Immune Response Of Macrophages Research Articles

Modulation of Human Macrophage Responses to Mycobacterium tuberculosis by Silver Nanoparticles of Different Size and Surface Modification

Exposure to silver nanoparticles (AgNP) used in consumer products carries potential health risks including increased susceptibility to infectious pathogens. Systematic assessments of antimicrobial macrophage immune responses in the context of AgNP exposure are important because uptake of AgNP by macrophages may lead to alterations of innate immune cell functions. In this study we examined the effects of exposure to AgNP with different particle sizes (20 and 110 nm diameters) and surface chemistry (citrate or polyvinlypyrrolidone capping) on cellular toxicity and innate immune responses against Mycobacterium tuberculosis (M.tb) by human monocyte-derived macrophages (MDM). Exposures of MDM to AgNP significantly reduced cellular viability, increased IL8 and decreased IL10 mRNA expression. Exposure of M.tb-infected MDM to AgNP suppressed M.tb-induced expression of IL1B, IL10, and TNFA mRNA. Furthermore, M.tb-induced IL-1β, a cytokine critical for host resistance to M.tb, was inhibited by AgNP but not by carbon black particles indicating that the observed immunosuppressive effects of AgNP are particle specific. Suppressive effects of AgNP on the M.tb-induced host immune responses were in part due to AgNP-mediated interferences with the TLR signaling pathways that culminate in the activation of the transcription factor NF-κB. AgNP exposure suppressed M.tb-induced expression of a subset of NF-κB mediated genes (CSF2, CSF3, IFNG, IL1A, IL1B, IL6, IL10, TNFA, NFKB1A). In addition, AgNP exposure increased the expression of HSPA1A mRNA and the corresponding stress-induced Hsp72 protein. Up-regulation of Hsp72 by AgNP can suppress M.tb-induced NF-κB activation and host immune responses. The observed ability of AgNP to modulate infectious pathogen-induced immune responses has important public health implications.

Lipocalin-2 ensures host defense against Salmonella Typhimurium by controlling macrophage iron homeostasis and immune response.

Lipocalin-2 (Lcn2) is an innate immune peptide with pleiotropic effects. Lcn2 binds iron-laden bacterial siderophores, chemo-attracts neutrophils and has immunomodulatory and apoptosis-regulating effects. In this study, we show that upon infection with Salmonella enterica serovar Typhimurium, Lcn2 promotes iron export from Salmonella-infected macrophages, which reduces cellular iron content and enhances the generation of pro-inflammatory cytokines. Lcn2 represses IL-10 production while augmenting Nos2, TNF-α, and IL-6 expression. Lcn2(-/-) macrophages have elevated IL-10 levels as a consequence of increased iron content. The crucial role of Lcn-2/IL-10 interactions was further demonstrated by the greater ability of Lcn2(-/-) IL-10(-/-) macrophages and mice to control intracellular Salmonella proliferation in comparison to Lcn2(-/-) counterparts. Overexpression of the iron exporter ferroportin-1 in Lcn2(-/-) macrophages represses IL-10 and restores TNF-α and IL-6 production to the levels found in wild-type macrophages, so that killing and clearance of intracellular Salmonella is promoted. Our observations suggest that Lcn2 promotes host resistance to Salmonella Typhimurium infection by binding bacterial siderophores and suppressing IL-10 production, and that both functions are linked to its ability to shuttle iron from macrophages.

