The Research Of Abnormal Promoter Methylation Of Multiple Genes In The Human Bronchial Epithelial Malignant Cells

Carcinoma of the lung is the most common cause of cancer death worldwide. Many researchers have focused on its pathogenesis and early diagnosis.For more than fifteen years, it has been recognized that the methylation patterns of tumor cells are significantly altered compared to those normal cells. Hypermethylation of certain CpG islands is seen in most tumor cells and is argued to be associated with gene silencing. DNA methylation changes can not only yield insight in to the complex molecular pathways leading to cancer, but also promise to provide a highly sensitive and specific diagnostic tool.To detect pathogenesis of radicalization inducing lung cancer and then provide evidence to clinical application, we have established a model of the transformed human bronchial epithelial cells (BEP2D) induced by -particles. We used methylation special polymerase chain reaction (MSP) to detect aberrant promoter methylation of p14ARP and p16INK4a (cell cycle regulators), O6-methylguanine-DNA methyltransferase and glutathione S-transferase P1 (DNA repair), BUB3 (mitosis spindle check-point) and death-associated protein kinase (cell apoptosis) genes in the transformed human bronchial epithelial cells (BEP2D) and its malignant transformant BERP35T1induced by -particles . At the same time, the level of p14ARP gene transcription was analyzed using RT-PCR.The CpG island in 5' promoter regions of p14ARP gene was unmethylated in BEP2D cells, but was methylated in the malignant transformant BERP35T1. To discuss further influence of CpG island methylaion, we used RT-PCR to extend the distance of exon 1 and one part of exon 2 of p14ARP and we found the level of BERP35T1 transcription was depressed remarkably. However p16INK4a gene, which shares two exons with p14ARP gene, is not methylated in CpG islands. MGMT gene was methylated in BEP2D and its malignant transformant BERP35T1. DAPK gene was methylated partly in BEP2D cells and methylated completely in BERP35T1. GSTP1 gene was unmethylated in BEP2D cells and was methylated partly in BERP35T1. BUB3 gene was unmethylated in BEP2D and BERP35T1 cells.To prove the reliability of our operation, we also surveyed the extend sequences of BUB3 gene. The result showed that all cytosines in our expending regions had changed to thymine. Therefore our method is approved credible.In conclusion, according our work, we found some tumor suppressor genes that controled cell cycle, proliferation, DNA repair and apoptosis had aberrant methylation in 5'- promoter CpG islands and therefore repressed transcription of these genes.