Myeloid-Derived Suppressor Cells Modulate Immune Responses Independently of NADPH Oxidase in the Ovarian Tumor Microenvironment in Mice

The phagocyte NADPH oxidase generates superoxide anion and downstream reactive oxidant intermediates in response to infectious threat, and is a critical mediator of antimicrobial host defense and inflammatory responses. Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells that are recruited by cancer cells, accumulate locally and systemically in advanced cancer, and can abrogate anti-tumor immunity. Prior studies have implicated the phagocyte NADPH oxidase as being an important component promoting MDSC accumulation and immunosuppression in cancer. We therefore used engineered NADPH oxidase-deficient (p47^phox−/−) mice to delineate the role of this enzyme complex in MDSC accumulation and function in a syngeneic mouse model of epithelial ovarian cancer. We found that the presence of NADPH oxidase did not affect tumor progression. The accumulation of MDSCs locally and systemically was similar in tumor-bearing wild-type (WT) and p47^phox−/− mice. Although MDSCs from tumor-bearing WT mice had functional NADPH oxidase, the suppressive effect of MDSCs on ex vivo stimulated T cell proliferation was NADPH oxidase-independent. In contrast to other tumor-bearing mouse models, our results show that MDSC accumulation and immunosuppression in syngeneic epithelial ovarian cancer is NADPH oxidase-independent. We speculate that factors inherent to the tumor, tumor microenvironment, or both determine the specific requirement for NADPH oxidase in MDSC accumulation and function.     

IL‐18 Cytokine Levels Modulate Innate Immune Responses and Cryptosporidiosis in Mice

IL‐18 is known to play a key role limiting Cryptosporidium parvum infection. In this study, we show that IL‐18 depletion in SCID mice significantly exacerbates C. parvum infection, whereas, treatment with recombinant IL‐18 (rIL‐18), significantly decreases the parasite load, as compared to controls. Increases in serum IFN‐γ levels as well as the up‐regulation of the antimicrobial peptides, cathelicidin antimicrobial peptide and beta defensin 3 (Defb3) were observed in the intestinal mucosa of mice treated with rIL‐18. In addition, C. parvum infection significantly increased mRNA expression levels (> 50 fold) of the alpha defensins, Defa3 and 5, respectively. Interestingly, we also found a decrease in mRNA expression of IL‐33 (a recently identified cytokine in the same family as IL‐18) in the small intestinal tissue from mice treated with rIL‐18. In comparison, the respective genes were induced by IL‐18 depletion. Our findings suggest that IL‐18 can mediate its protective effects via different routes such as IFN‐γ induction or by directly stimulating intestinal epithelial cells to increase antimicrobial activity.     

Altered Innate Immune and Glial Cell Responses to Inflammatory Stimuli in Amyloid Precursor Protein Knockout Mice

Amyloid precursor protein (APP) and its cleaved products have been reported to have important functions in CNS health, including in memory and synapse formation, cell survival and neuroprotection. Furthermore APP and its cleaved products have been shown to be transiently increased in response to various CNS stressors, suggesting a role in response to acute cellular injury. In an attempt to further understand the function of APP in response to CNS injury, we have used intracranial LPS injection as an inflammatory injury model in APP knock out mice (APPKO). Our data show that innate immune responses to LPS injection is significantly blunted in APPKO mice compared to APP sufficient wild type (BL6) mice. Morphologically, glial cells in APPKO mice appear less reactive, with shorter ramified processes and smaller cell bodies in response to LPS. Additionally, quantitative RT-PCR analysis for several glia markers and innate immune cytokine levels (e.g. TNFα, IL-6, IL-1β and IL-10) showed significantly reduced expression levels in LPS injected APPKO mice. In vitro cell culture assays confirmed this attenuated response to LPS stimulation by primary microglial cells isolated from APPKO mice. Our data suggests that APP full length protein and/or its cleaved products are necessary to mount a complete and effective innate immune cell response to inflammatory injury.     

