This study compared the metabolism of the tobacco procarcinogen, benzo(a)pyrene (BP), in monolayer cultures vs. 3-dimensional culture models using rat mammary epithelial (MEC) and fibroblast (MFB) cells derived from a lacl Fischer rat and determined whether MFBs could enhance metabolism of BP by MECs. Also, the metabolism of BP by MFBs and MECs was compared.
We hypothesized that MFBs would metabolize BP to genotoxic (DNA damaging) products at a higher rate when cultured as aggregates (3-D culture) compared to monolayers, and in a co-culture with MECs , would lead to additive/synergistic interaction.
We assessed genotoxic metabolism of BP by measuring metabolites of BP in culture medium, and in DNA from cells. This was done using HPLC. First we compared the effects of monolayer versus aggregates of MFBs. Next, we compared the effects of BP in co-cultures of varying ratios of epithelial and fibroblast cells. Lastly, we used a chamber system to determine whether, in the presence of BP, MFB aggregates in one chamber led to increased damage to DNA in the MEC chamber.
When MFBs were grown in aggregates their genotoxic metabolism of BP was slower than that of cells in monolayers. When MECs were grown in co-culture with MFBs, metabolism was enhanced, but we cannot yet determine whether any synergism between the cell types occurred. In the chamber assay, an increase in BP concentration applied to the MFBs led to an increased overall rate of genotoxic metabolism cells, but DNA from the MECs was not yet analyzed.
MFBs metabolize BP to genotoxic products more efficiently than MECs, consistent with parallel experiments on mutagenesis induced by BP. 3-D cultures of MFBs did not appear to be more effective than monolayers at metabolizing BP. Co-cultures and chamber cultures may prove useful in investigating interactions between MFBs and MECs.
Keywords: Biochemistry, Epithelium/epithelial, Fibroblasts, Histology - ultrastructure and Tobacco