Impact of donation mode on the proportion and function of T lymphocytes in the liver

Background Liver T-cells respond to the inflammatory insult generated during organ procurement and contribute to the injury following reperfusion. The mode of liver donation alters various metabolic and inflammatory pathways but the way it affects intrahepatic T-cells is still unclear. Methods We investigated the modifications occurring in the proportion and function of T-cells during liver procurement for transplantation. We isolated hepatic mononuclear cells (HMC) from liver perfusate of living donors (LD) and donors after brain death (DBD) or cardiac death (DCD) and assessed the frequency of T-cell subsets, their cytokine secretion profile and CD8 T-cell cytotoxicity function, responsiveness to a danger associated molecular pattern (High Mobility Group Box1, HMGB1) and association with donor and recipient clinical parameters and immediate graft outcome. Results We found that T-cells in healthy human livers were enriched in memory CD8 T-cells exhibiting a phenotype of non-circulating tissue-associated lymphocytes, functionally dominated by more cytotoxicity and IFN-γ-production in DBD donors, including upon activation by HMGB1 and correlating with peak of post-transplant AST. This liver-specific pattern of CD8 T-cell was prominent in DBD livers compared to DCD and LD livers suggesting that it was influenced by events surrounding brain death, prior to retrieval. Conclusion Mode of liver donation can affect liver T-cells with increased liver damage in DBD donors. These findings may be relevant in designing therapeutic strategies aimed at organ optimization prior to transplantation

Conducting retrospective impact analysis to inform a medical research charity’s funding strategies: The case of Asthma UK

© 2013 Hanney et al.; licensee BioMed Central Ltd. 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 cited.This article has been made available through the Brunel Open Access Publishing Fund.BACKGROUND: Debate is intensifying about how to assess the full range of impacts from medical research. Complexity increases when assessing the diverse funding streams of funders such as Asthma UK, a charitable patient organisation supporting medical research to benefit people with asthma. This paper aims to describe the various impacts identified from a range of Asthma UK research, and explore how Asthma UK utilised the characteristics of successful funding approaches to inform future research strategies. METHODS: We adapted the Payback Framework, using it both in a survey and to help structure interviews, documentary analysis, and case studies. We sent surveys to 153 lead researchers of projects, plus 10 past research fellows, and also conducted 14 detailed case studies. These covered nine projects and two fellowships, in addition to the innovative case studies on the professorial chairs (funded since 1988) and the MRC-Asthma UK Centre in Allergic Mechanisms of Asthma (the ‘Centre’) which together facilitated a comprehensive analysis of the whole funding portfolio. We organised each case study to capture whatever academic and wider societal impacts (or payback) might have arisen given the diverse timescales, size of funding involved, and extent to which Asthma UK funding contributed to the impacts. RESULTS: Projects recorded an average of four peer-reviewed journal articles. Together the chairs reported over 500 papers. All streams of funding attracted follow-on funding. Each of the various categories of societal impacts arose from only a minority of individual projects and fellowships. Some of the research portfolio is influencing asthma-related clinical guidelines, and some contributing to product development. The latter includes potentially major breakthroughs in asthma therapies (in immunotherapy, and new inhaled drugs) trialled by university spin-out companies. Such research-informed guidelines and medicines can, in turn, contribute to health improvements. The role of the chairs and the pioneering collaborative Centre is shown as being particularly important. CONCLUSIONS: We systematically demonstrate that all types of Asthma UK’s research funding assessed are making impacts at different levels, but the main societal impacts from projects and fellowships come from a minority of those funded. Asthma UK used the study’s findings, especially in relation to the Centre, to inform research funding strategies to promote the achievement of impact.This study was funded by Asthma UK

