As the number of patients on the waiting list for organ transplantation is increasing, the expansion of the donor spectrum is of utmost importance. Steatosis is a common finding and in most cases the reason for organ refusal or dysfunction after transplantation. Because apoptotic cell death plays a pivotal role in the pathogenesis of the preservation-reperfusion injury of the liver, the aim of our study was to investigate whether transient inhibition of p53-dependent apoptosis by the reversible p53-inhibitor pifithrin-α (PFT-α) is able to reduce apoptotic cell death in fatty livers. After induction of steatosis using dietary manipulation, fatty livers were harvested, stored for 24h in PFT-α-(20 µM) or DMSO-containing histidine-tryptophane-ketoglutarate (HTK) solution at 4 °C, and then reperfused for 2 h using an isolated perfused liver model. Apoptotic cell death was analyzed by means of epi-illumination fluorescence microscopy. In addition, bile flow, liver enzyme release in the effluent, O2-consumption and CO2-production were determined to indicate hepatocellular function, integrity and metabolism. The 2-h reperfusion following cold storage induced significant reperfusion injury. Transient inhibition of p53 was capable of ameliorating this injury by reducing apoptotic cell death and liver enzyme release, and by increasing bile flow, however, without affecting liver metabolic parameters. Therefore, we conclude that the inhibition of p53 during cold preservation represents a valid strategy for improving the impaired organ function of fatty livers subjected to cold preservation and reperfusion.