Analysis of benzo(a)pyrene-induced gene expression changes in Fisher 344 rat liver by Affymetrix microarray and quantitative real-time PCR

Benzo(a)pyrene (BaP), an environmental contaminant produced of combustion, is associated with a variety of adverse health effects, including cancer. Gene expression changes associated with early molecular level events occurring in response to BaP dietary exposures were measured using the Affymetrix microarray GeneChips in order to elucidate the underlying mechanisms of health effects prior to cell transformation, and to determine biomarkers of exposure to the compound. Microarray data revealed the differential expression (≥3-fold) of a large number of genes in response to BaP exposure after both 2 and 12-weeks exposures. A variety of clustering techniques such as hierarchical clustering, self organizing maps, and principal component analysis were used to explore the information contained in the gene expression data and to reveal similarities in the temporal and dose-related expression patterns of groups of genes. Based on the microarray data, Real-Time Quantitative Taqman PCR was used to validate the expression of six target genes of interest. The candidate genes selected (Cytochrome p450 1a1, Cyp1a1; Cytochrome p450 1a2, Cyp1a2; Glutathione-s-transferase, Gst-yc2; Tumor suppressor protein 53, p53; B-cell lymphoma, Bcl-2 ; Interleukin-1 beta, IL-1b) were based on their association with known health effects or their involvement in disease-related pathways. In addition, potential modes of action of BaP-induced toxicity or carcinogenicity were deduced through functional analysis of the genes differentially expressed and correlated with other known mechanisms of action of BaP on these cellular processes. Also, functional analysis revealed differences in temporal expression of genes associated with specific pathways, suggesting distinct temporal phases in response to short versus long-term exposure.