Activation and Long‐Term Maintenance of Adaptive Immunity in the Central Nervous System: A Double‐Edged Sword?

Negative checkpoint regulators (NCR) are intensely pursued as targets to modulate the immune response in cancer and autoimmunity. A large variety of NCR is expressed by central nervous system (CNS)-resident cell types and is associated with CNS homeostasis, interactions with peripheral immunity and CNS inflammation and disease. Immunotherapy blocking NCR affects the CNS as patients can develop neurological issues including encephalitis and multiple sclerosis (MS). How these treatments affect the CNS is incompletely understood, since expression and function of NCR in the CNS are only beginning to be unravelled. V-type immunoglobulin-like suppressor of T cell activation (VISTA) is an NCR that is expressed primarily in the haematopoietic system by myeloid and T cells. VISTA regulates T cell quiescence and activation and has a variety of functions in myeloid cells including efferocytosis, cytokine response and chemotaxis. In the CNS, VISTA is predominantly expressed by microglia and macrophages of the CNS. In this review, we summarize the role of NCR in the CNS during health and disease. We highlight expression of VISTA across cell types and CNS diseases and discuss the function of VISTA in microglia and during CNS ageing, inflammation and neurodegeneration. Understanding the role of VISTA and other NCR in the CNS is important considering the adverse effects of immunotherapy on the CNS, and in view of their therapeutic potential in CNS disease.

In this study, we investigate the expression of fractalkine (CX3CL1) and the fractalkine receptor (CX3CR1) in the naive rat and mouse central nervous system (CNS). We determine if the expression of this chemokine and its receptor are altered during chronic or acute inflammation in the CNS. In addition, we determine if CX3CL1, which has been reported to be chemoattractant to leukocytes in vitro, is capable of acting as a chemoattractant in the CNS in vivo. Immunohistochemistry was performed using primary antibodies recognizing soluble and membrane-bound CX3CL1 and the N-terminus of the CX3CR1. We found that neurons in the naive rodent brain are immunoreactive for CX3CL1 and CX3CR1, both showing a perinuclear staining pattern. Resident microglia associated with the parenchyma and macrophages in the meninges and choroid plexus constituitively express CX3CR1. In a prion model of chronic neurodegeneration and inflammation, CX3CL1 immunoreactivity is upregulated in astrocytes and CX3CR1 expression is elevated on microglia. In surviving neurons, expression of CX3CL1 appears unaltered relative to normal neurons. There is a decrease in neuronal CX3CR1 expression. Acute inflammatory responses in the CNS, induced by stereotaxic injections of lipopolysaccharide or kainic acid, results in activation of microglia and astrocytes but no detectable changes in the glial expression of CX3CL1 or CX3CR1. The expression of CX3CL1 and CX3CR1 by glial cells during inflammation in the CNS may be influenced by the surrounding cytokine milieu, which has been shown to differ in acute and chronic neuroinflammation.

In this study, we investigate the expression of fractalkine (CX3CL1) and the fractalkine receptor (CX3CR1) in the naive rat and mouse central nervous system (CNS). We determine if the expression of this chemokine and its receptor are altered during chronic or acute inflammation in the CNS. In addition, we determine if CX3CL1, which has been reported to be chemoattractant to leukocytes in vitro, is capable of acting as a chemoattractant in the CNS in vivo. Immunohistochemistry was performed using primary antibodies recognizing soluble and membrane-bound CX3CL1 and the N-terminus of the CX3CR1. We found that neurons in the naive rodent brain are immunoreactive for CX3CL1 and CX3CR1, both showing a perinuclear staining pattern. Resident microglia associated with the parenchyma and macrophages in the meninges and choroid plexus constituitively express CX3CR1. In a prion model of chronic neurodegeneration and inflammation, CX3CL1 immunoreactivity is upregulated in astrocytes and CX3CR1 expression is elevated on microglia. In surviving neurons, expression of CX3CL1 appears unaltered relative to normal neurons. There is a decrease in neuronal CX3CR1 expression. Acute inflammatory responses in the CNS, induced by stereotaxic injections of lipopolysaccharide or kainic acid, results in activation of microglia and astrocytes but no detectable changes in the glial expression of CX3CL1 or CX3CR1. The expression of CX3CL1 and CX3CR1 by glial cells during inflammation in the CNS may be influenced by the surrounding cytokine milieu, which has been shown to differ in acute and chronic neuroinflammation. GLIA 37:314–327, 2002. © 2002 Wiley-Liss, Inc.

