Extracellular matrix-modulated expression of human cell surface glycoproteins A42 and J143. Intrinsic and extrinsic signals determine antigenic phenotype.

Extracellular matrix (ECM)' plays an important regulatory role in cellular growth, migration, and differentiation (1-4). Pathologic processes such as tumor cell invasion and metastasis are also determined by cellular interactions with ECM (5, 6). Biochemical studies have identified collagens, fibronectin, laminin, proteoglycans, and several other proteins as major ECM components (1-3), and have shown that ECM composition varies between different normal and tumor tissues. The complexity and heterogeneity of ECM composition have hampered the molecular analysis of ECM-cell interactions . However, a range of phenotypic changes has been described for cultured cells after transfer from plastic surfaces to substrates coated with native ECM (7, 8) or with purified ECM components; ECM-induced phenotypic changes include enhanced substrate adhesiveness, cell spreading and migration, changes in cell morphology and proliferative activity, and expression of differentiated cellular functions (I-4). Some of these effects, e.g., increased substrate adhesion, may result directly from the binding of specialized cell surface structures to ECM molecules. Others are likely mediated by an active cellular response triggered by the interaction of ECM with cell surface receptors. Thus, ECM-derived signals (9) may activate a cascade of molecular changes within the cell and on the cell surface that account for the pleiotropic effects observed with ECM

Methylation of the imprinted GNAS1 gene in cell-free plasma DNA : equal steady-state quantities of methylated and unmethylated DNA in plasma

Background Genomic DNA sequences in cell-free plasma are biomarkers of cancer prognosis, where characteristic changes in methylation of tumour suppressor or oncogene DNA regions are indicative of changes in gene activity. Also, cell-free fetal DNA can be distinguished, by its methylation status, from the maternal DNA in the plasma of pregnant women, hence providing DNA biomarkers for the proposed minimally-invasive diagnosis of fetal aneuploidies, including Down's syndrome. However, the production and clearance of cell-free DNA from plasma in relation to its methylation status, are poorly understood processes. Methods We studied the methylation status of DNA derived from the imprinted GNAS1 locus, in cell-free plasma DNA of healthy adults. Heterozygotes were identified that carried the SNP rs1800905 in the imprinted region. The parent-of-origin-dependent DNA methylation was analysed by bisulfite conversion, followed by cloning and sequencing. Results Genomic DNA molecules derived from both the methylated, maternal, allele and the unmethylated, paternal, allele were found in plasma. Methylated and unmethylated DNA molecules were present in equal numbers. Conclusions Our data indicate that the methylation status of a DNA sequence has no effect on its steady state concentration in the cell-free DNA component of plasma, in healthy adults

Results of a randomized, double-blind phase II clinical trial of NY-ESO-1 vaccine with ISCOMATRIX adjuvant versus ISCOMATRIX alone in participants with high-risk resected melanoma.

BACKGROUND: To compare the clinical efficacy of New York Esophageal squamous cell carcinoma-1 (NY-ESO-1) vaccine with ISCOMATRIX adjuvant versus ISCOMATRIX alone in a randomized, double-blind phase II study in participants with fully resected melanoma at high risk of recurrence. METHODS: Participants with resected stage IIc, IIIb, IIIc and IV melanoma expressing NY-ESO-1 were randomized to treatment with three doses of NY-ESO-1/ISCOMATRIX or ISCOMATRIX adjuvant administered intramuscularly at 4-week intervals, followed by a further dose at 6 months. Primary endpoint was the proportion free of relapse at 18 months in the intention-to-treat (ITT) population and two per-protocol populations. Secondary endpoints included relapse-free survival (RFS) and overall survival (OS), safety and NY-ESO-1 immunity. RESULTS: The ITT population comprised 110 participants, with 56 randomized to NY-ESO-1/ISCOMATRIX and 54 to ISCOMATRIX alone. No significant toxicities were observed. There were no differences between the study arms in relapses at 18 months or for median time to relapse; 139 vs 176 days (p=0.296), or relapse rate, 27 (48.2%) vs 26 (48.1%) (HR 0.913; 95% CI 0.402 to 2.231), respectively. RFS and OS were similar between the study arms. Vaccine recipients developed strong positive antibody responses to NY-ESO-1 (p≤0.0001) and NY-ESO-1-specific CD4+ and CD8+ responses. Biopsies following relapse did not demonstrate differences in NY-ESO-1 expression between the study populations although an exploratory study demonstrated reduced (NY-ESO-1)+/Human Leukocyte Antigen (HLA) class I+ double-positive cells in biopsies from vaccine recipients performed on relapse in 19 participants. CONCLUSIONS: The vaccine was well tolerated, however, despite inducing antigen-specific immunity, it did not affect survival endpoints. Immune escape through the downregulation of NY-ESO-1 and/or HLA class I molecules on tumor may have contributed to relapse

