The role of innate lymphoid cells in mucosal inflammation in paediatric viral bronchiolitis

Background Human respiratory syncytial virus (RSV) is the most common cause of bronchiolitis and acute lower respiratory tract infections (LRTI) among children worldwide. RSV is the leading cause of childhood hospitalisation and a financial burden on national health systems. No vaccine that protects against RSV infection is available. Children with bronchiolitis are at high risk of developing wheeze and asthma later in childhood. Recent data suggest that innate immunity plays a vital role in childhood, especially when maternal antibodies' levels are low and adaptive immune responses are immature. Innate lymphoid cells (ILCs) are among the most recently discovered innate immune cell types that could play a pivotal role in innate responses to RSV bronchiolitis. ILCs have been classified into three groups: ILC1, ILC2 and ILC3 depending on their function, transcription factor expression and phenotype. Although ILC2 numbers have been found to be elevated in bronchiolitis, the phenotypic and functional differences between ILCs from healthy infants and those with bronchiolitis have not been explored. Hypothesis I propose that ILC frequencies and function are associated with disease severity and aetiology in children with acute lower respiratory tract infection. Sub-hypothesis 1 ILCs from neonates are functionally different from healthy adult subjects. Sub-hypothesis 2 Alterations in ILC frequencies and numbers in blood and airway samples are associated with disease severity, pathogen type and load. Sub-hypothesis 3 Alterations in ILC function are associated with disease severity, pathogen type and load. Methods To determine differences in ILCs in early life, lymphocyte populations, including ILC subset frequencies and numbers in blood samples collected from adults and neonates were investigated by flow cytometry. To study ILCs in bronchiolitis, blood and upper and lower airway samples were collected from infants with and without a respiratory viral infection. An optimised multicolour immunophenotyping panel (OMIP), intracellular staining panels (ICS) and other systems of analytical approach such as UMAP (Uniform Manifold Approximation and Projection), heatmaps, correlograms and principal component analysis (PCA) were used. ILCs were defined using the following gating strategy: live, single, CD45+, lineage negative (CD3, CD14, CD16, CD56, CD1a, CD123 FcεR1α)- and CD127+ cells. ILC1 cells were defined as CD117−CRTH2−, ILC2 cells were defined as CRTH2+ CD117int and ILC3s were defined as CD117+CRTH2−. At the same time, other lymphocyte and granulocyte populations were enumerated. Alongside cellular work, using the Meso Scale Discovery (MSD) platform, 29 immune mediators and cytokines were measured in airway samples. Results I successfully optimised flow cytometry panels to determine frequencies and function of ILCs in small volumes of difficult to obtain clinical samples. Results suggest that ILC numbers were higher in whole blood obtained from neonates compared to healthy adults (P=0.0032), however, they produced significantly less IFN-γ (P=0.0015). Furthermore, there were significantly lower numbers of NKT (P=0.0001) and MAIT cells (P=0.0031) in neonates compared to healthy adults. In bronchiolitis, ILC numbers were significantly decreased in peripheral blood (P=0.005) and airways samples (P=0.025) obtained from bronchiolitic compared to control subjects; however, their capacity for IFN-γ production was similar between those groups. Interestingly, there was a significantly decreased production of IFN-γ in CD4+ (P=0.042) and CD8+ T cells (P=0.006) from bronchiolitic compared to control patients. Finally, levels of almost all mediators were higher in bronchiolitic babies than controls, including both type-1, type-2 and type-17 associated cytokines. Only IL-4 production in lower and upper airways was increased in rhinovirus- (RV) but not RSV-associated bronchiolitis (P=0.006). Conclusions Small volumes of whole blood and paediatric respiratory samples can be used for enumerating ILCs (NK cells, ILC1, ILC2, ILC3), granulocytes (eosinophils and neutrophils), T – cells (CD4+ and CD8+), MAIT cells and NKT-like cells. The comparison of neonate and adult functional responses suggests that innate lymphoid cells are mature in early life and might play a pivotal role in the neonatal type-2 bias via the production of IL-13. Bronchiolitis was characterised not only by CD3+ T cell but also by CD3- lymphopenia. ILCs numbers were much lower in infants with bronchiolitis compared to healthy controls; however, ILC had the ability to produce IFN-γ. My data suggest that RSV and RV can lead to different phenotypes of bronchiolitis. Finally, I demonstrated the importance of studying the lower and upper airways simultaneously, discovering that inflammatory signatures were different between these two compartments. For the first time all members of the ILC family were studied in viral bronchiolitis. Determining why some infants develop bronchiolitis and the pathogenesis of lower airway disease in infants could lead to a better understanding of early life immunity and the development of new biomarkers and clinical prophylactics.Open Acces