Neurodegeneration is a critical problem in aging populations and is characterized by severe central nervous system (CNS) inflammation. Macrophages closely regulate inflammation in the CNS and periphery by taking on different activation states. The source of inflammation in many neurodegenerative diseases has been preliminarily linked to a decrease in the CNS M2 macrophage population and a subsequent increase in M1-mediated neuroinflammation. The Recepteur D’Origine Nantais (Ron) is a receptor tyrosine kinase expressed on tissue-resident macrophages including microglia. Activation of Ron by its ligand, macrophage-stimulating protein, attenuates obesity-mediated inflammation in the periphery. An in vivo deletion of the ligand binding domain of Ron (Ron−/−) promotes inflammatory (M1) and limits a reparative (M2) macrophage activation. However, whether or not this response influences CNS inflammation has not been determined. In this study, we demonstrate that in homeostasis Ron−/− mice developed an inflammatory CNS niche with increased tissue expression of M1-associated markers when compared to age-matched wild-type (WT) mice. Baseline metabolic analysis of CNS tissue indicates exacerbated levels of metabolic stress in Ron−/− CNS. In a disease model of multiple sclerosis, experimental autoimmune encephalomyelitis, Ron−/− mice exhibit higher disease severity when compared to WT mice associated with increased CNS tissue inflammation. In a model of diet-induced obesity (DIO), Ron−/− mice exhibit exacerbated CNS inflammation with decreased expression of the M2 marker Arginase-1 (Arg-1) and a robust increase in M1 markers compared to WT mice following 27 weeks of DIO. Collectively, these results illustrate that activation of Ron in the CNS could be a potential therapeutic approach to treating various grades of CNS inflammation underlying neurodegeneration.

Central nervous system (CNS) inflammation is a common cause of neurological dysfunction in dogs. Most dogs with CNS inflammation are diagnosed with presumptive autoimmune disease. A smaller number are diagnosed with an infectious etiology. Additionally, at necropsy, a subset of dogs with CNS inflammation do not fit previously described patterns of autoimmune disease and an infectious cause is not readily identifiable. Because viral infection is a common cause of meningoencephalitis in people, we hypothesize that a subset of dogs presented with CNS inflammation have an occult viral infection either as a direct cause of CNS inflammation or a trigger for autoimmunity. The goal of this research was to screen cerebrospinal fluid from a large number dogs with CNS inflammation for occult viral infection. One hundred seventy-two dogs with neurological dysfunction and cerebrospinal fluid (CSF) pleocytosis were identified. Of these, 42 had meningoencephalitis of unknown origin, six had steroid-responsive meningitis-arteritis, one had eosinophilic meningoencephalitis, five had documented infection, 21 had and undetermined diagnosis, and 97 had a diagnosis not consistent with primary inflammatory disease of the CNS (e.g., neoplasia). CSF samples were subsequently screened with broadly reactive PCR for eight viral groups: adenovirus, bunyavirus, coronavirus, enterovirus, flavivirus, herpesvirus, paramyxovirus, and parechovirus. No viral nucleic acids were detected from 168 cases screened for eight viral groups, which does not support occult viral infection as a cause of CNS inflammation in dogs. La Crosse virus (LACV) nucleic acids were detected from four cases in Georgia. Subclinical infection was supported in two of these cases but LACV could not be ruled-out as a cause of infection in the other two cases, suggesting further research is warranted to determine if LACV is an occult cause of CNS inflammation in dogs.

