Boven, L.A.

Myelin ingestion alters macrophage antigen-presenting function in vitro and in vivo (Article)

2011-07-01T00:00:00Z

During MS, phagocytosing myelin-containing macrophages arise and lie in close proximity to T cells. To date, it has not been addressed whether these myelinladen macrophages have the capacity to present antigens to T cells and whether this contributes to inflammation in disease. We demonstrate that in vitro-generated human and mouse myelin-laden macrophages expressed MHC class I and II and costimulatory molecules and are thus well equipped for antigen presentation.uman myelin-laden macrophages exhibited normal endocytosis of particulate and soluble antigens. In addition, human myelin-laden macrophages elicited active T cell proliferation of nai{dotless}̈ve as well as memory T cells. Furthermore, mouse myelin-laden macrophages induced primary antigen-specific CD4+T cell proliferation in vivo but transiently diminished IFN-γ release. Functionally, MOG peptide-loaded myelin-laden mouse macrophages modestly but significantly reduced the severity of MOG peptide-induced EAE. These data show that myelin uptake results in the induction of a population of macrophages that retains antigen-presenting capacity and limits autoimmune-mediated disease.

Highly pathogenic avian influenza virus H5N1 infects alveolar macrophages without virus production or excessive TNF-alpha induction (Article)

2011-06-01T00:00:00Z

Highly pathogenic avian influenza virus (HPAIV) of the subtype H5N1 causes severe, often fatal pneumonia in humans. The pathogenesis of HPAIV H5N1 infection is not completely understood, although the alveolar macrophage (AM) is thought to play an important role. HPAIV H5N1 infection of macrophages cultured from monocytes leads to high percentages of infection accompanied by virus production and an excessive pro-inflammatory immune response. However, macrophages cultured from monocytes are different from AM, both in phenotype and in response to seasonal influenza virus infection. Consequently, it remains unclear whether the results of studies with macrophages cultured from monocytes are valid for AM. Therefore we infected AM and for comparison macrophages cultured from monocytes with seasonal H3N2 virus, HPAIV H5N1 or pandemic H1N1 virus, and determined the percentage of cells infected, virus production and induction of TNF-alpha, a pro-inflammatory cytokine. In vitro HPAIV H5N1 infection of AM compared to that of macrophages cultured from monocytes resulted in a lower percentage of infected cells (up to 25% vs up to 84%), lower virus production and lower TNF-alpha induction. In vitro infection of AM with H3N2 or H1N1 virus resulted in even lower percentages of infected cells (up to 7%) than with HPAIV H5N1, while virus production and TNF-alpha induction were comparable. In conclusion, this study reveals that macrophages cultured from monocytes are not a good model to study the interaction between AM and these influenza virus strains. Furthermore, the interaction between HPAIV H5N1 and AM could contribute to the pathogenicity of this virus in humans, due to the relative high percentage of infected cells rather than virus production or an excessive TNF-alpha induction.

Myelin ingestion by macrophages promotes their motility and capacity to recruit myeloid cells (Article)

2010-08-01T00:00:00Z

Myelin-laden macrophages reside within the CNS, the CSF and in the CNS-draining lymph nodes during MS and EAE, suggesting migration of these macrophages between these compartments and interaction with other cells. Since chemokines and their receptors are pivotal for leukocyte trafficking, we addressed whether myelin ingestion affects chemotaxis of mouse macrophages in vitro. Myelin ingestion enhanced expression of CCR7 and CXCR3 on macrophages and migration towards CCL21 and CXCL10. Furthermore, myelin-laden macrophages released chemoattractants resulting in enhanced migration of myeloid cells in vitro. Our data demonstrate that myelin-laden macrophages have increased motility and suggest trafficking between anatomical compartments in vivo. © 2010 Elsevier B.V.

