Gene expression profiles of different clinical stages of colorectal carcinoma: toward a molecular genetic understanding of tumor progression

Background and aims Colorectal cancer is one of the leading causes of cancer deaths in the Western world. A better understanding of the development and progression of colorectal carcinoma is needed to define novel targets and strategies for treatment.Patients/methods Gene expression profiles were determined for primary tumors of 10 locally restricted (T3N0M0), 8 lymphatically metastasized (T3N+M0), 7 systemically metastasized (T3N+M1) colorectal carcinomas, and 6 specimens of normal colorectal tissue by histology-guided oligonucleotide microarray analysis.Results A total of 1,995 genes were differently regulated in primary tumors of colorectal carcinoma compared with normal colorectal tissue. Besides common features of dedifferentiation and different expression of genes involved in cell division, cell adhesion, angiogenesis, signal transduction and metabolism we observed a deregulation of genes with an as yet unclear function. We identified 126 genes that were subsequently up- and 204 genes down-regulated during tumor progression. Furthermore, we found a cluster of five genes exclusively up-regulated in primary tumors of systemically metastasized colorectal carcinomas. A comparison of locally restricted (T3N0M0) and systemically metastasized (T3N+M1) primary tumors showed 50 deregulated genes with a massive down-regulation of immune-modulatory genes in primary tumors of systemically metastasized carcinomas. Primary tumors of lymphatically (T3N+M0) and systemically metastasized (T3N+M1) carcinomas differed in the expression of 19 genes.Conclusion These results provide an additional step toward the identification of crucial genes for the progression of colorectal cancer and the identification of novel treatment targets or strategies.     

外源质粒DNA对小鼠肠道基因表达谱的影响

目的:研究外源质粒通过胃肠道途径吸收对小鼠肠道基因表达谱的影响.方法:给Balb/c小鼠灌胃质粒pcDNA3200μg,在灌胃后4h后分离空肠一段,提取肠组织的总RNA.利用寡核苷酸芯片对灌胃质粒pcDNA3后的Balb/c小鼠肠道进行基因表达谱研究.结果:灌胃外源质粒DNA后,所检测的17667基因中有61条基因产生差异表达,其中36条基因表达上调,25条基因表达下调.这些差异表达的基因主要涉及免疫应答、抗氧化及解毒功能、脂质代谢、阴离子转运蛋白、细胞凋亡及信号转导等过程.结论:外源质粒DNA通过胃肠道途径可广泛调控肠道多种基因表达.     

系统性红斑狼疮的基因表达谱及其免疫调控通路的研究

目的 通过基因表达谱勾画系统性红斑狼疮（SLE）发病机制中可能的免疫调控通路。方法 用寡核苷酸基因芯片研究了10例SLE（包括2例SLE同胞对）和10份正常对照的外周血白细胞基因表达谱。进行了基因表达谱和临床免疫表型的聚类分析和比较，筛选统计学显著性差异表达的SLE相关基因，并在转录和蛋白表达水平验证芯片表达谱结果。结果 聚类分析揭示SLE临床免疫表型与基因表达谱特征存在关联。获得25个显著性表达的SLE相关基因，其中大部分未经报道。转录水平直接验证了其中C／EBPD，LY6E，OAS2三个基因的表达，较大样本独立验证了C／EBPD，LY6E在SLE中的表达上调；蛋白质水平验证了C／EBPD的高表达。结论 SLE基因表达谱可能是其临床免疫表型的分子基础，探讨了通过基因表达谱进行SLE分型乃至鉴别诊断的可能性。通过显著性差异表达的SLE相关基因，推测干扰素及其免疫调控通路在SLE发病中可能扮演重要角色。     

Modulation of gene expression in MHCC97 cells by interferon alpha

AIM: To elucidate the molecular mechanisms of the inhibitory effects of IFN-α on tumor growth and metastasis in MHCC97 xenografts. METHODS: Three thousand international units per milliliter of IFN-α-treated and -untreated MHCC97 cells were enrolled for gene expression analysis using cDNA microarray. The mRNA levels of several differentially expressed genes in cDNA microarray were further identified by Northern blot and RT-PCR. RESULTS: A total of 190 differentially expressed genes including 151 IFN-α-repressed and 39 -stimulated genes or expressed sequence tags from 8 464 known human genes were found to be regulated by IFN-α in MHCC97. With a few exceptions, mRNA levels of the selected genes in RT-PCR and Northern blot were in good agreement with those in cDNA microarray. CONCLUSION: IFN-α might exert its complicated anti-tumor effects on MHCC97 xenografts by regulating the expression of functional genes involved in cell metabolism, proliferation, morphogenesis, angiogenesis, and signaling.     

