胃癌组织ING1抑癌基因表达及其生物学意义探讨

目的:探讨生长抑制因子1(ING1)基因表达及其与胃癌临床病理特征的关系。方法:荧光定量PCR技术和免疫印迹技术检测70例胃腺癌组织及癌旁对照胃黏膜组织中ING1基因的表达水平,并分析其与胃癌患者临床特征的关系。结果:70例癌组织中有57例(81.43%)ING1基因mRNA表达明显下调,其中4例组织为ING1表达缺失,与癌旁正常胃黏膜比较差异有统计学意义,P<0.01。ING1基因表达与胃癌患者的性别无明显相关性,与胃癌的分化程度、肿瘤累及深度以及淋巴结转移有关。结论:ING1在人胃癌细胞中表达下调,其表达水平与分化程度、肿瘤累及深度以及淋巴结转移有关。     

组织芯片检测p16蛋白在不同胃组织中的表达意义

[目的]分析胃癌相关差异表达蛋白p16在不同胃组织中的表达意义,为临床早期发现胃癌及评估胃癌患者预后提供有价值的资料.[方法]采用免疫组织化学染色,检测胃组织芯片(包括正常胃黏膜、癌旁、非典型增生、胃癌及淋巴结转移癌组织)中p16蛋白的表达,分析其在胃癌组织中表达与临床病理特征的关系.[结果] p16蛋白在正常胃黏膜、癌旁、非典型增生和胃癌组织中的表达率分别为76.47％ (26/34)、79.59％( 39/49)、34.62％ (9/26)和8.64％(7/81);p16蛋白在胃癌组织中阳性表达率较正常胃黏膜、癌旁组织和非典型增生组织降低(P＜0.01),而非典型增生组织中阳性表达率较正常胃黏膜和癌旁组织降低(P＜0.05);p16蛋白表达与胃癌患者年龄、肿块大小和淋巴结转移有关(P＜0.05).[结论] p16蛋白表达与胃癌的发生发展、患者年龄、肿块大小及淋巴结转移有关.     

应用微矩阵表达谱基因芯片筛选食管癌新的相关基因

[目的]应用微矩阵表达谱基因芯片筛选食管癌新的相关基因.[方法]微矩阵表达谱基因芯片(14114种基因)筛选3例食管癌及其对照癌旁组织的差异表达基因,用Northern印迹证实,用末端终止法测序,将测序结果在GenBank数据库进行同源性检索.[结果]检测的3例临床标本中,共有的差异表达基因31条,上调基因10条,下调基因21条,其序列与Genbank数据库比较,从中筛选出2条无明显同源的人类已知基因或片段,即新基因,并列出其序列.[结论]微矩阵表达谱基因芯片可应用于筛选食管癌新的相关基因,其具备高通量、特异性好、快速的特点;食管癌和对照癌旁组织间存在2条新的差异表达基因.     

错配修复基因hMSH6在胃癌中表达特点的研究

目的:了解错配修复基因hMSH6在胃癌中的表达特点.方法:用免疫组织化学SP法检测45例胃癌组织、36例癌旁黏膜和30例正常胃黏膜hMSH6蛋白的表达情况.结果:胃癌组、癌旁组与正常胃黏膜组hMSH6蛋白细胞核表达阳性率分别为24.44%(11/45)、13.89%(5/36)和53.33%(16/30);细胞质表达阳性率分别为66.67%(30/45)、58.33%(21/36)和96.67%(29/30);细胞核和细胞质表达阳性率胃癌组和癌旁组均显著低于正常胃黏膜组(P＜0.05);而胃癌组与癌旁组hMSH6蛋白在细胞核及细胞质中表达差异均无显著性意义(P＞0.05).胃癌组织中,hMSH6蛋白在细胞核中表达与患者性别、年龄,肿瘤分化程度及分期、淋巴结转移均无显著相关性(P＞0.05),但在细胞质的表达与肿瘤分化程度及分期正相关(P＜0.05),而与患者年龄、性别、淋巴结转移无显著相关性(P＞0.05).结论:胃癌及癌旁组织均出现错配修复基因hMSH6表达下调,这可能将成为胃癌发生的预警信号.     

