Avertano Noronha

Interferon β decreases T cell activation and interferon γ production in multiple sclerosis

Abstract Interferons (IFN) are biological molecules with anti-viral, anti-proliferative and immunomodulatory actions. There is evidence that IFN-γ increases the frequency of exacerbations of multiple sclerosis (MS) whereas IFN-β may reduce their frequency. Here we present evidence that IFN-β significantly decreases concanavalin A (Con A)-induced proliferation of peripheral blood mononuclear cells (PBMC) of MS patients and healthy individuals. Similar results were obtained when PBMS were activated through the T cell receptor (TcR) by anti-CD3 monoclonal antibody or independently of it by phorbol ester and Ca 2+ ionophore. These effects of IFN-β were also noted when IFN-γ and IFN-β were added together. Furthermore, IFN-β decreased proliferation when added to cells that were already pre-activated. Activated CD4 + and CD8 + T cells were downregulated to approximately the same extent. Analysis of cytokine production showed that IFN-γ production by Con A activated PBMC was increased in MS when compared to controls. IFN-β significantly decreased IFN-γ production in MS patients and control individuals. Con A activated cultures treated with IFN-β showed decreased IL2R expression and accumulation of IL2. These results show that IFN-β decreases T cell activation and IFN-γ production in vitro, effects that may be beneficial in MS.

Improved delayed visual reproduction test performance in multiple sclerosis patients receiving interferon beta-1b

We assessed neuropsychological function longitudinally in 30 MS patients who participated in the pivotal trial of interferon beta-1b (IFN-beta-1b). Nine patients received high-dose IFN-beta-1b (8.0 million units), eight low-dose IFN-beta-1b (1.6 MIU), and 13 placebo. There was significant improvement in Wechsler Memory Scale Visual Reproduction-Delayed Recall scores between years 2 and 4 of the trial in MS subjects receiving high-dose IFN-beta-1b. Motoric performance, MRI lesion area, and depression rating scores did not correlate with this finding. Comparison of MRI at baseline and at years 2 and 4 revealed significant changes over time for the total cohort (p < 0.02). Mean lesion area in the high-dose group did not change over time, whereas the low-dose and placebo groups had increases in total lesion area of 28 and 36%, respectively, at year 4. Expanded disability status scale scores did not change significantly between years 2 and 4 of the trial, nor did they correlate with MRI lesion area at any assessment point. We conclude that high-dose IFN-beta-1b improves delayed visual reproduction test performance in MS patients, a finding unlikely to be explained by practice effects or brain lesion area. NEUROLOGY 1996;47: 1463-1468

Interfer beta augments supperssor cell function in multiple sclerosis

Suppressor cell function has been previously reported to be decreased in patients with progressive multiple sclerosis (MS). The abnormality could not be corrected in vitro and was present even after patients were treated with immunosuppressive agents. We now report that interferon beta augments suppressor function in vitro in progressive MS. Nonspecific suppressor cell function as measured in a concanavalin A (Con A) suppressor assay was reduced in 24 MS patients (mean percent suppression, 19.6 ± 2.2) when compared to 19 normal subjects (mean percent suppression, 35.0 ± 3.3). The data are highly significant (p < 0.001). When recombinant human interferon beta (103 units/ml) was added to lymphocyte cultures with Con A, Suppressor activity improved from 19.6 ± 2.2 to 37.8 ± 2.6 in MS (p < 0.001) and from 35.0 ± 3.3 to 46.2 ± 3.5 (p < 0.025) in control subjects. This study shows that recombinant interferon beta improves suppressor function in humans, an effect that is particularly significant in progressive MS.

