Enhanced DNA adduct formation by benzo[a]pyrene in human liver cells lacking cytochrome P450 oxidoreductase

Diet is a major source of human exposure to polycyclic aromatic hydrocarbons (PAHs),
of which benzo[a]pyrene (BaP) is the most commonly studied and measured. BaP has
been considered to exert its genotoxic effects after metabolic activation by cytochrome
P450 (CYP) enzymes whose activity can be modulated by cytochrome P450
oxidoreductase (POR), the electron donor to CYP enzymes. Previous studies showed
that BaP-DNA adduct formation was greater in the livers of Hepatic Reductase Null
(HRN) mice, in which POR is deleted specifically in hepatocytes, than in wild-type (WT)
mice. In the present study we used human hepatoma HepG2 cells carrying a knockout
(KO) in the POR gene as a human in vitro model that can mimic the HRN mouse
model. Treatment to BaP for up to 48 hours caused similar cytotoxicity in POR KO and
WT HepG2 cells. However, levels of BaP activation (i.e. BaP-7,8-dihydrodiol formation)
were higher in POR KO HepG2 cells than in WT HepG2 cells after 48 hours. This also
resulted in substantially higher BaP-DNA adduct formation in POR KO HepG2 cells
indicating that BaP metabolism is delayed in POR KO HepG2 cells thereby prolonging
the effective exposure of cells to unmetabolized BaP. As was seen in the HRN mouse
model, these results suggest that cytochrome b5, another component of the mixedfunction oxidase system, which can also serve as electron donor to CYP enzymes
along with NADH:cytochrome b5 redutase, contributes to the bioactivation of BaP in
POR KO HepG2 cells. Collectively, these findings indicate that CYPs play a more
important role in BaP detoxication as opposed to activation.