarring의 마우스 모델

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

Digital Planimetry for Assessing Wound Closure Kinetics in a Mouse Model

Chronic wounds, due to their high prevalence, are a serious global health concern. Effective therapeutic strategies can significantly accelerate healing, thereby reducing the risk of complications and alleviating the economic burden on healthcare systems. Although numerous experimental studies have investigated wound healing, most rely on qualitative observations or quantitative direct measurements. The objective of this study was to standardize an indirect wound measurement method using digital planimetry, incorporating digital scaling and segmentation. This approach addresses the lack of detailed, step-by-step methodologies for accurate wound assessment. A photodocumentation booth was designed and constructed, and computer-assisted digital planimetry tools were employed to minimize variability in measurements of the wound area, perimeter, and the distance from the wound center to its edges. A circular traumatic wound (5 mm in diameter) was created on the dorsal midline at the shoulder blade level of male CD1 mice (n = 4, 10 weeks old, 30-35 g). Wound evolution was photodocumented for 14 days using the custom-designed photo booth, which controlled lighting conditions, focal distance, and subject positioning. Scaling and wound measurements were performed using segmentation in ImageJ software, and statistical analysis was conducted using statistical analysis software. The kinetics of wound closure showed a slight increase in wound size and perimeter between day 0 and day 2, followed by a gradual decrease until complete closure by day 14. The photodocumentation booth and computer-assisted digital planimetry enabled quantitative measurements with minimal variability. In conclusion, these tools provide a reliable and reproducible method for evaluating wound closure kinetics in pre-clinical models.

Wounds represent a global health challenge. This study developed a standardized photo booth utilizing digital planimetry to minimize wound measurement variability. Monitoring wounds in mice over 14 days revealed an initial increase in wound area and perimeter, followed by gradual closure. This methodology may aid in evaluating wound closure kinetics in pre-clinical models.

마우스 모델에서 상처 봉합 동역학을 평가하기 위한 디지털 플레니메트리

Chronic wounds, due to their high prevalence, are a serious global health concern. Effective therapeutic strategies can significantly accelerate healing, ...

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 ...

생물학

Optimized Protocol for Mouse Wound Healing Model with Stem Cell-Derived Exosomes

Mouse Wound Models and Preparation of Single-Cell Suspensions 

The management of acute full-thickness skin injuries presents a considerable challenge in clinical practice. The complexity of wound healing, involving diverse cell populations and the intricate wound microenvironment, complicates the assessment of treatment effects and their interactions using traditional experimental approaches. Single-cell transcriptome sequencing technology has revolutionized the understanding of cellular functions and mechanisms during wound healing. However, the variability in methodologies for creating mouse wound models introduces confounding factors that can impact experimental results. Furthermore, the process of dissociating mouse skin tissues presents significant technical hurdles, highlighting the critical need for high-quality single-cell suspensions for accurate transcriptome sequencing. Therefore, this study aims to optimize the construction of mouse wound models to accurately assess changes in wound closure while eliminating variables that could influence the subsequent sequencing result. Additionally, the preparation of single-cell suspensions was performed using both enzyme preparation protocols and test kit protocols to optimize cost and effectiveness.

Mouse Wound Models and Preparation of Single-Cell Suspensions

This study presents two crucial experimental techniques: the construction of a murine wound model for assessing wound closure and the preparation of single-cell suspensions from murine skin to preserve cellular viability.

The management of acute full-thickness skin injuries presents a considerable challenge in clinical practice. The complexity of wound healing, involving ...

흉터 상처 수리의 마우스 태아의 피부 모델

요약

더 많은 비디오 탐색

3 차원의 인간 피부 모델을 재구성 : 도구는 일반 피부와 흑색증의 진행을 공부하기

전체 피부 결함 모델은 생체의 혈관 신생을 평가하기

상처 치유의 생쥐 모델

3 차원 전체 두께 피부의 세대 동등한 및 자동화 부상

마우스의 등쪽 피하 지방 상공 회의소에서 허혈성 조직 손상 : 피부 플랩 모델은 급성 영구 허혈을 조사하기

등지성 폴리비닐 알코올 스폰지 이식 및 절제 꼬리 피부 상처 모델을 사용하여 급성 상처 치유의 평가.

Mechanotransduction-driven, human-like hypertrophic scarring의 마우스 모델

마우스 모델에서 상처 봉합 동역학을 평가하기 위한 디지털 플레니메트리

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

Mouse Wound Models and Preparation of Single-Cell Suspensions