On January 18, 2002, Elan Corporation and American Home Products reported their decision to temporarily suspend dosing in a phase 2a study of their experimental antiAlzheimer’s disease vaccine (AN-1792) after 4 patients showed clinical signs of central nervous system (CNS) inflammation. AN-1792, also known as AIP-001, is a 42amino acid peptide ( -amyloid) vaccine coupled with an immune adjuvant (QS-21), a purified saponin derivative. This double-blind, placebo-controlled study is being conducted in the United States and four European countries and was designed to measure the immune response to AN-1792 in patients with mild to moderate Alzheimer’s disease. To date, approximately 360 patients have received multiple doses of AN-1792. The presence of a virus within the cerebrospinal fluid was reported in 1 of the 4 patients with CNS inflammation under investigation. However, the cause of inflammation remains to be determined. The 4 reported cases occurred in France, where 97 patients have received the study drug. Dramatic results obtained with AN-1792 in transgenic animal models of Alzheimer’s disease raised unprecedented hope for an effective treatment of this devastating disorder. The vaccine increases -amyloid clearance through mechanisms not fully understood. Centrally, the vaccine appears to activate -amyloid phagocytosis by microglial monocytes. Peripherally, serum antibodies appear to bind and sequester -amyloid, thereby altering its equilibrium between the CNS and plasma. There are many agents and mechanisms that can result in CNS inflammation. One explanation for the encephalitis could be external contamination during lumbar punctures, which are required as part of the protocol. It is unlikely that the QS-21 adjuvant used in this trial is the source of the viruses, as it is not of animal origin but is a highly purified saponin derived from the bark of the Quillaja saponaria Molina tree. Viral, bacterial, fungal, and parasitic pathogens may breach the blood–brain barrier and enter the CNS through paracellular or transcellular mechanisms. Recent studies appear to indicate that -amyloid vaccination may alter the permeability of the blood–brain barrier. In the mouse, the complex formed by -amyloid and immunoglobulins appears to have a 3.9to 4.6-fold greater permeability coefficient than nonspecific monoclonal antibodies. It has been also reported that in transgenic mice the passively administered antibodies can cross the blood–brain barrier to act directly in the CNS. This suggests that -amyloid immunization may cause abnormal leakage of the blood–brain barrier and increase the chance that pathogenic microorganisms will reach the CNS. It is also possible and perhaps more likely that the adverse events of AN-1792 observed in patients with Alzheimer’s disease could reflect an autoimmune inflammatory response triggered by T-lymphocyte activation after immune-system stimulation with -amyloid. T-cell lines specific to -amyloid contain a high percentage of CD8-positive cytotoxic T cells, which are capable of lysing cells that overproduce -amyloid. It is of paramount importance to clarify the nature and mechanism of the observed adverse effects on the CNS associated with AN-1792. As other therapeutic anti-amyloid vaccination strategies are being pursued, we urgently need to understand if the observed toxic effects are specific to AN1792 or also occur with other immunization approaches.

Neurodegeneration is characterized by severe central nervous system (CNS) inflammation. Macrophages are immune cells that closely regulate inflammation by taking on different activation states. The Ron receptor tyrosine kinase is expressed on tissue resident macrophages including microglia. An in vivo deletion of Ron (Ron KO) promotes inflammatory (M1) and limits reparative (M2) macrophage activation. Whether this response influences CNS tissue inflammation is not characterized. The objective of this study was to elucidate whether Ron expression plays a critical role in CNS inflammation in homeostasis and subsequently in disease models. During homeostasis with no interventions, Ron KO mice developed an inflammatory CNS niche with increased tissue expression of M1-associated markers TNFα, Cox-2 and iNOS when compared to WT-mice (P<0.05). In a diet induced obesity(DIO) model of chronic inflammation, Ron KO mice exhibit exacerbated CNS inflammation with decreased expression of M2(Arg-1) and a robust increase in M1 markers compared to WT-mice following 27 weeks of high-fat-diet intervention(P<0.05). Lastly, we evaluated the role of Ron in a disease model of Multiple Sclerosis, experimental autoimmune encephalitis (EAE). Ron KO-EAE mice exhibited higher disease severity from peak disease state and onward when compared to WT-EAE mice (P<0.05). Ron KO mice had greater CNS expression of M1 markers relative to higher peripheral inflammation with increased IFNγ secretion from lymphoid organs during peak disease state(P<0.05). Collectively these results illustrate how maintenance of Ron expression in the CNS could then be a potential therapeutic approach to treating various grades of inflammation underlying neurodegeneration.

