Bu çalışma 1R03DK096151 tarafından kısmen desteklenmiştir.

Tüm fareler Jackson Laboratory (Bar Harbor, ME) satın alındı ​​ve NIH Rehberi ve Colorado Denver IACUC Üniversitesi onayıyla Colorado Denver, Barbara Davis Merkezi Hayvan Tesis Üniversitesi ile patojensiz şartlar altında muhafaza edildi. 1. Donör Böbrek Hasat <ol><li> Tüm aletler sterilize işlem boyunca steril eldiven giyin ve steril bir alan sağlamak. Bir ameliyat mikroskobu kullanımı ile tüm ameliyatları gerçekleştirmek. </li><li> Cerrahi pentobarbital ile anestezi uygulanmış (60 mg / kg IP) vericilerden fare verici böbrek çıkarın. Ayak-tutam anestezi derinliği tespit ve solunum hızını gözlemlemek. </li><li> Daha sonra 4-yollu sınırlamalar ile fare hareketsiz kürk Klip. Povidon-iyot ile cildi hazırlayın ve steril bir şekilde fare asmak. </li><li> 2 cm orta hat vertikal karın kesi yapın ve karın boşluğuna girin. Superior barsak geri çekin ve dışagöğüs üzerinde. Prosedür boyunca steril gazlı bez nemli sarılmış bağırsak tutun. </li><li> Büyük karın damarları tespit ve bunları harekete, herhangi bir lomber dalları tanımlamak ve dağlamak veya 10/0 naylon sütür ile lige ya. </li><li> Hemen sol renal damarlara uzak yaklaşık 2mm üzerinde künt diseksiyon ile inferior vena kava (IVC) ve ayrı abdominal aorta (AA) kızdırmak. 10/0 naylon ile bağlanır ve böbrek damarlarının küçük arteriyel ve venöz dalları bölün. </li><li> Şimdi bu yapıların künt diseksiyon ile dikkatlice renal arterden renal ven ayırın. Bu alıcıya IVC bir uç-yan anastamozu oluşturulması için kullanılacak renal ven proksimal ucunda bir Carrel yama doğru oluşturulması için izin verir. </li><li> Tanımlayın, ligatı ve sol adrenal damarları bölün. Bu böbreküstü aorta erişime izin ve 6-0 ipek sütür ligasyon için hazırlık aort etrafına yerleştirilen, ancak şu anda bağlı değildir. </li&gt; <li> Çevredeki facia gelen böbrek, üreter ve damarları harekete geçmek. Sağ ve Arter böbrek döndürün / herhangi posterior dalları bölün. Sonra sola böbrek dönün. </li><li> Şimdi üreter dikkat yönlendirir. Böbrek hilum üreter damarları korumak için çevredeki facia alarak bakımdan üreteri özgür bozmadan. Duktus deferens düzeyinde üreter bölün. Böbrek hemen geri kazanılması için hazırdır. </li><li> Yavaş yavaş, böylece donör heparinizing distal IVC içine heparin 300 birimlerini enjekte edilir. Böbreküstü AA etrafına yerleştirilen 6-0 ipek sütür aşağı kravat ve heparin tuzlu su çözeltisi (100 U / ml) 0.8ml distal abdominal aortun yoluyla sol böbrek serpmek. </li><li> Perfüzyon venöz Carrel yama oluşturmak kesildi kez hemen böbrek içine akmasını önlemek için. AA ve altındaki böbrek arteri açığa böbrek doğru renal ven çekin. </li><li> Renal arter bitişik aorta bir topuk ve yaratma DivideŞekil 1&#39;de gösterildiği gibi e manşet. donörden böbrek, üreter ve damarları çıkartın. </li><li> Implantasyonu kadar önceden hazırlanmış alıcı (0 dakika soğuk işemiya süresi) ya da önceden belirlenmiş bir süre için tercih edilen bir çözelti içinde 4 o C&#39;de saklanabilir doğrudan böbrek çıkarın. Exsanguanation ve servikal dislokasyon ile vericilerden Ötenazi. Donör böbreği kurtarmak için Toplam süresi yaklaşık. 15-20 dk. </li></ol> 2. Böbrek İmplant Tekniği <ol><li> Pentobarbital alıcı fare anestezi (60 mg / kg IP başlangıç ​​dozu, 25 mg / kg IP ek doz gerekli ise). Ayak-tutam anestezi derinliği tespit ve solunum hızını gözlemlemek. Kürk sonra 4-yollu sınırlamalar ile fare hareketsiz ve gözleri oftalmik merhem uygulayın Clip. Povidon-iyot ile cildi hazırlayın ve steril bir şekilde fare asmak. </li><li> 2 cm orta hat vertikal karın kesi yapın ve karın boşluğuna girin. Bağırsak geri çekinsuperior ve göğüs üzerinde dışa. Prosedür boyunca steril gazlı bez nemli sarılmış bağırsak tutun. </li><li> Şu anda sağ nefrektomi gerçekleştirin. 6-0 ipek dikişlerle renal arter ve ven Arter. Dikişlerle böbrek distalinden tüketim. Böbrek proksimal bölmek, sonra 6-0 ipek ile üreter bağlanır ve. </li><li> Böbrek damarlarının altında abdominal aorta ve inferior vena kava (IVC) izole edin. Anastomoz sitesine 4-0 pamuk aort etrafında bağları ve IVC üstün sonra aşağı yerleştirin. 