Adenosine and its receptors : translational studies in asthma and COPD

We were the first to report studies regarding adenosine receptor expression on inflammatory cells in peripheral blood and sputum in asthma patients and healthy controls, as well as sputum of COPD patients and asymptomatic smokers. With these results we partly addressed the difficulties presented in the conclusions part of chapter 2 of this thesis (29). Here we stated: "Thus far, the exact distribution and function of the different adenosine receptor subtypes in human airways remains largely unknown and this makes it difficult to establish which (combination of) receptors should be targeted" Our studies have increased our knowledge about adenosine receptor expression and distribution in healthy and disease situations which could already give some indications about treatment directions. With respect to asthma, we showed as anticipated that asthma patients have an increased number of eosinophils in peripheral blood and induced sputum, and that these numbers further increased in sputum after allergen challenge. However, AR expression on blood eosinophils in asthma did not differ from that in healthy controls nor did it change upon allergen challenge, thus not explaining this increased eosinophil migration. Furthermore, we were not able to evaluate AR expression on sputum eosinophils because only few asthma patients had sufficient numbers of eosinophils in sputum to perform reliable scoring. Thus, it appears that allergen challenge does not directly affect eosinophil recruitment via adenosine receptor stimulation to the lung tissue, and other indirect factors like mediator release from mast cells, epithelial cells and other resident or inflammatory cells thus likely contribute to eosinophils attraction. Since an important role in chemotaxis is described especially for A3R on eosinophils (29), it would be of importance to include determination of the expression of this receptor in future studies (e.g. by performing eosinophil purification of the sputum samples before cytospin preparation)

HDAC inhibition increases HLA class I expression in uveal melanoma

The treatment of uveal melanoma (UM) metastases or adjuvant treatment may imply immunological approaches or chemotherapy. It is to date unknown how epigenetic modifiers affect the expression of immunologically relevant targets, such as the HLA Class I antigens, in UM. We investigated the expression of HDACs and the histone methyl transferase EZH2 in a set of 64 UMs, using an Illumina HT12V4 array, and determined whether a histone deacetylase (HDAC) inhibitor and EZH2 inhibitor modified the expression of HLA Class I on three UM cell lines. Several HDACs (HDAC1, HDAC3, HDAC4, and HDAC8) showed an increased expression in high-risk UM, and were correlated with an increased HLA expression. HDAC11 had the opposite expression pattern. While in vitro tests showed that Tazemetostat did not influence cell growth, Quisinostat decreased cell sur

Benchmarked performance charts using principal components analysis to improve the effectiveness of feedback for audit data in HIV care

Abstract Background Feedback tools for clinical audit data that compare site-specific results to average performance over all sites can be useful for quality improvement. Proposed tools should be simple and clearly benchmark the site’s performance, so that a relevant action plan can be directly implemented to improve patient care services. We aimed to develop such a tool in order to feedback data to UK HIV clinics participating in the 2015 British HIV Association (BHIVA) audit assessing compliance with the 2011 guidelines for routine investigation and monitoring of adult HIV-1- infected individuals. Methods HIV clinic sites were asked to provide data on a random sample of 50–100 adult patients attending for HIV care during 2014 and/or 2015 by completing a self-audit spreadsheet. Outcomes audited included the proportion of patients with recorded resistance testing, viral load monitoring, adherence assessment, medications, hepatitis testing, vaccination management, risk assessments, and sexual health screening. For each outcome we benchmarked the proportion for a specific site against the average performance. We produced performance charts for each site using boxplots for the outcomes. We also used the mean and differences from the mean performance to produce a dashboard for each site. We used principal components analysis to group correlated outcomes and simplify the dashboard. Results The 106 sites included in the study provided information on a total of 7768 patients. Outcomes capturing monitoring of treatment of HIV-infection showed high performance across the sites, whereas testing for hepatitis, and risk assessment for cardiovascular disease and smoking, management of flu vaccination, sexual health screening, and cervical cytology for women were very variable across sites. The principal components analysis reduced the original 12 outcomes to four factors that represented HIV care, hepatitis testing, other screening tests, and resistance testing. These provided simplified measures of adherence to guidelines which were presented as a 4 bar dashboard of performance. Conclusion Our dashboard performance charts provide easily digestible visual summaries of locally relevant audit data that are benchmarked against the overall mean and can be used to improve feedback to HIV services. Feedback from clinicians indicated that they found these charts acceptable and useful