Classification of the Mediterranean lowland to submontane pine forest vegetation

Vegetation SurveyAim: Vegetation types of Mediterranean thermophilous pine forests dominated by Pinus brutia, Pinus halepensis, Pinus pinaster and Pinus pinea were studied in various areas. However, a comprehensive formal vegetation classification of these forests based on a detailed data analysis has never been developed. Our aim is to provide the first broad-scale classification of these pine forests based on a large data set of vegetation plots. Location: Southern Europe, North Africa, Levant, Anatolia, Crimea and the Caucasus. Methods: We prepared a data set of European and Mediterranean pine forest vegetation plots. We selected 7,277 plots dominated by the cold-sensitive Mediterranean pine species Pinus brutia, Pinus halepensis, Pinus pinaster and Pinus pinea. We classified these plots using TWINSPAN, interpreted the ecologically and biogeographically homogeneous TWINSPAN clusters as alliances, and developed an expert system for automatic vegetation classification at the class, order and alliance levels. Results: We described Pinetea halepensis as a new class for the Mediterranean lowland to submontane pine forests, included in the existing Pinetalia halepensis order, and distinguished 12 alliances of native thermophilous pine forests, including four newly described and three informal groups merging supposedly native stands and old-established plantations. The main gradients in species composition reflect elevational vegetation belts and the west–east, and partly north–south, biogeographical differences. Both temperature and precipitation seasonality co-vary with these gradients. Conclusions: We provide the first formal classification at the order and alliance levels for all the Mediterranean thermophilous pine forests based on vegetation-plot data. This classification includes traditional syntaxa, which have been critically revised, and a new class and four new alliances. We also outline a methodological workflow that might be useful for other vegetation classification syntheses. The expert system, which is jointly based on pine dominance and species composition, is a tool for applying this classification in research and nature conservation survey, monitoring and managementinfo:eu-repo/semantics/publishedVersio

GITR signaling potentiates airway hyperresponsiveness by enhancing Th2 cell activity in a mouse model of asthma

<p>Abstract</p> <p>Background</p> <p>Allergic asthma is characterized by airway hyperresponsiveness (AHR) and allergic inflammation of the airways, driven by allergen-specific Th2 cells. The asthma phenotypes and especially AHR are sensitive to the presence and activity of regulatory T (Treg) cells in the lung. Glucocorticoid-induced tumor necrosis factor receptor (GITR) is known to have a co-stimulatory function on effector CD4⁺T cells, rendering these cells insensitive to Treg suppression. However, the effects of GITR signaling on polarized Th1 and Th2 cell effector functions are not well-established. We sought to evaluate the effect of GITR signaling on fully differentiated Th1 and Th2 cells and to determine the effects of GITR activation at the time of allergen provocation on AHR and airway inflammation in a Th2-driven mouse model of asthma.</p> <p>Methods</p> <p>CD4⁺CD25^-cells were polarized <it>in vitro </it>into Th1 and Th2 effector cells, and re-stimulated in the presence of GITR agonistic antibodies to assess the effect on IFNγ and IL-4 production. To evaluate the effects of GITR stimulation on AHR and allergic inflammation in a mouse asthma model, BALB/c mice were sensitized to OVA followed by airway challenges in the presence or absence of GITR agonist antibodies.</p> <p>Results</p> <p>GITR engagement potentiated cytokine release from CD3/CD28-stimulated Th2 but not Th1 cells <it>in vitro</it>. In the mouse asthma model, GITR triggering at the time of challenge induced enhanced airway hyperresponsiveness, serum IgE and <it>ex vivo </it>Th2 cytokine release, but did not increase BAL eosinophilia.</p> <p>Conclusion</p> <p>GITR exerts a differential effect on cytokine release of fully differentiated Th1 and Th2 cells <it>in vitro</it>, potentiating Th2 but not Th1 cytokine production. This effect on Th2 effector functions was also observed <it>in vivo </it>in our mouse model of asthma, resulting in enhanced AHR, serum IgE responses and Th2 cytokine production. This is the first report showing the effects of GITR activation on cytokine production by polarized primary Th1 and Th2 populations and the relevance of this pathway for AHR in mouse models for asthma. Our data provides crucial information on the mode of action of the GITR signaling, a pathway which is currently being considered for therapeutic intervention.</p

A microRNA profile of human CD8(+) regulatory T cells and characterization of the effects of microRNAs on Treg cell-associated genes.

