Comparison of NMP22 BladderChek test and urine cytology for the detection of recurrent bladder cancer

OBJECTIVE: To assess the clinical performance of the NMP22 BladderChek test, which is a qualitative test, and to compare it with voided urine cytology for the detection of recurrent bladder cancer. We also evaluated whether cystoscopy can be omitted from the surveillance protocol by combining the two tests. METHODS: A total of 131 patients with a history of superficial transitional cell carcinoma of the bladder provided urine samples before a cystoscopic examination. Urine samples were assayed for the presence of NMP22 using the NMP22 BladderChek test and cytology was performed by a cytopathologist. Selected patients underwent a biopsy, with appropriate additional therapy. Results of the two tests were compared with that the results of cystoscopy, which was retained as the gold standard. For positive biopsies, the results of the NMP22 test and cytology were also correlated with the tumor stage and grade. RESULTS: Of the 46 recurrences detected by cystoscopy, the NMP22 test was positive in 39 cases and cytology in 19 cases. The sensitivity of the NMP22 test was 85%, which was significantly greater than that of cytology (41%). In particular, for low-risk tumors it was eight times more sensitive than cytology. The specificities of the NMP22 test and cytology were 77 and 96%, respectively. Combining the two tests increased overall sensitivity to 91%. However, 9% of the tumors were still not detected. CONCLUSION: The NMP22 BladderChek test is an in vitro qualitative test that is easily available and cheap; it can be performed by a urologist in the office and results can be interpreted within 30 min. The NMP22 test is superior to cytology for all grades and stages in the detection of recurrence in patients with a history of superficial bladder cancer. Our study indicates that the NMP22 test can be used as a substitute for urine cytology. The NMP22 test cannot replace cystoscopy, but it can be used as an adjunct to cystoscopy in the surveillance protocol for patients with superficial bladder cancer.     

Comparison of fluorescence in situ hybridization,NMP22 bladderchek,and urinary liquid‐based cytology in the detection of bladder urothelial carcinoma

The aim of this study is to evaluate the diagnostic values of the fluorescence in situ hybridization (FISH), NMP22 BladderChek, and liquid‐based cytology (LBC) in the detection of bladder urothelial carcinoma (UC). Consecutive voided urine samples were collected from 138 in‐house patients with a variety of urologic conditions and 37 healthy individuals as negative controls. FISH, NMP22 BladderChek, and LBC were performed on the specimens. All three tests were evaluated independently in a blinded fashion. In all, 104 out of the 175 patients enrolled in this study had histologically proven UC. LBC, FISH, and NMP22 BladderChek were successfully performed on 175, 149, and 119 cases, respectively. The three tests revealed overall sensitivities of 73.1%, 86.5%, and 67.6%, respectively. FISH was more sensitive than LBC (P=0.022) and NMP22 BladderChek (P=0.004). Combination of all the tests yielded a superior sensitivity of 96.7% compared with LBC (P<0.001), NMP22 BladderChek (P<0.001), and FISH (P=0.016), with the specificity only decreased slightly. Sensitivities of the three tests enhanced significantly with increasing UC grade (P<0.05). The positive rates of FISH and NMP22 BladderChek in equivocal cytologic diagnoses were 85.7% and 61.9% in UC, and 37.5% and 50.0% in non‐UC (FISH: P=0.021; NMP22 BladderChek: P=0.683). FISH was more sensitive than LBC and NMP22 BladderChek. FISH had the ability to clarify equivocal cytologic diagnoses. Combination of all three tests showed an improvement in the sensitivity compared to any single test alone in detecting UC with the specificity slightly decreased. Diagn. Cytopathol. 2013;41:852–857. © 2013 Wiley Periodicals, Inc.     

Cellular uptake of cyclotide MCoTI-I follows multiple endocytic pathways

Cyclotides are plant-derived proteins that naturally exhibit various biological activities and whose unique cyclic structure makes them remarkably stable and resistant to denaturation or degradation. These attributes, among others, make them ideally suited for use as drug development tools. This study investigated the cellular uptake of cyclotide, MCoTI-I in live HeLa cells. Using real time confocal fluorescence microscopy imaging, we show that MCoTI-I is readily internalized in live HeLa cells and that its endocytosis is temperature-dependent. Endocytosis of MCoTI-I in HeLa cells is achieved primarily through fluid-phase endocytosis, as evidenced by its significant colocalization with 10K-dextran, but also through other pathways as well, as evidenced by its colocalization with markers for cholesterol-dependent and clathrin-mediated endocytosis, cholera toxin B and EGF respectively. Uptake does not appear to occur only via macropinocytosis as inhibition of this pathway by Latrunculin B-induced disassembly of actin filaments did not affect MCoTI-I uptake and treatment with EIPA which also seemed to inhibit other pathways collectively inhibited approximately 80% of cellular uptake. As well, a significant amount of MCoTI-I accumulates in late endosomal and lysosomal compartments and MCoTI-I-containing vesicles continue to exhibit directed movements. These findings demonstrate internalization of MCoTI-I through multiple endocytic pathways that are dominant in the cell type investigated, suggesting that this cyclotide has ready access to general endosomal/lysosomal pathways but could readily be re-targeted to specific receptors through addition of targeting ligands.     

