title: Vincristine-induced bystander effect in human lymphocytes.
abstract: Bystander effect is a known radiobiological effect, widely described using ionizing radiations and which, more recently, has also been related to chemical mutagens. In this study, we aimed to assess whether or not a bystander response can be induced in cultured human peripheral lymphocytes by vincristine, a chemotherapeutic mutagen acting as spindle poison, and by mitomycin-C, an alkylating agent already known to induce this response in human lymphoblastoid cells. Designing a modified ad hoc protocol for the cytokinesis blocked micronucleus (MN) assay, we detected the presence of a dose-dependent bystander response in untreated cultures receiving the conditioned medium (CM) from mitomycin-C (MMC) or vincristine (VCR) treated cultures. In the case of MMC, MN frequencies, expressed as micronucleated binucleates, were: 13.5±1.41 at 6μM, 22±2.12 at 12μM or 28.25±5.13 at 15μM vs. a control value of 4.75±1.59. MN levels for VCR, expressed as micronucleated mononucleates were: 2.75±0.88 at 0.0μM, 27.25±2.30 at 0.4μM, 46.25±1.94 at 0.8μM, 98.25±7.25 at 1.6μM. To verify that no mutagen residual was transferred to recipient cultures together with the CM, we evaluated MN levels in cultures receiving the medium immediately after three washings following the chemical treatment (unconditioned medium). We further confirmed these results using a cell-mixing approach where untreated lymphocytes were co-cultured with donor cells treated with an effect-inducing dose of MMC or VCR. A distinct production pattern of both reactive oxygen species and soluble mediator proteins by treated cells may account for the differences observed in the manifestation of the bystander effect induced by VCR. In fact, we observed an increased level of ROS, IL-32 and TGF-β in the CM from VCR treated cultures, not present in MMC treated cultures.
authors.affiliation: Unità di Genetica, Dipartimento di Biologia, Pisa University, Via Derna 1, 56126 Pisa, Italy.
authors.forename: Serena
authors.initials: S
authors.lastname: Testi
authors.affiliation: Unità di Genetica, Dipartimento di Biologia, Pisa University, Via Derna 1, 56126 Pisa, Italy.
authors.forename: Alessia
authors.initials: A
authors.lastname: Azzarà
authors.affiliation: Unità di Genetica, Dipartimento di Biologia, Pisa University, Via Derna 1, 56126 Pisa, Italy.
authors.forename: Caterina
authors.initials: C
authors.lastname: Giovannini
authors.affiliation: Unità di Genetica, Dipartimento di Biologia, Pisa University, Via Derna 1, 56126 Pisa, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Lombardi
authors.affiliation: Dipartimento di Ricerca Traslazionale e delle Nuove Tecnologie in Medicina e Chirurgia, Pisa University, Via Savi 10, 56126 Pisa, Italy.
authors.forename: Simona
authors.initials: S
authors.lastname: Piaggi
authors.affiliation: Unità di Genetica, Dipartimento di Biologia, Pisa University, Via Derna 1, 56126 Pisa, Italy.
authors.forename: Maria Sole
authors.initials: MS
authors.lastname: Facioni
authors.affiliation: Unità di Genetica, Dipartimento di Biologia, Pisa University, Via Derna 1, 56126 Pisa, Italy; Research Center of Nutraceuticals and Food for Health, University of Pisa, Pisa, Italy. Electronic address: roberto.scarpato@unipi.it.
authors.forename: Roberto
authors.initials: R
authors.lastname: Scarpato
mesh_terms: D000328:Adult; D024201:Bystander Effect / Q000187:drug effects* / Q000235:genetics; D002478:Cells, Cultured; D018920:Coculture Techniques; D017077:Culture Media, Conditioned; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007378:Interleukins / Q000378:metabolism; D008214:Lymphocytes / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008297:Male; D048629:Micronuclei, Chromosome-Defective / Q000139:chemically induced*; D015162:Micronucleus Tests / Q000379:methods; D008856:Microscopy, Fluorescence; D016685:Mitomycin / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000378:metabolism; D014750:Vincristine / Q000494:pharmacology*; D055815:Young Adult
chemical_list: D017077:Culture Media, Conditioned; C496971:IL32 protein, human; D007378:Interleukins; D017382:Reactive Oxygen Species; D053773:Transforming Growth Factor beta1; D016685:Mitomycin; D014750:Vincristine
keywords: Bystander effect; Cytokines; Human lymphocytes; Micronuclei; ROS; Spindle poisons