Recently, regulatory T (Treg) cells have gained interest in the fields of immunopathology, transplantation and oncoimmunology. Here, we investigated the microRNA expression profile of human natural CD8(+)CD25(+) Treg cells and the impact of microRNAs on molecules associated with immune regulation. We purified human natural CD8(+) Treg cells and assessed the expression of FOXP3 and CTLA-4 by flow cytometry. We have also tested the ex vivo suppressive capacity of these cells in mixed leukocyte reactions. Using TaqMan low-density arrays and microRNA qPCR for validation, we could identify a microRNA 'signature' for CD8(+)CD25(+)FOXP3(+)CTLA-4(+) natural Treg cells. We used the 'TargetScan' and 'miRBase' bioinformatics programs to identify potential target sites for these microRNAs in the 3'-UTR of important Treg cell-associated genes. The human CD8(+)CD25(+) natural Treg cell microRNA signature includes 10 differentially expressed microRNAs. We demonstrated an impact of this signature on Treg cell biology by showing specific regulation of FOXP3, CTLA-4 and GARP gene expression by microRNA using site-directed mutagenesis and a dual-luciferase reporter assay. Furthermore, we used microRNA transduction experiments to demonstrate that these microRNAs impacted their target genes in human primary Treg cells ex vivo. We are examining the biological relevance of this 'signature' by studying its impact on other important Treg cell-associated genes. These efforts could result in a better understanding of the regulation of Treg cell function and might reveal new targets for immunotherapy in immune disorders and cancer

Calcitriol modulates the CD46 pathway in T cells

The complement regulator CD46 is a costimulatory molecule for human T cells that induces a regulatory Tr1 phenotype, characterized by large amounts of IL-10 secretion. Secretion of IL-10 upon CD46 costimulation is largely impaired in T cells from patients with multiple sclerosis (MS). Vitamin D can exert a direct effect on T cells, and may be beneficial in several pathologies, including MS. In this pilot study, we examined whether active vitamin D (1,25(OH)2D3 or calcitriol) could modulate the CD46 pathway and restore IL-10 production by CD46-costimulated CD4+ T cells from patients with MS. In healthy T cells, calcitriol profoundly affects the phenotype of CD46-costimulated CD4+ T cells, by increasing the expression of CD28, CD25, CTLA-4 and Foxp3 while it concomitantly decreased CD46 expression. Similar trends were observed in MS CD4+ T cells except for CD25 for which a striking opposite effect was observed: while CD25 was normally induced on MS T cells by CD46 costimulation, addition of calcitriol consistently inhibited its induction. Despite the aberrant effect on CD25 expression, calcitriol increased the IL-10:IFNc ratio, characteristic of the CD46-induced Tr1 phenotype, in both T cells from healthy donors and patients with MS. Hence, we show that calcitriol affects the CD46 pathway, and that it promotes anti-inflammatory responses mediated by CD46. Moreover, it might be beneficial for T cell responses in MS

The Hookworm Tissue Inhibitor of Metalloproteases (Ac-TMP-1) Modifies Dendritic Cell Function and Induces Generation of CD4 and CD8 Suppressor T Cells

Hookworm infection is a major cause of disease burden for humans. Recent studies have described hookworm-related immunosuppression in endemic populations and animal models. A Tissue Inhibitor of Metalloproteases (Ac-TMP-1) has been identified as one of the most abundant proteins released by the adult parasite. We investigated the effect of recombinant Ac-TMP-1 on dendritic cell (DC) and T cell function. Splenic T cells from C57BL/6 mice injected with Ac-TMP-1 showed reduced proliferation to restimulation with anti CD3 or bystander antigens such as OVA. Incubation of bone marrow-derived DCs with Ac-TMP-1 decreased MHC Class I and, especially, Class II expression but increased CD86 and IL-10 expression. Co-incubation of splenic T cells with DCs pulsed with Ac-TMP-1 induced their differentiation into CD4+ and, particularly, CD8+ CD25+Foxp3+ T cells that expressed IL-10. These cells were able to suppress proliferation of naïve and activated CD4+ T cells by TGF-Β-dependent (CD4+ suppressors) or independent (CD8+ suppressors) mechanisms. Priming of DCs with non-hookworm antigens, such as OVA, did not result in the generation of suppressor T cells. These data indicate that Ac-TMP-1 initiates the development of a regulatory response through modifications in DC function and generation of suppressor T cells. This is the first report to propose a role of suppressor CD8+ T cells in gastrointestinal helminthic infections

