端粒酶活性的原位检测在大肠癌早期诊断中的意义

目的：研究端粒酶活性的原位检测在大肠癌早期诊断中的意义。方法：运用原位杂交技术检测30例大肠癌组织、癌旁组织及其正常黏膜中端粒酶mRNA的表达情况。结果：在30例大肠癌中,端粒酶mRNA的阳性表达率为90.0%,明显高于癌旁组织、正常黏膜组织（P〈0.005）。端粒酶活性的表达强度与肿瘤的浸润深度、临床分期以及淋巴结转移显著相关（P〈0.05）,与性别、年龄以及肿瘤的分化程度无相关性。结论：端粒酶的激活在大肠癌的发生、发展中起到一定的作用,可以将端粒酶作为一种新的肿瘤标记,通过对细胞端粒酶活性的原位检测来协助大肠癌的早期诊断。     

端粒酶活性的原位检测在大肠癌早期诊断中的意义

目的:研究端粒酶活性的原位检测在大肠癌早期诊断中的意义。方法:运用原位杂交技术检测30例大肠癌组织、癌旁组织及其正常黏膜中端粒酶mRNA的表达情况。结果:在30例大肠癌中,端粒酶mRNA的阳性表达率为90.0%,明显高于癌旁组织、正常黏膜组织(P<0.005)。端粒酶活性的表达强度与肿瘤的浸润深度、临床分期以及淋巴结转移显著相关(P<0.05),与性别、年龄以及肿瘤的分化程度无相关性。结论:端粒酶的激活在大肠癌的发生、发展中起到一定的作用,可以将端粒酶作为一种新的肿瘤标记,通过对细胞端粒酶活性的原位检测来协助大肠癌的早期诊断。     

微阵列技术用于检测乳腺癌人表皮生长因子受体2基因状态

目的探讨细胞核微阵列技术及组织微阵列技术用于检测乳腺癌组织中HER-2基因扩增和蛋白表达状态。方法将248例乳腺癌普通组织蜡块制成组织微阵列组,应用免疫组织化学法（IHC）和荧光原位杂交法（FISH）分别检测HER-2基因和蛋白表达。两种检测方法的-致性采用检验,并以FISH检测结果为金标准,绘制IHC检测乳腺癌HER-2表达的ROC曲线图。结果248例乳腺癌FISH与IHC检测结果-致性分析显示：Kappa=0．711,P=0．000,两种检测方法的-致性尚可。IHC检测乳腺癌HER-2ROC曲线下面积为0．888,P=0．000,约登指数为0．700,准确率为87．9％。IHC（＋＋）中4例为17号染色体单体型,占FISH阳性病例总数的5．26％（4／76）。IHC（＋＋＋）中5例为17号染色体多倍体型,FISH检测均为阴性,占FISH阴性总例数的2．9％（5／172）。IHC的检测可以较好地反映出FISH的结果。结论细胞核微阵列及组织微阵列技术用于检测乳腺癌HER．2基因状态有着节约实验成本、同一性好、结果可靠的优点。     

p16基因mRNA及其编码蛋白在人乳腺癌中的表达

目的：研究人乳腺癌中ｐ１６基因ｍＲＮＡ及其编码蛋白的表达水平与肿瘤发生中预后的相关性。方法：采用免疫组化和原位杂交方法分别对１１例乳腺良性肿瘤和５９例乳腺癌中的ｐ１６基因及其编码蛋白进行检测。结果：ｐ１６基因蛋白表达水平与乳腺肿瘤的良、恶性显著相关（Ｐ〈０．０５）;ｐ１６基因ｍＲＮＡ及其蛋白表达水平与乳腺癌的腋窝淋巴结转移呈现显著负相关（Ｐ〈０．０５）,而与乳腺癌的术后自下而上期呈显著正相关（Ｐ〈     

The role of tissue microarray in the era of target-based agents

Tissue microarray (TMA) technologies have been developed over the last years, mainly to identify biomarkers useful for the correct identification and characterization of tumors. Moreover, TMA has been implemented in retrospective studies in order to identify predictive biomarkers of response to a given therapy and/or to find potential new targets for biological therapy. We analyzed the fields of application of TMA technology and the design of TMA varying according to the objectives to be studied. In this article, the reader will learn how to design TMAs in order to cover the objectives of clinical trials based upon the use of target-based agents. The main limits and advantages of TMA and the results achieved in cancer diagnosis will be also described. Tissue microarray technology should be systematically applied to define critical markers, in retrospective studies and in the screening of most human tumors in order to find new possible molecular targets and to molecularly define the diagnosis of the neoplastic diseases. TMAs have substantially improved the field of translational studies, even in the design and follow-up of studies based upon the use of target-based agents in cancer therapy.     

