Abstract

Benzo(a)pyrene [B(a)P] is a potent environmental carcinogen, which induces cell cycle changes. All-trans retinoic acid (ATRA) is a promising agent in prevention and treatment of human cancers. In the present study, we investigated the inhibition of B(a)P-induced cell cycle progression by ATRA in human embryo lung fibroblast (HELF). Our results showed that after treatment with B(a)P, the expression of cyclin D1 and E2F-1 were both increased significantly in HELF. There were almost no changes of CDK4 and E2F-4 expression by treatment with B(a)P. As expected, pretreatment with ATRA could efficiently decrease B(a)P-induced overexpression of cyclin D1 and E2F-1. In a further study, we stably transfected antisense cyclin D1 and antisense CDK4 plasmid into HELF. The inhibition of cyclin D1 expression and the inhibition of CDK4 expression significantly impaired the B(a)P-induced overexpression of E2F-1 respectively. Pretreatment with ATRA, cells expressing antisense cyclinD1 or antisense CDK4 showed a lesser decrease of B(a)P-induced overexpression of E2F-1 compared with similarly treated HELF. Furthermore, flow cytometry analysis showed that B(a)P promoted cell cycle progression from G₁ phase to S phase, while pretreatment with ATRA could inhibit B(a)P-induced cell cycle progression by an accumulation of cells in the G₁ phase. It was suggested that ATRA could block B(a)P-induced cell cycle promotion partly through the cyclin D1/E2F-1 pathway in HELF.

Keywords: All-trans retinoic acid; Benzo(a)pyrene; Cyclin D1; CDK4; E2F; Cell cycle

Abbreviations: ATRA, all-trans retinoic acid; RA, retinoic acid; B(a)P, benzo(a)pyrene; CDK4, cyclin-dependent kinase 4; pRb, retinoblastoma protein; HELF, human embryo lung fibroblast; FBS, fetal bovine serum; DTT, 1,4-dithiothreitol; SDS, sodium dodecyl sulfate; DMSO, dimethyl sulfoxide

Article Outline

1. Introduction

2. Materials and methods

2.1. Chemicals

2.2. Cell lines and treatment with chemicals

2.3. Generation of stable transfectants

2.4. Western blot analysis

2.5. Cell cycle analysis

2.6. Statistical analysis

3. Results

3.1. B(a)P increased levels of cyclin D1 and E2F-1

3.2. Pretreatment with ATRA blocked B(a)P-induced overexpression of cyclin D1 and E2F-1 in HELF

3.3. Treatment with antisense of cyclin D1 and antisense CDK4

3.4. Pretreatment with ATRA blocked B(a)P-induced cell cycle progression

4. Discussion

Acknowledgements

References