Comparison of the f-HPV typing™ and Hybrid Capture II® assays for detection of high-risk HPV genotypes in cervical samples

Human papillomavirus genotyping is being considered in cervical screening programs and for monitoring the effectiveness of HPV vaccination. Both approaches require access to fast, easy and high-throughput technology. The aim of this study was to compare a new commercial assay (f-HPV typing?) with the Hybrid Capture II? (HC2) to detect HPV infection. The F-HPV typing is a multiplex fluorescent PCR method recognizing E6 and E7 regions of 13 high-risk (HR) HPV types, the same set of HR-types targeted HC2 test. A subset of 157 cervical samples was tested with both assays. The percentage of positive HR-HPV DNA samples was 24% (37/155) by HC2 and 33% (49/155) by f-HPV typing. Concordant results were found in 133/155 (overall agreement, 85.8%; Cohen's kappa=0.65). The analytical sensitivity and specificity of f-HPV were 97.6 and 93, respectively. In conclusion, this study shows that the f-HPV assay provides a good alternative to HC2 to detect HPV infection, allowing simple and rapid HPV genotyping and detecting multiple infections.     

Is there a "bladder sex"? The relation of different sex hormones and sex hormone receptors in bladder in childhood

The aim of this study was to review the classical concept of bladder physiology in regard to sex hormone effects and to focus on a new concept. Sex hormones affect bladder functions and the classical concept tries to explain these effects via alpha and beta adrenoreceptors. The effects of hormones had been investigated in this understanding. To obtain a biological response to steroids in target tissues, specific proteins--so-called receptors--are warranted. After the demonstration of the sex hormone receptors in bladder first in adults and in children urge questioning the effects of hormones in this regard. The hypothesis is based on the fact that, if a special receptor is present, the direct (via sex hormone receptors) effect is more likely to occur as an indirect effect (via alpha and beta adrenoreceptors). There is evidence that there is a sex difference and the sex receptors and hormones play an important role in some bladder disorders. Therefore we stress the importance of investigating especially the pediatric patients with bladder problems in regard to sex hormone receptors and hormonal status.     

Glutathione S-transferase M1 gene polymorphism in bladder cancer patients. a marker for invasive bladder cancer?

The glutathione S-transferase M1 (GSTM1) gene is polymorphic in humans, and the deficiency in enzyme activity of GSTM1 is caused by the inherited homozygous absence of the GSTM1 gene, or the "null" genotype (GSTM1, 0/0). The increased risk of bladder cancer has been shown to correspond with this gene defect. No reports, however, have been found in the literature regarding GSTM1 gene deficiency with superficial and invasive bladder cancer. In this study, we examined the association of the GSTM1-null genotype with superficial and invasive bladder cancer. Using a polymerase chain reaction (PCR)-based method, we examined the frequency of the GSTM1 gene defect in Turkish patients with superficial bladder cancer (N = 61), invasive bladder cancer (N = 42), and control subjects (N = 202) who had no history of cancer. The GSTM1 null genotype was observed in 34.7% of the control subjects and in 54.3% of total bladder cancer patients (OR: 2.246; 95% CI: 1.384-3.645, P: 0.00094). In other words, the presence of the GSTM1-null genotype significantly increased the risk of bladder cancer development. Among invasive bladder cancers, the frequency of the GSTM1-null genotype was 64.3% (OR: 0.294, 95% CI: 0.147-0.590, P: 0.0003). This was also significantly higher than control subjects, indicating that patients carrying this genotype were at increased risk for developing invasive bladder cancer. This relationship was not statistically significant in the superficial bladder cancer group (OR: 0.585, 95% CI: 0.327-1.045, P: 0.06). Our results indicate that GSTM1 gene polymorphism should be considered as an important risk modifier in the development of bladder cancer and might be used as a predictive marker for invasive bladder cancer.     

Are volunteers delivering semen samples in fertility studies a biased population?