食管癌组织中HER-2/neu蛋白表达和基因扩增的检测及其临床意义

目的:检测食管癌组织中HER-2/neu蛋白表达及其基因扩增情况,并探讨其与食管癌患者临床病理指标的关系。方法:回顾性分析2000～2006年武汉大学人民医院收治的167例食管癌患者的临床资料。采用免疫组织化学和荧光原位杂交方法检测167例食管癌患者癌组织中HER-2/neu蛋白表达及基因扩增情况。结果:食管癌组织中HER-2/neu蛋白表达水平与基因扩增存在一定相关性(P<0.05),HER-2/neu蛋白的表达与食管癌分化程度和临床分期有关系(P<0.05),而HER-2/neu基因扩增仅与食管癌临床分期有关系(p<0.05)。HER-2/neu蛋白过表达和基因扩增阳性均可缩短食管癌患者的生存期(P<0.05)。结论:HER-2/neu蛋白过表达及基因扩增可能是食管癌患者的一个预后指标,联合检测HER-2/neu蛋白表达水平及基因扩增程度对于食管癌的靶向治疗可能具有一定指导意义。     

宫颈癌组织中癌基因c-myc与抑癌基因p16的对比分析

背景与目的：原癌基因的激活和抑癌基因的失活是许多肿瘤发生的机制;c-mvc与p16的异常表达是否共同参与宫颈癌的发生、发展,以及两种基因的表达与宫颈癌细胞对化疗药物反应的相关性均未明确。本研究旨在探讨c-myc与p16在宫颈癌发生、发展中的作用及其与化疗的关系。方法：采用原位分子杂交技术,用地高辛标记探针检测37例宫颈鳞癌组织(其中11例为化疗后组织)、21例癌前病变组织及5例正常宫颈组织中c-myc mRNA与p16mRNA的表达情况,并用图像分析仪对阳性信号的灰度值作定量分析。结果：正常宫颈组织、宫颈上皮内瘤样病变(cerlvical intraepithelial neoplasia,CIN)组织、宫颈鳞癌组织中p16的表达率分别为100．0％、71．4％及21．6％(P=0．0001),c-myc的表达率分别为0％、42．9％、75．7％(P=0．0011),p16及c-myc的表达在三组问差异均有统计学意义,c-myc表达强度随恶性程度的增高而增强,且CINⅢ级比CINⅡ级和CINⅠ级表达强,化疗前c-mvc表达率比化疗后高,c-myc与p16的表达呈负相关(rs=-0．907)。对化疗敏感者与不敏感者的c-myc和p16表达均无显著性差异。结论：癌基因c-mvc的过表达与抑癌基因p16的失表达在宫颈癌的发生、发展过程中可能起着重要的作用;c-myc在不同级别CIN增生细胞中的表达变化趋势提示细胞癌变是一个渐进的过程。     

Endoluminal tumor implant of a colorectal cancer in an anal fistula detected by FISH techniques: a case report

Intraluminal shedding of tumor cells is a rare infrequent sporadic colorectal cancer (sCRC) mechanism of spreading. Less than 30 cases of sCRC metastasis into anal fistula have been reported. Here, we study a 72-year-old male with an adenocarcinoma arising in an anal fistula. Subsequent studies revealed another tumor in the rectum without distant metastatic disease; therefore, a curative-intent abdominoperineal resection was performed. The histologic study showed a moderately differentiated adenocarcinoma in both locations. No perineural or lymphovascular invasion was observed, and all the lymphatic nodes resected were negative for malignancy. Both tumors showed positive CK20 and negative CK7 immunostaining, but KRAS G12D mutation was only detected in the rectal tumor. After those conventional studies, a cytogenetic profile of both tumors was performed by interphase fluorescence in situ hybridization (iFISH) techniques. The FISH study displayed an identical genetic profile in both tumors, loss of the chromosomes 8 and 18q, and no alteration in chromosome 7 and 13q. Based on pathological and genetic findings, we established the same clonal origin of both tumors. Currently, the diagnosis of an intraluminal CRC metastasis relies on histologic and immunohistochemistry findings. We suggest that genetic studies at the individual cell level by FISH techniques may be useful in order to differentiate synchronous from intraluminal metastasis.     

