저자는 그녀의 공헌을 위해 크리스티안 Pahrmann 감사드립니다. 봉합 재료를 제공 Ethicon, Norderstedt, 함부르크 (독일)에 대한 특별 감사합니다. 기금 소냐 Schrepfer는 도이치 Forschungsgemeinschaft (DFG) (SCHR992 / 3 1 SCHR992/4-1)에서 연구 보조금을 받았습니다.] 작품 ISHLT Shumway 경력 개발 그랜트 2010 포크 연구 기금 (스탠포드 대학)에 의해 지원되었다.

1. 내유 동맥 (IMA) 준비 <ol><li> 동맥 내피는 2 - 프랑스어 포가 티 동맥 embolectomy의 카테터 (박스터 헬스케어, 디어 필드, 일리노이, 미국)의 흐름에 의해 denuded됩니다. 카테터에 두번 내피 손상을 보장하기 위해 전체 선박의 길이 빠져있다. </li><li> 8mm의 길이와 직경 2.5 mm-3mm (예 Translumina, 유콘 스텐 트)의 인간의 스텐 트를 사용합니다. 주의 : 스텐 트의 직경은 사전 및 사후 스텐 트 협착증을 피하기 위해 10 % 이상에 의해 혈관 직경을 초과해서는 안됩니다. 주의 : 같은 연구 내에 스텐 트의 길이를 전환하지 마십시오. </li><li> 원하는 직경을 달성하기위한 적절한 풍선 압력 (어떤은 스텐 트 패키지에 언급되는)를 사용하여 스텐 트를 배포합니다. </li><li> 쇼핑몰 ° C RPMI + 헤파린 (500 IE/10 ML) 얼음 이식까지 4 IMA를 stented. </li></ol> 2. 동물 준비 RNU 누드 (CRL : NIH-Foxn1rnu) 쥐 (3백~3백50g)는 는걸 있습니다scantainer에서 종래의 조건에서 SED는 캐비닛, 수사관 표준 쥐 차우와 autoclaved 물 광고 libidum를 통풍이. <ol><li> 유도 챔버를 사용하여 isoflurane와 쥐 (2.5-3 %)를 마취. </li><li> 복부 면도를하고 뒷면에​​있는 쥐 배치하고 코와 마취를 유지하는 입안을 통해 안면을 넣으십시오. </li><li> 두번이 단계를 반복합니다 - 널리 프로보 - 요오드, 다음 사용을 80 % 에탄올을 사용하여 복부 지역을 소독. 쥐가 anesthetized 충분한지 확인하기 위해 뒷다리를 곤란 반사를 확인하십시오. </li><li> 미세한 관점에서 infrarenal 복부 대동맥을 노출하기 위해 상단 중앙 분리대 개복술을 수행합니다. </li><li> 식염수 moisturized 장갑 안에 창자를 놓습니다. 수분 손실을 방지하기 위해 내장 주위에 장갑을 접으십시오. </li><li> infrarenal 지역에서 분기점으로 대동맥 해부, 조심스럽게 선박의 지점에 손상을 줄 수 없습니다. </li><li> 대동맥 혈류를 중지 마이크로 클램프를 사용합니다. 를 삽입근위 클램프 먼저 말초 겸자로 따라갔다. </li><li> 응용 프로그램을 제거합니다. 0.5-0.7 밀리미터 대동맥 세그먼트 및 헤파린 (200 세대)와 나머지 대동맥를 플러시. </li><li> stented IMA를 타고 적절한 길이로 단축하고 간격으로 위치를. </li><li> 8-0 prolene 봉합사를 (Ethicon, Norderstedt, 독일)를 사용하여 봉합을 실행하여,받는 대동맥으로 IMA를 연결합니다. </li><li> 조심스럽게 첫 두개골 클램프 다음 꼬리 클램프를 엽니다. </li><li> 이식된 IMA과 대동맥의 말초 끝에 가시 펄스가 있어야합니다. </li><li> 복부에 다시​​ 창자를 놓습니다. </li><li> 사전 예열 멸균 식염수에 복부를 플러시. </li><li> 6-0 prolene 실행 봉합을 (Ethicon, Norderstedt, 독일)를 사용하여 복벽의 근육 레이어를 닫습니다. </li><li> 쥐가 마취 아직이지만, subcutaneously 4-5 밀리그램 / kg Carprofen 투입. </li><li> 삼일 게시물 surg 동안 (예 : Carprofen 또는 Meloxicam) 적절한 사후 수술 진통제를 관리ery. </li></ol> 3. 