Ye Meng, Liu Bing-ci, Du Hong-ju, You Bao-rong
National Institute of Occupation Health and Poison Control, Chinese Center for Disease Control and Prevention, Beijing 100050, China.
Wei Sheng Yan Jiu. 2006 Mar;35(2):135-8.
To establish Benzo (a) pyrene-treated human embryo lung fibroblasts (HELF), which were provided with some characteristics of transformed cells (T-HELF), and observe the changes of cyclin D, CDK4 and E2F-1/4 expression in T-HELF cells.
Using 200, 100, 50, 25, 5 and 1 micromol/L B(a)P treated HELF cells for 24 h. Using MTT detected the cytotoxicity of B(a)P. 100,200 micromol/L B(a)P was chosen for the treatment of HELF cells. HELF cells was treated with B(a)P three times. The identification of T-HELF was investigated. Changes of cell cycle and the expression of cyclin D1,CDK4 and E2F-1/4 were checked using the flow cytometer and Western blot analysis.
The survival rate of HELF cells treated with 25-200 micromol/L B(a)P was 78%-80%. 4 w after B(a)P treatment, cells were all dead in 200/mol/L groups. 6 w after 100 micromol/L B(a) P treatment, the morphological changes could be observed. 12 w after treatment, there were colonies of T-HELF cells in soft agar. In T-HELF cells, cyclin D1 expression was higher than normal HELF, and cell cycle progressed through G, to S without serum. There were not significant changes of CDK4 and E2F-1/4 expression in T-HELF cells compared with normal HELF.
The T-HELF was established. This model provided a potential tool for investigating the molecular mechanism of carcinogenesis of B(a)P. The higher expression of cyclin D1 may contribute to the cell cycle changes of T-HELF cells.
建立具有转化细胞某些特征的苯并（a）芘处理的人胚肺成纤维细胞（T-HELF），并观察T-HELF细胞中细胞周期蛋白D、细胞周期蛋白依赖性激酶4（CDK4）和E2F-1/4表达的变化。
用200、100、50、25、5和1微摩尔/升苯并（a）芘处理人胚肺成纤维细胞24小时。用MTT检测苯并（a）芘的细胞毒性。选择100、200微摩尔/升苯并（a）芘处理人胚肺成纤维细胞。用苯并（a）芘处理人胚肺成纤维细胞三次。对T-HELF进行鉴定。用流式细胞仪和蛋白质免疫印迹分析检测细胞周期变化及细胞周期蛋白D1、CDK4和E2F-1/4的表达。
用25-200微摩尔/升苯并（a）芘处理的人胚肺成纤维细胞存活率为78%-80%。苯并（a）芘处理4周后，200微摩尔/升组细胞全部死亡。100微摩尔/升苯并（a）芘处理6周后，可观察到形态学变化。处理12周后，软琼脂中有T-HELF细胞集落。在T-HELF细胞中，细胞周期蛋白D1表达高于正常的人胚肺成纤维细胞，且在无血清条件下细胞周期从G1期进入S期。与正常的人胚肺成纤维细胞相比，T-HELF细胞中CDK4和E2F-1/4表达无明显变化。
成功建立了T-HELF。该模型为研究苯并（a）芘致癌的分子机制提供了潜在工具。细胞周期蛋白D1的高表达可能导致T-HELF细胞的细胞周期变化。
