The authors would like to thank the many individuals who have worked on and perfected this technique and have been responsible for the generation of many manuscripts both in this lab and others. This work was supported by National Institutes of Health Grant EY12707 (PMS) and an unrestricted grant from Research to Prevent Blindness to Department of Ophthalmology.

NOT: Bu prosedürde kullanılan tüm hayvanlar Vizyon Araştırma Derneği ve Göz açıklamada Oftalmik ve Vizyon Araştırma Hayvanların Kullanımı yanı sıra Saint Louis Üniversitesi&#39;nde hayvan gözetim komitesi tarafından belirlenen esaslara uygun olarak tedavi edilir. NOT: Tüm cerrahi aletler ve çözümleri göz mikrobik enfeksiyonu sınırlamak için ameliyattan önce sterilize edilir. Bu hayvanların bu prosedürden bazı ağrı yaparken, biz analjezik istihdam yok unutulmamalıdır. Tüm analjezikler, anti-inflamatuar ve kornea nakli tepkiler iltihabı içeren bu yana, anti-inflamatuar ilaçların kullanılması kornea greft yetmezliği katılan hangi faktörlerin belirlemek için yeteneği taviz verecek çünkü bu nedeni. 1. Anestezi <ol style="margin-left: 40px;"><li> Ketamin (86 İP enjeksiyonları, genel anestezi altında donör ve alıcı fareler yerleştirin.98 mg / kg) ve ksilazin (13.04 mg / kg). </li><li> Genellikle bir saat 30 dk sürer işlem boyunca anestezi altında alıcı fare tutun. Sonuç olarak, sürekli bilinç kazanmak herhangi bir işaret için fareyi izlemek. </li><li> Hayvan kuruluğunu önlemek için uyuşturulduktan sonra ameliyat olmaz göze puralube merhem sürün. </li></ol> 2. Kornea Tırnak <ol style="margin-left: 40px;"><li> Donör kornea düğmesine elde edilmesi. <ol><li> Hayvan tamamıyla anestezi sonra,% 1 tropikamid ve% 2.5 fenilefrin hidroklorid göz damlası, bir çift uygulama yoluyla yeterli midriasis elde eder. </li><li> Yatay sağlam bir hareketli destek üzerine yerleştirilen bir gemide donör hayvanın başını yerleştirin. Göz tüm işlem boyunca yatay konumda olduğundan emin olmak için boyunda bant şeridi ile baş sabitleyin. </li><li> Üzerinden, Ucu metil mavisi ile boyanmış, bir 2 mm çapında trepan kullanılarak,Santral kornea greft sıralanıyor. </li><li> Keskin bir bıçak ile, kornea nüfuz ve donör endotel ve altta yatan merceğine zarar şansını azaltmak için bunu derinleştirmek için ön kamaraya Healon® enjekte. </li><li> Vannas makas ile donör graft pulu ve kullanılana kadar Hanks dengeli tuz çözeltisi içeren bir kabın içine koyun. </li><li> Donör greft çıkarılmamış sonra, CO 2 inhalasyon yoluyla donör fare euthanize. </li></ol></li><li> Greft yatağı hazırlamak. <ol><li> Alıcı için 2.1.2&#39;de yoluyla 2.1.1 açıklandığı gibi aynı adımları tekrarlayın. </li><li> 1.5 mm çaplı trefin kullanarak, alıcı greft özetlemektedir. </li><li> Keskin bir bıçak ile, kornea nüfuz ve altta yatan merceğine zarar şansını azaltmak için bunu derinleştirmek için ön kamaraya Healon® enjekte. </li><li> Vannas makas kullanılarak alıcıdan özetlenen santral kornea düğmesini çıkarın ve atın. </li></ol></li><li> Sütüreaşı <ol><li> Alıcının kornea yatak greft üzerinde donör kornea yerleştirin. Yeterli Healon lens ile doğrudan temas yoluyla zararlardan donör endotel hücreleri korumak için donör kornea altında olduğundan emin olun. </li><li> Süper ince uçlu microforceps kullanarak, daha sonra kapalı kravat, alıcının tarafında tam kalınlığının% 90 derinliğe sahip donör yoluyla, donör tarafına 11-0 naylon dikiş ilk ısırık yerleştirin. </li><li> Kornea sabitlenebilir sonra, kornea 8-10 toplam dikiş sahiptir ve verici kornea güvenli dizilmiş ve alıcı korneal aşı yatağa bağlanmasına imkan verecek şekilde midcardinal sütürler gerçekleştirin. </li></ol></li><li> Anteryor odacığını derinleştirme <ol><li> Ön kamaraya HBSS veya hava kabarcığı enjekte ederek ön kamara derinleştirmek ve yavaşça selüloz bir süngerle sızıntı kornea greft bütünlüğünü kontrol edin. NOT: Ön kamara reforme edilemez Eğer c yüksek bir olasılık varnakledilen korneanın gelecekteki değerlendirmeler çok zor hale getirecek ve aynı zamanda potansiyel donör kornea endotel disfonksiyonu ve böylece greft yetmezliğine yol açacaktır ataract. </li></ol></li><li> Nihai değerlendirme <ol><li> Öğrenci yuvarlak ve ön kamara derinliği normal olduğunu belirlemek için göz gözlemleyin. NOT: Bu iris dikilmesi sırasında zarar görmüş olduğunu gösterir ve böylece greft teknik bir arıza olarak kabul edilir yuvarlak