There is growing evidence that long-term central nervous system (CNS) inflammation exacerbates secondary deterioration of brain structures and functions and is one of the major determinants of disease outcome and progression. In acute CNS injury, brain microglia are among the first cells to respond and play a critical role in neural repair and regeneration. However, microglial activation can also impede CNS repair and amplify tissue damage, and phenotypic transformation may be responsible for this dual role. Mesenchymal stem cell (MSC)-derived exosomes (Exos) are promising therapeutic agents for the treatment of acute CNS injuries due to their immunomodulatory and regenerative properties. MSC-Exos are nanoscale membrane vesicles that are actively released by cells and are used clinically as circulating biomarkers for disease diagnosis and prognosis. MSC-Exos can be neuroprotective in several acute CNS models, including for stroke and traumatic brain injury, showing great clinical potential. This review summarized the classification of acute CNS injury disorders and discussed the prominent role of microglial activation in acute CNS inflammation and the specific role of MSC-Exos in regulating pro-inflammatory microglia in neuroinflammatory repair following acute CNS injury. Finally, this review explored the potential mechanisms and factors associated with MSC-Exos in modulating the phenotypic balance of microglia, focusing on the interplay between CNS inflammation, the brain, and injury aspects, with an emphasis on potential strategies and therapeutic interventions for improving functional recovery from early CNS inflammation caused by acute CNS injury.

During the era of highly active antiretroviral therapy (HAART), the prevalence of HIV-associated central nervous system (CNS) disease has increased despite suppression of plasma viremia. In a simian immunodeficiency virus (SIV) model system in which all animals develop AIDS and 90% develop CNS disease by 3 months after inoculation, pigtailed macaques were treated with a regimen of tenofovir disoproxil fumarate, saquinavir, atazanavir, and an integrase inhibitor starting at 12 days after inoculation and were euthanized at approximately 175 days after inoculation. Plasma and cerebrospinal fluid (CSF) viral loads declined rapidly after the initiation of HAART. Brain viral RNA was undetectable at necropsy, but viral DNA levels were not different from those in untreated SIV-infected macaques. CNS inflammation was significantly reduced, with decreased brain expression of major histocompatibility complex class II and glial fibrillary acidic protein and reduced levels of CSF CCL2 and interleukin 6. Brain from treated macaques had significantly lower levels of interferon beta, type 1 interferon-inducible gene myxovirus (influenza) resistance A, and indolamine 2,3-dioxygenase messenger RNA, suggesting that immune hyperactivation was suppressed, and fewer CD4(+) and CD8(+) T cells, suggesting that trafficking of T cells from peripheral blood was reduced. Brain levels of CD68 protein and tumor necrosis factor alpha and interferon gamma RNA were reduced but were not significantly lower, indicating continued CNS inflammation. These data, generated in a rigorous, high-viral-load SIV-infected macaque model, showed that HAART provided benefits with respect to CNS viral replication and inflammation but that no change in the level of viral DNA and continued CNS inflammation occurred in some macaques.

