膀胱癌组织端粒酶活性定量分析及临床应用的研究

目的：组装简便实用的端粒酶活性相对定量检测试剂盒,建立了膀胱肿瘤端粒酶活性非同位素相对定量检测技术。方法：用银染－TRAP略行改良,对54例膀胱癌组织、癌旁组织、12例正常膀胱黏膜,其中46例膀胱癌患者同时收集了尿液及膀胱冲洗液的脱落细胞进行端粒酶活性相对定量值分析。结果：54例膀胱癌组织中端粒酶活性相对定量值阳性率与正常膀胱黏膜和癌旁组织端粒酶活性相对定量值相比,差异有显著性（P＜0．001,P＜0．001）;癌旁组织略高于正常膀胱黏膜,但差异无显著性（P＞0．05）,46例膀胱癌膀胱冲洗液与尿液相比差异有显著性（P＜0．01）;54例膀胱癌与癌旁组织中端粒酶活性相对定量均值及标准差相比,差异有显著性（P＜0．001）;46例膀胱癌冲洗液与尿液端粒酶活性相对定量均值及标准差相比差异有显著性（P＜0．05）。结论：建立了膀胱肿瘤端粒酶活性的非同位素相对定量分析方法,具有特异性强、灵敏性高的特点,该方法可作为恶性肿瘤诊断和预后的指标之一。     

膀胱癌组织端粒酶活性定量分析及临床应用的研究

目的 :组装简便实用的端粒酶活性相对定量检测试剂盒 ,建立了膀胱肿瘤端粒酶活性非同位素相对定量检测技术。方法 :用银染 -TRAP略行改良 ,对 54例膀胱癌组织、癌旁组织、12例正常膀胱黏膜 ,其中 4 6例膀胱癌患者同时收集了尿液及膀胱冲洗液的脱落细胞进行端粒酶活性相对定量值分析。结果 :54例膀胱癌组织中端粒酶活性相对定量值阳性率与正常膀胱黏膜和癌旁组织端粒酶活性相对定量值相比 ,差异有显著性 (P <0 0 0 1,P <0 0 0 1) ;癌旁组织略高于正常膀胱黏膜 ,但差异无显著性 (P >0 0 5) ,4 6例膀胱癌膀胱冲洗液与尿液相比差异有显著性 (P <0 0 1) ;4 6例膀胱癌尿液与 10例正常尿液相比差异有显著性 (P <0 0 1) ;54例膀胱癌与癌旁组织中端粒酶活性相对定量均值及标准差相比 ,差异有显著性 (P <0 0 0 1) ;4 6例膀胱癌冲洗液与尿液端粒酶活性相对定量均值及标准差相比差异有显著性 (P <0 0 5)。结论 :建立了膀胱肿瘤端粒酶活性的非同位素相对定量分析方法 ,具有特异性强、灵敏性高的特点 ,该方法可作为恶性肿瘤诊断和预后的指标之一。     

尿脱落细胞端粒酶活性检测在膀胱癌中的诊断价值

目的：探讨通过检测尿脱落细胞端粒酶活性早期诊断膀胱癌。方法：采用改进的银染－TRAP（端粒重复序列扩增）法,分别对45例膀胱移行细胞癌患者,10例膀胱良性病变患者及8例正常人尿脱落细胞进行了端粒酶活性检测。结果：24例G1级（分化良好）的膀胱癌患者,端粒酶活性强阳性12例,弱阳性6例（阳性率75．0％）,21例G2,G3级膀胱癌患者,强阳性16例,弱阳性3例（阳性率94．5％）,膀胱良性病变患者及正常人尿脱落细胞只有1例端粒酶活性呈弱阳性,其余全部阴性。结论：尿脱落细胞端粒酶活性检测特异性强,敏感性高,与病理分类,组织分级等无明显关系,提示有可能成为膀胱癌临床早期诊断指标之一。     