대표 결과 조직학의 경우 표본은 알코올의 등급 시리즈, 포르말린 탈수 4퍼센트에서 수정되었으며, 1 % 건조 과산화 벤 조일 1 개 일간 dibutylphthalate 80 % methylmethacrylate와 20 %, MMA 나의 혼합물 (MMA 전)에 침입 4에 1-2일위한 3% 건조 벤조일의 과산화수소 (MMA III) : 1 개 날, MMA 나는 ° C. 이후 표본은 미리 polymerized 기지에있는 물을 욕조에 유리 튜브 안에 신선한 MMA III에 polymerized되었다. 느린 중합은 26 살에 튜브를 유지함으로써 달성되었다 ° C에서 하룻밤, 중합이 발생하기 전까지 ° C ~ 다음날 아침, 그리고 점차적으로 0.5 ° C ~ 12여 H에 의해 온도를 증가 28 온도를 증가. polymerized 블록 텅스텐 초경 나이프를 갖춘 MICROM HM 360 마이크로톰를 사용하여 5 μm의 두께로 sectioned 있었다. 초, proliferating 세포의 기원 (그림 1)를 식별하려면lides이 중 녹색 형광 단백질 (GFP) 또는 쥐를 식별하는 항체 물들일되었다 MHC-I와 인간의 평활근 세포. 이러한 연구 내용은, 인간 IMA는 리포터 유전자에 GFP가 하룻밤 IMA 전지의 안정적인 도입을 위해 lentiviral 입자를 이용하여 incubated되었습니다. 인간의 기원으로부터 분리 딸 세포가 GFP를 표현하여 식별 수 있습니다. deparaffinization 후, 열을 유발 에피토프 검색은 증기선을 사용하여 항원 복구 솔루션에 슬라이드를 가열하여 수행됩니다. 이미지-IT FX 신호 확장기가 차단 단계에 사용될 수 있습니다. 인간의 근본 세포 GFP (주 항체 희석액 (Dako)에 희석 1:100)에 대해 마우스 단클론 항체를 사용하여 확인하고, 추가로 염소 - 안티 - 마우스 IgG, 알렉사 형석 488 (1:1000 이차 항체 희석액에 희석으로 표시됩니다 ). 평활근 세포는 염소 - 안티 - 토끼 IgG, 알렉사 Fluo 다음, 토끼 polycolonal 안티 평활근 α-actin (주 항체 희석액에 희석 1:100)로 표시된R 555 (이차 항체 diluents에 희석 1:1000). 각 항체 배양 단계는 세 번 PBS는 중간에 세척으로 1 시간 37 ° C에서 수행됩니다. 핵은 10 분 DAPI 물들일 수 있습니다. 골드 antifade 시약을 연장 사용하여 슬라이드를 장착 후, 샘플은 공촛점 현미경을 사용하여 분석되었다. <img alt="그림 1" src="/files/ftp_upload/3663/3663fig1.jpg" /> 1 그림. 인간의 평활근 세포와 같은 neointimal 세포 :.. 그린 = 안티 GFP, 인간의 근본 세포를 레이블링, 빨간색 = 안티 인간의 평활근 세포 actin, 파란색 = DAPI, 세포 핵을 식별 B : 녹색 = 안티 쥐 MHC-I, 레이블링 쥐 원산지 세포, 빨간색 = 안티 인간의 평활근 세포 actin, 파란색 = DAPI, 세포 핵 식별. Proliferating 세포가 GFP를위한 평활근 세포와 긍정적인 것으로 판별하지만, 쥐 MHC-I 분자에 대한 부정하고 있습니다. 따라서 proliferating 세포는 인간의 근원입니다.

