Zhou Hongning, Calaf Gloria M, Hei Tom K
Center for Radiological Research, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA.
Int J Cancer. 2003 Oct 10;106(6):821-6. doi: 10.1002/ijc.11319.
Sufficient evidence has demonstrated that cigarette smoking is causally associated with various types of human cancers. In the United States, about 90% of deaths from lung cancer among men and 79% of those among women are associated with smoking. Tobacco-specific nitrosamines are formed from nicotine and related tobacco alkaloids and are the most carcinogenic compounds of tobacco smoke. The most potent N-nitrosamine contained in tobacco smoke is 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, NNK. In the our study, the oncogenic transforming effects of graded doses of NNK were examined using papillomavirus-immortalized human bronchial epithelial cells. Growth kinetics, saturation density, resistance to serum-induced terminal differentiation, anchorage independent growth and tumorigenicity in nude mice were used to investigate the various stages of transformation in bronchial epithelial cells. We show here that immortalized human bronchial epithelial cells in culture can be malignantly transformed by treatment with NNK (100 microg/ml or 400 microg/ml) for 7 days. Transformed cells produced progressively growing subcutaneous tumors upon inoculation into nude mice. Immunofluorescence staining for keratin expression confirmed the epithelial nature of the tumor cells. Increased expression of p16, beta-catenin and PCNA in the established cell lines were detected by immunofluorescence staining and quantified by confocal microscopy. These data suggested that NNK can induce malignant transformation of human bronchial epithelial cells, and the tumor cell lines established are useful models in investigating the carcinogenic mechanism(s) of NNK.
充分的证据表明，吸烟与多种人类癌症存在因果关联。在美国，男性肺癌死亡病例中约90%、女性肺癌死亡病例中约79%与吸烟有关。烟草特异性亚硝胺由尼古丁和相关烟草生物碱形成，是烟草烟雾中最具致癌性的化合物。烟草烟雾中所含最具活性的N-亚硝胺是4-(甲基亚硝氨基)-1-(3-吡啶基)-1-丁酮，即NNK。在我们的研究中，使用乳头瘤病毒永生化的人支气管上皮细胞检测了不同剂量NNK的致癌转化作用。利用生长动力学、饱和密度、对血清诱导的终末分化的抗性、非贴壁依赖性生长以及在裸鼠中的致瘤性来研究支气管上皮细胞转化的各个阶段。我们在此表明，培养的永生化人支气管上皮细胞经NNK（100微克/毫升或400微克/毫升）处理7天可发生恶性转化。将转化细胞接种到裸鼠中后可产生逐渐生长的皮下肿瘤。对角蛋白表达的免疫荧光染色证实了肿瘤细胞的上皮性质。通过免疫荧光染色检测并通过共聚焦显微镜定量已建立细胞系中p16、β-连环蛋白和增殖细胞核抗原（PCNA）表达的增加。这些数据表明，NNK可诱导人支气管上皮细胞发生恶性转化，所建立的肿瘤细胞系是研究NNK致癌机制的有用模型。