öğrenci değilse. </li><li> Göze antibiyotik merhem sürün. İsteğe bağlı: 7-0 ipek sütür ile göz kapağını kapatın. </li><li> Tamamen uyanık ve daha sonra tek tek ameliyat sonrası iki gün en az onları barındıracak kadar fareler gözlemleyin. </li></ol></li></ol> 3. Dikiş Kaldırma <ol style="margin-left: 40px;"><li> Kapak dikiş kullanılır Bu durumda, yukarıda tarif edildiği gibi fareler anestezi ve 48 saatte kapak dikiş çıkarın. </li><li> Postoperatif 7. günde fareler uyutmak. Sütür çıkarınes kornea grefti güvence. Dikişler ve hayvan tamamen uyanmış sonra, kendi kafesine geri. </li></ol> 4. Klinik Değerlendirme <ol style="margin-left: 40px;"><li> (Ön kamarada kan) katarakt (lens bulanıklıklarına), hifemayı dahil komplikasyon endikasyonları göz, uygun derinlik, ya da kornea önemli donukluk değil bir ön kamara inceleyin. &quot;Taviz&quot; olarak bu komplikasyonların arttığını olanlar düşünün ve CO 2 inhalasyon yoluyla onları euthanize. <ol><li> Unanesthetized fareler üzerinde tüm sınavları gerçekleştirmek. Öte yandan gözün daha iyi bir görünüm sağlamak için göz proptose böylece bir yandan böylece fareyi frenleyici ile fare tuşunu basılı tutun. Gözlemler tamamlandıktan sonra, kendi kafesine hayvan dönün. </li></ol></li><li> Tedavi grupları ile yabancı bir gözlemci kornea greft belirtileri kornealar 2-3 kez bir hafta nakledilen değerlendirebilir mırejeksiyon atakları veya kornea greft yetmezliği. Cerrahi mikroskop veya bu gözlemler için bir yatay yarık-lamba biyomikroskop kullanın. <ol><li> Aşağıdaki gibi ölçek tanımlanır 5&#39;e kadar 0 bir ölçek kullanılarak donukluk her korneayı değerlendirin: <ol><li> Donukluk emaresi var bu kornealar için 0 puan atayın. </li><li> Minimal yüzeyel opaklık gösterir bu kornealara 1 puan atayın. </li><li> Hafif ve derin donukluk görüntülemek ancak altta yatan öğrenci ve iris hala discernable kornealara 2 puan atayın. </li><li> Iris öğrenci marjlarının haricinde detaylı olarak görülemez burada stromal donukluk görüntülemek korneaya 3 puan atayın. </li><li> Yoğun stromal donukluk gösterilecek ve altta yatan yapılar görülebilir eğer kornea 4 puan atayın. </li><li> Tamamen gizlenmiş öğrenci ve iris ile tam donukluk ve yoğun stromal ödem, görüntülemek korneaya 5 puan atayın. </li></ol></li><li><html> Aynı zamanda, damar infiltrasyon (neovaskülarizasyon) Bunun için 1 ile 8 arasında bir derecelendirme ölçeği kullanarak derecesi her kornea değerlendirilmesi 4 eşit kadranın oluşan olarak kornea görebilir ve bu kadranın her kan damarlarının miktarının belirlenmesi Bu kadranda geniş vaskularizasyonu 2 0 (gemiler) o irade aralığı puan. Son neovaskülarizasyon puanı hesaplamak için her kadrandan bireysel puanları ekleyin. </li><li> Onlar 5 hafta kadar puan süre üst üste iki gözlem için 3 puan var sanki akut reddedilen kornealar sınıflandırır. </li><li> Olan korneaları 5 hafta netti fareler sınıflandırmak ama tabi geç dönem kornea alograf sahip olarak,&gt; 45 gün üst üste iki kez puan için 3 puan ile, engrafmanda sonrası zamanlarda donuklaşmalarını geliştirmek. Greft sağ kalımı analiz Kaplan-Meier sağkalım eğrileri kullanın. </li></ol></li></ol> Modelin 5. Manipülasyon in sol: 40px; &quot;&gt; <li> Dalaktan tek hücre hazırlanması. <ol><li> Tek hücre hazırlamak için, öncelikle donör fare euthanize. Sonra, dalak çıkarın. </li><li> Bir hücre süzgecinden dalak yerleştirin ve 3 ml şırınga bir şırınga pistonu ile bozabilir. </li><li> Hanks Dengelenmiş Tuz Çözeltisi, 10 ml hücreleri ve tekrar süspansiyon yıkayın. </li><li> Hücre süspansiyonu 10 ul çıkarın ve% 0.4 tripan 10 ul mavi ilave edilir ve karıştırılır. Merkezi ızgara hücreleri hemositometrede bu ekleyin ve saymak. Tüp içinde hücre sayısı ve hücre sayısı x 10 4 x 2 (tripan mavi seyreltme faktörü) 10 (tüp içinde hacmi) X. </li></ol></li><li> Ön kamaraya enjeksiyonu. <ol><li> Daha önce tarif edildiği gibi fareler anestezi. </li><li> Bir mikroskop kullanarak enjeksiyonları gerçekleştirin. Her intrakameral enjeksiyon için, 10 6, 0.005 ml hacim içinde dalak hücreleri ve bir 33 G iğne ile donatılmış 0.25 mi mikrolitre şırınga kullanın. NOT: Diğer manipülasyonlarModelin, özel bir faktör ortotopik kornea alograft ameliyat sonrası oynayabilir rolü belirlemek için ya da antagonistleri ya da agonistleri olarak hareket eden reaktif olan hayvanların muamele etmek suretiyle gerçekleştirilebilir. </li></ol></li></ol>