肝癌伴门静脉癌栓的基因表达研究

目的用cDNA芯片研究原发性肝癌(HCC)伴和不伴门静脉癌栓(PVTT)形成的基因表达差异.方法利用含有5075个功能基因cDNA的人表达谱芯片,将原发性肝癌患者分成AB、CD两组,AB组为不伴门静脉癌栓组(n=3),CD组为伴有门静脉癌栓组(n=3),分别提取标本癌、门静脉癌栓和癌旁组织,抽提总mRNA,反转录为cDNA与芯片进行杂交,对比mRNA表达的差异.结果两组相比较AB组的癌组织与CD组的癌及癌栓组织上调表达基因分别有39个和35个不同,两组下调表达基因分别有59个和46个不同.在不重复的两组的上调和下调表达基因中,表达明显上调的基因(ratio＞3)AB组有3个、CD组有5个,而明显下调的(ratio＜0.4)AB组有4个、CD组有10个.结论 cDNA芯片是检测伴和不伴门静脉癌栓肝癌的不同基因表达差异的有效方法.这些不同的基因表达与门静脉癌栓的形成、维持、发展有关.进一步分析这些基因将有助于门静脉癌栓生物学特性的了解.     

胃癌组织及相关淋巴结中端粒酶逆转录酶基因启动子区域甲基化检测的临床意义

[目的]检测胃癌患者肿瘤组织及其相应的癌旁组织和淋巴结组织中端粒酶逆转录酶(hTERT)基因启动子区域甲基化状态,并探讨其甲基化状态的改变与临床病理特征的关系.[方法]运用甲基化特异性PCR(MSP)方法,检测52例手术切除胃癌组织、癌旁组织及相关淋巴结中hTERT基因启动子区域甲基化状念、以同一标本正常组织作为阴性对照.[结果]正常胃黏膜组织未检测出hTERT表达、胃癌组织及癌旁组织、转移淋巴结中均检测出hTERT表达.转移淋巴结、胃癌组织中hTERT基因的甲基化阳性率分别为81.6％(31/38)、71.1％(37/52),明显高于癌旁组织的29.5％(13/52)(P＜0.01).胃癌组织hTERT基因甲基化阳性率与胃癌的临床分期、组织分化程度、肿瘤大小有相关性(P＜0.05).癌旁组织hTERT基因甲基化阳性率和胃癌的临床分期、肿瘤大小、组织分化程度、淋巴结转移具有相关性(P＜0.05).转移淋巴结hTERT基因 甲基化阳性率则与临床及病理特征无关.[结论]胃癌组织及转移淋巴结中存在hTERT基因启动子区域的异常甲基化调控、可能参与了胃癌的发生与发展.     

cDNA微阵列对22例弥漫型胃癌基因的检测

目的从转录组水平识别弥漫型胃癌与正常胃黏膜间的基因表达差异,探讨胃癌分子发生发展机制。方法收集22例弥漫型胃癌患者的新鲜冻存胃癌组织及同例相应正常胃黏膜。杂交芯片采用含14592个点的cDNA表达谱芯片。差异表达基因的筛选标准为该基因在50％以上样本中的肿瘤与正常组织荧光强度比（ratio比值）〉2或〈0．5。采用系统聚类法进行基因表达的相似性分析,标本组间比较采用方差分析。应用实时定量RT—PCR方法对芯片结果进行验证。结果胃癌组织与正常胃黏膜间的差异表达基因共357个,其中表达上调者153个,下调者204个。上调基因功能主要与细胞骨架运动、基质重建、细胞增殖及信号传导等相关;下调基因功能则主要与细胞免疫防御、毒理代谢、功能分化、核一浆转运及凋亡抑制等相关。TNM分期的Ⅰ＋Ⅱ期组与Ⅲ＋Ⅳ期组间,有7个基因的表达差异有统计学意义（P〈0．05）。RT—PCR验证结果与芯片表达结果一致。结论运用cDNA芯片进行弥漫型胃癌基因表达谱分析,有助于从分子水平全方位阐明弥漫型胃癌的发病机制及生物学特性,也有助于进一步发现新的分子诊断指标和基因治疗靶标。     

