Establishment of an Ex Vivo Lung Perfusion Rat Model for Translational Insights in Lung Transplantation

Summary

Here, we describe the necessary steps for establishing a rat EVLP model and show the inflammatory profile associated with the perfused lungs. The aim is to propagate knowledge and experiences about the rat EVLP model, enabling the integral understanding of the biological responses associated with that revolutionary technique.

Abstract

Since the establishment of lung transplantation as a therapeutic strategy for advanced lung diseases, the scientific community is faced with the problem of a low number of lungs considered viable for the donation process. In recent decades, however, this scenario has been positively changed, given the development of ex vivo lung perfusion (EVLP) as a strategy for evaluating and reconditioning marginal lungs. The establishment of EVLP in large transplant centers has favored an increase in the number of lung transplants, both by increasing the diagnostic accuracy of lung function and by constituting an effective platform for the reconditioning of lung grafts. In this context, faced with ethical and logistical issues, as well as in the study of immunological factors associated with lung transplantation, the development of rodent EVLP models has become important, given their reliability, the possibility of genetic manipulation, and lower costs. This paper describes a protocol for establishing a rat EVLP model and shows the inflammatory profile associated with the perfused lungs. This will help propagate knowledge about the rat EVLP model, promoting our understanding of the biological responses associated with that revolutionary technique.

Introduction

Lung transplantation has been recognized as a therapeutic strategy for end-stage diseases with improvements in surgical methods and immunosuppression during the last decades. Despite the high demand, the number of acceptable deceased lung donors is lower than for other solid organs, converging to the lower number of lung transplants performed^1,2,3. To address the donor pool shortage, the medical community has expanded the criteria for lung donation, turning organs previously considered nonviable into potential organs for transplantation. However, the extended criteria require....

Protocol

Animal experiments were performed in compliance with the Animal Use Protocol approved by the Animal Care Committee at the University Health Network. Male Lewis rats (255-330 g) were given ad libitum access to food and water. Then, they were maintained in a controlled environment (18-22 °C) with a 12 h day-night cycle. See the Table of Materials for details related to all materials, solutions, and instruments used in this protocol.

1. Initialization of the  ex vivo lung perfusion system

1. Ensure that all transducers are connected to the isolated perfusion organ equi....

Results

All lungs with CIT ranging from 20 min to 18 h could be perfused for 4 h (Figure 2)¹³. Compliance was stable in most groups with the exception of 18 h CIT, which gradually decreased over the 4 h perfusion period (Figure 2A). Despite this, no significant differences in vascular resistance, lung graft oxygenation, and glucose levels were observed for the groups (Figure 2B-D). L.......

Discussion

This study described the necessary steps for establishing a rat EVLP protocol. Here, we show that donor lungs can be perfused for 4 h after cold static preservation of up to 18 h at 4 °C. This was demonstrated by assessing lung compliance, pulmonary vascular resistance, perfusate glucose/lactate concentration, and P/F ratio.

The rise of the EVLP platform as an important tool in the evaluation of lung function and for graft reconditioning has motivated several groups to develop a rat EVLP .......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
14 G intravenous catheter	BD	381167	Orotracheal intubation
16 G intravenous catheter	BD	381157	Catheter to flush the lung
2-0 Suture Sofsilk	Covidien	S-305	Tracheal tube fixation
3-0 Suture Sofsilk	Covidien	S303	Fixation of arterial and atrial cannulas
IPL-2 Core Isolated Perfused Lung System for Rat	Harvard Apparatus	734276	Ex Vivo Lung Perfusion (EVLP) system
Ketamine (Narketan)	Vetoquinol	440894	Sedation/anesthesia
Large Pulmonary Artery Cannula	Harvard Apparatus	73-0711
Left Atrial Cannula	Harvard Apparatus	73-0712
Low potassium dextran glucose solution (Perfadex)	XVIVO	19811	Preservation solution
Methylprednisolone sodium succinate (SOLU-MEDROL)	Pfizer	14705	Anti-inflammatory
Prostaglandin E1 (Prostin VR)	Pfizer	RX297945
Small Animal Ventilator (Model 683)	Harvard Apparatus	55-0000
Sodium heparin	Leo Pharma	453811
Steen Solution	XVIVO	19004	Buffered extracellular solution to perfuse lungs during EVLP
Sugita Aneur clip curv	Mizuho	07-940-86	Tracheal clamp
Tracheal Cannula	Harvard Apparatus	73-3384
Xylazine (Rompun)	Bayer Healthcare	2169592	Sedation/anesthesia