Paramyxovirus Activation and Inhibition of Innate Immune Responses

Paramyxoviruses represent a remarkably diverse family of enveloped nonsegmented negative-strand RNA viruses, some of which are the most ubiquitous disease-causing viruses of humans and animals. This review focuses on paramyxovirus activation of innate immune pathways, the mechanisms by which these RNA viruses counteract these pathways, and the innate response to paramyxovirus infection of dendritic cells (DC). Paramyxoviruses are potent activators of extracellular complement pathways, a first line of defense that viruses must face during natural infections. We discuss mechanisms by which these viruses activate and combat complement to delay neutralization. Once cells are infected, virus replication drives type I interferon (IFN) synthesis that has the potential to induce a large number of antiviral genes. Here we describe four approaches by which paramyxoviruses limit IFN induction: by limiting synthesis of IFN-inducing aberrant viral RNAs, through targeted inhibition of RNA sensors, by providing viral decoy substrates for cellular kinase complexes, and through direct blocking of the IFN promoter. In addition, paramyxoviruses have evolved diverse mechanisms to disrupt IFN signaling pathways. We describe three general mechanisms, including targeted proteolysis of signaling factors, sequestering cellular factors, and upregulation of cellular inhibitors. DC are exceptional cells with the capacity to generate adaptive immunity through the coupling of innate immune signals and T cell activation. We discuss the importance of innate responses in DC following paramyxovirus infection and their consequences for the ability to mount and maintain antiviral T cells.     

流感病毒先天免疫应答和先天免疫逃逸的机制

流感病毒引起人类和动物的呼吸道感染已是全世界严重的经济和公共卫生问题。在感染早期,流感病毒会导致机体的先天免疫信号被激活,起到防御、清除病毒以及辅助适应性免疫应答的作用。但在与宿主共进化的过程中,流感病毒形成了多种逃逸策略,主要是通过病毒自身蛋白质阻断宿主天然免疫通路,抑制干扰素和炎性因子的生成。基于现有的研究成果,本文针对流感病毒先天免疫应答和先天免疫逃逸的机制做一扼要综述,这有助于加强流感病毒抗原进化的监测、探索疫苗和抗病毒药物的合理靶标,为更好地预防和控制该病提供有效的策略。     

人类冠状病毒调节宿主抗病毒天然免疫分子机制

SARS冠状病毒和正在全球流行的猪源H1N1型流感病毒等人类新发呼吸道病毒对人类生命健康构成严重威胁.人类重要呼吸道病毒与宿主抗病毒天然免疫的关系是近年来研究热点.SARS冠状病毒等很多RNA病毒能够编码某种蛋白质,抑制干扰素表达以及干扰素介导的抗病毒信号通路.人类冠状病毒木瓜样蛋白酶(papain-like protease,PLP)利用其自身去泛素化酶(DUB)活性,使干扰素表达通路中重要调节蛋白发生去泛素化,从而抑制干扰素信号传导.同时,PLP蛋白酶通过阻碍干扰素表达信号通路中最新发现的重要调节蛋白ERIS(也称MITA/STING)二聚化,使其失活并丧失激活干扰素通路的功能,这些发现对于阐明人类重要呼吸道病毒对宿主细胞抗病毒天然免疫反应的调节作用及其机制具有重要意义,为人类新发病毒致病机理、免疫防治以及抗病毒药物研究提供新的思路.     

Heightened Innate Immune Responses in the Respiratory Tract of COVID-19 Patients

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Salicylic Acid Regulates Plasmodesmata Closure during Innate Immune Responses in Arabidopsis

In plants, mounting an effective innate immune strategy against microbial pathogens involves triggering local cell death within infected cells as well as boosting the immunity of the uninfected neighboring and systemically located cells. Although not much is known about this, it is evident that well-coordinated cell–cell signaling is critical in this process to confine infection to local tissue while allowing for the spread of systemic immune signals throughout the whole plant. In support of this notion, direct cell-to-cell communication was recently found to play a crucial role in plant defense. Here, we provide experimental evidence that salicylic acid (SA) is a critical hormonal signal that regulates cell-to-cell permeability during innate immune responses elicited by virulent bacterial infection in Arabidopsis thaliana. We show that direct exogenous application of SA or bacterial infection suppresses cell–cell coupling and that SA pathway mutants are impaired in this response. The SA- or infection-induced suppression of cell–cell coupling requires an ENHANCED DESEASE RESISTANCE1– and NONEXPRESSOR OF PATHOGENESIS-RELATED GENES1–dependent SA pathway in conjunction with the regulator of plasmodesmal gating PLASMODESMATA-LOCATED PROTEIN5. We discuss a model wherein the SA signaling pathway and plasmodesmata-mediated cell-to-cell communication converge under an intricate regulatory loop.     

