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Use of human peripheral blood lymphocytes to evaluate the cyto/genotoxicity profile of oxime K048

Oximes are pharmacologically important nucleophylic agents acting as antidotes against poisoning by organophosphorus compounds. In this study, effect of K048 oxime on cell viability and chromosome stability in vitro was evaluated in primary cell cultures of isolated human peripheral blood lymphocytes. K048 oxime at concentrations of 730, 200, and 7.3 nmol/dm3 was tested in 30 minutes treatment. Cytotoxic effect was tested by using differential viability staining with a mixture of acridine orange and ethydium bromide, while in evaluation of genotoxic potential hOGG1 modified comet assay, cytokinesis-blocked micronucleus assay, and chromosome aberration analysis were used. Viability staining showed that oxime treatment does not induce apoptosis (8.0% highest concentration vs. 7.0% control) or necrosis indicating lack of its cytotoxic effect (9.6% highest concentration vs. 7.3% control). HOGG1 modified comet assay revealed dose dependant increase in the level of oxidative primary damage to DNA, significant at all doses tested. Expressed as % of DNA in comet tail, oxidative damage level in K048 treated lymphocytes ranged from 7.73-11.46% compared to 2.12% in control lymphocytes. However, oxime treatment did not affect micronucleus frequency nor induced significant increase in formation of structural chromosomal aberrations. Micronucleus frequency in oxime treated lymphocytes ranged from 13-19 per 3000 binucleated cells compared to 15 micronuclei per 3000 cells in control cultures. Chromatid breaks were detected in both, control and oxime treated lymphocytes with no difference in frequency, while formation of acentric fragments was observed only in lymphocytes treated with lowest K048 concentration (7.3 nmol/dm3). However, observed level of acentric formation (2 per 200 cells) was not statistically significant. In applied battery of cyto/gentoxicity assays, considering applied testing conditions K048 oxime showed acceptable biocompatibility at the level of cell viability and chromatin/chromosome integrity. Since no increase in secondary genome damage was detected, induced primary oxidative DNA damage might be result of cell stress induced by treatment and are rather repaired than fixed as chromosome damage.

Chromosome aberrations; DNA damage; In vitro; Lymphocyte; Micronuclei; Oxime

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