胃癌组织中Kiss-1 mRNA与Notch1 mRNA的表达及意义

目的探讨Kiss-1 mRNA与Notch1 mRNA在胃癌组织中的表达及意义。方法采用实时荧光定量聚合酶链反应检测胃癌和癌旁正常组织中Kiss-1 mRNA与Notch1 mRNA的表达情况,分析其与临床病理特征之间的关系。结果 Kiss-1 mRNA在胃癌组织中的表达较癌旁正常组织中降低(P0.05);Kiss-1 mRNA的表达与胃癌的淋巴结转移、TNM分期密切相关(P0.05),与胃癌的浸润深度、分化程度、远处转移无关(P0.05);Notch1 mRNA在胃癌组中的表达较癌旁正常组织升高(P0.05);Notch1 mRNA的表达与胃癌的淋巴结转移、TNM分期、分化程度密切相关(P0.05),与胃癌的浸润深度、远处转移无关(P0.05)。结论 Kiss-1在胃癌的发病过程中发挥抑癌基因作用,并通过抑制转移来影响胃癌的预后;Notch1在胃癌的发病过程中发挥癌基因作用,有潜力成为早期诊断胃癌的新型肿瘤标记物。     

Survivin基因在胃癌组织中的表达及临床意义

为研究Survivin基因在胃癌组织中的表达及其与胃癌发生、发展的关系,2003年5月至2004年5月,我们应用逆转录聚合酶链反应(RT—PCR)技术检测胃癌组织中Survivin基因的表达情况,并评价其与临床病理指标的关系。现报告如下。     

白血病患者中胰岛素样生长因子结合蛋白相关蛋白1基因的表达

目前关于胰岛素样生长因子结合蛋白相关蛋白1(IGFBP rP1)基因的研究报道主要见于实体瘤[13],尚无与白血病发生、发展的相关报道。我们利用实时定量逆转录酶式聚合反应(QRT PCR)技术,检测白血病患者不同病程中IGFBP rP1基因mRNA的表达水平,并与WT1、RbAp46基因进行相关分析,探讨     

ATP结合盒转运蛋白A1与动脉粥样硬化

对Tangier病病因的研究,发现ATP结合盒转运蛋白A1(ATP binding cassette transport protein A1,ABCA1)在胆固醇逆向转运(reverse cholesterol transport,RCT)中起重要作用。ABCA1主要通过核受体PPARs途径发挥作用。ABCA1基因变异影响其功能。     

ATP结合盒转运蛋白A1研究进展

动脉粥样硬化是心脑血管疾病的重要基础病理改变,而胆固醇在巨噬细胞内聚集和泡沫细胞形成是动脉粥样硬化的始动环节。ATP结合盒转运蛋白Al（ABCAl）是一种重要的胆固醇流出调节蛋白,能介导细胞内胆固醇逆向转运到细胞外,使之与载脂蛋白A-Ⅰ结合并包装形成高密度脂蛋白（HDL）的膜转运蛋白。因此,ABCAl是调节血浆HDL及细胞内胆固醇水平的重要膜蛋白,ABCAl的表达水平与动脉粥样硬化的发生关系密切。同时,细胞内脂质、核受体PPAR、LXR、RXR和细胞因子等对ABCAl蛋白表达具有凋控作用。本文将ABCAl研究进展作一综述。     

实时荧光定量RT-PCR对胃癌组织中hPMS2 mRNA 的检测及其临床意义

目的:探讨DNA错配修复基因hPMS2在胃癌组织中的表达水平及其临床意义.方法:应用实时荧光定量逆转录聚合酶链反应(RT-PCR)技术对41例胃癌患者的癌组织、37例癌旁萎缩性胃炎组织、25例慢性萎缩性胃炎组织及20例慢性浅表性胃炎组织中hPMS2 mRNA进行定量检测,以三磷酸甘油醛脱氢酶基因(hGAPDH)为内参照.结果:胃癌组织、癌旁萎缩性胃炎组织、慢性萎缩性胃炎组织及慢性浅表性胃炎组织中hPMS2 mRNA的含量分别是28.33±14.05,16.88±10.67,7.25±8.72和1.78±1.34,四组相比差异有显著性(F=31.82,P=0.00).胃癌组织中hPMS2 mRNA含量明显高于其他三组;癌旁萎缩性胃炎组织中含量明显高于非胃癌患者的胃炎组织,差异均有显著性(P＜0.01);而非胃癌患者的慢性萎缩性胃炎组织中hPMS2 mRNA的含量与慢性浅表性胃炎组织相比差别无显著性.hPMS2 mRNA含量与肿鬃瘤的大小、浸润深度、有无淋巴结转移关系不明显,差异均无显著性.结论:hPMS2基因的异常转录可能与胃癌的发生有关,而与胃癌的发展关系不大.     

