Brain heterogeneity leads to differential innate immune responses and modulates pathogenesis of viral infections

The central nervous system (CNS) is a highly complex organ with highly specialized cell subtypes. Viral infections often target specific structures of the brain and replicate in certain regions. Studies in mice deficient in type I Interferon (IFN) receptor or IFN-β have highlighted the importance of the type I IFN system against viral infections and non-viral autoimmune disorders in the CNS. Direct antiviral effects of type I IFNs appear to be crucial in limiting early spread of a number of viruses in CNS tissues. Increased efforts have been made to characterize IFN expression and responses in the brain. In this context, it is important to identify cells that produce IFN, decipher pathways leading to type I IFN expression and to characterize responding cells. In this review we give an overview about region specific aspects that influence local innate immune responses. The route of entry is critical, but also the susceptibility of different cell types, heterogeneity in subpopulations and micro-environmental cues play an important role in antiviral responses.Recent work has outlined the tremendous importance of type I IFNs, particularly in the limitation of viral spread within the CNS. This review will address recent advances in understanding the mechanisms of local type I IFN production and response, in the particular context of the CNS.     

Role of type I interferons in the activation of autoreactive B cells

Type I interferons (IFNs) are a family of cytokines involved in the defense against viral infections that play a key role in the activation of both the innate and adaptive immune system. IFNs both directly and indirectly enhance the capacity of B lymphocytes to respond to viral challenge and produce cytotoxic and neutralizing antibodies. However, prolonged type I IFN exposure is not always beneficial to the host. If not regulated properly IFN can drive autoantibody production as well as other parameters of systemic autoimmune disease. Type I IFNs impact B-cell function through a variety of mechanisms, including effects on receptor engagement, Toll-like receptor expression, cell migration, antigen presentation, cytokine responsiveness, cytokine production, survival, differentiation and class-switch recombination. Type I IFNs are also cytotoxic for a variety of cell types and thereby contribute to the accumulation of cell debris that serves as a potential source for autoantigens. Type I IFN engagement of a variety of accessory cells further promotes B-cell survival and activation, as exemplified by the capacity of type I IFNs to increase the level of B-cell survival factors, such as B lymphocyte stimulator, produced by dendritic cells. Therefore, it is not surprising that the loss of expression of the type I IFN receptor can have dramatic effects on the production of autoantibodies and on the clinical features of systemic autoimmune diseases such as systemic lupus erythematosus.     

Type I interferons directly down-regulate BCL-6 in primary and transformed germinal center B cells: differential regulation in B cell lines derived from endemic or sporadic Burkitt's lymphoma

Type I interferons (IFN) exert multiple effects on both the innate and adaptive immune system in addition to their antiviral and antiproliferative activities. Little is known, however about the direct effects of type I IFNs on germinal center (GC) B cells, the central components of adaptive B cell responses. We used Burkitt's lymphoma (BL) lines, as a model system of normal human GC B cells, to examine the effect of type I IFNs on the expression of BCL-6, the major regulator of the GC reaction. We show that type I IFNs, but not IFNγ, IL-2 and TNFα rapidly down-regulate BCL-6 protein and mRNA expression, in cell lines derived from endemic, but not from sporadic BL. IFNα-induced down-regulation is specific for BCL-6, independent of Epstein-Barr virus and is not accompanied by IRF-4 up-regulation. IFNα-induced BCL-6 mRNA down-regulation does not require de novo protein synthesis and is specifically inhibited by piceatannol. The proteasome inhibitor MG132 non-specifically prevents, while inhibitors of alternate type I IFN signaling pathways do not inhibit IFNα-induced BCL-6 protein downregulation. We validate our results with showing that IFNα rapidly down-regulates BCL-6 mRNA in purified mouse normal GC B cells. Our results identify type I IFNs as the first group of cytokines that can down-regulate BCL-6 expression directly in GC B cells.     

