Large-scale preparation and function of porcine hepatocyte spheroids

To obtain a large number of porcine hepatocyte aggregates (spheroids) that have great potential in a bioartificial liver (BAL), we performed spheroid formation at a high cell density in a 1-L-scale spinner flask fitted with a silicon tubing apparatus for oxygen supply. We thereby obtained, within 24 hours, approximately fifty times more porcine hepatocyte spheroids as compared with the results of previous reports. The amount obtained corresponds to 2.5x10(9) cells and to roughly one-sixth of the cell number required for a BAL for a human patient. When we cultured spheroids continuously in suspension, they expressed three times more albumin secretion and twice the ammonium removal as compared with conventional monolayers during 10 days culture. Collagen gel entrapment of spheroids particularly lowered albumin secretion. We therefore conclude that the supension culture vessel of porcine hepatocyte spheroids is one of the most promising module types.