Differential expression of adenosine A3 receptors controls adenosine A2A receptor-mediated inhibition of TLR responses in microglia (Article)

2009-06-15T00:00:00Z

Microglia activation is a prominent feature in many neuroinflammatory disorders. Unrestrained activation can generate a chronic inflammatory environment that might lead to neurodegeneration and autoimmunity. Extracellular adenosine modulates cellular activation through adenosine receptor (ADORA)-mediated signaling. There are four ADORA subtypes that can either increase (A2Aand A2Breceptors) or decrease (A1and A3receptors) intracellular cyclic AMP levels. The expression pattern of the subtypes thus orchestrates the cellular response to extracellular adenosine. We have investigated the expression of ADORA subtypes in unstimulated and TLR-activated primary rhesus monkey microglia. Activation induced an up-regulation of A2Aand a down-regulation of A3receptor (A3R) levels. The altered ADORA-expression pattern sensitized microglia to A2Areceptor (A2AR)- mediated inhibition of subsequent TLR-induced cytokine responses. By using combinations of subtype-specific agonists and antagonists, we revealed that in unstimulated microglia, A2AR-mediated inhibitory signaling was effectively counteracted by A3R-mediated signaling. In activated microglia, the decrease in A3R-mediated signaling sensitized them to A2AR-mediated inhibitory signaling. We report a differential, activation state-specific expression of ADORA in microglia and uncover a role for A3R as dynamically regulated suppressors of A2AR- mediated inhibition of TLR-induced responses. This would suggest exploration of combinations of A2AR agonists and A3R antagonists to dampen microglial activation during chronic neuroinflammatory conditions. Copyright

Surgical excision of CNS-draining lymph nodes reduces relapse severity in chronic-relapsing experimental autoimmune encephalomyelitis (Article)

2009-03-01T00:00:00Z

Despite lack of classical lymphatic vessels in the central nervous system (CNS), cells and antigens do reach the CNS-draining lymph nodes. These lymph nodes are specialized to mediate mucosal immune tolerance, but can also generate T- and B-cell immunity. Their role in multiple sclerosis and experimental autoimmune encephalomyelitis (EAE) therefore remains elusive. We hypothesized that drainage of CNS antigens to the CNS-draining lymph nodes is vital for the recurrent episodes of CNS inflammation. To test this, we surgically removed the superficial cervical lymph nodes, deep cervical lymph nodes, and the lumbar lymph nodes prior to disease induction in three mouse EAE models, representing acute, chronic, and chronic-relapsing EAE. Excision of the CNS-draining lymph nodes in chronic-relapsing EAE reduced and delayed the relapse burden and EAE pathology within the spinal cord, which suggests initiation of CNS antigen-specific immune responses within the CNS-draining lymph nodes. Indeed, superficial cervical lymph nodes from EAE-affected mice demonstrated proliferation against the immunizing peptide, and the deep cervical lymph nodes, lumbar lymph nodes, and spleen demonstrated additional proliferation against other myelin antigen epitopes. This indicates that intermolecular epitope spreading occurs and that CNS antigen-specific immune responses are differentially generated within the different CNS-draining lymphoid organs. Proliferation of splenocytes from lymphadenectomized and sham-operated mice against the immunizing peptide was similar. These data suggest a role for CNS-draining lymph nodes in the induction of detrimental immune responses in EAE relapses, and conclusively demonstrate that the tolerance-inducing capability of cervical lymph nodes is not involved in EAE. Copyright

Brain antigens in functionally distinct antigen-presenting cell populations in cervical lymph nodes in MS and EAE (Article)