BACKGROUND: It is not known to what extent the results of epidemiological studies on male fertility and semen quality based on voluntary participation in the general population are relevant. METHODS: In a study on the reproductive health of male partners of pregnant women, information was obtained from a group of men agreeing to collect a semen sample and to complete a questionnaire (group A), a group only completing the questionnaire (group B) and from men refusing to participate altogether (group C). RESULTS: The participation rate (group A) was 15.8% for 1409 men approached. Ages and socio-professional status were similar in the three groups. Time to pregnancy (TTP) was not significantly different in groups A and B, although there appeared to be an insignificantly higher proportion of couples taking longer than 12 months to conceive in group A than in group B. A history of urogenital disease appeared to be more frequent in groups A and B than in the general population. However, comparable semen characteristics were found for men with or without a history of urogenital disease. Pregnancy outcomes were similar in the three groups. CONCLUSIONS: The present study does not suggest major selection bias, although the social and reproductive histories of these men may prompt them to participate. Such factors need to be accounted for in similar studies.     

Does polyomavirus infection interfere with bladder cancer fluorescence in situ hybridization?

Urine cytology is a proven and widely used screening tool for the detection of urothelial carcinoma. However, morphologic features of polyomavirus infected cells, characterized by nuclear inclusions (decoy cells) are a known source of diagnostic confusion with malignancy. Fluorescence in situ hybridization (FISH) is now routinely used to support the cytological diagnosis of urothelial carcinoma and monitor for recurrence. We sought to determine whether polyomavirus infection could result in positive FISH results (aneuploidy). This study deals with retrospective study of 100 polyomavirus‐infected urine samples from patients with no history of urothelial carcinoma or organ transplantation. All cases were stained with Papanicolaou and acid hematoxylin stain. One slide from each sample was de‐stained and FISH was performed using chromosome enumeration probes 3, 7, 17, and locus‐specific probe 9p21. Adequate cells for FISH analysis (25 cells) were present in 81 cases; 19 cases were insufficient due to loss of cells during de‐staining and FISH preparation process. All polyomavirus‐infected cells (decoy cells) exhibited a normal chromosome pattern. Four cases were FISH positive, but there were no positive decoy cells. Decoy cells did not exhibit aneuploidy by FISH. The presence of decoy cells does not exclude the possibility of concurrent urothelial carcinoma. Acid hematoxylin stain appeared to supplement the Papanicolou stain in identifying and confirming the presence of polyomavirus infection. Diagn. Cytopathol. 2014;42:225–229. © 2013 Wiley Periodicals, Inc.     

Aggregation of α-synuclein in brain samples from subjects with glucocerebrosidase mutations

Recent studies show an increased frequency of mutations in the glucocerebrosidase gene (GBA1) in patients with α-synucleinopathies including Parkinson disease. Some patients with Gaucher disease (GD) develop parkinsonism with α-synuclein-positive inclusions post mortem. Proteins were extracted from the cerebral cortex of subjects with synucleinopathies with and without GBA1 mutations, controls and patients with GD. Patients with GBA1-associated synucleinopathies showed aggregation of oligomeric forms of α-synuclein in the SDS-soluble fraction, while only monomeric forms of α-synuclein were seen in subjects with GBA1 mutations without parkinsonism. Thus, brains from patients with GBA1-associated parkinsonism show biochemical characteristics typical of Lewy body disorders.     

Bias in binge eating disorder: how representative are recruited clinic samples?

The aim of this study was to investigate sampling bias as it affects recruited clinic samples of binge eating disorder (BED). Demographic and clinical characteristics of a recruited clinic sample were compared with a community sample. The 2 groups met the same operational definition of BED and were assessed using the same primarily interview-based methods. Ethnicity, severity of binge eating, and social maladjustment were found to increase treatment seeking among participants with BED rather than levels of psychiatric distress or comorbidity. These findings suggest that previous studies using recruited clinic samples have not biased estimates of psychiatric comorbidity in BED.     

Langerhans cell histiocytosis of the urinary bladder in a patient with bladder cancer previously treated with intravesical Bacillus Calmette–Guérin therapy