胆汁脱落细胞端粒酶活性检测在胆道恶性肿瘤诊断中的价值

背景与目的：已有学者对尿液,胰液,肠腔冲洗液,胸水等脱落细胞端粒酶活性进行了检测,探索其对恶性肿瘤的诊断价值。而胆汁脱落细胞端粒酶活性的检测尚少见报道。本文的目的是探讨胆汁脱落细胞端粒酶活性检测及其对胆道恶性梗阻的诊断价值。方法：应用TRAP银染法检测胆汁脱落细胞的端粒酶活性。结果：44例恶性肿瘤胆道梗阻病人获得的胆汁中有33例检测到端粒酶活性阳性,阳性率75．0％,而良性梗阻组19例中只有1例检测到端粒酶弱阳性,阳性率5．3％。胆汁脱落细胞端粒酶活性阳性率与淋巴结转移,远处转移以及分化程度等肿瘤临床病理学特征无关,但胰腺癌和胆管癌病人胆汁中端粒酶活性检测阳性率高。端粒酶活性检测与脱落细胞学检查对比结果显示：44例胆道恶性梗阻的病人中31例进行了胆汁脱落细胞学检查,结果只有3例癌细胞阳性,阳性率9．7％,这3例脱落细胞学检查阳性的胆汁脱落细胞端粒酶活性检测均为阳性。结论：TRAP银染法可有效检测到胆汁脱落癌细胞的端粒酶活性。端粒酶可以作为诊断恶性胆道疾病的分子标志物。胆汁中脱落细胞端粒酶活性检测可作为细胞学检查的补充诊断手段。     

泌尿系统疾病粒酶活性及临床意义

目的：检测泌尿系统疾病尿脱落细胞和相关肿瘤组织端粒酶活性，了解泌尿系统肿瘤发生可能的分子生物学基础，方法应用TRAP－PCR ELISA法对54例膀胱癌，15例肾癌肿瘤组织和尿脱落细胞及非癌患者和尿脱落细胞进行端粒酶活性测定。结果90．2％（50．54）膀胱癌组织，。93．3％（14／15）肾癌组织端粒酶呈阳性，相应尿脱落细胞端粒酶阳性率分别为90．1％，20．0％，其它疾病和尿脱落细胞均未检出端粒酶。结论：端粒酶活性与肿瘤的发生有关。并可为某些肿瘤的早期诊断与治疗提供可靠的指标与新的线索。     

新疆子宫颈癌的端粒酶活性研究

目的：探讨端粒酶活性与宫颈癌发生发展的关系及其在宫颈癌中的临床诊断价值。方法：用TRAP－ELISA法检测34例宫颈癌和21例正常宫颈的新鲜组织及脱落细胞标本的端粒酶活性,并进行比较。结果：宫颈癌细胞中端粒酶阳性检出率为88．2％,显著高于正常宫颈细胞的阳性检出率（9．5％）,端粒酶活性水平与宫颈癌临床分期,组织学分化无关。脱落细胞检测宫颈细胞的端粒酶活性与新鲜组织检测的结果一致。结论：端粒酶活性可作为宫颈癌的肿瘤标记物。用TRAP－ELISA法对宫颈脱落细胞行端粒酶检测有助于宫颈癌的筛查。     

子宫颈病变中端粒酶活性的表达

目的：探讨宫颈糜烂，宫颈上皮内瘤样变（CIN）及宫颈癌中端粒酶的表达及其意义，方法：采用TRAP法检测30例宫颈糜烂，16例CIN组织，20例宫颈癌及20例正常宫颈组织中端粒酶活性。结果：宫颈糜烂者端粒酶活性阳性率为3．0％（1／30），CIN端粒酶活性阳性率为56％（9／16），20例宫颈癌组织中端粒酶阳性表达率为805（16／20），正常宫颈组织中端粒酶表达阴性。恶性肿瘤与正常或良性病变组织中端粒酶活性差异有显著性（P＜0．05）。结论：端粒酶的活化在宫颈癌的发生过程中起重要作用，端粒酶活性检测可能成为宫颈癌及宫颈癌前病变早期诊断诊断和病情监测的指标。     

人肺癌端粒酶活性的研究

目的 探讨端粒酶活性表达与肺癌发生发展的关系。方法 应用ＰＣＲ－ＴＲＡＰ－银染法检测５０例肺癌组织及细胞和２０例相应癌旁组织、５例肺结核瘤组织及细胞的端粒酶活性。结果 肺癌组：３６例手术切除肺癌组织中端粒酶阳性率为９１．７％,７例纤维支气管镜活检肺癌组织和７例癌性胸水细胞中端粒酶阳性例数分别为６例和５例;总检出率为８８％（４４／５０）。对照组：癌旁组织中端粒酶阳性率为１０％（２／２０）,肺结核瘤组     

膀胱癌组织中端粒酶活性的检测及其意义

目的 研究端粒酶与膀胱移行细胞癌（TCC）生物学行为的关系。方法 方法 采用端粒酶重复序列扩增法及酶联免疫吸附性检测58例TCC组织、10例癌旁膀胱粘膜及10例正常膀胱粘膜组织的端粒酶活性。结果 TCC组织端粒酶阳性率为86．2（50／58）,不同病理分级、临床分期之间无显著性差异（P＞0．05）;癌旁组织中只有1例端粒酶阳性;正常组织中端粒酶均为阴性。结论 TCC组织中端粒酶活性阳性率明显高于正常膀胱粘膜组织及癌旁膀胱粘膜组织。各病理分级和临床分期之间端粒酶活性虽然无显著性差异（P＞0．05）,但端粒酶活性的强弱分布不同。端粒酶活性可作为TCC早期诊断、鉴别诊断及预测复发的肿瘤标志物之一。     

