MULTIPOROUS CELLULOSE MICROCARRIER FOR THE DEVELOPMENT OF A HYBRID ARTIFICIAL LIVER USING ISOLATED HEPATOCYTES

Background. This study was aimed at developing an optimal method for i mmobilizing isolated hepatocytes in cellulose multiporous microcarrier s (MCs) and evaluating the metabolic activity of MC-immobilized hepato cytes. Materials and Methods. Hepatocytes isolated from the livers of male Wistar rats were immobilized in collagen-coated MCs by intermitte nt stirring (30 rpm for 2 min per 15 min) for 180 min or accumulation methods. The accumulation method was performed by pouring aliquots of hepatocyte suspension (8 x 10(5)) and MC suspension (1 mg) in turn ont o a nylon mesh (pore size: 100 mu m). The metabolic activity of MC-imm obilized hepatocytes in floating culture and in a newly developed bior eactor was evaluated. The metabolic activity of MC-immobilized hepatoc ytes in the bioreactor was also evaluated in in vitro perfusion of a h ollow-fiber-based hybrid artificial liver support system. Results. The accumulation method immobilized 20 times more hepatocytes in collagen coated MCs than the intermittent stirring method (P < 0.01). Morpholo gical observation of hepatocyte-immobilized MCs revealed that many hep atocytes were immobilized deep within the MCs maintaining a spherical shape and normal microvilli on their surface. MC-immobilized hepatocyt es in floating culture revealed similar NH3 metabolism and glucose syn thesis to monolayer-cultured hepatocytes, and this metabolic activity was maintained during 9 h of floating culture. MC-immobilized hepatocy tes in a bioreactor also showed similar NH3 metabolism to monolayer-cu ltured hepatocytes. The NH3 metabolism of MC-immobilized hepatocytes i n in vitro perfusion of a hybrid artificial liver support system was 2 41.5 mu g/h/mg protein/m(2) membrane surface. Conclusions. The results of this study indicate that the accumulation method was optimal for i mmobilizing isolated rat hepatocytes in MCs and that MC-immobilized he patocytes maintained their metabolic activity for a long period. (C) 1 998 Academic Press.