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.

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			<td style="width:167px;">Comments/Description</td>
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			<td height="21" style="height:21px;width:216px;">Zeiss Surgical Microscope</td>
			<td style="width:71px;">Zeiss</td>
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			<td height="21" style="height:21px;">1 mL Syringe</td>
			<td>BD</td>
			<td>305122</td>
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			<td height="21" style="height:21px;">3 mL Syringe</td>
			<td>BD</td>
			<td>309657</td>
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			<td height="21" style="height:21px;">10 mL Syringe</td>
			<td>BD</td>
			<td>309602</td>
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			<td height="21" style="height:21px;width:216px;">Vannus Scissors</td>
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			<td height="21" style="height:21px;width:216px;">11-0 Sutures</td>
			<td style="width:71px;">Alcon</td>
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			<td height="21" style="height:21px;width:216px;">Trephine 2.0mm</td>
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			<td height="21" style="height:21px;width:216px;">Trephine 1.5 mm</td>
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			<td style="width:119px;">K 2-7510</td>
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			<td height="41" style="height:41px;width:216px;">Tricaine Hydrochloride 0.5%</td>
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			<td height="21" style="height:21px;width:216px;">7-0 Sutures</td>
			<td style="width:71px;">Alcon</td>
			<td align="right" style="width:119px;">8065</td>
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			<td height="41" style="height:41px;width:216px;">2.5% Phenylephrine HCl</td>
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			<td height="41" style="height:41px;width:216px;">1% Tropicamide</td>
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			<td height="21" style="height:21px;width:216px;">Hamilton Syringe</td>
			<td style="width:71px;">Hamilton</td>
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			<td height="21" style="height:21px;width:216px;">33 gauge needle</td>
			<td style="width:71px;">Hamilton</td>
			<td align="right" style="width:119px;">90033</td>
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			<td height="41" style="height:41px;width:216px;">Cell Strainer (100 &#956;m nylon)</td>
			<td style="width:71px;">BD Falcon</td>
			<td align="right" style="width:119px;">352360</td>
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			<td height="41" style="height:41px;width:216px;">Hemocytometer</td>
			<td style="width:71px;">Cardinal Health</td>
			<td style="width:119px;">B3175</td>
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			<td height="21" style="height:21px;width:216px;">Trypan Blue</td>
			<td style="width:71px;">Sigma</td>
			<td style="width:119px;">T8154</td>
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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

Research

JoVE Journal

Medicine

9.6K 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

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.

Ortotopik Aort Nakli: Rat Modeli Kronik Vaskülopati Geliştirme Çalışması

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

Tıp

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 ...

Development of Obliterative Bronchiolitis in a Murine Model of Orthotopic Lung Transplantation

Orthotopic lung transplantation in rats was first reported by Asimacopoulos and colleagues in 1971 1. Currently, this method is well accepted and standardized not only for the study of allo-rejection but also between syngeneic strains for examining mechanisms of ischemia-reperfusion injury after lung transplantation. Although the application of the rat and other large animal model 2 contributed significantly to the elucidation of these studies, the scope of those investigations is limited by the scarcity of knockout and transgenic rats. Due to no effective therapies for obliterative bronchiolitis, the leading cause of death in lung transplant patients, there has been an intensive search for pre-clinical models that replicate obliterative bronchiolitis. The tracheal allograft model is the most widely used and may reproduce some of the histopathologic features of obliterative bronchiolitis 3. However, the lack of an intact vasculature with no connection to the recipient's conducting airways, and incomplete pathologic features of obliterative bronchiolitis limit the utility of this model 4. Unlike transplantation of other solid organs, vascularized mouse lung transplants have only recently been reported by Okazaki and colleagues for the first time in 2007 5. Applying the basic principles of the rat lung transplant, our lab initiated the obliterative bronchiolitis model using minor histoincompatible antigen murine orthotopic single-left lung transplants which allows the further study of obliterative bronchiolitis immunopathogenesis6.

Obliterative bronchiolitis is the key impediment to the long-term survival of lung transplant recipients and the lack of a robust preclinical model precludes examining obliterative bronchiolitis immunopathogenesis. Unlike other solid organ transplants, vascularized mouse lung transplantation has only recently been developed. Here we show our independently developed obliterative bronchiolitis model after murine orthotopic single-lung transplantation.