10-0 naylon sütür ile alanında herhangi bir lomber gemileri tespit etmek ve bağlamak. </li><li> Üstün ardından ilk alt, pamuk bağları düğüm. Bu şekilde bazı kan aortatomi yapımında kolay aort tutulur. </li><li> Aort lümeni girmek için bir 30G iğne ile aortatomi Form. Yaklaşık 2mm uzunluğunda ince mikro makas ile kesi uzatın. </li><li> Donör aort topuk-ve-ayak manşet yan anastomoz son yapınAşağıdaki şekilde alıcı aorta. Donör aorta ve alıcı aort ve kravat kesinin alt açı bir 10-0 naylon dikiş kalmak dikiş yerleştirin. </li><li> Donör aort ilk ve abdominal aorta ve kravat kesi üstün köşesinde karşısında ikinci 10-0 naylon yerleştirin. Aortun yan duvarında aşağı üstün bir çalışan sütür hattını yapın ve daha önce yerleştirilen kalmak dikiş karşı kravat. Sonra çalışan moda ve kravat medial duvar dikin. </li><li> Aşağıdaki şekilde alıcıya IVC donör renal venin yan anastomoz son yapın. Bir 30G iğne ile IVC delinme ve ince mikro makasla kesi uzatmak. 10-0 naylon IVC içinde kesi alt köşesine donör renal ven kravat. Renal ven ile İVK ve kravat arasına çalışan sütür hattını olun. NOT: Bu tam kalınlıkta vasküler adventit dahil dikiş iğnesinin geçtiği de hayati önem taşımaktadıria ve intima elde edilir. Kenarların Eversion de sızdırmazlık ve anastomuzuna yardımcıları intima-to-intima temas var olmasını sağlar. Hemostatik pıhtılaşma ajan kaçağının azaltılmasında yararlı olabilir, biz bir cerrah yerine iyi bir teknik güveniyor öneririz. </li><li> Anastomozlar &quot;temiz&quot; olduğundan emin olun. Yani dikiş yerleştirirken karşıt duvarları yakalanmış değil olmasıdır. Bu başarısız bir greft ve arka bacak felç aşırı durumlarda neden olacağı akışına karşı ciddi bir daralma neden olur. NOT: Başka bir hayati önemli faktör anastomoz sütür hatlarının gerilim optimal olmasını sağlamak olduğunu. Çok gevşek ve geri dönüşümsüz sızıntı, çok sıkı ve neden olacaktır akış darlık olacaktır. Arteriyel tarafta bu greft zayıf perfüzyon neden olacaksa, venöz tarafında kalabalık böbrek neden olacaktır eğer. </li><li> Distal 4-0 pamuk kravat yeniden kurulması venöz akış bırakın. Veno ve hemostaz kezBize anastomoz proksimal 4-0 pamuk kravat gevşetin ve hemostaz için arteriyel anastomoz gözlemlemek giderek gözlenmiştir. </li><li> Her iki anastomoz güvenli kabul edilir bir kez fare pamuk bağlarını çıkarın. Pierce iki delik yaratarak bir 20 G iğne ile mesane. </li><li> Deliklerden kavisli bir forseps ipuçları Pass ve iki 10-0 naylon sütür ile mesane duvarına tutturulmuş olan üreter proksimal ucu ile forseps ile mesane yoluyla donör üreter çekin. Üreter mesane içinde geri ve iki 10-0 naylon sütür ile deliği kapatmak için izin ikinci delikten çıkan üreter aşırı uzunluğunu kesin. </li><li> Karın barsak dönün. Çalışan bir moda 5-0 İpek sütür kullanarak iki kat karın duvarı kapatın. </li><li> Kapanış üzerine sıvı resüsitasyon olarak karın içine steril sıcak normal serum fizyolojik 1.0 ml bolus yönetmek ve serum fizyolojik subkutan operasyon sonrası bir 0.8 ml enjekte edilir. Başka bir destek olacakziyaretinde önlemler ameliyat sırasında gerekmektedir. </li><li> Bir ısınma battaniye hayvan kurtarın. Toplam implant süresi yaklaşık. 35-45 dk. Böyle buprenorfın, 0.05 mg / kg, SC, prosedürün başlangıcında ve 72 saat sonrası op her 6-12 saatte 0.1-0.2 ml olarak analjezikler yönetin. </li></ol> 3. Kontralateral Nefrektomi <ol><li> Protokol şartlarına bağlı olarak, birkaç gün implant işleminden sonra, izofluran anestezisi altında bir kontralateral nefrektomi (indüksiyon için% 5 inhale izofluran, bakım için 1.5-2.0%) gerçekleştirin. </li><li> Karın boşluğu girin ve yavaşça hayvanın sağa barsak geri çekin. Çevre fasya sol böbreği ve künt incelemek Açığa. </li><li> 6-0 ipek dikişlerle renal arter, ven ve üreter bağlanır ve sonra dikişler üzerinde böbrek tüketim ve böbrek çıkarın. Toplam ameliyat süresi yaklaşık. 10 dk. </li></ol> 4. Graft Değerlendirmesi <ol><li> Tedbir serum creatialkali pikrat yöntemi (Jaffe reaksiyonu) ile dokuz. </li></ol>