多重耐药蛋白-1在胃癌中的表达及意义

目的:研究胃癌组织中多重耐药蛋白(MRP)-1的表达规律.方法:用免疫组织化学染色(SP法)检测30例慢性浅表性胃炎(CSG)、30例胃黏膜肠上皮化生(IM)、45例异型增生(Dys)和65例胃癌(GC)组织标本中MRP-1阳性表达情况,RTPCR技术检测MRP-1 mRNA的表达情况.均数的比较采用t检验,样本率的比较采用χ2检验,两因素的相关分析用直线相关分析.结果:在CSG→IM→Dys→GC的过程中,MRP-1的表达呈逐步递增趋势,MRP-1的阳性表达率GC组与Dys,IM组,均具有显著性差异(43/65 vs 19/45,6/30,P＜0.05),GC组MRP-1以及MRP-1 mRNA阳性表达率均高于相应癌旁组,差异具有非常显著性(66.2% vs 33.3%,63.1% vs 28.9%,P＜0.01);MRP-1与MRP-1mRNA表达强度呈正相关(r=0.598,P＜0.05).结论:胃癌形成过程中,MRP-1与MRP-1mRNA表达逐渐上调.     

RADIL基因在胰腺癌组织中的表达及其临床意义

目的 探讨胰腺癌组织中RADIL基因的表达及其与临床病理特征之间的相关性.方法 采用实时定量PCR方法检测40例胰腺癌及相应癌旁组织和5例正常胰腺组织中RADIL mRNA相对表达量,分析RADIL mRNA表达与胰腺癌临床病理特征之间的相关性.结果 RADIL mRNA在胰腺癌组织、癌旁组织及正常胰腺组织中均有表达,相对表达量分别为2.263±3.826、5.425±8.858和8.559±4.214,三组之间存在显著差异(P＜0.05).癌组织中RADIL mRNA高表达与肿瘤远处转移、分化程度呈负相关(r为-0.312和-0.294,P值均＜0.05),与肿瘤部位、肿瘤大小,血清CA19-9浓度、TNM分期及患者的性别和年龄均无显著相关性.结论 RADIL基因对胰腺癌可能具有一定的抑制作用.     

细胞毒素相关蛋白基因A与胃粘膜组织恶变的研究

目的:研究细胞毒素相关蛋白基因A(cagA)阳性幽门螺杆菌(Hp)在胃粘膜组织恶性化转变中的作用。方法:以含Hp细胞毒素相关蛋白基因A的质粒(P~(MC3))为外参照品,在微孔板上对PCR扩增cagA的产物作探针杂交及辣根过氧化物酶酶促显色分析,定量榆测不同组织中cagA阳性Hp。结果:往50份Hp尿素酶B基因阳性的临床标本中检测到28份cagA亦阳性(56％),其中5份浅表性胃炎组织,12份癌前病变组织及11份胃癌组织。浅表性胃炎组织cagA阳性的Hp感染率(31％)及每毫克组织感染的平均拷贝数(12.8)均显著低于(P<0.01或P<0.05)癌前病变组织(60％;77.1)或胃癌组织(79％;57.6),二者在后两种组织间均无显著性差别(P>O.05)。结论:cagA阳性的Hp感染可能有助于胃粘膜组织的恶性化转变,这种转变与cagA阳性Hp的感染量有关。     

RASSF1A基因启动子区甲基化在胃癌组织中的检测

目的:检测胃癌组织中RASSF1A基因启动子区甲基化状况,并探讨其与临床病理特征的关系.方法:采用甲基化特异性聚合酶链反应(methylation-specific PCIL MSP)检测39例胃癌组织,及相应癌旁组织和30例对照组织中RASSF1A基因启动子区甲基化水平.结果:胃癌组织中RASSF1A基因启动子区甲基化率显著高于癌旁组织甲基化率及对照组(64.1%vs 7.7%,0%,均P＜0.01).胃癌组织中不同年龄、性别、分化程度及淋巴结转移与否的RASSF1A基因甲基化率的差异均无统计学意义.结论:RASSF1A基因启动子区甲基化与胃癌的发生密切相关,MSP法是一种快速、敏感的基因甲基化检测方法,可用于胃癌的辅助诊断.     

