Pathway and time-resolved benzo[a]pyrene toxicity on Hepa1c1c7 cells at toxic and subtoxic exposure.
Stefan Kalkhof, Franziska Dautel, Salvatore Loguercio, Sven Baumann, Saskia Trump, Harald Jungnickel, Wolfgang Otto, Susanne Rudzok, Sarah Potratz, Andreas Luch, Irina Lehmann, Andreas Beyer, Martin von Bergen
Benzo[a]pyrene (B[a]P) is an environmental contaminant mainly studied for its toxic/carcinogenic effects. For a comprehensive and pathway orientated mechanistic understanding of the effects directly triggered by a toxic (5 μM) or a subtoxic (50 nM) concentration of B[a]P or indirectly by its metabolites, we conducted time series experiments for up to 24 h to study the effects in murine hepatocytes. These cells rapidly take up and actively metabolize B[a]P, which was followed by quantitative analysis of the concentration of intracellular B[a]P and seven representative degradation products. Exposure with 5 μM B[a]P led to a maximal intracellular concentration of 1604 pmol/5 × 10(4) cells, leveling at 55 pmol/5 × 10(4) cells by the end of the time course. Changes in the global proteome (>1000 protein profiles) and metabolome (163 metabolites) were assessed in combination with B[a]P degradation. Abundance profiles of 236 (both concentrations), 190 (only 5 μM), and 150 (only 50 nM) proteins were found to be regulated in response to B[a]P in a time-dependent manner. At the endogenous metabolite level amino acids, acylcarnitines and glycerophospholipids were particularly affected by B[a]P. The comprehensive chemical, proteome and metabolomic data enabled the identification of effects on the pathway level in a time-resolved manner. So in addition to known alterations, also protein synthesis, lipid metabolism, and membrane dysfunction were identified as B[a]P specific effects.
