title: Comparative Proteomic Analysis of Human Liver Tissue and Isolated Hepatocytes with a Focus on Proteins Determining Drug Exposure.
abstract: Freshly isolated human hepatocytes are considered the gold standard for in vitro studies of liver functions, including drug transport, metabolism, and toxicity. For accurate predictions of the in vivo outcome, the isolated hepatocytes should reflect the phenotype of their in vivo counterpart, i.e., hepatocytes in human liver tissue. Here, we quantified and compared the membrane proteomes of freshly isolated hepatocytes and human liver tissue using a label-free shotgun proteomics approach. A total of 5144 unique proteins were identified, spanning over 6 orders of magnitude in abundance. There was a good global correlation in protein abundance. However, the expression of many plasma membrane proteins was lower in the isolated hepatocytes than in the liver tissue. This included transport proteins that determine hepatocyte exposure to many drugs and endogenous compounds. Pathway analysis of the differentially expressed proteins confirmed that hepatocytes are exposed to oxidative stress during isolation and suggested that plasma membrane proteins were degraded via the protein ubiquitination pathway. Finally, using pitavastatin as an example, we show how protein quantifications can improve in vitro predictions of in vivo liver clearance. We tentatively conclude that our data set will be a useful resource for improved hepatocyte predictions of the in vivo outcome.
authors.forename: Anna
authors.initials: A
authors.lastname: Vildhede
authors.affiliation: §Biochemical Proteomics Group, Department of Proteomics and Signal Transduction, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany.
authors.forename: Jacek R
authors.initials: JR
authors.lastname: Wiśniewski
authors.forename: Agneta
authors.initials: A
authors.lastname: Norén
authors.forename: Maria
authors.initials: M
authors.lastname: Karlgren
authors.affiliation: ∥Uppsala University Drug Optimization and Pharmaceutical Profiling Platform (UDOPP), Chemical Biology Consortium, Science for Life Laboratory, 750 03 Uppsala, Sweden.
authors.forename: Per
authors.initials: P
authors.lastname: Artursson
mesh_terms: D000368:Aged; D001692:Biological Transport; D002478:Cells, Cultured; D002853:Chromatography, Liquid; D005260:Female; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000493:pharmacokinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D008875:Middle Aged; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D011804:Quinolines / Q000493:pharmacokinetics; D015398:Signal Transduction; D053719:Tandem Mass Spectrometry
chemical_list: D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D008565:Membrane Proteins; D020543:Proteome; D011804:Quinolines; C108475:pitavastatin
keywords: ADME proteins; CYP enzymes; UGT enzymes; drug transporters; human hepatocytes; in vitro−in vivo extrapolation; membrane proteome; oxidative stress; quantitative proteomics; total protein approach