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Size-Based Enrichment of Exfoliated Tumor Cells in Urine Increases the Sensitivity for DNA-Based Detection of Bladder Cancer  [PDF]
Elin Andersson, Kenneth Steven, Per Guldberg
PLOS ONE , 2014, DOI: 10.1371/journal.pone.0094023
Abstract: Bladder cancer is diagnosed by cystoscopy, a costly and invasive procedure that is associated with patient discomfort. Analysis of tumor-specific markers in DNA from sediments of voided urine has the potential for non-invasive detection of bladder cancer; however, the sensitivity is limited by low fractions and small numbers of tumor cells exfoliated into the urine from low-grade tumors. The purpose of this study was to improve the sensitivity for non-invasive detection of bladder cancer by size-based capture and enrichment of tumor cells in urine. In a split-sample set-up, urine from a consecutive series of patients with primary or recurrent bladder tumors (N = 189) was processed by microfiltration using a membrane filter with a defined pore-size, and sedimentation by centrifugation, respectively. DNA from the samples was analyzed for seven bladder tumor-associated methylation markers using MethyLight and pyrosequencing assays. The fraction of tumor-derived DNA was higher in the filter samples than in the corresponding sediments for all markers (p<0.000001). Across all tumor stages, the number of cases positive for one or more markers was 87% in filter samples compared to 80% in the corresponding sediments. The largest increase in sensitivity was achieved in low-grade Ta tumors, with 82 out of 98 cases positive in the filter samples (84%) versus 74 out of 98 in the sediments (75%). Our results show that pre-analytic processing of voided urine by size-based filtration can increase the sensitivity for DNA-based detection of bladder cancer.
Urine and bladder washing cytology for detection of urothelial carcinoma: standard test with new possibilities
Margareta Strojan Flezar
Radiology and Oncology , 2010, DOI: 10.2478/v10019-010-0042-8
Abstract: Background. Light microscopic evaluation of cell morphology in preparations from urine or bladder washing containing exfoliated cells is a standard and primary method for the detection of bladder cancer and also malignancy from other parts of the urinary tract. The cytopathologic examination is a valuable method to detect an early recurrence of malignancy or new primary carcinoma during the follow-up of patients after the treatment of bladder cancer. Conclusions. Characteristic cellular and nuclear signs of malignancy indicate invasive or in situ urothelial carcinoma or high-grade papillary urothelial carcinoma. However, low sensitivity of the method reflects the unreliable cytopathologic diagnosis of low-grade urothelial neoplasms as cellular and nuclear signs of malignancy in these neoplasms are poorly manifested. Many different markers were developed to improve the diagnosis of bladder carcinoma on urinary samples. UroVysion test is among the newest and most promising tests. By the method of in situ hybridization one can detect specific cytogenetic changes of urothelial carcinoma.
Detection of human telomerase reverse transcriptase messenger RNA in urine of high risk patients as a non invasive molecular diagnostic tool for urinary bladder cancer in Egyptians
SF Helal, HSH Kassem, WM Sameh, SN El- Achy
Egyptian Journal of Medical Human Genetics , 2009,
Abstract: Introduction and Objectives: Bladder cancer is an important national health problem as it is the leading cancer in men in Egypt. Cystoscopy and biopsy, currently remains the gold standard procedure for diagnosis, yet, it is invasive and costly. Urinary cytopathology remains to be the only non-invasive alternative method for diagnosis. Although it is tumour specifi c, yet it has a poor sensitivity, especially for low grade tumours. Detection of Telomerase enzyme in exfoliated urinary cells is a potentially good molecular diagnostic marker in bladder cancer, since the catalytic subunit of this enzyme (hTERT) proved to be essential for cellular immortality and oncogenesis. Subjects and Methods: The study comprised 39 patients (36 with urothelial carcinomas and 3 cases were squamous cell carcinoma) with bladder cancer and 22 non cancer control (including 14 patients with benign urological disorders and 8 healthy volunteers). The urine sample was split into two aliquots one was used to undertake RNA extraction and hTERT/GAPDH RT-PCR semiquantitative assay and the second for cytological examination. Cystoscopy was considered the reference standard for the identifi cation of bladder cancer. Results: The hTERT/GAPDH RT-PCR test showed signifi cantly higher diagnostic sensitivity than cytology (84% Vs. 75% p<0.008) for confi rmed UCC, particularly for low grade non-muscle invasive UCC (82% Vs. 64% p<0.005). On combining the two tests a sensitivity of 95% was obtained. A positive hTERT expression was detected 4-5 months earlier than cystoscopic evidence of recurrence in 2 patients during their follow up. Conclusion: In this pilot study, detection of hTERT expression in urine has shown to be a more sensitive marker for diagnosis of bladder cancer than cytology. The combination of urinary hTERT mRNA with cytological testing augments the sensitivity for the non-invasive early diagnosis of bladder cancer. This finding warrants further extended study to validate the potential role of hTERT expression as a diagnostic non invasive tool for high risk patients and detection of recurrence in bladder cancer in Egypt.
