INAIL-Italian Workers' Compensation Authority, Research Area - Department of Occupational Medicine, Monteporzio Catone, Rome, Italy.
Mutat Res. 2013 Jan 20;750(1-2):1-11. doi: 10.1016/j.mrgentox.2012.06.013. Epub 2012 Sep 23.
Cigarette smoke is a complex mixture of chemicals, some of which are known as carcinogens. The cyto-genotoxic effects of cigarette-smoke extract (CSE) from commercial cigarettes without (A and B) and with filter (C and D) were evaluated at different CSE concentrations on A549 and BEAS-2B cells. The particle content of the cigarette smoke and the metal composition of the CSE were also analyzed. The cells were exposed to 1-10% of the CSE from one cigarette per experiment. Cytotoxicity was evaluated by use of the MTT assay after 24h, and the lactate dehydrogenase (LDH) assay after 30min and 24h. The Fpg-modified comet assay was used to evaluate direct-oxidative DNA damage on cells exposed for 30min. As expected, unfiltered cigarette smoke (particularly from the B cigarette) contained a higher number of particles than filtered smoke. With smoke extract from the B cigarette we found a decrease in cell viability only in BEAS-2B cells. The results of the LDH test showed membrane damage for B-cigarette smoke extract, particularly in BEAS-2B cells. Extracts from unfiltered cigarette smoke induced significant direct DNA damage, to a larger extent in A549 cells. Filtered cigarette-smoke extract induced a significant direct DNA damage at 5-10%. A significant induction of oxidative DNA damage was found at the highest CSE concentration in both cell types (by smoke extracts from B and C cigarettes in A549 cells, and from A and D cigarettes in BEAS-2B cells). Smoke extracts from filter cigarettes induced less direct DNA damage than those from unfiltered cigarettes in A549 cells, probably due to a protective effect of filter. In BEAS-2B cells the smoke extract from the B-cigarette showed the highest genotoxic effect, with a concentration-dependent trend. These findings show a higher cyto-genotoxicity for smoke extracts from the B-cigarette and oxidative effects for those from the A and D cigarettes, particularly in BEAS-2B cells. Moreover, there was a higher responsiveness of A549 cells to genotoxic insult of CSE, and a cigarette-dependent genotoxicity in BEAS-2B cells. Our experimental model demonstrated to be suitable to sensitively detect early genotoxic response of different lung-cell types to non-cytotoxic concentrations of complex inhalable mixtures.
香烟烟雾是一种复杂的化学物质混合物，其中一些被称为致癌物质。评估了来自商业香烟的无过滤（A 和 B）和有过滤（C 和 D）香烟烟雾提取物（CSE）在不同 CSE 浓度下对 A549 和 BEAS-2B 细胞的细胞遗传毒性效应。还分析了香烟烟雾的颗粒含量和 CSE 的金属成分。将细胞暴露于每个实验 1-10%的一支香烟的 CSE。24 小时后用 MTT 法评估细胞毒性，30 分钟和 24 小时后用乳酸脱氢酶（LDH）法评估。使用 Fpg 修饰的彗星试验评估暴露 30 分钟后细胞的直接氧化 DNA 损伤。正如预期的那样，无过滤香烟（特别是 B 香烟）的颗粒数高于过滤香烟。使用 B 香烟的烟雾提取物，我们发现只有 BEAS-2B 细胞的细胞活力下降。LDH 试验结果表明 B 香烟烟雾提取物对细胞膜造成损害，尤其是在 BEAS-2B 细胞中。未过滤的香烟烟雾提取物在 A549 细胞中引起显著的直接 DNA 损伤，程度更大。过滤后的香烟烟雾提取物在 5-10%的浓度下诱导显著的直接 DNA 损伤。在两种细胞类型中（A549 细胞中来自 B 和 C 香烟的烟雾提取物，以及 BEAS-2B 细胞中来自 A 和 D 香烟的烟雾提取物），在最高 CSE 浓度下发现了显著的氧化 DNA 损伤诱导。与未过滤的香烟相比，A549 细胞中的过滤香烟烟雾提取物引起的直接 DNA 损伤更少，这可能是由于过滤的保护作用。在 BEAS-2B 细胞中，B 香烟的烟雾提取物显示出最高的遗传毒性作用，呈浓度依赖性趋势。这些发现表明，B 香烟的烟雾提取物具有更高的细胞遗传毒性，A 和 D 香烟的烟雾提取物具有氧化作用，尤其是在 BEAS-2B 细胞中。此外，A549 细胞对 CSE 的遗传毒性更敏感，而 BEAS-2B 细胞对香烟的遗传毒性具有依赖性。我们的实验模型被证明能够灵敏地检测不同肺细胞类型对非细胞毒性浓度复杂可吸入混合物的早期遗传毒性反应。