Microglial Interferon Signaling and White Matter

Microglia, the resident immune cells of the CNS, are primary regulators of the neuroimmune response to injury. Type I interferons (IFNs), including the IFNαs and IFNβ, are key cytokines in the innate immune system. Their activity is implicated in the regulation of microglial function both during development and in response to neuroinflammation, ischemia, and neurodegeneration. Data from numerous studies in multiple sclerosis (MS) and stroke suggest that type I IFNs can modulate the microglial phenotype, influence the overall neuroimmune milieu, regulate phagocytosis, and affect blood–brain barrier integrity. All of these IFN-induced effects result in numerous downstream consequences on white matter pathology and microglial reactivity. Dysregulation of IFN signaling in mouse models with genetic deficiency in ubiquitin specific protease 18 (USP18) leads to a severe neurological phenotype and neuropathological changes that include white matter microgliosis and pro-inflammatory gene expression in dystrophic microglia. A class of genetic disorders in humans, referred to as pseudo-TORCH syndrome (PTS) for the clinical resemblance to infection-induced TORCH syndrome, also show dysregulation of IFN signaling, which leads to severe neurological developmental disease. In these disorders, the excessive activation of IFN signaling during CNS development results in a destructive interferonopathy with similar induction of microglial dysfunction as seen in USP18 deficient mice. Other recent studies implicate “microgliopathies” more broadly in neurological disorders including Alzheimer’s disease (AD) and MS, suggesting that microglia are a potential therapeutic target for disease prevention and/or treatment, with interferon signaling playing a key role in regulating the microglial phenotype.     

Interferons and interferon inhibitory activity in disease and therapy

Interferon (IFN) resistance is an important factor in the pathophysiology of neoplastic disorders, certain viral infections (e.g., AIDS), and autoimmune diseases (e.g., lupus erythematosus and Wegner's granulomatosis). In addition, in some of these disorders, there is also decreased ability to produce IFNs. The capacity of viruses and neoplastic processes to interfere with the IFN system are thought to represent a "virus-against-host" or "cancer-against-host" defense mechanism. Four resistance factors have been identified: 1) release of free IFN-alpha/beta type 1 receptors into the circulation that, at appropriate concentrations, capture and inactivate IFNs; 2) a new IFN inhibitory protein has been isolated and its chemical structure is under study; 3) prostaglandin E2, which is produced by certain tumor cells, inhibits IFN production; and 4) high levels of cAMP phosphodiesterases present, for example in certain tumor cells, reduces cAMP, an important second messenger in IFN synthesis. Studies are under way to reverse these inhibitory effects and to increase endogenous interferon production.     

An important role for type III interferon (IFN-lambda/IL-28) in TLR-induced antiviral activity

Type III IFNs (IFN-lambda/IL-28/29) are cytokines with type I IFN-like antiviral activities, which remain poorly characterized. We herein show that most cell types expressed both types I and III IFNs after TLR stimulation or virus infection, whereas the ability of cells to respond to IFN-lambda was restricted to a narrow subset of cells, including plasmacytoid dendritic cells and epithelial cells. To examine the role of type III IFN in antiviral defense, we generated IL-28Ralpha-deficient mice. These mice were indistinguishable from wild-type mice with respect to clearance of a panel of different viruses, whereas mice lacking the type I IFN receptor (IFNAR(-/-)) were significantly impaired. However, the strong antiviral activity evoked by treatment of mice with TLR3 or TLR9 agonists was significantly reduced in both IL-28RA(-/-) and IFNAR(-/-) mice. The type I IFN receptor system has been shown to mediate positive feedback on IFN-alphabeta expression, and we found that the type I IFN receptor system also mediates positive feedback on IFN-lambda expression, whereas IL-28Ralpha signaling does not provide feedback on either type I or type III IFN expression in vivo. Finally, using bone-marrow chimeric mice we showed that TLR-activated antiviral defense requires expression of IL-28Ralpha only on nonhemopoietic cells. In this compartment, epithelial cells responded to IFN-lambda and directly restricted virus replication. Our data suggest type III IFN to target a specific subset of cells and to contribute to the antiviral response evoked by TLRs.     

The interferon system of teleost fish

Interferons (IFNs) are secreted proteins, which induce vertebrate cells into an antiviral state. In mammals, three families of IFNs (type I IFN, type II IFN and IFN-lambda) can be distinguished on the basis of gene structure, protein structure and functional properties. Type I IFNs, which include IFN-alpha and IFN-beta, are encoded by intron lacking genes and have a major role in the first line of defense against viruses. The human IFN-lambdas have similar biological properties as type I IFNs, but are encoded by intron containing genes. Type II IFN is identical to IFN-gamma, which is produced by T helper 1 cells in response to mitogens and antigens and has a key role in adaptive cell mediated immunity. IFNs, which show structural and functional properties similar to mammalian type I IFNs, have recently been cloned from Atlantic salmon, channel catfish, pufferfish, and zebrafish. Teleost fish appear to have at least two type I IFN genes. Phylogenetic sequence analysis shows that the fish type I IFNs form a group separated from the avian type I IFNs and the mammalian IFN-alpha, -beta and -lambda groups. Interestingly, the fish IFNs possess the same exon/intron structure as the IFN-lambdas, but show most sequence similarity to IFN-alpha. Recently, IFN-gamma genes have also been cloned from several fish species and shown to have the same exon/intron structure as mammalian IFN-gamma genes. The antiviral effect of mammalian type I IFN is exerted through binding to the IFN-alpha/beta-receptor, which triggers signal transduction through the JAK-STAT signal transduction pathway resulting in expression of Mx and other antiviral proteins. Putative IFN receptor genes have been identified in pufferfish. Several interferon regulatory factors and members of the JAK-STAT pathway have also been identified in various fish species. Moreover, Mx and several other interferon stimulated genes have been cloned and studied in fish. Furthermore, antiviral activity of Mx protein from Atlantic salmon and Japanese flounder has recently been demonstrated.     