GNAQ and GNA11 mutations and downstream YAP activation in choroidal nevi

Background: Mutations in GNAQ/11 genes are considered an early event in the development of uveal melanoma that may derive from a pre-existing nevus. The Hippo pathway, by way of YAP activation, rather than MAP kinase, has a role in the oncogenic capacity of GNAQ/11 mutations.Methods: We investigated 16 nevi from 13 human eyes for driver GNAQ/11 mutations using droplet digital PCR and determined whether nevi are clonal by quantifying mutant nevus cell fractions. Immunohistochemistry was performed on 15 nevi to analyse YAP activation.Results: For 15 out of 16 nevi, a GNAQ/11 mutation was detected in the nevus cells albeit at a low frequency with a median of 13%. Nuclear YAP, a transcriptional co-activator in the Hippo tumour-suppressor pathway, was detected in 14/15 nevi.Conclusions: Our analysis suggests that a mutation in GNAQ/11 occurs in a subset of choroidal nevus cells. We hypothesise that GNAQ/11 mutant-driven extracellular mitogenic signalling involving YAP activation leads to accumulation of wild-type nevus cells

Soluble HLA in the aqueous humour of uveal melanoma is associated with unfavourable tumour characteristics

A high HLA expression in uveal melanoma (UM) is part of the prognostically unfavorable inflammatory phenotype. We wondered whether the presence of soluble HLA (sHLA) in the aqueous humour is associated with clinical, histopathological or genetic tumour characteristics, and represents tumour HLA expression and intratumoural inflammation. Aqueous humour from 108 UM patients was analysed for the presence of sHLA, using a Luminex assay specific for HLA Class I. Clinical and genetic parameters were compared between sHLA-positive and negative eyes. A qPCR analysis was performed on tumour tissue using a Fluidigm assay. In 19/108 UM-containing eyes, the sHLA level in the aqueous was above the detection limit. Tumours in sHLA-positive eyes were significantly larger, more frequently involved the ciliary body, and more often showed monosomy 3, gain of chromosome 8q and loss of BAP1 staining. Melanoma-related survival was worse in patients with sHLA-positive aqueous humour. sHLA in the aqueous did not represent the tumour's HLA expression and did not relate to immune cell infiltration in the tumour. We conclude that UM-containing eyes may contain sHLA in the aqueous humour, where it is a prognostically-unfavourable sign and may influence local immune responses

Effects of IL-4 and IL-13 on adenosine receptor expression and responsiveness of the human mast cell line 1

Background: Inhalation of adenosine-5'-monophosphate (AMP) causes bronchoconstriction in asthma but not in healthy subjects. Bronchoconstriction upon AMP inhalation is thought to occur by histamine release and subsequent binding to receptors on airway smooth muscle cells. Methods: To explain enhanced sensitivity to AMP in asthma, mast cell expression of the adenosine A(2A) and A(2B) receptors and histamine release were measured after incubation of human mast cell tine 1 (HMC-1) cells with AMP and the non-specific adenosine receptor agonist 5'-N-ethylcarboxamidoadenosine (NECA) for 1.5 and 6 h. To establish a Thelper-2 environment resembling the asthma phenotype, HMC-1 cells were additionally cultured with IL-4 and IL-13 atone or stimulated with the combination of both cytokines; and AMP and NECA. To study effects of prolonged presence of the inflammatory environment, the cells were pre-incubated overnight (18 h) with IL-4 and IL-13 and additionally stimulated with AMP and NECA for 1.5 or 6 h. Results: AMP and NECA hardly affected adenosine receptor expression but increased IL-8 secretion. Incubation with IL-4 and IL-13 for 6 h increased adenosine A(2A) receptor expression and histamine secretion, but decreased IL-8 secretion. The combination of IL-4, IL-13, and AMP/NECA for 6 h increased A(2B) receptor expression and IL-8 secretion. Overnight stimulation with IL-4, IL-13 and subsequent stimulation with AMP/NECA for 1.5 h decreased A(2A)R expression which was accompanied by increased histamine secretion. Conclusion: These results suggest a role for decreased A(2A)R expression in enhanced adenosine responsiveness as observed in asthma. (C) 2008 Elsevier B.V. All rights reserved

Inflammatory Cytokines in Eyes with Uveal Melanoma and Relation with Macrophage Infiltration

Eyes with uveal melanoma have been shown to have multiple cytokines and chemokines in the aqueous humor. This study was conducted to determine whether a relation exists between the presence of these cytokines and the prognostic factors and survival of uveal melanoma

Ischemia is related to tumour genetics in uveal melanoma

Hypoxia-inducible factor 1-alpha (HIF1a) and its regulator von Hippel–Lindau protein (VHL) play an important role in tumour ischemia. Currently, drugs that target HIF1a are being developed to treat malignancies. Although HIF1a is known to be expressed in uveal melanoma (UM), it is as yet unknown which factors, such as tumour size or genetics, determine its expression. Therefore, we aimed to determine which tumour characteristics relate to HIF1a expression in UM. Data from 64 patients who were enucleated for UM were analysed. Messenger RNA (mRNA) expression was determined with the Illumina HT-12 v4 chip. In 54 cases, the status of chromosomes 3 and 8q, and BRCA1-associated protein 1 (BAP1) protein expression (immunohistochemistry) were determined. Findings were corrobora