Clearance of apoptotic cells by macrophages induces regulatory phenotype and involves stimulation of cd36 and platelet-activating factor receptor

Phagocytosis of apoptotic cells (efferocytosis) induces macrophage differentiation towards a regulatory phenotype (IL-10high/IL-12p40low). CD36 is involved in the recognition of apoptotic cells (AC), and we have shown that the platelet-activating factor receptor (PAFR) is also involved. Here, we investigated the contribution of PAFR and CD36 to efferocytosis and to the establishment of a regulatory macrophage phenotype. Mice bone marrow-derived macrophages were cocultured with apoptotic thymocytes, and the phagocytic index was determined. Blockage of PAFR with antagonists or CD36 with specific antibodies inhibited the phagocytosis of AC (~70–80%). Using immunoprecipitation and confocal microscopy, we showed that efferocytosis increased the CD36 and PAFR colocalisation in the macrophage plasma membrane; PAFR and CD36 coimmunoprecipitated with flotillin-1, a constitutive lipid raft protein, and disruption of these membrane microdomains by methyl-β-cyclodextrin reduced AC phagocytosis. Efferocytosis induced a pattern of cytokine production, IL-10high/IL-12p40low, that is, characteristic of a regulatory phenotype. LPS potentiated the efferocytosis-induced production of IL-10, and this was prevented by blocking PAFR or CD36. It can be concluded that phagocytosis of apoptotic cells engages CD36 and PAFR, possibly in lipid rafts, and this is required for optimal efferocytosis and the establishment of the macrophage regulatory phenotype

Oxidized LDL induces alternative macrophage phenotype through activation of CD36 and PAFR

OxLDL is recognized by macrophage scavenger receptors, including CD36; we have recently found that Platelet-Activating Factor Receptor (PAFR) is also involved. Since PAFR in macrophages is associated with suppressor function, we examined the effect of oxLDL on macrophage phenotype. It was found that the presence of oxLDL during macrophage differentiation induced high mRNA levels to IL-10, mannose receptor, PPARγ and arginase-1 and low levels of IL-12 and iNOS. When human THP-1 macrophages were pre-treated with oxLDL then stimulated with LPS, the production of IL-10 and TGF-β significantly increased, whereas that of IL-6 and IL-8 decreased. In murine TG-elicited macrophages, this protocol significantly reduced NO, iNOS and COX2 expression. Thus, oxLDL induced macrophage differentiation and activation towards the alternatively activated M2-phenotype. In murine macrophages, oxLDL induced TGF-β, arginase-1 and IL-10 mRNA expression, which were significantly reduced by pre-treatment with PAFR antagonists (WEB and CV) or with antibodies to CD36. The mRNA expression of IL-12, RANTES and CXCL2 were not affected. We showed that this profile of macrophage activation is dependent on the engagement of both CD36 and PAFR. We conclude that oxLDL induces alternative macrophage activation by mechanisms involving CD36 and PAFR

Immune complex induced arthritis in rats: role of lipid mediators on cell infiltration

We investigated the participation of lipid mediators in an experimental immune complex (IC) arthritis model in rats. The animals were subjected to intraarticular injection of anti-bovine sertLm albumin (BSA) IgG antibodies followed by i.v. injection of BSA. Histopathological analysis of the synovial membranes disclosed infiltration of polymorphonuclear (PMN) cells and vascular congestion. Slight increase in vascular permeability, measured by Evans blue dye extravasation into the joints, was detected after 3 h of arthritis. Cellular influx into the articular cavities was most evident at the sixth hour of arthritis with predominance of PMN. Pretreatment with either indomethacin, a cyclooxygenase inhibitor, or L-660,711, a peptido-leukotriene antagonist, did not inhibit cell infiltration, whereas pretreatment with either L-663,536, a 5-lipoxygenase inhibitor, or L-655,240, a thromboxane antagonist, significantly inhibited the phenomenon. Pretreatment with WEB 2170, a platelet activating factor (PAF) antagonist, also significantly inhibited cell influx. These results suggest that thromboxane, LTB4 and PAF mediate cell infiltration in this IC arthritis model

Boosting Adaptive Immunity: A New Role for PAFR Antagonists.

