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1.
垂体腺瘤p16抑癌基因免疫组化和原位杂交的检测及与PCNA共表达的研究 总被引:4,自引:0,他引:4
研究良性肿瘤垂体腺瘤中p16抑癌基因的改变和可能的作用机制。方法用免疫组化和原位杂交技术研究p16在垂体腺瘤中的表达及其与临床病理的关系;并结合反映细胞增殖活性的指标PCNA的改变,以进一步了解p16在垂体腺瘤发生、发展过程中可能作用的分子机理。结果p16基因在垂体腺瘤中不发生缺失,而是存在不同程度的表达增强,其免疫组化阳性百分率的高低与肿瘤的大小、侵袭性、复发密切相关;p16的表达与PCNA呈高度正相关。结论p16、PCNA反映垂体腺瘤细胞的增殖活性,可能对临床预后有提示意义。 相似文献
2.
目的TRIO基因位于染色体5p扩增区、编码1种在细胞周期调节中起主导作用的蛋白,本研究旨在了解TRIO的扩增和表达是否与膀胱肿瘤细胞的快速增殖有关。方法Ki67标记染色(Ki67LI)、荧光原位杂交(FISH)和定量PCR法(LightCyclerTMPCR)被用于研究。结果TRIO基因扩增与膀胱肿瘤细胞的快速生长相关(P<0.0001)。膀胱肿瘤分化越差、临床分期越高,Ki67LI与TRIO基因扩增数目均越高。含2317例组织微阵列FISH结果显现TRIO扩增仅见于1.8%早期膀胱肿瘤(pTa)(9例/499例),而在膀胱癌(pT1-4)高达12.8%(62例/485例)。定量PCR法进一步证实TRIO在显示5p扩增的膀胱癌组织高表达。结论TRIO扩增与膀胱肿瘤细胞的快速增殖有关,TRIO扩增和高表达可能在膀胱癌的发展过程中发挥重要作用。 相似文献
3.
4.
Molecular detection of neoplastic cells in lymph nodes of metastatic colorectal cancer patients predicts recurrence. 总被引:11,自引:0,他引:11
M Sanchez-Cespedes M Esteller K Hibi F O Cope W H Westra S Piantadosi J G Herman J Jen D Sidransky 《Clinical cancer research》1999,5(9):2450-2454
Disseminated disease, especially to the liver, constitutes the major risk of recurrence for colorectal cancer patients. However, successful resection can still be achieved in 25-35% of colorectal cancer patients with isolated metastases. To evaluate the clinical value of occult micrometastatic disease detection in lymph nodes, we tested genetic (K-ras and p53 gene mutations) and epigenetic (p16 promoter hypermethylation) molecular markers in the perihepatic lymph nodes from colorectal cancer patients with isolated liver metastases. DNA was extracted from 21 paraffin-embedded liver metastases and 80 lymph nodes from 21 colorectal cancer patients. K-ras and p53 gene mutations were identified in DNA from liver metastases by PCR amplification followed by cycle sequencing. A sensitive oligonucleotide-mediated mismatch ligation assay was used to search for the presence of K-ras and p53 mutations to detect occult disease in 68 lymph nodes from tumors positive for these gene mutations. Promoter hypermethylation at the p16 tumor suppressor gene was examined in both liver lesions and lymph nodes by methylation-specific PCR. Sixteen of the 21 (76%) liver metastases harbored either gene point mutations or p16 promoter hypermethylation. Twelve of the 68 lymph nodes were positive for tumor cells by molecular evaluation and negative for tumor cells by histopathology and cytokeratin immunohistochemistry, whereas none were positive for tumor cells by histopathology or negative for tumor cells by molecular analysis (P = 0.0005, McNemar's test). Moreover, in three patients with lymph nodes that were histologically negative at all sites, molecular screening detected tumor DNA at one or more lymph nodes. Survival analysis showed a median survival of 1056 days for patients without evidence of lymph node involvement by molecular analysis and 165 days for patients with positive lymph nodes by this approach (P = 0.0005). These results indicate that lymph node metastasis screening in colorectal cancer patients by molecular-based techniques increases the sensitivity of tumor cell detection and can be a good predictor of recurrence in colorectal cancer patients with resectable liver metastases. 相似文献
5.
