血瘀证患者差异基因表达谱探究.docVIP

血瘀证患者差异基因表达谱探究 　 作者：马晓娟, 殷惠军, 陈可冀 【摘要】 目的：应用寡核苷酸基因芯片技术，研究血瘀证患者差异基因表达谱。方法：16例血瘀证患者经冠状动脉造影诊断后，分为冠心病血瘀证组和非冠心病血瘀证组，每组8例；并选年龄和性别相匹配的8名健康人为健康对照组。抽取静脉血，分离白细胞，抽提RNA，质控芯片（Test3芯片）对样本质量进行检测，然后与Affymetrix U133 Plus 2.0芯片进行杂交，通过扫描和软件分析，比较冠心病血瘀证组、非冠心病血瘀证组与健康对照组的基因表达谱，筛选血瘀证相关差异基因，并进一步进行基因本体（Gene Ontology, GO）和通路分析，使用实时荧光定量逆转录聚合酶链反应法对目标基因进行验证。结果：通过差异基因筛选，与血瘀证相关的差异基因共有48个，其中上调基因26个，下调基因22个；通过GO分析，其中与炎症免疫相关的基因有5个，占10.4%；通路分析结果显示，有意义的10条通路中有5个涉及炎症和免疫反应。经实时荧光定量逆转录聚合酶链反应验证了基因芯片准确可靠。结论：血瘀证基因表达谱研究显示了炎症和免疫相关基因的比例和显著性优势，说明炎症和免疫反应在一定程度上介导了血瘀证的发生发展。 【关键词】 血瘀证; 基因表达谱; 炎症; 免疫反应 　　Objective: To investigate the differential gene expression profiles in patients with blood stasis syndrome by oligonucleotide microarray technique. Methods: Sixteen patients with blood stasis syndrome were divided into patients with coronary heart disease (CAD) (n=8) and nonCAD patients (n=8) by using coronary angiography. The sex and agematched eight healthy persons were enrolled as control group. Venous bloods were collected for extracting RNA. Test3 chip was first employed to examine the quality of samples. Then the samples were hybridized with Affymetrix U133 Plus 2.0 array to compare the gene expression profiles among the three groups. Genearray scanner and gene chip operating software were applied to screen hybridization signals and analyze gene expression respectively. Based on the comparison of the three groups of samples, the differential genes related with blood stasis syndrome were analyzed by Gene Ontology (GO) and pathway, and confirmed by realtime reverse transcription polymerase chain reaction (RTPCR). Results: Fortyeight differential genes were found being associated with blood stasis syndrome, including 26 upregulated genes and 22 downregulated genes. Five of the fortyeight genes (10.4%) were related to inflammatory reaction and immune response through the GO analysis. In the pathway analysis, five of ten significant pathways were referred to inflammation and immune response. The results o