Abstract
Lung cancer is the most common malignant tumor in hunan,and smoking is the first cause.The lung carcinogenesis is related to gene mutation,so its important to study the cell reaction to smoking and tumor generation in gene level.
cDNA microarray was used to screen out smoking related lung cancer genes,and the result was checked by RT-PCR. Inhibitor and activator was used to control the gene expression. TG2 expression level were detected by Western-blot and enzyme activity test,and flow cytometry was used to analyze cell apoptosis ,cell proliferation and cell cycle in malignant transformation of bronchial epithelial cells induced by cigarette smoking.
The total of differential expressed gene is 124,79 was up-regulated and 45 was down-regulated.These differential expressed can be divided into 27 kinds of group such as metabolism,cell apoptosis,cell cycle, Notch signal pathway,p53 signal pathway, peroxisome Proliferator-activated receptor pathway. The coincidence rate of RT-PCR and Gene chip was above 80%,which suggest the reliability of the gene chip result.
TG2 expression was down regulated in malignant transformed bronchial epithelial cells and cells exposed to cigarette smoking.TG2 expression and activitie change affect cell apoptosis,cell prolifiction and cell cycle.TG2 may affect cell malignant transformation through the regulation of these events.
Key words:lung cacer; gene chip; differential expressed gene; cigarette smoking; malignant transformation
摘 要
肺癌是人类最常见的恶性肿瘤,吸烟是引起肺癌的首要因素。肺癌的发生与细胞基因突变有关,从基因水平上研究细胞对卷烟烟气的反应及肿瘤形成过程是肿瘤研究的关键所在。
运用人全基因组芯片技术检测吸烟致肺癌相关基因差异表达,通过RT-PCR验证基因芯片结果的真实性。采用抑制剂和激活剂调节基因的蛋白表达和活性,运用western-blot、酶活性检测、流式细胞术等检测TG2在卷烟烟气在支气管上皮细胞恶性转化过程中的的表达情况及对细胞凋亡、增殖、周期等的影响。
结果发现吸烟致肺癌差异表达基因中,差异倍数在两倍以上的共124个,其中表达上调基因79个,表达下调基因45个。这些差异表达基因大致可分代谢、细胞凋亡、细胞周期、Notch信号通路、p53信号通路、过氧化物酶体增生物激活受体信号通路等共27个功能组。RT-PCR验证基因芯片结果发现,RT-PCR结果和基因芯片结果吻合率80%以上,说明基因芯片的结果比较真实可靠。
对差异表达基因TG2在卷烟烟气致支气管上皮细胞恶性转化相关性研究中发现,TG2在恶性转化细胞中表达下调,单次染毒也能使TG2表达下调,TG2表达及活性改变对细胞凋亡、增殖、周期等产生影响。TG2可能通过调控细胞凋亡、增殖、周期等影响卷烟烟气对支气管上皮细胞的恶性转化。
关键词:肺癌;基因芯片;差异表达基因;卷烟烟气;恶性转化
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