Short-Term Storage of the Ischemically Damaged Human Pancreas by the Two-Layer Method Prior to Islet Isolation

Abstract

A two-layer cold storage method (TLM) allows sufficient oxygen delivery to pancreata during preservation and resuscitates the viability of ischemically damaged pancreata in the canine pancreas transplant model. In this study, we applied a short-term preservation of the TLM to human pancreata after prolonged cold ischemia prior to islet isolation, and investigated the mechanisms of resuscitation of the ischemically damaged human pancreas by the TLM. Human pancreata were procured from cadaveric donors and preserved by the TLM for 3.2 ± 0.5 h after 11.1 ± 0.9 h of cold storage in UW (TLM group), or by cold UW alone for 11.0 ± 0.3 h (UW group). Islet isolations of all pancreata were performed using the Edmonton protocol. Islet recovery and in vitro functional viability of isolated islets were significantly increased in the TLM group compared with the UW group. According to the criteria of the Edmonton protocol, 10/14 cases (71%) in the TLM group were transplanted to patients with type I diabetes mellitus compared with only 5/21 cases (24%) in the UW group. In the metabolic assessment of human pancreata, levels of energetic parameters (ATP, total adenylates, and energy charge) were significantly increased, and malondialdehyde (MDA) levels were significantly decreased after the TLM preservation. There was no observable change in the incidence or degree of mitochondrial injury after the TLM preservation. Additional short-term storage by the TLM resuscitates the ischemically damaged human pancreas by regenerating the energetic status and prevents further damage by oxidative stress, ultimately leading to improvements of islet recovery and in vitro function. Use of the TLM following prolonged storage in UW provides an excellent adjunctive protocol for treating human pancreata for the rigors of the islet isolation process.

Improved Outcomes in Islet Isolation and Transplantation by the Use of a Novel Hemoglobin-based O2 Carrier

“…Maintaining an appropriate O 2 level would seem important to prevent ischemic damage and reperfusion injury during organ preservation and pancreatic islet isolation. Indeed, artificial O 2 carriers, such as perfluorocarbons (PFC), have a beneficial effect on islet isolation and transplantation outcomes when used during pancreas preservation with UW solution in the two layer method (TLM) (24)(25)(26)(27).…”

Chemical Oxygenation of Pancreatic Tissue Prior to Islet Isolation and Transplantation

“…As reported, this was the case for the studies from the Edmonton group (Tsujimura et al, 2002a, Tsujimura et al, 2004a, Salehi et al, 2006, Kin et al, 2006a but not for the study from the Nordic Network (Caballero-Corbalan et al, 2007). This is of particular relevance when pancreases from obese donors are recovered.…”

“…One explanation for this phenomenon is that TLM re-initiates production of ATP in mitochondrial pathways by providing sufficient oxygen levels. One can speculate that maintaining the metabolic activity of mitochondria within a certain range seems to be efficient to prevent excessive generation of ROS in mitochondria as main subcellular producers of ROS (Li et al, 2008, Tsujimura et al, 2004a and to inhibit mitochondrial pathways of apoptotic cell death (Matsuda et al, 2003).…”

“…One important difference that exists between the larger trials and previously published smaller studies concerns the method of oxygenation used. While in the smaller trials pancreases had been subjected to continuous oxygenation for a short period of approximately 3 hours subsequent to cold storage (Tsujimura et al, 2002a, Tsujimura et al, 2004a, Salehi et al, 2006, the large scale trials utilized oxygen-precharged PFD for the entire period of cold ischemia. However, experiments in rats clearly suggested that oxygen-precharged PFD is equivalent to continuously oxygenated PFD with regard to pancreas ATP concentration and isolated islet yield after 24 hours of cold storage (Hiraoka et al, 2001).…”

Resolution of Severe Atopic Dermatitis after Tacrolimus Withdrawal

“…The most remarkhas led to increased success both in islet cell processing and clinical outcome in recent years (37,45). Very promable benefits of islet transplantation include improved glycemic control (with normalization of glycated hemoising approaches are under development aiming at improving the efficiency of organ recovery and islet isolation globin A1c and reduction of glycemic excursion throughout the day) (20) and prevention of severe hypoglycemic techniques (6,15,19,50,54,59,63,67), enhancing engraftment through cytoprotection of islet grafts (27,28,35, episodes (1,40,51,53), even when exogenous insulin is required (i.e., marginal islet graft function) (8, 24,34,40,38,68), islet implantation into alternative sites (7,23,39), implementation of immunomodulatory (38,43) and im-45,47,48,52,56,57). Improved health-related (5) and dia-munoisolation (30,31) protocols to reduce the need for mated digestion method followed by density gradient purification was used (15,26,46).…”