Ortotopik Akciğer Transplantasyonu bir Mürin Modelinde Obliteratif Bronşiolit Gelişimi

Orthotopic lung transplantation in rats was first reported by Asimacopoulos and colleagues in 1971 1. Currently, this method is well accepted and ...

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 ...

Cardiac Catheterization in Mice to Measure the Pressure Volume Relationship: Investigating the Bowditch Effect

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these strategies is to examine their effects on heart function. There are several techniques to measure in vivo cardiac mechanics (e.g., echocardiography, pressure/volume relations, etc.). Compared to echocardiography, real-time left ventricular (LV) pressure/volume analysis via catheterization is more precise and insightful in assessing LV function. Additionally, LV pressure/volume analysis provides the ability to instantaneously record changes during manipulations of contractility (e.g., &#946;-adrenergic stimulation) and pathological insults (e.g., ischemia/reperfusion injury). In addition to the maximum (+dP/dt) and minimum (-dP/dt) rate of pressure change in the LV, an accurate assessment of LV function via several load-independent indexes (e.g., end systolic pressure volume relationship and preload recruitable stroke work) can be attained. Heart rate has a significant effect on LV contractility such that an increase in the heart rate is the primary mechanism to increase cardiac output (i.e., Bowditch effect). Thus, when comparing hemodynamics between experimental groups, it is necessary to have similar heart rates. Furthermore, a hallmark of many cardiomyopathy models is a decrease in contractile reserve (i.e., decreased Bowditch effect). Consequently, vital information can be obtained by determining the effects of increasing heart rate on contractility. Our and others data has demonstrated that the neuronal nitric oxide synthase (NOS1) knockout mouse has decreased contractility. Here we describe the procedure of measuring LV pressure/volume with increasing heart rates using the NOS1 knockout mouse model.

This article describes the measurement of murine left ventricular function via pressure/volume analysis at different heart rates.

Bowditch Efekt incelenmesi: Basınç Hacim İlişkisi Tedbir Fare Kardiyak Kateterizasyon

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these ...

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 ...

Development of a Noninvasive, Laser-Assisted Experimental Model of Corneal Endothelial Cell Loss

Nd:YAG lasers have been used to perform noninvasive intraocular surgery, such as capsulotomy for several decades now. The incisive effect relies on the optical breakdown at the laser focus. Acoustic shock waves and cavitation bubbles are generated, causing tissue rupture. Bubble sizes and pressure amplitudes vary with pulse energy and position of the focal point. In this study, enucleated porcine eyes were positioned in front of a commercially available Nd:YAG laser. Variable pulse energies as well as different positions of the focal spots posterior to the cornea were tested. Resulting lesions were evaluated by two-photon microscopy and histology to determine the best parameters for an exclusive detachment of corneal endothelial cells (CEC) with minimum collateral damage. The advantages of this method are the precise ablation of CEC, reduced collateral damage, and above all, the non-contact treatment.

Here, we present a protocol to detach corneal endothelial cells (CEC) from Descemet&#8217;s membrane (DM) using a neodymium:YAG (Nd:YAG) laser as an ex vivo disease model for bullous keratopathy (BK).

Kornea Endotel Hücre Kaybının Noninvaziv, Lazer Destekli Deneysel Modelinin Geliştirilmesi

Nd:YAG lasers have been used to perform noninvasive intraocular surgery, such as capsulotomy for several decades now. The incisive effect relies on the ...

Reduced Complications after Arterial Reconnection in a Rat Model of Orthotopic Liver Transplantation

The rat orthotopic liver transplantation (OLT) model is a powerful tool to study acute and chronic rejection. However, it is not a complete representation of human liver transplantation due to the absence of arterial reconnection. Described here is a modified transplantation procedure that includes the incorporation of hepatic artery (HA) reconnection, leading to a marked improvement in transplant outcomes. With a mean anhepatic time of 12 min and 14 s, HA reconnection results in improved perfusion of the transplanted liver and an increase in long-term recipient survival from 37.5% to 88.2%. This protocol includes the use of 3D-printed cuffs and holders to connect the portal vein and infrahepatic inferior vena cava. It can be implemented for studying multiple aspects of liver transplantation, from immune response and infection to technical aspects of the procedure. By incorporating a simple and practical method for arterial reconnection using a microvascular technique, this modified rat OLT protocol closely mimics aspects of human liver transplantation and will serve as a valuable and clinically relevant research model.