目的 了解食管癌高发区揭阳市食管癌患者 p16基因的表达与缺失情况。方法 采用免疫组化、PCR及原位杂交技术检测 6 0例食管癌标本中 p16蛋白表达、基因纯合性缺失及其mRNA信号表达情况。结果 (1)有 2 6例 (43.3% ) p16蛋白表达阳性 ,且有淋巴结转移与无转移之间蛋白表达差别显著 (P =0 .0 4 3) ;(2 )PCR检测外显子 2缺失有 2 7例 (45 % ) ,外显子 1缺失有 15例 (2 5 % ) ;(3)原位杂交检测 10 / 13例与免疫组化结果一致。结论 p16基因表达缺失与该组食管癌的浸润、转移有相关 ;外显子 2的缺失可能是 p16基因产物丢失的主要机制。 相似文献
6.
血浆p16和FHIT基因甲基化在食管癌及癌前病变检测的临床意义 总被引:2,自引:0,他引:2
目的:研究食管癌及癌前病变患者血浆中p16和FHIT基因甲基化情况,探讨其对食管早期癌和癌前病变诊断的临床应用价值。方法:应用甲基化特异性聚合酶链反应(methylation-specific PCR,MSP)方法,对食管癌及癌前病变、慢性食管炎患者组织及血浆标本进行了p16和FHIT基因甲基化检测。结果:在44例癌前病变、14例原位癌、37例浸润癌和10例慢性食管炎共105例患者组织DNA中,分别发现18例、11例、24例和1例p16基因甲基化,15例、9例、25例和0例FHIT基因甲基化。癌前病变和慢性食管炎患者血浆中未发现两基因甲基化。51例食管癌患者血浆中共检出14例p16基因和16例FHIT基因甲基化,其中2例为原位癌。结论:p16及FHIT基因甲基化检测有望将食管癌的筛查提前到原位癌阶段,为食管癌的早期发现提供帮助。 相似文献
7.
Park YK Cho CH Chi SG Han CS Ushigome S Unni KK 《International journal of oncology》2001,19(4):749-753
Mutational inactivation of the cyclin-dependent kinase inhibitors (CDKIs) (p16CDKN2a) tumor suppressor gene has been found in a variety of human tumor types. To investigate the involvement of CDKI abnormality in clear cell chondrosarcoma, alterations of CDKIs were examined in clear cell chondrosarcoma tissues using a quantitative DNA/PCR, PCR-SSCP. Two of 38 specimens (5.2%) we analyzed showed abnormally low levels of p16CDKN2a amplification, suggesting that the allelic deletion of the gene might be low frequent event in progression of this tumor. For detection of subtle sequence alterations such as point mutations, we performed SSCP analysis of the entire coding region of the p16CDKN2a gene. No altered SSCP patterns were found in 38 clear cell chondrosarcoma specimens. This study reflects the very low incidence of genetic alterations of the p16CDKN2a gene in clear cell chondrosarcoma. Therefore, we conclude that the alteration of the p16CDKN2a gene is not involved significantly in the development of clear cell chondrosarcoma. 相似文献
8.
p16基因的表达上调对鼻咽癌细胞恶性表型的影响:p16基因在鼻咽癌中的改变 总被引:1,自引:0,他引:1
探讨上调P16基因的表达对鼻咽癌细胞恶性表型逆转的作用,为P16基因在处鼻咽癌发病过程中具有一定的作用提供直接的语气方法,本文采用电穿孔的方法向P16基因表达下调的鼻咽癌细胞系HNE1中导入全长野生型的P16cDNA,通过G418筛选获得可以稳定传代的转当细胞系,对其中4个转染细胞系以PCR和Norhern杂交鉴定转染细胞系中外源P16cDNA整合以及P16基因表达的状况,并借助比较细胞倍增时间, 相似文献
9.
