E-cigarettes induce toxicological effects that can raise the cancer risk

Abstract

Electronic cigarettes (e-cigs) are devices designed to deliver nicotine in a vaping solution rather than smoke and without tobacco combustion. Perceived as a safer alternative to conventional cigarettes, e-cigs are aggressively marketed as lifestyle-choice consumables, thanks to few restrictions and a lack of regulatory guidelines. E-cigs have also gained popularity among never-smokers and teenagers, becoming an emergent public health issue. Despite the burgeoning worldwide consumption of e-cigs, their safety remains largely unproven and it is unknown whether these devices cause in vivo toxicological effects that could contribute to cancer. Here we demonstrate the co-mutagenic and cancer-initiating effects of e-cig vapour in a rat lung model. We found that e-cigs have a powerful booster effect on phase-I carcinogen-bioactivating enzymes, including activators of polycyclic aromatic hydrocarbons (PAHs), and increase oxygen free radical production and DNA oxidation to 8-hydroxy-2'-deoxyguanosine. Furthermore, we found that e-cigs damage DNA not only at chromosomal level in peripheral blood, such as strand breaks in leucocytes and micronuclei formation in reticulocytes, but also at gene level such as point mutations in urine. Our results demonstrate that exposure to e-cigs could endanger human health, particularly among younger more vulnerable consumers.

Figure 1

Metabolic/antioxidant enzymes and free radical yield in e-cig-exposed rat lung. (a) Cytochrome P450 (CYPs) is a superfamily of major isoenzymes involved in drug metabolism. CYP activities lead to the bioactivation of ubiquitous pre-mutagens and pre-carcinogens as well as ROS generation linked to their catalytic cycle. Data were obtained through enzymatic assays performed on microsomal lung fractions using several specific probes: MROD (CYP1A2-like) increased up to 262%, PROD (2B1/2) 384%, APD (3A, 1A, 2A, 2D) 19% P < 0.05, 16-α TOH (2B1/2C11) 48% (P < 0.01), 17-TOH (3A1) 41% (P < 0.01). (b) EPR spectra of nitroxide radicals observed in rat lung tissues in control samples (green spectra), and in e-cig vapour-treated samples (red spectra). (c) EPR intensity of the first spectral line of the observed nitroxide radicals (arbitrary units). (d) Antioxidant enzymes: CAT, NQO1 and SOD were reduced more than 32% (P < 0.01). (e) Transferases shown here are involved in the detoxifying step of xenobiotic metabolism making drugs or toxins more water-soluble. They also contribute to preserving DNA from adduct formation converting carcinogens into inactive or less toxic compounds: UDP-GT unchanged, GST 28% loss (P < 0.01). Each bar represents the means ± S.D. of ten measurements performed on ten rats, ^* P < 0.05, ^** P < 0.01, two-tailed t-test.

Figure 2

Systemic antioxidant capacity, oxidative DNA damage and lipidomics. (a) FRAP lung, loss >33% (P < 0.05). (b) FRAP plasma (P = 0.059). (c) FRAP plasma from e-cig group inversely correlated vs CO (r = 0.930, P < 0.001). (d) FRAP plasma from control group positively correlated vs CO (r = 0.880, P < 0.01). (e) 8-OHdG lung levels markedly increased ~288% (P < 0.01). (f) FRAP lung from e-cig group inversely correlated vs 8-OHdG (r = 0.845, P < 0.05). Data (n = 5 measurements per group) are expressed as means ± standard error of the mean (SEM), analysed by one-way analysis of variance (ANOVA) (g) left side, content of esterified cholesterol, total cholesterol and triglycerides (mg/dL) determined by GC/MS and GC/FID for qualitative and quantitative analysis, respectively, on Control and E-cig groups. Right side, sum of C18:1 trans isomers, saturated fatty acids (SFA), polyunsaturated fatty acids (PUFA) and PUFA n-6 series (PUFA n-6) in percentage (%) of total fatty acids determined by GC/FID on Control and E-cig groups. Each bar represents the means ± S.D. of ten measurements performed on ten ^* P < 0.05, ^** P < 0.01, ^*** P < 0.001, two-tailed t-test.

Figure 3

Genotoxicity of e-cig vapour. (a) Distribution of individual median TI% for the alkaline Comet assay. (b) Box-plot of TI%: primary DNA damage increase (P < 0.001). (c) Representative image of micronuclei (yellow MN-RET, orange RET, green erythrocytes (E). (d,e) MN-RET vs RET and RET vs E plus RET; MN-RET vs RET increased (P < 0.05); hematopoietic depression up to 50% loss, as RT fraction of total red blood cells, (P < 0.001). Error bars ± S.D. ^* P < 0.05; ^*** P < 0.001 two-tailed t-test. (f,g) Urinary mutagenesis: TA100 and YG1024 S. typhimurium revertants/plate increased in a dose-dependent manner ± S9 mix. ^* P < 0.05; ^** P < 0.01; ^*** P < 0.001 Bonferroni’s test).