Distinct evolution process among type I interferon in mammals

Interferon (IFN) is thought to play an important role in the vertebrate immune system, but systemic knowledge of IFN evolution has yet to be elucidated. To evaluate the phylogenic distribution and evolutionary history of type I IFNs, 13gen omes were searched using BLASTn program, and a phylogenetic tree of vertebrate type I IFNs was constructed. In the present study, an IFNδ-like gene in the human genome was identified, refuting the concept that humans have no IFNδ genes, and other mammalian IFN genes were also identified. In the phylogenetic tree, the mammalian IFNβ, IFN?, and IFNκ formed a clad e sepa rate f rom the other mammalian type I IFNs, while piscine and avian IFNs formed distinct clades. Based on this phylogenetic analysis and the various characteristics of type I IFNs, the evolutionary history of type I IFNs was further evaluated. Our data indicate that an ancestral IFNα-like gene forms a core from which new IFNs divided during vertebrate evolution. In addition, the data suggest how the other type I IFNs evolved from IFNα and shaped the complex type I IFN system. The promoters of type I IFNs were conserved among different mammals, as well as their genic regions. However, the intergenic regions of type I IFN clusters were not conserved among different mammals, demonstrating a high selec tion pressure upon type I IFNs during their evolution.     

Type I IFN promotes IL-10 production from T cells to suppress Th17 cells and Th17-associated autoimmune inflammation

Whereas the immune system is essential for host defense against pathogen infection or endogenous danger signals, dysregulated innate and adaptive immune cells may facilitate harmful inflammatory or autoimmune responses. In the CNS, chronic inflammation plays an important role in the pathogenesis of neurodegenerative diseases such as multiple sclerosis (MS). Our previous study has demonstrated a critical role for the type I IFN induction and signaling pathways in constraining Th17-mediated experimental autoimmune encephalomyelitis (EAE), an animal model of human MS. However, it remains unknown if self-reactive Th17 cells can be reprogrammed to have less encephalitogenic activities or even have regulatory effects through modulation of innate pathways. In this study, we investigated the direct effects of type I IFN on Th17 cells. Our data show that IFNβ treatment of T cells cultured under Th17 polarizing conditions resulted in reduced production of IL-17, but increased production of IL-10. We also found that IFNβ induced IL-10 production by antigen specific T cells derived from immunized mice. Furthermore, IFNβ treatment could suppress the encephalitogenic activity of myelin-specific T cells, and ameliorate clinical symptoms of EAE in an adoptive transfer model. Together, results from this study suggest that IFNβ may induce antigen-specific T cells to produce IL-10, which in turn negatively regulate Th17-mediate inflammatory and autoimmune response.     

Type I IFN protects against murine lupus

Both the type I (IFN-alpha beta) and type II (IFN-gamma) IFNs have been heavily implicated in the pathogenesis of systemic lupus erythematosus. To test the relative roles of these systems, congenic lupus-prone MRL/CD95(lpr/lpr) (MRL/lpr) mice lacking the type I IFN receptor (IFN-RI), type II IFN receptor (IFN-RII), or both, were derived. As expected, deficiency for IFN-RII protected MRL/lpr mice from the development of significant autoimmune-associated lymphadenopathy, autoantibodies, and renal disease. However, deficiency for the IFN-RI surprisingly worsened lymphoproliferation, autoantibody production, and end organ disease; animals doubly deficient for IFN-RI and IFN-RII developed an autoimmune phenotype intermediate between wild-type and IFN-RII-deficient animals, all correlating with an ability of type I IFN to suppress MRL B cell activation. Thus, type I IFNs protect against both the humoral and end organ autoimmune syndrome of MRL/lpr mice, independent of IFN-gamma. These findings warrant caution in the use of type I IFN antagonists in the treatment of autoimmune diseases and suggest further investigation into the interplay between the types I and II IFNs during the ontogeny of pathogenic autoantibodies.     