We report an extremely rare case of Langerhans cell histiocytosis (LCH) of the urinary bladder. A 68-year-old man presented with gross hematuria. Cystoscopy showed multiple papillary tumors in the urinary bladder, and transurethral resection was performed. Pathological diagnosis was high-grade papillary urothelial carcinoma with lamina propria invasion. The patient received six treatments with intravesical Bacillus Calmette–Guérin (BCG) therapy. Seven months after surgery, follow-up cystoscopy showed three elevated lesions in the urinary bladder, two of which were identified histologically as recurrent urothelial carcinoma. Microscopic examination of the lesion at the anterior wall revealed diffuse infiltration of medium to large histiocytoid cells in the lamina propria, many of which had distorted nuclei and nuclear grooves. Dense eosinophilic infiltration was also observed. Immunohistochemically, the histiocytoid cells were diffusely positive for S-100 and CD1a, but negative for cytokeratin AE1/AE3 and melanosome-associated antigen recognized by HMB-45. Based on the histological and immunohistochemical features, we diagnosed the lesion as LCH of the urinary bladder. There was no evidence of recurrence of either bladder cancer or LCH after an 18-month follow-up. To avoid misdiagnosis, urologists and pathologists should be aware that LCH may develop in the urinary bladder after intravesical BCG therapy for bladder cancer.     

How to test at once six cytokines in samples as small as 25 microl?

Inflammatory and regulatory or anti-inflammatory cytokines (TNFalpha, IL-1beta, -6, -8, -10 and -12) regulate both the humoral and cellular immune responses. Cytokines have diverse peripheral and central functions. They are critical mediators of protective host responses, including defense against microbial invasion and tumorigenesis. However, the production of specific proinflammatory cytokines must be tightly regulated and compartmentalized to prevent the overexpression of these molecules that can end in chronic inflammation and tissue injury. Many diseases like autoimmune disease (rheumatoid arthritis, multiple sclerosis, arteriosclerosis, Crohn's disease), neurodegenerative disease (Alzheimer's and Parkinson's disease), tumor invasion and metastasis correlate with a deregulation in cytokine action. Thus, cytokines network provides an attractive and intensely competitive area of potential targets for therapeutic intervention. To monitor such secretion patterns in presence of putative drugs obtained by high throughput screening (HTS) some new techniques recently appeared on the market. We here compared results obtained by CBA (BD Cytometric Bead Array) to IC50 values obtained by classical sandwich Elisa. The complexity and cost of this new method is largely compensated by simultaneous testing of 6 cytokines in only 25 micro L of cell supernatant.     

Vdelta2 T-lymphocyte responses in cord blood samples from Italy and Côte d'Ivoire

Cord blood T lymphocytes are immature and their functional defect partially reflects a suboptimal level of costimulatory signals provided by neonatal antigen-presenting cells. Neonatal Vδ2 T lymphocytes, a small component of cellular immunity involved in the response against bacteria, protozoa, virus-infected cells and tumours, are also considered to be immature. Cord blood Vδ2 T lymphocytes are mostly naïve, proliferate poorly and do not produce cytokines in response to the model phosphoantigen isopentenyl pyrophosphate. We cultured cord blood mononuclear cells with the aminobisphosphonate Pamidronate or with live bacille Calmette–Guérin, and showed that both elicit a strong cord blood Vδ2 T-cell proliferative response, inducing the expression of activation markers and promoting the differentiation from naïve to memory cells. Our results suggest that cord blood Vδ2 T cells are not inherently unresponsive and can mount strong responses to aminobisphosphonates and mycobacteria. Neonatal Vδ2 T lymphocytes may be important participants in responses to microbial infections early in life.     

REBA HPV-ID® for efficient genotyping of human papillomavirus in clinical samples from Korean patients

This study was conducted to evaluate the overall performance of a reverse blot hybridization‐based assay, REBA HPV‐ID® (Molecules and Diagnostics, Wonju, Korea) for genotyping human papillomaviruses (HPV). HPV Genotyping on 356 specimens examined cytologically was performed using the REBA HPV‐ID®, and its results were compared with those obtained using the MyHPV DNA Chip® (Mygene, Seoul, Korea), DNA chip‐based HPV genotyping assay. The results from this study showed that the positivity rate of the REBA HPV‐ID® for abnormal cytological samples was higher (80.9%) than that of the MyHPV DNA chip (69.8%). In addition, the REBA HPV‐ID® positivity rate with normal cytological samples was higher (64.4%) than that obtained using DNA chips (34.4%). Subsequently, sequence analysis was performed with specimens that generated conflicting test results. Sequence analysis confirmed that the specimens which were positive by REBA HPV‐ID® did indeed contain HPV sequences. The results of this study suggest that the REBA HPV‐ID® is a sensitive test for genotyping HPV of clinical specimens. J. Med. Virol. 84: 1248–1253, 2012. © 2012 Wiley Periodicals, Inc.     