The goal of this study is to modify the rat orthotopic liver transplant model to better represent human liver transplantation and improve recipient survival. The presented method reestablishes hepatic arterial inflow by connecting the donor liver&#39;s common hepatic artery to the recipient liver&#39;s proper hepatic artery.

Ortotopik Karaciğer Transplantasyonunun Sıçan Modelinde Arteriyel Reconnection Sonrası Azaltılmış Komplikasyonlar

The rat orthotopic liver transplantation (OLT) model is a powerful tool to study acute and chronic rejection. However, it is not a complete representation ...

Orthotopic Kidney Auto-Transplantation in a Porcine Model Using 24 Hours Organ Preservation And Continuous Telemetry

In the present era of organ transplantation with critical organ shortage, various strategies are employed to expand the pool of available allografts for kidney transplantation (KT). Even though, the use of allografts from extended criteria donors (ECD) could partially ease the shortage of organ donors, ECD organs carry a potentially higher risk for inferior outcomes and postoperative complications. Dynamic organ preservation techniques, modulation of ischemia-reperfusion and preservation injury, and allograft therapies are in the spotlight of scientific interest in an effort to improve allograft utilization and patient outcomes in KT.
Preclinical animal experiments are playing an essential role in translational research, especially in the medical device and drug development. The major advantage of the porcine orthotopic auto-transplantation model over ex vivo or small animal studies lies within the surgical-anatomical and physiological similarities to the clinical setting. This allows the investigation of new therapeutic methods and techniques and ensures a facilitated clinical translation of the findings. This protocol provides a comprehensive and problem-oriented description of the porcine orthotopic kidney auto-transplantation model, using a preservation time of 24 hours and telemetry monitoring. The combination of sophisticated surgical techniques with highly standardized and state-of-the-art methods of anesthesia, animal housing, perioperative follow up, and monitoring ensure the reproducibility and success of this model.

Large animal models play an essential role in preclinical transplantation research. Due to its similarities to the clinical setup, the porcine model of orthotopic kidney auto-transplantation described in this article provides an excellent in vivo setting for the testing of organ preservation techniques and therapeutic interventions.

24 Saat Organ Koruma Ve Sürekli Telemetri Kullanılarak Bir Domuz Modelinde Ortotopik Böbrek OtoTransplantasyonu

In the present era of organ transplantation with critical organ shortage, various strategies are employed to expand the pool of available allografts for ...

Intramyocardial Transplantation of MSC-Loading Injectable Hydrogels after Myocardial Infarction in a Murine Model

One of the major issues facing current cardiac stem cell therapies for preventing postinfarct heart failure is the low retention and survival rates of transplanted cells within the injured myocardium, limiting their therapeutic efficacy. Recently, the use of scaffolding biomaterials has gained attention for improving and maximizing stem cell therapy. The objective of this protocol is to introduce a simple and straightforward technique to transplant bone marrow-derived mesenchymal stem cells (MSCs) using injectable hydroxyphenyl propionic acid (GH) hydrogels; the hydrogels are favorable as a cell delivery platform for cardiac tissue engineering applications due to their ability to be cross-linked in situ and high biocompatibility. We present a simple method to fabricate MSC-loading GH hydrogels (MSC/hydrogels) and evaluate their survival and proliferation in three-dimensional (3D) in vitro culture. In addition, we demonstrate a technique for intramyocardial transplantation of MSC/hydrogels in mice, describing a surgical procedure to induce myocardial infarction (MI) via left anterior descending (LAD) coronary artery ligation and subsequent MSC/hydrogels transplantation.

Stem cell-based therapy has emerged as an efficient strategy to repair injured cardiac tissues after myocardial infarction. We provide an optimal in vivo application for stem cell transplantation using gelatin hydrogels that are able to be enzymatically cross-linked.

Murine Modelinde Miyokard İnfarktüsünden Sonra MSC Yüklemeli Enjekte Edilebilir Hidrojellerin intramiyokardiyal Transplantasyonu

One of the major issues facing current cardiac stem cell therapies for preventing postinfarct heart failure is the low retention and survival rates of ...