Plasma membrane proteomes of differentially matured dendritic cells identified by LC-MS/MS combined with iTRAQ labelling

AbstractDendritic cells (DCs) play a pivotal role in polarising Th lymphocyte subsets but it is unclear what molecular events occur when DCs generate Th2-type responses. Here, we analysed plasma membrane-enriched fractions from immature, pro-Th1 and pro-Th2 DCs and used a combination of iTRAQ labelling and LC–MS/MS to quantify changes in the proteomes. Analysis was performed on triplicate biological samples and changes verified by flow cytometry. MHC class II molecules and CD29 were up-regulated in pro-Th1 DCs whilst CD18 and CD44 were up-regulated in pro-Th2 DCs. One of the most down-regulated molecules in pro-Th1 DCs was YM-1 whilst the greatest decrease in pro-Th2 DCs was NAP-22. Other molecules up-regulated in pro-Th2 DC compared to pro-Th1 DCs included some potentially involved in protein folding during antigen processing (clathrin and Rab-7), whilst other non-membrane proteins such as enzymes/transporters related to cell metabolism (malate dehydrogenase, pyruvate kinase, and ATPase Na+/K+) were also recorded. This suggests that pro-Th2 DCs are more metabolically active while pro-Th1 DCs have a mature ‘end state’. Overall, although several molecules were preferentially expressed on pro-Th2 DCs, our proteomics data support the view of a ‘limited maturation’ of pro-Th2 DCs compared to pro-Th1 DCs

Cellular and Molecular Mechanisms Underlying the Strong Neonatal IL-12 Response of Lamb Mesenteric Lymph Node Cells to R-848

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Circulating CD14brightCD16+ 'intermediate' monocytes exhibit enhanced parasite pattern recognition in human helminth infection.

Circulating monocyte sub-sets have recently emerged as mediators of divergent immune functions during infectious disease but their role in helminth infection has not been investigated. In this study we evaluated whether 'classical' (CD14brightCD16-), 'intermediate' (CD14brightCD16+), and 'non-classical' (CD14dimCD16+) monocyte sub-sets from peripheral blood mononuclear cells varied in both abundance and ability to bind antigenic material amongst individuals living in a region of Northern Senegal which is co-endemic for Schistosoma mansoni and S. haematobium. Monocyte recognition of excretory/secretory (E/S) products released by skin-invasive cercariae, or eggs, of S. mansoni was assessed by flow cytometry and compared between S. mansoni mono-infected, S. mansoni and S. haematobium co-infected, and uninfected participants. Each of the three monocyte sub-sets in the different infection groups bound schistosome E/S material. However, 'intermediate' CD14brightCD16+ monocytes had a significantly enhanced ability to bind cercarial and egg E/S. Moreover, this elevation of ligand binding was particularly evident in co-infected participants. This is the first demonstration of modulated parasite pattern recognition in CD14brightCD16+ intermediate monocytes during helminth infection, which may have functional consequences for the ability of infected individuals to respond immunologically to infection

Mangafodipir Protects against Hepatic Ischemia-Reperfusion Injury in Mice

Mangafodipir is a contrast agent used in magnetic resonance imaging that concentrates in the liver and displays pleiotropic antioxidant properties. Since reactive oxygen species are involved in ischemia-reperfusion damages, we hypothesized that the use of mangafodipir could prevent liver lesions in a mouse model of hepatic ischemia reperfusion injury. Mangafodipir (MnDPDP) was compared to ischemic preconditioning and intermittent inflow occlusion for the prevention of hepatic ischemia-reperfusion injury in the mouse.Mice were subjected to 70% hepatic ischemia (continuous ischemia) for 90 min. Thirty minutes before the ischemic period, either mangafodipir (10 mg/kg) or saline was injected intraperitoneally. Those experimental groups were compared with one group of mice preconditioned by 10 minutes' ischemia followed by 15 minutes' reperfusion, and one group with intermittent inflow occlusion. Hepatic ischemia-reperfusion injury was evaluated by measurement of serum levels of aspartate aminotransferase (ASAT) activity, histologic analysis of the livers, and determination of hepatocyte apoptosis (cytochrome c release, caspase 3 activity). The effect of mangafodipir on the survival rate of mice was studied in a model of total hepatic ischemia.<0.01), and by higher rates of survival in treated than in untreated animals (P<0.001). The level of protection by mangafodipir was similar to that observed following intermittent inflow occlusion and higher than after ischemic preconditioning.Mangafodipir is a potential new preventive treatment for hepatic ischemia-reperfusion injury

Chronic Helminth Infections Protect Against Allergic Diseases by Active Regulatory Processes

Developed countries are suffering from an epidemic rise in immunologic disorders, such as allergy-related diseases and certain autoimmunities. Several studies have demonstrated a negative association between helminth infections and inflammatory diseases (eg, allergy), providing a strong case for the involvement of helminth infections in this respect. However, some studies point in the opposite direction. The discrepancy may be explained by differences in frequency, dose, time, and type of helminth. In this review, new studies are discussed that may support the concept that chronic helminth infections in particular—but not acute infections—are associated with the expression of regulatory networks necessary for downmodulating allergic immune responses to harmless antigens. Furthermore, different components of regulatory networks are highlighted, such as the role of regulatory T and B cells, modulation of dendritic cells, early innate signals from structural cells (eg, epithelial cells), and their individual contributions to protection against allergic diseases. It is of great interest to define and characterize specific helminth molecules that have profound immunomodulatory capacities as targets for therapeutic application in the treatment or prophylaxis of allergic manifestations