Tissue expression of TGF-β1 in uterine cervical samples from HIV/AIDS patients

Case-control study based on the immunohistochemistry for TGF-β1 evaluation of cervical samples obtained from two groups of women: CIN/HIV− and CIN/HIV+. Eleven women infected with HIV and with a histopathological diagnosis of CIN were included. The control group consisted of 12 patients with CIN. Cervical tissue samples obtained from all patients were submitted to histopathology and semiquantitative analysis of immunostaining for TGF-β1 protein. In addition, the peripheral CD4+ cell count and viral load were evaluated in HIV + patients. Tissue expression of the cytokine was higher in the CIN/HIV+ group compared to control (p = 0.0023). In addition, higher TGF-β1 expression was observed in higher grade cervical lesions in the two groups. There was a trend toward a direct correlation between peripheral CD4+ T cell count and tissue TGF-β1, and toward an inverse correlation between viral load and cytokine expression. Thus, TGF-β1 was more marked in situations in which cervical lesions are known to present a more aggressive behavior, suggesting that this cytokine is involved in the pathogenesis of tumor growth in these lesions. Tissue expression of TGF-β1 is increased in cervical samples from HIV-infected women with CIN.     

Blastocystis surface antigen is stable in chemically preserved stool samples for at least 1 year

Blastocystis spp. refer to a group of prevalent enteric protists found in humans and animals. Detection of Blastocystis spp. in fecal samples is often performed by clinicians with direct microscopy, which provides low sensitivity, or with culture and polymerase chain reaction testing, a method which is problematic when used with formalin-preserved stool samples. Prior study of Blastocystis and other enteric protists suggests that immunofluorescence antibody (IFA) stain could provide sensitivity and compatibility with formalin, but no information is available on the longevity of Blastocystis sp.’s surface antigens in formalin. We collected fecal samples from animals at a country fair held in the summer of 2009 in Oregon, USA. Samples were tested for the presence of Blastocystis infection using an IFA stain shortly after collection, and again after 1 year, with samples stored refrigerated at 4–8 °C. Most samples collected from steer, pigs, and goats were found to be Blastocystis positive. All fecal samples that were Blastocystis positive initially remained positive after 1 year. Blastocystis-negative samples remained negative. Minimal degradation was observed in stained slides. Blastocystis surface antigens detected by a polyclonal stain remained stable in formalin for a period of at least 1 year.     

Loss of nuclear prothymosin-α expression is associated with disease progression in human superficial bladder cancer

In this paper, we report a study on the clinical relevance of prothymosin-α expression and its correlation with intratumoral Foxp3⁺ and CD8⁺ lymphocytes (Foxp3⁺TIL and CD8⁺TIL) in bladder cancer patients. We used immunohistochemical staining for prothymosin-α, Foxp3, and CD8 on 101 tumor specimens harvested by endoscopic resection. The results were correlated with clinicopathological variables and clinical outcome in bladder cancer patients, particularly in 73 patients with superficial disease, using the log-rank test and Cox proportional hazard model. Overall, of the tumors, 30 % were negative, 34 % showed nuclear, and 37 % showed cytoplasmic prothymosin-α expression. Foxp3⁺TILs were detected in 11 % of patients (nonnuclear vs. nuclear, p?=?0.096). Patients with a history of urothelial carcinoma have a higher frequency of nonnuclear prothymosin-α expression than those without (p?=?0.016, chi-square test). By univariate and multivariate analyses of cases with superficial disease, grade and stage were identified as independent predictors for recurrence-free survival (p?=?0.016 and 0.016, respectively). Higher stage and nonnuclear prothymosin-α expression independently predict shorter progression-free survival (p?=?0.006 and 0.043, respectively). The presence of Foxp3⁺TILs was significantly associated with disease progression by univariate analysis (p?=?0.022), but not by multivariate analysis (p?=?0.147). In vitro assays showed that J82 cells which express ectopically nuclear prothymosin-α exhibit higher growth rate and secrete less TGF-β1 than those with cytoplasmic expression or control cells. Altogether, prothymosin-α expression is a determinant of disease progression in superficial bladder cancer. Foxp3⁺TILs tend to be found more often in bladder cancer with nonnuclear prothymosin-α expression. Future study is required to unravel their interaction.     

