利用cDNA微阵列-CGH筛选支气管上皮细胞恶性转化中的扩增基因

目的:基因组不稳定是导致肺癌发生与发展的重要分子机理之一。本研究旨在筛选支气管上皮细胞恶性转化过程中拷贝数扩增的基因。方法:利用业已建立的支气管上皮细胞体外恶性转化模型,通过cDNA微阵列-CGH技术对支气管上皮来源的永生化细胞和癌变细胞的基因拷贝数进行了检测,并对部分结果进行了实时PCR验证。结果:永生化BEP2D细胞染色体中的某些区域存在不同程度的扩增,包括5q31.3、9q32-33.1、14q22.2-23.1、19p13.12-13.13、20q13.12-13.31;恶性转化BERP35T2细胞染色体中的扩增区域集中在1p12-13.1、5q33.1、5q31.3、9q32、19p13.12-13.13;5q31.3、9q32、19q13.12-13.13是以上2种细胞系中的共同扩增区域。共检测到201个基因的拷贝数发生扩增,其中PCNA、TP53及GADD45A基因的异常扩增已经实时PCR进一步验证。结论:在支气管上皮细胞恶性转化过程中,病毒与低剂量辐射的双重作用使得某些重要基因的拷贝数发生扩增,因基因剂量增加而导致某些癌基因高表达可能是细胞恶性转化的重要机制之一。

Identification of Amplified Genes in the Malignant Transformation of Bronchial Epithelial Cells cDNA Microarray-CGH

Objective: Genomic instability is one of the most important molecular mechanisms,which contribute to the development of lung cancer.To screen amplified genes in the malignant transformation of bronchial epithelial cells.Methods: By cDNA microarray-CGH,the alterations of gene copy in the immortalized cell line and cancer cell line that derived from bronchial epithelial cells was detected.Some results were conformed by real-time PCR.Results: The amplified chromosome regions found in the immortalized cell line included 5q31.3,9q32-33.1,14q22.2-23.1,19p13.12-13.13 and 20q13.12-13.31;the amplified regions in cancer cell line were found in 1p12-13.1,5q33.1,5q31.3,9q32 and 19p13.12-13.13.The amplified regions in the both two cells were 5q31.3,9q32 and 19q13.12-13.13.201 genes were amplified in total.The gene copies of PCNA,TP53 and GADD45A were conformed further by real-time PCR.Conclusion: Amplification of some important genes can be induced by HPV-18 and radiation in the malignant transformation process of bronchial epithelial cells.The increased gene dosage would result in the overexpression of some oncogenes,which may be one of the important molecular events involved in the malignant transformation process of bronchial epithelial cells.