T cell lymphocytes play an essential role in the adaptive immunity. They arise from the hematopoietic compartment and reach the thymus, where they achieve their development through specialized maturation and selection steps. Immune system comprises specific and complex mechanisms in the thymus and the periphery aiming at preventing the generation and the survival of self-reactive T cells that could lead to autoimmune disorders. These mechanisms are known as central and peripheral tolerance and rely on different cellular actors such as medullary thymic epithelial cells (mTECs), dendritic cells (DCs), lymph node stromal cells (LNSCs) and regulatory T and B cells. Self-reactive T cells escaping thymic central tolerance are kept in check in the periphery through specific mechanisms that include T cell anergy, T cell deletion and Treg induction. Although conventional DCs (cDCs) have been first in line in mediating peripheral tolerance, increasing evidence demonstrates the contribution of other actors such as plasmacytoid dendritic cells (pDCs) in this process. pDCs are important linkers of the innate and adaptive immunity. They are characterized by their ability to secrete pro-inflammatory cytokines and large amount of type I interferon (IFN-I) upon pathogenic infections, but also express major Histocompatibility complex (MHC) and costimulatory molecules enabling them to interact with T cells. pDCs have been involved in the control of infections, but are also important actors of peripheral tolerance. The diversity of their functions, depending on the context in which they will evolve, has associated pDCs to pro-immunogenic and tolerogenic scopes. In the lab, we are principally interested in the contribution of pDCs in Multiple sclerosis, a progressive inflammatory demyelinating disease of the central nervous system (CNS). To address the role of pDCs in MS, we performed our studies in the murine model of Experimental autoimmune encephalomyelitis (EAE). Using genetically modified mice harbouring specific abrogation of Ag-presenting function in pDCs, my co-workers previously demonstrated that myelin Ag presentation by pDCs promotes the expansion of regulatory T cells (Tregs) that inhibit encephalitogenic TH1 and TH17 cell priming in secondary lymphoid organs (SLOs). In this manuscript I report our recent findings supporting that pDCs display tolerogenic functions during the priming and the effector phase of EAE development and are able to control and dampen the disease. The first study investigates the interplays between pDCs and Tregs and shows that Ag-specific MHCII interactions with Tregs licenced tolerogenic features in steady-state pDCs by inducing their expression of the indoleamine-2,3 dioxygenase (IDO1). In EAE context, Treg-educated IDO+ pDCs are required to confer suppressive functions to Tregs which promote the inhibition of encephalitogenic T cell priming in draining LNs, resulting in attenuated EAE. In the second study, we explore the therapeutic effect of pDCs transfer in EAE mice after disease onset. We show that the transfer of immature MOG35-55 pre-loaded pDCs during EAE acute phase leads to substantial reduction of CNS inflammation and significant amelioration of disease clinical scores. We demonstrate that pDC-protection relies on the massive recruitment of endogenous immature pDCs in the inflamed spinal cord via the Chemerine/CMKLR1 axis. Endogenous pDC recruitment is required to down-modulate CNS inflammation, encephalitogenic TH1 and TH17 cell responses and EAE severity. Overall this work supports previous findings showing the importance of pDCs in the regulation of CNS autoimmunity and unravel these cells for potential use, by targeting different particular functions, in the development of future therapies to treat MS patients.

The present review discusses current concepts of HIV-associated neurocognitive disorders (HAND) in the era of antiretroviral therapy (ART). As the HIV epidemic enters its fourth decade (the second decade of ART), research must address evolving factors in HAND pathogenesis. These include persistent systemic and central nervous system (CNS) inflammation, aging in the HIV-infected brain, HIV subtype (clade) distribution, concomitant use of drugs of abuse, and potential neurotoxicity of ART drugs. Although the severest form of HAND, HIV-associated dementia (HAD), is now rare due to ART, the persistence of milder, functionally important HAND forms persist in up to half of HIV-infected individuals. HAND prevalence may be higher in areas of Africa where different HIV subtypes predominate, and ART regimens that are more effective in suppressing CNS HIV replication can improve neurological outcomes. HAND are correlated with persistent systemic and CNS inflammation, and enhanced neuronal injury due to stimulant abuse (cocaine and methamphetamine), aging, and possibly ART drugs themselves. Prevention and treatment of HAND requires strategies aimed at suppressing CNS HIV replication and effects of systemic and CNS inflammation in aging and substance-abusing HIV populations. Use of improved CNS-penetrating ART must be accompanied by evaluation of potential ART neurotoxicity.

