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Time- and concentration-dependent changes in gene expression induced by benzo(a)pyrene in two human cell lines, MCF-7 and HepG2.

Hockley SL, Arlt VM, Brewer D, Giddings I, Phillips DH - BMC Genomics (2006)

Bottom Line: In contrast, BeP did not induce consistent gene expression changes at the same concentrations.Phenotypic anchoring of the expression data to DNA adduct levels detected by 32P-postlabelling, cell cycle data and p53 protein expression identified a number of genes that are linked to these biological outcomes, thereby strengthening the identification of target genes.The overall response to BaP consisted of up-regulation of tumour suppressor genes and down-regulation of oncogenes promoting cell cycle arrest and apoptosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Section of Molecular Carcinogenesis, Institute of Cancer Research, Brookes Lawley Building, Cotswold Road, Sutton, Surrey SM2 5NG, UK. sarah.hockley@icr.ac.uk

ABSTRACT

Background: The multi-step process of carcinogenesis can be more fully understood by characterizing gene expression changes induced in cells by carcinogens. In this study, expression microarrays were used to monitor the activity of 18,224 cDNA clones in MCF-7 and HepG2 cells exposed to the carcinogen benzo(a)pyrene (BaP) or its non-carcinogenic isomer benzo(e)pyrene (BeP). Time and concentration gene expression effects of BaP exposure have been assessed and linked to other measures of cellular stress to aid in the identification of novel genes/pathways involved in the cellular response to genotoxic carcinogens.

Results: BaP (0.25-5.0 muM; 6-48 h exposure) modulated 202 clones in MCF-7 cells and 127 in HepG2 cells, including 27 that were altered in both. In contrast, BeP did not induce consistent gene expression changes at the same concentrations. Significant time- and concentration-dependent responses to BaP were seen in both cell lines. Expression changes observed in both cell lines included genes involved in xenobiotic metabolism (e.g., CYP1B1, NQO1, MGST1, AKR1C1, AKR1C3,CPM), cell cycle regulation (e.g., CDKN1A), apoptosis/anti-apoptosis (e.g., BAX, IER3), chromatin assembly (e.g., histone genes), and oxidative stress response (e.g., TXNRD1). RTqPCR was used to validate microarray data. Phenotypic anchoring of the expression data to DNA adduct levels detected by 32P-postlabelling, cell cycle data and p53 protein expression identified a number of genes that are linked to these biological outcomes, thereby strengthening the identification of target genes. The overall response to BaP consisted of up-regulation of tumour suppressor genes and down-regulation of oncogenes promoting cell cycle arrest and apoptosis. Anti-apoptotic signalling that may increase cell survival and promote tumourigenesis was also evident.

Conclusion: This study has further characterised the gene expression response of human cells after genotoxic insult, induced after exposure to concentrations of BaP that result in minimal cytotoxicity. We have demonstrated that investigating the time and concentration effect of a carcinogen on gene expression related to other biological end-points gives greater insight into cellular responses to such compounds and strengthens the identification of target genes.

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Principal component analysis of MCF-7 cells exposed to BaP and BeP using the 165 BaP-modulated genes from the 15 K microarray analysis. Black represents BaP samples and grey, BeP samples.
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Figure 5: Principal component analysis of MCF-7 cells exposed to BaP and BeP using the 165 BaP-modulated genes from the 15 K microarray analysis. Black represents BaP samples and grey, BeP samples.

Mentions: MCF-7 and HepG2 cells were also exposed to the non-carcinogenic isomer of BaP, BeP, which does not form DNA adducts, in order to distinguish genotoxic from toxic responses induced by BaP. MCF-7 cells were exposed to up to 2.5 μM BeP for 6, 24 and 48 h and HepG2 cells to 5 μM for 6 and 24 h. Gene expression changes induced by BeP were analysed using the 15 K cDNA microarray. In both cell lines few genes were significantly altered over 1.4-fold by BeP (Additional file 4) and comparison of the BaP and BeP expression data did not identify any common significant or consistent gene changes. Figure 5 illustrates that the BaP and BeP gene expression responses are clearly different.


