The increasing demand for transplantation has created a need for additional organ sources, and acceptance criteria have been widened, not only for kidney retrieval but increasingly for other organs with a lower tolerance of warm ischemia, such as the liver, pancreas, and lungs[18, 19]. This expansion of donor criteria to include the use of grafts from elderly donors, DCD, or organs from donors with other medical conditions has underlined the need for a technique that prevents any further damage during the preservation period. Previous reports strongly suggest that DCD donors are a valuable source of organs[3, 14, 20, 21]. Reconditioning using ANOR appears to give better results than in-situ cooling preservation, with a reduction of primary nonfunction and delayed graft function. In France, the national regulatory authority (Agence de la Biomédecine) has defined the conditions for organ removal, specifying the tolerated durations of warm and cold ischemia, for uncontrolled donors (type II of the Maastricht classification). However, no preliminary study has determined the optimal conditions for ANOR. Consequently, there is room for experimental studies to decipher the different steps of the protocol.
The development of a porcine preclinical experimental model of DCD donors allows for a study of the ANOR impact on organs without the limitations of a clinical study. Several models of DCD donors have previously been published in the literature to study the conditioning of kidney transplants by normothermic perfusion. Recently, using rats of 300 g and a warm ischemia period corresponding to clamping of the renal artery for 15 or 30 minutes, Moers et al. studied reperfusion for 1 to 2 hours. However, the restored pulsatile flow was far from comparable to a continuous flow of ANOR. In 2008, Hosgood et al. used a Large White pig model weighing between 40 and 50 kg. In this model, after 25 minutes of warm ischemia, kidneys were reperfused normothermically for 3 hours in an isolated organ preservation system using autologous blood. However, this experimental condition remains far from a clinical situation. In 2010, Rojas-Pena et al. used an extracorporeal membrane oxygenation circuit with pigs from 25 to 30 kg. Both external jugular veins were cannulated in place of the inferior vena cava. In addition, the absence of clamping of the descending thoracic aorta mimicked a systemic circulation model, which was not exactly the ANOR condition. The durations of warm ischemia were 10 and 30 minutes, respectively, and 90 and 100 minutes for reperfusion.
To our knowledge, the uncontrolled DCD model presented herein is the closest to the conditions encountered in clinical practice. Indeed, we use a large animal, the pig, well accepted as a preclinical model with a size, anatomy, and cardiovascular physiology very similar to those of human beings. Our model also used truly regional circulation between the inferior vena cava and the abdominal aorta, which was limited to the subdiaphragmatic region by clamping of the descending thoracic aorta. Moreover, our model is highly reproducible. In all cases, hemodynamic and respiratory objectives were achieved with very similar conditions from one animal to another. After 4 hours of ANOR, there was no significant difference compared with T0 for pH and plasma sodium levels. Lactose dehydrogenase levels were increased, a sign of tissue damage that was probably due to the stresses of surgery (incisions, muscular attrition, and so on); the lack of change between 1 and 4 hours of ANOR highlight the fact that this marker is not adapted for discriminant diagnosis determination. We observed an expected hemodilution, mainly due to the ANOR priming (about 700 to 800 cm3). However, hemoglobin levels remained correct, at 9.3 ± 0.6 g/dl after 4 hours of ANOR. Interestingly, in our hands, renal function likewise remained stable during ANOR and reached a steady state during the procedure.
However, the injection of heparin before cardiac arrest is a critical limit in this model. We aim to correct this easily in future protocols. Note that in previously described models, injection of heparin was likewise carried out before cardiac arrest.
It would also have been interesting to monitor urine production during ANOR. However, bladder catheterization of Large White male pigs is not possible, owing to a tortuous urethra and we consequently chose not to approach the bladder surgically, thereby avoiding the opening of the peritoneum and fluid, particularly lymph fluid, accumulation. However, we noted that at the end of ANOR, urine (about 500 cm3) was collected in the bladder. To better monitor kidney function, the implantation of a urine catheter during ANOR is envisioned for future experiments.