Energy metabolism and re-establishment of intercellular adhesion complexes of gel entrapped hepatocytes

We studied the effect of continuous medium flow on the viability and structural organization of hepatocytes high density entrapped in alginate gel beads in the first few hours after isolation. The metabolic energy status of the entrapped cells, monitored in vivo by ³¹P NMR spectroscopy, was stable during the experimental time and a physiological redox ratio was reached after the first three hours of culture. The morphological analysis revealed that the entrapped hepatocytes placed in a fixed-bed bioreactor under continuous flow showed a polyhedrical shape with numerous microvilli on cell surface and reconstituted tight junctions as well as bile canalicular structures, closely resembling those present in the liver. These results suggest that continuous flow allows the culture of hepatocytes at very high cell density within a matrix without loss of viability and accelerates cellular tissue reconstruction at very short times after isolation. This type of culture could represent a very useful model for physiological and toxicological studies as well as a promising approach toward the development of a bioartificial hybrid support device in acute liver failure.