DEVELOPMENT OF A BIOARTIFICIAL LIVER EMPLOYING XENOGENEIC HEPATOCYTES

Liver failure is a major cause of mortality. A bioartificial liver (BA L) employing isolated hepatocytes can potentially provide temporary su pport for liver failure patients. We have developed a bioartificial li ver by entrapping hepatocytes in collagen loaded in the luminal side o f a hollow fiber bioreactor. In the first phase of development, liver- specific metabolic activities of biosynthesis, biotransformation and c onjugation were demonstrated. Subsequently anhepatic rabbits were used to show that rat hepatocytes continued to function after the BAL was linked to the test animal. For scale-up studies, a canine liver failur e model was developed using D-galactosamine overdose. In order to secu re a sufficient number of hepatocytes for large animal treatment, a co llagenase perfusion protocol was established for harvesting porcine he patocytes at high yield and viability. An instrumented bioreactor syst em, which included dissolved oxygen measurement, pH control, flow rate control, an oxygenator and two hollow fiber bioreactors in series, wa s used for these studies. An improved survival of dogs treated with th e BAL was shown over the controls. In anticipated clinical application s, it is desirable to have the liver-specific activities in the BAL as high as possible. To that end, the possibility of employing hepatocyt e spheroids was explored. These self-assembled spheroids formed from m onolayer culture exhibited higher liver-specific functions and remaine d viable longer than hepatocytes in a monolayer. To ease the surface r equirement for large-scale preparation of hepatocyte spheroids, we suc ceeded in inducing spheroid formation in stirred tank bioreactors for both rat and porcine hepatocytes. These spheroids formed in stirred ta nks were shown to be morphologically and functionally indistinguishabl e from those formed from a monolayer. Collagen entrapment of these sph eroids resulted in sustaining their liver-specific functions at higher levels even longer than those of spheroids maintained in suspension. For use in the BAL, a mixture of spheroids and dispersed hepatocytes w as used to ensure a proper degree of collagen gel contraction. This mi xture of spheroids and dispersed cells entrapped in the BAL was shown to sustain the high level of liver-specific functions. The possibility of employing such a BAL for improved clinical performance warrants fu rther investigations.