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背景与目的:细胞逃避转化生长因子-β(TGF-β)诱导的对细胞生长、增殖的抑制是许多肿瘤发生的一个重要机制。Smad7是TGF-β信号转导通路的抑制型Smads,它可阻断TGF-β信号在胞浆内的传导,其紊乱是TGF-β信号转导通路紊乱的机制之一。本研究旨在分析在细胞恶性转化过程中,Smad7基因表达是否发生紊乱,TGF-β1对Smad7基因的调控功能有无发生变化,以探索细胞发生恶性转化的原因。方法:培养BEP2D细胞及BERP35T-2细胞,于收获前60min和90min加入不同剂量的TGF-β1,提取细胞总RNA,分别以未加TGF-β1的细胞组作为对照,用Northernblot杂交比较两组细胞Smad7mRNA表达的差异以及细胞对TGF-β1细胞因子刺激的反应性。同时提取BEP2D及BERP35T-2细胞蛋白,用Westernblot方法比较两组细胞内源性TGF-β1表达的差异。结果:Smad7mRNA表达水平恶性转化细胞高于永生化细胞;加了TGF-β1细胞因子后,BEP2D细胞Smad7mRNA表达增高,BERP35T-2细胞表达水平改变不明显。而内源性TGF-β1的表达水平,BERP35T-2细胞稍高于BEP2D细胞。结论:Smad7在辐射致肺癌细胞系中的过表达及对TGF-β1应答的降低可能是辐射诱发肺癌发生的机制之一。
BACKGROUND & AIM: The inhibition of cell growth and proliferation induced by transforming growth factor-β (TGF-β) is one of the most important mechanisms of tumorigenesis. Smad7 is an inhibitory Smads of TGF-β signal transduction pathway, which can block the transmission of TGF-β signal in the cytoplasm, which is one of the mechanisms of disorder of TGF-β signal transduction pathway. The purpose of this study was to investigate whether Smad7 gene expression is disturbed during malignant transformation and whether TGF-β1 regulates Smad7 gene function in order to explore the possible causes of malignant transformation. Methods: BEP2D cells and BERP35T-2 cells were cultured. Different doses of TGF-β1 were added 60 min and 90 min before harvest to extract the total RNA of the cells. The cells without TGF-β1 were used as control. Differences in Smad7 mRNA expression and cellular reactivity to TGF-β1 cytokine stimulation. The proteins of BEP2D and BERP35T-2 were extracted simultaneously, and the difference of endogenous TGF-β1 expression between the two groups was compared by Western blot. Results: Smad7 mRNA expression in malignant transformed cells was higher than that in immortalized cells. The expression of Smad7 mRNA in BEP2D cells was increased and the expression of BERP35T-2 cells was not changed after addition of TGF-β1. However, the expression of endogenous TGF-β1 was slightly higher in BERP35T-2 cells than in BEP2D cells. Conclusion: The overexpression of Smad7 in lung cancer cell lines induced by radiation and the decrease of TGF-β1 response may be one of the mechanisms of radiation-induced lung cancer.