Background. Nuclear factor-κB (NF-κB) and interleukin (IL)-8 play important roles in the pathophysiology of acute lung injury after lung transplantation. Because alveolar epithelium is one of the most important sites at which IL-8 production takes place after reperfusion of donor lungs, we examined the effects of cold/rewarming on NF-κB and IL-8 expression in alveolar epithelial cells. Methods. A549 cells were preserved at 4°C for 5 hr and then rewarmed for up to 20 hr. NF-κB was analyzed by electrophoretic mobility shift assay. IL-8 mRNA expression was examined by reverse transcription-polymerase chain reaction. IL-8 concentration in the cell culture medium after rewarming was measured by enzyme-linked immunosorbent assay. Results. NF-κB was increased in the nuclear extracts as early as 30 min after rewarming. There was a marked increase in the IL-8 mRNA expression at 1 and 3 hr after rewarming. IL-8 concentration in the cell culture medium was progressively increased during 20 hr following rewarming. The cell culture medium inhibited apoptosis of neutrophils significantly. The cold/rewarming-induced IL-8 production was reduced to approximately 50% by introducing an antisense oligonucleotide for the p65 subunit of NF-κB and by treatment with N-acetyl-leucinyl-leucinyl-norleucinal and pyrrolidine dithiocarbamate. The effect of dexamethasone treatment was dose dependent (reduced to approximately 30% at 10-5 M dexamethasone). Conclusions. Our results indicate that rewarming of cold-preserved alveolar epithelial cells itself may be an important initiator of the inflammatory cascades, including NF-κB activation and IL-8 release. Inhibition of NF-κB would be worth trying to control unnecessary IL-8 production and the inflammatory response in the donor lungs.
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