Dendritic Macrophages Research Articles

Abstract 4881: CD300f-regulated efferocytosis by dendritic cells and macrophages controls the initiation and resolution of experimental colitis

Abstract Phagocytic clearance of apoptotic cells (efferocytosis) is critical for resolution of inflammation and prevention of chronic inflammatory disorders, including inflammatory bowel diseases, and inflammation-associated cancer. CD300f (CLM-1), a cell surface receptor that recognizes phosphatidylserine exposed on apoptotic cells, has been shown to promote efferocytosis by macrophages. In the present study, we found that CD300f plays a critical role in controlling the severity and duration of dextran sulfate sodium (DSS)-induced colitis, and CD300f-expressing macrophages and dendritic cells (DC) were identified as key players controlling disease pathogenesis. Compared to Cd300f+/+ mice, Cd300f-/- mice showed increased susceptibility to DSS-induced colitis and impaired resolution of inflammation. In Cd300f-/- mice, macrophage-mediated efferocytosis was reduced, resulting in apoptotic cell accumulation in the gut mucosa. In contrast, Cd300f-/- DC engulfed apoptotic cells more efficiently than Cd300f+/+ DC. CD300f-deficient DC transfer exacerbated DSS-induced inflammation and delayed its resolution, whereas CD300f-expressing macrophage transfer attenuated DSS-induced inflammation. Hyperactive efferocytosis by CD300f-deficient gut DC resulted in excessive tumor necrosis factor-á (TNF-á) secretion, which induced secondary interferon-ã (IFN-ã) over-production, both of which impaired the resolution of colitis. TNF-á neutralization resulted in decreased levels of gut pro-inflammatory cytokines and significant disease amelioration. Collectively, these results suggest that CD300f is important for preventing chronic inflammation by enhancing macrophage efferocytosis and inhibiting TNF-á production induced by DC efferocytosis. From these findings, it is conceivable that increasing macrophage-mediated efferocytosis and suppressing DC-mediated efferocytosis through upregulation of CD300f expression would provide a new therapeutic strategy for inflammatory bowel disease patients. Citation Format: Ha-Na Lee, Linjie Tian, Jacquice Davis, Mariam Quinones, Yasmine Belkaid, Konrad Krzewski, John E. Coligan. CD300f-regulated efferocytosis by dendritic cells and macrophages controls the initiation and resolution of experimental colitis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4881.

Abstract 4134: Immune infiltration and PD-L1 expression in the tumor microenvironment are prognostic in osteosarcoma

Abstract Purpose: Over the past four decades, osteosarcoma (OS) survival rates have remained stagnant. There is a need to identify novel therapies to target OS. In this study we examined the expression of Programed Death Ligand 1 (PD-L1) and defined the tumor microenvironment in OS in order to assess the feasibility of utilizing immune checkpoint inhibitors as a treatment modality. Experimental Design: PD-L1 expression in OS was examined in two patient cohorts using immunohistochemistry (IHC) (n = 48, n = 59) and expression was validated using quantitative real time PCR (qRT-PCR) (n = 21) and western blotting (n = 9). IHC was used to determine presence of tumor infiltrating lymphocytes and antigen presenting cells (APCs) in the tumor. Expression of PD-L1 was correlated with the presence of immune cells and with event free survival in OS. Results: PD-L1 is expressed in up to 25% of primary OS tumors. Of the PD-L1 positive tumors the majority were also PD-1 positive (92% vs 8%, p = 0.002). In addition, the presence of immune cells in the tumor mass was significantly associated with PD-L1 expression. Although all immune cell types examined were present in OS, only infiltration by APCs, specifically CD1a positive dendritic cells (48.6% vs. 51.4%, p = 0.0010) and CD68 positive macrophages (70.3% vs. 29.7%, p = 0.0316), was associated with worse outcomes. PD-L1 expression was significantly associated with worsened survival (21.6% pos. vs. 78.4% neg., p = 0.0146). Conclusions: For the first time we have identified PD-L1 expression or presence of CD1a or CD68 positive antigen presenting cells as prognostic markers for worsened outcome in OS. Furthermore, we show that IHC is a valid detection method for PD-L1 in OS. With up to 25% of OS patients expressing PD-L1, this study provides rational for targeting the PD-L1:PD-1 axis for immunotherapy in OS. Citation Format: Pratistha Koirala, Michael Roth, Jonathan Gill, Jordan Chinai, Sajida Piperdi, David Geller, Bang Hoang, Vincent Poon, Xingxing Zang, Richard Gorlick. Immune infiltration and PD-L1 expression in the tumor microenvironment are prognostic in osteosarcoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4134.

The Phenotypic Characterization of the Human Renal Mononuclear Phagocytes Reveal a Co-Ordinated Response to Injury.