胆管癌组织及胆汁中端粒酶活性的检测及意义

目的：检测胆管癌组织和胆汁中的端粒酶活性，以研究与端粒酶与胆管癌的关系及对胆管癌的诊断意义。方法：用PCR—ELISA方法检测20例胆管癌的癌组织和胆汁中端粒酶活性，同时用相同的方法检测20例正常胆管组织和胆汁中的端粒酶活性以作对照。结果：20例胆管癌组织中，端粒酶活性阳性为80％（16／20），胆汁中端粒酶活性阳性率75％（15／20）。正常胆管组织端粒酶活性阳性表达率为0（0／20），胆汁中端粒酶活性阳性率为0（0／20）。结论：端粒酶可能参与了胆管癌的发生发展过程，检测胆汁中端粒酶活性可有助于胆管癌的诊断。     

Telomerase activity in urine sediments as a tool for noninvasive detection of bladder cancer

Telomerase is extensively investigated as potential diagnostic and prognostic marker in human tumors. In this study, we determined telomerase activity in histological specimens and voided urine of 52 human bladder cancers. Using the PCR-ELISA method telomerase activity was found in 21 (88%) of the 24 tumor tissues and in the corresponding sediments from voided urine of patients with superficial bladder carcinoma (Ta/T1). In case of muscle-invasive tumors (T2-T4), telomerase activity was found in 27 (96%) of the 28 tumor tissues and in 26 (93%) of the 28 urine sediments. Enzyme activity was not detected in 13 control urine sediments. Telomerase activity was not significantly associated with clinicopathological parameters supporting the diagnostic rather than prognostic value of this marker in bladder cancer. The present study demonstrates that telomerase activity detection in voided urine has high potential for noninvasive diagnosis of superficial bladder tumors.     

Clinical sensitivity of p53 mutation detection in matched bladder tumor, bladder wash, and voided urine specimens

BACKGROUND: Mutations in the p53 tumor suppressor gene may correlate with an increased risk of recurrence and disease progression in patients with bladder carcinoma. The ability to accurately and sensitively detect p53 mutations in cytology specimens may be of benefit in the treatment of bladder carcinoma patients with superficial, minimally invasive disease. METHODS: Genomic DNA was isolated from 49 cases, each of which was comprised of matched bladder tumor tissue, bladder wash, and voided urine specimens obtained concurrently at a single institution. The genomic DNA was analyzed for mutations in the p53 tumor suppressor gene using a p53 mutation detection assay. Automated dideoxy sequencing of mutant specimens also was performed. RESULTS: Of the 49 cases, 29 (59%) showed no evidence of p53 mutations in the tumor, bladder wash, or voided urine specimens. Of the remaining 20 cases, 19 showed evidence of mutations in the tumor. Of these 19 p53 mutant bladder tumors, 16 (84%) were detected in the matched bladder wash and 16 (84%) were detected in the matched voided urine specimens. One case resulted in the detection of mutant p53 in the voided urine and the bladder wash, but not in the tumor. Analysis of the results between tumor tissue and bladder wash or tumor and voided urine showed 84.2% sensitivity, 96.8% specificity, and 91.8% accuracy. Sequence analysis of the mutant cases showed that the mutations detected in the tumor tissue were the same mutations detected in the bladder wash and the voided urine specimens. CONCLUSIONS: Both voided urine and bladder wash specimens from patients with bladder carcinoma were found to provide a high rate of clinical accuracy for the determination of the p53 gene status in patients with bladder tumors.     

尿细胞角蛋白检测与尿脱落细胞学检查在膀胱癌诊断中的价值

目的:探讨尿细胞角蛋白检测与尿脱落细胞学检查在膀胱移行细胞癌诊断中的价值。方法:136例怀疑膀胱癌者,进行尿细胞角蛋白8和18的含量(UBC值)。检测与尿细胞学检查,其中87例经组织学证实为膀胱移行细胞癌。比较两者诊断膀胱癌的敏感性和特异性。结果:尿细胞角蛋白的敏感性为70.1%,特异性为73.3%;尿细胞学的敏感性为42.5%,特异性为83.7%。尿细胞角蛋白在膀胱癌不同分级和分期中的敏感性优于尿细胞学(P<0.05)。结论:尿细胞角蛋白的检测在早期诊断膀胱癌方面优于尿细胞学检查,可作为膀胱癌的早期检测指标。     

Hypermethylation of multiple genes in tumor tissues and voided urine in urinary bladder cancer patients.

