Ischemia/Reperfusion Injury

Renal transplantation model in the pig: the protective role of biliverdin (BV) and carbon monoxide (CO) in ischemia/reperfusion injury.

Transplantation and marginal donors

The limited availability of organs still influences the chance of transplantation for all the patients that are in waiting list. Developing innovative strategies and technologies that could permit the salvage of organs that are considered “marginal” for transplantation, is surely an interesting perspective but it has to be evaluated carefully. Even though the idea of “marginality” is still in development, the organs taken from non heart-beating donor and organs exposed to ischemia injuries are definitely marginal. With the purpose of salvaging marginal organs for clinical use, the preconditioning with some substances seems effective for the decrease of ischemia/reperfusion injury.

Heme Oxygenase 1 (HO-1)

Heme Oxygenase (HO-1), an inducible enzyme, expressed everywhere, plays a key role of protection, and it catalyses the oxidation of heme into biliverdin (BV), with release of iron (II) and carbon monoxide (CO), produced with antioxidant and anti-inflammatory properties. The gene expression of HO-1 is produced by stimulus connected to oxidative stress and inflammation, and the effects of heme’s catabolism are actually the real actors of cellular protection’s mechanism.

Pre-clinic models of ischemia/reperfusion injury

Protective effects of heme’s degradation products have been demonstrated for the first time at the Centre of Biotechnologies on big size animals, realizing two pre-clinic models of ischemia/reperfusion injury.

Cardiac model   

In a model of cardiopulmonary bypass the animals were preconditionedwith CO through inhalation. The hearts were covered and arrested with a Celsior cardioplegia solution saturated with CO, in deep hypothermia. The heart rate was reactivated through electrical defibrillation and the pigs were observed for the following 12 hours. CO’s treatment improved the clinical performance and the energetic condition of the organ, and decreasing the edema and the apoptosis, promoting organ’s recovery.

Hepatic model

These results were confirmed in a model of pig liver conserved in Celsior at 4° C for 18 hours (cold ischemia).After the liver was reperfused through an extracorporeal reperfusion circuit connected to a “reservoir” pig. Results gained with this second experimental model show that the preconditioning with CO and BV permits an improvement of hepatic functionality, and decrease of hepatic injury, apoptosis, intercellular acidosis and release of pro-inflammatory cytokine.

Pig’s kindney transplantation model

With the intent of confirming CO and BV’s protection role that was highlighted in the past studies, we developed, in collaboration with Centro Nazionale Trapianti (Transplantation National Centre) and Harvard Medical School – Beth Israel Deaconess Medical Center of Boston, a pig model of kidney transplantation. In this model we performed the transplantation of both kidneys after an extended ischemia and their following preservation in Celsior at 4° C for 24 hours. Then the kidneys were transplanted in allogenic pigs that were previously surgically prepared viaa bilateral nephrectomy. The animals that had the transplantation were observed for 7 days in which the renal function was analyzed (creatinine, BUN, sodium, potassium and total proteins). There was a daily evaluation of the animal’s health condition and were imparted to the pig drugs with  analgesic, immunosuppressive and antibiotic properties. A first phase of the project scheduled the standardizations of surgical, clinical and pharmacological procedures in order to develop the model and determine the time of ischemia. We are currently evaluating the protective effect of treatment, both donor and receiver, with carbon monoxide and/or biliverdin on ischemia/reperfusion injury in terms of renal functionality.

Future perspectives

Future perspectives are:

• Research on molecular and biochemical mechanism as reasons behind ischemia/reperfusion injury and on the effects of preconditioning factors on expressions’ level of genes involved  in this process.
• Evaluation about the possibility to realize a clinical trial, with a feasibility study, through the use of the substances tested on the cadaveric donor.