Role of interferons in the regulation of cell proliferation,differentiation, and development

There now appears to be evidence to support the view that the type I IFNs are naturally produced negative regulators of growth that also modify cell differentiation. Consistent with this, it appears that the ability to produce and respond to IFN is suppressed in early embryonic development when cell proliferation and differentiation are essential. In the later stages of fetal development, IFN production is de-repressed, and cells show increased sensitivity to IFN, which may be important in regulating cell proliferation and/or differentiation processes or the interaction between fetal and maternal tissues. Interestingly, the IFN system can also be suppressed in disease states such as the development of tumours or in the establishment of a (chronic) viral infection. Therefore, understanding the developmental regulation of the IFN system may be important to understanding and controlling the IFN system in disease. More extensive studies of the developmental stage and tissue-specific expression of type I IFNs and their receptors are necessary, as well as more direct in vivo experiments to further elucidate the role of the IFN system in reproduction and development. © 1994 Wiley-Liss, Inc.     

Modulation of lymphocyte proliferation and immunoglobulin synthesis by interferon-gamma and "type I" interferons

Interferon (IFN)-alpha and IFN-beta ("type I" IFNs), but not IFN-gamma reduced phytohemagglutinin- or pokeweed mitogen (PWM)-induced proliferation in cultures of human mononuclear leukocytes. Proliferation induced by specific antigens (tuberculin PPD or tetanus toxoid) or by exogenous interleukin 2 (IL-2) was strongly inhibited by type I IFNs and, to a lesser extent, by IFN-gamma as well. Inhibition of proliferation in mitogen-stimulated cultures was not due to a reduced production of IL-2 or to an inhibition of IL-2 receptor expression. Type I IFNs inhibited immunoglobulin (Ig) production in PWM-stimulated unseparated mononuclear cells, whereas IFN-gamma enhanced Ig production in such cultures. In cultures of purified B cells type I IFNs caused a stimulation of Ig production and this B-cell differentiation factor (BCDF)-like activity of IFNs was synergistically enhanced in the presence of IL-2. IFN-gamma produced less BCDF-like activity than type I IFNs. These results show that in some instances type I IFNs can be more potent in affecting functions of cells of the immune system than IFN-gamma.     

Role of type I interferons during macrophage activation by lipopolysaccharide

Activation of macrophages by bacterial lipopolysaccharide (LPS) is accompanied by the secretion of type I interferons (IFNs) which can act in an autocrine manner. We examined the role of type I IFNs in macrophage responses to LPS using bone marrow-derived macrophages (BMM) from IFNAR1-/- mice, which lack a component of the type I IFN receptor and do not respond to type I IFNs. We found that, unlike wild-type (WT) BMM, LPS-treated IFNAR1-/- cells failed to produce nitric oxide (NO), or express inducible NO synthase (iNOS), indicating that type I IFNs are essential for all LPS-stimulated NO production in BMM. Exogenously added type II IFN (IFNgamma) rescued these responses in LPS-treated IFNAR1-/- BMM. In contrast to effects on NO, type I IFNs negatively regulated respiratory burst activity in LPS-primed BMM. We also found that while type I IFNs mediated the anti-proliferative effects of lower concentrations of LPS, at higher concentrations LPS acted in a type I IFNs-independent manner. Finally, we report that type I IFNs are a survival factor for BMM. Despite this, the ability of LPS to also prevent apoptosis in BMM was independent of type I IFNs. These findings highlight the diverse roles of type I IFNs in mediating LPS-stimulated macrophage responses.     

Antiviral Responses in Mouse Embryonic Stem Cells: DIFFERENTIAL DEVELOPMENT OF CELLULAR MECHANISMS IN TYPE I INTERFERON PRODUCTION AND RESPONSE*

We have recently reported that mouse embryonic stem cells (mESCs) are deficient in expressing type I interferons (IFNs) in response to viral infection and synthetic viral RNA analogs (Wang, R., Wang, J., Paul, A. M., Acharya, D., Bai, F., Huang, F., and Guo, Y. L. (2013) J. Biol. Chem. 288, 15926–15936). Here, we report that mESCs are able to respond to type I IFNs, express IFN-stimulated genes, and mediate the antiviral effect of type I IFNs against La Crosse virus and chikungunya virus. The major signaling components in the IFN pathway are expressed in mESCs. Therefore, the basic molecular mechanisms that mediate the effects of type I IFNs are functional in mESCs; however, these mechanisms may not yet be fully developed as mESCs express lower levels of IFN-stimulated genes and display weaker antiviral activity in response to type I IFNs when compared with fibroblasts. Further analysis demonstrated that type I IFNs do not affect the stem cell state of mESCs. We conclude that mESCs are deficient in type I IFN expression, but they can respond to and mediate the cellular effects of type I IFNs. These findings represent unique and uncharacterized properties of mESCs and are important for understanding innate immunity development and ESC physiology.