Current therapy of multiple sclerosis (MS) with interferon-beta (IFN-b) or glatiramer acetate (GA) has modest effects on the course of MS. Both compounds affect several immune variables, like expression of cell surface molecules and cytokine levels. Here we compared untreated MS, therapy with IFN-b alone and combined with GA, and healthy controls (HC), regarding expression on HLA -DR+ blood mononuclear cells (MNC) of C D1a that is a cell surface molecule with capacity to present glycolipids to T cells, and of C D80 and C D86 which are costimulatory molecules that activate Th1 and Th2 responses. C ytokine production by MNC was also measured. Flow cytometry and ELISA were used. C ross-sectional comparisons revealed that untreated MS patients had higher C D1a+ HLA -DR+ MNC and lower IL-10 production compared to patients treated with IFN-b or IFN-b+G A or HC. Untreated MS patients also had higher spontaneous IFN-g and IL-12p70 production compared to MS patients treated with IFN-b+G A or HC, but not when compa...

Multiple sclerosis (MS) is a disabling, inflammatory, demyelinating disease of the central nervous system considered to be mediated by autoreactive T cells. Dendritic cells (DC), being professional antigen-presenting cells, play a pivotal role in the decision between T-cell activation and anergy. It has been suggested that mature DC (mDC) induce immunity, whereas immature DC (imDC) have the potential to induce tolerance. In this study, we investigated the effects of autologous imDC versus autologous mDC on lymphocytes with respect to the expression of functionally important cell-surface molecules and production of cytokines. Our aims were to investigate whether the maturation status of DC differs between MS and healthy controls (HC) and to explore whether the effects of DC on T-cell responses differ between MS and HC. DC were generated from adherent blood mononuclear cells from patients with MS and HC. imDC were obtained by culture with either granulocyte-macrophage colony-stimulating factor (GM-CSF) + interleukin-4 (IL-4) or GM-CSF + IL-4 + IL-10. mDC were obtained by adding lipopolysaccharide to DC cultures. Upon coculture with autologous lymphocytes, mDC activated the autologous T cells as reflected by increased CD25 and cytotoxic T-lymphocyte antigen-4 expression on CD4(+) T cells together with the increased production of both T helper 1 (Th1) (IL-2 and interferon-gamma) and Th2 (IL-10 and IL-4) cytokines. Unmodulated naïve imDC induced the production of only IL-4. An exposure of imDC to IL-10 induced the production of IL-4 as well as IL-10 by autologous lymphocytes. We hypothesize that such imDC are important in controlling the proinflammatory environment in vivo in patients with MS.

Current therapy of multiple sclerosis (MS) with interferon-beta (IFN-beta) or glatiramer acetate (GA) has modest effects on the course of MS. Both compounds affect several immune variables, like expression of cell surface molecules and cytokine levels. Here we compared untreated MS, therapy with IFN-beta alone and combined with GA, and healthy controls (HC), regarding expression on HLA-DR+ blood mononuclear cells (MNC) of CD1a that is a cell surface molecule with capacity to present glycolipids to T cells, and of CD80 and CD86 which are costimulatory molecules that activate Th1 and Th2 responses. Cytokine production by MNC was also measured. Flow cytometry and ELISA were used. Cross-sectional comparisons revealed that untreated MS patients had higher CD1a+ HLA-DR+ MNC and lower IL-10 production compared to patients treated with IFN-beta or IFN-beta+GA or HC. Untreated MS patients also had higher spontaneous IFN-gamma and IL-12p70 production compared to MS patients treated with IFN-beta+GA or HC, but not when compared to MS patients on monotherapy with IFN-beta. Low CD1a+ HLA-DR+ MNC and low spontaneous production of IL-12p70 and IFN-gamma were more pronounced in patients treated with IFN-beta+GA than with IFN-beta alone. In order to clarify whether these changes reflect disease activity or treatment effects, we performed a follow up study. Nineteen MS patients with disease progression, despite monotherapy with IFN-beta for more than one year; were re-examined after one to three and four to six months of treatment with IFN-beta+GA. This combination therapy was associated with normalization of CD1a+ HLA-DR+ MNC, IL-12p70 and IFN-gamma. It remains to be shown whether these immunological changes imply a clinical benefit. Follow up studies of immune variables versus clinical effects during combined therapy of MS with IFN-beta+GA are ongoing.