应用cDNA微阵列基因芯片筛选胃低分化腺癌相关基因的研究

背景与目的：胃低分化腺癌癌变的分子机制至今不清楚,关键是未找到与胃低分化腺癌密切相关的基因.本研究拟建立胃低分化腺癌基因表达谱,筛选差异表达基因,进一步分析差异表达基因与胃癌发生、发展关系.方法：用含10 000个已知基因的cDNA微阵列分析胃低分化腺癌和癌旁正常胃黏膜基因表达谱的变化,免疫组化研究差异表达基因与胃癌的关系.结果：二倍以上的差异表达基因212个,其中在胃低分化腺中表达上调169个,表达下调43个.S-P免疫组化结果显示：EMS1蛋白表达定位于胞质,呈黄色至棕黄色;EMS1蛋白在20例正常胃黏膜阳性表达率为20%（4/20）,在146 例胃癌中阳性表达率为89.72% （131/146）;EMS1蛋白在胃癌中的表达高于正常胃黏膜（P＜0.001）.结论：发现EMS1与胃癌有关,为进一步寻找胃癌相关基因提供了重要的研究线索.     

基因表达谱芯片胃癌差异表达基因的筛选

目的:利用基因表达谱芯片研究胃癌组织中差异表达的基因,从多基因角度研究胃癌发生的分子机制.方法:抽提6例胃癌组织和相应的癌旁组织的总RNA,反转录成cDNA同时进行标记.将标记的cDNA与基因表达谱芯片杂交,经过芯片的扫描和数据处理,分析出胃癌组织和癌旁组织之间差异表达的基因.结果:通过对胃癌组织和癌旁组织的基因表达谱的比较分析,发现在胃癌组织中表达差异＞2倍的基因共有696个,其中表达上调的基因318个,表达下调的基因378个.差异表达的基因主要参与信号转导、免疫反应和细胞运动等生物学过程.结论:胃癌组织与癌旁组织的表达谱存在较大差异,利用基因表达谱芯片可筛选出胃癌差异表达的基因,从而有利于在临床上对肿瘤的诊断和治疗.     

肺腺癌癌旁组织及胚胎肺的基因表达谱研究

目的 利用基因芯片技术研究肺腺癌组织、癌旁组织及胚胎肺组织中的基因表达差异。方法 分别抽取肺腺癌组织、癌旁组织和胚胎肺组织的总RNA,并纯化mRNA。采用逆转录的方法,制成cDNA链,并以两种荧光Cy5和Cy3标记后做探针,与含有1152条人类全长基因芯片进行杂交。以ScanArray 4000荧光扫描仪扫描芯片上两种荧光信号,并用计算机处理和分析。结果 4例肺腺癌组织和癌旁组织标本中,共同表达差异基因25个,其中上调基因3个,下调基因22个;胚胎肺组织和癌旁组织标本中,表达差异基因316个,其中上调基因192个,下调基因124个;胚胎肺组织和癌旁组织中表达差异基因与肺腺癌组织和癌旁组织比较,共同表达差异基因16个,其中上调基因12个,下调基因4个。结论 肺腺癌与癌旁组织共同表达差异的25个基因可能与肺癌的发生、发展有关;胚胎肺组织与癌旁组织表达差异的316个基因与生长发育环境有关;胚胎肺组织和癌旁组织与肺腺癌组织和癌旁组织共表达差异的16个基因可能与早期肺腺癌启动有关。     

