Protection by Carolina rinse solution, acidotic pH, and glycine against lethal reperfusion injury to sinusoidal endothelial cells of rat livers stored for transplantation

The critical injury causing graft failure after prolonged liver storage involves reperfusion-induced killing of sinusoidal endothelial cells and activation of Kupffer cells. Treatment of stored livers with Carolina rinse solution (CRS) prevents endothelial cell killing, reduces Kupffer cell activation, and improves graft survival. Accordingly, our aim was to evaluate the components of CRS and other agents for protection against reperfusion injury to rat livers stored 24 hr in University of Wisconsin solution. CRS virtually abolished endothelial cell killing, prevented denudation of the sinusoidal lining, and decreased structural changes in Kupffer cells indicative of activation. The only component of CRS preventing endothelial cell killing was acidic pH of 6.5. However, when pH was subsequently increased to 7.4, antioxidants (allopurinol, deferoxamine mesylate, and glutathione), vasodilators (adenosine and nicardipine), and possibly energy substrates (fructose, glucose, and insulin) partially blocked pH-dependent cell killing (pH paradox). Na+/H+ exchange inhibition, protease inhibition, and Ca2+-free buffer did not decrease reperfusion injury, but the amino acid glycine protected strongly. Strychnine, which binds to glycine receptors in the central nervous system, protected equally well. Protection by glycine and CRS was synergistic, virtually eliminating cell killing after 48 hr of storage. Unlike CRS and some of its components (adenosine and Ca2+ channel blocker), glycine did not block tumor necrosis factor-alpha release by cultured rat Kupffer cells, although gabexate mesylate, a protease inhibitor, strongly blocked tumor necrosis factor-alpha release. In conclusion, the combination of CRS and glycine enhanced protection against reperfusion injury to stored livers and may prove beneficial in improving graft function and survival after liver transplantation.