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R., Archer, B., Ambati, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vascular+Endothelial+Growth+Factor+Receptor+1+morpholino+increases+graft+survival+in+a+murine+penetrating+keratoplasty.">Vascular Endothelial Growth Factor Receptor 1 morpholino increases graft survival in a murine penetrating keratoplasty.</a> Invest. Ophthalmol. Vis. Sci. 53, 8458-8471 (2012).</li><li>Kim, H. K., Choi, J. A., Uehara, H., Zhang, X., Ambati, B. K., Cho, Y. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Presurgical+corticosteroid+treatment+improves+corneal+transplant+survival+in+mice.">Presurgical corticosteroid treatment improves corneal transplant survival in mice.</a> Cornea. 32, 1591-1598 (2013).</li><li>Yamazoe, K., Yamazoe, K., Shimazaki-Den, S., Shimazaki, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prognostic+factors+for+corneal+graft+recovery+after+severe+corneal+graft+rejection+following+penetrating+keratoplasty.">Prognostic factors for corneal graft recovery after severe corneal graft rejection following penetrating keratoplasty.</a> BMC Ophthalmol. 13, 5 (2013).</li><li>Panda, A., Vanathi, M., Kumar, A., Dash, Y., Priya, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Corneal+graft+rejection.">Corneal graft rejection.</a> Surv. Ophthalmol. 52, 375-396 (2007).</li><li>Patel, S. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Graft+survival+and+endothelial+outcomes+in+the+new+era+of+endothelial+keratoplasty.">Graft survival and endothelial outcomes in the new era of endothelial keratoplasty.</a> Exp. Eye Res. 95, 40-47 (2012).</li><li>Anshu, A., Price, M. O., Tan, D. T., Price, F. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endothelial+keratoplasty:+a+revolution+in+evolution.">Endothelial keratoplasty: a revolution in evolution.</a> Surv. Ophthalmol. 57, 236-252 (2013).</li></ol>

