Formation of multicellular spheroids composed of rat hepatocytes

A novel method for the preparation of spheroids containing two types of cells (hetero-spheroid) has been successfully developed by using a collagen-conjugated thermo-responsive polymer, poly-N-isopropyl acrylamide, as a cell substratum. The hetero-spheroid was prepared by detaching the confluent monolayer composed of parenchymal and non-parenchymal rat liver cells at a temperature below lower critical solution temperature and culturing it on the non-adhesive substratum. The hetero-spheroid had activity in albumin secretion and PNPA hydrolase activity over a period of 60 days in dishes. These findings suggest that this spheroid formation system is a useful model of an alternative to animal tests of hepatotoxicity.