脑胶质瘤相关新基因的聚类分析在其分子病理学分类中的意义

CLUSTER ANALYSIS AND IDENTIFY SIGNIFICANCE OF NOVEL GENES RELATED TO GLIOMA IN MOLECULE CLASSIFICATION

目的 探讨人脑胶质瘤发生发展中相关基因的表达及功能 ,初步建立基于基因表达谱的胶质瘤分子病理学分类方法。方法 用含 13939种人类基因的BioStarH14 0S型表达谱芯片 ,以正常脑组织及胶质瘤组织总RNA制备的探针杂交芯片 ;ScanArray 4 0 0 0扫描芯片 ,提取正常脑组织及胶质瘤组织差异基因并行生物信息分析 ,对部分新基因行Northern杂交、原位杂交等功能研究 ;Hierarchical聚类对差异基因进行特征提取 ,进行初步胶质瘤分子病理分类。结果 在正常脑组织与18例不同级别胶质瘤组织中筛选出 12 0 0条 (8 6 1% )差异基因 ,其中 395条 (2 83% )基因尚未登录GeneBank ,胶质组织瘤中 348条基因表达上调 ,85 2条下调 ,信息分析与细胞信号传导等多类基因密切相关 ;Northern、原位杂交显示新基因与胶质瘤密切相关 ;聚类发现微管相关蛋白、黏附素超家族蛋白、细胞外基质相关基因等与分子分型相关 ,基于基因表达谱的分子病理分类与依据细胞形态的病理学分类基本一致 ,且更能反映胶质瘤发生发展的内在本质。结论 胶质瘤发生发展存在多类基因表达改变 ,表达谱芯片对胶质瘤分类具有特异性高、重复性好、样本量少的特点 ,基因表达谱及相关新基因的发现和分子病理学分类有助于阐明胶质瘤分子病理学机制。

OBJECTIVE TO SCREEN DIFFERENTIALLY EXPRESSED GENES IN THE DEVELOPMENT OF HUMAN GLIOMA AND ESTABLISH MOLECULAR CLASSIFICATION OF GLIOMA PRELIMINARY BASED ON GENE EXPRESSION USING CDNA MICROARRAY.METHODS BRAIN SPECIMENS WERE OBTAINED FROM 18 PATIENTS WITH GLIOMA, 10 MALES AND 8 FEMALES, AGED 14~62 WITH AN AVERAGE AGE OF 44.4. THE TOTAL RNAS OF THESE GLIOMA SPECIMENS AND 2 SPECIMENS OF DONATED BRAIN OF NORMAL ADULTS WERE EXTRACTED. BIOSTARH140S MICROARRAY (INCLUDING 8347 OLD GENES AND 5592 NOVEL GENES) WERE ADOPTED AND HYBRIDIZED WITH PROBES WHICH WERE PREPARED FROM THE TOTAL RNAS. DIFFERENTIALLY EXPRESSED GENES BETWEEN THE NORMAL TISSUES AND GLIOMA TISSUES WERE ASSAYED AFTER SCANNING CDNA MICROARRAY WITH SCANARRAY 4000. NORTHERN HYBRIDIZATION, AND IN SITU HYBRIDIZATION (ISH) WERE USED TO IDENTIFY THE FUNCTIONS OF NOVEL GENES. THOSE DIFFERENTIALLY EXPRESSED GENES WERE STUDIED WITH HIERARCHICAL METHOD AND MOLECULAR CLASSIFICATION OF GLIOMA WAS PRELIMINARILY CARRIED OUT.RESULTS AMONG THE 13 939 TARGET GENES, THERE WERE 1200 (8.61%) DIFFERENTIALLY EXPRESSED GENES AND 395 (2.83%) NOVEL GENES. 348 GENES WERE UP-REGULATED AND 852 GENES WERE DOWN-REGULATED IN GLIOMA. THE RESULTS OF BIOINFORMATICAL ANALYSIS, NORTHERN HYBRIDIZATION AND ISH REVEALED THAT THOSE NOVEL GENES WERE HIGHLY ASSOCIATED WITH GLIOMA. THERE WERE MULTIPLE GENES WHICH WERE RELEVANCE TO CLASSIFICATION BY HIERARCHICAL METHOD, SUCH AS MAP GENE, CYTOSKELETON AND MATRIX MOTILITY GENES, ETC. MOLECULAR CLASSIFICATION OF GLIOMA WITH HIERARCHICAL CLUSTER WAS IN ACCORDANCE WITH PATHOLOGY AND REVEALED INTERNAL ESSENCE IN TUMORIGENESIS AND DEVELOPMENT.CONCLUSION MULTIPLE GENES PLAY IMPORTANT ROLES IN DEVELOPMENT OF GLIOMA. CDNA MICROARRAY TECHNOLOGY IS A POWERFUL TECHNIQUE IN SCREENING FOR DIFFERENTIALLY EXPRESSED GENES BETWEEN TWO DIFFERENT KINDS OF TISSUES.