PURPOSE: We aimed to investigate the methylation pattern in bladder cancer and assess the diagnostic potential of such epigenetic changes in urine. EXPERIMENTAL DESIGN: The methylation status of 7 genes (RARbeta, DAPK, E-cadherin, p16, p15, GSTP1, and MGMT) in 98 cases of bladder transitional cell carcinoma and 4 cases of carcinoma in situ was analyzed by methylation-specific PCR. Twenty-two cases had paired voided urine samples for analysis. RESULTS: In transitional cell carcinoma tumor tissues, aberrant methylation was frequently detected in RARbeta (87.8%), DAPK (58.2%), E-cadherin (63.3%), and p16 (26.5%), whereas methylation of p15 (13.3%), GSTP1 (5.1%), and MGMT (5.1%) is not common. No association between methylation status and grading or muscle invasiveness was demonstrated. In 22 paired voided urine samples of bladder cancer, methylation of DAPK, RARbeta, E-cadherin, and p16 could be detected in 45.5%, 68.2%, 59.1%, and 13.6% of the cases, respectively. The sensitivity of methylation analysis (90.9%) was higher than that of urine cytology (45.5%) for cancer detection. Methylation of RARbeta(50%), DAPK (75%), and E-cadherin (50%) was also detected in carcinoma in situ. In 7 normal urothelium samples and 17 normal urine controls, no aberrant methylation was detected except for RARbeta methylation in 3 normal urothelium samples (42.9%) and 4 normal urine samples (23.5%), respectively. CONCLUSIONS: Our results demonstrated a distinct methylation pattern in bladder cancer with frequent methylation of RARbeta, DAPK, E-cadherin, and p16. Detection of gene methylation in routine voided urine using selected markers appeared to be more sensitive than conventional urine cytology.     

联合检测尿液中CYFRA21-1及端粒酶活性在膀胱癌早期诊断中的意义

目的 通过联合检测尿液中细胞角蛋白19(CYFRA21-1)及端粒酶活性在膀胱癌患者中的水平,探讨其在膀胱癌术后复发早期诊断中的意义.方法 运用双单抗夹心酶联免疫法(ELISA)检测110例膀胱癌患者,52例良性患者及30名健康志愿者尿液样品中CYFRA21-1含量,以3.3 ng/mL检测临界值;采用聚合酶链反应的酶联免疫吸附试验(PCR-ELISA)方法 检测以上三组尿液样品中端粒酶活性.结果 膀胱癌患者尿液中CYFRA21-1水平显著高于良性患者组和正常对照组(P<0.01).与膀胱癌分期分级的相关性分析显示浸润性膀胱癌(T2、T3、T4)CYFRA21-1水平均高于表浅性膀胱癌(Tis、Ta、T1),相比有统计学意义(P<0.05).CYFRA21-1水平、端粒酶活性检测诊断膀胱癌的敏感度分别为92.2%、78.2%,特异度为66.7%、93.3%;而将上述方法 联合应用来筛检则敏感性可达到97.5%.结论 联合应用CYFRA21-1及端粒酶的检测敏感性高,特异性好,可用于膀胱癌的早期诊断及术后随访.     

Highly sensitive and specific novel biomarkers for the diagnosis of transitional bladder carcinoma

Transitional bladder carcinoma (BCa) is prevalent in developed countries, particularly among men. Given that these tumors frequently recur or progress, the early detection and subsequent monitoring of BCa at different stages is critical. Current BCa diagnostic biomarkers are not sufficiently sensitive for substituting or complementing invasive cystoscopy. Here, we sought to identify a robust set of urine biomarkers for BCa detection. Using a high-resolution, mass spectrometry-based, quantitative proteomics approach, we measured, compared and validated protein variations in 451 voided urine samples from healthy subjects, non-bladder cancer patients and patients with non-invasive and invasive BCa. We identified five robust biomarkers: Coronin-1A, Apolipoprotein A4, Semenogelin-2, Gamma synuclein and DJ-1/PARK7. In diagnosing Ta/T1 BCa, these biomarkers achieved an AUC of 0.92 and 0.98, respectively, using ELISA and western blot data (sensitivity, 79.2% and 93.9%; specificity, 100% and 96.7%, respectively). In diagnosing T2/T3 BCa, an AUC of 0.94 and 1.0 was attained (sensitivity, 86.4% and 100%; specificity, 100%) using the same methods. Thus, our multiplex biomarker panel offers unprecedented accuracy for the diagnosis of BCa patients and provides the prospect for a non-invasive way to detect bladder cancer.     