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	<li>Skoskiewicz, M., Chase, C., Winn, H. J., Russell, P. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kidney+transplants+between+mice+of+graded+immunogenetic+diversity.">Kidney transplants between mice of graded immunogenetic diversity.</a> Transplant Proc. 5 (1), 721-725 (1973).</li><li>Kalina, S. L., Mottram, P. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+microsurgical+technique+for+renal+transplantation+in+mice.">A microsurgical technique for renal transplantation in mice.</a> Aust N Z J Surg. 63 (3), 213-216 (1993).</li><li>Han, W. R., Murray-Segal, L. J., Mottram, P. 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The authors have nothing to disclose.

Bu nakli tekniği Mastering zordur, ancak başarılı kez çok güçlü bir araştırma aracı. Hasta cerrah / araştırmacı detay ve küçük hayvan modellerinde daha çok herhangi bir cerrahi işlem mastering için anahtar olduğunu tekniğin, tutarlılık dikkat tarafından ödüllendirilir. Fare böbrek nakli mastering teknik zorluklar birçok kıvrımlar vardır ve diğer küçük hayvan nakli modellerinde deneyimi bu yordamı mücadelede önce kazanılan gerektiğini kuvvetle muhtemeldir. <p class="jove_conte...

1973 yılından bu yana farelerde böbrek nakli modeli değerli bir araştırma aracı olmuştur, ama teknik sorunlar yaygın kullanımını engellemiştir. Yıllar boyunca birçok kağıtları yayınlanan bu prosedüre iyileştirmeler / iyileştirmeler ayrıntılarıyla oylandı. Öncelikle vaskülarize solid organ transplantasyonu bir model olarak bu prosedür sadece, aynı zamanda 1973 6 Russell laboratuvar tarafından icat edildi heterotopik kalp nakli modeli muhtemelen ikinci. Her iki model de allojenik ret yanıtları, gecikmiş greft fonksiyonu gelişimi içine araştırma kendilerini ödünç ve iskemi reperfüzyon hasarı. Böbrek nakli ile rapor edilecek en yaygın konulardan biri de laboratuvarda yaşanan arteriyel tromboz 4,5,7 nispeten yüksek insidansı. Bu nedenle trombüs oluşumunun bir literatür taraması gerçekleştirmek ve muhtemelen bu teknik sorunun nedenini bulmak için yola ve aynı zamanda hazırlamak içinolası çözüm. Tromboz en muhtemel nedeni kan bağışı renal artere daha sonra donör böbrek aorta içine alıcı aortadan alır biraz dolambaçlı yoldur. Bu yol, trombosit aktivasyonu ve trombus oluşumuna neden olabilir renal arter türbülans neden olur. Son gözlemler ve literatür 8-14 bir arama dayanarak% 0 tromboz azalttı yeni bir teknik ile geldi. Burada açıklanan teknik imkanları kan akışını geliştirilmiş bir arteriyel topuk-ve-ayak manşet oluşumunda önceden bildirilen teknikler arasında değişir ve anlamlı trombüs oluşumunu azaltır. Manşon az 45 ° aort (Şekil 1A ve 1B) uzunlamasına eksenine belirli bir açıda renal arter ostiyum yüzeyi boyunca alt böbrek aort bölünmesi ile meydana getirilir. Bu uzunluğu yaklaşık 2mm bir manşet ile sonuçlanır. Bir venöz Carrel yama r bölücü oluşturulmuşturIVC içine enal ven böylece manşet çapı artar. Kızıl-böbrek verici abdominal aorta topuk-and-toe manşet uç-yan alıcı abdominal aort ve donör renal venin anastomoz olduğunu / IVC yama (IVC) uç-yan alıcı karın inferior vena kava anastomoz olduğunu . Üreter sonra içine ve Han ve ark 3 tarafından açıklandığı gibi mesane bağlanır. Bu çalışmada sadece sıcak iskemi süresi ile tedavi edilmemiş nakli için (yani., Soğuk iskemi) karşılaştırılır. Bu durumda sıcak iskemi verici böbrek yoluyla kan akımının kesilmesinden zaman anlamına gelir (aşağıda 1.11 adıma) ve alıcı (aşağıda adım 2.11) &#39;de greft reperfüzyon. Soğuk iskemi böbrek perfüze değildir ve implant prosedürünün başına kadar soğuk hava depolarında muhafaza edilir zaman anlamına gelir.