Introduction -We have previously demonstrated that cold ischemia is the limiting factor in expanding donor criteria in kidneys from the uncontrolled-DCD (uDCD). In the early era of clinical kidney transplantation, retrieval of kidneys from uDCD donors involved a prolonged period of warm ischemic exposure, with a corresponding minimal period ex vivo since organ preservation was in its infancy. Today we have the inverse situation with limited warm ischemia and prolonged hypothermic preservation times due to mandated organ sharing algorithms. In the present study we demonstrate that oxidative metabolism of sufficient magnitude to support reparative processes can be accomplished ex vivo in human kidney allografts following as much as 3 hours of postmortem ischemia. Method -Human kidneys (n=5) were recovered following cardiac arrest with consent of next-of-kin with a mean ischemic period of 2.5 +/-0.70 hours. The ischemically damaged kidneys were flushed of blood and placed on an acellular near-normothermic perfusion at 32°C for 24-hours. During the ex vivo perfusion parameters evaluating resuscitated oxidative metabolism, cytoskeletal damage and reparative processes involving new synthesis were monitored. Controls (n=5) consisted of the paired human kidneys with equivalent ischemic damage that were hypothermically perfused at 4°C for 24-hours . Two critical regenerative pathways involving new synthesis were evaluated: The junctional integrity protein, ZO-1 was used to assess cytoskeletal integrity. The up-regulation of proliferating cell nuclear antigen (PCNA) was used to assess recovery of synthetic functions. Results -Test kidneys that were perfused at near-normothermia (32°C) demonstrated recovery in terms of restored oxidative metabolism (oxygen consumption (>0.14cc/min/g)), time-dependant normalization of cytoskeletal integrity and up-regulation of DNA synthesis. In contrast, the control kidneys that were hypothermically perfused (4°C) did not demonstrate recovery during the 24 hours of perfusion. Conclusions -The lack of recovery in the control kidneys is not surprising given that hypothermia inhibits oxidative metabolism by more than 96%, thereby preventing activation of cellular reparative processes. The ability to resuscitate oxidative metabolism leading to up-regulation of DNA synthesis observed in the warm perfused kidneys allows for regeneration of cytoskeletal integrity. The ability to "repair" ischemically damaged kidneys ex vivo may present opportunities for expanded donor criteria using uDCD.
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