Abstract
Sterigmatocystin (STC) and 5-methoxysterigmatocystin (5-M-STC) are structurally related mycotoxins with cytotoxic and genotoxic properties. In the present study, we hypothesized that DNA damage induced by non-cytotoxic concentrations of single and combined mycotoxins could alter the phosphorylation of the checkpoint proteins Chk2 and FANCD2 (ELISA) in HepG2 and A549 cells. The cytotoxic potential (MTT test) of single and combined STC and 5-M-STC, the nature of their interaction (additivity, antagonism, or synergy) and DNA damage level (alkaline comet assay) in HepG2 and A549 cells were also investigated. All experiments were performed after 24 h of mycotoxin treatment. 5-M-STC was 10-folds more cytotoxic than STC to both HepG2 and A549 cells. Both mycotoxins are genotoxic to HepG2 and A549 cells by inducing both double and single DNA strand breaks that activate Chk2 (especially in HepG2 cells) but not the FANCD2 protein. STC exerted higher genotoxic potential than 5-M-STC in HepG2 and A549 cells when both toxins were applied individually at the same concentration. Dual combinations of non-cytotoxic mycotoxin concentrations showed additive to antagonizing cytotoxic and genotoxic effects. The absence and low activation of checkpoint proteins during prolonged exposure to non-cytotoxic concentrations of STC and 5-M-STC could support cell proliferation and carcinogenesis.
Highlights
Streigmatocystin (STC) and 5-methoxysterigmatocystin (5-MET-STC) are closely related polyketide mycotoxins mainly produced by diverse species of Aspergillus section Nidulantes series Versicolores [1,2,3,4]
Cytotoxicity (MTT test), genotoxicity, and expression of Chk2 as well as FANCD2 phosphorylation and expression profiles were determined in human lung adenocarcinoma A549 cells and hepatocellular carcinoma HepG2 cells upon 24 h exposure to single and combined STC and 5-M-STC
According to cell viability upon 24 h treatment with mycotoxin concentrations ranging from 0.1 to 150 μM (Table 1) as well as the IC50 obtained in the MTT test, HepG2 cells were significantly more sensitive than A549 cells to both mycotoxins with 5-M-STC having about 10 times the cytotoxic potential of STC (Table 2)
Summary
Streigmatocystin (STC) and 5-methoxysterigmatocystin (5-MET-STC) are closely related polyketide mycotoxins mainly produced by diverse species of Aspergillus section Nidulantes series Versicolores [1,2,3,4]. As a precursor of aflatoxin, STC is structurally related to this carcinogenic mycotoxin Upon ingestion, both are activated by the liver cytochrome P450 system to reactive an epoxide that forms DNA adducts with guanine [5]. The serine/threonine kinase Chk is the transducer kinase involved in spreading the DNA damage signal through the phosphorylation of effector proteins involved in DNA repair, cell cycle regulation, p53 signalling, and apoptosis [19]. In BEAS-2B and A549 cells, STC induced DNA damage and cell cycle arrest related to altered expression of cyclindependent kinase-CDK1 and cyclin B1, which all may contribute to lung carcinogenesis [14]. Since STC and 5-M-STC are structurally related mycotoxins with cytotoxic and genotoxic properties in vitro, we hypothesized that DNA damage induced by sub-cytotoxic concentrations of individual and combined mycotoxins might alter Chk and FANCD2 phosphorylation and/or expression in HepG2 and A549 cells. The cytotoxic potential of single and combined STC and 5-M-STC, the type of their interaction (additivity, antagonism, or synergy) as well as DNA damage level evoked by their sub-cytotoxic concentrations in HepG2 and A549 cells were explored
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