<ol>
	<li>Deuse, T., Ikeno, F., Robbins, R. C., Schrepfer, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Imaging+In-Stent+Restenosis:+An+Inexpensive,+Reliable,+and+Rapid+Preclinical+Model.">Imaging In-Stent Restenosis: An Inexpensive, Reliable, and Rapid Preclinical Model.</a> J. Vis. Exp. (31), e1346 (2009).</li><li>Oyamada, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trans-iliac+rat+aorta+stenting:+a+novel+high+throughput+preclinical+stent+model+for+restenosis+and+thrombosis.">Trans-iliac rat aorta stenting: a novel high throughput preclinical stent model for restenosis and thrombosis.</a> J. Surg. Res. 166, e91-e95 (2011).</li><li>Chamberlain, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+mouse+model+of+in+situ+stenting.">A novel mouse model of in situ stenting.</a> Cardiovascular research. 85, 38-44 (2010).</li><li>Deuse, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Introducing+the+first+polymer-free+leflunomide+eluting+stent.">Introducing the first polymer-free leflunomide eluting stent.</a> Atherosclerosis. 200, 126-134 (2008).</li><li>Finn, A. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+healing+after+sirolimus,+paclitaxel,+and+bare+metal+stent+placement+in+combination+with+peroxisome+proliferator-activator+receptor+gamma+agonists:+requirement+for+mTOR/Akt2+in+PPARgamma+activation.">Differential healing after sirolimus, paclitaxel, and bare metal stent placement in combination with peroxisome proliferator-activator receptor gamma agonists: requirement for mTOR/Akt2 in PPARgamma activation.</a> Circulation research. 105, 1003-1012 (2009).</li><li>Tellez, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coronary+bare+metal+stent+implantation+in+homozygous+LDL+receptor+deficient+swine+induces+a+neointimal+formation+pattern+similar+to+humans.">Coronary bare metal stent implantation in homozygous LDL receptor deficient swine induces a neointimal formation pattern similar to humans.</a> Atherosclerosis. 213, 518-524 (2010).</li><li>Suzuki, Y., Yeung, A. C., Ikeno, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+pre-clinical+animal+model+in+the+translational+research+of+interventional+cardiology.">The pre-clinical animal model in the translational research of interventional cardiology.</a> JACC Cardiovasc. Interv. 2, 373-383 (2009).</li><li>Holt, C. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intimal+proliferation+in+an+organ+culture+of+human+internal+mammary+artery.">Intimal proliferation in an organ culture of human internal mammary artery.</a> Cardiovascular research. 26, 1189-1194 (1992).</li><li>Swanson, N., Javed, Q., Hogrefe, K., Gershlick, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+internal+mammary+artery+organ+culture+model+of+coronary+stenting:+a+novel+investigation+of+smooth+muscle+cell+response+to+drug-eluting+stents.">Human internal mammary artery organ culture model of coronary stenting: a novel investigation of smooth muscle cell response to drug-eluting stents.</a> Clin. Sci. (Lond). 103, 347-353 (2002).</li></ol>

