LONG-TERM CONTINUOUS-CULTURE OF HEPATOCYTES IN A PACKED-BED REACTOR UTILIZING POROUS RESIN

As part of our attempt to develop a hybrid artificial liver support sy stem using cultured hepatocytes, we investigated the long-term metabol ic function of hepatocytes incubated in a packed-bed type reactor usin g reticulated polyvinyl formal (PVF) resin as a supporting material. L ong-term (up to 1 week) perfusion culture experiments using the packed -bed reactor (20 mm i.d.) loaded with 500 PVF resin cubes (mean pore s ize 250 mu m, 2 x 2 x 2 mm), together with conventional monolayer cult ure experiments as controls, were performed in serum-free or serum-con taining medium. Ammonium metabolism and urea synthesis activities were evaluated quantitatively based on reaction kinetic analyses. Initial rates of ammonium metabolism and urea-N synthesis, as well as GPT enzy me activities, were adopted as indexes of the metabolic performance of the reactor and activities of the cultured hepatocytes. When serum-fr ee medium was used in the perfusion cultures, ammonium metabolic and u rea-N synthetic rates showed significant decay with elapse of the cult ure period, being less than 10% of those measured on day 1. This loss of activity was more prominent in the perfusion culture than in the mo nolayer cultures using this medium. In contrast, when serum-containing medium was used, approximately 50% of these activities obtained on da y 1 were maintained even at the end of the cultures both in the perfus ion and monolayer culture experiments. We concluded that the packed-be d reactor using PVF resin enabled high-density culture of hepatocytes, and showed a satisfactory ability to maintain the metabolic function of immobilized hepatocytes for relatively long periods of up to 1 week . This type of reactor is thus considered tb represent a breakthrough in overcoming the difficulties involved in the development of a hybrid -type artificial liver support system. (C) 1994 John Wiley and Sons, I nc.