Selenium Supplementation Restores Innate and Humoral Immune Responses in Footrot-Affected Sheep

Dietary selenium (Se) alters whole-blood Se concentrations in sheep, dependent upon Se source and dosage administered, but little is known about effects on immune function. We used footrot (FR) as a disease model to test the effects of supranutritional Se supplementation on immune function. To determine the effect of Se-source (organic Se-yeast, inorganic Na-selenite or Na-selenate) and Se-dosage (1, 3, 5 times FDA-permitted level) on FR severity, 120 ewes with and 120 ewes without FR were drenched weekly for 62 weeks with different Se sources and dosages (30 ewes/treatment group). Innate immunity was evaluated after 62 weeks of supplementation by measuring neutrophil bacterial killing ability. Adaptive immune function was evaluated by immunizing sheep with keyhole limpet hemocyanin (KLH). The antibody titer and delayed-type hypersensitivity skin test to KLH were used to assess humoral immunity and cell-mediated immunity, respectively. At baseline, FR-affected ewes had lower whole-blood and serum-Se concentrations; this difference was not observed after Se supplementation. Se supplementation increased neutrophil bacterial killing percentages in FR-affected sheep to percentages observed in supplemented and non-supplemented healthy sheep. Similarly, Se supplementation increased KLH antibody titers in FR-affected sheep to titers observed in healthy sheep. FR-affected sheep demonstrated suppressed cell-mediated immunity at 24 hours after intradermal KLH challenge, although there was no improvement with Se supplementation. We did not consistently prevent nor improve recovery from FR over the 62 week Se-treatment period. In conclusion, Se supplementation does not prevent FR, but does restore innate and humoral immune functions negatively affected by FR.     

Mouse Hepatitis Virus Infection Upregulates Genes Involved in Innate Immune Responses

Neurotropic recombinant strain of Mouse Hepatitis Virus, RSA59, induces meningo-encephalitis, myelitis and demyelination following intracranial inoculation. RSA59 induced neuropathology is partially caused by activation of CNS resident microglia, as demonstrated by changes in cellular morphology and increased expression of a microglia/macrophage specific calcium ion binding factor, Iba1. Affymetrix Microarray analysis for mRNA expression data reveals expression of inflammatory mediators that are known to be released by activated microglia. Microglia-specific cell surface molecules, including CD11b, CD74, CD52 and CD68, are significantly upregulated in contrast to CD4, CD8 and CD19. Protein analysis of spinal cord extracts taken from mice 6 days post-inoculation, the time of peak inflammation, reveals robust expression of IFN-γ, IL-12 and mKC. Data suggest that activated microglia and inflammatory mediators contribute to a local CNS microenvironment that regulates viral replication and IFN-γ production during the acute phase of infection, which in turn can cause phagolysosome maturation and phagocytosis of the myelin sheath, leading to demyelination.     

Influence of Phthalates on in vitro Innate and Adaptive Immune Responses

Phthalates are a group of endocrine disrupting chemicals, suspected to influence the immune system. The aim of this study was to investigate the influence of phthalates on cytokine secretion from human peripheral blood mononuclear cells. Escherichia coli lipopolysaccharide and phytohemagglutinin-P were used for stimulation of monocytes/macrophages and T cells, respectively. Cells were exposed for 20 to 22 hours to either di-ethyl, di-n-butyl or mono-n-butyl phthalate at two different concentrations. Both diesters were metabolised to their respective monoester and influenced cytokine secretion from both monocytes/macrophages and T cells in a similar pattern: the secretion of interleukin (IL)-6, IL-10 and the chemokine CXCL8 by monocytes/macrophages was enhanced, while tumour necrosis factor (TNF)-α secretion by monocytes/macrophages was impaired, as was the secretion of IL-2 and IL-4, TNF-α and interferon-γ by T cells. The investigated phthalate monoester also influenced cytokine secretion from monocytes/macrophages similar to that of the diesters. In T cells, however, the effect of the monoester was different compared to the diesters. The influence of the phthalates on the cytokine secretion did not seem to be a result of cell death. Thus, results indicate that both human innate and adaptive immunity is influenced in vitro by phthalates, and that phthalates therefore may affect cell differentiation and regenerative and inflammatory processes in vivo.     