用基因芯片筛选胃黏膜癌变相关基因

Objective： To study the difference of gene expression and screen the carcinogenesis associated gene in gastric mucosa by oligonucleotide microarray. Methods： Using the U133A gene chip to detect the gene expression profile difference between pericancerous mucosa （mucosa inside nearly 2 cm by cancer） and normal section of gastric mucosa. Bioinformatics was used to analyze the detected result on their localization and function in chromosome. Results＂ （1） A total of 150 genes with a difference of more than 3 times in expression levels by comparing the pericancerous mucosa with normal gastric mucosa, of 130 genes were up-regulated （SLR〉I.5）, and 20 were down- regulated （SLR〈 -1.5）. From the gene expression difference was to do the function classification, among those 22 enzyme and 6 enzyme regular genes were most one （18.7%）. The next were 17 nucleic acid binding associated genes （11.3%）. The third were 15 signal transduction associated genes （10%）. Fourth, were 13 protein binding associated genes （8.7%）. Besides the 40 genes were unknown their function, above mentioned 4 groups were 48.7% of the gene total number; （2） The pericancerous mucosa （P） and gastric cancer （T） were simultaneously compared with normal gastric mucosa, which had 71 genes with the same expression difference, of 61 genes were up-regulated （pericancerous SLR〉I.5）, and other 10 genes were down-regulated （pericancerous SLR〈 -1.5）. From their localization on the chromosome, there was simultaneously 71 genes appearance both in the pericancerous mucosa and in gastric cancer. The most one was 11 abnormal genes on the No. 19 chromosome. The next was No. 1, 2, 16 and 17 chromosomes which had 6 genes, respectively. It was not finding an abnormal gene on the No. 5, 14, 22 and Y chromosome. Conclusion： It suggested those genes may be related to the promotion in early gastric carcinogenesis and their progress. Four main groups （enzyme and enzyme regular, nucleic acid binding, signal transduction, protein binding） that associated gene＇s abnormality be played an importance role in studying the carcinogenesis of gastric cancer. The No. 19 and No. 1, 2, 16, 17 chromosomes are important sites of the oncogene transformation.     

食管癌和癌旁上皮基因表达谱差异

目的：用基因芯片技术研究食管癌和癌旁组织基因表达谱差异,筛选与早期癌变相关的基因,方法：分别抽提食管癌组织,癌旁组织和正常食管上皮的总RNA并纯化mRNA,将各mRNA逆转录合成以Cy5和Cy3标记的cDNA-一链做探针,分别混合后在2张含有4096条双点人类全长基因的芯片上进行杂交,用扫描仪扫描芯片荧光信号图像,用软件对扫描图像进行数字上处理和分析,结果：食管癌与正常食管上皮比较差异2倍以上共有135个基因,癌旁上皮与正常食管上皮比较差异2倍以上共有31个基因,其中13个基因与食管癌中出现相同,结论：通过三者的基因谱平行比较提示食管癌与正常食管上此比较差异2倍以上的135个基因可能与食管癌的发生和发展有关,癌旁上皮与正常食管上皮比较差异2倍以上的31个基因可能与早期食管癌变的启动和演化有关。     

PI3K/AKT2在胃癌组织表达及其与临床病理特征和生存期的关系

目的探讨PI3K、AKT2在胃癌、癌旁组织中的表达,分析其表达水平与临床病理特征和预后的关系。方法用组织芯片和免疫组织化学法检测189例胃癌组织、54例癌旁组织、32例正常胃黏膜中PI3K、AKT2的表达。结果胃癌、癌旁组织和正常胃黏膜PI3K阳性表达率分别是76.7%、25.9%和25.0%。AKT2阳性表达率分别是75.7%、27.8%和37.5%。胃癌组织PI3K表达与有无淋巴结转移、浸润深度、分化程度和Lauren分型有关（P〈0.05）,与肿瘤大小无关（P〉0.05）。AKT2表达与肿瘤大小、有无淋巴结转移、浸润深度、分化程度、Lauren分型有关（P〈0.05）。单因素生存分析显示,PI3K、AKT2阳性组对胃癌患者生存期的影响有统计学意义（P〈0.05）,Cox多元回归分析显示,AKT2的异常表达可以作为影响胃癌预后的独立因素（P〈0.05）。结论 PI3K/AKT2细胞信号传导通路参与胃癌的发生、发展、浸润转移,是胃癌发生的晚期事件。AKT2的表达可以作为胃癌预后的独立指标。     