2009-03-01T00:00:00Z

Drainage of central nervous system (CNS) antigens to the brain-draining cervical lymph nodes (CLN) is likely crucial in the initiation and control of autoimmune responses during multiple sclerosis (MS). We demonstrate neuronal antigens within CLN of MS patients. In monkeys and mice with experimental autoimmune encephalomyelitis (EAE) and in mouse models with non-inflammatory CNS damage, the type and extent of CNS damage was associated with the frequencies of CNS antigens within the cervical lymph nodes. In addition, CNS antigens drained to the spinal-cord-draining lumbar lymph nodes. In human MS CLN, neuronal antigens were present in pro-inflammatory antigen-presenting cells (APC), whereas the majority of myelin-containing cells were anti-inflammatory. This may reflect a different origin of the cells or different drainage mechanisms. Indeed, neuronal antigen-containing cells in human CLN did not express the lymph node homing receptor CCR7, whereas myelin antigen-containing cells in situ and in vitro did. Nevertheless, CLN from EAE-affected CCR7-deficient mice contained equal amounts of myelin and neuronal antigens as wild-type mice. We conclude that the type and frequencies of CNS antigens within the CLN are determined by the type and extent of CNS damage. Furthermore, the presence of myelin and neuronal antigens in functionally distinct APC populations within MS CLN suggests that differential immune responses can be evoked.

Differentiation of primary adult microglia alters their response to TLR8-mediated activation but not their capacity as APC (Article)

2007-11-15T00:00:00Z

Activated microglia are found in a variety of neuroinflammatory disorders where they have attributed roles as effector as well as antigen-presenting cells (APC). Critical determinants for the multifaceted role of microglia are the differentiation potential of microglia and their mode of activation. In this study, we have investigated the effects of M-CSF and GM-CSF-mediated differentiation of adult primate microglia on their cellular phenotype, antigen presentation, and phagocytic function as well as on Toll-like receptor (TLR)-mediated responses. We show that although cell morphology and expression levels of activation markers were markedly different, differentiation with either factor yielded microglia that phenotypically and functionally resemble macrophages. Both M-CSF and GM-CSF-differentiated microglia were responsive to TLR1/2, 2, 3, 4, 5, 6/2, and 8-mediated activation, but not to TLR7 or 9-mediated activation. Intriguingly, M-CSF-differentiated microglia expressed higher levels of TLR8-encoding mRNA and protein, and produced larger amounts of proinflammatory cytokines in response to TLR8-mediated activation as compared to GM-CSF-differentiated microglia. While differentiation of adult microglia by growth factors that can be produced endogenously in the central nervous system is thus unlikely to change their APC function, it can alter their innate responses to infectious stimuli such as ssRNA viruses. Resident primate microglia may thereby help shape rather than initiate adaptive immune responses.

Brain-derived human immunodeficiency virus-1 Tat exerts differential effects on LTR transactivation and neuroimmune activation (Article)

2007-03-01T00:00:00Z

Molecular diversity within brain-derived HIV-1 sequences is highly variable depending on the individual gene examined and the neurological status of the patient. Herein, we examined different brain-derived human immunodeficiency virus (HIV)-1 tat sequences in terms of their effects on LTR transactivation and host gene induction in neural cells. Astrocytic and monocytoid cells co-transfected with prototypic tat clones derived from non-demented (ND) (n = 3) and demented (HAD) (n = 3) AIDS patients and different HIV-LTR constructs revealed that LTR transactivation mediated by tat clones derived from HAD patients was decreased (p < 0.05). A Tat-derived peptide containing the amino acid 24-38 domain from a ND clone caused down-regulation of the LTR transactivation (p < 0.05) in contrast to peptides from other Tat regions derived from HAD and ND tat clones. Both brain-derived HAD and ND tat constructs were able to induce the host immune genes, MCP-1 and IL-1β. Microarray analysis revealed several host genes were selectively upregulated by a HAD-derived tat clone including an enzyme mediating heparan sulphate synthesis, HS3ST3B1 (p < 0.05), which was also found to be increased in the brains of patients with HAD. Expression of the pro-apoptotic gene, PDCD7, was reduced in cells transfected with the HAD-derived tat clone and moreover, this gene was also suppressed in monocytoid cells infected with a neurotropic HIV-1 strain. Thus, mutations within the HIV-1 tat gene may exert pathogenic effects contributing to the development of HAD, which are independent of its effects on LTR transactivation.