TREM2 mRNA Expression in Leukocytes Is Increased in Alzheimer’s Disease and Schizophrenia

TREM2 and TYROBP are causal genes for Nasu–Hakola disease (NHD), a rare autosomal recessive disease characterized by bone lesions and early-onset progressive dementia. TREM2 forms a receptor signaling complex with TYROBP, which triggers the activation of immune responses in macrophages and dendritic cells, and the functional polymorphism of TREM2 is reported to be associated with neurodegenerative disorders such as Alzheimer’s disease (AD). The objective of this study was to reveal the involvement of TYROBP and TREM2 in the pathophysiology of AD and schizophrenia. Methods: We investigated the mRNA expression level of the 2 genes in leukocytes of 26 patients with AD and 24 with schizophrenia in comparison with age-matched controls. Moreover, we performed gene association analysis between these 2 genes and schizophrenia. Results: No differences were found in TYROBP mRNA expression in patients with AD and schizophrenia; however, TREM2 mRNA expression was increased in patients with AD and schizophrenia compared with controls (P < 0.001). There were no genetic associations of either gene with schizophrenia in Japanese patients. Conclusion: TREM2 expression in leukocytes is elevated not only in AD but also in schizophrenia. Inflammatory processes involving TREM2 may occur in schizophrenia, as observed in neurocognitive disorders such as AD. TREM2 expression in leukocytes may be a novel biomarker for neurological and psychiatric disorders.

660 nm Red LED Induces Secretory Leukocyte Protease Inhibitor (SLPI) in Lipopolysaccharide-Stimulated RAW264.7 Cell

SLPI acts as a modulator of the innate immune responses of macrophages, neutrophils and odontoblasts, and LPS-inducible anti-inflammatory cytokine to suppress the production of pro-inflammatory products by macrophages. Many studies have revealed the effects of light emitting diodes (LEDs) on the tissue repair and inflammatory responses. Although the anti-inflammatory mechanisms of irradiation with LEDs in gingival fibroblasts are known, the effects of 660 nm red LEDs on the inflammation remain unclear. Moreover, there is no report regarding the molecular mechanism for the relationship between SLPI and biological effects of LEDs. The effects of 660 nm red LEDs on inflammation with SLPI were investigated by examining the effects of 660 nm LED on the SLPI expression of RAW264.7 cells after LPS stimulation. This paper reports that the 660 nm red LED induced SLPI expression or reduced the LPS response, and inhibited NF-κB activation directly, leading to the suppression of pro-inflammatory cytokines, such as TNF-α and IL-1β, suggesting that it might be a useful wavelength LED for inflammation therapy.

IRAK-M regulates the inhibition of TLR-mediated macrophage immune response during late in vitro Leishmania donovani infection.

Intramacrophage protozoan parasite Leishmania donovani, causative agent of visceral leishmaniasis, escapes Toll-like receptor (TLR) dependent early host immune response by inducing the deubiquitinating enzyme A20, which is sustained up to 6 h postinfection only. Therefore, Leishmania must apply other means to deactivate late host responses. Here, we elucidated the role of IL-1 receptor-associated kinase M (IRAK-M), a negative regulator of TLR signaling, in downregulating macrophage proinflammatory response during late hours of in vitro infection. Our data reveal a sharp decline in IRAK1 and IRAK4 phosphorylation at 24 h postinfection along with markedly reduced association of IRAK1-TNF receptor associated factor 6, which is mandatory for TLR activation. In contrast, IRAK-M was induced after A20 levels decreased and reached a maximum at 24 h postinfection. IRAK-M induction coincided with increased stimulation of TGF-β, a hallmark cytokine of visceral infection. TGF-β-dependent signaling-mediated induction of SMAD family of proteins, 2, 3, and 4 plays important roles in transcriptional upregulation of IRAK-M. In infected macrophages, siRNA-mediated silencing of IRAK-M displayed enhanced IRAK1 and IRAK4 phosphorylation with a concomitant increase in downstream NF-κB activity and reduced parasite survival. Taken together, the results suggest that IRAK-M may be targeted by L. donovani to inhibit TLR-mediated proinflammatory response late during in vitro infection.

Analysis of porcine macrophage immune response to antigenic molecules and short chain fatty acids

Analysis of porcine macrophage immune response to antigenic molecules and short chain fatty acids

Stk38 protein kinase preferentially inhibits TLR9-activated inflammatory responses by promoting MEKK2 ubiquitination in macrophages.