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Instrument</td>
			<td>Roboz #</td>
			<td>Fine Science Tools #</td>
			<td>Arosurgical #</td>
		</tr>
		<tr>
			<td>Straight micro-dissecting forcep #5</td>
			<td>RS-5015</td>
			<td>11295-51</td>
			<td></td>
		</tr>
		<tr>
			<td>Curved micro-dissecting forcep #7</td>
			<td>RS-5047</td>
			<td>11297-00</td>
			<td></td>
		</tr>
		<tr>
			<td>Curved serrated forcep</td>
			<td>RS-5137</td>
			<td>11052-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Vannas micro-dissecting scissors, short</td>
			<td>RS-5610</td>
			<td></td>
			<td>09.140.08</td>
		</tr>
		<tr>
			<td colspan="2">Micro-dissecting scissors, straight, sharp, long</td>
			<td></td>
			<td>11.602.11</td>
		</tr>
		<tr>
			<td>Micro spring handle needle holder</td>
			<td></td>
			<td></td>
			<td>11.549.15</td>
		</tr>
		<tr>
			<td>Straight mosquito forcep</td>
			<td></td>
			<td>91308-12</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro-dissecting scissors, straight, blunt</td>
			<td>RS-5962</td>
			<td>14078-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro-dissecting scissors, curved, blunt</td>
			<td>RS-5981</td>
			<td>14079-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro retractor</td>
			<td>RS-6540</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Instrument tray, 10&#8221; x 6 &#189;&#8221; x &#190;&#8221;</td>
			<td>RT-1350S</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Silk suture, 5/0, 22.5m spool</td>
			<td></td>
			<td>18020-50</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Suture</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>10/0 nylon</td>
			<td></td>
			<td></td>
			<td>T4A10Q07</td>
		</tr>
		<tr>
			<td>5/0 silk</td>
			<td></td>
			<td></td>
			<td>E19A05N</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Gloves</td>
			<td></td>
			<td>Drapes</td>
			<td></td>
		</tr>
		<tr>
			<td>Biogel from Medex Supply</td>
			<td></td>
			<td>Precept, #64-9012-9</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Syringes</td>
			<td></td>
			<td>Cotton applicators</td>
			<td></td>
		</tr>
		<tr>
			<td>B-D 1cc insulin, #329424</td>
			<td></td>
			<td colspan="2">Fisher-brand, #23-400-100</td>
		</tr>
		<tr>
			<td></td>
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			<td>Povidone-Iodine swabs</td>
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			<td>PDI, #B40600</td>
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			<td>4/0 Cotton ties</td>
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			<td colspan="2">Domestic cotton autoclaved with instruments</td>
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murine kidney transplant technique

