The Study Of Cigarette Smoking Condensates (CSC) Inducing Malignant Transformation Of Immortalized Human Bronchial Epithelial Cells (BEP2D)

Objection: To detect the ability of malignant transformation of immortalized human bronchial epithelial cells (BEP2D) using cigarette smoking condensates (CSC) alone in vitro and establish a model of lung cancer caused by smoking alone. At the same time, we detected different expression genes in different stage cells treated by CSC. So this study is to provide an experimental groundwork for understanding cell and molecule mechanisms in more phases of lung cancers caused by smoking. Method: At first, we tested the slowly cytotoxicity of BEP2D cells treated by CSC. So we can select the does of slowly transformation according to the relative survive rate of BEP2D cells by MTT test. Endpoints such as growth kinetics, resistance to serum-induced terminal differentiation, anchorage-independence growth and tumorigenicity in nude mice were used to assess the transformation stage of these cells. And then, we detected difference expression genes in all groups using human genome oligonucleotide array. In addition, we also explored the oxidation damage of BEP2D cells caused by CSC utilizing the tag technique of reactive oxygen species (ROS)—DCFH-DA and HE. Results: We selected 0.001 per cigarette/ml (LHC-8) as transformat does at 60-80% relative survive rates. At the same time, we also selected same alcohol solvent content in transformat dose—0.0005ml /ml (95% alcohol/ ml LHC-8) as alcohol solvent control group. Picking up the 10^th,20^th,30^th,40^thpassage cells after CSC treated as inspect point for cell aggregates. The speed of growth increases and double time decreases appearly. Resistances to serum and anchorage independence growth were also enhanced in 30^th,40^th passage cells. The majority genes are down regulation, the minority genes are up regulation of difference expression genes. The down regulation genes in promoter domain have significant relation with the methylation of critical genes in lung cancer. This study still discovered that CSC can cause a plenty of ROS in BEP2D cells and has dose-effect, but the cells after treated with CSC have got some resisting capacity to oxidation damage. What is this? Expect later discover. Conclusion: CSC alone may result in BEP2D cells slowly malignant transformation. The tumorigenicity was depent on growthing tumor in nude mice. The mechanism of transformation is likely to relate to the cells gaining anti-oxidation damage. This research provided a good base for later deep study.