Sign In

A subscription to JoVE is required to view this content. Sign in or start your free trial.

In This Article

  • Summary
  • Abstract
  • Introduction
  • Protocol
  • Representative Results
  • Discussion
  • Acknowledgements
  • Materials
  • References
  • Reprints and Permissions

Summary

The purpose of this manuscript and protocol is to explain and demonstrate in detail the surgical procedure of orthotopic kidney transplantation in rats. This method is simplified to achieve the correct perfusion of the donor kidney and shorten the reperfusion time by using the venous and ureteral cuff anastomosis technique.

Abstract

Kidney transplantation offers increased survival rates and improved quality of life for patients with end-stage renal disease, as compared to any type of renal replacement therapy. Over the past few decades, the rat kidney transplantation model has been used to study the immunological phenomena of rejection and tolerance. This model has become an indispensable tool to test new immunomodulatory pharmaceuticals and regimens prior to proceeding with expensive preclinical large animal studies.

This protocol provides a detailed overview of how to reliably perform orthotopic kidney transplantation in rats. This protocol includes three distinctive steps that increase the probability of success: perfusion of the donor kidney by flushing through the portal vein and the use of a cuff system to anastomose the renal veins and ureters, thereby decreasing cold and warm ischemia times. Using this technique, we have achieved survival rates beyond 6 months with normal serum creatinine in animals with syngeneic or tolerant kidney transplants. Depending on the aim of the study, this model can be modified by pre- or posttransplant treatments to study the acute, chronic, cellular, or antibody-mediated rejection. It is a reproducible, reliable, and cost-effective animal model to study different aspects of kidney transplantation.

Introduction

Historically, the first transplant rejection studies were performed by Brent and Medawar using skin transplants in rodents1. It soon became clear that skin has distinct immunological features, making it a highly immunogenic organ that is different in rejection from other vascularized solid organs2. Rat studies of solid organ transplant rejection are habitually limited to heart, liver, and kidney transplants. Although each of these organs is suitable to study rejection, there are advantages and disadvantages to each of them. Heart transplants are often transplanted into the abdomen and anastomosed to the aorta and vena ca....

Protocol

Lewis (RT11) and Dark Agouti (DA) (RT1Aa) rats were purchased from commercial vendors (see the Table of Materials). These fully MHC-mismatched strains are often used to study acute renal allograft rejection. All animals were housed and maintained according to the National Institutes of Health’s (NIH) guidelines in a specific pathogen-free facility at the Johns Hopkins University. All procedures were approved by the institutional animal care and use committee.

Representative Results

We performed syngeneic (N = 5) and allogeneic kidney transplants (N = 5). Animals with a syngeneic transplant achieved long-term survival without any immunosuppressive treatment. Animals that received an allogeneic transplant without immunosuppression rejected their graft and succumbed to renal failure with a median survival of 8 days (Figure 4A). Mean serum creatinine increased modestly in the syngeneic group while it increased by 14-fold in the allogeneic.......

Discussion

In this manuscript, we describe the surgical method for orthotopic KT in rats in detail, including all the necessary equipment needed to perform this procedure (Figure 5). In 1965, Fisher and Lee published the first report on KT in rats, which became the start of an exciting investigative field18. Since then, many modifications have been introduced to improve the reproducibility of this model. It has served as an effective animal model for studying ischemia-reperfusio.......

Acknowledgements

This work was funded by a generous gift from the Bombeck Family Estate.

....

Materials

NameCompanyCatalog NumberComments
Buprenorphine HCLReckitt Benckiser Healthcare UKNDC12496-0757-5
Dissecting forceps, curvedZhenbang, China11cm Flat handle 
Heparin sodium injection USPSagent Pharmaceuticals NDC25021-400-10
Micro-forceps, straight, smoothJingzhong, ChinaWA3010
Micro needle holderJingzhong, ChinaWA2010
Micro vessel clampsJingzhong, ChinaWA40120
Micro spring sciccor 1ROBOZRS-5620
Micro spring sciccor 2F.S.T.91501-09
Micro spring sciccor 3Zhenbang, China8.5cm Vannas,curved
Prograf (Tacrolimus/FK506)Astellas
RatsCharles River & Taconic Biosciences LEW/Crl & DA-M 
ShaverWahl79600-2101
Suture 4-0EthiconJ304H
Suture, 4-0 Ethicon683G
Suture, 10-0 Ethicon2820G
Syringes & NeedlesBD
Thread, 8-0Ashaway75290
Ureteral cuffMicrolumen160-1Polymide Tubing, Diameter 0.41 mm 
Venous cuffIntramedic BD7441PE-200 Non-radiopaque polyethylene tubing ID: 1.4 mm, OD: 1.9 mm

References

  1. Billingham, R. E., Brent, L., Medawar, P. B. Actively acquired tolerance of foreign cells. Nature. 172 (4379), 603-606 (1953).
  2. Murray, J. E. Organ transplantation (skin, kidney, heart) and the plastic surgeon.

Explore More Articles

Orthotopic Kidney TransplantationRatSurgical ProcedurePortal Vein PerfusionCuff SystemRenal Vein AnastomosisUreter AnastomosisMicrosurgical Technique

This article has been published

Video Coming Soon

JoVE Logo

Privacy

Terms of Use

Policies

Research

Education

ABOUT JoVE

Copyright © 2024 MyJoVE Corporation. All rights reserved