The first mouse kidney transplant technique was published in 19731 by the Russell laboratory. Although it took some years for other labs to become proficient in and utilize this technique, it is now widely used by many laboratories around the world. A significant refinement to the original technique using the donor aorta to form the arterial anastomosis instead of the renal artery was developed and reported in 1993 by Kalina and Mottram 2 with a further advancement coming from the same laboratory in 1999 3. While one can become proficient in this model, a search of the literature reveals that many labs still experience a high proportion of graft loss due to arterial thrombosis. We describe here a technique that was devised in our laboratory that vastly reduces the arterial thrombus reported by others 4,5. This is achieved by forming a heel-and-toe cuff of the donor infra-renal aorta that facilitates a larger anastomosis and straighter blood flow into the kidney.

Bu cerrahi teknik, basit greft sağkalım / ret çalışmaları, ya da oldukça karmaşık deneysel protokollerin biri için izin verir. Rakamlar biz bu gelişmiş arter anastomoz tekniği kullanılarak avantajlarını göstermek aşağıda. Biz önemli ölçüde% 0, böylece artan verimlilik% 35 den arteriyel tromboz insidansını azaltmıştır bu tekniği kullanarak. Biz tutulan Aynı% 0 tromboz sonucu bir yıldan bu tekniği kullandık. 1, bu yeni tekniğin temeli olan arteriyel topuk-ve-ayak manşe...

Watch this Scientific Journal Video about Murine Kidney Transplant Technique at JoVE.com

Murine Kidney Transplant Technique

The goal of this manuscript is to describe the steps required to perform a kidney transplant in a mouse, paying particular attention to the details of the arterial anastomosis.

Murin Böbrek nakli Tekniği

The first mouse kidney transplant technique was published in 19731 by the Russell laboratory. Although it took some years for other labs to become ...

Research

JoVE Journal

Medicine

15.8K Görüntüleme.  University of Colorado, Denver. The goal of this manuscript is to describe the steps required to perform a kidney transplant in a mouse, paying particular attention to the details of the arterial anastomosis. 

Video: Murine Kidney Transplant Technique

A Method for Murine Islet Isolation and Subcapsular Kidney Transplantation

Since the early pioneering work of Ballinger and Reckard demonstrating that transplantation of islets of Langerhans into diabetic rodents could normalize their blood glucose levels, islet transplantation has been proposed to be a potential treatment for type 1 diabetes 1,2. More recently, advances in human islet transplantation have further strengthened this view 1,3. However, two major limitations prevent islet transplantation from being a widespread clinical reality: (a) the requirement for large numbers of islets per patient, which severely reduces the number of potential recipients, and (b) the need for heavy immunosuppression, which significantly affects the pediatric population of patients due to their vulnerability to long-term immunosuppression. Strategies that can overcome these limitations have the potential to enhance the therapeutic utility of islet transplantation.
Islet transplantation under the mouse kidney capsule is a widely accepted model to investigate various strategies to improve islet transplantation. This experiment requires the isolation of high quality islets and implantation of islets to the diabetic recipients. Both procedures require surgical steps that can be better demonstrated by video than by text. Here, we document the detailed steps for these procedures by both video and written protocol. We also briefly discuss different transplantation models: syngeneic, allogeneic, syngeneic autoimmune, and allogeneic autoimmune.

Transplantation of isolated islets has been proposed to be a potential treatment for type 1 diabetes. Here we describe a method to isolate islets from mouse pancreata and transplant them to the subcapsular space of the kidney.

Murin Islet İzolasyon ve subkapsüler Böbrek Nakli Bir Yöntem

Since the early pioneering work of Ballinger and Reckard demonstrating that transplantation of islets of Langerhans into diabetic rodents could normalize ...

Tıp

Murine Cervical Heart Transplantation Model Using a Modified Cuff Technique

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are available to perform mechanistic in vivo studies. While heart transplantation models using a suture technique were first successfully developed in rats, the translation into an equally widespread used murine equivalent was never achieved due the technical complexity of the microsurgical procedure. In contrast, non-suture cuff techniques, also developed initially in rats, were successfully adapted for use in mice1-3. This technique for revascularization involves two major steps I) everting the recipient vessel over a polyethylene cuff; II) pulling the donor vessel over the formerly everted recipient vessel and holding it in place with a circumferential tie. This ensures a continuity of the endothelial layer, short operating time and very high patency rates4.
Using this technique for vascular anastomosis we performed more than 1,000 cervical heart transplants with an overall success rate of 95%. For arterial inflow the common carotid artery and the proximal aortic arch were anastomosed resulting in a retrograde perfusion of the transplanted heart. For venous drainage the pulmonary artery of the graft was anastomosed with the external jugular vein of the recipient5.
Herein, we provide additional details of this technique to supplement the video.

