A number of compounds found in particulate matter with an aerodynamic diameter <2.5 (PM2.5) can interact with DNA either directly or after enzymatic transformation to induce DNA modifications. These particulate matter (PM)- induced alterations in DNA may be associated with increased frequencies of pollution-associated diseases, such as lung cancer. In the present study, we applied different methods to assess the mutagenicity and genotoxicity of monthly PM2.5 organic extracts collected over a full year. We used the Salmonella assay, exposed cultured human embryonic lung fibroblasts and applied extracellular lactate dehydrogenase (LDH) and 2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium- 5-carboxyanilide inner salt (XTT) assays to assess the cytotoxicity of PM2.5 on the cells. We assessed both the expression levels of a number of DNA repair genes (using qRT-qPCR) and the genetic profile of the treated cells compared to the control. The expression levels of XRCC1 and APE1, which are involved in the first steps of base excision repair, as well as ERCC1, XPA and XPF, which encode nucleotide excision repair subunits, were analysed. The monthly mean of the PM2.5 collected was 35.16±22.06 μg/ m3. The mutagenicity of PM2.5 to TA98 was 46±50 net revertants/m3, while the mutagenicity to TA98+S9 was 17± 19 net revertants/m3. The mean IC50 values were 2.741± 1.414 and 3.219±2.764 m3 of equivalent air in the XTT and LDH assays, respectively. A marked and significant increase in APE1 expression levels was observed in the exposed cells. This effect was also significantly correlated with mutagenicity (p<0.01). No induced AFLP fragment profile alterations were detected. The proposed approach seems to be useful for integrated evaluation and for highlighting the mechanisms inducing DNA damage.

Evaluating the genotoxicity of urban PM2.5 using PCR-based methods in human lung cells and the Salmonella TA98 reverse test

TRAVERSI, Deborah;CERVELLA, Piero;GILLI, Giorgio
2015

Abstract

A number of compounds found in particulate matter with an aerodynamic diameter <2.5 (PM2.5) can interact with DNA either directly or after enzymatic transformation to induce DNA modifications. These particulate matter (PM)- induced alterations in DNA may be associated with increased frequencies of pollution-associated diseases, such as lung cancer. In the present study, we applied different methods to assess the mutagenicity and genotoxicity of monthly PM2.5 organic extracts collected over a full year. We used the Salmonella assay, exposed cultured human embryonic lung fibroblasts and applied extracellular lactate dehydrogenase (LDH) and 2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium- 5-carboxyanilide inner salt (XTT) assays to assess the cytotoxicity of PM2.5 on the cells. We assessed both the expression levels of a number of DNA repair genes (using qRT-qPCR) and the genetic profile of the treated cells compared to the control. The expression levels of XRCC1 and APE1, which are involved in the first steps of base excision repair, as well as ERCC1, XPA and XPF, which encode nucleotide excision repair subunits, were analysed. The monthly mean of the PM2.5 collected was 35.16±22.06 μg/ m3. The mutagenicity of PM2.5 to TA98 was 46±50 net revertants/m3, while the mutagenicity to TA98+S9 was 17± 19 net revertants/m3. The mean IC50 values were 2.741± 1.414 and 3.219±2.764 m3 of equivalent air in the XTT and LDH assays, respectively. A marked and significant increase in APE1 expression levels was observed in the exposed cells. This effect was also significantly correlated with mutagenicity (p<0.01). No induced AFLP fragment profile alterations were detected. The proposed approach seems to be useful for integrated evaluation and for highlighting the mechanisms inducing DNA damage.
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Particulate matter; DNA Damage; mutation analysis; Gene expression; AFLP; Air pollution
Deborah Traversi; Piero Cervella; Giorgio Gilli
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2318/150158
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