The authors have no competing financial interests.

Burada açıklanan kornea transplantasyonu mürin modeli faktörler en iyi, hem reddi 15,17,18,20, 26-30 ve kornea kabul 21-25 ile ilişkili olduğunu prediktif olan bir modelinde insan kornea alograft reddini çalışma araştırmacı sağlar allogrefti. Hastalar tedavi etmek ya da önlemek ya da red 31 &#39;e ya da topikal veya sistemik steroid tedavisi verilmiş olup, burada insan kornea transplantasyonu, farklı olarak, bu model, tipik olarak böyle bir tedavinin yokluğunda ret alog...

Kornea nakli insanlarda yapılan transplantasyon en başarılı ve yaygın türlerinden biridir. Bu ameliyatı yapılır neden nedenleri yaralanma, bulaşıcı hastalık, 1 ya da bulaşıcı olmayan kornea hastalığının 2 diğer formları bir sonucudur. Amerika&#39;nın Göz Bankası Derneği rakamlar 46.000 (: restoresight.org/eye_banks/eye_banks.html at web sitesine bakınız) 2011 yılında yapılmıştır göstermektedir. Başarısının bir göstergesi allojenik kornea greft için bir yıllık başarısızlık oranları 10 ila% 15 ve başarı% 70 3-8 aşan 5 yıl olarak değişir olmasıdır. Birçok araştırma göstermiştir ki gibi, kornea allograftlann başarısı doğrudan göz immünolojik olarak ayrıcalıklı bir site olduğunu gerçeği ile ilgilidir. Bir bağışıklık ayrıcalık site olarak kornea durumundan sorumlu faktörler suppres kornea tarafından üretilen korneada ikisi de kan ve lenf damarlarının eksikliği, antijen sağlayan hücreler bir nispi olmaması, faktörlerini içeririmmün efektör fonksiyonlar 9-15 s, MHC düşük sentezleme 16 antijenleri ve FasL 17-20 ifadesi. Ancak, başarı için bu greft predispozan bu faktörlerin rağmen, onlar ret 3-7 tabi yoktur. Sonuç olarak, bu ret arabuluculuk bu mekanizmaları anlamak yanı sıra reddini önlemek için çeşitli terapiler test kritik önem taşımaktadır. Bu amaçla, biz burada kontrollü bir deney ortamında kornea nakli incelemek için 20 yıldır kullanılmakta olan kornea naklinin bir fare modeli tarif. Transplantasyon tepkiler nihai nakledilen doku başarısız veya başarılı olup olmadığını belirleyecek uyum içinde çalışan birçok farklı faktörü içeren bu yana, herhangi bir in vitro modelde bu faktörlerin önemini anlamak mümkün değildir. Sonuç olarak, sağlam hayvanları ile yapılan çalışmalarda başarı veya failu ya önemli olan hangi faktörlerin belirlenmesi gerekmektedirnakledilen dokuların yeniden. Diğer hayvan türlerinin kornea nakli incelemek için kullanılmış olsa da, diğer türler ile karşılaştırıldığında, sıçangil modeli birçok avantajı vardır. İlki, belli transjenleri veya fonksiyon nakli daha iyi ele alınabilir spesifik immünolojik faktörlerin ifadesini yoksun gen hedef alınmıştır farelerin birçok suşları varlığıdır. Buna ek olarak, hayvan bir çok diğer türler için mevcut olmayan fareler için spesifik olan ve bir çok tepkin madde (faktörlerini nötralize rekombinant faktörler ve antikorlar, hem de) vardır. Çünkü bu faktörlerin varlığı, bu model akut korneal allogreft cevapları 15, 17,18,20 -29 dahil ilgili faktörleri belirlemek için yaygın olarak kullanılır olmuştur. Ayrıca, kornea nakli yer faktörlerinin çoğu, diğer dokuların naklinden işlevsel olduğu da bilinmektedir.