Telomerase activity in bladder cancer, bladder washings and in urine

With only a few exceptions, the ribonucleoprotein telomerase has been found in malignant, but not in benign tissues. Telomerase is thus a potentially new diagnostic marker. Carcinoma of the urinary bladder is the most frequent malignant tumor of the urinary tract and, after prostatic carcinoma, the second most common malignancy of the genitourinary system. In order to evaluate the diagnostic capabilities of telomerase in bladder carcinomas, four cell lines derived from human urothelial carcinomas of the bladder, 75 tissue samples from bladder carcinomas, eight tissue samples of normal bladder urothelium, 40 bladder washings and 30 urine samples were examined for telomerase activity. The four cell lines derived from urothelial carcinoma of the bladder (F975, 582, SCaBER, UM-UC-3) all exhibited high telomerase activity and were thus used as positive controls. Telomerase activity was found in nearly all (96%) tissue samples obtained from histologically confirmed urothelial carcinoma of the bladder. None of the normal tissue samples examined showed telomerase activity. Telomerase activity was similarly found in 73% of bladder washings in patients with histologically confirmed bladder carcinoma. There were no false positive results. The determination of telomerase activity in bladder washing samples thus represents a new diagnostic method for detection of tumor cells in rinsing media. Because of the early inactivation or degradation of telomerase there was no detection of the enzyme in native urine in the present study.     

细针穿刺肿瘤组织端粒酶活性检测在乳腺癌诊断中的意义

目的 :探讨细针穿刺乳腺肿瘤组织端粒酶活性检测在乳腺癌诊断中的意义。方法 :用PCR ELISA法检测79例术前乳腺肿瘤穿刺活检标本和大体标本的端粒酶活性并与病理诊断进行对照。结果 :乳腺癌 65例 ,穿刺组织端粒酶阳性 5 7例 ,阳性率为 87 7% ;大体组织端粒酶阳性 5 4例 ,阳性率83 1% ;淋巴结有转移者端粒酶活性高于无淋巴结转移者 ;乳腺良性疾病 14例 ,端粒酶阳性 2例 ,阳性率 14 3 %。结论 :术前乳腺肿瘤穿刺组织端粒酶活性检测有利于乳腺肿瘤的早期诊断及鉴别诊断 ,可以间接了解乳腺癌的进展程度     

检测尿脱落细胞中微卫星不稳定性的临床意义

目的:探讨膀胱癌患者尿脱落细胞微卫星不稳定性及临床意义.方法:将42例患者尿液标本经适当处理后,采用PCR-PAGE-银染方法检测尿脱落细胞中微卫星不稳定性的情况,分析其与临床病理参数的关系.结果:膀胱癌患者尿中MSI阳性率为59.52%(25/42),浸润性肿瘤患者尿中阳性率较高,但无统计学意义(P＞0.05);复发病例中阳性率(80.00%)比初发者(40.91%)明显要高(P＜0.05).结论:检测尿脱落细胞微卫星不稳定性可以作为膀胱癌的一种无创诊断及筛查手段.     

乳腺肿瘤组织中端粒酶的表达及其对诊治的意义

背景与目的：研究表明,端粒酶的活化可使细胞转化为癌细胞。本研究拟分析端粒酶在乳腺良性和恶性组织中的表达及其对乳腺癌早期诊断的价值。方法：应用PCR－ELISA方法检测178例乳腺癌、乳腺纤维瘤等患者术后组织中端粒酶的表达。结果：端粒酶阳性表达率在原发性乳腺癌组织为86．72％（111／128）;乳腺癌癌前病变组织为46．67％（7／15）;乳腺纤维瘤组织为26．66％（4／15）;乳腺非癌组织、囊性增生组织全阴性。乳腺癌在不同的临床分期中,端粒酶阳性表达率为临床I期63．16％,Ⅱ期85．96％,Ⅲ期为965％,Ⅳ期为100％。原发性乳腺癌伴腋窝淋巴结阳性者端粒酶阳性表达率为95．83％（69／72）,腋窝淋巴结阴性者端粒酶阳性表达率75％（42／56）。结论：端粒酶的表达与乳腺癌的临床分期（P＜0．005）,腋淋巴结转移密切相关（P＜0．01）。特别是它在乳腺癌癌前病变中有表达。端粒酶的检测对乳腺癌的早期诊断具有重要价值。