NDR/LATS kinase family is highly conserved from yeast to human. It remains unknown whether the members of this family function in innate immune responses. Here we demonstrate that Stk38 negatively regulates TLR9-mediated immune responses in macrophages. Stk38 constitutively associates with ubiquitin E3 ligase Smurf1, and facilitates Smurf1-mediated MEKK2 ubiquitination and degradation. MEKK2 is required for CpG-induced ERK1/2 activation, TNF-α and IL-6 production but not required for LPS-induced TNF-α and IL-6 production. Accordingly, Stk38 deficiency increases CpG-induced ERK1/2 activation, TNF-α and IL-6 production without significantly affecting LPS-induced TNF-α and IL-6 production. Stk38-deficient mice produce more TNF-α and IL-6, and display increased lethality than control wild-type mice upon E. coli infection. Stk38-deficient mice are also more susceptible to CLP-induced sepsis than control mice. Thus, Stk38 is important in limiting inflammatory cytokine production and necessary for protecting host from inflammatory injury during infection, possibly by negatively regulating TLR9 signalling.

An Adenoviral Vector Encoding Full-Length Dectin-1 Promotes Aspergillus-Induced Innate Immune Response in Macrophages

The incidence of invasive pulmonary aspergillosis (IPA) has increased significantly over the last two decades. Alveolar macrophages (AMs) represent the first line of pulmonary host response to Aspergillus conidia. Recognition of conidia by AMs involves Dectin-1 (CLEC7A), which is a conserved structure to combine β-glucans. The deficiency of Dectin-1 results in impaired fungal killing and uncontrolled growth of Aspergillus fumigatus. Thus, we hypothesized that high expression of Dectin-1 would enhance the host recognition and fungal killing. We set out to develop an adenoviral vector encoding full-length Dectin-1 (Ad-Dectin-1-EGFP) and then transfect it to MH-S cells. Transfect cell model was verified by using real-time RT-PCR, Western blot, flow cytometric, and confocal microscopic assays. And also, the function of Dectin-1 was explored by measuring cytokine release and killing ability during the course of A. fumigatus infection. We constructed a recombinant adenovirus which could upregulate the expression of Dectin-1 and verified that Dectin-1 was expressed on cell membrane. The function of Dectin-1 was also demonstrated by its ability in promoting the production of cytokines and increasing the killing ability during the course of A. fumigatus infection. An adenoviral vector was successfully applied to the production of a recombinant adenovirus encoding full-length Dectin-1, and also, its function in Aspergillus-induced innate immune response was demonstrated.

Role of hydrogen sulfide in myocardial infarction via immue response of macrophages (IRM5P.645)

Abstract Macrophages are critical players in the inflammation, and are of central importance for healing after Myocardial infarction (MI). Hydrogen sulfide (H2S) has been demonstrated to possess cardioprotection in MI. However, it is unclear whether and how macrophage modulation by H2S can affect infarct repair. Thus, the present study was to evaluate the roles and mechanisms of H2S on macrophage post-MI. The cardiac function were improved in both WT mice and CSE KO mice, pretreated with different concentrations of NaHS (Exogenous sources of H2S). Meanwhile, the cardiac tissues that treated with NaHS exerted an increased immunohistochemical staining for macrophage marker (Galectin-3). Enhanced M2 macrophage polarization was observed in the infarcted heart treatment with NaHS following MI, along with increased production of M2 signature cytokines, such as interleukin-10 and CD163. In vitro, NaHS accelerated the migration of macrophage cell line RAW264.7. While, the inhibitors (refer to Pyk2, FAK, Src, and Rac1), not only significantly restored the migratory ability in response to the NaHS, but also blocked the activation of phospho-Src, -Pyk2, -FAK397, and -FAK925. Furthermore, the internalization of integrin β1 on the macrophage surface was induced by NaHS. Whereas, integrin β1 silencing inhibited the macrophage migration and the Src signaling activation. The results suggest that H2S may have the potential as an anti-infarct of MI by immune response of macrophages.