We have previously shown that the Platelet-Activating Factor Receptor (PAFR) engagement in murine macrophages and dendritic cells (DCs) promotes a tolerogenic phenotype reversed by PAFR-antagonists treatment in vitro. Here, we investigated whether a PAFR antagonist would modulate the immune response in vivo. Mice were subcutaneously injected with OVA or OVA with PAFR-antagonist WEB2170 on days 0 and 7. On day 14, OVA-specific IgG2a and IgG1 were measured in the serum. The presence of WEB2170 during immunization significantly increased IgG2a without affecting IgG1 levels. When WEB2170 was added to OVA in complete Freund's adjuvant, enhanced IgG2a but not IgG1 production was also observed, and CD4+ FoxP3+ T cell frequency in the spleen was reduced compared to mice immunized without the antagonist. Similar results were observed in PAFR-deficient mice, along with increased Tbet mRNA expression in the spleen. Additionally, bone marrow-derived DCs loaded with OVA were transferred into naïve mice and their splenocytes were co-cultured with fresh OVA-loaded DCs. CD4(+) T cell proliferation was higher in the group transferred with DCs treated with the PAFR-antagonist. We propose that the activation of PAFR by ligands present in the site of immunization is able to fine-tune the adaptive immune response

Modulation of Leishmania (L.) amazonensis Growth in Cultured Mouse Macrophages by Prostaglandins and Platelet Activating Factor

The role of endogenously synthesized PAF and prostaglandins on the infection of mouse macrophages by Letsbmanta (L.) amazonensis was investigated, as well as the possible correlation between the effects of these inflammatory mediators with nitric oxide production. It was found that pretreatment of macrophages with 10−5 M of the PAF antagonists, BN-52021 or WEB-2086, increased macrophage infection by 17 and 59%, respectively. The cyclooxygenase inhibitor, indomethacin (10 μg/ml), induced a significant inhibition which was reversed by addition of PGE (10-3 M) to the culture medium. These results suggested that the infection of macrophages by leisbmanla is inhibited by PAF and enhanced by prostaglandins and that these mediators are produced by macrophages during this infection. This was confirmed by addition of these mediators to the culture medium before infection; PAF (10−6, 10−9 and 10−12M) reduced significantly the infection whereas PGE2 (10−5 M) induced a marked enhancement. This effect of exogenous PAF on macrophage infection was reversed by the two PAF antagonists used in this study as well as by the inhibitor of nitric oxide synthesis, L-arginine methyl ester (100 mM). Taken together the data suggest that endogenous production of PAF and PGE2 exert opposing effects on Lesbmana–macrophage interaction and that nitric oxide may be involved in the augmented destruction of parasites induced by PAF

Reachability in Higher-Order-Counters

Higher-order counter automata (\HOCS) can be either seen as a restriction of higher-order pushdown automata (\HOPS) to a unary stack alphabet, or as an extension of counter automata to higher levels. We distinguish two principal kinds of \HOCS: those that can test whether the topmost counter value is zero and those which cannot. We show that control-state reachability for level $k$ \HOCS with $0$-test is complete for \mbox{$(k-2)$}-fold exponential space; leaving out the $0$-test leads to completeness for \mbox{$(k-2)$}-fold exponential time. Restricting \HOCS (without $0$-test) to level $2$, we prove that global (forward or backward) reachability analysis is \PTIME-complete. This enhances the known result for pushdown systems which are subsumed by level $2$ \HOCS without $0$-test. We transfer our results to the formal language setting. Assuming that \PTIME \subsetneq \PSPACE \subsetneq \mathbf{EXPTIME}, we apply proof ideas of Engelfriet and conclude that the hierarchies of languages of \HOPS and of \HOCS form strictly interleaving hierarchies. Interestingly, Engelfriet's constructions also allow to conclude immediately that the hierarchy of collapsible pushdown languages is strict level-by-level due to the existing complexity results for reachability on collapsible pushdown graphs. This answers an open question independently asked by Parys and by Kobayashi.Comment: Version with Full Proofs of a paper that appears at MFCS 201

Thin-film flow in helically wound rectangular channels with small torsion

Laminar gravity-driven thin-film flow down a helically-wound channel of rectangular cross-section with small torsion in which the fluid depth is small is considered. Neglecting the entrance and exit regions we obtain the steady-state solution that is independent of position along the axis of the channel, so that the flow, which comprises a primary flow in the direction of the axis of the channel and a secondary flow in the cross-sectional plane, depends only on position in the two-dimensional cross-section of the channel. A thin-film approximation yields explicit expressions for the fluid velocity and pressure in terms of the free-surface shape, the latter satisfying a non-linear ordinary differential equation that has a simple exact solution in the special case of a channel of rectangular cross-section. The predictions of the thin-film model are shown to be in good agreement with much more computationally intensive solutions of the small-helix-torsion Navier–Stokes equations. The present work has particular relevance to spiral particle separators used in the mineral-processing industry. The validity of an assumption commonly used in modelling flow in spiral separators, namely that the flow in the outer region of the separator cross-section is described by a free vortex, is shown to depend on the problem parameters