The murine cervical heart transplantation model is well suited for immunological as well as ischemia reperfusion injury studies. We modified the procedure using a non-suture cuff technique and performed more than 1,000 successful transplants with this approach.
Herein, we provide additional details of this technique to supplement the video.

Modifiye Manşet Tekniği Sıçangiller Servikal Kalp Nakli Modeli

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are ...

Technique of Subnormothermic Ex Vivo Liver Perfusion for the Storage, Assessment, and Repair of Marginal Liver Grafts

The success of liver transplantation has resulted in a dramatic organ shortage. In most transplant regions 20-30% of patients on the waiting list for liver transplantation die without receiving an organ transplant or are delisted for disease progression. One strategy to increase the donor pool is the utilization of marginal grafts, such as fatty livers, grafts from older donors, or donation after cardiac death (DCD). The current preservation technique of cold static storage is only poorly tolerated by marginal livers resulting in significant organ damage. In addition, cold static organ storage does not allow graft assessment or repair prior to transplantation.
These shortcomings of cold static preservation have triggered an interest in warm perfused organ preservation to reduce cold ischemic injury, assess liver grafts during preservation, and explore the opportunity to repair marginal livers prior to transplantation. The optimal pressure and flow conditions, perfusion temperature, composition of the perfusion solution and the need for an oxygen carrier has been controversial in the past.
In spite of promising results in several animal studies, the complexity and the costs have prevented a broader clinical application so far. Recently, with enhanced technology and a better understanding of liver physiology during ex vivo perfusion the outcome of warm liver perfusion has improved and consistently good results can be achieved.
This paper will provide information about liver retrieval, storage techniques, and isolated liver perfusion in pigs. We will illustrate a) the requirements to ensure sufficient oxygen supply to the organ, b) technical considerations about the perfusion machine and the perfusion solution, and c) biochemical aspects of isolated organs.

Technique of Subnormothermic Ex Vivo Liver Perfusion for the Storage, Assessment, and Repair of Marginal Liver Grafts

Marginal grafts, such as fatty livers, grafts from older donors, or livers retrieved after cardiac death (DCD) tolerate conventional, cold static storage only poorly. We developed a novel model of subnormothermic ex vivo liver perfusion for preservation, assessment, and repair of marginal liver grafts prior to transplantation.

Subnormothermic Tekniği<em&gt; Ex vivo</em&gt; Karaciğer Depolama, Değerlendirme Perfüzyon ve Marjinal Karaciğer Greft Onarımı

The success of liver transplantation has resulted in a dramatic organ shortage. In most transplant regions 20-30% of patients on the waiting list for ...

Murine Heterotopic Heart Transplant Technique

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become proficient in and utilize this technique, it is now widely used by many laboratories around the world. A significant refinement to the original technique was developed and reported in 2001 by Niimi. Described here are the techniques that have evolved over more than a decade in the hands of three surgeons (Plenter, Grazia, Pietra) in our center. These techniques are now being passed on to a younger generation of surgeons and researchers.
Based largely on the Niimi experience, the procedures used have evolved in the fine details - details which we will endeavor to relate here in such a way that others may be able to use this very useful model. Like Niimi, we have found that a video aid to learning is a priceless resource for the beginner.

The manuscript describes the steps required to perform the heterotopic heart transplant in the mouse.

Mürin Heterotopik Kalp Nakli Tekniği

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become ...

Technique of Porcine Liver Procurement and Orthotopic Transplantation using an Active Porto-Caval Shunt

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation waiting list die without receiving an organ transplant or are delisted due to disease progression. Even after a successful transplantation, rejection and side effects of immunosuppression remain major concerns for graft survival and patient morbidity. 
Experimental animal research has been essential to the success of liver transplantation and still plays a pivotal role in the development of clinical transplantation practice. In particular, the porcine orthotopic liver transplantation model (OLTx) is optimal for clinically oriented research for its close resemblance to human size, anatomy, and physiology. 
Decompression of intestinal congestion during the anhepatic phase of porcine OLTx is important to guarantee reliable animal survival. The use of an active porto-caval-jugular shunt achieves excellent intestinal decompression. The system can be used for short-term as well as long-term survival experiments. The following protocol contains all technical information for a stable and reproducible liver transplantation model in pigs including post-operative animal care.