Mammalian tissues contain networks of mononuclear phagocytes (MPh) that sense injury and orchestrate the response to it. In mice, this is affected by distinct populations of dendritic cells (DC), monocytes and macrophages and recent studies suggest the same is true for human skin and intestine but little is known about the kidney. Here we describe the analysis of MPh populations in five human kidneys and show they are highly heterogeneous and contain discrete populations of DC, monocytes and macrophages. These include: plasmacytoid DC (CD303+) and both types of conventional DC—cDC1 (CD141+ cells) and CD2 (CD1c+ cells); classical, non-classical and intermediate monocytes; and macrophages including a novel population of CD141+ macrophages clearly distinguishable from cDC1 cells. The relative size of the MPh populations differed between kidneys: the pDC population was bi-modally distributed being less than 2% of DC in two kidneys without severe injury and over 35% in the remaining three with low grade injury in the absence of morphological evidence of inflammation. There were profound differences in the other MPh populations in kidneys with high and low numbers of pDC. Thus, cDC1 cells were abundant (55 and 52.3%) when pDC were sparse and sparse (12.8–12.5%) when pDC were abundant, whereas the proportions of cDC2 cells and classical monocytes increased slightly in pDC high kidneys. We conclude that MPh are highly heterogeneous in human kidneys and that pDC infiltration indicative of low-grade injury does not occur in isolation but is part of a co-ordinated response affecting all renal DC, monocyte and macrophage populations.

Abstract A16: The crosstalk between Indoleamine 2,3 Dioxygenase (IDO) and cancer stem cells in murine melanoma: A potential modality of local control

Abstract Introduction: Indoleamine 2,3-dioxygenase (IDO), the rate-limiting enzyme of tryptophan catabolism, has been strongly associated with the progression of malignancy and poor survival in melanoma patients. As a result, IDO is a leading target for interventions aimed at restoring melanoma immune surveillance. IDO, which is produced by cells including dendritic cells, macrophages, stem cells, and most tumor cells, potently suppresses immune responses by depleting local tryptophan and allowing the accumulation of tryptophan metabolites. However, the actual cellular and molecular mechanisms underlying the immunoregulatory function of IDO remain largely unknown. The relatively newly described cancer stem cells (CSCs) are tumor-initiating cells (TICs) that often survive therapy and give rise to second-line tumors by self-renewal and differentiation into multiple cell types. CSCs are proposed to persist in tumors as a distinct population and to cause relapse and metastasis by giving rise to new tumors. Therefore, development of specific therapies targeted at CSCs holds hope for improvement of survival and quality of life of cancer patients, especially those with metastatic disease. Methods: C57/BL6 mice were injected with B16F10 murine melanoma cell line (1 x 106 cells/mouse, subcutaneous). Tumor growth monitored. At 3mm, tumors were injected with an IDO Inhibitor (1 MT) (Group 1) twice weekly (20 mg/mouse per injection). Controls (Group 2) received vehicle only (PBS), Group 3 weekly Cisplatin (5mg/kg), and Group 4 both Cisplatin and 1MT (IDO inhibitor). Once the first tumor reached 12mm in size, the mice were sacrificed and tumor cells subjected to flow-cytometry, and statistical analysis, and p-values <0.05 considered significant. Results: Our data showed for the first time that IDO was expressed by CSCs in a murine melanoma model. These IDO-expressing CSCs may be capable of suppressing T lymphocyte infiltration into the tumor microenvironment, which was reversed by the IDO inhibitor, 1-methyl-tryptophan. Conclusion: Our findings offer an important new line of evidence that interventional targeting of IDO activity could be used to restore tumor immunity by relieving IDO-mediated immune suppression of CSCs in the tumor microenvironment as well as in tumor cells themselves. Citation Format: Marsha Kocherla, Blake Christianson, James Walker, Xu Qin, Jack C. Yu, Babak Baban. The crosstalk between Indoleamine 2,3 Dioxygenase (IDO) and cancer stem cells in murine melanoma: A potential modality of local control. [abstract]. In: Proceedings of the Fourth AACR International Conference on Frontiers in Basic Cancer Research; 2015 Oct 23-26; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2016;76(3 Suppl):Abstract nr A16.

The PCome of Ascaris suum as a model system for intestinal nematodes: identification of phosphorylcholine-substituted proteins and first characterization of the PC-epitope structures.

