A novel full-scale flat membrane bioreactor utilizing porcine hepatocytes:Cell viability and tissue-specific functions

When designing an extracorporeal hybrid liver support device, special atten tion should be paid to providing the architectural basis for reconstructing a proper cellular microenvironment that ensures highest and prolonged func tional activity of the liver cells. The common goal is to achieve high cell density culture and to design the bioreactor for full-scale primary liver cell cultures under adequate mass transfer conditions. An important aim of this study was to evaluate the biochemical performance of a flat membrane b ioreactor that permits high-density hepatocyte culture and simultaneously t o culture cells under sufficient oxygenation availability conditions compar able to the in vive-like microenvironment. In such a bioreactor pig liver c ells were cultured within an extracellular matrix between oxygen-permeable flat-sheet membranes. In this investigation we used a novel scaled-up proto type consisting of up to 20 modules in a parallel mode. Each module was see ded with 2 x 10(8) cells. Microscopic examination of the hepatocytes reveal ed morphological characteristics as found in vivo. Cell concentration incre ased in the first days of culture, as indicated by DNA measurements. The pe rformance of the bioreactor was monitored for 18 days in terms of albumin s ynthesis, urea synthesis, ammonia elimination, and diazepam metabolism. The ability of the hepatocytes to synthesize albumin and urea increased during the first days of culture. Higher rates of albumin synthesis were obtained at day 9 and remained at a value of 1.41 pg/h/cell until day 18 of culture . The rate of urea synthesis increased from 23 ng/h/cell to 28 ng/h/cell an d then remained constant. Cells eliminated ammonia at a rate of about 56 pg /h/cell, which was constant over the experimental period. Hepatocytes in th e bioreactor metabolized diazepam and generated three different metabolites : nordiazepam, temazepam, and oxazepam. The production of such metabolites was sustained until 18 days of culture. These results demonstrated that the scale-up of the bioreactor was assessed, and it could be demonstrated that the device design aimed at the reconstruction of the liver-specific tissue architecture supported the expression of liver-specific functions of prima ry pig liver cells.