应用基因表达谱芯片技术筛选GABA作用下肝星状细胞差异表达基因
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肖 凡, 魏红山, 李国力, 张剑平, 宋 川
γ-氨基丁酸;肝星状细胞;基因芯片技术肖凡,魏红山,李国力,张剑平,宋川.应用基因表达谱芯片技术筛选GABA作用下肝星状细胞差异表达基因. 世界华人消化杂志2007;15(6)629-632,肖凡,魏红山,李国力,张剑平,宋
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肖凡, 魏红山, 李国力, 张剑平, 宋川, 北京地坛医院病毒研究所 北京市 100011
通讯作者: 魏红山, 100011, 北京市安外大街地坛公园13号, 北京地坛医院病毒研究所. cindy2358@163.com
电话: 010-64211031-2358 传真: 010-64211031-2306
收稿日期: 2006-12-13 接受日期: 2007-01-04
Screening of genes differentially expressed in hepatic stellate cells acted by gamma aminobutyric acid with microarray assay
Fan Xiao, Hong-Shan Wei, Guo-Li Li, Jian-Ping Zhang, Chuan Song
Fan Xiao, Hong-Shan Wei, Guo-Li Li, Jian-Ping Zhang, Chuan Song,Institute of Infectious Diseases, Beijing Ditan Hospital, Beijing 100011, China
Correspondence to:Hong-Shan Wei, Institute of Infectious Diseases, Beijing Ditan Hospital, 13 Ditan Park, Anwai Street, Beijing 100011, China. cindy2358@163.com
Received:2006-12-13 Accepted:2007-01-04
Abstract
AIM:To screen genes regulated by gamma aminobutyric acid (GABA) in hepatic stellate cells (HSC) using microarray assay, and to investigate GABA biological function.
METHODS: HSC-T6 was co-cultured with 100 mmol/L GABA for 48 hours, and then mRNA was extracted from the cells to reversely transcribe into cDNA. HSC-T6 co-cultured with phosphate buffer saline (PBS) was used as controls. The cDNA profile was analyzed by microarray assay.
RESULTS:For the profile of 4096 genes, 37 genes were found differentially expressed, of which 11 were up-regulated while 26 were down-regulated significantly.
CONCLUSION: Differentially expressed genes under the action of GABA are successfully screened by microarray technique, demonstrating that GABA can affect the gene profile of HSC.
Key Words: Gamma aminobutyric acid; Hepatic stellate cell; Microarray assay
Xiao F, Wei HS, Li GL, Zhang JP, Song C. Screening of genes differentially expressed in hepatic stellate cells acted by gamma aminobutyric acid with microarray assay. Shijie Huaren Xiaohua Zazhi 2007;15(6):629-632
摘要目的:应用基因芯片技术检测g-氨基丁酸(GABA)对肝星状细胞(HSC-T6) 基因表达谱的影响.
方法: 10mmol/L的GABA作用于HSC-T6细胞48 h, 提取mRNA, 逆转录为cDNA. 以PBS作用的HSC-T6细胞为对照, 进行cDNA 芯片分析.
结果: 在4096个基因表达谱的筛选中, 发现有11个基因表达水平显著上调, 26个基因表达水平显著下调.
结论: 成功地应用基因芯片筛选GABA作用下肝星状细胞差异表达基因, 证明GABA对肝星状细胞的表达谱有显著作用.
关键词: γ-氨基丁酸; 肝星状细胞; 基因芯片技术
肖凡, 魏红山, 李国力, 张剑平, 宋川. 应用基因表达谱芯片技术筛选GABA作用下肝星状细胞差异表达基因. 世界华人消化杂志 2007;15(6):629-632
RNA电泳图. 1, 2: GABA-10 mmol/L; 3,4: Control.
2.2 芯片杂交体系验证及结果分析在芯片上共有4096个cDNA.为了监控芯片杂交技术体系的整个过程, 在芯片上设置阴性对照(8条水稻基因,共8个点), 这些点的杂交信号均很低, 证实了数据的可靠性.由于实验组探针标记Cy5荧光素(呈红色),对照组探针标记Cy3荧光素(呈绿色),红绿颜色的差异就显示该基因在实验组和对照组中基因表达水平上的差异, 黄色代表表达水平无差异. 按阳性标准, 从4096个基因中筛选出差异表达基因共27条,其中11条基因表达增强, 26条基因表达降低(图2,图3).
图2 双色荧光标记叠加图.
图3 杂交信号强度散点图.