The human CMV-UL86 peptide 981-1003 shares a crossreactive T-cell epitope with the encephalitogenic MOG peptide 34-56, but lacks the capacity to induce EAE in rhesus monkeys (Article)

2007-01-01T00:00:00Z

Rhesus monkeys immunized with MOG34-56, a dominant T-cell epitope from myelin/oligodendrocyte glycoprotein, develop an acute neurological disease resembling acute disseminated encephalomyelitis (ADEM) in humans. The typical large demyelinated lesions and mononuclear infiltrates in the monkey brains are caused by MOG34-56T-cells. We show that MOG34-56-reactive CD4+ and CD8+ T-cells are induced in monkeys immunized with a peptide from the human CMV major capsid protein (UL86; 981-1003), that shares sequence similarity with MOG34-56. Monkeys sensitized against the viral peptide and subsequently challenged with MOG34-56display histological signs of encephalitis, but do not show overt neurological signs.

Proinflammatory bacterial peptidoglycan as a cofactor for the development of central nervous system autoimmune disease (Article)

2005-01-01T00:00:00Z

Upon stimulation by microbial products through TLR, dendritic cells (DC) acquire the capacity to prime naive T cells and to initiate a proinflammatory immune response. Recently, we have shown that APC within the CNS of multiple sclerosis (MS) patients contain peptidoglycan (PGN), a major cell wall component of Gram-positive bacteria, which signals through TLR and NOD. In this study, we report that Staphylococcus aureus PGN as a single component can support the induction of experimental autoimmune encephalomyelitis (EAE) in mice, an animal model for MS. Mice immunized with an encephalitogenic myelin oligodendrocyte glycoprotein peptide in IFA did not develop EAE. In contrast, addition of PGN to the emulsion was sufficient for priming of autoreactive Th1 cells and development of EAE. In vitro studies demonstrate that PGN stimulates DC-mediated processes, reflected by increased Ag uptake, DC maturation, Th1 cell expansion, activation, and proinflammatory cytokine production. These data indicate that PGN-mediated interactions result in proinflammatory stimulation of Ag-specific effector functions, which are important in the development of EAE. These PGN-mediated processes may occur both within the peripheral lymph nodes as well as in the CNS and likely involve recognition by TLR on DC. Thus, PGN may provide a physiological trigger of DC maturation, and in this way disrupt the normal tolerance to self Ag. As such, PGN signaling pathways may serve as novel targets for the treatment of MS.

Transfer of central nervous system autoantigens and presentation in secondary lymphoid organs (Article)

2002-01-01T00:00:00Z

Dendritic cells are thought to regulate tolerance induction vs immunization by transferring Ags and peripheral signals to draining lymph nodes (LN). However, whether myelin Ag transfer and presentation in LN occurs during demyelinating brain disease is unknown. In this study, we demonstrate redistribution of autoantigens from brain lesions to cervical LN in monkey experimental autoimmune encephalomyelitis (EAE) and in multiple sclerosis (MS). Immunohistochemical analysis revealed significantly more cells containing myelin Ags in cervical LN of monkeys with EAE compared with those of healthy control monkeys. Myelin Ags were observed in cells expressing dendritic cell/macrophage-specific markers, MHC class II, and costimulatory molecules. Moreover, these cells were directly juxtaposed to T cells, suggesting that cognate interactions between myelin-containing APC and T cells are taking place in brain-draining LN. Indeed, myelin Ag-reactive T cells were observed in cervical LN from marmosets and rhesus monkeys. Importantly, these findings were paralleled by our findings in human tissue. We observed significantly more myelin Ag-containing cells in LN of individuals with MS compared with those of control individuals. These cells expressed APC markers, as observed in marmosets and rhesus monkeys. These findings suggest that during MS and EAE, modulation of T cell reactivity against brain-derived Ags also takes place in cervical LN and not necessarily inside the brain. A major implication is that novel therapeutic strategies may be targeted to peripheral events, thereby circumventing the blood-brain barrier.