Comparison of microglia and infiltrating CD11c⁺ cells as antigen presenting cells for T cell proliferation and cytokine response.

BACKGROUND: Tissue-resident antigen-presenting cells (APC) exert a major influence on the local immune environment. Microglia are resident myeloid cells in the central nervous system (CNS), deriving from early post-embryonic precursors, distinct from adult hematopoietic lineages. Dendritic cells (DC) and macrophages infiltrate the CNS during experimental autoimmune encephalomyelitis (EAE). Microglia are not considered to be as effective APC as DC or macrophages. METHODS: In this work we compared the antigen presenting capacity of CD11c⁺ and CD11c⁻ microglia subsets with infiltrating CD11c⁺ APC, which include DC. The microglial subpopulations (CD11c⁻ CD45dim CD11b⁺ and CD11c⁺ CD45dim CD11b⁺) as well as infiltrating CD11c⁺ CD45high cells were sorted from CNS of C57BL/6 mice with EAE. Sorted cells were characterised by flow cytometry for surface phenotype and by quantitative real-time PCR for cytokine expression. They were co-cultured with primed T cells to measure induction of T cell proliferation and cytokine response. RESULTS: The number of CD11c⁺ microglia cells increased dramatically in EAE. They expressed equivalent levels of major histocompatibility complex and co-stimulatory ligands CD80 and CD86 as those expressed by CD11c⁺ cells infiltrating from blood. CD11c⁺ microglia differed significantly from blood-derived CD11c⁺ cells in their cytokine profile, expressing no detectable IL-6, IL-12 or IL-23, and low levels of IL-1ß. By contrast, CD11c⁻ microglia expressed low but detectable levels of all these cytokines. Transforming growth factor ß expression was similar in all three populations. Although CNS-resident and blood-derived CD11c⁺ cells showed equivalent ability to induce proliferation of myelin oligodendrocyte glycoprotein-immunised CD4⁺ T cells, CD11c⁺ microglia induced lower levels of T helper (Th)1 and Th17 cytokines, and did not induce Th2 cytokines. CONCLUSIONS: Our findings show distinct subtypes of APC in the inflamed CNS, with a hierarchy of functional competence for induction of CD4⁺ T cell responses.

Inflammation in the central nervous system and Th17 responses are inhibited by IFN-gamma-Induced IL-18 binding protein.

Inflammatory responses are essential for immune protection but may also cause pathology and must be regulated. Both Th1 and Th17 cells are implicated in the pathogenesis of autoimmune inflammatory diseases, such as multiple sclerosis. We show in this study that IL-18-binding protein (IL-18bp), the endogenous inhibitor of the Th1-promoting cytokine IL-18, is upregulated by IFN-gamma in resident microglial cells in the CNS during multiple sclerosis-like disease in mice. Test of function by overexpression of IL-18bp in the CNS using a viral vector led to marked reduction in Th17 responses and robust inhibition of incidence, severity, and histopathology of disease, independently of IFN-gamma. The disease-limiting action of IL-18bp included suppression of APC-derived Th17-polarizing cytokines. IL-18bp thus acts as a sensor for IFN-gamma and can regulate both Th1 and Th17 responses in the CNS.

Interferon regulatory factor-7 modulates experimental autoimmune encephalomyelitis in mice.

BACKGROUND: Multiple sclerosis (MS) is an inflammatory disease of the central nervous system (CNS) with unknown etiology. Interferon-ß (IFN-ß), a member of the type I IFN family, is used as a therapeutic for MS and the IFN signaling pathway is implicated in MS susceptibility. Interferon regulatory factor 7 (IRF7) is critical for the induction and positive feedback regulation of type I IFN. To establish whether and how endogenous type I IFN signaling contributes to disease modulation and to better understand the underlying mechanism, we examined the role of IRF7 in the development of MS-like disease in mice. METHODS: The role of IRF7 in development of EAE was studied by immunizing IRF7-KO and C57BL/6 (WT) mice with myelin oligodendrocyte glycoprotein using a standard protocol for the induction of EAE. We measured leukocyte infiltration and localization in the CNS using flow cytometric analysis and immunohistochemical procedures. We determined levels of CD3 and selected chemokine and cytokine gene expression by quantitative real-time PCR. RESULTS: IRF7 gene expression increased in the CNS as disease progressed. IRF7 message was localized to microglia and infiltrating leukocytes. Furthermore, IRF7-deficient mice developed more severe disease. Flow cytometric analysis showed that the extent of leukocyte infiltration into the CNS was higher in IRF7-deficient mice with significantly higher number of infiltrating macrophages and T cells, and the distribution of infiltrates within the spinal cord was altered. Analysis of cytokine and chemokine gene expression by quantitative real-time PCR showed significantly greater increases in CCL2, CXCL10, IL-1ß and IL17 gene expression in IRF7-deficient mice compared with WT mice. CONCLUSION: Together, our findings suggest that IRF7 signaling is critical for regulation of inflammatory responses in the CNS.

