Objectives Evaluate the cytotoxicity and genotoxicity of short- and long-term e-cigarette vapor exposure on a panel of normal epithelial and head and neck squamous cell carcinoma (HNSCC) cell lines. Materials and Methods HaCaT, UMSCC10B, and HN30 were treated with nicotine-containing and nicotine-free vapor extract from two popular e-cigarette brands for periods ranging from 48 hours to 8 weeks. Cytotoxicity was assessed using Annexin V flow cytometric analysis, trypan blue exclusion, and clonogenic assays. Genotoxicity in the form of DNA strand breaks was quantified using the neutral comet assay and γ-H2AX immunostaining. Results E-cigarette-exposed cells showed significantly reduced cell viability and clonogenic survival, along with increased rates of apoptosis and necrosis, regardless of e-cigarette vapor nicotine content. They also exhibited significantly increased comet tail length and accumulation of γ-H2AX foci, demonstrating increased DNA strand breaks. Conclusion E-cigarette vapor, both with and without nicotine, is cytotoxic to epithelial cell lines and is a DNA strand break-inducing agent. Further assessment of the potential carcinogenic effects of e-cigarette vapor is urgently needed.

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We have measured the yield of single-strand breaks (SSBs), induced by 137Cs gamma radiation as assayed by agarose gel electrophoresis, for three plasmids and SV40 DNA irradiated in aerobic aqueous solution. DNA SSBs are caused mainly by the hydroxyl radical under these conditions. To characterize the reactivity of DNA with the hydroxyl radical, we investigated the variation of the G value for SSBs [G(SSB)] with the concentration of added hydroxyl radical scavengers. We find that simple competition kinetics does not describe our results, but that a nonhomogeneous kinetics model is in good agreement. At a DNA concentration of 50 micrograms cm-3, G(SSB) for the direct effect is about 1 x 10(-5) mumol J-1 for the DNA substrates studied. This is equivalent to 2 x 10(-10) SSB Gy-1 Da-1. Estimates of the efficiency of SSB induction per OH. radical interaction with DNA (0.32-0.44) reveal that all plasmids are essentially equal in reactivity.

The ability of the potential chemopreventive agent S-2-(3-aminopropylamino)ethylphosphorothioic acid (WR-2721) to protect against radiation-induced mutagenesis at the hprt locus and cell killing was studied using CHO-AA8 cells incubated for 30 min at 37 degrees C in growth medium containing its active thiol 2-[(aminopropyl)amino]ethane-thiol (WR-1065). In parallel experiments, the thiol and disulfide forms of the drug present in cells and incubation medium were determined in order to identify which, if either, of the components were associated with the observed protective effects. Treatment with 4 mM WR-1065 produced significant intracellular levels of the thiol (WRSH) and disulfide (WRSS) forms of the drug, but also caused dramatic elevation of cellular glutathione (GSH) and cysteine levels, accompanied by marked protection against 60Co gamma-photon- and neutron-induced cell killing and mutagenesis. When drug-treated cells were transferred to drug-free medium and incubated for 4 h at 37 degrees C, levels of WRSH and WRSS and protection against cell killing decreased markedly, whereas levels of GSH and cysteine and protection against mutagenesis showed little change. GSH and cysteine levels were not associated with protection against radiation-induced mutagenesis, as established by experiments performed with buthionine sulfoximine to block GSH synthesis. These data do not support the hypothesis that modulation of GSH or cysteine levels by WR-1065 is a major mechanism accounting for protection. Protection against mutagenesis was seen for cells incubated in medium with concentrations of added WR-1065 as low as 10 microM, where cellular levels of WRSH and WRSS became difficult to measure (< or = 5 microM) and no protection against cell killing was found. An unexpected observation was that cells incubated in 40 microM WR-1065 incorporated the drug much more rapidly than expected for uptake by passive diffusion and concentrated the drug to a marked degree; this indicates that a cell-mediated transport system is involved in the uptake of WR-1065 at low drug concentrations.

Guanyl radicals, the product of the removal of a single electron from guanine, are produced in DNA by the direct effect of ionizing radiation. We have produced guanyl radicals in DNA by using the single electron oxidizing agent (SCN)2-, itself derived from the indirect effect of ionizing radiation via thiocyanate scavenging of OH. We have examined the reactivity of guanyl radicals in plasmid DNA with the six most easily oxidized amino acids cysteine, cystine, histidine, methionine, tryptophan and tyrosine and also simple ester and amide derivatives of them. Cystine and histidine derivatives are unreactive. Cysteine, methionine, tyrosine and particularly tryptophan derivatives react to repair guanyl radicals in plasmid DNA with rate constants in the region of approximately 10(5), 10(5), 10(6) and 10(7) dm3 mol(-1) s(-1), respectively. The implication is that amino acid residues in DNA binding proteins such as histones might be able to repair by an electron transfer reaction the DNA damage produced by the direct effect of ionizing radiation or by other oxidative insults.