Gene expression profile differences in gastric cancer, pencancerous epithelium and normal gastric mucosa by gene chip

AIM: To study the difference of gene expression in gastric cancer (T), pericancerous epithelium (P) and normal tissue of gastric mucosa (C), and to screen an associated novel gene in early gastric carcinogenesis by oligonudeotide microarray. METHODS: U133A (Affymetrix, Santa Clara, CA) gene chip was used to detect the gene expression profile difference in T, P and C, respectively. Bioinformatics was used to analyze the detected results. RESULTS: When gastric cancer was compared with normal gastric mucosa, 766 genes were found, with a difference of more than four times in expression levels. Of the 766 genes, 530 were up-regulated (Signal Log Ratio [SLR]>2), and 236 were down-regulated (SLR<-2). When pericancerous epithelium was compared with normal gastric mucosa, 64 genes were found, with a difference of more than four times in expression levels. Of the 64 genes, 50 were up-regulated (SLR>2), and 14 were down-regulated (SLR<-2). Compared with normal gastric mucosa, a total of 143 genes with a difference in expression levels (more than four times, either in cancer or in pericancerous epithelium) were found in gastric cancer (T) and pericancerous epithelium (P). Of the 143 genes, 108 were up-regulated (SLR>2), and 35 were down-regulated (SLR<-2). CONCLUSION: To apply a gene chip could find 143 genes associated with the genes of gastric cancer in pericancerous epithelium, although there were no pathological changes in the tissue slices. More interesting, six genes of pericancerous epithelium were up-regulated in comparison with genes of gastric cancer and three genes were down-regulated in comparison with genes of gastric cancer. It is suggested that these genes may be related to the carcinogenesis and development of early gastric cancer.     

Gene expression profile differences in gastric cancer, pericancerous epithelium and normal gastric mucosa by gene chip

AIM: To study the difference of gene expression in gastric cancer (T), pericancerous epithelium (P) and normal tissue of gastric mucosa (C), and to screen an associated novel gene in early gastric carcinogenesis by oligonucleotide microarray. METHODS: U133A (Affymetrix, Santa Clara, CA) gene chip was used to detect the gene expression profile difference in T, P and C, respectively. Bioinformatics was used to analyze the detected results. RESULTS: When gastric cancer was compared with normal gastric mucosa, 766 genes were found, with a difference of more than four times in expression levels. Of the 766 genes, 530 were up-regulated (Signal Log Ratio (SLR) >2), and 236 were down-regulated (SLR<-2). When pericancerous epithelium was compared with normal gastric mucosa, 64 genes were found, with a difference of more than four times in expression levels. Of the 64 genes, 50 were up-regulated (SLR>2), and 14 were down-regulated (SLR<-2). Compared with normal gastric mucosa, a total of 143 genes with a difference in expression levels (more than four times, either in cancer or in pericancerous epithelium) were found in gastric cancer (T) and pericancerous epithelium (P). Of the 143 genes, 108 were up-regulated (SLR>2), and 35 were down-regulated (SLR<-2). CONCLUSION: To apply a gene chip could find 143 genes associated with the genes of gastric cancer in pericancerous epithelium, although there were no pathological changes in the tissue slices. More interesting, six genes of pericancerous epithelium were up-regulated in comparison with genes of gastric cancer and three genes were down-regulated in comparison with genes of gastric cancer. It is suggested that these genes may be related to the carcinogenesis and development of early gastric cancer.     

应用人类全基因组寡核苷酸芯片筛查自身免疫性肝炎相关基因

为研究自身免疫性肝炎发病过程中基因表达改变与免疫反应的关系,我们采用人类全基因寡核菅酸芯片筛查自身免疫性肝炎差异表达基因,并对结果进行聚类分析,筛选出与疾病关系最密切的基因功能群,为进一步阐明发病机制,寻找致病基因提供参考。     

以基因芯片技术研究胸腺类癌引起异位ACTH综合征的基因表达

目的　研究引起异位ACTH综合征的胸腺类癌组织与正常胸腺组织的基因表达差异,探讨胸腺类癌引起异位ACTH综合征的发病机制。方法　利用基因芯片技术,通过不同荧光(Cy3和Cy5)标记的对照和胸腺类癌组织RNA样本与芯片杂交,观察两种组织间的基因差异表达。结果　研究发现,基因芯片的4 224个基因中403个基因在胸腺类癌组织中表达下调, 394个基因表达上调2倍以上(其中23个与细胞分裂有关), 51个基因上调5倍以上(其中与细胞分裂有关的基因1个,即PAK3)。结论　引起异位ACTH综合征的胸腺类癌组织与正常胸腺组织存在多个差异表达的基因,PAK3可能参与了该肿瘤的发生。     