大肠癌组织中Survivin和p73的表达及其意义

目的：探讨大肠癌组织中Survivin和p73的表达特点及其相互关系。方法：运用S—P免疫组织化学法染色结合图像分析,探讨Survivin和p73在大肠癌组织、癌旁组织及正常组织中的表达特点。结果：在60例大肠癌的癌、癌旁、正常组织标本中,Survivin表达的平均光密度值分别为：0．5443±0．0385、0．3263±0．0284、0．1467±0．0227;癌组织中Survivin的表达水平显著升高,与癌旁组织、正常组织中的表达比较存在显著差异（P〈0．01）。p73表达的平均光密度值分别为：0．7617±0．0251、0．5327±0．0318、0．3460±0．0223;癌组织中p73蛋白的表达水平显著升高,与癌旁组织、正常组织中的表达比较存在显著差异（P〈0．01）Survivin和p73在大肠癌组织中的表达存在依从关系（P〈0．01）,二者呈直线相关,相关系数为0．783。结论：Survivin和p73在大肠癌组织中高表达,与癌旁、正常组织中的表达比较有显著性差异,提示二者在大肠癌的发生发展中起一定的作用,是细胞恶变的指标;Survivin和p73在大肠癌的发生发展中具有协同作用,检测大肠癌患者Survivin和p73对控制大肠癌进展具有重要临床意义。     

High-throughput tissue microarray analysis of CMYC amplificationin urinary bladder cancer

Alterations of chromosome 8, preferentially deletions of 8p and gains of 8q, belong to the most frequent cytogenetic changes in bladder cancer. CMYC on 8q24 is a candidate oncogene in this region. Little is known about the clinical significance of CMYC copy number changes in urinary bladder cancer because its frequency is low and a limited numbers of tumors were analyzed so far. To investigate the impact of CMYC alterations on tumor progression and patient prognosis in bladder cancer, we applied FISH to a tissue microarray containing 2317 bladder cancer samples. Presence of CMYC copy number increase was associated with advanced stage and high grade. CMYC amplifications were seen in 3 of 467 pTa (0.6%), 10 of 247 pT1 (4%) and 11 of 201 pT2-4 urothelial carcinomas (5.5%; p < 0.0001), as well as in 1 of 123 G1 (0.8%), 8 of 470 G2 (1.7%) and 17 of 365 G3 urothelial carcinomas (4.7%; p < 0.0001). CMYC gains were present in 49 of 467 pTa (10.5%), 39 of 247 pT1 (15.8%) and 43 of 201 pT2-4 urothelial carcinoma (21.4%; p < 0.0001), as well as in 7 of 123 G1 (5.7%), 56 of 470 G2 (11.9%) and 72 of 365 G3 urothelial carcinomas (19.7%; p < 0.0001). CMYC copy number changes were unrelated to prognosis of bladder cancer patients. We conclude that alterations of the CMYC gene, including copy number gains and amplifications, are linked to genetically unstable bladder cancers that are characterized by a high histologic grade and/or invasive growth. Patient prognosis was not affected by CMYC gene copy number changes.     

HPV16/18DNA和p53、RB、MCM7蛋白在乳腺癌中的表达及意义

目的:探讨HPV16/18DNA和p53、RB、MCM7蛋白在乳腺癌中的表达及意义.方法:用原位杂交和免疫组织化学EnVision法检测30例正常乳腺组织、30例乳腺腺病、30例乳腺不典型导管上皮增生、20例乳腺导管原位癌和68例乳腺癌组织标本中HPV16/18DNA和p53、RB、MCM7蛋白的表达.结果:乳腺癌组织中HPV16/18DNA的阳性率为58.8％,明显高于正常乳腺、乳腺腺病和乳腺不典型导管上皮增生组,差异有统计学意义(P＜0.05).在不典型导管上皮增生、导管原位癌和乳腺癌组织中HPV16/18DNA与p53和RB蛋白的表达均呈负相关(P＜0.05),而癌组中HPV 16/18DNA与MCM7蛋白表达呈正相关(r=0.750,P＜0.05).癌组中组织学G2、G3级,pTNM Ⅱ、Ⅲ期以及有淋巴结转移组中HPV16/18DNA和MCM7蛋白的阳性表达率均显著高于组织学G1级和pTNM Ⅰ期以及无淋巴结转移组(P＜0.05);与之相反,p53和RB蛋白的阳性表达率则显著降低(P＜0.05).结论:HPV16/18感染通过对p53、RB和MCM7蛋白表达调控的影响可能在高危型HPV感染后乳腺癌的发生、发展及转移进程中起到重要的作用.通过联合检测它们在乳腺癌及癌前病变中的表达以评价临床疾病进展和肿瘤生物学行为.     