Abolishing Cell Wall Glycosylphosphatidylinositol-Anchored Proteins in Candida albicans Enhances Recognition by Host Dectin-1.

Fungi can shield surface pathogen-associated molecular patterns (PAMPs) for evading host immune attack. The most common and opportunistic human pathogen, Candida albicans, can shield β-(1 3)-glucan on the cell wall, one of the major PAMPs, to avoid host phagocyte Dectin-1 recognition. The way to interfere in the shielding process for more effective antifungal defense is not well established. In this study, we found that deletion of the C. albicans GPI7 gene, which was responsible for adding ethanolaminephosphate to the second mannose in glycosylphosphatidylinositol (GPI) biosynthesis, could block the attachment of most GPI-anchored cell wall proteins (GPI-CWPs) to the cell wall and subsequently unmask the concealed β-(1,3)-glucan. Neutrophils could kill the uncloaked gpi7 mutant more efficiently with an augmented respiratory burst. The gpi7 mutant also stimulated Dectin-1-dependent immune responses of macrophages, including activation of nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) pathways and secretion of specific cytokines, such as tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), and IL-12p40. Furthermore, the gpi7 null mutant could induce an enhanced inflammatory response through promoting significant recruitment of neutrophils and monocytes and could stimulate stronger Th1 and Th17 cell responses to fungal infections in vivo. These in vivo phenotypes also were Dectin-1 dependent. Thus, we assume that GPI-CWPs are involved in the immune mechanism of C. albicans escaping from host recognition by Dectin-1. Our studies also indicate that the blockage of GPI anchor synthesis is a strategy to inhibit C. albicans evading host recognition.

CRH knockout inhibits the murine innate immune responses in association with endoplasmic reticulum stress after thermal injury

Previous studies in our laboratory have demonstrated the hypothalamus destruction and adrenalectomy could blunt the innate immunity while boosting the excessive inflammation after injury. We aimed to investigate the effects of corticotrophin-releasing hormone knockout (CRH KO) on the innate immune responses in macrophages as well as to elucidate the underlying mechanism. The chemotaxis and phagocytosis activities of macrophages, bacteria translocation, plasma tumor necrosis factor (TNF)-α secretion, and intestinal injury were observed in the presence of the endoplasmic reticulum stress after thermal injury in CRH KO mice. Meanwhile, the messenger RNA (mRNA) and protein expression of glucose response protein 78 (GRP78), X-box binding protein 1 (XBP1), and activating transcription factor 6 (ATF6) in macrophages was also determined. After thermal injury, the chemotaxis and phagocytosis of peritoneal macrophages were increased, which were both reversed by CRH gene deficiency. The gut-derived bacteria translocation to liver tissues, lung tissues and mesenteric lymph nodes was significantly strengthened in CRH KO mice compared with CRH wild-type littermates. Circulating TNF-α level was increased markedly in response to thermal injury and CRH KO further increased its secretion. Furthermore, the mRNA and protein levels of GRP78, XBP1, and ATF6 in peritoneal macrophages increased, while their expressions in CRH KO mice all decreased significantly. CRH KO mice showed enhancement of inflammatory responses and severe tissue injuries after thermal injury. CRH exerted immune defensive actions on immune cells and organs in the early phase of injury, suggesting that the underlying mechanisms are related to endoplasmic reticulum stress.

Mo1828 Reduced Intestinal Leukocyte Exit Heightens the Severity of DSS Colitis in a Murine Model of Lymphovascular Deficiency

