PMID- 20654426
OWN - NLM
STAT- PubMed-not-MEDLINE
DCOM- 20121002
LR  - 20190918
IS  - 0887-2333 (Print)
IS  - 0887-2333 (Linking)
VI  - 12
IP  - 4
DP  - 1998 Aug
TI  - Use of Primary Rat and Human Hepatocyte Sandwich Cultures for Activation of 
      Indirect Carcinogens: Monitoring of DNA Strand Breaks and Gene Mutations in 
      Co-cultured Cells.
PG  - 431-44
AB  - Loss of cytochrome P-450 content is a common feature in conventional culture 
      systems of primary hepatocytes. In contrast to the standard in vitro situation, 
      in vivo each hepatocyte is exposed to an extracellular matrix (space of Disse) at 
      two opposing basolateral surfaces. This in vivo symmetry has been reconstructed 
      in vitro by culturing rat or human hepatocytes within two layers of collagen, 
      thus forming a sandwich configuration. Activation of dimethylbenzanthracene 
      (DMBA) or benzo[a]pyrene (BaP) was studied in rat and human hepatocytes. 
      Genotoxic effects were studied in a three-dimensional co-culture model between 
      sandwich hepatocytes and mammalian cells using the comet assay for detection of 
      DNA strand breaks, and the HPRT test for detection of gene mutations. Sandwich 
      hepatocytes generated active metabolites. The maintenance of metabolic properties 
      in hepatocytes was dependent on extracellular matrix geometry. The number of 
      DMBA- or BaP-induced genotoxic effects tended to be higher than in standard S-9 
      mix assays. While the ability to activate indirect carcinogens disappears within 
      hours in primary hepatocytes, hepatocyte sandwich cultures enhance their ability 
      to activate indirect carcinogens within 1 wk and retain this activity for up to 2 
      wk. This is the main advantage of the sandwich method over the more simple and 
      conventional assays. While freshly isolated hepatocytes, regardless of whether in 
      sandwich culture or in conventional assays, are injured by the isolation 
      procedure and possess a corresponding reduced activation ability, hepatocytes in 
      sandwich cultures recover over the course of a few days, and acquire a much 
      higher ability to activate indirect carcinogens. Consequently, the indirect 
      carcinogens BaP and DMBA, which were ineffective (BaP) or exhibited only weak 
      effects (DMBA) at a concentration of 160nmol/ml in 1-2-day-old hepatocytes, were 
      clearly effective (BaP) or showed about a threefold increase in genotoxicity 
      (DMBA) in 8-day-old hepatocytes in sandwich co-culture. In contrast to the 
      experiments with S-9 mix, which is toxic to mammalian cells and does not allow 
      treatment times of more than 2-3hr, cells in co-culture with human or rat 
      hepatocytes can be treated for at least 24hr. The use of sandwich cultures has 
      not yet been described for genotoxicity studies. The results of the present study 
      may perhaps facilitate the acceptance of this method as a co-culture model for 
      the field of genetic toxicology. Use of hepatocytes alone for genotoxicity 
      studies cannot be recommended for difficulties in isolating intact cells from the 
      sandwich cultures. The use of human hepatocytes in sandwich co-culture should 
      enable a more relevant evaluation of potential human genotoxicity with specific 
      chemicals and should put the extrapolation of genetic toxicology data from animal 
      species to humans on a more scientific basis. Beyond that, experiments with 
      animals in vivo could be avoided.
FAU - Fahrig, R
AU  - Fahrig R
AD  - Fraunhofer-Institute for Toxicology and Aerosol Research, Department of Genetics, 
      Nikolai-Fuchs-Strasse 1, D-30625 Hannover, Germany.
FAU - Rupp, M
AU  - Rupp M
FAU - Steinkamp-Zucht, A
AU  - Steinkamp-Zucht A
FAU - Bader, A
AU  - Bader A
LA  - eng
PT  - Journal Article
PL  - England
TA  - Toxicol In Vitro
JT  - Toxicology in vitro : an international journal published in association with 
      BIBRA
JID - 8712158
EDAT- 1998/08/01 00:00
MHDA- 1998/08/01 00:01
CRDT- 2010/07/27 06:00
PHST- 1997/10/21 00:00 [accepted]
PHST- 2010/07/27 06:00 [entrez]
PHST- 1998/08/01 00:00 [pubmed]
PHST- 1998/08/01 00:01 [medline]
AID - S0887-2333(98)00005-8 [pii]
AID - 10.1016/s0887-2333(98)00005-8 [doi]
PST - ppublish
SO  - Toxicol In Vitro. 1998 Aug;12(4):431-44. doi: 10.1016/s0887-2333(98)00005-8.