Smad4基因沉默对胰腺上皮内瘤变细胞基因表达谱影响的初步研究

Preliminary Study of Smad4 Gene Silencing on Gene Expression Profile of Pancreatic Intraepithelial Neoplasia Cells

背景:抑癌基因Smad4失活可使由Kras基因突变启动的胰腺上皮内瘤变(PanIN)细胞发生恶性转化,但其具体机制尚未阐明。目的:探讨Smad4基因沉默的PanIN细胞(PanIN-S细胞)基因表达谱的变化,分析Smad4基因沉默致PanIN细胞增殖和恶性转化的可能分子机制。方法:通过RNA干扰沉默PanIN细胞的内源性Smad4基因表达以构建PanIN-S细胞,小鼠全基因表达谱芯片筛查PanIN细胞与PanIN-S细胞的基因表达谱差异,实时荧光定量RT-PCR验证基因芯片筛选出的细胞周期相关差异表达基因。结果:基因芯片共筛选出237个差异表达基因,其中148个基因在PanIN-S细胞中表达上调,89个表达下调。差异表达基因主要涉及细胞周期、增殖、凋亡、黏附、转录活性等。实时荧光定量RT-PCR验证示细胞周期相关基因p27、p19、细胞周期蛋白(cyclin)D1表达在PanIN细胞与PanIN-S细胞间存在显著差异,与基因芯片筛选结果一致。结论:PanIN-S细胞p27、p19和cyclin D1基因表达发生明显改变,可能参与了PanIN-S细胞的增殖和恶性转化。

Background： Inactivation of tumor suppressor gene Smad4 can promote the malignant transformation of pancreatic intraepithelial neoplasia （PanIN） cells initiated by Kras mutation, but its mechanism has not yet been clarified. Aims： To investigate the alteration of gene expression profile of Smad4-silenced PanIN cells （PanIN-S ceils）, and to assess the possible mechanism of Smad4 gene silencing induced proliferation and malignant transformation of PanIN cells. Methods： PanIN-S ceils were obtained by silencing the endogenous Smad4 gene in PanIN cells through RNA interference. Differential gene expression profile of PanIN cells and PanIN-S cells were detected by mouse pan-gene chip array analysis, and real- time fluorescent quantitative RT-PCR was performed to further identify the differentially expressed cell cycle-related genes detected by gene chip array amalysis. Results： Gene chip array analysis detected 237 differentially expressed genes, of which 148 were up-regulated and 89 were down-regulated in PanIN-S cells, mostly involved in cell cycle, proliferation, apoptosis, adhesion, transcription, etc. Real-time fluorescent quantitative RT-PCR demonstrated the identity of differential expression of cell cycle-related genes p27, p19 and cyclin D1 detected by gene chip array analysis in PanIN cells and PanIN-S cells. Conclusions： Gene expressions of p27, p19 and cyclin D1 in PanIN-S cells alter significantly, which may be involved in the proliferation and malignant transformation of PanIN-S ceils.