Summary

Murin hornhindetransplantation: en model til at studere den mest almindelige form for organtransplantation

Published: November 17, 2014
doi:

Summary

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.

Abstract

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.

Introduction

Hornhindetransplantation er en af ​​de mest vellykkede og almindelige typer af transplantation udføres i mennesker. Hvorfor denne operation udføres, er et resultat af skade, infektionssygdomme 1, eller andre former for ikke-infektiøs corneasygdom 2. Tal fra Eye Bank Association of America viser, at over 46.000 blev udført i 2011 (se hjemmeside på: restoresight.org/eye_banks/eye_banks.html). En indikation af dens succes er at et år fejlrater for allogene hornhinde podninger spænder fra 10 til 15%, og efter 5 år succesen er over 70% 3-8. Så mange undersøgelser har vist, er succes hornhinde allotransplantater direkte relateret til det faktum, at øjet er et immunologisk privilegeret sted. Faktorer, der er ansvarlige for hornhinder status som en immun privilegium websted omfatter manglen på både blod- og lymfekar i hornhinden, en relativ mangel af antigen-præsenterende celler, faktorer produceret af hornhinden at suppress immuneffektorceller funtions 9-15, lav ekspression af MHC antigener 16, og ekspressionen af FasL 17-20.

Men på trods af disse faktorer, som disponerer disse podninger for succes, må de gennemgå afvisning 3-7. Derfor forstå de mekanismer, der medierer denne afvisning samt teste de forskellige behandlingsformer til at forhindre afstødning er af afgørende betydning. Med henblik herpå vi beskriver her en murin model for hornhinde-transplantation, der har været i brug i over 20 år til at undersøge corneal transplantation i et kontrolleret eksperimentelle miljø. Da transplantationscentre reaktioner involverer mange forskellige faktorer, der arbejder i koncert, der vil endelige afgøre, om det transplanterede væv svigter eller lykkes, er det ikke muligt at forstå betydningen af disse faktorer i en in vitro model. Derfor er undersøgelser med anvendelse af levende dyr for at afgøre, hvilke faktorer er vigtige for enten succes eller failure af transplanteret væv.

Mens andre dyrearter er blevet anvendt til at studere hornhindetransplantation den murine model har flere fordele i forhold til anvendelse af andre arter. Den første er, at der findes mange stammer af mus, der udtrykker bestemte transgener eller er blevet gen-målrettede at mangle ekspression af specifikke immunologiske faktorer, hvis funktion i transplantation bedre kan undersøges. Derudover er der mange reagenser (både rekombinante faktorer og antistoffer, der neutraliserer faktorer), der er specifikke for mus, og som ikke findes for mange andre arter af dyr. På grund af eksistensen af disse faktorer, er denne model blevet brugt i udstrakt grad at identificere relevante faktorer involveret i akutte hornhinde allogen svar 15, 17,18,20 -29. Desuden er mange af de faktorer, der er involveret i hornhinde transplantation også kendt for at være funktionelle i transplantation af andre væv.

Protocol

BEMÆRK: Alle dyr, der anvendes i denne procedure behandles i overensstemmelse med Foreningen for Forskning i Vision og Oftalmologi redegørelse for brugen af ​​dyr i Ophthalmic og Vision Research samt de retningslinjer, der er fastsat af dyret tilsynsudvalg ved Saint Louis University. BEMÆRK: Alle kirurgiske instrumenter og opløsninger steriliseres forud for operation for at begrænse mikrobiel infektion i øjet. Det skal bemærkes, at mens dyrene gøre oplever nogen smerte fra denne procedure, vi ikke bes…

Representative Results

Den murine model af hornhindetransplantation har været anvendt i over 20 år med held karakterisere mekanismer både hornhinde allograftafstødning 19-23 og hornhinden allograft accept 13, 15,16,18, 24-27. Denne model blev brugt til at fastslå betydningen af FasL ekspression i hornhinde allograft accept, i, at dyr, der mangler FasL ikke var i stand til at acceptere hornhinde allografter 15. Det er også blevet anvendt til at påvise, at vaskulær endotel vækstfaktor receptor 1 morphol…

Discussion

Den murine model af hornhinde transplantation beskrevet her giver investigator for at studere menneskets hornhinde allograftafstødning i en model, der er prædiktive for, hvilke faktorer der bedst forbundet med både afvisning 15,17,18,20, 26-30 og accept 21-25 af hornhinde allografter. I modsætning til den menneskelige hornhinde transplantation, hvor patienterne får enten topisk eller systemisk steroidbehandling til enten at behandle eller forhindre afstødning 31, er denne model anv…

Disclosures

The authors have nothing to disclose.

Acknowledgements

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.

Materials

Name of Material/ Equipment Company Catalog Number Comments/Description
Zeiss Surgical Microscope Zeiss Rebuilt
1 mL Syringe BD 305122
3 mL Syringe BD 309657
10 mL Syringe BD 309602
Vannus Scissors Stortz E-3387
11-0 Sutures Alcon 717939M
Trephine 2.0mm Katena K 2-7520
Trephine 1.5 mm Katena K 2-7510
Tricaine Hydrochloride 0.5% Alcon NDC 0065-0741-12
Healon Abbott Healon OVD
Forceps FST 11251-20
7-0 Sutures Alcon 8065
2.5% Phenylephrine HCl Alcon NDC 61314-342-02
1% Tropicamide Bausch & Lomb NDC-24208-585-59
Hamilton Syringe Hamilton 7654-01
33 gauge needle Hamilton 90033
Cell Strainer (100 μm nylon) BD Falcon 352360
Hemocytometer Cardinal Health B3175
Trypan Blue Sigma T8154

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Cite This Article
Yin, X., Tajfirouz, D. A., Stuart, P. M. Murine Corneal Transplantation: A Model to Study the Most Common Form of Solid Organ Transplantation. J. Vis. Exp. (93), e51830, doi:10.3791/51830 (2014).

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