David J Simpson Anne M McNicol David C Murray Adil Bahar Helen E Turner John A H Wass Margaret M Esiri Richard N Clayton William E Farrell 《Clinical cancer research》2004,10(5):1780-1788
PURPOSE: The majority of cases of Cushing's disease are due to the presence of a corticotroph microadenoma. Less frequently no adenoma is found and histology shows either corticotroph hyperplasia, or apparently normal pituitary. In this study we have used molecular pathology to determine whether the tissue labeled histologically as "normal" is indeed abnormal. EXPERIMENTAL DESIGN: Tissue from 31 corticotroph adenomas and 16 nonadenomatous pituitaries were subject to methylation-sensitive PCR to determine the methylation status of the p16 gene CpG island. The proportion of methylated versus unmethylated CpG island was determined using combined bisulphite restriction analysis. Methylation status was correlated with immunohistochemical detection of p16. RESULTS: Seventeen of 31 adenomas (54.8%), 4 of 6 cases of corticotroph hyperplasia, and 7 of 10 apparently normal pituitaries showed p16 methylation. Ten of 14 (71%; P = 0.01) adenomas and 2 of 3 cases of corticotroph hyperplasia, which were methylated, failed to express p16 protein. However, only 2 of 7 apparently normal pituitaries that were methylated failed to express p16 protein. Quantitative analysis of methylation using combined bisulphite restriction analysis showed only unmethylated CpG islands in postmortem normal pituitaries; however, in adenomas 80-90% of the cells within a specimen were methylated. The reverse was true for corticotroph hyperplasia and apparently normal pituitaries where only 10-20% of the cells were methylated. Thus, the decreased proportion of cells that were methylated, particularly in those cases of apparently normal pituitary, is the most likely explanation for the lack of association between this change and loss of cognate protein in these cases. CONCLUSIONS: To our knowledge this is the first report that describes an intrinsic molecular change, namely methylation of the p16 gene CpG island, common to all three histological patterns associated with Cushing's disease. Thus, the use of molecular pathology reveals abnormalities undetected by routine pathological investigation. In cases of "apparently" normal pituitaries it is not possible to determine whether the change is associated with adenoma cells "scattered" throughout the gland, albeit few in number, or with the ancestor-clonal origin of these tumor cells. 相似文献
10.
Analytical methods for evaluation on whole cells of human papillomavirus infection. Human papillomavirus (HPV) infection is currently identified by the presence of viral DNA using molecular biology. As in situ hybridization is valuable for HPV-DNA detection mainly with non-isotopic probes, we evaluated the sensitivity of various techniques, using as models three cell lines containing different copy numbers of HPV DNA/cell (CaSki with 600 copies of HPV 16, SiHa with 1-2 copies of HPV 16, HeLa with 10-50 copies of HPV 18). Epifluorescence microscopy and flow cytometry allowed detection of 600 copies in CaSki cells; in addition, cell fixation was found to influence the fluorescent intensity. Several procedures were assayed to increase the sensitivity of in situ hybridization. The use of biotinylated HPV-16 oligonucleotides as probes was not effective, because only CaSki cells were positive. After amplification of HPV-16 or -18 DNA sequences with polymerase chain reaction (PCR) on whole cells in suspension and hybridization with plasmid probes, fluorescent hybridization spots were found in CaSki and HeLa cells by both epifluorescence microscopy and flow cytometry. The various procedures applied for revelation of DNA-DNA hybrids (use of phycoerythrin or cyanine instead of fluorescein, Pinkel's 3-step amplified system of fluorescein) did not enhance the sensitivity of in situ hybridization. HPV DNA was very effectively detected by cell examination under a laser-scanning confocal microscope, since 1-2 copies of HPV 16 were observed in SiHa cells without previous PCR amplification. Thus, the efficacy of in situ hybridization for HPV detection may be conditioned by different factors. Laser-scanning microscopy represents an alternative to the use of PCR amplification. These techniques are potentially useful to study single genes. 相似文献
11.
