engleski

The role of oxidative stress in bee venom induced DNA damage

Bee venom (BV) is a complex mixture of substances which posses broad spectrum of biological activities. Several studies reported different mechanisms of action where BV exhibited toxic effects by damaging cell membrane structures, DNA molecule or caused cell death. It is already known that the exposure to various toxic substances can increase production of reactive oxygen species (ROS) with the consequent disturbance of the oxidative balance in the cell, called oxidative stress. The oxidative stress causes the damage to membrane lipids which manifests as the increase of the malondialdehyde (MDA) concentration. The main antioxidant in the organism is glutathione (GSH) that plays central role in defence against cell insults and the decrease of GSH concentration depends on the exposure to oxidative stress. The aim of the present study was to examine DNA damage in human peripheral blood lymphocytes (HPBL) and its relation to oxidative stress after treatment with BV in different concentrations and lengths of time. For that purpose, we combined alkaline comet assay, a simple and sensitive technique for detection of basal DNA damage, with a restriction enzyme formamidopyrimidine-DNA glycosylase (FPG), which recognizes oxidized purines (products of DNA oxidation) and some alkylated DNA products. In addition, we assessed the levels of GSH and MDA as sensitive biomarkers of oxidative stress after BV treatment. Moreover, N-acetyl-L-cysteine (NAC) a source of cysteine for the synthesis of the endogenous antioxidant GSH was used as additional biomarker for evaluation of possible involvement of oxidative stress in bee venom mediated genotoxicity. Our results demonstrated that the intracellular GSH level decreased and MDA level increased after BV exposure in a dose and time-dependent manner. Subsequently, alkaline comet assay parameters revealed that bee venom induced DNA damage in HPBL. Digestion of DNA from BV-treated HPBL with purified FPG, which recognizes specific oxidized purines, displayed a greater extent of DNA strand breaks than non-digested DNA, providing evidence that BV induced oxidation of purines. Moreover, pretreatment with the antioxidant NAC prior to BV incubation significantly protected HPBL against BV-induced DNA damage. Those results provide evidence that the genotoxicity of BV is at least partly mediated by oxidative stress.

Bee venom; Human leukocytes; DNA damage; Oxidative stress; Glutathione; Malondialdehyde; N-acetyl-L-cysteine

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