Pro- and anti-inflammatory effects of Nitric Oxide on Dendritic Cells


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PhD thesis: a PhD thesis.
Pro- and anti-inflammatory effects of Nitric Oxide on Dendritic Cells
Graf Lukas Stefan
Carolina Obregon/Laurent Nicod 
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Université de Berne
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Dendritc cells are key players in initiating primary immune responses. In an immature state they reside in nonlymphoid organs, capturing and processing foreign antigens and microbial pathogens. Once they are activated, they migrate to the lymph nodes and the maturation program is triggered leading to the expression of cell surface molecules (such as CD83) and co-stimulatory molecules (such as CD40, CD80 and CD86), as well as to the release of cytokines such as IL-12 and TNF-α. Once in the lymph nodes a potent interaction with T cells initiates the acquired immune response. In the lung, DCs are continually challenged not only with antigens, but also with noxious particles, which are going to influence lung redox state and in consequence influence lung immunity. Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are the most highly reactive molecules involved in numerous physiological processes. Superoxide anions (O2-), hydroxyl radicals (OH-), hydrogen peroxide (H2O2) are most known molecules among ROS and nitric oxide (NO.), Nitrogen dioxide (NO2), Nitrite ion (NO2-), nitrate ion (NO3-) and peroxynitrites (ONOO-) are the most know molecules among the RNS. In this work we have investigated the effect that RNS such as nitric oxide (NO.) can play in the LPS-activation of DCs. NO. (DPTA-NONOate donor) was added to DC-culture in different concentrations as well as in different time points of DC maturation. Modulation of DC response was measured by the release of cytokines (IL-12, IL-10, TNF-α), the expression of maturation markers (CD86, CD83, CD80) and the capacity to present antigensto T cells. Furthermore, to analyze intracellular changes in DCs, three intracellular pathways were studied, NFB, p38 and ERK. The viability of DCs during the experiments was analyzed by annexin-V and PI. The results have shown that NO. is an important factor able to modulate an inflammatory or anti-inflammatory process depending on the concentration used: IL-10 and TNF-α were released in a dose dependent manner while the release of IL-12 was inhibited in a dose dependent manner. NO. also modulates inflammation depending on the maturation state of the cells. NO. have shown to have an inhibitory role on immature DCs, modulating the inflammation induced by LPS by decreasing the level of IL-12 and IL-10. In contrast, if NO. is added after maturation, it seems to strengthen inflammatory response of mature DCs increasing the release of TNF-α and IL-12. Interestingly, NO. is not able to induce any changes in the phenotype of mature DCs. As expected, NO. alone in high concentrations (1mM) was toxic for DCs, LPS induced to some extend apoptosis, however it do not induce an extensive cell death of DCs. Interestingly, when NO. was added to DCs either before or after LPS maturation, NO. was protective on the DC population decreasing the necrosis and apoptosis rate. Concerning the intracellular changes, a slight modulation of intracellular pathways such as MAPK ERK 1/2 p38 and NF-κB was observed. NO. is able to modulate more specifically the phosphorylation state of MAPK ERK1/2, when added before LPS-maturation, but MAPK p38 and NF-κB seems to be more involved after LPS maturation, Summarising we have shown, that NO. potently modulates LPS induced DCs activation, depending on their concentration and the time point used during the maturation of DCs. NO. is able to lead a Th2 response and high TNF-α release when used in high concentration, in contrast to low concentration of NO. which leads to enhanced IL-12 release favouring rather the development of Th1 subset. NO. co-modulates the LPS activation in DCs probably by inducing a differential kinetics of activation ERK1/2, NF-κB and p38.
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12/02/2010 16:10
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21/08/2019 5:33
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