Screening and identification of mimotope of gastric cancer associated antigen MGb1-Ag

AIM: Using a monoclonal antibody against gastric cancer antigen named MGb1 to screen a phage-displayed random peptide library fused with coat protein pⅢ in order to get some information on mimotopes.METHODS: Through affinity enrichment and ELISA screening,positive clones of phages were amplified. 10 phage clones were selected after three rounds of biopanning and the ability of specific binding of the positive phage clones to MGb1-Ab were detected by ELISA assay (DNA sequencing was performed and the amino acid sequences were deduced)By blocking test, specificity of the mimic phage epitopes was identified.RESULTS: There were approximately 200 times ofenrichment about the titer of bound phages after three rounds of biopanning procedures. DNA of 10 phage clones after the third biopanning was assayed and the result showed that the positive clones had a specific binding activity to MGb1-Ab and a weak ability of binding to control mAb or to mouse IgG. DNA sequencing of 10 phage clones was performed and the amino acid sequences were deduced.According to the homology of the amino acid sequences of the displayed peptides, most of the phage clones had motifs of H(x)Q or L(x)S. And these 10 phage clones could also partly inhibit the binding of MGb1-Ab to gastric cancer cell KATO-Ⅲ. The percentage of blocking was from (21.0±1.6)%to (39.0±2.7)%.CONCLUSION: Motifs of H(x)Q and L(x)S selected and identified show a high homology in the mimic epitopes of gastric cancer associated antigen. There may be one or more clones which can act as candidates of tumor vaccines.     

Screening and identification of mimotope of gastric cancer associated antigen MGb1-Ag

AIM: Using a monoclonal antibody against gastric cancer antigen named MGbl to screen a phage-displayed random peptide library fused with coat protein plII in order to get some information on mimotopes.lV~37BODS: Through affinity enrichment and EUSA screening,positive clones of phages were amplified. 10 phage clones were selected after three rounds of biopanning and the ability of specific binding of the positive phage clones to MGb1-Ab were detected by ELISA assay (DNA sequencing was performed and the amino acid sequences were deduced)By blocking test, specificity of the mimic phage epitopes was identified.RESULTS: There were approximately 200 times of enrichment about the titer of bound phages after three rounds of biopanning procedures. DNA of 10 phage clones after the third biopanning was assayed and the result showed that the positive clones had a specific binding activity to MGbl-Ab and a weak ability of binding to control mAb or to mouse IgG. DNA sequencing of 10 phage clones was performed and the amino acid sequences were deduced.According to the homology of the amino acid sequences of the displayed peptides, most of the phage clones had motifs of H(x)Q or L(x)S. And these 10 phage clones could also partly inhibit the binding of MGbl-Ab to gastric cancer cell KATO-Ⅲ. The percentage of blocking was from (21.0&#177;1.6) %to (39.0&#177;2.7) %.CONCLUSION: Motifs of H(x)Q and L(x)S selected and identified show a high homology in the mimic epitopes of gastric cancer associated antigen. There may be one or more clones which can act as candidates of tumor vaccines.     

应用基因表达谱芯片筛选胃腺癌相关基因

目的:利用基因芯片技术筛选胃腺癌组织和癌旁正常组织间的差异表达基因．方法:分别抽取胃腺癌组织和癌旁正常组织的总RNA．采用逆转录的方法,制成cDNA链, 并以两种荧光Cy5和Cy3标记后作为探针,与含有14 784条人类14KcDNA基因表达谱芯片进行杂交．以Agilent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:在14 784条基因中,4例胃腺癌组织和癌旁正常组织共同差异表达基因29条,其中上调基因10条,下调基因19条,上调的基因中有2 条功能信息不明．结论:胃腺癌发生过程中有多基因的参与,胃腺癌与癌旁正常组织共同差异表达的29条基因可能与胃癌的发生、发展有关．     

Subcellular distribution of small GTP binding proteins in pancreas: identification of small GTP binding proteins in the rough endoplasmic reticulum.