This study evaluates levels of cerebrospinal fluid (C SF) brain-specific proteins (BSP) in subjects with optic neuritis (O N) who are at high risk of progression to multiple sclerosis (MS). Forty-one subjects had acute O N and 17 subjects with other neurological diseases (OND) served as controls. Twenty-o ne subjects with O N had white matter lesions on magnetic resonance imaging (MRI) and intrathecal synthesis of oligoclonal IgG bands (OB) consistent with being at high risk of progression to MS; eight of whom later were diagnosed with clinically definite MS (C DMS). Levels of S100B, ferritin and two neurofilament heavy chain phosphoforms (NfH SMI34 and NfH SMI35 ) were analysed using ELISA technique. A putative index of ‘axonal health’ was expressed as a ratio of NfH SMI34 to NfH SMI35 . NfH SMI34 and the NfHSMI34:SMI35 were significantly elevated in subjects with O N compared to controls. No significant differences in levels of C SF BSP were seen between O N subjects with C DMS plus those at high risk of progression to MS and O N subjects with normal MRI and negative C SF analysis. In conclusion, there is evidence of axonal damage in subjects who present with O N, which is independent of the diagnosis of C DMS.

Using multivariate analyses, individual risk of clinically definite multiple sclerosis (CDMS) after monosymptomatic optic neuritis (MON) was quantified in a prospective study with clinical MON onset during 1990-95 in Stockholm, Sweden. During a mean follow-up time of 3.8 years, the presence of MS-like brain magnetic resonance imaging (MRI) lesions and oligoclonal immunoglobulin (Ig) G bands in cerebrospinal fluid (CSF) were strong prognostic markers of CDMS, with relative hazard ratios of 4.68 [95% confidence interval (CI) 2.21-9.91] and 5.39 (95% CI 1.56-18.61), respectively. Age and season of clinical onset were also significant predictors, with relative hazard ratios of 1.76 (95% CI 1.02-3.04) and 2.21 (95% CI 1.13-3.98), respectively. Based on the above two strong predictors, individual probability of CDMS development after MON was calculated in a three-quarter sample drawn from a cohort, with completion of follow-up at three years. The highest probability, 0.66 (95% CI 0.48-0.80), was obtained for individuals presenting with three or more brain MRI lesions and oligoclonal bands in the CSF, and the lowest, 0.09 (95% CI 0.02-0.32), for those not presenting with these traits. Medium values, 0.29 (95% CI 0.13-0.53) and 0.32 (95% CI 0.07-0.73), were obtained for individuals discordant for the presence of brain MRI lesions and oligoclonal bands in the CSF. These predictions were validated in an external one-quarter sample.

Magnetic resonance imaging (MRI) remains the most valuable tool for monitoring disease activity and progression in patients with multiple sclerosis (MS), a chronic demyelinating disease of the central nervous system (CNS) with presumably autoimmune etiology. Chemokine receptors have been implicated in MS as key molecules directing inflammatory cells into the CNS. Regulatory (CD4+CD25+) T cells (Tr cells) are important in suppressing autoimmunity, and their absolute or functional deficit could be expected in MS. In the present study, venous blood was obtained from MS patients concurrent with MRI examination of the brain, and expression of chemokine receptors CCR1, CCR2, CCR5, CXCR3 and CXCR4 by CD4 T cells and monocytes, proportions of Tr cells, as well as expression of CD45RO, CD95, CTLA-4, HLA-DR and interleukin (IL)-10 by Tr cells and non-Tr (CD25-) CD4 T cells was analyzed by flow cytometry. Surface expression of CXCR3 by CD4 T cells was downregulated in the group of patients with high lesion load (LL) on T2-weighted images and gadolinium (Gd)-enhancing lesions on T1-weighted images, compared to the group with high LL and no Gd-enhancing lesions, and to the group with low LL, suggesting internalization of CXCR3 due to the release of its chemokine ligand (IP-10/CXCL10) from active MS lesions. Proportions of Tr cells amongst all CD4 T cells, and expression of IL-10 by Tr cells were increased in the patients with high LL and Gd-enhancing lesions. These results suggest that there is correlation between MRI parameters, chemokine receptor expression and the status of circulating Tr cells in MS, but further studies need to discriminate between pathogenetically relevant and bystander phenomena.