In multicellular parasites (e.g., nematodes and protozoa), proteins and glycolipids have been found to be decorated with phosphorylcholine (PC). PC can provoke various effects on immune cells leading to an immunomodulation of the host's immune system. This immunomodulation allows long-term persistence but also prevents severe pathology due to downregulation of cellular immune responses. PC-containing antigens have been found to interfere with key proliferative signaling pathways in B and T cells, development of dendritic cells and macrophages, and mast cell degranulation. These effects contribute to the observed modulated cytokine levels and impairment of lymphocyte proliferation. In contrast to glycosphingolipids, little is known about the PC-epitopes of proteins. So far, only a limited number of PC-modified proteins from nematodes have been identified. In this project, PC-substituted proteins and glycolipids in Ascaris suum have been localized by immunohistochemistry in specific tissues of the body wall, intestine, and reproductive tract. Subsequently, we investigated the PCome of A. suum by 2D gel-based proteomics and detection by Western blotting using the PC-specific antibody TEPC-15. By peptide-mass-fingerprint matrix-assisted laser-desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS), we could identify 59 PC-substituted proteins, which are in involved multiple cellular processes. In addition to membrane proteins like vitellogenin-6, we found proteins with structural (e.g., tubulins) and metabolic (e.g., pyruvate dehydrogenase) functions or which can act in the defense against the host's immune response (e.g., serpins). Initial characterization of the PC-epitopes revealed a predominant linkage of PC to the proteins via N-glycans. Our data form the basis for more detailed investigations of the PC-epitope structures as a prerequisite for comprehensive understanding of the molecular mechanisms of immunomodulation.

Immunosuppressive arenaviral exoribonuclease

Arenaviruses cause up to 500,000 zoonotic infections per year in endemic areas of Africa and South America and can lead to severe and lethal hemorrhagic fever (HF) symptoms [1]. Currently, there is no specific antiviral drug or vaccine available for the treatment of these infections, with the exception of the Candid #1 vaccine that has successfully been used to prevent infections of Junin virus in the endemic areas of Argentina. Progression and severity of arenaviral HF disease generally correlates with viremia and host immune suppression, the molecular mechanisms of which have only recently been revealed [2]. Mounting an early and effective induction of the host innate-immune response to the infection, which is followed by robust cell-mediated responses, is thought to be essential to control the outcome of the infection. Many arenaviruses, especially those belonging to the Old World virus group, have developed different strategies to effectively suppress the host immune responses to infections. Our laboratory has been involved in characterizing the molecular mechanisms behind innate immune suppression mediated by arenaviral nucleoprotein (NP). Our high-resolution structure of the Lassa virus (LASV) NP reveals a conserved DEDDh exonuclease in its C terminal domain [3], which can degrade double-stranded RNA that would normally trigger the type I interferons (IFN) signaling pathway upon viral infections. NP has also been shown to interfere with the proper functions of several factors in the RIG-I-like receptor (RLR) pathway, namely IKKe to prevent IRF3 activation, and the Nuclear Factor Kappa B (NF-κB) pathway, and therefore suppressing the production of IFN-β and potentially other cytokines [4]. Initial studies using NPs expressed from plasmid DNAs have attributed the non-pathogenic nature of the Tacaribe virus (TCRV) to a change of amino acids around the DEDDh exonuclease site and the observed lack of IRF3 inhibition. Our efforts to resequence TCRV and to perform a comprehensive structure-function analysis of the TCRV NP have revealed discrepancies with the GenBank deposited sequence and shown that the IFN-antagonism displayed by the NP exoribonuclease (RNase) activity is a conserved mechanism among different arenaviruses, including but not necessarily limited to TCRV, LASV and Pichinde virus (PICV) [5]. Using recombinant viruses, we have recently shown the importance of the IFN-suppressive activity of NP and its role in mediating optimal viral replication in cell culture as well as in a surrogate animal model of LASV infection that is based on PICV-infected guinea pigs [5]. Specifically, we show that mutating each of the NP's catalytic DEDDh residues to alanines results in significantly reduced levels of IFN-β inhibition and virus grow potentials in immune-competent cells, and in an attenuated phenotype of the mutant viruses in infected animals. The important role of the NP RNase function in suppressing immune response in order to favor virus replication is evidenced by the appearance of wild-type virus revertants in many of the sick animals with detectable levels of viremia [5]. Our findings are supported by recent studies with recombinant LASV carrying the NP catalytic double mutations (D389T/G392A and D389A/G392A), which lose the ability to suppress the type I IFN-induction pathway in dendritic cells or macrophages, which are thought to be primary target cells for virus infection [6, 7]. The inability of the LASV NP catalytic mutants to suppress type I IFNs results in upregulated expression levels of major histocompatibility complex (MHC) class I gene pathways as well as important macrophage-derived cytokines that activate natural killer (NK) cells to participate in the direct killing of the infected cells. Thus, the proper function of the NP RNase appears to be essential in suppressing the early innate immune response, which directly correlates with the priming of cellular immunity and therefore impacts virus clearance in vivo. Taken together, studies conducted in our laboratory as well as by other laboratories have provided strong evidence to support the role of the arenaviral NP exoribonuclease function in mediating immune suppression upon viral infections. It is important to highlight the fact that this is the first time that a viral exoribonuclease function has been attributed to mediating host immune evasion. Because there is currently no specific antiviral therapy available against arenaviruses, a better level of understanding of how arenaviruses are able to evade host immunity is warranted as it may offer new ways to evaluate future treatment options and to design and test vaccine candidates against these deadly human viral pathogens.