Subfractionation of a canine pancreatic homogenate was performed by several differential centrifugation steps, which gave rise to fractions with distinct marker profiles. Specific binding of guanosine 5'-[gamma-[35S]thio]triphosphate (GTP[gamma-35S]) was assayed in each fraction. Enrichment of GTP[gamma-35S] binding was greatest in the interfacial "smooth" microsomal fraction, expected to contain Golgi and other smooth vesicles. There was also marked enrichment in the rough microsomal fraction. Electron microscopy and marker protein analysis revealed the rough microsomes (RMs) to be highly purified rough endoplasmic reticulum (RER). The distribution of small (low molecular weight) GTP binding proteins was examined by a [alpha-32P]GTP blot-overlay assay. Several apparent GTP binding proteins of molecular masses 22-25 kDa were detected in various subcellular fractions. In particular, at least two such proteins were found in the Golgi-enriched and RM fractions, suggesting that these small GTP binding proteins were localized to the Golgi and RER. To more precisely localize these proteins to the RER, native RMs and RMs stripped of ribosomes by puromycin/high salt were subjected to isopycnic centrifugation. The total GTP[gamma-35S] binding, as well as the small GTP binding proteins detected by the [alpha-32P]GTP blot overlay, distributed into fractions of high sucrose density, as did the RER marker ribophorin I. Consistent with a RER localization, when the RMs were stripped of ribosomes and subjected to isopycnic centrifugation, the total GTP[gamma-35S] binding and the small GTP binding proteins detected in the blot-overlay assay shifted to fractions of lighter sucrose density along with the RER marker.     

人类抗凋亡基因Survivin的克隆及其在胃癌细胞中的表达定位

目的 从人胃癌细胞中克隆Survivin(SVV)基因，研究其在胃癌细胞中的表达定位。方法 RT-PCR技术自人胃癌细胞SGC7901中克隆Survivin基因。建立真核表达载体pEGFP-N1-SVV。将含SVV序列的阳性重组子克隆瞬时转染人胃癌细胞SGC7901、激光共聚焦技术检测SVV基因转染胃癌细胞后的细胞内定位。结果 获得人SVV基因的全长cDNA片段，经序列测定、核苷酸序列比对，证实所获片段与SVV基因已知序列完全一致。含SVV基因序列的阳性重组子瞬时转染人胃癌细胞SGC7901后，激光共聚焦显示，SVV基因转染成功，并且表明，SVV基因在胃癌细胞的细胞浆中表达。结论 SVV基因在胃癌细胞的细胞浆中表达。     

胃癌相关基因及其蛋白分子的研究进展

近十年来的大量研究已证实胃癌的发生、发展和预后是多基因改变的结果。胃癌可分成两种临床病理型 :肠型 (或良好分化型 ,即腺管型 )及弥漫型 (或差分化型 ,即无腺管形成的腺癌及印戒细胞癌和硬癌 )。这两种类型的病因、好发部位、发病率、生物学行为及分子改变均明显不同。现已证实基因突变及相关蛋白分子的改变在肿瘤细胞粘附、信号传递、识别、生长及ＤＮＡ修复中起着重要的作用。研究胃癌相关基因及其蛋白分子不仅对于理解胃癌的发病机制十分重要 ,而且对于发现在诊断和评价预后中有临床意义的分子标记物也同样重要。本文就近年来在胃癌…     

丙型肝炎病毒非结构基因5A反式激活基因4A结合蛋白基因的筛选

目的 筛选并克隆HCV非结构基因NS5A反式激活基因4剪切体NS5ATP4A的结合蛋白基因,为研究NS5ATP4A的生物学功能提供线索. 方法 构建HCV NS5ATP4A蛋白诱饵酵母质粒,转化酵母AH109后与含文库质粒的酵母Y187进行配合,在营养缺陷培养基上进行双杂交筛选.选择既能在4重营养缺陷培养基(SD/-Trp/-Leu/-Ade/-His)上生长,又能在涂有X-α-半乳糖的四缺培养平皿上生长的蓝色菌落,提取此酵母克隆的质粒,转化大肠杆菌后进行测序,并进行生物信息学分析.结果 筛选出7个基因,其中已知功能基因6个,未知功能基因1个,这些基因与RNA合成、蛋白质翻译、细胞周期及肿瘤免疫有关. 结论 HCV NS5ATP4A结合蛋白基因的成功筛选,提示了HCV NS5ATP4A新的信号转导途径,为HCV致病机制的进一步研究提供了依据.     