Clinical significance of adenoviral infections
Brki? Sne?ana V.,Vukobratov Zita,Preveden Tomislav A.,Jovanovi? Jovana R.
Medicinski Pregled , 2002, DOI: 10.2298/mpns0208337b
Abstract: Introduction The aim of this article was to point to ubiquitous adenoviral infections and to give a literature overview. Adenoviral infections present with a variety of clinical manifestations, causing many differential diagnosis problems. Diagnosis and epidemiology In our country diagnosis is made using the complement fixation test (CFT), which detects antibodies due to solubile group specific antigen. In acute infections, we need two sera samples given in 2 weeks period with 4-fold rise or fall in titers. Due to many asymptomatic infections which can give increased titer by CFT, many physicians think that patients have a persistent infection. Persistent adenoviral infections need not be accompanied by any special clinical symptomatology. However, adenoviral infections still play an important role in acute respiratory infections. The most severe respiratory infection is pneumonia which can be associated with acute respiratory distress syndrome and death. Disseminated adenoviral diseases appear in 2.5% of all adenoviral infections with the same percentage between immunocompetent and immunocompromised persons. In immunocompromised persons adenoviral infections manifest as haemorrhagic cystitis, fulminant or acute hepatitis or meningoencephalitis. Therapy and conclusion We still don't have a special treatment for these kinds of infections. Variety of antiviral drugs with controversial effects have been reported in management of adenoviral infections in immunocompromised persons. In USA adenoviral vaccine has been excluded from vaccine schedules among military personnel, but morbidity and the first two deaths due to these infections in the last 30 years reported by MMWR in 2000, may change this policy.
Epigenetic Markers for Bladder Cancer in Urine
Wun-Jae Kim and Yong-June Kim
Translational Oncogenomics , 2012,
Abstract: Many tumor markers for bladder cancer have been evaluated for use in detecting and monitoring bladder cancers tissue specimens, bladder washes, and urine specimens. However, none of the biomarkers reported to date has shown sufficient sensitivity and specificity to detect the entire spectrum of bladder cancers in routine clinical practice. The limited value of the established prognostic markers demands analysis of new molecular parameters having the potential to predict the prognosis of bladder cancer patients, particularly, the high-risk patients at risk of cancer progression and recurrence. Abnormal methylation of CpG islands can efficiently repress transcription of the associated gene in a manner akin to mutations and deletions. Several tumor suppressor genes correlated with bladder cancer contain CpG islands in their promoters. Markers for aberrant methylation may be a potential gateway for monitoring bladder cancer. Hypermethylation of several gene promoters was detected in urine sediment DNA from bladder cancer patients. Detection of DNA methylation in voided urine is feasible and noninvasive. Methylation is an important molecular mechanism in the development of bladder cancer and could be used as a prognostic and diagnostic marker. Aberrant patterns of epigenetic modification could, in the near future, be crucial indicators in cancer diagnosis, prognosis, and may additionally be good targets for developing novel therapies while maintaining quality of life.
Epigenetic Markers for Bladder Cancer in Urine
Wun-Jae Kim,Yong-June Kim
Translational Oncogenomics , 2007,
Abstract: Many tumor markers for bladder cancer have been evaluated for use in detecting and monitoring bladder cancers tissue specimens, bladder washes, and urine specimens. However, none of the biomarkers reported to date has shown sufficient sensitivity and specificity to detect the entire spectrum of bladder cancers in routine clinical practice. The limited value of the established prognostic markers demands analysis of new molecular parameters having the potential to predict the prognosis of bladder cancer patients, particularly, the high-risk patients at risk of cancer progression and recurrence. Abnormal methylation of CpG islands can efficiently repress transcription of the associated gene in a manner akin to mutations and deletions. Several tumor suppressor genes correlated with bladder cancer contain CpG islands in their promoters. Markers for aberrant methylation may be a potential gateway for monitoring bladder cancer. Hypermethylation of several gene promoters was detected in urine sediment DNA from bladder cancer patients. Detection of DNA methylation in voided urine is feasible and noninvasive. Methylation is an important molecular mechanism in the development of bladder cancer and could be used as a prognostic and diagnostic marker. Aberrant patterns of epigenetic modification could, in the near future, be crucial indicators in cancer diagnosis, prognosis, and may additionally be good targets for developing novel therapies while maintaining quality of life.
