Abstract

In vitro (geno)toxicity assessment of electronic vapour products (EVPs), relative to conventional cigarette, currently uses assays, including the micronucleus and Ames tests. Whilst informative on induction of a finite endpoint and relative risk posed by test articles, such assays could benefit from mechanistic supplementation. The ToxTracker and Aneugen Clastogen Evaluation analysis can indicate the activation of reporters associated with (geno)toxicity, including DNA damage, oxidative stress, the p53-related stress response and protein damage. Here, we tested for the different effects of a selection of neat e-liquids, EVP aerosols and Kentucky reference 1R6F cigarette smoke samples in the ToxTracker assay. The assay was initially validated to assess whether a mixture of e-liquid base components, propylene glycol (PG) and vegetable glycerine (VG) had interfering effects within the system. This was achieved by spiking three positive controls into the system with neat PG/VG or phosphate-buffered saline bubbled (bPBS) PG/VG aerosol (nicotine and flavour free). PG/VG did not greatly affect responses induced by the compounds. Next, when compared to cigarette smoke samples, neat e-liquids and bPBS aerosols (tobacco flavour; 1.6% freebase nicotine, 1.6% nicotine salt or 0% nicotine) exhibited reduced and less complex responses. Tested up to a 10% concentration, EVP aerosol bPBS did not induce any ToxTracker reporters. Neat e-liquids, tested up to 1%, induced oxidative stress reporters, thought to be due to their effects on osmolarity in vitro. E-liquid nicotine content did not affect responses induced. Additionally, spiking nicotine alone only induced an oxidative stress response at a supraphysiological level. In conclusion, the ToxTracker assay is a quick, informative screen for genotoxic potential and mechanisms of a variety of (compositionally complex) samples, derived from cigarettes and EVPs. This assay has the potential for future application in the assessment battery for next-generation (smoking alternative) products, including EVPs.

中文翻译：

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体外 ToxTracker 和 Aneugen Clastogen 评估扩展测定作为评估电子液体及其气溶胶相对遗传毒性潜力的工具

 抽象的

相对于传统卷烟，电子蒸气产品 (EVP) 的体外（基因）毒性评估目前使用的分析方法包括微核和艾姆斯测试。虽然提供了关于有限终点的诱导和测试物品造成的相对风险的信息，但此类测定可以从机械补充中受益。 ToxTracker 和 Aneugen Clastogen 评估分析可以表明与（基因）毒性相关的报告基因的激活，包括 DNA 损伤、氧化应激、p53 相关应激反应和蛋白质损伤。在这里，我们在 ToxTracker 检测中测试了精选的纯电子烟油、EVP 气雾剂和肯塔基州参考 1R6F 香烟烟雾样本的不同影响。该测定最初经过验证，以评估电子烟液基础成分、丙二醇 (PG) 和植物甘油 (VG) 的混合物是否在系统内产生干扰作用。这是通过将三个阳性对照加入纯 PG/VG 或磷酸盐缓冲盐水泡 (bPBS) PG/VG 气雾剂（不含尼古丁和香料）的系统中来实现的。 PG/VG 对化合物诱导的反应没有太大影响。接下来，与香烟烟雾样品相比，纯电子液体和 bPBS 气雾剂（烟草味；1.6% 游离碱尼古丁、1.6% 尼古丁盐或 0% 尼古丁）表现出减少且不太复杂的反应。经过测试，浓度高达 10%，EVP 气溶胶 bPBS 不会诱导任何 ToxTracker 报告基因。测试浓度高达 1% 的纯电子烟液会诱导氧化应激报告基因，这被认为是由于它们对体外渗透压的影响。电子烟液尼古丁含量不影响诱发的反应。此外，单独添加尼古丁仅诱导超生理水平的氧化应激反应。 总之，ToxTracker 检测是一种快速、信息丰富的筛查方法，可用于筛查来自香烟和 EVP 的各种（成分复杂）样品的遗传毒性潜力和机制。该测定法未来有可能应用于下一代（吸烟替代品）产品（包括 EVP）的评估电池。