Accounting for European funds in budget entities

European funds as well as non-returnable foreign funding received by local government entities as part of projects, are classified as income of those entities, the accounting of which is similar, and in accord with existing regulations. The aim of this study is to examine some practical resolutions pertaining to financial records used in accounting departments, as well as practical examples of accounting and presenting information on projects co-funded by European resources

Accounting for European funds in budget entities

European funds as well as non-returnable foreign funding received by local government entities as part of projects, are classified as income of those entities, the accounting of which is similar, and in accord with existing regulations. The aim of this study is to examine some practical resolutions pertaining to financial records used in accounting departments, as well as practical examples of accounting and presenting information on projects co-funded by European resources

Europa som et forandringsprojekt

Valget til Europa-Parlamentet står for døren. Her med en tale afholdt af Paweł Swieboda som er formand for demosEUROPA – Centre for European Strategy. Talen blev afholdt 31. marts 2014 på en EU-koference i København.&nbsp

Rozliczanie srodkуw europejskich przez jednostki budzetowe

Środki europejskie to środki finansowe, które są wykorzystywane, aby zrealizować cel, który ma na celu wspieranie i restrukturyzację gospodarek krajów członkowskich Unii Europejskiej. Budżet Unii Europejskiej tworzą dochody, które przede wszystkim pochodzą z tych państw. Budżety roczne są ustalane w granicach, które wyznaczane są przez długoterminowy plan finansowy. Skutkiem tego jest przewidywalność ponoszonych wydatków. Obecny plan finansowy obowiązuje do 2020 r

Early IL-6 signalling promotes IL-27 dependent maturation of regulatory T cells in the lungs and resolution of viral immunopathology.

Interleukin-6 is a pleiotropic, pro-inflammatory cytokine that can promote both innate and adaptive immune responses. In humans with respiratory virus infections, such as Respiratory Syncytial Virus (RSV), elevated concentrations of IL-6 are associated with more severe disease. In contrast the polymorphisms in the Il6 promoter which favour lower IL-6 production are associated with increased risk of both RSV and Rhinovirus infections. To determine the precise contribution of IL-6 to protection and pathology we used murine models of respiratory virus infection. RSV infection resulted in increased IL-6 production both in the airways and systemically which remained heightened for at least 2 weeks. IL-6 depletion early, but not late, during RSV or Influenza A virus infection resulted in significantly increased disease associated with an influx of virus specific TH1 and cytotoxic CD8+ T cells, whilst not affecting viral clearance. IL-6 acted by driving production of the immunoregulatory cytokine IL-27 by macrophages and monocytes, which in turn promoted the local maturation of regulatory T cells. Concordantly IL-27 was necessary to regulate TH1 responses in the lungs, and sufficient to limit RSV induced disease. Overall we found that during respiratory virus infection the prototypic inflammatory cytokine IL-6 is a critical anti-inflammatory regulator of viral induced immunopathology in the respiratory tract through its induction of IL-27

Rozliczanie srodkуw europejskich przez jednostki budzetowe

Środki europejskie to środki finansowe, które są wykorzystywane, aby zrealizować cel, który ma na celu wspieranie i restrukturyzację gospodarek krajów członkowskich Unii Europejskiej. Budżet Unii Europejskiej tworzą dochody, które przede wszystkim pochodzą z tych państw. Budżety roczne są ustalane w granicach, które wyznaczane są przez długoterminowy plan finansowy. Skutkiem tego jest przewidywalność ponoszonych wydatków. Obecny plan finansowy obowiązuje do 2020 r

Human Rhinovirus 16 Causes Golgi Apparatus Fragmentation without Blocking Protein Secretion