Ki-67、p53及HER-2在结直肠癌组织中表达的临床意义

目的:了解Ki-67、p53及HER-2在结直癌组织中表达的临床意义.方法:采用免疫组化法,观察Ki-67、p53、HER-2在39例结直肠癌组织中的表达特点,并与Dukes分期、组织分化等临床病理特征进行研究探讨.结果:Ki-67、p53及HER-2在结直肠癌组织中的阳性表达率分别为82.1％、56.4％及10.3％;在结直肠癌中,Ki-67在年龄大小和有无淋巴结转移表达中有显著差异(P＜0.05);p53和HER-2在患者性别、年龄、部位、分化程度、Dukes分期、淋巴结转移表达无明显差异(P＞0.05);结直肠腺癌组织中Ki-67与p53的表达呈正相关.结论:联合检测Ki-67和p53在结直肠腺癌中的表达有利于对结直肠腺癌恶性程度及患者预后进评估;而HER-2基因在结直肠肿瘤的表达及相关性治疗,仍需要进一步研究、探讨及验证.     

Nuclear thymidylate synthase expression, p53 expression and 5FU response in colorectal carcinoma.

Thymidylate synthase (TS) is a key enzyme in DNA synthesis and is inhibited by metabolites of the chemotherapeutic agent 5-fluorouracil (5FU). Nuclear expression of TS in human tissue in vivo has not been characterised and its clinicopathological correlates in malignancy are unknown. 52 cases of primary colorectal carcinoma (CRC) and 24 cases of matched metastatic carcinoma were studied immunohistochemically using the monoclonal antibody TS106. The degree of nuclear TS immunostaining correlated closely with levels of TS mRNA expression amongst 10 CRCs studied. Strong nuclear immunostaining was seen in normal basal crypt colonocytes and germinal centre cells, and in a varying proportion of adenocarcinoma cells. Amongst the primary carcinomas, higher TS nuclear expression was associated with prominent extracellular mucin production and right-sided location. Higher TS nuclear expression also showed a significant association with poorer response to protracted venous infusional 5FU therapy. There was no clear association between TS nuclear expression and Ki67 or p53 expression assessed immunohistochemically. There was a strong positive correlation between TS nuclear expression in primary and metastatic CRC but the latter generally showed higher expression than matched primary tumour tissue. These findings confirm the nuclear expression of TS protein in human cells in vivo and provide new insight into how such expression may relate to the behaviour of CRCs.     

胃癌组织芯片快速制备及RegIV编码产物检测

目的：建立快速胃癌组织芯片（tissuemicroarray,TMA）制作方法,探讨其在RegⅣ编码产物检测和研究中的应用。方法：收集日本富山大学医学部1993-04-20-2010-11-29胃癌石蜡标本372例和相应癌旁黏膜标本（距离癌组织〉4cm）198例,应用AZUMAYA芯片机构建48阵列组织芯片,利用免疫组化和原位杂交检测RegIV蛋白和mRNA的原位表达。在OlympusBX41显微镜下选定切取部位,利用2mm的穿刺针切除供体蜡块,不用制备受体蜡块,直接将切除组织放入金属包埋皿的固定盒上制备48阵列芯片。结果：组织芯片经HE染色确定了组织病理学诊断,免疫组化和原位杂交均发现在肠上皮化生和黏液分泌性肿瘤细胞质中存在RegⅣ蛋白和tuRNA表达。胃黏膜肠化生中RegⅣ蛋白表达为100％（63／63）,高于胃炎的29．0％（27／93,χ^2=73．67,P〈0．001）、腺瘤的85．7％（36／42,χ^2=9．54,P〈0．005）和胃癌的45．2％（168／372,χ^2=65．06,P〈0．001）。胃癌组织中,印戒细胞癌阳性率为95．3％（41／43）,高于低分化癌的27．4％（32／117）,χ^2=55．89,P〈0．001。原位杂交发现,癌旁黏膜肠化生上皮细胞中RegIVmRNA表达阳性率为95．2％（40／42）,高于癌旁正常黏膜的38．6％（17／44,χ^2=30．63,P〈0．001）和胃癌的20．3％（14／69,X2=55．74,P〈0．001）。结论：制备TMA不需要制备受体蜡块,提高了TMA制作速度。异常RegⅣ表达与胃黏膜肠化生球样异型增生-印戒细胞癌的发生途径关系密切。     

Deletions of the region 17p11-13 in advanced melanoma revealed by cytogenetic analysis and fluorescence in situ hybridization.