<table><tbody><tr><td>Zeiss Surgical Microscope</td><td>Zeiss</td><td>Rebuilt</td><td></td></tr><tr><td>1 ml Syringe</td><td>BD</td><td>305122</td><td></td></tr><tr><td>3 ml Syringe</td><td>BD</td><td>309657</td><td></td></tr><tr><td>10 ml Syringe</td><td>BD</td><td>309602</td><td></td></tr><tr><td>Vannus Scissors</td><td>Stortz</td><td>E-3387</td><td></td></tr><tr><td>11-0 Sutures</td><td>Alcon</td><td>717939M</td><td></td></tr><tr><td>Trephine 2.0 mm</td><td>Katena</td><td>K 2-7520</td><td></td></tr><tr><td>Trephine 1.5 mm</td><td>Katena</td><td>K 2-7510</td><td></td></tr><tr><td>Tricaine Hydrochloride 0.5%</td><td>Alcon</td><td>NDC 0065-0741-12</td><td></td></tr><tr><td>Healon</td><td>Abbott</td><td>Healon OVD</td><td></td></tr><tr><td>Forceps</td><td>FST</td><td>11251-20</td><td></td></tr><tr><td>7-0 Sutures</td><td>Alcon</td><td>8065</td><td></td></tr><tr><td>2.5% Phenylephrine HCl</td><td>Alcon</td><td>NDC 61314-342-02</td><td></td></tr><tr><td>1% Tropicamide</td><td>Bausch &amp;amp; Lomb</td><td>NDC-24208-585-59</td><td></td></tr><tr><td>Hamilton Syringe</td><td>Hamilton</td><td>7654-01</td><td></td></tr><tr><td>33 gauge needle</td><td>Hamilton</td><td>90033</td><td></td></tr><tr><td>Cell Strainer (100 &amp;mu;m nylon)</td><td>BD Falcon</td><td>352360</td><td></td></tr><tr><td>Hemocytometer</td><td>Cardinal Health</td><td>B3175</td><td></td></tr><tr><td>Trypan Blue</td><td>Sigma</td><td>T8154</td><td></td></tr></tbody></table>

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.

Kornea naklinin fare modeli başarıyla kornea allogreft reddi 19-23 ve kornea allogreft kabul 13, 15,16,18, 24-27 hem mekanizmasını karakterize etmek için 20 yılı aşkın süredir kullanılmaktadır. Bu model FasL&#39;yi eksikliği hayvanlarda, kornea allogreft kabul FasL ifade önemini kurmak için kullanılan kornea alograftlarını 15 kabul etmek mümkün değildi. Aynı zamanda, vasküler endotelyal büyüme faktörü reseptörü 1 morfolino tedavisi, korneal aşı sağkalımı...

Watch this Scientific Journal Video about Murine Corneal Transplantation: A Model to Study the Most Common Form of Solid Organ Transplantation at JoVE.com

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

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.