관심의 어떠한 충돌 선언 없습니다.

생체내 연구 모델에서 서로 다른가 스텐 트 배치 후 내막 증식증의 발전을 조사하기 위해 존재하고 있지만, 이러한 모델은 여전히 극복하기 위해 translational 장애물에 직면. 또한, 대형 동물 모델은 고가의 주택 및 특수 조건뿐만 아니라 수술 장비 모든 실험실을 사용할 수 없습니다입니다. 인간의 내막 증식 및 인 - 스텐 트 restenosis의 개발을 연구하는 인간 IMA를 사용하...

<table border="1"><tbody><tr><td> 시약의 이름 </td><td> 회사 </td><td> 카탈로그 번호 </td><td> 댓글 </td></tr><tr><td> 2 프랑스어 포가 티의 카테터 </td><td> 박스터 헬스케어, 디어 필드, 일리노이, 미국 </td><td> 120602F </td><td></td></tr><tr><td> 유콘 스텐 트 </td><td> Translumina 게엠베하, Hechingen, 독일 </td><td></td><td> 학업 프로토콜에 따라 원하는 스텐 트를 사용하여 </td></tr><tr><td> RPMI 미디어 </td><td> Biochrom </td><td> Nr.F1275 </td><td></td></tr><tr><td> 헤파린 </td><td> 벡스터 </td><td> 2B0953 </td><td></td></tr><tr><td> isoflurane </td><td> 대수도 원장 </td><td> B506 </td><td></td></tr><tr><td> 프로보 - 요오드 </td><td> Betadine Puredue 파마 </td><td> EAN : 5995327165830 </td><td></td></tr><tr><td> 80% ethano리터 </td><td> Geyer </td><td> ETV 80 / 0500 </td><td></td></tr><tr><td> 마이크로 클램프 </td><td> 하버드 장치 </td><td> PY2 - 61-0186 </td><td></td></tr><tr><td> 봉합 8-0 </td><td> 존슨 앤 존슨, </td><td> 2808G </td><td></td></tr><tr><td> 봉합 6-0 </td><td> 존슨 앤 존슨, </td><td> 1,698 H </td><td></td></tr><tr><td> Carprofen </td><td> Feizer의 투어뇌려 </td><td> PZN : 0,110,208 </td><td></td></tr><tr><td> Metamizol </td><td> Ratiopharm </td><td></td><td></td></tr><tr><td> 대상 검색 솔루션, pH9 </td><td> Dako </td><td> S2368 </td><td></td></tr><tr><td> 이미지 IT FX 신호 확장기 </td><td> Invitrogen </td><td> I36933 </td><td></td></tr><tr><td> 마우스 단클론 항 GFP 항체 </td><td> BD 생활 색상 </td><td> 632,381 </td><td></td></tr><tr><td> 일차 항체 희석액 </td><td> Dako </td><td> S3022 </td><td></td></tr><tr><td> 염소 - 안티 - 마우스 IgG, 알렉사 형석 488 </td><td> Invitrogen </td><td> A11017 </td><td></td></tr><tr><td> 이차 항체 희석액 </td><td> Dako </td><td> S0809 </td><td></td></tr><tr><td> 토끼 polycolonal 안티 평활근 α-actin </td><td> Abcam, </td><td> ab5694 </td><td></td></tr><tr><td> 염소 - 안티 - 토끼 IgG, 알렉사 형석 555 </td><td> Invitrogen </td><td> A21430 </td><td></td></tr><tr><td> 골드 antifade 시약을 연장 </td><td> Invitrogen </td><td> P36930 </td><td></td></tr></tbody></table>

human internal mammary artery (ima) transplantation and stenting: a human model to study the development of in-stent restenosis

선박 stenting 후 내막 증식증의 개발에 pathobiological 및 pathophysiological 프로세스를 조사 생체내 연구 모델의 잠복기는 translational 방법 1,2를위한 열쇠입니다. 일반적으로 사용되는 동물 모델 마우스, 쥐, 토끼 및 돼지 3-5 포함되어 있습니다. 이러한 생물 학적 과정이 이미 동물의 혈관에서 공부하지만, 인간의 연구 모델 6,7 년 전에는 수행되지 않습니다 때문에 그러나, 임상 설정으로 이러한 모델의 번역이 어려운 남아 있습니다. 이 동영상에서 우리는이 translational 격차를 극복하는 새로운 인간화 모델을 보여줍니다. 표시된 절차, 재현할 쉬우며, 빠르게 수행할 수 있으며 내막 증식증과 다양한 stents의 적용의 개발을 공부하기 적합합니다. 이 비디오는 immunodeficient 쥐에 이식 다음에 인간의 혈관에 스텐 트 기술을 수행하는 방법을 보여줍니다, 그리고인간과 같은 proliferating 세포의 출처를 식별합니다.

Watch this Scientific Journal Video about Human Internal Mammary Artery IMA Transplantation and Stenting: A Human Model to Study the Development of In-Stent Restenosis at JoVE.com

Human Internal Mammary Artery IMA Transplantation and Stenting: A Human Model to Study the Development of In-Stent Restenosis

이 비디오는 immunodeficient 쥐 모델에서 인간의 함대 (IMA)를 사용하여 스텐 트를 배포 후 내막 증식증의 발전을 연구하는 모델을 보여줍니다.

인간 내유 동맥 (IMA) 이식과 Stenting : 인 Restenosis 스텐 트의 개발을 공부하는 인간 모델

Preclinical in vivo research models to investigate pathobiological and pathophysiological processes in the development of intimal hyperplasia after vessel ...

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Human Internal Mammary Artery (IMA) Transplantation and Stenting: A Human Model to Study the Development of In-Stent Restenosis

11.2K 조회수.  TSI-Lab, Germany. 이 비디오는 immunodeficient 쥐 모델에서 인간의 함대 (IMA)를 사용하여 스텐 트를 배포 후 내막 증식증의 발전을 연구하는 모델을 보여줍니다.