Time- and concentration-dependent changes in gene expression induced by benzo(a)pyrene in two human cell lines, MCF-7 and HepG2.

Hockley SL, Arlt VM, Brewer D, Giddings I, Phillips DH - BMC Genomics (2006)

Principal component analysis of MCF-7 cells exposed to BaP and BeP using the 165 BaP-modulated genes from the 15 K microarray analysis. Black represents BaP samples and grey, BeP samples.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1621085&req=5

Figure 5: Principal component analysis of MCF-7 cells exposed to BaP and BeP using the 165 BaP-modulated genes from the 15 K microarray analysis. Black represents BaP samples and grey, BeP samples.
Mentions: MCF-7 and HepG2 cells were also exposed to the non-carcinogenic isomer of BaP, BeP, which does not form DNA adducts, in order to distinguish genotoxic from toxic responses induced by BaP. MCF-7 cells were exposed to up to 2.5 μM BeP for 6, 24 and 48 h and HepG2 cells to 5 μM for 6 and 24 h. Gene expression changes induced by BeP were analysed using the 15 K cDNA microarray. In both cell lines few genes were significantly altered over 1.4-fold by BeP (Additional file 4) and comparison of the BaP and BeP expression data did not identify any common significant or consistent gene changes. Figure 5 illustrates that the BaP and BeP gene expression responses are clearly different.

Bottom Line: In contrast, BeP did not induce consistent gene expression changes at the same concentrations.Phenotypic anchoring of the expression data to DNA adduct levels detected by 32P-postlabelling, cell cycle data and p53 protein expression identified a number of genes that are linked to these biological outcomes, thereby strengthening the identification of target genes.The overall response to BaP consisted of up-regulation of tumour suppressor genes and down-regulation of oncogenes promoting cell cycle arrest and apoptosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Section of Molecular Carcinogenesis, Institute of Cancer Research, Brookes Lawley Building, Cotswold Road, Sutton, Surrey SM2 5NG, UK. sarah.hockley@icr.ac.uk

ABSTRACT

Background: The multi-step process of carcinogenesis can be more fully understood by characterizing gene expression changes induced in cells by carcinogens. In this study, expression microarrays were used to monitor the activity of 18,224 cDNA clones in MCF-7 and HepG2 cells exposed to the carcinogen benzo(a)pyrene (BaP) or its non-carcinogenic isomer benzo(e)pyrene (BeP). Time and concentration gene expression effects of BaP exposure have been assessed and linked to other measures of cellular stress to aid in the identification of novel genes/pathways involved in the cellular response to genotoxic carcinogens.

Results: BaP (0.25-5.0 muM; 6-48 h exposure) modulated 202 clones in MCF-7 cells and 127 in HepG2 cells, including 27 that were altered in both. In contrast, BeP did not induce consistent gene expression changes at the same concentrations. Significant time- and concentration-dependent responses to BaP were seen in both cell lines. Expression changes observed in both cell lines included genes involved in xenobiotic metabolism (e.g., CYP1B1, NQO1, MGST1, AKR1C1, AKR1C3,CPM), cell cycle regulation (e.g., CDKN1A), apoptosis/anti-apoptosis (e.g., BAX, IER3), chromatin assembly (e.g., histone genes), and oxidative stress response (e.g., TXNRD1). RTqPCR was used to validate microarray data. Phenotypic anchoring of the expression data to DNA adduct levels detected by 32P-postlabelling, cell cycle data and p53 protein expression identified a number of genes that are linked to these biological outcomes, thereby strengthening the identification of target genes. The overall response to BaP consisted of up-regulation of tumour suppressor genes and down-regulation of oncogenes promoting cell cycle arrest and apoptosis. Anti-apoptotic signalling that may increase cell survival and promote tumourigenesis was also evident.

Conclusion: This study has further characterised the gene expression response of human cells after genotoxic insult, induced after exposure to concentrations of BaP that result in minimal cytotoxicity. We have demonstrated that investigating the time and concentration effect of a carcinogen on gene expression related to other biological end-points gives greater insight into cellular responses to such compounds and strengthens the identification of target genes.

Show MeSH
Related in: MedlinePlus