A compact three-dimensional (3D) module is needed for hepatocyte culture in order to develop an effective hybrid artificial liver system that can retain hepatocellular structure and differentiated functions. We treated the 3D module with collagen gel to entrap rat hepatocytes. This method yielded a high hepatocellular density (2 x 10(7) cells/ml) over a period of 14 days and maintained the secretion of albumin and ureogenesis at the same level as the control monolayer method. The ammonia removal remained at 43% of the Day 0 value over 8 days of perfusion. Our data show that this approach may be useful for liver support therapy in an extracorporeal circuit.