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

This study established a haproducible rat EVOP model to investigate ischemia hyperfusion injury, inflammatory signally, and donor graft function, aiming to evaluate, preserve and repair lungs ex vivo for improved transplantation outcomes. The most recent developments include using EVOP to access, recondition, and even treat donor lungs before transplantation. Our protocol addresses the need for cost-effective reproducible models for translational lung transplant research.

Our findings provide a reliable small MOEVOP model to test new therapies and improve donor lung evaluation and preservation. To begin, check that all transducers are connected to the isolated perfusion organ equipment and the data acquisition system, then launch the data acquisition software. Prior to perfusion, fill the ex vivo lung perfusion or EVLP circuit with 150 milliliters of supplemented steam solution.

Set the warm water bath to 20 degrees Celsius and circulate the Perfusate solution to warm up the EVLP system. Orotracheal intubate an anesthetized rat. Next, connect the tracheal tube to the small animal ventilation system to ventilate the rat.

Place the rat in a supine position. With a pair of scissors and forceps. Enter the peritoneal cavity and carry the incision cranially to perform a median laparotomy.

Now inject sodium heparin into the inferior vena cava. Enter the thoracic cavity through the xiphoid process and cautiously open the diaphragm radially without damaging the lung. Then cranially resect the sternum.

To retrieve the heart and lung block, make an incision on the inferior vena cava along the apex of the left heart. With a pair of micro scissors, make an anterior incision in the right ventricular outflow tract. Then insert an 18 gauge intravenous catheter into the pulmonary trunk and flush the lungs with 20 milliliters of low potassium dextran or LPD solution.

Immediately after flushing, clamp the lower third of the trachea at the end of inspiration to preserve the lungs in an inflated state. Then harvest the lung and place it in LPD solution for storage. To begin lung perfusion, first, secure a pre-tied zero silk ligature underneath the main pulmonary artery and around the ventricles.

Connect the pulmonary artery cannula to the inflow line of the EVLP system. Start the peristaltic pump at 10%of maintenance flow, ensuring the removal of any air in the cannula. Secure the inflow cannula into the main pulmonary artery.

Then insert the drainage cannula from the apex of the heart into the left atrium. Connect the left atrial cannula to the outflow line of the system. Now make a small hole in the trachea and insert the tracheal cannula.

Connect the tracheal cannula to the ventilation line of the system. Gradually increase the perfusion flow rate to attain 20%of the cardiac output. Release the tracheal clamp 20 minutes after perfusion initiation, then start lung ventilation followed by EVLP gas flow.

Record the dynamic lung compliance and pulmonary vascular resistance every hour during EVLP. Take Perfusate samples from the sample port every hour. Flash, freeze the samples in liquid nitrogen for further analysis.

After ceasing perfusion, clamp the trachea to maintain the lungs in an inflated state. Then isolate lung samples and tubes, flash freeze the samples in liquid nitrogen. All lungs with CIT ranging from 20 minutes to 18 hours could be perfused for four hours.

Compliance gradually decreased in the 18 hour CIT group over the perfusion period. No significant differences in vascular resistance lung graft oxygenation, and glucose levels were observed for the groups. Lactate values tended to increase over time with higher values observed at longer CIT.

All groups showed similar values of perfusion electrolytes, and level of edema formation with the 24 hour group showing severe edema.