Central nervous system (CNS) infection and inflammation severely reduce the capacity of cytochrome P-450 metabolism in the liver. We developed a mouse model to examine the effects of CNS inflammation on hepatic cytochrome P-450 metabolism. FVB, C57BL/6, and C3H/HeouJ mice were given Escherichia coli LPS (2.5 microg) by intracerebroventricular (ICV) injection. The CNS inflammatory response was confirmed by the elevation of TNF-alpha and/or IL-1beta proteins in the brain. In all mouse strains, LPS produced a 60-70% loss in hepatic Cyp3a11 expression and activity compared with saline-injected controls. Adrenalectomy did not prevent the loss in Cyp3a11 expression or activity, thereby precluding the involvement of the hypothalamic-adrenal-pituitary axis. Endotoxin was detectable (1-10 ng/ml) in serum between 15 and 120 min after ICV dosing of 2.5 microg LPS. Peripheral administration of 2.5 microg LPS by intraperitoneal injection produced similar serum endotoxin levels and a similar loss (60%) in Cyp3a11 expression and activity in the liver. The loss of Cyp3a11 in response to centrally or peripherally administered LPS could not be evoked in Toll-like receptor-4 (TLR4)-mutant (C3H/HeJ) mice, indicating that TLR4 signaling pathways are directly involved in the enzyme loss. In summary, we conclude that LPS is transferred from the brain to the circulation in significant quantities in a model of CNS infection or inflammation. Subsequently, LPS that has reached the circulation stimulates a TLR4-dependent mechanism in the periphery, evoking a reduction in Cyp3a11 expression and metabolism in the liver.

BackgroundTwo-photon laser scanning microscopy (TPLSM) has become a powerful tool in the visualization of immune cell dynamics and cellular communication within the complex biological networks of the inflamed central nervous system (CNS). Whereas many previous studies mainly focused on the role of effector or effector memory T cells, the role of naïve T cells as possible key players in immune regulation directly in the CNS is still highly debated.MethodsWe applied ex vivo and intravital TPLSM to investigate migratory pathways of naïve T cells in the inflamed and non-inflamed CNS. MACS-sorted naïve CD4+ T cells were either applied on healthy CNS slices or intravenously injected into RAG1 -/- mice, which were affected by experimental autoimmune encephalomyelitis (EAE). We further checked for the generation of second harmonic generation (SHG) signals produced by extracellular matrix (ECM) structures.ResultsBy applying TPLSM on living brain slices we could show that the migratory capacity of activated CD4+ T cells is not strongly influenced by antigen specificity and is independent of regulatory or effector T cell phenotype. Naïve T cells, however, cannot find sufficient migratory signals in healthy, non-inflamed CNS parenchyma since they only showed stationary behaviour in this context. This is in contrast to the high motility of naïve CD4+ T cells in lymphoid organs. We observed a highly motile migration pattern for naïve T cells as compared to effector CD4+ T cells in inflamed brain tissue of living EAE-affected mice. Interestingly, in the inflamed CNS we could detect reticular structures by their SHG signal which partially co-localises with naïve CD4+ T cell tracks.ConclusionsThe activation status rather than antigen specificity or regulatory phenotype is the central requirement for CD4+ T cell migration within healthy CNS tissue. However, under inflammatory conditions naïve CD4+ T cells can get access to CNS parenchyma and partially migrate along inflammation-induced extracellular SHG structures, which are similar to those seen in lymphoid organs. These SHG structures apparently provide essential migratory signals for naïve CD4+ T cells within the diseased CNS.

The role of mast cells in virus-induced inflammation in the murine central nervous system

Microglial NLRP3 inflammasome activation mediates IL-1β-related inflammation in prefrontal cortex of depressive rats.