Experimental animal research plays a pivotal role in the development of clinical transplantation practice. The porcine orthotopic liver transplantation model (OLTx) closely resembles human conditions and is frequently used in clinically oriented research. The following protocol contains all information for a reliable porcine OLTx model using an active porto-caval-jugular shunt.

Active Porto-Caval Shunt kullanarak Domuz Karaciğer Tedarik ve Ortotopik Nakli Tekniği

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation ...

Mouse Model of Alloimmune-induced Vascular Rejection and Transplant Arteriosclerosis

Vascular rejection that leads to transplant arteriosclerosis (TA) is the leading representation of chronic heart transplant failure. In TA, the immune system of the recipient causes damage of the arterial wall and dysfunction of endothelial cells and smooth muscle cells. This triggers a pathological repair response that is characterized by intimal thickening and luminal occlusion. Understanding the mechanisms by which the immune system causes vasculature rejection and TA may inform the development of novel ways to manage graft failure. Here, we describe a mouse aortic interposition model that can be used to study the pathogenic mechanisms of vascular rejection and TA. The model involves grafting of an aortic segment from a donor animal into an allogeneic recipient. Rejection of the artery segment involves alloimmune reactions and results in arterial changes that resemble vascular rejection. The basic technical approach we describe can be used with different mouse strains and targeted interventions to answer specific questions related to vascular rejection and TA.

We describe a protocol for aortic interposition grafting in mice. The goal of the protocol is to provide a model with which to study pathological processes and therapeutic strategies relevant to alloimmune reactions in arteries and the resultant arterial changes that contribute to organ transplant failure.

Aloimmün kaynaklı Vasküler Reddi ve Nakli arterioskleroz Fare Modeli

Vascular rejection that leads to transplant arteriosclerosis (TA) is the leading representation of chronic heart transplant failure. In TA, the immune ...

Normothermic Ex Vivo Kidney Perfusion for the Preservation of Kidney Grafts prior to Transplantation

Kidney transplantation has become a well-established treatment option for patients with end-stage renal failure. The persisting organ shortage remains a serious problem. Therefore, the acceptance criteria for organ donors have been extended leading to the usage of marginal kidney grafts. These marginal organs tolerate cold storage poorly resulting in increased preservation injury and higher rates of delayed graft function. To overcome the limitations of cold storage, extensive research is focused on alternative normothermic preservation methods.
Ex vivo normothermic organ perfusion is an innovative preservation technique. The first experimental and clinical trials for ex vivo lung, liver, and kidney perfusions demonstrated favorable outcomes.
In addition to the reduction of cold ischemic injury, the method of normothermic kidney storage offers the opportunity for organ assessment and repair. This manuscript provides information about kidney retrieval, organ preservation techniques, and isolated ex vivo normothermic kidney perfusion (NEVKP) in a porcine model. Surgical techniques, set up for the perfusion solution and the circuit, potential assessment options, and representative results are demonstrated.

Normothermic Ex Vivo Kidney Perfusion for the Preservation of Kidney Grafts prior to Transplantation

The severe organ shortage has resulted in increased use of marginal kidney grafts for transplantation. This has triggered interest in alternative storage methods, since marginal grafts especially tolerate cold storage poorly. The technique of normothermic ex vivo kidney perfusion (NEVKP) represents a novel preservation method for kidney grafts prior to &#160;transplantation.

Normotermik<em&gt; Ex Vivo</em&gt; Böbrek Koruma Böbrek Nakli Perfüzyon önce greftler

Kidney transplantation has become a well-established treatment option for patients with end-stage renal failure. The persisting organ shortage remains a ...

An &#8220;All-laser&#8221; Endothelial Transplant

The &#8220;all laser&#8221; assisted endothelial keratoplasty is a procedure that is performed with a femtosecond laser used to cut the donor tissue at an intended depth, and a near infrared diode laser to weld the corneal tissue. The proposed technique enables to reach the three main goals in endothelial keratoplasty: a precise control in the thickness of the donor tissue; its easy insertion in the recipient bed and a reduced risk of donor lenticule dislocation. The donor cornea thickness is measured in the surgery room with optical coherence tomography (OCT), in order to correctly design the donor tissue dimensions. A femtosecond laser is used to cut the donor cornea. The recipient eye is prepared by manual stripping of the descemetic membrane. The donor endothelium is inserted into a Busin-injector, the peripheral inner side is stained with a proper chromophore (a water solution of Indocyanine Green) and then it is pulled in the anterior chamber. The transplanted tissue is placed in the final and correct location and then diode laser welding is induced from outside the eyeball. The procedure has been performed on more than 15 patients evidencing an improvement in surgery performances, with a good recovery of visual acuity and a reduced donor lenticule dislocation event.

