accumulation of leukocytes, 14,17,20 are the primary causes of Liver sinusoidal endothelial cell impairment and/or storage-related graft failure. However, in previous investigamicrocirculatory disturbances are thought to induce tions, the microcirculatory disturbances have not been tested storage-related graft failure; however, the respective in an adequate experimental setting of cold ischemia-reperfuroles of changes induced by extended cold preservation sion with a clear separation of the effects of transplantation and transplantation, as well as their interactions, are from those of extended cold preservation on the liver microcirstill unknown. To this end, the alterations of the liver culation.
microcirculation and of hepatocyte and sinusoidal endo-
The present study was designed to evaluate the effects of thelial cell functions induced by extended cold preservacold preservation-reperfusion injury on rat liver microcirculation and/or transplantation were assessed using an isotion, particularly on the various vascular and extravascular lated perfused rat liver model combined to an orthotopic spaces of the sinusoidal architecture. To this end, the alterrat liver transplantation model. Liver microcirculation ations induced by extended cold preservation and/or tranremained minimally altered following extended cold plantation were investigated on livers using an isolated perischemia alone, despite a marked deterioration of sinufused rat liver model combined to an orthotopic rat liver soidal endothelial cell function, while liver microcirculatransplantation model. Liver microcirculation was detertion alterations were mainly characterized by areas of mined using the multiple-indicator dilution curve technique, no-reflow following transplantation alone. It was only the Goresky technique. 21 In addition, taurocholate (TC) and when both procedures were associated that hepatocyte antipyrine (AT) elimination was used to determine hepatofunction became markedly compromised, without furcyte function, while hyaluronic acid (HA) elimination was ther deterioration in liver microcirculation. It is conused to establish sinusoidal endothelial cells function. The cluded that extended cold preservation and transplantaunderstanding of the respective alterations induced by extion as such are not associated with lethal liver injury.
tended cold preservation and/or transplantation, as well as However, the sinusoidal cell impairment and the liver their interactions when combined, revealed new information microcirculatory disturbances, induced by both condion the current concept of the mechanisms leading to cold tions combined, are important factors leading to secondischemia-reperfusion liver injury. ary hepatic nonfunction, which might be triggered by extrahepatic events. (HEPATOLOGY 1997;25:664-671.)
Methods
Animals. Inbred male Lewis rats (Charles River, St.-Constant, Liver transplantation has become an accepted therapy for Quebec, Canada) weighing 275 to 300 g at the time of surgery were patients with end-stage liver diseases. 1 The development of used both for donors and recipients to avoid immunologic interfer- University of Wisconsin (UW) solution has extended the sucence. Donor and recipient animals were housed in Plexiglas cages in cessful preservation time up to 24 hours, thereby increasing a temperature-and humidity-controlled environment and allowed organ availability and sharing. 2,3 Nevertheless, preservation free access to water and normal rat chow until 12 hours preceding surgery, during which they had free access to water only. The experi-time remains relatively short compared with kidney graft, ments described in this report were conducted according to the Guide and preservation injury continues to be a major clinical probfor the Care and the Use of Laboratory Animals. lem with a 10% incidence of primary nonfunction. Materials. For isolated perfused rat liver experiments, the follow-Although primary nonfunction is ultimately characterized ing materials were obtained: bovine serum albumin fraction V (Boehby severe hepatocellular failure, with impaired consciousringer, Dorval, Quebec, Canada), TC (Sigma Chemical Co., St. Louis, ness, coagulopathy, and reduced bile flow, 6,7 the underlying MO), HA (prepared from rooster comb, Sigma Chemical Co. (ca. no. mechanism of cold ischemia-reperfusion injury that leads to H-5388), and AT (BDH Chemicals, Saint-Laurent, Quebec, Canada). primary nonfunction is still unknown. It is currently hypothe-For multiple-indicator dilution curve studies, the following sub- sized that sinusoidal endothelial cell impairment [10][11][12] and/or stances were used: 51 C-labeled bovine red blood cells (RBCs) prepared with Na 2 51 CrO 2 (Frosst, Kirkland, Quebec, Canada); 3 H-water and liver microcirculatory disturbances, 13-15 mediated by oxygen 14 C-sucrose purchased from Du Pont-New England Nuclear (Boston, radicals, 13,15,16 activation of Kupffer cells, 17-19 or sinusoidal MA); 99m Tc-albumin, obtained by adding 99m Tc (St.-Luc Hospital) to human serum albumin-stannous chloride complex (Frosst). The UW solution was purchased from Dupont Critical Care (Mississauga, Ontario, Canada). Abbreviations: UW, University of Wisconsin; TC, taurocholate; AT, antipyrine; HA, hyal-Experimental Design. Animals were randomly assigned into four uronic acid; RBC, red blood cell; Tr, transplantation; LDH, lactate dehydrogenase; Cl, experimental groups (n Å 8 in each group) as follows: 1) a control hepatic clearance; Pr, preservation duration. From the Andre ´-Viallet Clinical Research Center, Ho ˆpital Saint-Luc and Universite ´de group (control) in which the experiments were performed immedi-Montre ´al,