A gain-of-function CCR4 mutations in adult T-cell leukemia/lymphoma (ATL) enhance the chemotactic abilities and P13K/AKT activation

High expression of the CC chemokine receptor 4 (CCR4) has been identified as a hallmark gene in ATL-an aggressive peripheral T-cell neoplasm. CCR4 is a chemokine receptor, which has a critical role in immune cell trafficking including Th2, T-regs and skin homing memory T-cells. CCR4 ligands, CCL17 and CCL22 are produced in lymph nodes and skin from dendritic cells, macrophages and Langerhans cells. Most ATL cases express surface CCR4 (90%) and infiltrate lymph nodes and skin. We performed RNA-Seq on two primary ATL cases and discovered recurrent nonsense mutations in CCR4. Through an extended analysis using Sanger sequencing, CCR4 mutations were detected in 14/53 ATL samples (26%) and consisted exclusively of nonsense or frameshift mutations that truncated the coding region at C329, Q330 or Y331 in the carboxyl-terminus. We hypothesized that the mutant CCR4 isoforms might enhance chemotaxis of the affected cells to CCR4 ligands. A chemotaxis assay using 32D beta, a mouse myeloid cell line and ED40515 (+), an ATL cell line, clarified that the ectopic expression of CCR4-Q330* but not CCR4-WT transduced cells enhanced the chemotactic ability of the transduced cells toward CCL17 and CCL22. To define the mechanism of the enhanced chemotactic ability, we examined the change in surface CCR4 levels after CCL22 exposure in CCR4-WT-and CCR4-Q330*-reconstituted ED40515 (+) cells. Compared with CCR4-WT, CCR4 internalization in CCR4-Q330*-reconstituted cells was significantly impaired. Thus, the ATL CCR4 mutants manifested impaired desensitization by ligand, which likely contributed to the enhanced chemotaxis of cells bearing these mutants. We explored the influence of the ATL CCR4 mutants on PI(3) kinase (PI3K)-dependent activation of AKT since it has been reported that binding of CCL22 to CCR4 activates AKT in CEM leukemic T-cells and in human Th2 cells. CCR4-Q330* reconstituted ED40414 (+) cells showed strong activation of AKT with CCL22 ligation compared with CCR4-WT reconstituted cells. The AKT activation was inhibited by the pan-P13K inhibitor, BKM120, indicating that the CCR4-mediated AKT activation was PI3K dependent. Lastly, we tested whether the acquisition of CCR4 mutations by ATL cells imparts a selective growth advantage relative to cells with wild-type CCR4. CCR4-Q330*-reconstituted cells had a selective growth advantage in the presence of CCL22, supporting, at least in part, the hypothesis that CCR4 mutations provided the affected cells with a positive selection pressure through CCL22 ligation that contributed to the ATL pathogenesis. Our findings provide a rationale for a clinical trial to test whether inhibition of CCR4 signaling might have a therapeutic potential for patients with ATL.

History and perspectives of the monocyte-macrophage system

The macrophage cell system was identified by Metchnikoff on the basis of its phagocytic ability. Later on, the reticulohistiocytic system was defined as being composed of antigen-presenting reticulum cells and macrophages. Van Furth proposed that the mononuclear phagocyte system includes all tissue macrophages as well as antigen-presenting cells and blood monocytes as their precursors. Recent findings have shown that blood monocytes are not just transient forms involved in the recruitment of macrophages but that different dendritic and monocytic subpopulations can be observed in blood. In tissue, self-renewing macrophages derived from the yolk sac as well as monocyte-derived dendritic cells and monocyte-derived macrophages can be distinguished. Due to their plasticity and polarization, under inflammatory conditions monocyte-derived macrophages may be beneficial for the reestablishment of homeostasis or may contribute to mostly chronic diseases. Because of their ubiquitous distribution, monocytes and macrophages are increasingly considered to be possible therapeutic targets.