Secondary lymphoid organ chemokines have been implicated in chronic inflammation. Their expression in the central nervous system (CNS) has not been studied. Here, levels of secondary lymphoid organ chemokines CCL19 (Exodus-3, MIP-3beta), CCL21 (Exodus-2, 6Ckine, SLC) and CXCL12 (SDF-1alpha) were analysed by ELISA in cerebrospinal fluid (CSF) and plasma from patients with multiple sclerosis (MS); acute optic neuritis (ON) with oligoclonal IgG in the CSF (i.e., first bout of MS); acute ON without oligoclonal IgG (non-MS-type ON); other inflammatory neurological diseases (OIND); and non-inflammatory neurological diseases (NIND). NIND CSF contained CCL19 and CXCL12, while CCL21 was not detected. Intrathecal production of CCL19 and CCL21 was elevated in MS, MS-type ON, and OIND, but not in non-MS-type ON. In MS, CSF levels of CCL19 weakly correlated with CSF cell counts. Intrathecal production of CXCL12 was elevated only in OIND. The role of elevated CCL19 and CCL21 in MS could be retention of mature dendritic cells (DC) in the CNS, recruitment of nai;ve T cells and activated B cells, as well as de novo formation of secondary lymphoid structures in MS plaques.

CD4(+)CD25(high) T regulatory (Tr) cells, representing high IL-2 receptor alpha chain expressing cells, have been shown to inhibit proliferation and cytokine secretion by CD4(+) T cells that are assumed to represent important effector cells in auto-aggressive immunity. Tr cells may therefore be considered of importance in the pathogenesis of multiple sclerosis (MS). Glatiramer acetate (GA; Copaxone) is approved as a disease-modulating agent that ameliorates the course of MS. The goal of this study was to examine in vitro effects of GA on Tr cells from MS patients subgrouped according to treatment without or with disease-modulating drugs, and healthy controls (HC). Three-colour flow cytometry was used to investigate in vitro influence of GA, and of the encephalitogenic myelin basic protein (MBP) peptide 83-89 as control, on the blood Tr cell proportion and on their functionally important cell surface molecules CD45RO, CD69, CD95 and HLA-DR, and on intracellular CTLA-4 and IL-10. Irrespective of exposure to GA or MBP((83-99)), levels of blood Tr cells expressing HLA-DR remained low in untreated MS patients and HC compared to the three treated MS patient groups. In vitro exposure to GA resulted in elevated levels of IL-10 producing Tr cells in all MS patient groups irrespective of receiving treatment as well as in HC. Exposure to GA or MBP((83-99)) had no effects on levels of Tr cells expressing other above-mentioned molecules. We conclude that GA induces elevated IL-10 production by Tr cells that is uniform and independent of ongoing MS treatment with IFN-beta or GA or IFN-beta+GA.

Myeloid and plasmacytoid dendritic cells (DC) are present in cerebrospinal fluid (CSF) in non-inflammatory neurological diseases (NIND) and elevated in clinically definite multiple sclerosis (MS) and in early MS - acute monosymptomatic optic neuritis (ON). Here, we show that expression of CCR5, a chemokine receptor for regulated on activation, normal T cell expressed and secreted (RANTES) and macrophage inflammatory protein (MIP)-1alpha/beta, is elevated on blood myeloid (CD11c(+)) DC in MS and ON compared to non-inflammatory controls. In contrast, expression of CXCR4, a receptor for stromal cell-derived factor (SDF)-1alpha, is similar in all groups. Blood myeloid DC from MS patients respond chemotactically to RANTES and MIP-1beta, which are expessed in MS lesions. In active MS and ON, expression of CCR5 by myeloid DC in blood correlates with numbers of these cells in CSF. Thus, elevation of CCR5 may contribute to recruitment of myeloid DC to CSF in MS and ON. Recruitment of plasmacytoid DC to CSF appears to be CCR5-independent.