Proteome Profiling of Breast Tumors by Gel Electrophoresis and Nanoscale Electrospray Ionization Mass Spectrometry

We have conducted proteome-wide analysis of fresh surgery specimens derived from breast cancer patients, using an approach that integrates size-based intact protein fractionation, nanoscale liquid separation of peptides, electrospray ion trap mass spectrometry, and bioinformatics. Through this approach, we have acquired a large amount of peptide fragmentation spectra from size-resolved fractions of the proteomes of several breast tumors, tissue peripheral to the tumor, and samples from patients undergoing noncancer surgery. Label-free quantitation was used to generate protein abundance maps for each proteome and perform comparative analyses. The mass spectrometry data revealed distinct qualitative and quantitative patterns distinguishing the tumors from healthy tissue as well as differences between metastatic and non-metastatic human breast cancers including many established and potential novel candidate protein biomarkers. Selected proteins were evaluated by Western blotting using tumors grouped according to histological grade, size, and receptor expression but differing in nodal status. Immunohistochemical analysis of a wide panel of breast tumors was conducted to assess expression in different types of breast cancers and the cellular distribution of the candidate proteins. These experiments provided further insights and an independent validation of the data obtained by mass spectrometry and revealed the potential of this approach for establishing multimodal markers for early metastasis, therapy outcomes, prognosis, and diagnosis in the future. © 2008 American Chemical Society

The development and validation of the Dementia Quality of Life Scale for Older Family Carers (DQoL-OC)

Purpose: Little is known about how caregiving affects the quality of life (QoL) of older family carers and no dementia and age-specific QoL scale is available for use with this population. This study aimed to develop and validate a unique dementia caregiving- and age-specific tool – the ‘Dementia Quality of Life Scale for Older Family Carers’ (DQoL-OC). Methods: The scale items were identified in focus groups with older family carers in the UK. Content and face validity were evaluated by a panel of six experts. A set of 100 items assessed on a 5-point Likert scale was tested with 182 older family carers. Test–re-test reliability was conducted with 18 individuals. Exploratory factor analysis was used to identify the QoL model and reduce the number of scale items. Convergent construct validity and internal consistency were also established. Results: A one-factor solution containing 22 items was obtained. Test–re-test reliability (lower bound r = 0.835; p < 0.001), internal consistency (Cronbach's α = 0.936), and convergent construct validity were established. Significantly lower levels of QoL were found in female older carers; those who perceived their relatives with dementia as being at the earlier stages of the disease and with unstable dementia symptoms; those providing care more hours per day and more days per week; and those in younger-old age. Conclusions: The DQoL-OC is a valid and reliable scale that will be useful for research and in clinical practice with older family carers of people with dementia. These study results will inform future health and social care aiming to improve life quality for this overlooked population of carers

Logics, rhetoric and 'the blob': populist logic in the Conservative reforms to English schooling