应用寡核苷酸芯片对乙型肝炎病毒转染HepG2细胞基因表达谱的研究

HBV与肝细胞之间相互作用的分子机制还不清楚，对HBV蛋白调控宿主细胞基因表达情况也了解甚少。本实验选用基因芯片进行HBV基因表达谱改变的研究，筛选HBV感染应答基因，探讨HBV感染的分子致病机制，寻找预防和治疗HBV感染的靶点。     

Gene expression profile of esophageal cancer in North East India by cDNA microarray analysis

AIM: To identify alterations in genes and molecular functional pathways in esophageal cancer in a high incidence region of India where there is a widespread use of tobacco and betel quid with fermented areca nuts. METHODS: Total RNA was isolated from tumor and matched normal tissue of 16 patients with esophageal squamous cell carcinoma. Pooled tumor tissue RNA was labeled with Cy3-dUTP and pooled normal tissue RNA was labeled with Cy5-dUTP by direct labeling method. The labeled probes were hybridized with human 10K cDNA chip and expression profiles were analyzed by Genespring GX V 7.3 (Silicon Genetics). RESULTS: Nine hundred twenty three genes were differentially expressed. Of these, 611 genes were upregulated and 312 genes were downregulated. Using stringent criteria (P ≤ 0.05 and ≥ 1.5 fold change), 127 differentially expressed genes (87 upregulated and 40 downregulated) were identified in tumor tissue. On the basis of Gene Ontology, four different molecular functional pathways (MAPK pathway, G-protein coupled receptor family, ion transport activity, and serine or threonine kinase activity) were most significantly upregulated and six different molecular functional pathways (structural constituent of ribosome, endopeptidase inhibitor activity, structural constituent of cytoskeleton, antioxidant activity, acyl group transferase activity, eukaryotic translation elongation factor activity) were most significantly downregulated. CONCLUSION: Several genes that showed alterations in our study have also been reported from a high incidence area of esophageal cancer in China. This indicates that molecular profiles of esophageal cancer in these two different geographic locations are highly consistent.     

应用肿瘤基因解剖工程数据库检测大肠癌基因表达谱

目的进一步了解大肠癌与正常组织间基因表达谱差异，寻找可能用于临床诊断和治疗的基因标志。方法应用肿瘤基因解剖工程(CGAP)数据库中基因表达短序列分析(SAGE)数据筛查大肠癌和正常组织差异表达基因，并用RT-PCR方法进一步在大肠癌细胞株(SW1116、Lovo、HCT-8、HCe-8693)和20例组织标本中验证。结果在SACE数据库中共分析大肠癌和正常组织短序列195160个，发现差异基因216个。对其中17个基因进行RT-PCR验证分析，在细胞株中基因检出阳性率为35．3％～76．5％，组织标本中阳性率为88．2％。对其中转化生长因子β1、蛋白酶体26S亚单位、热休克蛋白1进行半定量RT-PCR，基因差异表达率分别为60％、50％、35％。结论利用CGAP数据库中SAGE数据能快捷、可靠地筛查大肠癌差异基因表达谱，对这些差异基因进一步分析可能为临床诊治大肠癌提供基因标志。     

肝癌相关基因DLK1转基因小鼠的构建与鉴定

目的 构建并鉴定肝脏特异性表达的DLKl转基因小鼠.方法 通过基因重组方法,将小鼠DLK1 cDNA片段置于小鼠白蛋白基因增强子和启动子序列下游,构建肝脏特异性表达的DLKl重组质粒,酶切重组质粒得到转基因片段,转基因在体外进行表达鉴定后,显微注射获得DLK1转基因首建小鼠,对首建小鼠进行传代,利用F1代小鼠进行DLK1转基因表达的鉴定.结果 逆转录(RT)-PCR和细胞免疫荧光显示,转基因DLK1片段可在小鼠肝癌细胞系Hep1-6中表达.RT-PCR和免疫组化结果显示,DLK1在F1代成年转基因小鼠肝脏中特异性表达.结论 成功构建了肝脏特异性表达的DLK1转基因小鼠.     

长期高脂饮食对C57BL/6小鼠骨骼肌基因表达谱的影响

目的 运用基因芯片技术研究长期高脂饮食对C57BL/6小鼠骨骼肌基因表达谱的影响,探究长期高脂饮食导致小鼠肥胖、胰岛素抵抗(IR)和2型糖尿病(T2DM)的分子机制.方法 20只雄性4周龄C57 BL/6小鼠随机分为正常饮食组和高脂饮食组,各10只,分别饲以基础和高脂饲料.16周后,称量各组小鼠体重;测定血糖、空腹血清胰岛素(FINs)、高密度脂蛋白(HDL)、甘油三酯(TG)、总胆固醇(TC)和游离脂肪酸(FFAs)水平;随后,每组随机选取4只小鼠处死,分离股四头肌,提取总RNA进行基因芯片分析.结果 高脂饮食组与正常饮食组相比,体重显著增加;FINs水平显著升高;HDL水平无显著性差异,而TC、TG和FFA水平均显著升高.采用基因芯片相关统计软件分析数据,结果发现正常饮食组与高脂饮食组相比,骨骼肌共出现590个差异表达基因.其中表达上调基因有321个,表达下调基因有269个.结论长期高脂饮食可以引起C57BL/6小鼠骨骼肌基因谱发生显著变化.     

