Recent studies in our laboratory indicated that
arsenic trioxide has the ability to cause significant cytotoxicity, and induction of a significant number of stress genes in human liver
carcinoma cells, HepG2. However, similar investigations with
atrazine did not show any significant effects of this chemical on HepG2 cells, even at its maximum solubility of 100 microg/mL in 1%
dimethyl sulfoxide (
DMSO). Further cytogenetic studies were therefore carried out to investigate the combined effects of
arsenic trioxide and
atrazine on cell viability and gene expression in immortalized human hepatocytes. Cytotoxicity was evaluated using the MTT-assay for cell viability, while the CAT-Tox (L) assay was performed to measure the induction of stress genes in thirteen different recombinant cell lines generated from human liver
carcinoma cells (HepG2), by creating stable transfectants of different mammalian promoter-
chloramphenicol acetyltransferase (CAT) gene fusions. Cytotoxicity experiments yielded LC50 values of 11.9 +/- 2.6 microg/mL for
arsenic trioxide in de-ionized water, and 3.6 +/- 0.4 microg/mL for
arsenic trioxide in 100 microg/mL
atrazine; indicating a 3 fold increase in
arsenic toxicity associated with the
atrazine exposure. Co-exposure of HepG2 cells to
atrazine also resulted in a significant increase in the potency of
arsenic trioxide to upregulate a number of stress genes including those of the
glutathione-S-transferase Ya subunit--GST Ya, metallothioneinIIa--HMTIIA, 70-kDa heat shock protein--HSP70, c-fos, 153-kDa growth arrest and DNA damage (GADD153), 45-kDa growth arrest and DNA damage (GADD45), and 78-kDa
glucose regulated protein--GRP78 promoters, as well as the
xenobiotic response element--XRE,
tumor suppressor protein response element--p53RE, cyclic
adenosine monophosphate response element--CRE, and
retinoic acid response element--RARE. No significant changes were observed with respect to the influence of
atrazine on the modulation of
cytochrome P450 1A1-CYP 1A1, and
nuclear factor kappa (B site) response element--NFkappaBRE by
arsenic trioxide. These results indicate that co-exposure to
atrazine strongly potentiates
arsenic trioxide-induced cytotoxicity and transcriptional activation of stress genes in transformed human hepatocytes.