Abstract 3936: Frequent gain-of-function CCR4 mutations in adult T-cell leukemia/lymphoma (ATLL)

Abstract High expression of CC chemokine receptor 4 (CCR4) has been identified as a hallmark gene in ATLL, an aggressive peripheral T-cell neoplasm. CCR4 is a chemokine receptor, which has a critical role in immune cell trafficking. CCR4 ligands, CCL17 and CCL22, were produced in lymph nodes and skin from dendritic cells, macrophages and Langerhans cells. Most ATLL cases express surface CCR4 (90%) and infiltrate to lymph nodes and skin. These observations suggest that CCR4 could have a role in ATLL biology, but it is still unclear whether dysregulation of CCR4 function contributes to ATLL pathogenesis. We performed RNA-Seq for two primary ATLL cases and discovered recurrent non-sense mutations in CCR4. Though an extended analysis using Sanger sequencing, CCR4 mutations were detected in 14/53 ATLL samples (26%) and consisted exclusively of nonsense or frameshift mutations that truncated the coding region at C329, Q330 or Y331 in the carboxy-terminus. In 5 cases for which paired normal DNA was available, three different CCR4 mutations were detected only in the ATLL cells (Q330*; Q330 frameshift; Y331*), demonstrating that they were acquired somatically during malignant transformation or progression. Chemotaxic assay using 32Dβ, a mouse myeloid cell line, and ED40515(+), an ATLL cell line, clarified that the ectopic expression of CCR4-Q330* enhanced the chemotactic ability of the transduced cells toward CCL17 and CCL22 rather than CCR4-WT transduced cells. To understand the mechanism of this enhanced chemotactic ability, we studied the change in surface CCR4 levels after CCL22 exposure in CCR4-WT-and CCR4-Q330*-reconstituted ED40515(+) cells. Compared with CCR4-WT, CCR4 internalization in CCR4-Q330*-reconstituted cells was significantly impaired. Thus, the ATLL CCR4 mutants impair desensitization by ligand, which likely contributes to the enhanced chemotaxis of cells bearing these mutants. We explored the influence of the ATLL CCR4 mutants on PI3K/AKT signaling by immunoblot analysis and phosphor-flow analysis. CCR4-Q330*-reconstituted ED40515(+) showed strong activation of AKT with CCL22 ligation compared with CCR4-WT-reconstituted cell. Furthermore, we tested whether the acquisition of CCR4 mutations by ATLL cells imparts a selective growth advantage relative to cells with wild type CCR4. CCR4-Q330*-reconstituted cells had a selective growth advantage in the presence of CCL22, supporting at least in part the hypothesis that CCR4 mutation are able to provide the affected cells a positive selection pressure through CCL22 ligation and contributes to ATLL pathogenesis. Our findings implicate somatic gain-of-function CCR4 mutations in the pathogenesis of ATLL and suggest that inhibition of CCR4 signaling might have therapeutic potential in this refractory malignancy. Citation Format: Masao Nakagawa, Roland Schmitz, Wenming Xiao, Carolyn K. Goldman, Weihong Xu, Yandan Yang, Xin Yu, Thomas A. Waldmann, Louis M. Staudt. Frequent gain-of-function CCR4 mutations in adult T-cell leukemia/lymphoma (ATLL). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3936. doi:10.1158/1538-7445.AM2015-3936

Abstract 4289: Targeting of phosphatidylserine by monoclonal antibodies enhances the activity of immune checkpoint inhibitors in breast tumors

Abstract Phosphatidylserine (PS) is a phospholipid normally residing in the inner leaflet of the plasma membrane that becomes exposed on tumor vascular endothelial cells (ECs) and tumor cells. PS exposure becomes enhanced in response to chemotherapy, irradiation, and oxidative stresses in the tumor microenvironment. PS exposure in tumors promotes an immunosuppressive microenvironment which includes the recruitment of myeloid derived suppressor cells (MDSCs), immature dendritic cells and M2-like macrophages, as well as, the production of anti-inflammatory cytokines. Binding of PS targeting antibodies on tumor endothelial cells, tumor cells and their secreted microparticles triggers an Fc-FcR mediated pro-inflammatory cellular and cytokine response that reverses the immunosuppressive PS meditated checkpoint, thereby enhancing anti-tumor immunity. A chimeric PS-targeting antibody, bavituximab, is being used in combination with chemotherapy to treat patients with solid tumors in multiple late-stage clinical trials. Using breast tumors in immune competent mice, we demonstrate PS targeting antibodies enhance the anti-tumor activity of combination therapies including anti-PD-1 antibodies. Tumor growth inhibition correlates with statistically significant increases in the infiltration of CD8+ T cells and a reduction of myeloid-derived suppressor cells (MDSCs). The combination of these mechanisms promotes strong, localized, anti-tumor responses without the side-effects of systemic immune activation. Citation Format: Jian Gong, Shen Yin, Van Nguyen, Jeff Hutchins, Bruce D. Freimark. Targeting of phosphatidylserine by monoclonal antibodies enhances the activity of immune checkpoint inhibitors in breast tumors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4289. doi:10.1158/1538-7445.AM2015-4289

Abstract 4056: Recognition of tumor cells by Dectin-1 orchestrates innate immune cells for anti-tumor responses

