title: The mechanism of action of the two-layer (Euro-Collins' solution/perfluorochemical) cold-storage method in canine pancreas preservation--the effect of 2,4 dinitrophenol on graft viability and adenosine triphosphate tissue concentration.
abstract: Tissue concentrations of adenosine triphosphate have been previously associated with successful pancreas preservation using the two-layer cold storage method in a canine autotransplantation model. To clarify the role of ATP vs. oxygenation per se, we used 2,4 dinitrophenol, an uncoupler of mitochondrial oxidative phosphorylation. DNP caused no toxicity in pancreas grafts preserved for 24 hr in Euro-Collins' solution or 48 hr in University of Wisconsin solution. Tissue concentration of ATP and viability of pancreas grafts, defined as maintenance of normoglycemia for 5 days following transplantation, were compared among six groups after a preservation interval of 24 or 48 hr. After 24 hr all grafts were viable, whether preserved using simple cold storage in EC (group 1a), two-layer (EC/perfluorochemical [PFC]) method (group 2a), or two-layer (EC+DNP/PFC) method (group 3a); respective graft survival was 4/4 (100%), 5/5 (100%), and 4/5 (80%); one of five dogs in group 3a died of a cause unrelated to the pancreas. ATP levels were higher in group 2a compared with group 1a (7.47 +/- 0.47 vs. 1.41 +/- 0.53 mumol/g dry weight, P less than 0.01) and lower in group 3a compared with group 2a (1.25 +/- 0.37 vs. 7.47 +/- 0.47, P less than 0.01). After 24 hr, we observed no difference in viability despite ATP concentration differences. However after 48 hr preservation, graft viability varied among the groups: 0/4 (0%), 4/4 (100%), and 0/3 (0%) in groups 1b, 2b, and 3b, respectively. ATP tissue concentration was again higher in group 2b after two-layer (EC/PFC) method preservation (7.91 +/- 1.21 vs. 1.21 +/- 0.31 mumol/g dry weight, P less than 0.01) compared with EC preservation (group 1b). DNP again caused a significant decrease in tissue ATP in group 3b (0.61 +/- 0.07 vs. 7.91 +/- 1.21, P less than 0.01). The two-layer (EC/PFC) method clearly protected pancreas viability, and inhibition of ATP production using DNP caused loss of viability in this model. We conclude that oxygenation of the pancreas during preservation by the two-layer method allows continued ATP production within the graft. Metabolic processes vital to cellular integrity can be maintained, which produces an extended period of preserved pancreatic viability.
authors.affiliation: First Department of Surgery, Kobe University School of Medicine, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Kuroda
authors.forename: Y
authors.initials: Y
authors.lastname: Fujino
authors.forename: A
authors.initials: A
authors.lastname: Morita
authors.forename: Y
authors.initials: Y
authors.lastname: Tanioka
authors.forename: Y
authors.initials: Y
authors.lastname: Ku
authors.forename: Y
authors.initials: Y
authors.lastname: Saitoh
mesh_terms: D019297:2,4-Dinitrophenol; D000255:Adenosine Triphosphate / Q000032:analysis; D000818:Animals; D015925:Cryopreservation; D004140:Dinitrophenols / Q000494:pharmacology; D004285:Dogs; D005260:Female; D006085:Graft Survival / Q000187:drug effects; D006982:Hypertonic Solutions; D008297:Male; D009926:Organ Preservation / Q000379:methods*; D010100:Oxygen / Q000502:physiology; D010179:Pancreas / Q000737:chemistry; D016035:Pancreas Transplantation
chemical_list: D004140:Dinitrophenols; C037096:Euro-Collins' solution; D006982:Hypertonic Solutions; D000255:Adenosine Triphosphate; D019297:2,4-Dinitrophenol; D010100:Oxygen