비디오: Human Internal Mammary Artery IMA Transplantation and Stenting: A Human Model to Study the Development of In-Stent Restenosis

Orthotopic Aortic Transplantation: A Rat Model to Study the Development of Chronic Vasculopathy

Research models of chronic rejection are essential to investigate pathobiological and pathophysiological processes during the development of transplant vasculopathy (TVP).
The commonly used animal model for cardiovascular chronic rejection studies is the heterotopic heart transplant model performed in laboratory rodents. This model is used widely in experiments since Ono and Lindsey (3) published their technique. To analyze the findings in the blood vessels, the heart has to be sectioned and all vessels have to be measured.
Another method to investigate chronic rejection in cardiovascular questionings is the aortic transplant model (1, 2). In the orthotopic aortic transplant model, the aorta can easily be histologically evaluated (2). The PVG-to-ACI model is especially useful for CAV studies, since acute vascular rejection is not a major confounding factor and Cyclosporin A (CsA) treatment does not prevent the development of CAV, similar to what we find in the clinical setting (4). A7-day period of CsA is required in this model to prevent acute rejection and to achieve long-term survival with the development of TVP.
This model can also be used to investigate acute cellular rejection and media necrosis in xenogeneic models (5).

This video demonstrates the orthotopic aortic transplant model as a simple model to study the development of transplant vasculopathy (TVP) in rats.

Orthotopic 대동맥 이식 : 만성 Vasculopathy의 개발 연구에 생쥐 모델

Research models of chronic rejection are essential to investigate pathobiological and pathophysiological processes during the development of transplant ...

연구

의학

Utilizing Transcranial Magnetic Stimulation to Study the Human Neuromuscular System

Transcranial magnetic stimulation (TMS) has been in use for more than 20 years 1, and has grown exponentially in popularity over the past decade. While the use of TMS has expanded to the study of many systems and processes during this time, the original application and perhaps one of the most common uses of TMS involves studying the physiology, plasticity and function of the human neuromuscular system. Single pulse TMS applied to the motor cortex excites pyramidal neurons transsynaptically 2 (Figure 1) and results in a measurable electromyographic response that can be used to study and evaluate the integrity and excitability of the corticospinal tract in humans 3. Additionally, recent advances in magnetic stimulation now allows for partitioning of cortical versus spinal excitability 4,5. For example, paired-pulse TMS can be used to assess intracortical facilitatory and inhibitory properties by combining a conditioning stimulus and a test stimulus at different interstimulus intervals 3,4,6-8. In this video article we will demonstrate the methodological and technical aspects of these techniques. Specifically, we will demonstrate single-pulse and paired-pulse TMS techniques as applied to the flexor carpi radialis (FCR) muscle as well as the erector spinae (ES) musculature. Our laboratory studies the FCR muscle as it is of interest to our research on the effects of wrist-hand cast immobilization on reduced muscle performance6,9, and we study the ES muscles due to these muscles clinical relevance as it relates to low back pain8. With this stated, we should note that TMS has been used to study many muscles of the hand, arm and legs, and should iterate that our demonstrations in the FCR and ES muscle groups are only selected examples of TMS being used to study the human neuromuscular system.

Transcranial magnetic stimulation (TMS) is a non-invasive tool to gain insight on the physiology and function of the human nervous system. Here, we present our TMS techniques to study cortical excitability of the upper limb and lumbar musculature.

인간 신경근육학 시스템을 공부하기 Transcranial 자기 자극을 활용

Transcranial magnetic stimulation (TMS) has been in use for more than 20 years 1, and has grown exponentially in popularity over the past decade.  While ...

Stem Cell Transplantation in an in vitro Simulated Ischemia/Reperfusion Model

Stem cell transplantation protocols are finding their way into clinical practice1,2,3. Getting better results, making the protocols more robust, and finding new sources for implantable cells are the focus of recent research4,5. Investigating the effectiveness of cell therapies is not an easy task and new tools are needed to investigate the mechanisms involved in the treatment process6. We designed an experimental protocol of ischemia/reperfusion in order to allow the observation of cellular connections and even subcellular mechanisms during ischemia/reperfusion injury and after stem cell transplantation and to evaluate the efficacy of cell therapy. H9c2 cardiomyoblast cells were placed onto cell culture plates7,8. Ischemia was simulated with 150 minutes in a glucose free medium with oxygen level below 0.5%. Then, normal media and oxygen levels were reintroduced to simulate reperfusion. After oxygen glucose deprivation, the damaged cells were treated with transplantation of labeled human bone marrow derived mesenchymal stem cells by adding them to the culture. Mesenchymal stem cells are preferred in clinical trials because they are easily accessible with minimal invasive surgery, easily expandable and autologous. After 24 hours of co-cultivation, cells were stained with calcein and ethidium-homodimer to differentiate between live and dead cells. This setup allowed us to investigate the intercellular connections using confocal fluorescent microscopy and to quantify the survival rate of postischemic cells by flow cytometry. Confocal microscopy showed the interactions of the two cell populations such as cell fusion and formation of intercellular nanotubes. Flow cytometry analysis revealed 3 clusters of damaged cells which can be plotted on a graph and analyzed statistically. These populations can be investigated separately and conclusions can be drawn on these data on the effectiveness of the simulated therapeutical approach.