Identification of coffee components that stimulate dopamine release from pheochromocytoma cells (PC-12)

Coffee and caffeine are known to affect the limbic system, but data on the influence of coffee and coffee constituents on neurotransmitter release is limited. We investigated dopamine release and Ca(2+)-mobilization in pheochromocytoma cells (PC-12 cells) after stimulation with two lyophilized coffee beverages prepared from either Coffea arabica (AR) or Coffea canephora var. robusta (RB) beans and constituents thereof. Both coffee lyophilizates showed effects in dilutions between 1:100 and 1:10,000. To identify the active coffee compound, coffee constituents were tested in beverage and plasma representative concentrations. Caffeine, trigonelline, N-methylpyridinium, chlorogenic acid, catechol, pyrogallol and 5-hydroxytryptamides increased calcium signaling and dopamine release, although with different efficacies. While N-methylpyridinium stimulated the Ca(2+)-mobilization most potently (EC(200): 0.14±0.29μM), treatment of the cells with pyrogallol (EC(200): 48±14nM) or 5-hydroxytryptamides (EC(200): 10±3nM) lead to the most pronounced effect on dopamine release. In contrast, no effect was seen for the reconstituted biomimetic mixture. We therefore conclude that each of the coffee constituents tested stimulated the dopamine release in PC-12 cells. Since no effect was found for their biomimetic mixture, we hypothesize other coffee constituents being responsible for the dopamine release demonstrated for AR and RB coffee brews.     

Impact of risk factors on the performance of the nuclear matrix protein 22 point-of-care test for bladder cancer detection

OBJECTIVES

To evaluate the impact of different risk factors and symptoms on the performance characteristics of the nuclear matrix protein 22 (NMP22) BladderChek® test (Matritech Inc., Newton, MA, USA) for the detection of bladder cancer, as this is a point‐of‐care assay that measures NMPs in voided urine.

PATIENTS AND METHODS

In all, 23 academic, private practice and veterans’ facilities in 10 states prospectively enrolled consecutive patients from September 2001 to May 2002. Participants included 1328 patients at elevated risk of bladder cancer from factors such as a history of smoking or symptoms including haematuria and/or dysuria. Patients at risk of malignancy of the urinary tract provided a voided urine sample for analysis of NMP22 protein and cytology before cystoscopy.

RESULTS

Of the 1328 patients: no urinary disease, benign disease and malignancy were found in 545 (41%), 704 (53%) and 79 (6%) patients, respectively. Overall, the positive predictive value (PPV) for detection of bladder cancer was 20.3% (45/222) and negative predictive value (NPV) was 96.9% (1072/1106). The PPV was higher in men (24.0%) than women (13.2%). In men, the PPV increased with smoking (35.4%), gross haematuria (51.2%) and a combination of both factors (70.6%). The impact on the PPV of smoking (9.7%) and gross haematuria (28.6%) was less dramatic in women. The PPV increased from 16.8% in patients aged <65 years to 23.5% in those aged >65 years. The NPV remained almost always >95% except in men with gross haematuria where it decreased to 77% in smokers and 94% in non‐smokers.

CONCLUSION

The PPV of the BladderChek test improves in patients at higher risk of bladder cancer reaching 77% in men presenting with gross haematuria who are aged >65 years and smoke. The NPV is highest in women aged <65 years, up to 100%.     

核基质蛋白在膀胱癌中的研究进展

膀胱癌是人类最常见的恶性肿瘤之一。其复发率高,尤其是高级别膀胱癌,早期诊断和规律随访是减少病患死亡率、提高生活质量的主要措施。尿脱落细胞和膀胱镜检查是传统的诊断膀胱癌及监测术后复发的"金标准"。近年来,随着分子生物学的飞速发展,众多膀胱癌肿瘤标志物得以发现和研究,其中核基质蛋白(NMPs)因高度敏感性和特异性、简便无创而受到临床广泛关注。本文就膀胱癌新型肿瘤标志物NMPs的研究进展作一综述。     

