Successful nonfreezing, subzero preservation of rat liver with 2,3-butanediol and type I antifreeze protein

Background. Organ cryopreservation is hindered by ice inflicted damage. Nonfreezing preservation of livers at subzero temperatures might offer advantages over current preservation. Methods. Sprague-Dawley rats were divided into three groups. UW livers (n = 6) were stored in University of Wisconsin (LW) solution at +4 degreesC. UWB livers (n = 6) were perfused ex vivo with UW + 10% 2,3-butanediol at less than or equal to7 degreesC and stored at -4 degreesC. AFP livers (n = 4) were preserved identical to UWB livers, except for addition of 1 mg/ml of type I antifreeze protein. After 24 h livers were perfused with Krebs-Henseleit buffer (37 degreesC) for 60 min. Bile production, O-2 consumption (O2C), taurocholate extraction, and lactate dehydrogenase (LDH) release during perfusion and liver adenine nucleotide content and energy charge at the end of perfusion were measured. Cell membrane integrity was determined by trypan blue infusion. Results. Ice formation was prevented in all livers stored at -4 degreesC. Bile production, O2C, and taurocholate extraction were similar among three groups. Livers stored at -4 degreesC contained significantly more adenine nucleotides than livers stored at +4 degreesC but the energy charge was similar. LDH release was significantly greater (P < 0.05) in the AFP group vs UWB and UW (63 vs 28 and 21 mU/min/g liver, respectively). Hepatocyte and sinusoidal cell trypan blue uptake was similar in all three groups. Conclusions. Butanediol with or without AFP was effective in preventing ice formation up to 24 h in rat livers stored at -4<degrees>C. Although as effective as current +4 degreesC protocols, subzero preservation for longer periods needs to be achieved prior to clinical application. (C) 2001 Academic Press.