Stem Cell Transplantation in an in vitro Simulated Ischemia/Reperfusion Model

We demonstrate how to set up an in vitro ischemia/reperfusion model and how to evaluate the effect of stem cell therapy on postischemic cardiac cells.

에서 간세포 이식 체외에서 시뮬레이션 국소 빈혈 / Reperfusion 모델

Stem cell transplantation protocols are finding their way into clinical practice1,2,3. Getting better results, making the protocols more robust, and ...

Human Neuroendocrine Tumor Cell Lines as a Three-Dimensional Model for the Study of Human Neuroendocrine Tumor Therapy

Neuroendocrine tumors (NETs) are rare tumors, with an incidence of two per 100,&#x200A;000 individuals per year, and they account for 0.5% of all human malignancies.1 Other than surgery for the minority of patients who present with localized disease, there is little or no survival benefit of systemic therapy. Therefore, there is a great need to better understand the biology of NETs, and in particular define new therapeutic targets for patients with nonresectable or metastatic neuroendocrine tumors. 3D cell culture is becoming a popular method for drug screening due to its relevance in modeling the in vivo tumor tissue organization and microenvironment.2,3 The 3D multicellular spheroids could provide valuable information in a more timely and less expensive manner than directly proceeding from 2D cell culture experiments to animal (murine) models.
To facilitate the discovery of new therapeutics for NET patients, we have developed an in vitro 3D multicellular spheroids model using the human NET cell lines. The NET cells are plated in a non-adhesive agarose-coated 24-well plate and incubated under physiological conditions (5% CO2, 37 &deg;C) with a very slow agitation for 16-24 hr after plating. The cells form multicellular spheroids starting on the 3rd or 4th day. The spheroids become more spherical by the 6th day, at which point the drug treatments are initiated. The efficacy of the drug treatments on the NET spheroids is monitored based on the morphology, shape and size of the spheroids with a phase-contrast light microscope. The size of the spheroids is estimated automatically using a custom-developed MATLAB program based on an active contour algorithm. Further, we demonstrate a simple method to process the HistoGel embedding on these 3D spheroids, allowing the use of standard histological and immunohistochemical techniques. 
This is the first report on generating 3D spheroids using NET cell lines to examine the effect of therapeutic drugs. We have also performed histology on these 3D spheroids, and displayed an example of a single drug's effect on growth and proliferation of the NET spheroids. Our results support that the NET spheroids are valuable for further studies of NET biology and drug development.

We present a simple agarose overlay platform to grow 3D multicellular spheroids using neuroendocrine cancer cell lines. This method provides a very convenient way to examine the effect of therapeutic drugs on the neuroendocrine tumor cells. It could also help us establish human neuroendocrine tumor spheroids for cancer therapy.

인간 Neuroendocrine 종양 치료의 연구를위한 3 차원 모델로 인간 Neuroendocrine 종양 세포 라인

Neuroendocrine tumors (NETs) are rare tumors, with an incidence of two per 100,&#x200A;000 individuals per year, and they account for 0.5% of all human ...

A Murine Model of Stent Implantation in the Carotid Artery for the Study of Restenosis

Despite the considerable progress made in the stent development in the last decades, cardiovascular diseases remain the main cause of death in western countries. Beside the benefits offered by the development of different drug-eluting stents, the coronary revascularization bears also the life-threatening risks of in-stent thrombosis and restenosis. Research on new therapeutic strategies is impaired by the lack of appropriate methods to study stent implantation and restenosis processes. Here, we describe a rapid and accessible procedure of stent implantation in mouse carotid artery, which offers the possibility to study in a convenient way the molecular mechanisms of vessel remodeling and the effects of different drug coatings.

A model of stent implantation in mouse carotid artery is described. Compared to other similar methods, this procedure is very rapid, simple and accessible, offering the possibility to study in a convenient way the vascular wall reaction to different drug-eluting stents and the molecular mechanisms of restenosis.

재 협착의 연구에 대한 경동맥 스텐트 시술의 쥐과 모형

Despite  the considerable progress made in the stent development in the last decades,  cardiovascular diseases remain the main cause of death in western ...