Inhibition of bacillus Calmette-Guérin-induced nitric oxide in bladder tumor cells may improve BCG treatment

Bacillus Calmette-Guérin (BCG) is considered to be one of the most effective treatments for superficial and in situ bladder cancer. However, either failure to respond initially or relapse within the first 5 years of treatment has been observed in some patients. As nitric oxide (NO) has been detected in the bladder of BCG-treated patients, we analyzed the role of endogenous NO generated after BCG treatments on human (T24) and murine (MB49 and MBT2) bladder tumor cells in the viability of tumor and immune cells, both in vitro and in vivo. In vitro inhibition of cancer cells after BCG treatment was evaluated by cell titer assay. NO production was determined as nitrite by Griess reagent. The death of immunocytes was evaluated by 51Cr release. Tumor histology with hematoxylin and eosin and Masson's trichrome staining was performed. BCG induced a direct inhibition of tumor cell growth in vitro, independently of NO levels. Besides, BCG-mediated NO production by tumor cells induced the death of spleen and peritoneal cells in syngeneic mice. The in vivo inhibition of NO synthase (NOS) activity by NG-nitro-L-arginine methyl ester in combination with BCG, improved tumor regression by generating a healing tissue. The increase of NO generated after BCG administration may induce the death of immunocytes. The in vivo inhibition of NO ameliorated immunotherapy with BCG by additional tumor growth inhibition. Our results suggested the possibility that the final outcome of patients with bladder tumors may improve by modulating NOS activity concomitantly with BCG therapy.     

Can bladder adenocarcinomas be distinguished from schistosomiasis-associated bladder cancers by using array comparative genomic hybridization analysis?

Bladder cancer is the most common malignancy in many tropical and subtropical areas, correlating well with the endemicity of schistostomiasis. The majority of schistostomiasis-associated (SA) bladder cancers are squamous cell cancers, whereas the majority of non-SA cases in the Western world are transitional cell cancers, suggesting different carcinogenetic mechanisms. Approximately 6% of SA and 1% of non-SA cases are adenocarcinomas. To achieve fine-resolution information of DNA copy number changes in SA adenocarcinomas, 10 tumor samples were analyzed on an oligonucleotide-based CGH array. The frequency of aberrations ranged from 2 to 17, with an average of 10 alterations per sample. The most frequently gained regions were 20q and 8q (in 70 and 60% of the cases, respectively), whereas the most frequently lost regions were 5q and 8p (both in 40% of the cases). In addition, six regions of amplification were found in three samples, containing both well characterized and novel regions. Comparison of the DNA copy number profiles to previously reported profiles of SA transitional cell carcinoma and squamous cell carcinoma revealed similarities (e.g., gains at 5p and 8q), as well as differences (e.g., TCC- and SCC-associated losses at 18p and 20p, and adenocarcinoma-associated gains at 20q). The results suggest that although SA cancers share genetic features, there also exist histology-specific regions of gain and loss.     

Increases in HIV-related sexual risk behavior among community samples of gay men in London and Glasgow: how do they compare?

OBJECTIVE: In this paper, we compare trends in sexual risk among gay men in the largest city in England (London) and the largest city in Scotland (Glasgow). METHODS: Self-complete questionnaires administered to representative samples of men visiting the commercial gay scenes in London and Glasgow in 1996, 1999, and 2002 (N = 8247). RESULTS: Multivariate logistic regression was used to assess the trends in unprotected anal intercourse (UAI), UAI with partners of unknown/discordant HIV status, and UAI with more than 1 partner. Each increased significantly in 1999 and 2002 in London, but only in 2002 in Glasgow. Testing for HIV also increased significantly in London, but not in Glasgow. Overall, HIV testing levels were considerably lower in Glasgow (in 2002, 49.1% vs. 74.6% in London). Overall, sexual risk was higher in London, but UAI with partners of unknown/discordant HIV status was higher in Glasgow (in 2002, 27.4% vs. 21.3%). CONCLUSIONS: Although the same pattern of increase in HIV-related sexual risk behavior was apparent in the cities, differences in HIV testing and nonconcordant UAI suggest different HIV prevention needs and that targeted health promotion interventions are required in London and Glasgow. City-specific factors should be considered in the development of appropriate sexual health interventions.