High density culturing of porcine hepatocytes immobilized on nonwoven polyurethane-based biomatrices

Objective: Hepatocytes are increasingly used as functional units in bioarti ficial liver devices. The objective of the present study was to investigate the feasibility of culturing porcine hepatocytes in high density on a nove l polyurethane-based nonwoven three-dimensional matrix. We investigated (1) the optimal cell density within this culture configuration, (2) the mainte nance of liver-specific morphology and cell functions over long-term period s and (3) the necessity to apply an additional extracellular matrix compone nt (collagen gel). Methods: Nonwoven polyurethane matrices were manufactured by a specially de veloped fiber extrusion technology. Pig hepatocytes were cultured at variou s cell densities of 0.1, 0.25, 0.5, 0.75, 1 and 2 x 10(6) cells/cm(2) on th ree-dimensional networks of nonwoven polyurethane matrices and cell adhesio n as well as functional parameters (DNA of nonattached/attached cells, lact ate dehydrogenase release and cytochrome P450 activity) were determined. To assess the performance of cells within this configuration albumin and urea excretion was measured over 8 days. The potentially beneficial effect of a n additional extracellular matrix configuration was evaluated by comparing the average albumin synthesis in groups of identical cell numbers. Results: The optimal cell density in this three-dimensional culture configu ration was 1 x 10(6) cells/cm(2). The functional capacity of hepatocytes wa s stable for 8 days at an average level of 53.7 +/- 5.6 ng/h/mug DNA and of 1.8 +/- 0.14 mug/h/mug DNA for albumin and urea excretion, respectively. T he supplementation of an extracellular matrix configuration did not improve functional activity of cells. Average albumin synthesis was 35.6 ng/h/mug DNA (28.7, 42.8) and 32.7 ng/h/mug DNA (23.4, 49.2) for collagen-immobilize d and control cultures, respectively, Conclusion: The results of the study indicate that nonwoven polyurethane sh eets supply a biocompatible support structure for functionally active high density cultures. Thus, nonwoven polyurethane matrices should be further in vestigated on with respect to their role in the development, optimization a nd design of bioartificial liver systems. Copyright (C) 2001 S.Karger AG, B asel.