The replication of picornaviruses has been described to cause fragmentation of the Golgi apparatus that blocks the secretory pathway. The inhibition of major histocompatibility complex class I upregulation and cytokine, chemokine and interferon secretion may have important implications for host defense. Previous studies have shown that disruption of the secretory pathway can be replicated by expression of individual nonstructural proteins; however the situation with different serotypes of human rhinovirus (HRV) is unclear. The expression of 3A protein from HRV14 or HRV2 did not cause Golgi apparatus disruption or a block in secretion, whereas other studies showed that infection of cells with HRV1A did cause Golgi apparatus disruption which was replicated by the expression of 3A. HRV16 is the serotype most widely used in clinical HRV challenge studies; consequently, to address the issue of Golgi apparatus disruption for HRV16, we have systematically and quantitatively examined the effect of HRV16 on both Golgi apparatus fragmentation and protein secretion in HeLa cells. First, we expressed each individual nonstructural protein and examined their cellular localization and their disruption of endoplasmic reticulum and Golgi apparatus architecture. We quantified their effects on the secretory pathway by measuring secretion of the reporter protein Gaussia luciferase. Finally, we examined the same outcomes following infection of cells with live virus. We demonstrate that expression of HRV16 3A and 3AB and, to a lesser extent, 2B caused dispersal of the Golgi structure, and these three nonstructural proteins also inhibited protein secretion. The infection of cells with HRV16 also caused significant Golgi apparatus dispersal; however, this did not result in the inhibition of protein secretion. IMPORTANCE The ability of replicating picornaviruses to influence the function of the secretory pathway has important implications for host defense. However, there appear to be differences between different members of the family and inconsistent results when comparing infection with live virus to expression of individual nonstructural proteins. We demonstrate that individual nonstructural HRV16 proteins, when expressed in HeLa cells, can both fragment the Golgi apparatus and block secretion, whereas viral infection fragments the Golgi apparatus without blocking secretion. This has major implications for how we interpret mechanistic evidence derived from the expression of single viral proteins

Natural killer cells and innate lymphoid cells but not NKT cells are mature in their cytokine production at birth.

Early life is a time of increased susceptibility to infectious diseases and development of allergy. Innate lymphocytes are crucial components of the initiation and regulation of immune responses at mucosal surfaces, but functional differences in innate lymphocytes early in life are not fully described. We aimed to characterise the abundance and function of different innate lymphocyte cell populations in cord blood in comparison to that of adults. Blood was collected from adult donors and umbilical vessels at birth. Multicolour flow cytometry panels were used to identify and characterise lymphocyte populations and their capacity to produce hallmark cytokines. Lymphocytes were more abundant in cord blood compared to adults, however, mucosal-associated invariant T (MAIT) cells and Natural Killer T (NKT)-like cells, were far less abundant. The capacity of NKT-like cells to produce cytokines and their expression of the cytotoxic granule protein granzyme B and the marker of terminal differentiation CD57 were much lower in cord blood than in adults. In contrast, Natural Killer (NK) cells were as abundant in cord blood as in adults, they could produce IFNγ, and their expression of granzyme B was not significantly different to that of adult NK cells, although CD57 expression was lower. All innate lymphoid cell (ILC) subsets were more abundant in cord blood, and ILC1 and ILC2 were capable of production of IFNγ and IL-13, respectively. In conclusion, different innate lymphoid cells differ in both abundance and function in peripheral blood at birth and with important implications for immunity in early life

Introduction to methods of modelling information wars as a 21st century threat

Purpose: Using System Dynamics approach together with Lanchester and SIR models for modeling information war. Theoretical considerations. Approach: Due to the theoretical form of conducted research the main research methodso were a literature review and simulations based on developed model. Conclusions: The result of the research is the model of information war based on System Dynamics approach. The model focuses on how socjety wealth and counterdisinformation campaings affect on war efficiency. One of the key conclusion from the simulations results is that one of the main goals of attacking side should be elimination or taking control over public media of attacked one. Practical implications: The model of information war which was developed during the researche, gives a possibility to get new knowledge about war information procesess. It allows to predict causes and effects of disinformation campaings and helps to make proper decisions connected with countermeasuers that are taken. Presented article appoints directions which needs to be explored in connection with information wars. Orginality: Presented researches are pioneerign and in such form on this field was not conducted so far. Using Lanchester equations, connected with epidemic spread model and System Dynamics approach they provide new knowledge about the phenomenon of information war.peer-reviewe