tandard preservation of donor livers is performed by flushing the organ with a cold solution at the time of procurement, which is followed by static storage on ice. This approach reduces liver metabolic activity, allowing safe storage for up to 12-18 h (refs. 1,2). Recently, the combination of supercooling with subzero storage (−4 °C) and hypothermic, as well as subnormothermic, machine perfusion achieved an extension of the preservation time of human livers up to 27 h (ref. 2). By contrast, human livers can also be kept metabolically fully active for up to 24 h by supplying normothermic blood or oxygenated fluids in a controlled environment by machine perfusion 1,3-5. The possibility of repairing poor-quality livers sufficiently to enable transplantation requires preserving metabolically active livers 1,3 for several days. Accordingly, the need for long-term organ preservation technology has been endorsed by multiple private and governmental initiatives 1. However, currently used normothermic machine perfusion technologies have been used only for a relatively short time period (for example, a median perfusion time of 9 h (ref. 5)), to successfully maintain hemodynamics, perfusate oxygenation and temperature. We sought to extend perfusion time beyond 24 h by engineering a perfusion machine that recapitulates additional core body functions that are critical to liver health. We chose 7-d as a target because this time period has been shown to offer a credible time frame for inducing clinically relevant liver regeneration in patients undergoing complex liver resection 6-8. Under physiologic conditions, the liver, which constitutes 2.5% of body weight, receives 25% of the blood output of the heart and performs >5,000 functions 9. It has a unique dual vascular supply with high-pressure, oxygen-rich arterial blood entering through the hepatic artery and low-pressure, oxygen-reduced portal vein blood draining the abdominal viscera. Its high metabolic activity produces waste products that are excreted in the bile or removed by hepatic macrophages or the kidneys. The metabolic profile is largely controlled by pancreatic hormones, including insulin and glucagon. Our perfusion technology, developed in the "Liver4Life project", includes automated control of glucose levels by injection of insulin and glucagon, a dialysis membrane for waste-product removal, regulation of oxygenation and liver movement to prevent pressure necrosis. Results Liver perfusion machine. Our perfusion machine (Fig. 1a,b) recapitulates blood supply through the two vascular entries of the liver, the hepatic artery and portal vein. The hepatic artery is supplied with oxygen-rich blood at elevated pressure (mean arterial pressure (MAP) ≥ 65 mmHg) in a pulsatile manner (Fig. 1c), whereas the portal vein receives blood at low pressure (around 5-10 mmHg) with a reduced oxygen content (venous blood, non-pulsatile). The system maintains oxygen saturation of 65% in the vena cava by continuously adjusting oxygen content in the portal vein (Fig. 1d). In vivo, nutr...