Quantitative Proteomics Reveals the Roles of Peroxisome-associated Proteins in Antiviral Innate Immune Responses

Compared with whole-cell proteomic analysis, subcellular proteomic analysis is advantageous not only for the increased coverage of low abundance proteins but also for generating organelle-specific data containing information regarding dynamic protein movement. In the present study, peroxisome-enriched fractions from Sendai virus (SeV)-infected or uninfected HepG2 cells were obtained and subjected to quantitative proteomics analysis. We identified 311 proteins that were significantly changed by SeV infection. Among these altered proteins, 25 are immune response-related proteins. Further bioinformatic analysis indicated that SeV infection inhibits cell cycle-related proteins and membrane attack complex-related proteins, all of which are beneficial for the survival and replication of SeV within host cells. Using Luciferase reporter assays on several innate immune-related reporters, we performed functional analysis on 11 candidate proteins. We identified LGALS3BP and CALU as potential negative regulators of the virus-induced activation of the type I interferons.One of the most significant evolutionary features in eukaryotes is the appearance of a membrane system to separate enzymatic reactions and to provide scaffolds for signal transduction. The small genome of viruses requires that they use the host''s cellular machinery, especially host intracellular membranes, to assemble their replication complexes and to complete their replication cycle (1). Therefore, it is not surprising to find that the eukaryotic membrane system is also involved in antiviral responses (2–4).Subcellular organelles with extensive membrane systems include the endoplasmic reticulum (ER)¹, mitochondria, endosomes and peroxisomes. Despite their well-recognized functions in cell metabolism, these organelles and their related membranes have been identified in recent years as important innate immune platforms (5). The ER is a key organelle for maintaining cellular homeostasis. Several recent studies have linked the ER to antiviral immune responses and have elucidated the related mechanisms (6–10). The mitochondria serve mainly as the power plants of eukaryotic cells but also participate in numerous crucial cellular processes, such as calcium homeostasis, apoptosis and aging (11–13). In recent years, the mitochondria and mitochondrial-associated membranes (MAM) have also emerged as fundamental hubs for innate antiviral immunity. Several important antiviral immune-related proteins, such as VISA (also referred to as MAVS/CARDIF/IPS-1) and MITA (also referred to as STING), have been found to localize to the mitochondrial membrane (14–19). Peroxisomes are monolayer-membrane organelles present in nearly all types of human cells, with a particularly high abundance in hepatocytes and nephrocytes, which are involved in various oxidative enzymatic reactions. It has been reported that the peroxisomal-associated protein VISA induces a rapid interferon (IFN)-independent expression that provides short-term protection, whereas the mitochondrial-associated VISA activates an IFN-dependent signaling pathway with delayed kinetics (20). The peroxisomal VISA can activates IRF1-mediated IFN-λ production (21). The endosomes, although known as players in cellular endocytosis and vascular transport, have functions that extend to the antigen presentation of major histocompatibility complex (MHC) class I, MHC class II and CD1 molecules of the adaptive immune system, as well as pattern recognition of innate immune-related receptors, such as Toll-like receptors (TLRs) and NOD-like receptors (NLRs) (22, 23).Subcellular fractionation is an effective experimental strategy for isolating/enriching specific organelles. Compared with whole-cell-based proteomic analysis, combining the isolation of subcellular components with mass spectrometry-based proteomic analysis is advantageous not only for characterizing low abundance proteins but also for monitoring protein abundance changes at the organelle level. To systematically analyze the role of peroxisomal-related proteins in innate immune responses, we used a modified two-step gradient centrifugation method to enrich the peroxisomes from cells with or without SeV infection, followed by a quantitative proteomic analysis. A total of 2946 proteins were quantified among which 311 proteins were found to be significantly changed by SeV infection. A statistical enrichment test was used to reveal that 13 protein groups were changed significantly (p < 0.05) compared with the entire protein list. Cell cycle-related proteins and membrane attack complex (MAC)-related proteins were down-regulated, which may facilitate virus survival and replication in host cells. Luciferase reporter assays were performed to further screen for the significantly changed proteins that could affect SeV-induced activation of the type I IFN signaling pathway. Not only does our data provide new and unbiased protein-level information regarding viral infection processes, we also provide direct evidence for the involvement of two proteins (CALU and LGALS3BP) as potential negative regulators in the virus-triggered induction of the type I IFNs.     

