Liver Progenitor Cell Line HepaRG Differentiated in a Bioartificial Liver Effectively Supplies Liver Support to Rats with Acute Liver Failure

Abstract

A major roadblock to the application of bioartificial livers is the need for a human liver cell line that displays a high and broad level of hepatic functionality. The human bipotent liver progenitor cell line HepaRG is a promising candidate in this respect, for its potential to differentiate into hepatocytes and bile duct cells. Metabolism and synthesis of HepaRG monolayer cultures is relatively high and their drug metabolism can be enhanced upon treatment with 2% dimethyl sulfoxide (DMSO). However, their potential for bioartificial liver application has not been assessed so far. Therefore, HepaRG cells were cultured in the Academic Medical Center bioartificial liver (AMC-BAL) with and without DMSO and assessed for their hepatic functionality in vitro and in a rat model of acute liver failure. HepaRG-AMC-BALs cultured without DMSO eliminated ammonia and lactate, and produced apolipoprotein A-1 at rates comparable to freshly isolated hepatocytes. Cytochrome P450 3A4 transcript levels and activity were high with 88% and 37%, respectively, of the level of hepatocytes. DMSO treatment of HepaRG-AMC-BALs reduced the cell population and the abovementioned functions drastically. Therefore, solely HepaRG-AMC-BALs cultured without DMSO were tested for efficacy in rats with acute liver failure (n = 6). HepaRG-AMC-BAL treatment increased survival time of acute liver failure rats ∼50% compared to acellular-BAL treatment. Moreover, HepaRG-AMC-BAL treatment decreased the progression of hepatic encephalopathy, kidney failure, and ammonia accumulation. These results demonstrate that the HepaRG-AMC-BAL is a promising bioartificial liver for clinical application.

Phase 1 and Phase 2 Drug Metabolism and Bile Acid Production of HepaRG Cells in a Bioartificial Liver in Absence of Dimethyl Sulfoxide

“…2B). The basal activity was 11.2 6 4.0 nmol/h/mg protein, 37% the rate of PHHs (historical data) (Nibourg et al, 2012).…”

“…HepaRG cells were kindly provided by C. Guguen-Guillouzo (INSERM, Rennes, France). At day 0, laboratory model (9 ml) AMC-BALs were loaded with~750 million cells and cultured in 500 ml recirculating HepaRG-CG medium, i.e., HepaRG medium without DMSO, but supplemented with 1 mM N-carbamoyl-L-glutamate (Sigma-Aldrich, St. Louis, MO) (Nibourg et al, 2012).…”

“…The total cell mass is stable under the given bioreactor culture conditions (Nibourg et al, 2012), and therefore the bilirubin glucuronidation data obtained from HepaRG-AMC-BALs at day 11 were normalized for protein content of the cell lysate of HepaRG-AMC-BALs at day 14. …”

“…Culturing HepaRG in three dimensions in a bioreactor, the AMCbioartificial liver (AMC-BAL), for only 14 days without DMSO generated viable cultures mimicking the human liver to high extent (Nibourg et al, 2012): the HepaRG-AMC-BALs eliminated ammonia and lactate and produced apolipoprotein A-1 at rates comparable with freshly isolated hepatocytes. Moreover, CYP3A4 transcript level was high with 88% of the level of PHHs.…”

“…The transcript levels of nuclear receptors hepatocyte nuclear factor 4 and pregnane X receptor, both central players in the regulation of drug metabolism (Tirona et al, 2003), were between 100 and 150% of human liver. Furthermore, a study in rats with acute liver failure showed that the HepaRG-AMC-BAL replaced the liver function, leading to increased survival (Nibourg et al, 2012).…”