人髓系细胞触发受体-1模拟多肽的筛选和鉴定

目的寻找能特异性与人髓系细胞触发受体-1（TREM-1）蛋白结合并抑制其传导通路的多肽。方法克隆、表达和纯化TREM-1功能区蛋白，并以之为诱饵蛋白，筛选随机噬菌体展示肽库。经过4轮生物淘洗，通过ELISA方法和单核细胞ELISA方法分析噬菌体克隆与TREM-1蛋白的亲和力。化学合成模拟多肽，检测其对盲肠结扎穿刺（CLP）小鼠的治疗效果。结果成功找到5种噬菌体克隆，ELISA法和细胞ELISA均阳性。应用化学方法合成的模拟多肽HYGMTHPNTMsH能降低CLP小鼠的死亡率。结论模拟多肽HYGMTHPNTMSH对脓毒症小鼠有保护作用。     

Atrogin-1, a muscle-specific F-box protein highly expressed during muscle atrophy

Muscle wasting is a debilitating consequence of fasting, inactivity, cancer, and other systemic diseases that results primarily from accelerated protein degradation by the ubiquitin-proteasome pathway. To identify key factors in this process, we have used cDNA microarrays to compare normal and atrophying muscles and found a unique gene fragment that is induced more than ninefold in muscles of fasted mice. We cloned this gene, which is expressed specifically in striated muscles. Because this mRNA also markedly increases in muscles atrophying because of diabetes, cancer, and renal failure, we named it atrogin-1. It contains a functional F-box domain that binds to Skp1 and thereby to Roc1 and Cul1, the other components of SCF-type Ub-protein ligases (E3s), as well as a nuclear localization sequence and PDZ-binding domain. On fasting, atrogin-1 mRNA levels increase specifically in skeletal muscle and before atrophy occurs. Atrogin-1 is one of the few examples of an F-box protein or Ub-protein ligase (E3) expressed in a tissue-specific manner and appears to be a critical component in the enhanced proteolysis leading to muscle atrophy in diverse diseases.     

胃癌及癌前病变粘膜中呈高表达6个基因片段的筛选及其临床意义

目的　筛选数个在胃癌及癌前病变粘膜中检出率高的差异表达基因片段并探讨其临床意义．方法　以筛选出的ｇｃｙｓ１ 至ｇｃｙｓ１８ 的新基因片段作为研究对象,采用定量ＲＴＰＣＲ 技术检测４８ 例胃癌、１６ 例其他肿瘤及１１０ 例活检的胃粘膜标本．结果　①在４８ 例胃癌中,表达检出率高的６ 个片段依次为：ｇｃｙｓ１０ 为９０－５ ％ ,ｇｃｙｓ１ 为８０－９ ％ ,ｇｃｙｓ１８ 为６９－５ ％ ,ｇｃｙｓ１１为６１－９ ％ ,ｇｃｙｓ４ 为４２－８ ％ ,ｇｃｙｓ８ 为３５－７ ％ ,在癌旁组织中检出率＜ １０ ％ ;６ 个片段的检出率在胃癌及癌旁组织中存在显著性差异（ Ｐ ＜ ０－０１） ;②在１６ 例其他肿瘤中, 表达率依次为：ｇｃｙｓ１０ 为３７－５ ％ ,ｇｃｙｓ１ 为１４－３ ％ ,ｇｃｙｓ１８ 为２７－５ ％ ,ｇｃｙｓ１１为４３－７ ％ ,ｇｃｙｓ４ 为１４－３ ％ ,ｇｃｙｓ８ 为３１－３ ％ ;③３３ 例萎缩性胃炎中,表达率依次为：ｇｃｙｓ１０ 是４０－５ ％ ,ｇｃｙｓ１ 是１６－７ ％ ,ｇｃｙｓ１８ 为４４－５ ％ ,ｇｃｙｓ１１ 为４３－３ ％ ,ｇｃｙｓ４ 为３３－３ ％ ,ｇｃｙｓ８为４８－３ ％ ;２８ 例肠腺化生