A Human Ex Vivo Atherosclerotic Plaque Model to Study Lesion Biology

Atherosclerosis is a chronic inflammatory disease of the vasculature. There are various methods to study the inflammatory compound in atherosclerotic lesions. Mouse models are an important tool to investigate inflammatory processes in atherogenesis, but these models suffer from the phenotypic and functional differences between the murine and human immune system. In vitro cell experiments are used to specifically evaluate cell type-dependent changes caused by a substance of interest, but culture-dependent variations and the inability to analyze the influence of specific molecules in the context of the inflammatory compound in atherosclerotic lesions limit the impact of the results. In addition, measuring levels of a molecule of interest in human blood helps to further investigate its clinical relevance, but this represents systemic and not local inflammation. Therefore, we here describe a plaque culture model to study human atherosclerotic lesion biology ex vivo. In short, fresh plaques are obtained from patients undergoing endarterectomy or coronary artery bypass grafting and stored in RPMI medium on ice until usage. The specimens are cut into small pieces followed by random distribution into a 48-well plate, containing RPMI medium in addition to a substance of interest such as cytokines or chemokines alone or in combination for defined periods of time. After incubation, the plaque pieces can be shock frozen for mRNA isolation, embedded in Paraffin or OCT for immunohistochemistry staining or smashed and lysed for western blotting. Furthermore, cells may be isolated from the plaque for flow cytometry analysis. In addition, supernatants can be collected for protein measurement by ELISA. In conclusion, the presented ex vivo model opens the possibility to further study inflammatory lesional biology, which may result in identification of novel disease mechanisms and therapeutic targets.

A Human Ex Vivo Atherosclerotic Plaque Model to Study Lesion Biology

Atherosclerosis is a chronic inflammatory process. This manuscript illustrates an easy to use ex vivo model to investigate fresh carotid or coronary artery plaques. The ex vivo model allows for the investigation of potential substances on the inflammatory milieu in human atherosclerotic lesions and results can be analyzed by various methods.

인간의<em&gt; 전의 VIVO</em&gt; 죽상 경화 모델 병변 생물학을 연구하는

Atherosclerosis is a chronic inflammatory disease of the vasculature. There are various methods to study the inflammatory compound in atherosclerotic ...

Excitotoxic Stimulation of Brain Microslices as an In vitro Model of Stroke

Examining molecular mechanisms involved in neuropathological conditions, such as ischemic stroke, can be difficult when using whole animal systems. As such, primary or &#39;neuronal-like&#39; cell culture systems are commonly utilized. While&#160;these systems are relatively easy to work with, and are useful model systems in which various functional outcomes (such as cell death) can be readily quantified, the examined outcomes and pathways in cultured immature neurons (such as excitotoxicity-mediated cell death pathways) are not necessarily the same as those observed in mature brain, or in intact tissue. Therefore, there is the need to develop models in which cellular mechanisms in mature neural tissue can be examined. We have developed an in vitro technique that can be used to investigate a variety of molecular pathways in intact nervous tissue. The technique described herein utilizes rat cortical tissue, but this technique can be adapted to use tissue from a variety of species (such as mouse, rabbit, guinea pig, and chicken) or brain regions (for example, hippocampus, striatum, etc.). Additionally, a variety of stimulations/treatments can be used (for example, excitotoxic, administration of inhibitors, etc.). In conclusion, the brain slice model described herein can be used to examine a variety of molecular mechanisms involved in excitotoxicity-mediated brain injury.

Excitotoxic Stimulation of Brain Microslices as an In vitro Model of Stroke

We have developed a brain slice model which can be used to examine molecular mechanisms involved in excitotoxicity-mediated brain injury.&#160;This technique generates viable mature brain tissue and reduces animal numbers required for experimentation, whilst keeping the neuronal circuitry, cellular interactions, and postsynaptic compartments partly intact.

같은 뇌 Microslices의 흥분 독성 자극<em&gt; 체외</em&gt; 스트로크의 모델

Examining molecular mechanisms involved in neuropathological conditions, such as ischemic stroke, can be difficult when using whole animal systems. As ...