Abstract The eradication of cancer cells requires communication between cells which constitute the complex immune system. Natural killer (NK) cells are essential tumor-killing effector cells of the innate arm of the immune system; however, little is known about whether or how other immune cells recognize tumor cells to assist NK cells. In addition, although signal-transducing pattern recognition receptors are known to play important roles in innate immune responses against invading pathogens through recognition of pathogen-associated molecular patterns (PAMPs), it is still enigmatic whether and how these receptors contribute to anti-tumor immune responses. In this study, we show that the innate immune pattern recognition receptor Dectin-1 expressed on dendritic cells (DCs) and macrophages is critical to NK-mediated killing of tumor cells. We also show that tumor cell-mediated Dectin-1 signaling is instigated by the receptor recognition of N-glycan structures on tumor cells, which we term tumor-associated molecular patterns (TAMPs). As such, the TAMP-Dectin-1 signaling causes activation of the Interferon Regulatory Factor 5 (IRF5) transcription factor and subsequent induction of genes required for the full-blown killing activity of NK cells. The importance of these events is underscored by the observation of massive tumor metastasis in mice genetically deficient in either Dectin-1 or IRF5. Thus, these results reveal a hitherto unrecognized facet of pattern recognition receptors in the orchestration of anti-tumor innate immune responses; recognition of TAMPs. This study is the first demonstration that an innate immune pattern recognition receptor potentiates anti-tumor immune responses, offering new insight into anti-tumor activity of the innate immune system with implications for anti-tumor immunotherapy. Citation Format: Hiroaki IKUSHIMA, Tadatsugu TANIGUCHI. Recognition of tumor cells by Dectin-1 orchestrates innate immune cells for anti-tumor responses. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4056. doi:10.1158/1538-7445.AM2015-4056

Analysis of immune cell populations and cytokine profiles in murine splenocytes exposed to whole-body low-dose irradiation

Purpose: In contrast to high-dose therapeutic irradiation, definitive research detailing the physiological effects of low-dose irradiation is limited. Notably, the immunological response elicited after low-dose irradiation remains controversial.Materials and methods: Female C57BL/6 mice were whole- body-irradiated with a single or three daily fractions up to a total dose of 0.1, 1, or 10 cGy. Blood and spleen were harvested 2, 7 and 14 days after irradiation.Results: The splenic CD4+ T cell subpopulations were temporarily increased at 2 days after single or fractionated irradiation, whereas the percentage of dendritic cells (DC) and macrophages was decreased. Whereas CD8+ T cell populations were decreased in single-dose irradiated mice at day 7, early and sustained reduction of CD8+ T cell numbers was observed in fractionated- dose-irradiated mice from day 2 until day 14. In addition, single-dose irradiation resulted in a Th1 cytokine expression profile, whereas fractionated-dose irradiation drove a Th2 shift. Additionally, increased expression of immune-related factors was observed at early time-points with single-dose irradiation, in contrast to the dose-independent induction following fractionated-dose irradiation.Conclusions: Our results demonstrate that low-dose irradiation modulates the immune response in mice, where the sensitivity and kinetics of the induced response vary according to the dosing method.

Innate Immune Sensing of Fusarium culmorum by Mouse Dendritic Cells

Chronic inhalation of grain dust is associated with asthma and chronic bronchitis in grain worker populations. Exposure to fungal particles was postulated to be an important etiologic agent of these pathologies. Fusarium species frequently colonize grain and straw and produce a wide array of mycotoxins that impact human health, necessitating an evaluation of risk exposure by inhalation of Fusarium and its consequences on immune responses. Data showed that Fusarium culmorum is a frequent constituent of aerosols sampled during wheat harvesting in the Vaud region of Switzerland. The aim of this study was to examine cytokine/chemokine responses and innate immune sensing of F. culmorum in bone-marrow-derived dendritic cells and macrophages. Overall, dendritic cells and macrophages responded to F. culmorum spores but not to its secreted components (i.e., mycotoxins) by releasing large amounts of macrophage inflammatory protein (MIP)-1α, MIP-1β, MIP-2, monocyte chemoattractant protein (MCP)-1, RANTES, and interleukin (IL)-12p40, intermediate amounts of tumor necrosis factor (TNF), IL-6, IL-12p70, IL-33, granulocyte colony-stimulating factor (G-CSF), and interferon gamma-induced protein (IP-10), but no detectable amounts of IL-4 and IL-10, a pattern of mediators compatible with generation of Th1 or Th17 antifungal protective immune responses rather than with Th2-dependent allergic responses. The sensing of F. culmorum spores by dendritic cells required dectin-1, the main pattern recognition receptor involved in β-glucans detection, but likely not MyD88 and TRIF-dependent Toll-like receptors. Taken together, our results indicate that F. culmorum stimulates potently innate immune cells in a dectin-1-dependent manner, suggesting that inhalation of F. culmorum from grain dust may promote immune-related airway diseases in exposed worker populations.

