Protective effect of donor pretreatment with carbon monoxide on ischemia/reperfusion injury after rat liver transplantation; possible role of Hsp 70 expression

Abstract

Introduction: Stress response genes, such as heme oxygenase-1 (HO-1) and heat shock protein 70 (HSP70), are known to be involved in ischemia/reperfusion (I/R) injury and have beneficial effects against oxidative stress. Carbon monoxide (CO), the byproduct of heme degradation by HO-1, has been shown to provide protection against I/R injury. This study was designed to examine whether donor pretreatment with CO has beneficial effects on hepatic cold I/R injury. Methods: Orthotopic syngenic LEW rat liver transplantation (OLT) was performed after 18 hrs preservation in cold UW. Donor rats were exposed to a non-toxic dose of CO (250 ppm) in a CO chamber for 24 hrs before harvesting. Controls donors were maintained in room air. Hepatic I/R injury was evaluated by serum AST levels and histopathology. Expression of HSP70 protein in donor liver was investigated before harvest. After OLT, expression of TNF-α and ICAM-1 mRNA, iNOS protein, and cleaved caspase-3 were measured. Results (Table):TABLE—ABSTRACT 107AST (IU/L) 24 hNeutrophil (/mm2) 48 hTNF-α (fold increase) 1 hiNOS (band denity) 3 hCaspase-3 (band denity) 3 hAir5547 ± 97987.1 ± 15.214.7 ± 5.326.3 ± 5.33.8 ± 0.3CO1981 ± 381∗23.6 ± 8.6∗4.8 ± 1.5∗7.5 ± 2.1∗2.8 ± 0.1∗Mean ± SD, n = 2–6 animals per group,∗P < 0.05. CO-inhalation for 24 hrs significantly increased HSP70 expression in the donor liver. After OLT, donor CO pretreatment markedly attenuated hepatic I/R injury assessed by low AST levels, reduced necrosis, and decreased neutrophil infiltration. Hepatic expression of TNF-α and ICAM-1 mRNA, and iNOS protein was significantly reduced in CO-treated group. Additionally, cleaved caspase-3 expressing apoptotic cells were significantly decreased with donor CO-inhalation. Conclusion: These results demonstrate that donor CO pretreatment leads to attenuation of hepatic cold I/R injury through the inhibition of iNOS induction, pro-inflammatory responses, and pro-apoptotic reactions. The protective effect of donor CO pretreatment is in part mediated by HSP70 induction in the donor liver. Thus, low-dose CO inhalation would be a novel therapeutic strategy to combat hepatic cold I/R injury.