A lot has been written about the lasting implications of the Conservative reforms to English schooling, particularly changes made by Michael Gove as Education Secretary (2010–2014). There is a lot less work, however, on studying the role that language, strategy and the broader political framework played in the process of instituting and winning consent for these reforms. Studying these factors is important for ensuring that any changes to education and schooling are not read in isolation from their political context. Speeches particularly capture moments where intellectual and strategic political traditions meet, helping us to form a richer understanding of the motives behind specific reform goals and where they fit into a political landscape. This article analyses speeches and policy documents from prominent politicians who led the Conservative education agenda between 2010–2014 to illustrate how politicians mobilised a deliberate populist strategy and argumentation to achieve specific educational goals, but which have had broader social and political implications. Concepts from interpretive political studies are used to develop a case analysis of changes to teacher training provision and curriculum reform, illustrating how politicians constructed a frontier between ‘the people’ (commonly teachers or parents) and an illegitimate ‘elite’ (an educational establishment) that opposed change. This anti‐elite populist rhetoric, arguably first tested in the Department for Education, has now become instituted more widely in our current British politics

Current challenges in software solutions for mass spectrometry-based quantitative proteomics

This work was in part supported by the PRIME-XS project, grant agreement number 262067, funded by the European Union seventh Framework Programme; The Netherlands Proteomics Centre, embedded in The Netherlands Genomics Initiative; The Netherlands Bioinformatics Centre; and the Centre for Biomedical Genetics (to S.C., B.B. and A.J.R.H); by NIH grants NCRR RR001614 and RR019934 (to the UCSF Mass Spectrometry Facility, director: A.L. Burlingame, P.B.); and by grants from the MRC, CR-UK, BBSRC and Barts and the London Charity (to P.C.

Neuroinflammation, Mast Cells, and Glia: Dangerous Liaisons

The perspective of neuroinflammation as an epiphenomenon following neuron damage is being replaced by the awareness of glia and their importance in neural functions and disorders. Systemic inflammation generates signals that communicate with the brain and leads to changes in metabolism and behavior, with microglia assuming a pro-inflammatory phenotype. Identification of potential peripheral-to-central cellular links is thus a critical step in designing effective therapeutics. Mast cells may fulfill such a role. These resident immune cells are found close to and within peripheral nerves and in brain parenchyma/meninges, where they exercise a key role in orchestrating the inflammatory process from initiation through chronic activation. Mast cells and glia engage in crosstalk that contributes to accelerate disease progression; such interactions become exaggerated with aging and increased cell sensitivity to stress. Emerging evidence for oligodendrocytes, independent of myelin and support of axonal integrity, points to their having strong immune functions, innate immune receptor expression, and production/response to chemokines and cytokines that modulate immune responses in the central nervous system while engaging in crosstalk with microglia and astrocytes. In this review, we summarize the findings related to our understanding of the biology and cellular signaling mechanisms of neuroinflammation, with emphasis on mast cell-glia interactions

Shedding Light on Vampires: The Phylogeny of Vampyrellid Amoebae Revisited

With the advent of molecular phylogenetic techniques the polyphyly of naked filose amoebae has been proven. They are interspersed in several supergroups of eukaryotes and most of them already found their place within the tree of life. Although the ‘vampire amoebae’ have attracted interest since the middle of the 19th century, the phylogenetic position and even the monophyly of this traditional group are still uncertain. In this study clonal co-cultures of eight algivorous vampyrellid amoebae and the respective food algae were established. Culture material was characterized morphologically and a molecular phylogeny was inferred using SSU rDNA sequence comparisons. We found that the limnetic, algivorous vampyrellid amoebae investigated in this study belong to a major clade within the Endomyxa Cavalier-Smith, 2002 (Cercozoa), grouping together with a few soil-dwelling taxa. They split into two robust clades, one containing species of the genus Vampyrella Cienkowski, 1865, the other containing the genus Leptophrys Hertwig & Lesser, 1874, together with terrestrial members. Supported by morphological data these clades are designated as the two families Vampyrellidae Zopf, 1885, and Leptophryidae fam. nov. Furthermore the order Vampyrellida West, 1901 was revised and now corresponds to the major vampyrellid clade within the Endomyxa, comprising the Vampyrellidae and Leptophryidae as well as several environmental sequences. In the light of the presented phylogenetic analyses morphological and ecological aspects, the feeding strategy and nutritional specialization within the vampyrellid amoebae are discussed