Sıçangiller Kornea Nakli: Katı Organ Nakli En Yaygın Formu Eğitim için Bir Model

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

murine-corneal-transplantation-model-to-study-most-common-form-solid

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9.7K Görüntüleme.  Saint Louis University. 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.

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

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 ...

Tıp

Mouse Kidney Transplantation: Models of Allograft Rejection

Rejection of the transplanted kidney in humans is still a major cause of morbidity and mortality. The mouse model of renal transplantation closely replicates both the technical and pathological processes that occur in human renal transplantation. Although mouse models of allogeneic rejection in organs other than the kidney exist, and are more technically feasible, there is evidence that different organs elicit disparate rejection modes and dynamics, for instance the time course of rejection in cardiac and renal allograft differs significantly in certain strain combinations. This model is an attractive tool for many reasons despite its technical challenges. As inbred mouse strain haplotypes are well characterized it is possible to choose donor and recipient combinations to model acute allograft rejection by transplanting across MHC class I and II loci. Conversely by transplanting between strains with similar haplotypes a chronic process can be elicited were the allograft kidney develops interstitial fibrosis and tubular atrophy. We have modified the surgical technique to reduce operating time and improve ease of surgery, however a learning curve still needs to be overcome in order to faithfully replicate the model. This study will provide key points in the surgical procedure and aid the process of establishing this technique.

Here, we present a protocol to study the immunology of rejection. The surgical model presented reports a short operating time and a concise technique. Depending on the donor-recipient strain combination, the transplanted kidney may develop acute cellular rejection or chronic allograft damage, defined by interstitial fibrosis and tubular atrophy.

Fare Böbrek Transplantasyonu: Allogreft Reddi Modelleri

Rejection of the transplanted kidney in humans is still a major cause of morbidity and mortality. The mouse model of renal transplantation closely ...

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.

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 ...

Murine Full-thickness Skin Transplantation

Murine full-thickness skin transplantation is a well-established in vivo model to study alloimmune response and graft rejection. Despite its limited application to humans, skin transplantation in mice has been widely employed for transplantation research. The procedure is easy to learn and perform, and it does not require delicate microsurgical techniques nor extensive training. Moreover, graft rejection in this model occurs in a very reproducible immunological reaction and is easily monitored by direct inspection and palpation. In addition, secondary skin transplantation with donor-matched or third-party skin grafts can be performed on more complex transplant models as an alternative and uncomplicated method to assess donor-specific tolerance. The complications are low and are in general limited to anesthesia overdose or respiratory distress after the procedure. Graft failure, on the other hand, occurs commonly as a result of poor preparation of the graft, incorrect positioning in the graft bed, or inappropriate placement of the bandage. In this article, we present a protocol for full-thickness skin transplantation in mice and describe the important steps necessary for a successful procedure.

Murine full-thickness skin transplantation is a well-established model to study rejection in an alloimmune setting. Here, we provide a tutorial of each step involved in performing a BALB/c--&#62;C57BL/6 full-thickness skin transplant.

Mürin Tam kat Cilt Nakli

Murine full-thickness skin transplantation is a well-established in vivo model to study alloimmune response and graft rejection. Despite its limited ...

İmmünoloji ve Enfeksiyon

A Novel Microsurgical Model for Heterotopic, En Bloc Chest Wall, Thymus, and Heart Transplantation in Mice

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression must be pursued. Employing vascularized bone marrow transplantation and co-transplantation of the thymus have shown promise in this regard in various animal models.1-11 Vascularized bone marrow transplantation allows for the uninterrupted transfer of donor bone marrow cells within the preserved donor microenvironment, and the incorporation of thymus tissue with vascularized bone marrow transplantation has shown to increase T-cell chimerism ultimately playing a supportive role in the induction of immune regulation. The combination of solid organ and vascularized composite allotransplantation can uniquely combine these strategies in the form of a novel transplant model. Murine models serve as an excellent paradigm to explore the mechanisms of acute and chronic rejection, chimerism, and tolerance induction, thus providing the foundation to propagate superior allograft survival strategies for larger animal models and future clinical application. Herein, we developed a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique. The experience in syngeneic and allogeneic transplant settings is described for future broader immunological investigations via an instructional manuscript and video supplement.