】 ObjectiveTo study the
biological effect of As2O3 on human pulmonary adenocarcinoma GLC-82
cells and its mechanisms. MethodsMTT reduction assay,morphology
investigation,flowcytometric analysis,DNA gel electrophoresis and TUNEL,RT-PCR,Northern
blot, Western blot methods were perfomed. ResultsAs2O3
inhibited the growth of GLC-82 cell line. The cells treated with As2O3
showed a typical apoptotic morphology and hypodiploid peak before G1 phase. DNA of treated
GLC-82 cells appeared a ladder pattern characteristic of apoptosis. In situ cell death
detection analysis also revealed the DNA fragmentation. Besides,As2O3
inhibited c-myc gene expression and increased p53 and p16 gene expression. ConclusionAs2O3
can induce GLC-82 cell apoptosis mainly with regulation of c-myc,p53 and p16 gene
expression. 相似文献
12.
外源性p16基因表达可抑制人胃癌细胞的恶性增殖 总被引:20,自引:1,他引:19
目的探讨mtsl/p16基因在肿瘤发生发展过程中的作用及在临床基因诊断和基因治疗中的应用前景。方法构建mtsl/p16基因的表达载体并导入表达水平下调的PAMC82细胞,用PCR、Northern杂交、mRNA原位杂交和Western杂交对获得G418抗性的细胞进行外源性mtsl/p16基因整合及表达的鉴定。对导入外源性p16基因的细胞进行裸鼠致瘤能力及病理学特性分析。结果转染p16基因的PAMC82细胞(命名为PAMCp16)有外源性mtsl/p16基因的整合及表达,在裸鼠中的致瘤性显著降低。肿瘤组织病理分析结果显示,PAMCp16细胞形成的肿瘤,其分化程度优于PAMC82细胞。结论导入外源性mtsl/p16基因可抑制肿瘤细胞的恶性增殖和促进细胞分化。 相似文献
13.
The presence of comedo carcinoma in the intraductal component is an important pathologic feature for predicting local recurrence after breast conserving treatment. Furthermore, the c-erbB-2 gene is frequently overexpressed in comedo carcinoma and it correlates well with gene amplification. This study revealed that 77.0% of the c-erbB-2 amplification positive tumors had intraductal lesions of the comedo type; while only 6.7% of the amplification negative tumors had comedo carcinoma (X(2)=106.6, p<0.0001). Accordingly, when c-erbB-2 amplification is detected in the invasive tumor, intraductal lesions which may co-exist in the tumor are likely to be comedo carcinoma. To predict the presence of comedo type intraductal carcinoma, c-erbB-2 gene amplification was analysed by means of polymerase chain reaction (PCR) from small samples of fine-needle aspiration biopsies. Breast tumors from ten patients were analysed and one had a 3.4-fold amplification of c-erbB-2 gene. The pathological appearance of this case was pure ductal carcinoma in situ with comedo carcinoma. The preoperative detection of c-erbB-2 gene amplification by PCR will be a useful means of selecting patients at high risk for local recurrence after breast conserving treatment. 相似文献
14.
