Hepatic cold hypoxia and oxidative stress: implications for ICAM-1 expression and modulation by glutathione during experimental isolated liver preservation.
Cold preservation and reperfusion of liver during transplantation are necessary steps in the procedure but which are also associated with damage to the organ. One aspect of this damage is thought to concern up-regulation of inflammatory markers, such as the adhesion molecule intercellular adhesion molecule 1 (ICAM-1) on target cells in the liver. This aids sequestration of activated leucocytes, which promote inflammation, by a complex sequence of events, including free radical mediated damage. We have studied changes in ICAM-1 in rat liver as a consequence of cold preservation for various times, and also after warm reperfusion during isolated liver perfusion. We have also investigated the effects of the free radical scavenging agent (reduced glutathione-GSH) on the modulation of ICAM-1 expression after cold hypoxia and reperfusion. Livers were subjected to various regimes of cold preservation and reperfusion. Liver biopsies were taken at three time points (initial baseline on liver exposure; after organ flushing and post-storage at 0, 8, 16, and 24h cold hypoxia in University of Wisconsin solution; in the same livers after 1h warm reperfusion). The tissues were processed for frozen biopsy work, and frozen sections were stained using immunohistochemical methods, for blinded scoring by an independent observer. Positive controls were obtained by exposure to endotoxin lipopolysaccharide before liver flushing. ICAM-1 expression was low in control livers (0.33+/-0.21), and increased to near maximal (2.83+/-0.17) after endotoxin exposure. ICAM-1 expression increased progressively with cold preservation, reaching values of 1.17+/-0.31 and 1.83+/-0.31 after 16 and 24h, respectively (P<0.05 and 0.02 versus controls). Warm reperfusuion increased ICAM-1 expression in all flushed groups and with longer cold preservation was close to maximal (2.67+/-0.21 after 16h and 2.98+/-0.02 after 24h; P<0.001 in both cases). Addition of the free radical scavenger GSH prevented up-regulation of ICAM-1 in livers reperfused after flushing and cold storage for up to 8h; beyond this time, ICAM-1 expression still increased, such that by 24h cold preservation and reperfusion absence (2.98+/-0.02) or presence (2.67+/-0.21) made no difference. We conclude that liver ICAM-1 expression is demonstrably increased by progressive cold preservation and reperfusion, and is only marginally affected by addition of GSH during reperfusion. The model can be used to investigate other agents which might be more successful in preventing post-storage inflammatory damage.
University Department of Surgery and Liver Transplant Unit, Royal Free and University College Medical School, London, United Kingdom.
M
M
El-Wahsh
B
B
Fuller
B
B
Davidson
K
K
Rolles
D000818:Animals; D002448:Cell Adhesion; D003080:Cold Temperature; D005609:Free Radicals; D005978:Glutathione / Q000378:metabolism*; D000860:Hypoxia; D007249:Inflammation; D018799:Intercellular Adhesion Molecule-1 / Q000096:biosynthesis* / Q000378:metabolism; D007511:Ischemia; D007962:Leukocytes / Q000378:metabolism; D008099:Liver / Q000378:metabolism / Q000473:pathology*; D008297:Male; D009926:Organ Preservation / Q000379:methods*; D018384:Oxidative Stress; D010477:Perfusion; D051381:Rats; D017207:Rats, Sprague-Dawley; D015424:Reperfusion; D013997:Time Factors; D015854:Up-Regulation
D005609:Free Radicals; D018799:Intercellular Adhesion Molecule-1; D005978:Glutathione