Hattori Naoko, Nakagawa Takeya, Yoneda Mitsuhiro, Nakagawa Kaori, Hayashida Hiromi, Ito Takashi
Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan.
Department of Biochemistry, Nagasaki University School of Medicine, Nagasaki, Japan.
Biochem Biophys Rep. 2020 Nov 21;24:100865. doi: 10.1016/j.bbrep.2020.100865. eCollection 2020 Dec.
Heat-Not-Burn (HNB) products, generating vapor without combusting tobacco leaves, have been developed with the expectation that the number and quantity of chemicals in the vapor of these products would be reduced compared with the smoke from conventional combustible cigarettes. However, whether the lower chemical levels correlate with lower toxicity remains to be determined. Here we examined differences in the biological effects of conventional cigarette smoke (CS) and two HNB products, Ploom TECH and Ploom TECH+, using the cultured cancer cell line A549 and the normal bronchial epithelium cell line BEAS-2B. The conventional CS 3R4F extract (0.5%) markedly decreased cell proliferation of both A549 and BEAS-2B cells; however, 0.5% extracts of these commercially available HNB products did not affect cell growth. To determine the cause of decreased cell proliferation, a TUNEL assay was performed, and the results indicated that apoptosis had occurred in both A549 and BEAS-2B cells at 24 h after exposure to 3R4F. To further explore the effect of CS on epigenetics, we performed western blotting to detect histone H2A phosphorylation, which is known to affect transcriptional regulation. Only the 3R4F extract decreased histone H2A phosphorylation in both A549 and BEAS-2B cells. Next, we examined alterations in gene expression after treatment of A549 cells with Ploom TECH, Ploom TECH+, or 3R4F extracts. It was found that 339, 107, and 103 genes were upregulated more than 2 fold in A549 cells treated with 3R4F, Ploom TECH, or Ploom TECH + extracts, respectively. Among the 339 genes that were upregulated in response to 3R4F, we focused on , , and , since they were upregulated more than 100 fold, which was confirmed using RT-qPCR. These results suggest that CS, but not HNB products, cause epigenetic disruption and cell apoptosis, possibly by elevating transcription of genes such as .
加热不燃烧（HNB）产品通过不燃烧烟叶产生蒸汽，其研发目的是期望这些产品蒸汽中的化学物质数量和含量相比传统可燃香烟的烟雾会减少。然而，较低的化学物质水平是否与较低的毒性相关仍有待确定。在此，我们使用培养的癌细胞系A549和正常支气管上皮细胞系BEAS-2B，研究了传统香烟烟雾（CS）与两种HNB产品Ploom TECH和Ploom TECH+的生物效应差异。传统CS 3R4F提取物（0.5%）显著降低了A549和BEAS-2B细胞的增殖；然而，这些市售HNB产品的0.5%提取物并未影响细胞生长。为确定细胞增殖减少的原因，进行了TUNEL检测，结果表明在暴露于3R4F后24小时，A549和BEAS-2B细胞均发生了凋亡。为进一步探究CS对表观遗传学的影响，我们进行了蛋白质印迹法以检测组蛋白H2A磷酸化，已知其会影响转录调控。只有3R4F提取物降低了A549和BEAS-2B细胞中的组蛋白H2A磷酸化。接下来，我们在用Ploom TECH、Ploom TECH+或3R4F提取物处理A549细胞后检查了基因表达的变化。结果发现，在用3R4F、Ploom TECH或Ploom TECH +提取物处理的A549细胞中，分别有339、107和103个基因上调超过2倍。在响应3R4F而上调的339个基因中，我们重点关注了[具体基因名称未给出]，因为它们上调超过100倍，这通过逆转录定量聚合酶链反应（RT-qPCR）得到了证实。这些结果表明，CS而非HNB产品会导致表观遗传破坏和细胞凋亡，可能是通过提高诸如[具体基因名称未给出]等基因的转录来实现的。