To study combined solid organ and vascularized composite allotransplantation, we describe a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique.

Farelerde A Novel Mikrocerrahi Heterotopik Modeli, En Bloku Göğüs Duvarı, Thymus ve Kalp Nakli

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression ...

Orthotopic Hind Limb Transplantation in the Mouse

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and translational research in the field of transplantation medicine. A vast array of reagents is available exclusively in this setting, including mono- and polyclonal antibodies for both diagnostic and interventional applications. In addition, a vast number of genotyped, inbred, transgenic, and knock out strains allow detailed investigation of the individual contributions of humoral and cellular components to the complex interplay of an immune response and make the mouse the gold standard for immunological research. 
Vascularized Composite Allotransplantation (VCA) delineates a novel field of transplantation using allografts to replace "like with like" in patients suffering traumatic or congenital tissue loss. This surgical methodological protocol shows the use of a non-suture cuff technique for super-microvascular anastomosis in an orthotopic mouse hind limb transplantation model. The model specifically allows for comparison between established paradigms in solid organ transplantation with a novel form of transplants consisting of various different tissue components. Uniquely, this model allows for the transplantation of a viable vascularized bone marrow compartment and niche that have the potential to exert a beneficial effect on the balance of immune acceptance and rejection. This technique provides a tool to investigate alloantigen recognition and allograft rejection and acceptance, as well as enables the pursuit of functional nerve regeneration studies to further advance this novel field of transplantation.

This novel model for orthotopic hind limb transplantation in the mouse, applying a non-suture cuff technique for super-microvascular anastomosis, provides a powerful tool for in&#160;vivo mechanistic immunological research related to vascularized composite allotransplantation (VCA).

Fare Ortotopik Hind Ekstremite nakli

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and ...

An Immunological Model for Heterotopic Heart and Cardiac Muscle Cell Transplantation in Rats

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications have been reported since the first description in 1964. After 30 years of performing heterotopic heart transplantation in rats, we have developed a simplified surgical approach, which can be easily taught and performed without further surgical training or background.
After dissection of the ascending aorta and the pulmonary artery and ligation of superior and inferior caval and pulmonary veins, the donor heart is harvested and subsequently perfused with ice-cold saline solution supplemented with heparin. After clamping and incising the recipient abdominal vessels, the donor ascending aorta and pulmonary artery are anastomosed to the recipient abdominal aorta and inferior vena cava, respectively, using continuous running sutures.
Depending on different donor-recipient combinations, this model allows analyses of either acute or chronic rejection of allografts. The immunological significance of this model is further enhanced by a novel approach of in-ear injection of vital cardiac muscle cells and subsequent analysis of draining cervical lymphatic tissue.

We describe a model of heterotopic abdominal heart transplantation in rats, implying modifications of current strategies, which lead to a simplified surgical approach. Additionally, we describe a novel rejection model by in-ear injection of vital cardiac muscle cells, allowing further transplant immunological analyses in rats.

Sıçanlarda Heterotopik Kalp ve Kardiyak Kas Hücre Nakli için İmmünolojik Model

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications ...

A Modified Murine Heterotopic Heart Transplant Protocol Matching Contemporary Standards of Aseptic Technique, Anesthesia, and Analgesia

The development of experimental models of cardiac transplantation in animals has contributed to many advances in the fields of immunology and solid organ transplantation. While the heterotopic vascularized murine cardiac transplantation model was initially utilized in studies of graft rejection using combinations of mismatched inbred mouse strains, access to genetically modified strains and therapeutic modalities can provide powerful new preclinical insights. Fundamentally, the surgical methodology for this technique has not changed since its development, especially with respect to important factors such as aseptic technique, anesthesia, and analgesia, which make material impacts on postsurgical morbidity and mortality. Additionally, improvements in perioperative management are expected to provide improvements in both animal welfare and experimental outcomes. This paper reports upon a protocol developed in collaboration with a subject matter expert in veterinary anesthesia and describes the surgical technique with an emphasis on perioperative management. Additionally, we discuss the implications of these refinements and provide details on troubleshooting critical surgical steps for this procedure.