Ubiquitous Over-Expression of Chromatin Remodeling Factor SRG3 Ameliorates the T Cell-Mediated Exacerbation of EAE by Modulating the Phenotypes of both Dendritic Cells and Macrophages.

Although SWI3-related gene (SRG3), a chromatin remodeling factor, is critical for various biological processes including early embryogenesis and thymocyte development, it is unclear whether SRG3 is involved in the differentiation of CD4+ T cells, the key mediator of adaptive immune responses. Because it is known that experimental autoimmune encephalomyelitis (EAE) development is determined by the activation of CD4+ T helper cells, here, we investigated the role of SRG3 in EAE development using SRG3 transgenic mouse models exhibiting two distinct SRG3 expression patterns: SRG3 expression driven by either the CD2 or β-actin promoter. We found that the outcome of EAE development was completely different depending on the expression pattern of SRG3. The specific over-expression of SRG3 using the CD2 promoter facilitated EAE via the induction of Th1 and Th17 cells, whereas the ubiquitous over-expression of SRG3 using the β-actin promoter inhibited EAE by promoting Th2 differentiation and suppressing Th1 and Th17 differentiation. In addition, the ubiquitous over-expression of SRG3 polarized CD4+ T cell differentiation towards the Th2 phenotype by converting dendritic cells (DCs) or macrophages to Th2 types. SRG3 over-expression not only reduced pro-inflammatory cytokine production by DCs but also shifted macrophages from the inducible nitric oxide synthase (iNOS)-expressing M1 phenotype to the arginase-1-expressing M2 phenotype during EAE. In addition, Th2 differentiation in β-actin-SRG3 Tg mice during EAE was associated with an increase in the basophil and mast cell populations and in IL4 production. Furthermore, the increased frequency of Treg cells in the spinal cord of β-actin-SRG3 Tg mice might induce the suppression of and accelerate the recovery from EAE symptoms. Taken together, our results provide the first evidence supporting the development of a new therapeutic strategy for EAE involving the modulation of SRG3 expression to induce M2 and Th2 polarization, thereby inhibiting inflammatory immune responses.

Alterations of dendritic cells, inflammatory monocytes and macrophages in mice during Pneumocystis murina infection

Objective To investigate the alterations and phenotypes of dendritic cells, inflammatory monocytes and macrophages in immunocompetent mice during Pneumocystis murina (P.murina) infection for further analysis of the function of these cells during P. murina infection. Methods Wild type male C57BL/6 mice at age 6-8 weeks were randomly divided into two groups including the group with P. murina infection and the group receiving sham surgery. The mice without any intervention were used to set up the blank control group. The loads of P. murina strains in lung tissues of each mouse were quantified by TaqMan real-time fluorescence polymerase chain reaction after the infection. Histopathological examination was performed to evaluate the degree of inflammation in lung tissues. The numbers of dendritic cells, inflammatory monocytes and macrophages in lung tissues, peripheral blood and bone marrow samples, and the changes of inflammatory monocytes in spleen tissues were measured by flow cytometry analysis. The expression of major histocompatability complex Ⅱ (MHCⅡ), CX3C chemokine receptor 1 (CX3CR1) and CC chemokine receptor 2 (CCR2) by dendritic cells, inflammatory monocytes and macrophages in lung tissues during P. murina infection were analyzed by flow cytometry analysis. All of the data were collected one, two, three and four weeks after the corresponding treatments. Results The loads of P. murina strains in P. murina infected mice were elevated after two and three weeks infection, but decline at week 4 (P>0.05). Significant pathological changes including the alveolar destruction, inflammatory cell infiltration and thickened alveolar septum in mice with P. murina infection were observed under a microscope at week 3 and week 4. Compared to the sham surgery treatment group, the number of CD11c+ CD11b+ dendritic cells were increased in lung tissues, but decreased in blood samples during P. murina infection (P 0.05). The CD11c+ CD11b+ dendritic cells in lung tissues of mice with P. murina infection expressed high levels of MHCⅡ and CX3CR1, and low levels of CCR2. The inflammatory monocytes in lung tissues of mice expressed high levels of CCR2, moderate levels of MHCⅡ and low levels of CX3CR1 during P. murina infection. High levels of CX3CR1 and low levels of MHCⅡ and CCR2 were observed in macrophages from lung tissues of mice with P. murina infection. Conclusion Highly expressed CD11c+ CD11b+ dendritic cells and MHCⅡ were detected in lung tissues of mice during P. murina infection, indicating that CD11c+ CD11b+ dendritic cells were involved in the host defense against P. murina infection. Key words: Pneumocystis murina infection; Dendritic cells; Inflammatory monocytes; Macrophages

Correction: Oxidation by Neutrophils-Derived HOCl Increases Immunogenicity of Proteins by Converting Them into Ligands of Several Endocytic Receptors Involved in Antigen Uptake by Dendritic Cells and Macrophages

Correction: Oxidation by Neutrophils-Derived HOCl Increases Immunogenicity of Proteins by Converting Them into Ligands of Several Endocytic Receptors Involved in Antigen Uptake by Dendritic Cells and Macrophages

Protective immune responses to dengue virus infection and vaccines: perspectives from the field to the bench.

