The effect of preservation temperature and period on resuscitation of the ischemically damaged canine pancreas during preservation by the two-layer (University of Wisconsin solution/perfluorochemical) method.

We have shown that 24 to 48 hour-preservation by the two-layer (University of Wisconsin solution (UW)/perfluorochemical (PFC)) method at 4 degrees C resuscitates a canine pancreas subjected to 90 min of warm ischemia. However, it is necessary to shorten preservation period for resuscitation of the ischemically damaged pancreas in a clinical simultaneous pancreas-kidney transplantation. The purpose of this study was to clarify the effect of preservation temperature and period on the resuscitation of the ischemically damaged pancreas during preservation by the two-layer method. First of all, we examined the possibility of resuscitation of the ischemically damaged pancreas during short-term preservation by the two-layer method. After 90 minutes of warm ischemia, canine pancreases were preserved by the two-layer method at 4, 20, or 37 degrees C. In control group, the pancreas graft was autotransplanted without preservation. Graft viability was judged by graft survival after autotransplantation. Pancreas grafts subjected to 90 min of warm ischemia were not viable (0/5). At 4 degrees C, 5 to 12 hr-preservation did not resuscitate the grafts (0/3 and 0/3 respectively). At 20 degrees C, 3 and 5 hr-preservation resuscitated the grafts (3/5 and 5/5 respectively), although 1 and 8 hr-preservation were not successful (0/3 and 0/3 respectively). At 37 degrees C, all the grafts were not resuscitated irrespective of preservation period. It was clear that the ischemically damaged pancreas was resuscitated during shot-term preservation by the two-layer method only at 20 degrees C. Secondly, we measured tissue adenine nucleotide levels by high performance liquid chromatography and pancreatic tissue perfusions using H2 clearance technique on reperfusion and examined the viability of vascular endothelium by nuclear trypan blue staining to make clear the necessary conditions for resuscitation of the ischemically damaged pancreas at 20 degrees C. ATP tissue levels in one hr-preserved grafts were 2.55 +/- 0.38 mumol/g dry weight and were significantly lower compared with the levels in 5 and 8 hr-preserved grafts, 9.40 +/- 2.09 (P < 0.01) and 7.37 +/- 1.06 mumol/g dry weight (P < 0.01) respectively. On the other hand, nuclear trypan blue uptakes of endothelial cells in 8 hr-preserved grafts were 37.6 +/- 11.6% and were significantly higher than 1 hr- and 5 hr-preserved grafts 5.6 +/- 4.5 (P < 0.01) and 5.0 +/- 3.0% (P < 0.01) respectively. As a consequence, pancreatic tissue perfusions in 8 hr-preserved grafts, 31.0 +/- 3.5 ml/min/g, were significantly lower than 1 hr- and 5 hr-preserved graft 72.0 +/- 11.6 (P < 0.01), 63.9 +/- 13.3 ml/min/g (P < 0.01) respectively. However, thromboxane A2 synthesis inhibitor (OKY046 0.1 mM/L) decreased the percentage of trypan blue uptake (8.2 +/- 3.6%) without interfering ATP synthesis (8.44 +/- 0.92 mumol/g dry weight) in 8 hr-preserved pancreas and tissue perfusions after reperfusion were dramatically improved (99.6 +/- 11.8 ml/min/g). As a result the ischemically damaged pancreas was resuscitated (4/5, 80%). It was suggested that 1 hr-preservation was not enough to synthesize ATP, which was essential to repair damaged cells, although vascular endothelial cells were maintained. Eight hr-preservation incurs endothelial cell damage although ATP tissue levels were maintained and consequently microcirculation was disturbed at reperfusion but OKY046 protects endothelial cells against preservation/reperfusion injury. As a consequence, the ischemically damaged pancreas was resuscitated. We conclude that short-term (3 to 8 hr) preservation at 20 degrees C by the two-layer method with OKY046 accelerates ATP synthesis, which is essential for repairing damaged cells and protects microvascular endothelial cells. This makes it possible to resuscitate the canine pancreas graft subjected to 90 min warm ischemia. This method holds promise for pancreas-kidney transplantation from cardiac arrest donors.