process of intestinal inflammation is less understood. While we could demonstrate that HDAC inhibitors induce amelioration of colitis in mice, we are further interested in how far specific HDAC are responsible for inflammatory responses and what are the mechanism behind these effects. Methods: The macrophage cell line RAW 264.7 and the monocyte cell line U937 were analysed for HDAC5-expression using quantitative PCR and Western-blot. The cells were transfected with a HDAC5-encoding plasmid for overexpression or small interfering (si)RNA for knock-down studies. HDAC5 knock out mice were used in dextran sulphate sodium (DSS) induced colitis model. Colon tissue was analysed by histology and lamina propria mononuclear cells (LPMCs) were isolated. All cells were analysed by flow cytometry. Supernatant of colon and cell cultures were analysed by cytometric bead assay. Results: In RAW 264.7 and U937 expression of HDAC5 mRNA down regulated for up to 7 h but presented baseline levels after 24 h. HDAC5-overexpression in RAW 264.7 resulted in an increased LPS-dependent secretion of the pro-inflammatory cytokines tumor necrosis factor alpha, the monocyte chemoattractant protein-1 and interleukin 6. Subsequently, knock-down of HDAC5-mRNA expression by specific siRNA significantly reduced the secretion of these cytokines. These effects were accompanied by increased nuclear factor kappa B activity. Further experimental colitis experiments demonstrate a differential disease pattern for HDAC5 knockout mice compared to wild type mice. This was shown by an increased histological disease score, higher weight loss and severe intestinal bleeding. LPMC isolation followed by extensive FACS analysis of the LPMC confirmed these results by demonstrating changes in macrophage polarization. When the colonic barrier function was analysed in DSS induced colitis experiments, we detected a decreased trans-epithelial resistance and modulated flux of mannitol, HRP and FITC-Dextran. Conclusion: Our data demonstrate a critical function of HDAC5 in the pro-inflammatory immune response of macrophages in the lamina propria as well as a role within the physiological barrier of the gut epithelium, proposing a potential new target for the treatment of inflammatory bowel disease

Targeted Disruption of Ig-Hepta/Gpr116 Causes Emphysema-like Symptoms That Are Associated with Alveolar Macrophage Activation

Ig-Hepta/GPR116 is a member of the G protein-coupled receptor family predominantly expressed in the alveolar type II epithelial cells of the lung. Previous studies have shown that Ig-Hepta is essential for lung surfactant homeostasis, and loss of its function results in high accumulation of surfactant lipids and proteins in the alveolar space. Ig-Hepta knock-out (Ig-Hepta(-/-)) mice also exhibit emphysema-like symptoms, including accumulation of foamy alveolar macrophages (AMs), but its pathogenic mechanism is unknown. Here, we show that the bronchoalveolar lavage fluid obtained from Ig-Hepta(-/-) mice contains high levels of inflammatory mediators, lipid hydroperoxides, and matrix metalloproteinases (MMPs), which are produced by AMs. Accumulation of reactive oxygen species was observed in the AMs of Ig-Hepta(-/-) mice in an age-dependent manner. In addition, nuclear factor-κB (NF-κB) is activated and translocated into the nuclei of the AMs of Ig-Hepta(-/-) mice. Release of MMP-2 and MMP-9 from the AMs was strongly inhibited by treatment with inhibitors of oxidants and NF-κB. We also found that the level of monocyte chemotactic protein-1 is increased in the embryonic lungs of Ig-Hepta(-/-) mice at 18.5 days postcoitum, when AMs are not accumulated and activated. These results suggest that Ig-Hepta plays an important role in regulating macrophage immune responses, and its deficiency leads to local inflammation in the lung, where AMs produce excessive amounts of reactive oxygen species and up-regulate MMPs through the NF-κB signaling pathway.

Morphological and proteomic analyses reveal that unsaturated guluronate oligosaccharide modulates multiple functional pathways in murine macrophage RAW264.7 cells.

