SERIAL ANALYSIS OF GENE EXPRESSION (SAGE) IN HUMAN STOMACH CANCER

本项目的目的是对人胃癌进行基因表达序列分析（SAGE）。SAGE最初是在组织培养系统中开发的，应用于临床胃癌基因表达的全局分析，需要从含有最少量基质细胞的组织样本中纯化高质量的mRNA。1999年，在检查组织样本之前，我们用SAGE分析了从我们研究所未分化的胃癌中建立的癌细胞系KKLS的基因表达。初步分析可定量鉴定出5182个SAGE标签中表达的2969个独特基因，连接率为1.02%。在高表达基因中包括线粒体基因和一些未在基因库中登记的未知基因。我们认为KKLS的基因表达谱将为进一步分析不同组织学类型的人胃癌的基因表达奠定基础。2000年，人类胃癌组织进行了SAGE。在分析过程中，我们在从手术材料中制备高纯度mRNA时遇到了困难，后来通过改进mRNA的提取和纯化方法克服了这一困难。与KKLS细胞系的基因表达模式相似，在人胃癌样本SAGE标签文库中定量检测到未知的核基因和线粒体基因。最近的报道显示，线粒体DNA中频繁的基因改变表明，线粒体基因表达的改变参与了胃癌的发生和进展。虽然我们不能在规定的期限内（1999 - 2000）完成这个项目，但我们的实验室正在进行针对人类胃癌的SAGE，目标是核基因和线粒体基因。

The purpose of this project is to conduct serial analysis of gene expression (SAGE) for human stomach cancer. Applying SAGE, that was originally developed in the tissue culture system, for global analysis of gene expression in clinical stomach cancer requires purification of high quality mRNA from tissue samples that contain minimal amount of stromal cells. In 1999, prior to examine tissue samples, we analyzed by SAGE for gene expression in a cancer cell line, KKLS that was established from undifferentiated stomach carcinoma in our institute. This preliminary analysis could quantitatively identify 2969 unique genes expressed among 5182 SAGE tags obtained from the cell line, with a linker ratio of 1.02%. Among highly expressed genes were included mitochondrial genes and several unknown genes unregistered to the Gene Bank. We assume that the gene expression profile of KKLS would be fundamental for further analysis of gene expression in human stomach cancer showing different histological types.In 2000, human stomach cancer tissues has been subjected to SAGE.During this analysis we have had difficulty in preparation of highly purified mRNA from the surgical materials, which was later overcome by modification of our method for extraction and purification of mRNA.Similar to the gene expression pattern in KKLS cell line, unknown nuclear genes and mitochondrial genes were quantitatively detected in the library of SAGE tags yielded from human stomach cancer samples. Recent reports showing frequent genetic alteration in mitochondrial DNA suggest an involvement of altered expression of mitochondrial genes in development and progression of stomach cancer. Although we could not complete this project in the given term (1999 to 2000), SAGE for human stomach cancer is being in progress in our laboratory, targeting nuclear as well as mitochondrial genes.