| Lung cancer is the one of most common malignant tumor in world wide and in recent twenty years, the incidence of lung cancer has increased quickly in our country. Till now, the morbidity and mortality of lung cancer have occupied the top of the cancers. Early diagnosis is the most important key to increase the five-year survival rate. The behavior and outcome of lung cancers are highly variable, and not only is the molecular basis of this variability unknown, but neither standard histopathology nor currently available molecular markers can predict these characteristics. Accordingly, the identification of novel molecular biomarkers to differentiate tumor from normal cells and predict tumor behavior such as pathologic stage, response to chemotherapy, and site of relapse, is of great importance in clinical practice. In present study , we use arrayed human placenta cDNA library to make gene DNA microarray chip by which gene expression profile associated with NSCLC was screened. We found a down-regulated gene in NSCLC comparede with adjacent normal lung tissue. It was then confirmed as glypican-3 gene by sequencing. Biochemical studies and cell culure assays have implied gpc3 serve as adhesion, motility, proliferation, differentiation, morphogenesis, carcinogensis and development such as ovarian cancer, breast cancer, mesotheliomas, kidney cancer, gastric cancer, hepatpcarcinoma and Wilm's tumor.Part 1. Investigation of gene screening of NSCLC associated gene using arrayed library.Objective To screen gene assiociated with NSCLC . Methods The arrayed library was prepared from human's placenta cDNA library phurchased from Invitrogen company and was carried out by the Biorobatic TAS multifunction DNA microarray machine. Total RNA was purified from two paired NSCLC tissues and adjacent normal lung tissues andreverse-transcripted to cDNA with biotin probe-labeled and then hybridized with the cDNA arrays respectively. Results We found a clone which have an at least 5-fold decreased gene in NSCLC tissues compared with that hybridized with adjacent normal lung tissues. After sequencing and comparing to the GenBank, it was identified as gpc3 gene, encoding a member of Glypican family. Conclusion The expression of gpc3 down-regulated in NSCLC was first discovered. Gene chip made from arrayed library can screen genes associated with NSCLC effectually and economically.Part 2. Gpc3 gene expression in human NSCLCObjective To investigated the gpc3 gene expression in human NSCLC . Methods Gpc3 gene expression in 38 NSCLC with paired tumor tissues and adjacent normal lung tissues was detected by RT-PCR , Northern Blot analysis and real-time RT-PCR. Results Gpc3 was detected expressing in adjacent normal lung tissues but not detected in tumor tissues by RT-PCR and Northern Blot analysis. Real-time RT-PCR quantitively examine the expression level of gpc3 reveal that the average comparative expression level in lung tissues was 0.0265+0.0119, while in NSCLC was 0.0026+0.0024, the defference between them was significant(P<0.0001).Of one hundred percent cases, the expression ofgpc3 was more than 2-fold in adjacent normal lung tissues compared with in tumor tissues,of eighty-four percent cases more than 5-fold. Analysized with the case history and pathological materials. In non-smokers, the ratio of gpc3 in tumor tissues compared with in adjacent normal lung tissues was significantly higher than that in smokers(P<0.05); In small tumor(<3cm), the ratio of gpc3 in tumor tissues compared with in adjacent normal lung tissues was significantly higher than that in large tumor(>3cm,P<0.05); The disversity expression of gpc3 was not correlated with age, sex, pathology type(squamose NSCLC and adenocarcinoma NSCLC), differentiated degree(well differentiated and poorly differentiated), clinic type(centrality NSCLC and vicinity NSCLC), fibrocapsule and lymphnode metastasis. Conclusion The down-regulated expression of gpc3 in NSCLC was universal. The disversity expression of gpc3 was correlated to cigarette smoking and tumor size.Part 3. G...
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