Adenoviral Producer Cells  [PDF]
Imre Kovesdi,Susan J. Hedley
Viruses , 2010, DOI: 10.3390/v2081681
Abstract: Adenovirus (Ad) vectors, in particular those of the serotype 5, are highly attractive for a wide range of gene therapy, vaccine and virotherapy applications (as discussed in further detail in this issue). Wild type Ad5 virus can replicate in numerous tissue types but to use Ad vectors for therapeutic purposes the viral genome requires modification. In particular, if the viral genome is modified in such a way that the viral life cycle is interfered with, a specific producer cell line is required to provide trans-complementation to overcome the modification and allow viral production. This can occur in two ways; use of a producer cell line that contains specific adenoviral sequences incorporated into the cell genome to trans-complement, or use of a producer cell line that naturally complements for the modified Ad vector genome. This review concentrates on producer cell lines that complement non-replicating adenoviral vectors, starting with the historical HEK293 cell line developed in 1977 for first generation Ad vectors. In addition the problem of replication-competent adenovirus (RCA) contamination in viral preparations from HEK293 cells is addressed leading to the development of alternate cell lines. Furthermore novel cell lines for more complex Ad vectors and alternate serotype Ad vectors are discussed.
Diagnosis of Bladder Cancer by Urine Survivin, an Inhibitor of Apoptosis: A Preliminary Report  [cached]
Seyed Amirmohsen Ziaee,Seyed Javad Moula,Seyed Mohammadmehdi Hosseini Moghaddam,Darioush Eskandar-Shiri
Urology Journal , 2006,
Abstract: Introduction: Survivin is an inhibitor of apoptosis that is expressed in undifferentiated tissues like tumors. Detection of survivin in urine has been proposed as a diagnostic marker for bladder cancer. We evaluated the urine samples of patients with bladder cancer for survivin and compared them with healthy controls. Materials and Methods: The urine specimens of 20 patients with transitional cell carcinoma (TCC) of the bladder (group 1) and 18 controls without cancer (group 2) were collected before cystoscopy and assessed for survivin by reverse transcriptase polymerase chain reaction. Results: All patients except 1 in group 1 were men. Urine specimens were positive for survivin in 18 (90%) and 9 (50%) patients of groups 1 and 2, respectively (P = .007). Sixteen patients with TCC had urine cytology, of which 6 (37.5%) were positive. Urine survivin was positive in all 10 patients with negative cytology. Nine patients in this group had low-grade tumors. Conclusion: Urine survivin seems to have a higher sensitivity than urine cytology, especially in low-grade bladder cancer. The quantitative measurement of survivin in urine by advanced techniques may provide a better diagnostic and prognostic tool. However, the clinical use of survivin and its association with different stages and grades of TCC still requires more studies.
Frictional characteristics of exfoliated and epitaxial graphene  [PDF]
Young Jun Shin,Ryan Stromberg,Rick Nay,Han Huang,Andrew T. S. Wee,Hyunsoo Yang,Charanjit S. Bhatia
Physics , 2012,
Abstract: To determine the friction coefficient of graphene, micro-scale scratch tests are conducted on exfoliated and epitaxial graphene at ambient conditions. The experimental results show that the monolayer, bilayer, and trilayer graphene all yield friction coefficients of approximately 0.03. The friction coefficient of pristine graphene is less than that of disordered graphene, which is treated by oxygen plasma. Ramping force scratch tests are performed on graphene with various numbers of layers to determine the normal load required for the probe to penetrate graphene. A very low friction coefficient and also its high pressure resistance make graphene a promising material for antiwear coatings.
Synthesis and Characterization of Exfoliated Graphite/ABS Composites  [PDF]
Bhardwaj Neha, K. S. Manjula, B. Srinivasulu, Shit C. Subhas
Open Journal of Organic Polymer Materials (OJOPM) , 2012, DOI: 10.4236/ojopm.2012.24011
Abstract: Exfoliated graphite was prepared by chemical route and then further subjected to thermal oxidation and sonication for size reduction and increased interlayer spacing in natural flake graphite. Exfoliated graphite/ABS composites were pre-pared with varying filler concentration by solution casting method. Exfoliated graphite and composites were character-ised by scanning electron microscopy and energy dispersive X-ray analysis, Fourier transform infrared spectroscopy and X-ray diffractometry techniques. After thermal exfoliation significant peak of graphite at 2? = 26.4? disappeared completely, confirming successful exfoliation of graphite. SEM images revealed homogeneous dispersion of exfoliated graphite in the matrix and EDAX confirmed successful reduction of graphite oxide.
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