胃癌患者腹水及外周血中PTN、IL-16表达及其意义

[目的]探讨胃癌患者腹水及外周血中PTN、IL-16表达水平及其临床意义。[方法]以52名胃癌伴腹水患者、48名胃癌不伴腹水患者以及20名健康者为研究对象,采用ELISA法测定PTN、IL-16的表达水平。[结果]胃癌伴腹水组患者腹水及外周血中PTN、IL-16表达水平均明屁高于胃癌不伴腹水组和健康对照组,差异有统计学意义（P〈0．05）;PTN与IL-16在胃癌患者中表达呈正相关（P〈0．01）。[结论]胃癌患者腹水及外周血中PTN、IL-16表达升高,且具有高度相关性。     

Prognostic impact of detecting viable circulating tumour cells in gastric cancer patients using a telomerase-specific viral agent: a prospective study

ABSTRACT: BACKGROUND: The identification of circulating tumour cells (CTCs) in peripheral blood is a useful approach to estimate prognosis, monitor disease progression, and measure treatment effects in various malignancies. However, clinical relevance of CTCs is controversial. We attempted to detect viable CTCs in the peripheral blood of gastric cancer patients using a telomerase-specific viral agent. We took a 7.5-ml blood sample from 65 treatment-negative gastric cancer patients before surgery and 10 healthy volunteers. We detected viable CTCs in the blood samples after incubating them with a telomerase-specific, replication-selective, oncolytic adenoviral agent carrying the green fluorescent protein (GFP) gene (OBP-401). GFP-positive CTCs were defined as having a diameter of at least 7.735 mum; this threshold was determined by receiver operating characteristic curve analysis. GFP-positive cells were counted under a fluorescence microscope. RESULTS: There was a significant difference in overall survival among the patients with 0[EN DASH]4 and those with [GREATER-THAN OR EQUAL TO]5 GFP-positive CTCs in the stage I[EN DASH]IV disease group and stage II[EN DASH]IV advanced disease group. The number of GFP-positive CTCs was not related to cancer stage. Among the pathological findings, the number of GFP-positive CTCs was only significantly related to venous invasion, although there were trends towards more GFP-positive CTCs with disease progression (tumour depth, lymph node metastasis, distant metastasis, lymphatic invasion, and histological type). CONCLUSIONS: There was a significant relationship between the number of GFP-positive CTCs and overall survival in the patients with gastric cancer. The detection of CTCs using OBP-401 may be useful for prognostic evaluation.Trial registrationUniversity Hospital Medical Information Network in Japan, UMIN000002018.     

CAGE? MAGE-A1和MAGE-A3去甲基化在胃癌发生发展中的作用

探讨肿瘤相关抗原基因（CAGE）、黑色素瘤抗原基因-A1（MAGE-A1）和黑色素瘤抗原基因-A3（MAGE-A3）启动子区去甲基化状态改变在胃癌早期发生、发展中的作用及意义。方法:87例内镜活检标本分为胃癌组、癌前病变组和正常对照组,应用甲基化特异性PCR方法分别检测各组中3种基因启动子区的去甲基化状态。结果:胃癌组、癌前病变组和正常对照组中,各基因启动子区的去甲基化阳性率分别为:CAGE:80.0%（24/30）、37.5%（12/32）和4.0%（1/25）;MAGE-A1:60.0%（18/30）、21.9%（7/32）和0（0/25）;MAGE-A3:46.7%（14/30）、12.5%（4/32）和8.0%（2/25）,呈下降趋势。3种基因启动子区去甲基化阳性率在3组间比较,差异有统计学意义（P<0.05）。30例胃癌至少发生1种基因启动子区去甲基化的有27例;至少发生2种基因启动子区去甲基化23例;7例同时发生了3种基因启动子区的去甲基化。结论:这3种基因启动子区去甲基化改变可能发生于胃癌发病早期,且贯穿于胃癌发生、发展的全过程并起到一定的作用,联合检测可能有助于提高胃癌去甲基化诊断的阳性率。      