Murine Corneal Transplantation: A Model to Study the Most Common Form of Solid Organ Transplantation

Corneal transplantation is the most common form of organ transplantation in the United States with between 45,000 and 55,000 procedures performed each year. While several animal models exist for this procedure and mice are the species that is most commonly used. The reasons for using mice are the relative cost of using this species, the existence of many genetically defined strains that allow for the study of immune responses, and the existence of an extensive array of reagents that can be used to further define responses in this species. This model has been used to define factors in the cornea that are responsible for the relative immune privilege status of this tissue that enables corneal allografts to survive acute rejection in the absence of immunosuppressive therapy. It has also been used to define those factors that are most important in rejection of such allografts. Consequently, much of what we know concerning mechanisms of both corneal allograft acceptance and rejection are due to studies using a murine model of corneal transplantation. In addition to describing a model for acute corneal allograft rejection, we also present for the first time a model of late-term corneal allograft rejection.

Mice have been used as a model for studying many forms of transplantation, including corneal transplantation. We describe in this report a murine model for both acute and late-term corneal transplantation.

쥐의 각막 이식 : 고체 장기 이식의 가장 일반적인 형태를 연구하기위한 모델

Corneal transplantation is the most common form of organ transplantation in the United States with between 45,000 and 55,000 procedures performed each ...

Establishment of a Human Multiple Myeloma Xenograft Model in the Chicken to Study Tumor Growth, Invasion and Angiogenesis

Multiple myeloma (MM), a malignant plasma cell disease, remains incurable and novel drugs are required to improve the prognosis of patients. Due to the lack of the bone microenvironment and auto/paracrine growth factors human MM cells are difficult to cultivate. Therefore, there is an urgent need to establish proper in vitro and in vivo culture systems to study the action of novel therapeutics on human MM cells. Here we present a model to grow human multiple myeloma cells in a complex 3D environment in vitro and in vivo. MM cell lines OPM-2 and RPMI-8226 were transfected to express the transgene GFP and were cultivated in the presence of human mesenchymal cells and collagen type-I matrix as three-dimensional spheroids. In addition, spheroids were grafted on the chorioallantoic membrane (CAM) of chicken embryos and tumor growth was monitored by stereo fluorescence microscopy. Both models allow the study of novel therapeutic drugs in a complex 3D environment and the quantification of the tumor cell mass after homogenization of grafts in a transgene-specific GFP-ELISA. Moreover, angiogenic responses of the host and invasion of tumor cells into the subjacent host tissue can be monitored daily by a stereo microscope and analyzed by immunohistochemical staining against human tumor cells (Ki-67, CD138, Vimentin) or host mural cells covering blood vessels (desmin/ASMA).
In conclusion, the onplant system allows studying MM cell growth and angiogenesis in a complex 3D environment and enables screening for novel therapeutic compounds targeting survival and proliferation of MM cells.

Human multiple myeloma (MM) cells require the supportive microenvironment of mesenchymal cells and extracellular matrix components for survival and proliferation. We established an in vivo chicken embryo model with engrafted human myeloma and mesenchymal cells to study effects of cancer drugs on tumor growth, invasion and angiogenesis.

닭에서 인간의 다발성 골수종 이종 이식 모델의 설립은 종양의 성장, 침략과 혈관 신생을 공부하기

Multiple myeloma (MM), a malignant plasma cell disease, remains incurable and novel drugs are required to improve the prognosis of patients. Due to the ...

Induction of Accelerated Atherosclerosis in Mice: The "Wire-Injury" Model

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. Moreover, by rupture of the defective vascular wall, atherosclerosis induces occlusive thrombus formation, which represents the main cause of myocardial infarction or stroke and the most frequent cause of death. Despite the advances in the cardiovascular field, many questions remain unanswered, and additional basic research is essential to improve our understanding of the molecular mechanisms during atherosclerosis and its effects. Due to limited clinical studies, there is a need for representative animal models recreating atherosclerotic conditions such as neointima formation after stent implantation, balloon angioplasty, or endarterectomy. Since the mouse presents many advantages and is the most frequently used model for studying molecular processes, the current study proposes an invasive procedure of endothelial denudation, also known as the wire-injury model, which is representative of the human condition of neointima formation in arteries after revascularization procedures.

Induction of Accelerated Atherosclerosis in Mice: The &quot;Wire-Injury&quot; Model

This study describes an invasive procedure for the induction of accelerated atherosclerosis in mice. In comparison to other methods using electric- or cryo-induced injury, mechanical-induced injury mimics the human condition of restenosis after revascularization therapies and is ideal for the study of the molecular mechanisms involved.

마우스에서 가속 죽상 경화증의 유도: "와이어 부상" 모델

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. ...