Three Pairs of Protease-Serpin Complexes Cooperatively Regulate the Insect Innate Immune Responses

Serpins are known to be necessary for the regulation of several serine protease cascades. However, the mechanisms of how serpins regulate the innate immune responses of invertebrates are not well understood due to the uncertainty of the identity of the serine proteases targeted by the serpins. We recently reported the molecular activation mechanisms of three serine protease-mediated Toll and melanin synthesis cascades in a large beetle, Tenebrio molitor. Here, we purified three novel serpins (SPN40, SPN55, and SPN48) from the hemolymph of T. molitor. These serpins made specific serpin-serine protease pairs with three Toll cascade-activating serine proteases, such as modular serine protease, Spätzle-processing enzyme-activating enzyme, and Spätzle-processing enzyme and cooperatively blocked the Toll signaling cascade and β-1,3-glucan-mediated melanin biosynthesis. Also, the levels of SPN40 and SPN55 were dramatically increased in vivo by the injection of a Toll ligand, processed Spätzle, into Tenebrio larvae. This increase in SPN40 and SPN55 levels indicates that these serpins function as inducible negative feedback inhibitors. Unexpectedly, SPN55 and SPN48 were cleaved at Tyr and Glu residues in reactive center loops, respectively, despite being targeted by trypsin-like Spätzle-processing enzyme-activating enzyme and Spätzle-processing enzyme. These cleavage patterns are also highly similar to those of unusual mammalian serpins involved in blood coagulation and blood pressure regulation, and they may contribute to highly specific and timely inactivation of detrimental serine proteases during innate immune responses. Taken together, these results demonstrate the specific regulatory evidences of innate immune responses by three novel serpins.     

Sphingolipid Pathway Regulates Innate Immune Responses at the Fetomaternal Interface during Pregnancy

For a successful pregnancy, the mother''s immune system has to tolerate the semiallogeneic fetus. A deleterious immune attack is avoided by orchestration of cellular, hormonal, and enzymatic factors. However, the precise mechanisms underlying fetomaternal tolerance are not yet completely understood. In this study, we demonstrate that sphingolipid metabolism constitutes a novel signaling pathway that is indispensable for fetomaternal tolerance by regulating innate immune responses at the fetomaternal interface. Perturbation of the sphingolipid pathway by disruption of the sphingosine kinase gene (Sphk) during pregnancy caused unusually high expression of neutrophil chemoattractants, CXCL1 and CXCL2, in the decidua, leading to a massive infiltration of neutrophils into the fetomaternal interface with enhanced oxidative damage, resulting in early fetal death. Sphk-deficient mice also exhibited neutrophilia in the peripheral blood, enhanced generation of granulocytes in the bone marrow, and a decrease in the number of decidual natural killer cells. The blockage of neutrophil influx protected Sphk-deficient mice against pregnancy loss. Notably, a similar result was obtained in human decidual cells, in which Sphk deficiency dramatically increased the secretion of CXCL1 and IL-8. In conclusion, our findings suggest that the sphingolipid metabolic pathway plays a critical role in fetomaternal tolerance by regulating innate immunity at the fetomaternal interface both in mice and humans, and it could provide novel insight into the development of therapeutic strategies to treat idiopathic pregnancy loss in humans.     

Inhibition of Innate Immune Responses Is Key to Pathogenesis by Arenaviruses

Mammalian arenaviruses are zoonotic viruses that cause asymptomatic, persistent infections in their rodent hosts but can lead to severe and lethal hemorrhagic fever with bleeding and multiorgan failure in human patients. Lassa virus (LASV), for example, is endemic in several West African countries, where it is responsible for an estimated 500,000 infections and 5,000 deaths annually. There are currently no FDA-licensed therapeutics or vaccines available to combat arenavirus infection. A hallmark of arenavirus infection (e.g., LASV) is general immunosuppression that contributes to high viremia. Here, we discuss the early host immune responses to arenavirus infection and the recently discovered molecular mechanisms that enable pathogenic viruses to suppress host immune recognition and to contribute to the high degree of virulence. We also directly compare the innate immune evasion mechanisms between arenaviruses and other hemorrhagic fever-causing viruses, such as Ebola, Marburg, Dengue, and hantaviruses. A better understanding of the immunosuppression and immune evasion strategies of these deadly viruses may guide the development of novel preventative and therapeutic options.