The significance of the p53 tumour-suppressor gene in the oncogenesis of a variety of malignant tumours has been demonstrated over recent years. However, the role of p53 in human malignant melanoma is still unclear. Therefore, we investigated melanoma metastases from 11 patients cytogenetically and with fluorescence in situ hybridization (FISH) after short-term culture, employing a p53 region-specific probe for 17p13.1 and a probe detecting the centromere of chromosome 17. Furthermore, paraffin-embedded tissue samples from nine of these patients were investigated immunohistochemically for expression of the p53 protein. Deletions of the short arm of chromosome 17 were seen in six melanomas in cytogenetic analysis. With FISH, three malignant melanomas had clones with only one p53-allele and an additional four malignant melanomas showed a reduced number of signals at the p53 tumour-suppressor gene locus compared with signals for the centromeric region of chromosome 17. This was confirmed by immunohistochemistry. Our results suggest that the 17p11-13 region is frequently deleted in malignant melanomas and that p53 or other genes located on this band might contribute to the malignant potential of advanced melanoma.     

血管免疫母细胞性T细胞淋巴瘤临床-病理-分子遗传学改变与预后相关性分析

目的 血管免疫母细胞性T细胞淋巴瘤(angioimmunoblatic T-cell lymphoma,AITL)是一种侵袭性淋巴瘤,本研究探讨AITL的临床病理特征、与EB病毒相关性及分子遗传学改变,并分析相关预后因素.方法 选取2003-01-03-2012-12-31新疆医科大学第一附属医院有详细临床病理资料的16例AITL患者石蜡样本,EnVision法进行CXCL13、PD-1、CD10和Bcl-6免疫标记;原位杂交技术检测肿瘤EB病毒编码mRNA表达情况;荧光原位杂交技术检测肿瘤组织中Bcl-2、Bcl-6和C-MYC基因异常情况,并对所有患者进行随访.结果 16例患者,男11例(68.8％),女5例(31.2％),男女比例2.2∶1.中位年龄65.5岁.75.0％(12/16)患者国际预后指标(international prognostic indicators,IPI)评分为中高危,56.3％(9/16)患者血清乳酸脱氢酶(lactic dehydrogenase,LDH)异常,20％(2/10)患者存在骨髓侵犯,81.3％(13/16)患者临床分期为ⅢB期及以上;免疫组织化学染色结果示,CXCL13、PD-1、CD10、Bcl-6阳性率分别为93.8％(15/16,95％CI为70％～100％)、75.0％(12/16,95％CI为48％～93％)、50.0％(8/16,95％CI为25％～75％)和81.3％(13/16,95％CI为54％～96％);AITL显示特征性的CD23和CD21滤泡树突状细胞网增生,阳性率均为87.5％(14/16,95％CI为62％～98％);原位杂交:EBER阳性率为56.3％(9/16,95％CI为30％～80％);FISH检测结果示,10例AITL患者Bcl-2/IgH、Bcl-6、C-myc检测均为阴性;术后电话随访,自病理确诊之日起开始计算生存期,15例患者有随访结果.随访1～84个月,全组中位生存期为17个月;其中5例在1年内死亡,1年生存率为66.7％(5/15),总共6例患者死亡,占全组的40.0％(6/15).单因素分析显示,年龄≥65岁、IPI评分高危组、LDH水平异常、临床病理分期ⅢB～Ⅳ期患者预后较差.结论 AITL好发于老年患者,具有高度侵袭性,预后差,CXCL13、PD-1、CD10、Bcl-6可作为一个免疫组化套餐,在该肿瘤的诊断具有重要作用,临床病理分期、IPI评分、LDH水平是影响预后的重要因素.     

乳腺癌组织芯片中BAG-1和p73及p16的表达及其意义

目的:检测BAG-1、p73和p16在乳腺癌、癌旁组织和正常组织中的表达,并讨论其意义。方法:利用组织芯片采用免疫组化DAKO Envision法检测乳腺癌、癌旁组织、正常组织中BAG-1、p73和p16的表达。结果:组织芯片中BAG-1、p73和p16在胞核、胞质均有表达,在正常组织中以胞质为主,但随着病变表达逐渐增加。在乳腺癌组织中的表达比乳腺癌癌旁组织、正常组织中高,差异有统计学意义,P<0·01;而p16的表达正好相反,差异亦有统计学意义,P<0·01。BAG-1、p73和p16的阳性率在Ⅰ级和Ⅱ、Ⅲ级乳腺癌之间差异无统计学意义,P>0·05。结论:BAG-1、p73和p16表达的变化提示它们可能参与了乳腺癌的发生。