2.3 差异表达基因分析 在基因芯片的扫描分析中, 如果荧光染料的Cy5/Cy3比值在2.0以上,就判断为上调基因, 在本研究中发现有11种基因的表达水平上调(表1).如果荧光染料的Cy5/Cy3比值在0.5以下,就判断为下调基因, 在本研究中发现有26种基因的表达水平下调(表2) ......
肖凡, 魏红山, 李国力, 张剑平, 宋川, 北京地坛医院病毒研究所 北京市 100011
通讯作者: 魏红山, 100011, 北京市安外大街地坛公园13号, 北京地坛医院病毒研究所. cindy2358@163.com
电话: 010-64211031-2358 传真: 010-64211031-2306
收稿日期: 2006-12-13 接受日期: 2007-01-04
Screening of genes differentially expressed in hepatic stellate cells acted by gamma aminobutyric acid with microarray assay
Fan Xiao, Hong-Shan Wei, Guo-Li Li, Jian-Ping Zhang, Chuan Song
Fan Xiao, Hong-Shan Wei, Guo-Li Li, Jian-Ping Zhang, Chuan Song,Institute of Infectious Diseases, Beijing Ditan Hospital, Beijing 100011, China
Correspondence to:Hong-Shan Wei, Institute of Infectious Diseases, Beijing Ditan Hospital, 13 Ditan Park, Anwai Street, Beijing 100011, China. cindy2358@163.com
Received:2006-12-13 Accepted:2007-01-04
Abstract
AIM:To screen genes regulated by gamma aminobutyric acid (GABA) in hepatic stellate cells (HSC) using microarray assay, and to investigate GABA biological function.
METHODS: HSC-T6 was co-cultured with 100 mmol/L GABA for 48 hours, and then mRNA was extracted from the cells to reversely transcribe into cDNA. HSC-T6 co-cultured with phosphate buffer saline (PBS) was used as controls. The cDNA profile was analyzed by microarray assay.
RESULTS:For the profile of 4096 genes, 37 genes were found differentially expressed, of which 11 were up-regulated while 26 were down-regulated significantly.
CONCLUSION: Differentially expressed genes under the action of GABA are successfully screened by microarray technique, demonstrating that GABA can affect the gene profile of HSC.
Key Words: Gamma aminobutyric acid; Hepatic stellate cell; Microarray assay
Xiao F, Wei HS, Li GL, Zhang JP, Song C. Screening of genes differentially expressed in hepatic stellate cells acted by gamma aminobutyric acid with microarray assay. Shijie Huaren Xiaohua Zazhi 2007;15(6):629-632
摘要目的:应用基因芯片技术检测g-氨基丁酸(GABA)对肝星状细胞(HSC-T6) 基因表达谱的影响.
方法: 10mmol/L的GABA作用于HSC-T6细胞48 h, 提取mRNA, 逆转录为cDNA. 以PBS作用的HSC-T6细胞为对照, 进行cDNA 芯片分析.
结果: 在4096个基因表达谱的筛选中, 发现有11个基因表达水平显著上调, 26个基因表达水平显著下调.
结论: 成功地应用基因芯片筛选GABA作用下肝星状细胞差异表达基因, 证明GABA对肝星状细胞的表达谱有显著作用.
关键词: γ-氨基丁酸; 肝星状细胞; 基因芯片技术
肖凡, 魏红山, 李国力, 张剑平, 宋川. 应用基因表达谱芯片技术筛选GABA作用下肝星状细胞差异表达基因. 世界华人消化杂志 2007;15(6):629-632
RNA电泳图. 1, 2: GABA-10 mmol/L; 3,4: Control.
2.2 芯片杂交体系验证及结果分析在芯片上共有4096个cDNA.为了监控芯片杂交技术体系的整个过程, 在芯片上设置阴性对照(8条水稻基因,共8个点), 这些点的杂交信号均很低, 证实了数据的可靠性.由于实验组探针标记Cy5荧光素(呈红色),对照组探针标记Cy3荧光素(呈绿色),红绿颜色的差异就显示该基因在实验组和对照组中基因表达水平上的差异, 黄色代表表达水平无差异. 按阳性标准, 从4096个基因中筛选出差异表达基因共27条,其中11条基因表达增强, 26条基因表达降低(图2,图3).
图2 双色荧光标记叠加图.
图3 杂交信号强度散点图.
2.3 差异表达基因分析 在基因芯片的扫描分析中, 如果荧光染料的Cy5/Cy3比值在2.0以上,就判断为上调基因, 在本研究中发现有11种基因的表达水平上调(表1).如果荧光染料的Cy5/Cy3比值在0.5以下,就判断为下调基因, 在本研究中发现有26种基因的表达水平下调(表2) ......
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