目的:探讨直接在石蜡切片上进行原位PCR的可行性,以此分析霍奇金淋巴瘤(hodgkinlymphoma,HL)H/R-S细胞的性质。方法:建立一种5′-标记引物直接原位PCR方法,对3例10%中性甲醛固定石蜡包埋HL的H/R-S细胞IgH基因重排情况进行检测。结果:2例结节硬化型霍奇金淋巴瘤(nodularsclerosistypehodgkinlymphoma,NSHL)H/R-S细胞中,1例有VH2、VH3和FR3基因重排,1例有VH1和VH3基因重排,1例混合细胞型霍奇金淋巴瘤(mixedcellularitytypehodgkinlymphoma,MCHL)的H/R-S细胞有VH2、VH3和FR2a基因重排。背景的部分小淋巴细胞核内也出现了阳性信号。结论:1)在石蜡切片上进行直接法原位PCR是可行和有效的。2)NSHL和MCHL的H/R-S细胞来源于不同分化阶段的B细胞,NSHL和MCHL的H/R-S细胞为多克隆性增生。周围部分背景小淋巴细胞有可能与H/R-S细胞来自同一B细胞克隆。 相似文献
15.
目的应用荧光原位杂交(fluorescence in situ hybridization,FISH)技术,了解膀胱尿路上皮肿瘤细胞核染色体畸变情况及其对病理诊断及鉴别诊断的应用价值。方法采用3、7、17号染色体着丝粒及p16基因探针,对33例不同级别膀胱尿路上皮肿瘤组织和10例正常对照膀胱组织进行FISH检测。结果 3、7、17染色体及p16扩增率与缺失率,浸润性尿路上皮癌与其他5组比较均有统计学意义(P〈0.01);而其他各组间比较无统计学意义(P〉0.05),4种探针联合检测≥2个指标出现异常,浸润性尿路上皮癌占100%,与其他5组比较有统计学意义(P〈0.01);该指标对浸润性尿路上皮癌诊断敏感性为81.82%、特异性为91.67%。结论应用FISH技术,可以了解膀胱尿路上皮肿瘤组织3、7、17染色体及p16基因的畸变情况,FISH技术还可作为尿路上皮肿瘤病理学诊断与鉴别诊断及术后监测的重要手段。 相似文献
16.
Wang X Mori I Tang W Nakamura M Nakamura Y Sato M Sakurai T Kakudo K 《Breast cancer (Tokyo, Japan)》2002,9(3):216-219
BACKGROUND: p63 is a homologue of the p53 tumor suppressor gene and its protein is selectively expressed in the basal cells of a variety of epithelial tissues. It has recently been confirmed that p63 is expressed in the basal cells of normal prostate glands but not in prostatic carcinomas. Whether expression of p63 in breast correlates with tumor progression is the focus of this study. METHODS: Forty cases, which all contained normal breast tissue, ductal hyperplasia, ductal carcinoma in situ and invasive ductal carcinoma in the same patient were included in this investigation using an indirect immunohistochemical method and double staining. RESULTS: p63 was exclusively expressed in the myoepithelial cells of normal breast, partially expressed in ductal hyperplasia, rarely expressed in carcinoma in situ and not expressed in invasive carcinomas. CONCLUSION: The results suggest an association between loss of p63 expression and progression of breast ductal carcinoma. p63 immunostaining might be of assistance for distinguishing invasive ductal carcinoma from ductal carcinoma in situ or rare questionable ductal hyperplastic lesions, leading to correct therapy clinically. 相似文献
17.
目的探讨肿瘤抑制基因p16在几种类型白血病中的改变及其频率。方法对77例原发性白血病及非霍奇金淋巴瘤标本分别采用Southern印迹杂交,PCR┐SSCP的方法检测缺失及点突变。结果Southernblot杂交发现p16基因异常共11例,其中纯合缺失7例,异常重组片段4例,SS┐CP发现p16第三外显子异常泳动带6例,是否为突变有待测序证实。结论p16基因改变主要集中在急性淋巴细胞性白血病,占47.7%(9/19),提示这一肿瘤抑制基因在淋巴细胞发生恶性变的机制中起一定作用 相似文献
18.