An &amp;#8220;All-laser&amp;#8221; Endothelial Transplant

An all laser procedure is proposed in the endothelial transplant. The surgical technique is based on the use of a femtosecond laser to prepare the donor tissue. Laser welding technique is then used to secure the donor endothelium in the correct position.

Bir &quot;All-lazer&quot; Endotelyal Nakli

The &#8220;all laser&#8221; assisted endothelial keratoplasty is a procedure that is performed with a femtosecond laser used to cut the donor tissue at an ...

Mouse Model for Pancreas Transplantation Using a Modified Cuff Technique

Mouse models have several advantages in transplantation research, including easy handling, a variety of genetically well-defined strains, and the availability of the widest range of molecular probes and reagents to perform in vivo as well as in vitro studies. Based on our experience with various murine transplantation models, we developed a heterotopic pancreas transplantation model in mice with the intent to analyze mechanisms underlying severe ischemia reperfusion injury-associated early graft damage. In contrast to previously described techniques using suture techniques, herein we describe a new procedure using a non-suture cuff technique. 
In recent years, we have performed more than 300 pancreas transplantations in mice with an overall success rate of &gt;90%, a success rate never described before in mouse pancreas transplantation. The backbone of this non-suture cuff technique for graft revascularization consists of two major steps: (I) pulling the recipient vessel over a polyethylene/polyamide cuff and fixing it with a circumferential ligature, and (II) placing the donor vessel over the everted recipient vessel and fixing it with a second circumferential ligature. The resultant continuity of the endothelial layer results in less thrombogenic lesions with high patency rates and, finally, high success rates.
In this model, arterial anastomosis is achieved by pulling the abdominal aorta of the donor graft over the everted common carotid artery of the recipient animal. Venous drainage of the graft is achieved by pulling the portal vein of the graft over the everted external jugular vein of the recipient. This manuscript provides details and crucial steps of the organ recovery and organ implantation procedures, which will allow researchers with microsurgical skills to perform the transplantation successfully in their laboratories.

Among abdominal solid organ transplantation, pancreatic grafts are prone to develop severe ischemia reperfusion injury-associated graft damage, leading eventually to early graft loss. This protocol describes a model of murine pancreas transplantation using a non-suture cuff technique, ideally suited for analyzing these early, deleterious damages.

Fare modeli değiştirilmiş manşet tekniği kullanarak pankreas nakli için

Mouse models have several advantages in transplantation research, including easy handling, a variety of genetically well-defined strains, and the ...

Murine Cervical Aortic Transplantation Model using a Modified Non-Suture Cuff Technique

With the introduction of powerful immunosuppressive protocols, distinct advances are possible in the prevention and therapy of acute rejection episodes. However, only minor improvement in the long-term results of transplanted solid organs could be observed over the past decades. In this context, chronic allograft vasculopathy (CAV) still represents the leading cause of late organ failure in cardiac, renal and pulmonary transplantation.
Thus far, the underlying pathogenesis of CAV development remains unclear, explaining why effective treatment strategies are presently missing and emphasizing a need for relevant experimental models in order to study the underlying pathophysiology leading to CAV formation. The following protocol describes a murine heterotopic cervical aortic transplantation model using a modified non-suture cuff technique. In this technique, a segment of the thoracic aorta is interpositioned in the right common carotid artery. With the use of the non-suture cuff technique, an easy to learn and reproducible model can be established, minimizing the possible heterogeneity of sutured vascular micro anastomoses.

Here, we present a protocol of heterotopic aortic transplantation in mice using the non-suture cuff technique in a cervical murine model. This model can be used to study the underlying pathology of chronic allograft vasculopathy (CAV) and can help evaluate new therapeutic agents in order to prevent its formation.

Modifiye Dikişsiz Manşet Tekniği Kullanılarak Murine Servikal Aort Transplantasyon Modeli

With the introduction of powerful immunosuppressive protocols, distinct advances are possible in the prevention and therapy of acute rejection episodes. ...