The present paper describes a modified technique for heterotopic vascularized cardiac transplantation with updated aseptic technique, analgesia, and anesthesia.

Çağdaş Aseptik Teknik, Anestezi ve Analjezi Standartlarına Uygun Modifiye Murin Heterotopik Kalp Nakli Protokolü

The development of experimental models of cardiac transplantation in animals has contributed to many advances in the fields of immunology and solid organ ...

A Porcine Corneal Endothelial Organ Culture Model Using Split Corneal Buttons

Experimental research on corneal endothelial cells is associated with several difficulties. Human donor corneas are scarce and rarely available for experimental investigations as they are normally needed for transplantation. Endothelial cell cultures often do not translate well to in vivo situations. Due to the biostructural characteristics of non-human corneas, stromal swelling during cultivation induces substantial corneal endothelial cell loss, which makes it difficult to perform cultivation for an extended period of time. Deswelling agents such as dextran are used to counteract this response. However, they also cause significant endothelial cell loss. Therefore, an ex vivo organ culture model not requiring deswelling agents was established. Pig eyes from a local slaughterhouse were used to prepare split corneal buttons. After partial corneal trephination, the outer layers of the cornea (epithelium, bowman layer, parts of the stroma) were removed. This significantly reduces corneal endothelial cell loss induced by massive stromal swelling and Descemet&#39;s membrane folding throughout longer cultivation periods and improves general preservation of the endothelial cell layer. Subsequent complete corneal trephination was followed by the removal of the split corneal button from the remaining eye bulb and cultivation. Endothelial cell density was assessed at follow-up times of up to 15 days after preparation (i.e., days 1, 8, 15) using light microscopy. The preparation technique used allows a better preservation of the endothelial cell layer enabled by less stromal tissue swelling, which results in slow and linear decline rates in split corneal buttons comparable to human donor corneas. As this standardized organo-typically cultivated research model for the first time allows a stable cultivation for at least two weeks, it is a valuable alternative to human donor corneas for future investigations of various external factors with regards to their effects on the corneal endothelium.

Here, a step-by-step protocol for the preparation and cultivation of porcine split corneal buttons is presented. As this organo-typically cultivated organ culture model shows cell death rates within 15 days, comparable to human donor corneas, it represents the first model allowing long-term cultivation of non-human corneas without adding toxic dextran.

Split Korneal Düğmeleri Kullanarak Bir Domuz Korneal Endotel Organ Kültür Modeli

Experimental research on corneal endothelial cells is associated with several difficulties. Human donor corneas are scarce and rarely available for ...

A Simplified Model for Heterotopic Heart Valve Transplantation in Rodents

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of transplant with the potential to deliver durable heart valves capable of somatic growth with no requirement for anticoagulation. However, the immunobiology of heart valve transplants remains unexplored, highlighting the need for animal models to study this new type of transplant. Previous rat models for heterotopic aortic valve transplantation into the abdominal aorta have been described, though they are technically challenging and costly. For addressing this challenge, a renal subcapsular transplant model was developed in rodents as a practical and more straightforward method for studying heart valve transplant immunobiology. In this model, a single aortic valve leaflet is harvested and inserted into the renal subcapsular space. The kidney is easily accessible, and the transplanted tissue is securely contained in a subcapsular space that is well vascularized and can accommodate a variety of tissue sizes. Furthermore, because a single rat can provide three donor aortic leaflets and a single kidney can provide multiple sites for transplanted tissue, fewer rats are required for a given study. Here, the transplantation technique is described, providing a significant step forward in studying the transplant immunology of heart valve transplantation.

This protocol describes a simple and efficient method for the transplantation of aortic valve leaflets under the renal capsule to allow for the study of alloreactivity of heart valves.