Xenogeneic hepatocytes have recently been used in a bioartificial liver device as a potential short-term extracorporeal support of acute liver failure. Scaling up the system requires large quantities of viable and highly active cells. Hepatocytes grown as spheroids manifest higher metabolic activities for longer time periods as compared to those in monolayer cultures. Use of hepatocyte spheroids for application in a bioartificial liver can possibly alleviate the need of scaling up. Porcine hepatocytes when cultured under stirred conditions, from multicellular spheroids in a defined culture medium. Spheroids were formed 24 h after cell inoculation with an efficiency of 80-90% and a mean diameter of about 135 microns. Scanning electron microscopy revealed numerous microvilli projecting from the entire surface of the spheroids. Transmission electron microscopy revealed differentiated hepatocytes which displayed well-developed cytoplasmic structures separated by bile canaliculus-like structures. The morphological studies show a resemblance between cells in the spheroids and in the liver in vivo. Urea-genesis by spheroids was twice as active and was sustained for a longer culture period than that by hepatocytes cultured as monolayers. Preparation of porcine hepatocyte spheroids in an agitated vessel is simple efficient and reproducible. It will allow for preparation of large quantities of spheroids to be employed in a bioartificial liver device as well as in liver metabolism studies.

The population dynamics of developing somatic embryos of carrot (Daucur carota L.) was investigated in batch and fed-batch cultures using modified Murashige and Skoog medium. These substrate limitations coincided not only with stoppage of biomass increase, but also with the increase in total concentration of embryos as well as the advancement of the embryo into a more mature stage. Both glucose and ammonium were depleted from the culture. Restoring either glucose, or ammonium and nitrate, as to approximately initial concentrations in fed-batch experiments, did not result in a significant increase of the total normal embryo concentration. On the other hand, medium replacement led to increase in biomass concentration, total embryo number, and improved embryo maturity. The addition of a mixture of glucose, ammonium, and nitrate to the spent medium resulted in variable increases in biomass and embryo number, but always less than those resulting from media replacement. Although the total number of embryos was higher after medium replacement, the fraction of embryos reaching torpedo stage was still only 50%. The need for a better means of population characterization for further kinetic studies is discussed.