title: Melittin induced cytogenetic damage, oxidative stress and changes in gene expression in human peripheral blood lymphocytes.
abstract: Melittin (MEL) is the main constituent and principal toxin of bee venom. It is a small basic peptide, consisting of a known amino acid sequence, with powerful haemolytic activity. Since MEL is a nonspecific cytolytic peptide that attacks lipid membranes thus leading to toxicity, the presumption is that it could have significant therapeutic benefits. The aim was to evaluate the cyto/genotoxic effects of MEL in human peripheral blood lymphocytes (HPBLs) and the molecular mechanisms involved using a multi-biomarker approach. We found that MEL was cytotoxic for HPBLs in a dose- and time-dependent manner. It also induced morphological changes in the cell membrane, granulation and lysis of exposed cells. After treating HPBLs with non-cytotoxic concentrations of MEL, we observed increased DNA damage including oxidative DNA damage as well as increased formation of micronuclei and nuclear buds, and decreased lymphocyte proliferation determined by comet and micronucleus assays. The observed genotoxicity coincided with increased formation of reactive oxygen species, reduction of glutathione level, increased lipid peroxidation and phospholipase C activity, showing the induction of oxidative stress. MEL also modulated the expression of selected genes involved in DNA damage response (TP53, CDKN1A, GADD45α, MDM), oxidative stress (CAT, SOD1, GPX1, GSR and GCLC) and apoptosis (BAX, BCL-2, CAS-3 and CAS-7). Results indicate that MEL is genotoxic to HPBLs and provide evidence that oxidative stress is involved in its DNA damaging effects. MEL toxicity towards normal cells has to be considered if used for potential therapeutic purposes.
authors.affiliation: Mutagenesis Unit, Institute for Medical Research and Occupational Health, 10000 Zagreb, Croatia. Electronic address: ggajski@imi.hr.
authors.forename: Goran
authors.initials: G
authors.lastname: Gajski
authors.affiliation: Department of Pharmaceutical Botany, Faculty of Pharmacy and Biochemistry, University of Zagreb, 10000 Zagreb, Croatia.
authors.forename: Ana-Marija
authors.initials: AM
authors.lastname: Domijan
authors.affiliation: Department for Genetic Toxicology and Cancer Biology, National Institute of Biology, 1000 Ljubljana, Slovenia.
authors.forename: Bojana
authors.initials: B
authors.lastname: Žegura
authors.affiliation: Department for Genetic Toxicology and Cancer Biology, National Institute of Biology, 1000 Ljubljana, Slovenia.
authors.forename: Alja
authors.initials: A
authors.lastname: Štern
authors.affiliation: Mutagenesis Unit, Institute for Medical Research and Occupational Health, 10000 Zagreb, Croatia.
authors.forename: Marko
authors.initials: M
authors.lastname: Gerić
authors.affiliation: Toxicology Unit, Institute for Medical Research and Occupational Health, 10000 Zagreb, Croatia.
authors.forename: Ivana
authors.initials: I
authors.lastname: Novak Jovanović
authors.affiliation: Molecular Toxicology Unit, Institute for Medical Research and Occupational Health, 10000 Zagreb, Croatia.
authors.forename: Ivana
authors.initials: I
authors.lastname: Vrhovac
authors.affiliation: Department of Molecular Biology, Division of Biology, Faculty of Science, University of Zagreb, 10000 Zagreb, Croatia.
authors.forename: Josip
authors.initials: J
authors.lastname: Madunić
authors.affiliation: Molecular Toxicology Unit, Institute for Medical Research and Occupational Health, 10000 Zagreb, Croatia.
authors.forename: Davorka
authors.initials: D
authors.lastname: Breljak
authors.affiliation: Department for Genetic Toxicology and Cancer Biology, National Institute of Biology, 1000 Ljubljana, Slovenia.
authors.forename: Metka
authors.initials: M
authors.lastname: Filipič
authors.affiliation: Mutagenesis Unit, Institute for Medical Research and Occupational Health, 10000 Zagreb, Croatia.
authors.forename: Vera
authors.initials: V
authors.lastname: Garaj-Vrhovac
mesh_terms: D000328:Adult; D017209:Apoptosis / Q000187:drug effects; D015415:Biomarkers / Q000378:metabolism; D002478:Cells, Cultured; D002869:Chromosome Aberrations / Q000139:chemically induced*; D020552:Comet Assay; D004249:DNA Damage; D005786:Gene Expression Regulation / Q000187:drug effects*; D005978:Glutathione / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D015227:Lipid Peroxidation / Q000187:drug effects; D008214:Lymphocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008555:Melitten / Q000633:toxicity*; D015162:Micronucleus Tests; D009153:Mutagens / Q000633:toxicity*; D016877:Oxidants / Q000633:toxicity*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000819:agonists / Q000378:metabolism
chemical_list: D015415:Biomarkers; D009153:Mutagens; D016877:Oxidants; D017382:Reactive Oxygen Species; D008555:Melitten; D005978:Glutathione
keywords: Cytotoxicity; Gene expression; Genotoxicity; Human lymphocytes; Melittin; Oxidative stress