Modular bioreactor for primary human hepatocyte culture: medium flow stimulates expression and activity of detoxification genes

Biotechnol J. 2011 May;6(5):554-64. doi: 10.1002/biot.201000326. Epub 2011 Jan 21.

Abstract

Down-regulation of detoxification genes, notably cytochrome P450 (CYPs), in primary hepatocyte cultures is a long-standing and major concern. We evaluated the influence of medium flow in this model. Hepatocytes isolated from 12 different liver donors were cultured either in a multichamber modular bioreactor (MCmB, flow rate 250-500 μL/min) or under standard/static conditions, and the expression of 32 genes, enzyme activities and biological parameters were measured 7-21 days later. mRNA expression of genes involved in xenobiotic/drug metabolism and transport, including CYP1A1, 1A2, 2B6, 2C9, 3A4 (and activities for some of them), UDP-glucuronosyltransferase (UGT) 1A1, UGT2B4, UGT2B7, glutathione S-transferase (GSTα), and multidrug resistance protein 1 (MDR1) and MRP2, were specifically up-regulated by medium flow as compared with static controls in all cultures tested. In 2-week-old cultures, expression of detoxification genes reached levels close to or higher than those measured in freshly isolated hepatocytes. In contrast, CYP2D6 and most of other tested genes were not affected by medium flow. We conclude that medium flow specifically interferes with, and up-regulates, the activity of xenosensors and/or the expression of detoxification genes in primary human hepatocytes. Down-regulation of detoxification genes in conventional (static) cultures is therefore partly a consequence of the absence of medium circulation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATP Binding Cassette Transporter, Subfamily B / genetics
  • ATP Binding Cassette Transporter, Subfamily B, Member 1 / genetics
  • Aryl Hydrocarbon Hydroxylases / genetics
  • Bioreactors*
  • Cells, Cultured
  • Cytochrome P-450 CYP1A1 / genetics
  • Cytochrome P-450 CYP1A2 / genetics
  • Cytochrome P-450 CYP2B6
  • Glucuronosyltransferase / genetics
  • Glutathione Transferase / genetics
  • Hepatocytes / cytology
  • Hepatocytes / metabolism*
  • Humans
  • Oxidoreductases, N-Demethylating / genetics
  • Reverse Transcriptase Polymerase Chain Reaction
  • Shear Strength

Substances

  • ATP Binding Cassette Transporter, Subfamily B
  • ATP Binding Cassette Transporter, Subfamily B, Member 1
  • P-glycoprotein 2
  • Aryl Hydrocarbon Hydroxylases
  • CYP2B6 protein, human
  • Cytochrome P-450 CYP1A1
  • Cytochrome P-450 CYP1A2
  • Cytochrome P-450 CYP2B6
  • Oxidoreductases, N-Demethylating
  • UGT1A1 enzyme
  • UGT2B7 protein, human
  • Glucuronosyltransferase
  • UGT2B4 protein, human
  • Glutathione Transferase