Protective immune responses to dengue virus infection and vaccines: perspectives from the field to the bench.

Cytokine release and endothelial dysfunction: a perfect storm in orbivirus pathogenesis.

Although bluetongue viruses (BTV) and epizootic haemorrhagic disease viruses (EHDV) are closely related, there are differences in susceptibility to these viruses both between and within a species. White‑tailed deer are susceptible to disease by both BTV and EHDV, sheep are susceptible to BTV, but resistant to EHDV, and cattle can be infected with both viruses but disease is usually subclinical. Host genetics probably play a role in the disease outcome, but cytokine and endothelial responses are likely to determine if subclinical or clinical disease develops. Dendritic macrophages deliver virus to lymph nodes following the bite of an infected Culicoides. The virus then disseminates to many organs replicating in mononuclear phagocytes and endothelium. Initially, an interferon‑1 response probably determines if the disease develops. Replication in mononuclear cells and endothelium results in the release of cytokines and vasoactive mediators, and may result in endothelial cell death leading to the clinical features of fever, hyperaemia, exudation of fluid, and haemorrhage. Disease outcome may also be linked to virus binding Toll‑like receptor‑3 and upregulation of endothelial surface receptors potentiating cytokine release and allowing transmigration of inflammatory cells, respectively. Despite a wealth of information, host genetics involved in resistance to BTV and EHDV and how variations in cytokines and endothelial responses determine clinical outcome still need further elucidation.

The Good, the Bad, and the Ugly of Dendritic Cells during Prion Disease.

Prions are a unique group of proteinaceous pathogens which cause neurodegenerative disease and can be transmitted by a variety of exposure routes. After peripheral exposure, the accumulation and replication of prions within secondary lymphoid organs are obligatory for their efficient spread from the periphery to the brain where they ultimately cause neurodegeneration and death. Mononuclear phagocytes (MNP) are a heterogeneous population of dendritic cells (DC) and macrophages. These cells are abundant throughout the body and display a diverse range of roles based on their anatomical locations. For example, some MNP are strategically situated to provide a first line of defence against pathogens by phagocytosing and destroying them. Conventional DC are potent antigen presenting cells and migrate via the lymphatics to the draining lymphoid tissue where they present the antigens to lymphocytes. The diverse roles of MNP are also reflected in various ways in which they interact with prions and in doing so impact on disease pathogenesis. Indeed, some studies suggest that prions exploit conventional DC to infect the host. Here we review our current understanding of the influence of MNP in the pathogenesis of the acquired prion diseases with particular emphasis on the role of conventional DC.

A study of serum interleukin-12 in a sample of autistic children in Egypt

Introduction Autistic spectrum disorders (ASDs) prevalence varies widely by sex and the racial/ethnic group. The male-to-female ratio ranged from 3 to 4 : 1. Some consider ASD to be an autoimmune disorder, in which the autoimmune response to the developing brain myelin may impair anatomical development of neural pathways in autistic children; this affects the speed of impulse transmission. Interleukin-12 (IL-12) is an interleukin that is naturally produced by dendritic cells and macrophages in response to antigenic stimulation; it plays an important role in the activities of natural killer cells and T lymphocytes involved in the immune system. Aim of the work The present study was conducted to compare the level of serum IL-12 between children with autistic disorder (AD) and healthy control children, and also to study the relation of serum IL-12 with the severity of autistic symptoms. Participants and methods A cross-sectional study was conducted on two groups; group I included 20 patients with AD and group II included 20 normal children matched for age and sex, recruited from the Child and Adolescent Outpatients Clinic at Al Hadra University Hospital. All children were subjected to a complete psychiatric history, physical and neurological examination, psychometric assessment by Childhood Autistic Rating Scale, and estimation of serum IL-12 using the enzyme-linked immunosorbent assay method. Results The mean serum level of IL-12 was significantly higher in AD children than in controls and was related to a younger age, male sex, a positive family history and ante/natal/postnatal history, nondevelopment of spoken language, the presence of comorbidities, and higher Childhood Autistic Rating Scale mean scores. Conclusion The study pointed out an immunological impairment in the form of an elevated serum level of IL-12 in autistic children and its positive relation to autistic symptom severity. This supports the immunological etiology of ASD.