107 research outputs found
Differential loss of chromosome 11q in familial and sporadic parasympathetic paragangliomas detected by comparative genomic hybridization
Parasympathetic paragangliomas (PGLs) represent neuroendocrine tumors
arising from chief cells in branchiomeric and intravagal paraganglia,
which share several histological features with their sympathetic
counterpart sympathoadrenal paragangliomas. In recent years, genetic
analyses of the familial form of PGL have attracted considerable interest.
However, the majority of paragangliomas occurs sporadically and it remains
to be determined whether the pathogenesis of sporadic paraganglioma
resembles that of the familial form. Furthermore, data on comparative
genetic aberrations are scarce. To provide fundamental cytogenetic data on
sporadic and hereditary PGLs, we performed comparative genomic
hybridization using directly fluorochrome-conjugated DNA extracted from 12
frozen and 4 paraffin-embedded tumors. The comparative genomic
hybridization data were extended by loss of heterozygosity analysis of
chromosome 11q. DNA copy number changes were found in 10 (63%) of 16
tumors. The most frequent chromosomal imbalance involved loss of
chromosome 11. Six of seven familial tumors and two of nine sporadic
tumors showed loss of 11q (86% versus 22%, P = 0.012). Deletions of 11p
and 5p were found in two of nine sporadic tumors. We conclude that overall
DNA copy number changes are infrequent in PGLs compared to sympathetic
paragangliomas and that loss of chromosome 11 may be an important event in
their tumorigenesis, particularly in familial paragangliomas
Simvastatin improves the sexual health-related quality of life in men aged 40 years and over with erectile dysfunction : Additional data from the Erectile Dysfunction and Statin trial
© 2014 Trivedi et al. This is an Open Access article distributed under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited.Background: Erectile dysfunction is prevalent in men over 40 years, affecting their quality of life and that of their partners. The aims of this study were:a)To evaluate the internal reliability of the male erectile dysfunction specific quality of life (MED-QoL) scale and explore its factor structure.b)To evaluate the effect of simvastatin on subscales of the MED-QoL in men over forty years with erectile dysfunction. Methods: This is a double blind randomised controlled trial of 40 mg simvastatin or placebo given once daily for six months to men over forty years with untreated erectile dysfunction, who were not at high cardiovascular risk and were not on anti-hypertensive or lipid-lowering medication. 173 eligible men were recruited from 10 general practices in East of England. Data were collected at two points over 30 weeks. We report on the factor structure of MED-QoL, the internal reliability of the scale and the derived subscales, and the effect of simvastatin on MED-QoL subscales. Results: An initial analysis of the MED-QoL items suggested that a number of items should be removed (MED-QoL-R). Exploratory factor analysis identified three subscales within the MED-QoL-R which accounted for 96% of the variance, related to feelings of Control, initiating Intimacy, and Emotional response to erectile dysfunction. The alpha value for the revised scale (MED-Qol-R) was >0.95 and exceeded .82 for each subscale. Regression analysis showed that patients in the placebo group experienced a significantly reduced feeling of Control over erectile dysfunction than those in the statin group. Those in the placebo group had significantly lower Emotional response than those in the statin group at the close of trial, but there was no significant treatment effect on Intimacy. Conclusions: Our revised MED-QoL-R identified three subscales. Secondary analysis showed a significant improvement in sexual health related quality of life, specifically in relation to perception of control and emotional health in men with untreated erectile dysfunction given 40 mg simvastatin for six months. Trial registration: Current Controlled Trials ISRCTN66772971.Peer reviewe
Losses of chromosomes 1p and 3q are early genetic events in the development of sporadic pheochromocytomas
Despite several loss of heterozygosity studies, a comprehensive genomic
survey of pheochromocytomas is still lacking. To identify DNA copy number
changes which might be important in tumor development and progression and
which may have diagnostic utility, we evaluated genetic aberrations in 29
sporadic adrenal and extra-adrenal pheochromocytomas (19 clinically benign
tumors and 10 malignant lesions). Comparative genomic hybridization was
performed using directly fluorochrome-conjugated DNA extracted from frozen
(16) and paraffin-embedded (13) tumor tissues. The most frequently
observed changes were losses of chromosomes 1p11-p32 (86%), 3q (52%), 6q
(34%), 3p, 17p (31% each), 11q (28%), and gains of chromosomes 9q (38%)
and 17q (31%). No amplification was identified and no difference between
adrenal and extra-adrenal tumors was detected. Progression to malignant
tumors was strongly associated with deletions of chromosome 6q (60% versus
21% in clinically benign lesions, P = 0.0368) and 17p (50% versus 21%).