Alginate is a natural polysaccharide extracted from various species of marine brown algae. Alginate-derived guluronate oligosaccharide (GOS) obtained by enzymatic depolymerization has various pharmacological functions. Previous studies have demonstrated that GOS can trigger the production of inducible nitric oxide synthase (iNOS)/nitric oxide (NO), reactive oxygen species (ROS) and tumor necrosis factor (TNF)-α by macrophages and that it is involved in the nuclear factor (NF)-κB and mitogen-activated protein (MAP) kinase signaling pathways. To expand upon the current knowledge regarding the molecular mechanisms associated with the GOS-induced immune response in macrophages, comparative proteomic analysis was employed together with two-dimensional electrophoresis (2-DE), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/TOF MS) and Western blot verification. Proteins showing significant differences in expression in GOS-treated cells were categorized into multiple functional pathways, including the NF-κB signaling pathway and pathways involved in inflammation, antioxidant activity, glycolysis, cytoskeletal processes and translational elongation. Moreover, GOS-stimulated changes in the morphologies and actin cytoskeleton organization of RAW264.7 cells were also investigated as possible adaptations to GOS. This study is the first to reveal GOS as a promising agent that can modulate the proper balance between the pro- and anti-inflammatory immune responses, and it provides new insights into pharmaceutical applications of polysaccharides.

Effect of poly(vinyl alcohol) (PVA) wear particles generated in water lubricant on immune response of macrophage

Abstract To develop a novel biotribological material for artificial cartilage implant with a lubricity supplement of the joint surface, we focused on two types of poly(vinyl alcohol) (PVA) hydrogel: repeated freeze–thawing (PVA-FT) and cast-drying (PVA-CD) gels. Here we observed the morphology of wear particles generated during a reciprocating wear test and assessed macrophage immune responses by applying hydrogel wear particles. As a result, PVA-CD had a significantly lower total amount of wear than did PVA-FT. The size distributions of PVA-FT and -CD wear particles were similar. Most of the particles were nanoparticles up to approximately 50 nm in diameter. Considering the particle volume distribution, there were very few micron- and submicron-sized wear particles around 1 μm in diameter. In SEM observations of dried PVA wear particles, both distributions of wear particles of PVA-FT and -CD were similar. Micron-sized wear particles were chiefly formed by close packing of 20- to 50-nm-sized particles. Biochemical and immunological evaluations revealed no cytotoxic effects of wear particles on macrophages. Cytokine synthesis of both wear particle-stimulated groups was significantly lower than that of the lipopolysaccharide-stimulated positive control. Therefore, it is suggested that PVA wear particles do not affect the macrophage immune response.

Increased PD-L1 expression and PD-L1/CD86 ratio on macrophages infected with Neisseria gonorrhoeae contributes to the functional Suppression of T-cell responses.

