Since epidemiological studies have implicated the co-exposition of iron oxides and polycyclic aromatic hydrocarbons as potential etiological factors involved in the excess of mortality from lung cancer in miners, experimental studies have been performed to investigate the role of iron on benzo[a]pyrene (B[a]P)-induced lung pathogenesis. We demonstrated previously that in vivo damage was higher when B[a]P was coated onto hematite than when B[a]P was administered alone. In order to determine the role of (i) different cell types and (ii) adsorption of hematite in this potentiation, in vitro studies were developed. The Comet assay was first used to measure DNA damage in four isolated cell types from Sprague-Dawley rats at 1, 2, 4, 8 and 24h after in vitro treatment with hematite (Fe2O3) or B[a]P or B[a]P coated onto hematite. For the two treatments with B[a]P, no damage was observed in alveolar macrophages, but significant increases in damage were seen in lymphocytes, hepatocytes and lung cells (where the effects of B[a]P coated onto hematite were stronger than those of B[a]P alone). In a second part of the study, the Comet assay was conducted with lung cells to measure the in vitro effect of (i) the coating and (ii) the role of the physical properties of Fe2O3. A statistically significant increase in damage was observed for the coating of B[a]P onto Fe2O3 compared (i) with their simple addition and (ii) with the coating of B[a]P onto graphite used as an inert compound. This study showed that (i) Fe2O3/B[a]P acts essentially in lung cells, (ii) the coating is a primordial step and (iii) the physical properties of Fe2O3 play a very minor role, which suggests another mechanism of action to explain the higher toxicity. Hence, our data may contribute to explain the excess of mortality in epidemiological studies and overall why exposures to B[a]P coated onto Fe2O3 resulted in higher toxicity in rodents compared to exposure to B[a]P alone.