实时荧光定量RT-PCR和寡核苷酸芯片方法分析食管腺癌组织中基因的表达

目的采用实时荧光定量RT—PCR和寡核苷酸芯片技术分析食管腺癌基因表达的特征。方法应用寡核苷酸芯片筛选6例食管腺癌基因表达谱,实时荧光定量RT—PCR验证其中8个基因的表达变化。结果2倍差异表达基因（DEGs）中,上调基因共212个,下调基因共126个;涉及细胞周期相关基因、信号转导相关基因、血管生成相关基因、细胞增殖相关基因、凋亡抑制基因、抑癌基因、黏附和代谢等。实时荧光定量RT-PCR和寡核苷酸芯片技术对8个基因的检测结果一致,不仅变化方向相同,而且表达值非常接近。结论食管腺癌的发生、发展涉及多基因多步骤,是一个复杂的过程。寡核苷酸芯片和实时荧光定量RT-PCR分析基因表达变化都是可信的,基因芯片筛选基因表达谱具有高通量大规模的特点,而实时荧光定量RT-PCR则适合单个基因表达变化的研究,两者互为补充和印证。     

应用基因芯片技术筛选胰腺癌相关基因

目的应用基因芯片技术筛选胰腺癌相关基因。方法将14000种人类基因PCR产物按微矩阵排列点样于化学涂层的载玻片上，制成基因芯片。按一步法抽提4例胰腺癌和癌旁正常胰腺组织的总RNA，将等量的RNA分别逆转录合成荧光分子掺人的cDNA一链作探针，混合后杂交上述基因芯片。经严格洗片后用ScanArray 4000扫描仪扫描芯片荧光信号图像，每点上两种荧光信号的强度分别代表Cy5-dCTP和Cy3-dCTP的量，获得的荧光信号图像用计算机分析。结果按差异显著性标准，从14000个基因中筛选出在胰腺癌组织中共同差异表达基因189条，其中已知基因101条，新基因88条。在筛选出的已知基因中，有50条表达上调，51条表达下调。结论基因芯片技术的肿瘤基因表达谱分析能够高通量筛选胰腺癌相关基因。并高效对基因功能进行研究。胰腺癌基因表达谱的分析有助于认识肿瘤发病机制。     

应用基因芯片技术研究原发性胆汁性肝硬化患者外周血单个核细胞基因表达谱特征

目的 应用基因芯片技术分析原发性胆汁性肝硬化患者外周血单个核细胞的基因表达谱特征.方法 选取9例原发性胆汁性肝硬化患者、9名健康对照为研究对象,提取其外周血单个核细胞总RNA,进行人类全基因组寡核苷酸芯片(约22000个基因)检测,筛选出差异表达基因及相关信号通路.结果 原发性胆汁性肝硬化患者外周血单个核细胞中差异表达的基因共有79个,其中21个表达上调,58个表达下调.这些基因对应着27条信号通路,其中有6条通路参与了免疫调控及细胞凋亡过程,分别是自然杀伤细胞介导的细胞毒通路、Toll样受体信号通路、抗原加工提呈通路、细胞因子相互作用通路、T细胞受体信号通路、细胞凋亡通路.结论 原发性胆汁性肝硬化患者外周血单个核细胞中存在特有的差异表达基因,针对这些基因及相关信号通路的进一步研究,将为探索发病机制和寻找生物标志物提供新的方向.     

Gene expression profiling and prediction of clinical outcome in ovarian cancer

Epithelial ovarian cancer is the most lethal gynaecological cancer. Despite debulking surgery and platinum/taxane-based chemotherapy, the prognosis remains poor with 25% 5-year survival. Current histo-clinical prognostic factors are insufficient to capture the complex cascade of events that drive the heterogeneous clinical behaviour of the disease. There is a crucial need to identify new prognostic subclasses of disease as well as new therapeutic targets. Today, DNA microarrays allow the simultaneous and quantitative analysis of the mRNA expression levels of thousands of genes in a tumour sample. They have been applied to ovarian cancer research for predicting initial surgical resectability, survival and response to first-line chemotherapy. The first results are promising. In this review, we describe recent applications of DNA microarrays in ovarian cancer research and discuss some issues to address in the near future to allow the technology to reach its full potential in clinical practice.