Influence of Short-Term Glucocorticoid Therapy on Regulatory T Cells In Vivo

Background: Pre- and early clinical studies on patients with autoimmune diseases suggested that induction of regulatory T(Treg) cells may contribute to the immunosuppressive effects of glucocorticoids(GCs). Objective: We readdressed the influence of GC therapy on Treg cells in immunocompetent human subjects and naı¨ve mice. Methods: Mice were treated with increasing doses of intravenous dexamethasone followed by oral taper, and Treg cells in spleen and blood were analyzed by FACS. Sixteen patients with sudden hearing loss but without an inflammatory disease received high-dose intravenous prednisolone followed by stepwise dose reduction to low oral prednisolone. Peripheral blood Treg cells were analyzed prior and after a 14 day GC therapy based on different markers. Results: Repeated GC administration to mice for three days dose-dependently decreased the absolute numbers of Treg cells in blood (100 mg dexamethasone/kg body weight: 2.861.86104 cells/ml vs. 336116104 in control mice) and spleen (dexamethasone: 2.861.96105/spleen vs. 956226105/spleen in control mice), which slowly recovered after 14 days taper in spleen but not in blood. The relative frequency of FOXP3+ Treg cells amongst the CD4+ T cells also decreased in a dose dependent manner with the effect being more pronounced in blood than in spleen. The suppressive capacity of Treg cells was unaltered by GC treatment in vitro. In immunocompetent humans, GCs induced mild T cell lymphocytosis. However, it did not change the relative frequency of circulating Treg cells in a relevant manner, although there was some variation depending on the definition of the Treg cells (FOXP3+: 4.061.5% vs 3.461.5%*; AITR+: 0.660.4 vs 0.560.3%, CD127low: 4.061.3 vs 5.063.0%* and CTLA4+: 13.8611.5 vs 15.6612.5%; * p,0.05). Conclusion: Short-term GC therapy does not induce the hitherto supposed increase in circulating Treg cell frequency, neither in immunocompetent humans nor in mice. Thus, it is questionable that the clinical efficacy of GCs is achieved by modulating Treg cell numbers

Self-tolerance in multiple sclerosis

During the last decade, several defects in self-tolerance have been identified in multiple sclerosis. Dysfunction in central tolerance leads to the thymic output of antigen-specific T cells with T cell receptor alterations favouring autoimmune reactions. In addition, premature thymic involution results in a reduced export of naïve regulatory T cells, the fully suppressive clone. Alterations in peripheral tolerance concern costimulatory molecules as well as transcriptional and epigenetic mechanisms. Recent data underline the key role of regulatory T cells that suppress Th1 and Th17 effector cell responses and whose immunosuppressive activity is impaired in patients with multiple sclerosis. Those recent observations suggest that a defect in self-tolerance homeostasis might be the primary mover of multiple sclerosis leading to subsequent immune attacks, inflammation and neurodegeneration. The concept of multiple sclerosis as a consequence of the failure of central and peripheral tolerance mechanisms to maintain a self-tolerance state, particularly of regulatory T cells, may have therapeutic implications. Restoring normal thymic output and suppressive functions of regulatory T cells appears an appealing approach. Regulatory T cells suppress the general local immune response via bystander effects rather than through individual antigen-specific responses. Interestingly, the beneficial effects of currently approved immunomodulators (interferons β and glatiramer acetate) are associated with a restored regulatory T cell homeostasis. However, the feedback regulation between Th1 and Th17 effector cells and regulatory T cells is not so simple and tolerogenic mechanisms also involve other regulatory cells such as B cells, dendritic cells and CD56bright natural killer cells