Macrophage cell lines use CD81 in cell growth regulation

CD81 is an integral membrane protein belonging to the tetraspanin superfamily. It has two extracellular domains that interact with cell surface proteins and two intracellular tails that contribute to cellular processes. Although there are considerable data about how CD81 affects T- and B-cell function, not much is known about how it impacts macrophages. To address this, we established four cell lines from mouse bone marrow in the presence of macrophage colony-stimulating factor and transfection with SV40 large T antigen. Two were CD81[superscript]−/− (ASD1 and ASD2) and two were CD81[superscript]+/− (2ASD1.10 and 2BSD1.10). Cells were Mac-2-, PU.1-, and c-fms-positive and all the cell lines were phagocytic indicating that they were macrophage-like. In mixtures of the two cell types in tissue culture, CD81[superscript]−/− cells out competed CD81[superscript]+/− cells with CD81-bearing cells being undetectable after 50 cell culture passages. Although cell divisions during log-phase growth were not significantly different between CD81[superscript]+/− macrophage cells and CD81[superscript]−/− macrophage cells, we found that CD81[superscript]−/− macrophage cells reached a higher density at confluency than CD81[superscript]+/− macrophage cells. CD81 transcript levels increased as cultures became confluent, but transcript levels of other tetraspanin-related molecules remained relatively constant. Transfection of CD81 into ASD1 (CD81[superscript]−/−) cells reduced the density of confluent cultures of transformants compared to cells transfected with vector alone. These data suggest that CD81 potentially plays a role in macrophage cell line growth regulation

Propionic fermentation by the probiotic Propionibacterium freudenreichii to functionalize whey

International audienceA new probiotic functionalized sweet whey was evaluated. Weaned healthy piglets consumed sweet whey (SW), unmodified or fermented by P. freudenreichii CIRM-BIA 129 (PF-SW). Fecal short chain fatty acids amounts remained unchanged. Bifidobacteria were enhanced in the PF-SW group, and so was the expression of T-bet, which orchestrates Th1 differentiation of T lymphocytes, in mesenteric lymph nodes immune cells (MLNC). This was consistent with ex vivo increased TNF-α secretion by MLNC in response to lipopolysaccharide (LPS). The consumption of the functionalized whey induced a different response in peripheral blood mononuclear cells (PBMC) to ex vivo stimulations, as the inhibition of TNF-α secretion in response to concanavalin A stimulation. Thus, by cultivating a probiotic GRAS bacterium in concentrated whey, prior to spray drying, it is possible to transform this by-product into a functional ingredient. This opens new avenues for the development of functional ingredients through enhanced valorisation of whey

Simulation of the large-area growth of homoepitaxial 4H-Sic by chemical vapor deposition

The growth of thick epitaxial 4H-SiC layers with low defect density is an essential step for the fabrication of SiC based devices. Cold- and hot-wall reactors using silane and propane diluted in hydrogen were used in this study. The typical growth temperature range is 1700-1900 K and total pressure range 10-100 kPa. The resulting epilayers exhibit low background doping, low defect density and good thickness uniformity. The main problem is that it is difficult with this first generation of reactors to ensure a good uniformity of deposition over large wafer dimension. A 3D simulation approach of heat and mass transfer was used with two objectives. The first one is to evaluate the electrical, thermal, transport and kinetic databases. The second one is to have a visualization and a quantification of the flow, temperature and gaseous species fields for a better understanding of engineering problems at elevated temperatures

Plasticity of migrating CD1b+ and CD1b- lymph dendritic cells in the promotion of Th1, Th2 and Th17 in response to Salmonella and helminth secretions.

Dendritic cells (DCs) are pivotal in the development of specific T-cell responses to control pathogens, as they govern both the initiation and the polarization of adaptive immunity. To investigate the capacities of migrating DCs to respond to pathogens, we used physiologically generated lymph DCs (L-DCs). The flexible polarization of L-DCs was analysed in response to Salmonella or helminth secretions known to induce different T cell responses. Mature conventional CD1b(+) L-DCs showed a predisposition to promote pro-inflammatory (IL-6), pro-Th1 (IL-12p40) and anti-inflammatory (IL-10) responses which were amplified by Salmonella, and limited to only IL-6 induction by helminth secretions. The other major population of L-DCs did not express the CD1b molecule and displayed phenotypic features of immaturity compared to CD1b(+) L-DCs. Salmonella infection reduced the constitutive expression of TNF-α and IL-4 mRNA in CD1b(-) L-DCs, whereas this expression was not affected by helminth secretions. The cytokine response of T cells promoted by L-DCs was analysed in T cell subsets after co-culture with Salmonella or helminth secretion-driven CD1b(+) or CD1b(-) L-DCs. T cells preferentially expressed the IL-17 gene, and to a lesser extent the IFN-γ and IL-10 genes, in response to Salmonella-driven CD1b(+) L-DCs, whereas a preferential IL-10, IFN-γ and IL-17 gene expression was observed in response to Salmonella-driven CD1b(-) L-DCs. In contrast, a predominant IL-4 and IL-13 gene expression by CD4(+) and CD8(+) T cells was observed after stimulation of CD1b(+) and CD1b(-) L-DCs with helminth secretions. These results show that mature conventional CD1b(+) L-DCs maintain a flexible capacity to respond differently to pathogens, that the predisposition of CD1b(-) L-DCs to promote a Th2 response can be oriented towards other Th responses, and finally that the modulation of migrating L-DCs responses is controlled more by the pathogen encountered than the L-DC subsets