Expression of Glutamine Synthetase and Carbamoylphosphate Synthetase I in a Bioartificial Liver: Markers for the Development of Zonation in vitroPoyck P.P.C.a · Hoekstra R.a, b · Vermeulen J.L.M.b · van Wijk A.C.W.A.a · Chamuleau R.A.F.M.c · Hakvoort T.B.M.b · van Gulik T.M.a · Lamers W.H.b
aDepartment of Surgery (Surgical Laboratory), bAMC Liver Center, and cDepartment of Gastroenterology and Hepatology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands
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Background: Mechanisms underlying hepatic zonation are not completely elucidated. In vitro test systems may provide new insights into current hypotheses. In this study, zonally expressed proteins, i.e. glutamine synthetase (GS; pericentral) and carbamoylphosphate synthetase (CPS; periportal), were tested for their expression patterns in the bioartificial liver of the Academic Medical Center (AMC-BAL). Methods: Distribution and organization of porcine hepatocytes inside the AMC-BAL as well as GS and CPS expression were analyzed (immuno-)histochemically in time. Ten zonally expressed proteins were analyzed by RT-PCR on cell isolate and bioreactor samples. General metabolic and hepatocyte-specific functions were determined as well. Results: Viable hepatocyte layers of approximately 150 µm were observed around gas capillaries, whereas inside the matrix, single cells or small aggregates were present. GS protein and mRNA levels were upregulated in time. GS protein was preferentially expressed in hepatocytes adjacent to oxygen-supplying capillaries and in previously CPS-positive hepatocytes. No shift towards a periportal or pericentral phenotype was observed from RT-PCR analysis. Conclusion: Induction of GS expression inside the AMC-BAL is not dependent of (low) oxygen tensions and hepatic nuclear factor 4α transcript levels. GS expression might be related to (1) low substrate levels and/or autocrine soluble factors, or (2) to cytoskeleton interactions, putatively associated with the β-catenin signaling pathway.
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