NPM1基因突变调控THP-1细胞体外增殖和侵袭及其机制

目的探讨NPM1基因突变对THP．1细胞体外增殖和侵袭的影响及其机制。方法将THP．1细胞分为THP-1．mA组、空载体转染组和未处理组,THP-1．mA组使用携带人NPMl-mA的重组质粒pEGFPCI．NPMl．mA转染THP-1细胞,建立稳定表达NPM1．mA的白血病细胞系（THP-1-mA）;空载体转染组使用空载体质粒pEGFPCI转染THP-1细胞;未处理组不进行质粒转染。采用反转录PCR、免疫细胞化学检测3组细胞NPMl．mA基因和蛋白的表达;使用细胞生长曲线观察细胞体外增殖能力;流式细胞术检测细胞周期分布;细胞体外侵袭实验观察细胞体外侵袭能力;实时荧光定量PCR检测血管生成素1（Ang-1）、Ang．2mRNA的表达。结果成功构建了稳定表达NPMl．mA的白血病细胞株。与空载体转染组和未处理组比较,稳定表达NPMl。mA蛋白的THP-1．mA细胞体外增殖能力明显增强,G1期细胞比例减少,S期细胞比例增加（均P〈0．01）。与空载体转染组和未处理组比较,细胞体外侵袭实验显示THP．1-mA组细胞体外侵袭能力增强,实时荧光定量PCR显示THP．1．mA组细胞Ang．1mRNA表达增高（均P〈0．01）。结论NPMl突变基因的表达能够促进THP-1细胞体外增殖和侵袭,而Ang-1可能在其中发挥重要作用。     

NMP22诊断膀胱移行细胞癌价值的评价

目的 :探讨核基质蛋白 2 2 (NMP2 2 )对膀胱移行细胞癌的临床诊断价值。方法 :选取 5 8例膀胱移行细胞癌患者为实验组 ,2 0例泌尿系统非尿路上皮肿瘤疾病的患者为对照组 1,10名健康志愿者为对照组 2 ,应用酶联免疫法测定三组受试者尿NMP2 2的含量。结果 :实验组尿NMP2 2中位含量为 2 3 .2× 10 3U/L ,明显高于对照组 1的 8.3× 10 3U/L和对照组 2的 2 .6× 10 3U/L (P <0 .0 1) ;尿NMP2 2检测膀胱移行细胞癌的敏感性为79% ,高于尿细胞学的 3 8% (P <0 .0 5 ) ;NMP2 2的阳性率与膀胱移行细胞癌的分期、分级以及肿瘤的形状、大小和数目无显著相关性。结论 :尿NMP2 2对膀胱癌的诊断有较高的敏感性 ,简便、无创 ,检测结果不受肿瘤的分化程度、生长方式、肿瘤大小和数目的影响     

尿NMP22及Cox-2蛋白对表浅性膀胱癌术后复发的预判表达及临床意义

目的 比较尿NMP22和Cox-2蛋白在表浅性膀胱癌患者治疗前后的表达水平,探讨二者在表浅性膀胱癌诊断与随访中的临床应用价值.方法 选择40例表浅性膀胱移行细胞癌和40例其他泌尿系统疾病患者,检测术前晨尿NMP22及Cox-2蛋白的表达水平.40例浅表性膀胱癌患者均行膀胱肿瘤电切术(TURBt),术后予规律灌注化疗及膀胱镜检随访.每次行膀胱镜检前,留取尿液检测尿NMP22及Cox-2蛋白的表达水平,比较术后二者阳性表达与膀胱镜检发现肿瘤复发时间的长短.结果 表浅性膀胱癌患者中尿NMP22蛋白和Cox-2的敏感性分别为62.5％和80.0％,特异性分别为85.0％和95.0％,Youden指数分别为47.5％和75.0％.尿Cox-2蛋白的敏感性和Youden指数均高于NMP22(P＜0.05).术后随访,NMP22及Cox-2蛋白阳性表达的时间较膀胱镜检发现肿瘤复发时间平均早4.4及5.8个月.结论 尿NMP22和Cox-2蛋白是诊断和随访膀胱移行细胞癌高特异性及敏感性的指标.     

NMP22在泌尿系统肿瘤中的临床应用研究

为了探讨尿液中核基质蛋白(NMP22)对泌尿系统恶性肿瘤诊断的临床意义.本文应用ELISA测定271例泌尿系恶性肿瘤患者和43例泌尿系良性病变患者尿液NMP22含量.结果表明: 检测NMP22对尿路上皮恶性肿瘤的敏感性为89.2%,特异性为93.3%.尿路上皮恶性肿瘤、非尿路上皮恶性肿瘤和泌尿系良性病变患者, 尿NMP22阳性率分别为: 89.2%、37.2%和34.5%, 其中膀胱移行细胞癌89.5%.泌尿系恶性肿瘤比良性病变阳性率明显增高(P＜0.05),尿路上皮恶性肿瘤比非尿路上皮恶性肿瘤阳性率增高(P＜0.05).另外, 尿中NMP22含量与膀胱移行细胞癌分期、分级和肿瘤生长方式以及瘤体数目均无显著差异(P＞0.05).提示尿液NMP22检测对诊断尿路上皮恶性肿瘤有较高的敏感性, NMP22对泌尿系肿瘤良、恶性鉴别诊断也有一定意义.