Event Abstract Back to Event Increased PD-L1 expression and PD-L1/CD86 ratio on macrophages infected with Neisseria gonorrhoeae contributes to the functional Suppression of T-cell responses. Alejandro Escobar1*, Carolina Ortiz1, Killen Garcia1, Paula Rodas2 and Claudio Acuña-Castillo3 1 Universidad de Chile, Instituto de Investigación en Ciencias Odontológicas, Chile 2 Universidad Andres Bello, Center for Integrative Medicine and Innovative Science, Chile 3 Universidad de Santiago, Chile Neisseria gonorrhoeae is the etiological agent of the sexually transmitted disease gonorrhoea, which is widespread throughout the world. Infections with Neisseria gonorrhoeae do not improve immune response in patients with re-infection, suggesting that gonococcus displays several mechanisms to evade immune response and survive in the host. Our data indicate that Neisseria gonorrhoeae is able to suppress the protective immune response in macrophages, polarized them towards an immunoregulatory profile characterized by up-regulation of M2-Macrophage-associated markers and IL-10. Considering there are numerous reports indicating the capacity of Intracellular pathogens like Mycobacteria to impair innate and adaptative immune response using inhibitory role played by Programmed Death 1 and Programmed Death Ligand 1 (PD-1/PD-L1) interactions, in this study, we evaluated surface co-stimulatory (CD86, CD40) and co-inhibitory (PD-L1) markers expression and allostimulatory capacity in macrophages infected with Neisseria gonorrhoeae. Results showed that Neisseria gonorrhoeae was not able to induce the co-stimulatory CD86 neither CD40. However, we found a significant up-regulation of PD-L1 in macrophages upon gonococcal infection. PD-L1/CD86 ratio was increased and positively correlated with PD-L1 expression on macrophages. Allostimulatory capacity of macrophages was impaired and inversely correlated with PD-L1 expression and PD-L1/CD86 ratio. Finally, to verify that the cell contact–dependent inhibition of proliferation was due to macrophage´s PD-L1, we set up a co-culture in the presence of blocking antibody to PD-L1 and to IL-10. We showed that only PD-L1 blocking restored T-cell proliferation. These findings suggested that the effect of inhibitory marker PD-L1 overwhelmed the effect of co-stimulatory markers and down-regulated macrophage-T activation in presence of Neisseria gonorrhoeae infection. This study has important implications for understanding the mechanisms of clearance versus long-term persistence of N. gonorroheae infection and might be applicable for the development of new therapeutic strategies. Acknowledgements Dirección Investigación Facultad Odontología Universidad de Chile. Keywords: Neisseria gonorrhoeae, Macrophages, PD-L1, alloprolifertion, Immune Evasion Conference: IMMUNOCOLOMBIA2015 - 11th Congress of the Latin American Association of Immunology - 10o. Congreso de la Asociación Colombiana de Alergia, Asma e Inmunología, Medellin, Colombia, 13 Oct - 16 Oct, 2015. Presentation Type: Oral Presentation Topic: Infectious and parasitic diseases Citation: Escobar A, Ortiz C, Garcia K, Rodas P and Acuña-Castillo C (2015). Increased PD-L1 expression and PD-L1/CD86 ratio on macrophages infected with Neisseria gonorrhoeae contributes to the functional Suppression of T-cell responses.. Front. Immunol. Conference Abstract: IMMUNOCOLOMBIA2015 - 11th Congress of the Latin American Association of Immunology - 10o. Congreso de la Asociación Colombiana de Alergia, Asma e Inmunología. doi: 10.3389/conf.fimmu.2015.05.00303 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 15 Apr 2015; Published Online: 15 Sep 2015. * Correspondence: Dr. Alejandro Escobar, Universidad de Chile, Instituto de Investigación en Ciencias Odontológicas, Santiago, Chile, janodvm@gmail.com Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Alejandro Escobar Carolina Ortiz Killen Garcia Paula Rodas Claudio Acuña-Castillo Google Alejandro Escobar Carolina Ortiz Killen Garcia Paula Rodas Claudio Acuña-Castillo Google Scholar Alejandro Escobar Carolina Ortiz Killen Garcia Paula Rodas Claudio Acuña-Castillo PubMed Alejandro Escobar Carolina Ortiz Killen Garcia Paula Rodas Claudio Acuña-Castillo Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Modulations of Macrophage Immune Responses by Mycobacterial PE/PPE Family of Proteins

Modulations of Macrophage Immune Responses by Mycobacterial PE/PPE Family of Proteins

Mycobacterium tuberculosis 19-kDa lipoprotein induces Toll-like receptor 2-dependent peroxisome proliferator-activated receptor γ expression and promotes inflammatory responses in human macrophages.

Mycobacterium tuberculosis (M.tb) enhances its survival in macrophages by suppressing immune responses, in part through its complex cell wall structures. M.tb 19‑kDa lipoprotein (P19), a component of the complex cell wall structures of M.tb, is a Toll‑like receptor (TLR) agonist, and may induce immune responses through TLR2. Furthermore, the activation of peroxisome proliferator‑activated receptor γ (PPARγ) is also involved in M.tb‑induced immune responses in macrophages. In the present study, specific agonists/antagonists and siRNA were used to investigate the role of PPARγ in P19‑induced immune responses in human macrophages, including TLR2 activation, p38 phosphorylation and cytokine production. In the present study, PPARγ expression, p38 phosphorylation and cytokine production were upregulated following M.tb H37Rv infection or P19 treatment. By pretreating macrophages with a specific PPARγ agonist or antagonist, it was demonstrated that phosphorylation and IL‑6 production are modulated in macrophages by PPARγ activity. Following TLR2 knockdown in macrophages, the expression of PPARγ was significantly decreased in the presence or absence of P19 treatment. Furthermore, p38 phosphorylation and cytokine production were significantly reduced in TLR2 knockdown macrophages following P19 treatment. It was demonstrated in the current study that PPARγ was induced and activated by M.tb infection and that P19‑induced PPARγ expression, p38 phosphorylation and cytokine production in macrophages are dependent on TLR2. These findings suggest a role for PPARγ and TLR2 in P19‑induced p38 phosphorylation and cytokine production, thereby potentially influencing M.tb pathogenesis.