Spontaneous complement activation on human B cells results in localized membrane depolarization and the clustering of complement receptor type 2 and C3 fragments.

While our previous studies have demonstrated that complement activation induced by complement receptors type 2 (CR2/CD21) and 1 (CR1/CD35) results in C3-fragment deposition and membrane attack complex (MAC) formation in human B cells, the consequences of these events for B-cell functions remain unknown. In the present study, we show that CR2-induced complement activation results in membrane depolarization, as indicated by annexin V binding, with kinetics similar to those of C3-fragment deposition and different from those of MAC formation. On the other hand, like MAC formation, depolarization requires activation of complement via the alternative pathway, as indicated by total inhibition upon neutralization of factor D, and is abrogated by combined blockade of CR1 and CR2, but not of either receptor alone. The membrane depolarization is not associated with the apoptosis of B cells, as examined by co-staining with APO-2.7 or by the TdT-mediated biotin-dUTP nick-end labelling (TUNEL) assay. Confocal microscopy revealed that depolarization and C3 deposition, unlike MAC deposition, are limited to restricted areas on the B-cell surface. Double staining revealed a close association between the C3-fragment patches and membrane depolarization, as well as redistribution of lipid rafts to these areas. We propose that these events may play a role in the regulation of B-cell signalling and cross-talk with T cells.

Decreased interleukin-2 responses to Fusobacterium nucleatum and Porphyromonas gingivalis in generalized aggressive periodontitis.

BACKGROUND: Compromised T-cell responses to periodontal pathogens may contribute to the pathogenesis of generalized aggressive periodontitis (GAgP). In this study, we attempted to characterize T-helper cell (Th1, Th2, and Th17) responses in patients with GAgP and healthy controls upon stimulation with disease-relevant pathogens. METHODS: Mononuclear cells (MNCs) from 10 white patients with GAgP and 10 white controls were stimulated with Porphyromonas gingivalis American Type Culture Collection (ATCC) 33277 (Pg), Prevotella intermedia ATCC 25611, Fusobacterium nucleatum ATCC 49256 (Fn), and similar bacteria isolated from the participants' inherent oral flora. Tetanus toxoid (TT) was used as control antigen. The resulting production of interferon-gamma (IFN-gamma) and interleukin (IL)-2, -4, -5 and -17 and the induced proliferation of CD4+ T cells were measured. RESULTS: MNCs from patients with GAgP exhibited decreased IL-2 responses to Pg and Fn. No difference was observed between patients with GAgP and controls with regard to CD4+ T-cell proliferation or the production of IFN-gamma and IL-4, -5, and -17, irrespective of whether type strains or bacteria isolated from the participants' oral cavity were used for stimulation. Moreover, similar proliferative and cytokine responses to TT were observed. Notably, smoking patients with GAgP exhibited significantly lower IFN-gamma responses to the bacteria and to TT than non-smoking patients or controls. CONCLUSIONS: The decreased IL-2 responses of patients with GAgP to Pg and Fn combined with adequate IL-2 responses to TT suggest an impaired antigen-specific T-cell reactivity with periodontal pathogens in GAgP. The decreased IFN-gamma responses of smokers within the patient group suggest that smoking may aggravate this impairment.

Enhancement of human adaptive immune responses by administration of a high-molecular-weight polysaccharide extract from the cyanobacterium Arthrospira platensis.

The effect of consumption of Immulina, a high-molecular-weight polysaccharide extract from the cyanobacterium Arthrospira platensis, on adaptive immune responses was investigated by evaluation of changes in leukocyte responsiveness to two foreign recall antigens, Candida albicans (CA) and tetanus toxoid (TT), in vitro. Consumption of Immulina by 11 healthy male volunteers caused an immediate, but temporary, increase of CA-induced CD4+ T-helper (Th) cell proliferation (P < .02). TT-induced Th cell proliferation was increased in individuals over 50 years of age (P < .05) and correlated with age (P < .02). Consumption for 8 days enhanced the CA-induced B cell proliferation (P < .02), but after 56 days a diminution was seen (P < .03). The CA-elicited production of the Th1 cytokines tumor necrosis factor (TNF)-alpha, interleukin (IL)-2, and interferon (IFN)-gamma was increased after Immunlina administration for 3 days (P < .001, < .03, and < .007, respectively), and increased IL-2 production persisted after 56 days (P < .004). The TNF-alpha, IFN-gamma, and IL-6 responses to TT were enhanced after 8 and 14 days (P < .002-.05), while IL-5 responses increased significantly within 3 days (P < .04) and fell below baseline levels after 14 days (P < .008). Conversely, consumption for 3 days inhibited the IL-4 responses to both CA and TT (P < .008 and P < .03, respectively). No effects on IL-10 responses were observed. Upon addition to normal mononuclear cells in vitro, Immulina elicited strong TNF-alpha, IL-1beta, and IL-6 responses, indicating that it acts by inducing a pro-inflammatory state. Taken together, the data suggest that Immulina causes an age-dependent, temporary enhancement of adaptive immune responses.