Fluorescence in situ hybridization confirmed the comparative genomic
hybridization data of chromosomes 1p, 3q, and 6q, and revealed aneuploidy
in some tumors. Our results suggest that the development of
pheochromocytomas is associated with specific genomic aberrations, such as
losses of 1p, 3q, and 6q and gains of 9q and 17q. In particular, tumor
suppressor genes on chromosomes 1p and 3q may be involved in early
tumorigenesis, and deletions of chromosomes 6q and 17p in progression to
malignancy
Highly Sensitive Determination of Hydrogen Peroxide and Glucose by Fluorescence Correlation Spectroscopy
BACKGROUND: Because H(2)O(2) is generated by various oxidase-catalyzed reactions, a highly sensitive determination method of H(2)O(2) is applicable to measurements of low levels of various oxidases and their substrates such as glucose, lactate, glutamate, urate, xanthine, choline, cholesterol and NADPH. We propose herein a new, highly sensitive method for the measurement of H(2)O(2) and glucose using fluorescence correlation spectroscopy (FCS). METHODOLOGY/PRINCIPAL FINDINGS: FCS has the advantage of allowing us to determine the number of fluorescent molecules. FCS measures the fluctuations in fluorescence intensity caused by fluorescent probe movement in a small light cavity with a defined volume generated by confocal illumination. We thus developed a highly sensitive determination system of H(2)O(2) by FCS, where horseradish peroxidase (HRP) catalyzes the formation of a covalent bond between fluorescent molecules and proteins in the presence of H(2)O(2). Our developed system gave a linear calibration curve for H(2)O(2) in the range of 28 to 300 nM with the detection limit of 8 nM. In addition, by coupling with glucose oxidase (GOD)-catalyzed reaction, the method allows to measure glucose in the range of 80 nM to 1.5 µM with detection limit of 24 nM. The method was applicable to the assay of glucose in blood plasma. The mean concentration of glucose in normal human blood plasma was determined to be 4.9 mM. CONCLUSIONS/SIGNIFICANCE: In comparison with commercial available methods, the detection limit and the minimum value of determination for glucose are at least 2 orders of magnitude more sensitive in our system. Such a highly sensitive method leads the fact that only a very small amount of plasma (20 nL) is needed for the determination of glucose concentration in blood plasma
Dutch National Round Robin Trial on Plasma-Derived Circulating Cell-Free DNA Extraction Methods Routinely Used in Clinical Pathology for Molecular Tumor Profiling
BACKGROUND: Efficient recovery of circulating tumor DNA (ctDNA) depends on the quantity and quality of circulating cell-free DNA (ccfDNA). Here, we evaluated whether various ccfDNA extraction methods routinely applied in Dutch laboratories affect ccfDNA yield, ccfDNA integrity, and mutant ctDNA detection, using identical lung cancer patient-derived plasma samples. METHODS: Aliquots of 4 high-volume diagnostic leukapheresis plasma samples and one artificial reference plasma sample with predetermined tumor-derived mutations were distributed among 14 Dutch laboratories. Extractions of ccfDNA were performed according to local routine standard operating procedures and were analyzed at a central reference laboratory for mutant detection and assessment of ccfDNA quantity and integrity. RESULTS: Mutant molecule levels in extracted ccfDNA samples varied considerably between laboratories, but there was no indication of consistent above or below average performance. Compared to silica membrane-based methods, samples extracted with magnetic beads-based kits revealed an overall lower total ccfDNA yield (-29%; P < 0.0001) and recovered fewer mutant molecules (-41%; P < 0.01). The variant allelic frequency and sample integrity were similar. In samples with a higher-than-average total ccfDNA yield, an augmented recovery of mutant molecules was observed. CONCLUSIONS: In the Netherlands, we encountered diversity in preanalytical workflows with potential consequences on mutant ctDNA detection in clinical practice. Silica membrane-based methodologies resulted in the highest total ccfDNA yield and are therefore preferred to detect low copy numbers of relevant mutations. Harmonization of the extraction workflow for accurate quantification and sensitive detection is required to prevent introduction of technical divergence in the preanalytical phase and reduce interlaboratory discrepancies
Implementation of Novel Molecular Biomarkers for Non-small Cell Lung Cancer in the Netherlands:How to Deal With Increasing Complexity
The diagnostic landscape of non-small cell lung cancer (NSCLC) is changing rapidly with the availability of novel treatments. Despite high-level healthcare in the Netherlands, not all patients with NSCLC are tested with the currently relevant predictive tumor markers that are necessary for optimal decision-making for today's available targeted or immunotherapy. An expert workshop on the molecular diagnosis of NSCLC involving pulmonary oncologists, clinical chemists, pathologists, and clinical scientists in molecular pathology was held in the Netherlands on December 10, 2018. The aims of the workshop were to facilitate cross-disciplinary discussions regarding standards of practice, and address recent developments and associated challenges that impact future practice. This paper presents a summary of the discussions and consensus opinions of the workshop participants on the initial challenges of harmonization of the detection and clinical use of predictive markers of NSCLC. A key theme identified was the need for broader and active participation of all stakeholders involved in molecular diagnostic services for NSCLC, including healthcare professionals across all disciplines, the hospitals and clinics involved in service delivery, healthcare insurers, and industry groups involved in diagnostic and treatment innovations. Such collaboration is essential to integrate different technologies into molecular diagnostics practice, to increase nationwide patient access to novel technologies, and to ensure consensus-preferred biomarkers are tested
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