食管癌高发区HPV检测及与p53的关系 总被引:21,自引:0,他引:21
目的 探讨食管癌高发区中人乳头瘤病毒(HPV)、爱泼斯坦-巴尔病毒(EBV)与食管癌发病的关系。并初步研究HPV感染与p53过表达之间的关系。方法 设计了多对引物进行PCR、原位杂交及原位PCR、免疫组化,并对30例高发区食管癌进行检测,结果 用PCR检出HPV-L1阳性率10.0%,HPV-16-E6阳性率60.0%,HPV-16-E7阳性率63.3%,HPV-18-E6及EBV的检出率分别为6.7%和0,p53蛋白的检出率为73.3%,用原位杂交和原位PCR检出HPV-16-E6阳性率均为53.3%.HPV-L1的低检出率及HPV-16E6、HPV-16-E7基因的较高检出率可能提示,在肿瘤细胞中HPV常以部分丢失的形式整合,而E6、E7常在整合中保留。结论 HPV-16型可能与高发区食管癌发生密切相关,而HPV-18、EBV则关系不大,p53突变在食管癌的发病中起重要作用。但其与HPV感染之间的关系不大。 相似文献
19.
背景与目的:近年研究表明,抑癌基因p16表达缺失及失活在人类多种肿瘤发生发展过程中发挥重要作用,但其与垂体腺瘤之间的关系研究甚少,本研究旨在探讨垂体腺瘤中p16基因表达情况及其与肿瘤临床病理特点、侵袭性、复发之间的关系。方法:用RT-PCR、Westernblot方法分别检测70例垂体腺瘤及10例正常脑组织中p16mRNA、p16蛋白表达情况,并结合临床病理进行探讨。结果:74.3%(52/70)垂体腺瘤中的p16mRNA、p16蛋白表达缺失或低下,侵袭性腺瘤、复发腺瘤p16表达缺失率均分别高于非侵袭腺瘤及非复发腺瘤,但统计学处理各组间无显著差异(P>0.05);p16表达缺失肿瘤的平均直径为(22.1±7.2)mm,明显大于阳性表达肿瘤的直径(8.1±4.5)mm(P<0.01)。此外,p16基因表达与垂体腺瘤其它临床病理特点均无显著相关。结论:在垂体腺瘤中存在较普遍的p16基因转录抑制及表达低下,其失活可能在垂体腺瘤形成早期发挥重要作用,并对肿瘤生长及生物学行为演变起一定促进作用。 相似文献
20.
Sekido Y Umemura S Takekoshi S Suzuki Y Tokuda Y Tajima T Osamura RY 《International journal of oncology》2003,22(6):1225-1232
The aim of the present study was to clarify differences in genetic events between primary breast cancers and asynchronous metastatic/recurrent lesions, by examining HER2 gene amplification and p53 mutation. The subjects were 44 breast cancer patients with asynchronous metastasis or recurrence. Synchronous metastases were excluded. HER2 overexpression and gene amplification were examined using immunohistochemistry and fluorescent in situ hybridization (FISH). P53 point mutation was examined by immunohistochemistry, laser-captured microdissection, PCR-single-strand conformation polymorphism, and a direct sequencing method. Immunohistochemistry showed that, for HER2, p53, ER and PgR, discordance rates between primary and recurrent tumor were 2 (4.5%), 1 (2.3%), 7 (15.9%) and 10 (22.7%), respectively. Two primary tumors with discordant HER2 overexpression were composed of at least two populations of carcinoma cells, with and without HER2 gene amplification. Distribution of HER2 gene amplification was consistent with protein overexpression. Corresponding recurrent tumors consisted of carcinoma cells without HER2 gene amplification. Of 6 recurrent tumors in which the primary carcinoma had a p53 point mutation, 3 tumors had identical mutations, 1 tumor had a different point mutation, and 2 tumors had no mutation. It was suspected that the latter 3 recurrent tumors comprised a minor component of the primary tumor. In the present study, we examined a large series of asynchronous recurrent tumors. A limited number of these tumors showed discordance between primary and recurrent tumors. Detailed observations revealed that cell populations present in recurrent tumors were also present in the primary tumors, although they comprised a minor component of the primary tumor. Heterogeneity of the primary tumor apparently contributed to discordance. 相似文献