Myeloid (CD11c(+)) dendritic cells (DC) are present in cerebrospinal fluid (CSF), as well as in the meninges and choroid plexus. Functional studies of these DC are hindered or impossible. To obviate this problem, we investigated the effects of CSF supernatants from patients with non-inflammatory neurological diseases (NIND), multiple sclerosis (MS), bacterial meningitis (BM) and Lyme meningoencephalitis (LM) on immature monocyte-derived DC (moDC) from healthy donors. CSF supernatants caused maturation of moDC (MS > LM > NIND > BM), as reflected by a decrease in CD1a, and an increase in HLA-DR, CD80 and CD86 expression. The maturation effect of MS CSF and LM CSF could be blocked by anti-TNF-alpha MoAb or recombinant human IL-10. moDC cultured with BM CSF either remained immature or turned into CD14(+) macrophage-like cells and were relatively inefficient at inducing T cell responses in vitro. In contrast, moDC cultured with LM CSF induced strong Th1 responses. Both BM CSF and LM CSF contained IFN-gamma, a cytokine that augments IL-12 production by moDC and hence should confer an ability to induce a Th1 response. However, BM CSF also contained high levels of IL-10, which could antagonize the effects of IFN-gamma on moDC. moDC cultured with MS CSF induced a higher production of IFN-gamma from T. cells compared to moDC cultured with NIND CSF or BM CSF. In summary, soluble factors present in the CSF may influence the phenotype and functions of meningeal, choroid plexus and CSF DC which, in turn, may have an impact on the character of intrathecal T. cell responses.

Abnormal immune responses to myelin associated glycoprotein (MAG), a component of myelin of the central and peripheral nervous system, have been suggested to play a role in the pathogenesis of multiple sclerosis (MS) and certain types of inflammatory polyneuropathy. To identify possible immunodominant MAG peptides in neuroinflammation, we examined T and B cell responses to five selected synthetic MAG peptides and myelin proteins in 21 patients with non-inflammatory polyneuropathy, 26 patients with MS, 10 optic neuritis patients and 17 healthy subjects. Enzyme-linked immunosorbent spot-forming cell assays were adopted, allowing the detection and enumeration of individual antigen responsive T and B cells in body fluids. Patients with polyneuropathy as well as those with MS had elevated levels of T and B cells recognizing MAG and its peptides. Any of the five MAG peptides under study functioned as immunodominant T and/or B cell epitope in individual subjects. None of the MAG peptides elicited a specific disease-associated T or B cell response. The enhanced T and B cell response to myelin components like MAG may play some role in initiation and/or progression of these diseases, but they could also represent secondary responses associated with myelin damage and indicate tolerization rather than autoaggressive immunity.

Glatiramer acetate (GA; Copolymer 1; Copaxone) and interferon-beta (IFN-beta) modulate the course of multiple sclerosis (MS), but probably by different mechanisms. GA, a mixture of synthetic peptides, is believed to act as an altered peptide ligand with inhibitory effects on autoreactive T cells and promoting Th2 cells. It is unknown whether GA affects dendritic cells (DCs), which, besides strong antigen presenting capacity, orchestrate Th1 and Th2 responses. IFN-beta inhibits IL-12 production by DCs over unknown mechanisms. This study was designed to investigate in vitro effects of GA and IFN-beta on the development and function of DCs from MS patients and healthy controls, and to explore their possible synergistic effects on DCs. DCs were generated from adherent blood mononuclear cells (MNCs). GA or IFN-beta or both, when added at initiation of DC cultures, rapidly promoted the development of adherent MNCs into floating, activated DCs as reflected by up-regulation of HLA-DR and CD86 expression. IFN-beta, but not GA, induced IL-3R expression on DCs. Compared to DCs from healthy controls, MS patients' DCs expressed higher levels of the myeloid DC marker CD1a and produced lower amounts of IL-10. GA reduced IL-12 production by DCs. IFN-beta also reduced IL-12, but increased IL-10 production by DCs from both MS patients and healthy controls. GA and IFN-beta synergistically suppressed CD1a and enhanced CD86 expression on MS DCs. These findings document novel mechanisms of action of GA and IFN-beta at the DC level in MS.