Helicobacter pylori infection on the risk of stomach cancer and chronic atrophic gastritis.

Helicobacter pylori infection is associated with gastric adenocarcinoma. However, the mechanisms of this interaction are still unclear. This study was conducted to explore the effects of H. pylori infection on early and late stage gastric carcinogenesis. This study included 134 patients with adenocarcinoma of the stomach (ACS), 67 patients with chronic atrophic gastritis (CAG), and 65 normal controls recruited at Memorial Sloan-Kettering Cancer Center (MSKCC) from November 1, 1992 to November 1, 1994. Epidemiologic data were collected by a modified National Cancer Institute Health Habits History Questionnaire. H. pylori infection was diagnosed by pathological evaluation. Risk factors were analyzed using logistic regression. The odds ratio (OR) associated with H. pylori infection was 10.4 [95% confidence interval (CI): 2.6-41.6] for CAG and 11.2 (95% CI: 2.5-50.3) for gastric cancer in comparison with normal controls, with adjustment for pack-years of smoking, alcohol drinking, body mass index, total caloric intake, dietary fat and fiber intake, and Barrett's esophagus. But H. pylori infection was not associated with risk of stomach cancer when patients with stomach cancer were compared with patients with CAG (OR = 0.6, 95% CI: 0.3-1.3) after controlling for potential confounding variables. This association was persistent when only patients with both gastric cancer and chronic gastritis were considered as cases and patients with CAG were considered as controls (OR = 0.7, 95% CI: 0.3-2.0) in the multivariate analysis. Our results suggest that H. pylori infection may be involved in the early stage of development of CAG, but not in the development of stomach cancer from CAG, and indicate that strategies for prevention of stomach cancer should target the early stage to eliminate H. pylori infection in high-risk populations.     

Promoter hypermethylation of tumor-related genes in gastric intestinal metaplasia of patients with and without gastric cancer

Promoter hypermethylation is an alternative mechanism of gene silencing in human cancers including gastric cancer. While intestinal metaplasia (IM) is generally regarded as a precancerous lesion of the stomach, our study examines the presence of gene promoter hypermethylation in IM of patients with and without gastric cancer. We examined 31 samples of gastric cancer, 36 gastric IM (21 associated with gastric cancer and 15 from noncancer patients) and 10 normal gastric biopsies. Tissues containing foci of IM were carefully microdissected from paraffin-embedded section. Bisulfite-modified DNA was examined for gene promoter hypermethylation in DAP-kinase, E-cadherin, GSTP1, p14, p15, p16, RASSF1A and hMLH1 by methylation-specific-PCR. None of the control gastric tissues had hypermethylation detected, but gene promoter hypermethylation was frequently detected in gastric cancer and IM. The mean number of methylated genes in cancer and IM was 3.0 and 1.4, respectively (p < 0.0001). Methylation in IM from cancer patients was all associated with concurrent methylation in the corresponding tumor samples. The numbers of methylated genes were similar in IM obtained from cancer and noncancer patients. By examining the methylation patterns of these genes, 3 differential methylation patterns were recognized: hypermethylation was more frequent in cancer than in IM (DAP-kinase, p14, p15 and p16); comparable frequencies of methylation in cancer and IM (E-cadherin and hMLH1); and no methylation (GSTP1). Aberrant methylation in tumor-related genes is frequently detected in gastric IM of both cancer and noncancer patients, suggesting their early involvement in the multistep progression of gastric carcinogenesis.