A novel murine model of late-phase reaction of immediate hypersensitivity

We describe here a novel experimental model of late-phase reaction of immediate hypersensitivity developed in mice. It consists of introducing small fragments of heat-coagulated hen egg white into the subcutaneous tissue of mice. After 14 days, animals challenged with purified ovalbumin into the footpad presented an immediate swelling of the paw peaking at 30 min, followed by two peaks of swelling at 6 and 24 h. Histological examination of the paws showed a massive eosinophil infiltration (more than 800 cells/5 microscopic fields). This intense infiltration persisted for more than 14 days after the challenge. Furthermore, in mice immunized with coagulated egg white the delayed swelling of the paws and eosinophilic infiltration were significantly higher than in mice immunized with the classical protocol of ovalbumin in alumen adjuvant. Transfer of lymph node cells obtained from mice implanted with heat-coagulated hen egg white induced footpad swelling and eosinophil infiltration in response to ovalbumin. High levels of ovalbuminspecific IgG1 but not of IgE were detected in the serum of these animals. The advantages of this model for the experimental study of late-phase reaction per se and its relevance to the study of allergic diseases such as asthma are discussed

Current issues in research on structure–property relationships in polymer nanocomposites

The understanding of the basic physical relationships between nano-scale structural variables and the macroscale properties of polymer nanocomposites remains in its infancy. The primary objective of this article is to ascertain the state of the art regarding the understanding and prediction of the macroscale properties of polymers reinforced with nanometer-sized solid inclusions over a wide temperature range. We emphasize that the addition of nanoparticles with large specific surface area to polymer matrices leads to amplification of a number of rather distinct molecular processes resulting from interactions between chains and solid surfaces. This results in a “non-classical” response of these systems to mechanical and electro-optical excitations when measured on the macroscale. For example, nanoparticles are expected to be particularly effective at modifying the intrinsic nano-scale dynamic heterogeneity of polymeric glass-formation and, correspondingly, recent simulations indicate that both the strength of particle interaction with the polymer matrix and the particle concentration can substantially influence the dynamic fragility of polymer glass-formation, a measure of the strength of the temperature dependence of the viscosity or structural relaxation time. Another basic characteristic of nanoparticles in polymer matrices is the tendency for the particles to associate into extended structures that can dominate the rheological, viscoelastic and mechanical properties of the nanocomposite so that thermodynamic factors that effect nanoparticle dispersion can be crucially important. Opportunities to exploit knowledge gained from understanding biomechanics of hierarchical biological protein materials and potential applications in materials design and nanotechnology are among future research challenges. Research on nanocomposites formed from block copolymers and nanoparticles offers huge promise in molecular electronics and photovoltaics. The surface functionalization of nanoparticles by the grafting of polymer brushes is expected to play important role in the designing of novel organic/inorganic nanocomposite materials. The formation of bulk heterojunctions at the nanometer scale leads to efficient dissociation of the charge pairs generated under sunlight. Based on the presentations and discussion, we make recommendations for future work in this area by the physics, chemistry, and engineering communities.Czech Republic. Ministry of Education, Youth, and Sports (MSM0021630501

Uptake of oxLDL and IL-10 production by macrophages requires PAFR and CD36 recruitment into the same lipid rafts

Macrophage interaction with oxidized low-density lipoprotein (oxLDL) leads to its differentiation into foam cells and cytokine production, contributing to atherosclerosis development. In a previous study, we showed that CD36 and the receptor for platelet-activating factor (PAFR) are required for oxLDL to activate gene transcription for cytokines and CD36. Here, we investigated the localization and physical interaction of CD36 and PAFR in macrophages stimulated with oxLDL. We found that blocking CD36 or PAFR decreases oxLDL uptake and IL-10 production. OxLDL induces IL-10 mRNA expression only in HEK293T expressing both receptors (PAFR and CD36). OxLDL does not induce IL-12 production. The lipid rafts disruption by treatment with βCD reduces the oxLDL uptake and IL-10 production. OxLDL induces co-immunoprecipitation of PAFR and CD36 with the constitutive raft protein flotillin-1, and colocalization with the lipid raft-marker GM1-ganglioside. Finally, we found colocalization of PAFR and CD36 in macrophages from human atherosclerotic plaques. Our results show that oxLDL induces the recruitment of PAFR and CD36 into the same lipid rafts, which is important for oxLDL uptake and IL-10 production. This study provided new insights into how oxLDL interact with macrophages and contributing to atherosclerosis development