Immunomodulatory Role of Mycobacterial PE/PPE Family of Proteins

Mycobacterium tuberculosis (M.tb) is an adept pathogen possessing unique strategies to exploit and corrupt the very core defense systems designed to curb its own survival and multiplication. Comparative analysis of mycobacterial genomes indicate that the pathogenic mycobacterium have acquired a multitude of genes which play important roles in modulating protective host-cell signaling that are triggered in response to an infection. Therefore, defining the “set of genes” expressed by intracellular pathogens as well as understanding modulations of host-signaling pathways during infection is extremely crucial to devise successful clinical intervention against the development of disease. The unique PE/PPE proteins, which constitute about 10% of coding capacity of M.tb genome, owing to their high abundance and expansion in pathogenic mycobacterial species warrants a better understanding of their role in mycobacterial pathogenesis. The PE/PPE families have been thought to play important roles in generating antigenic variation and immune evasion. Although a comprehensive understanding of their role is yet to be fully understood, emer ging evidence supports a role of PE/PPE proteins at multiple levels of infectious process. This review delineates the importance of these proteins in the modulation of macrophage immune effector responses, as well as importance of these proteins in clinical manifestations of TB.

The Tissue Factor/Thrombin/Protease-Activated Receptor 1 Pathway Enhances Double-Strand RNA Induced Immune Responses in Macrophages

Introduction: Co-regulation of the immune response and the coagulation cascade after infection is thought to be an ancient response to limit pathogen spread. Recently, we showed that activation of the thrombin receptor, protease-activated receptor 1 (PAR1), on fibroblasts enhanced the innate immune responses to RNA virus infection. Here, we investigated whether PAR1 activation by the extrinsic coagulation pathway contributes to dsRNA-induced innate immune responses in macrophages.Methods: Activation of the type-I interferon (IFN) pathway in the murine macrophage cell line RAW264.7 and bone-marrow derived macrophages (BMDM) from WT and PAR1-/- was analyzed after dsRNA (poly I:C) and/or PAR-1 stimulation. In addition, innate immune responses in the spleen were analyzed in vivo 4 hours after poly I:C (8mg/kg) injection in mice with reduced tissue factor expression (LowTF) or global PAR1 deletion (PAR1-/-) as well as in WT mice with a thrombin inhibitor (dabigatran etexilate, 10g/kg chow) or PAR-1 inhibitor (SCH79797, 25μg/kg). Lastly, we investigated the innate immune response in the spleen of WT and PAR1-/-mice after infection with the single-stranded RNA virus coxsackievirus B3.Results: RAW264.7 and BMDM exhibited a toll-like receptor 3 dependent induction of IFNβ and CXCL10 after poly I:C stimulation. Activation of PAR-1 with either thrombin or agonist peptide enhanced poly I:C induction of IFNβ and CXCL10. A deficiency of tissue factor levels, thrombin inhibition, PAR-1 inhibition or PAR1 deficiency resulted in reduced expression levels of type-I IFNs and IFN-response genes such as CXCL10 in the spleen and plasma in mice given poly I:C. Last, PAR1-/-mice exhibited impaired IFNβ immune response 4 days after coxsackievirus B3 infection compared to WT mice.Conclusion: Our study indicates that the coagulation dependent activation of PAR1 on macrophages is important for anti-viral responses to dsRNA. We speculate that PAR1 inhibition may interfere with anti-viral responses in humans. DisclosuresNo relevant conflicts of interest to declare.