The experimental autoimmune encephalomyelitis (EAE) model of MS: utility for understanding disease pathophysiology and treatment

While no single model can exactly recapitulate all aspects of multiple sclerosis (MS), animal models are essential in understanding the induction and pathogenesis of the disease and to develop therapeutic strategies that limit disease progression and eventually lead to effective treatments for the human disease. Several different models of MS exist, but by far the best understood and most commonly used is the rodent model of experimental autoimmune encephalomyelitis (EAE). This model is typically induced by either active immunization with myelin-derived proteins or peptides in adjuvant or by passive transfer of activated myelin-specific CD4+ T lymphocytes. Mouse models are most frequently used because of the inbred genotype of laboratory mice, their rapid breeding capacity, the ease of genetic manipulation, and availability of transgenic and knockout mice to facilitate mechanistic studies. Although not all therapeutic strategies for MS have been developed in EAE, all of the current US Food and Drug Administration (FDA)-approved immunomodulatory drugs are effective to some degree in treating EAE, a strong indicator that EAE is an extremely useful model to study potential treatments for MS. Several therapies, such as glatiramer acetate (GA: Copaxone), and natalizumab (Tysabri), were tested first in the mouse model of EAE and then went on to clinical trials. Here we discuss the usefulness of the EAE model in understanding basic disease pathophysiology and developing treatments for MS as well as the potential drawbacks of this model.

Detection of in vivo stimulated cerebrospinal-fluid lymphocytes by flow cytometry in patients with multiple sclerosis.

To examine the immune response in the central nervous system in patients with multiple sclerosis (MS), we characterized the cell-cycle phase of lymphocytes from cerebrospinal fluid. Cells were stained with acridine orange, and both RNA and DNA content were determined by flow cytometry. Although most cells were in the quiescent phase of the cycle, a significant increase was seen in the percentage of cells in the first stage of the proliferative cycle (G1 phase) when 21 samples of cerebrospinal fluid from 17 patients with MS were compared with samples from 21 controls (P < 0.001). Stimulated cells in the first stage of the cycle were seen in all categories of MS: active, progressive, or stable. In addition, increased numbers of cells in the second stage of the cycle (S phase) were seen in six specimens from patients with MS, five of whom had actie disease, but no increases were seen in controls (P < 0.02). These data indicate that stimulated lymphocytes were present in the cerebrospinal fluid durng all phases of MS and that stimulation becomes more intense during an acute exacerbation.

Circulating immune complexes in neurologic disease

Sera from patients with multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), subacute sclerosing panencephalitis (SSPE), and myasthenia gravis (MG) were assayed for immune complexes. Three techniques were used: a modified Raji cell assay, the 129-Clq polyethyleneglycol assay, and a solid-phase Clq assay. Immune complex levels were elevated in sera of some patients with MS, ALS, and SSPE, but the elevations were modest when compared with active systemic lupus erythematosus (SLE). In some cases, abnormalities were detected in only one assay system; in other cases, abnormalities were detected by two or three assay systems. In MS, immune complex elevations correlated with active disease and with decreased suppressor cell activity. Of two ALAS patients with antecedent poliomyelitis, one had markedly increased levels of immune complexes in two assays. In MG, levels of immune complexes did not differ from those of controls.

Tau fraction of transferrin is present in human aqueous humor and is not unique to cerebrospinal fluid.

In view of the many similarities between the aqueous humor and the cerebrospinal fluid (CSF), we investigated whether the tau fraction of transferrin, which is believed to be specific for CSF, is also present in aqueous humor. Samples of aqueous humor and CSF from clinically normal individuals were analyzed by SDS-PAGE and isoelectric focusing (IEF), and probed with anti-transferrin antiserum and the lectin Limax flavus agglutinin. Specific bands which corresponded to transferrin and its tau fraction were detected in both fluids. After isolation of these bands by affinity chromatography with anti-transferrin antiserum, we detected a single, diffuse fraction at an apparent molecular weight of 80 kDa in both aqueous humor and CSF. Peptide mapping revealed no detectable difference in this fraction between the two fluids. The detection of the tau fraction in human aqueous humor indicates that this molecule cannot be considered a specific cerebrogenic marker as had been thought previously. In view of the known growth-promoting properties of transferrin, its isolation from both ocular and cranial fluids attains special significance.

Preferential increase of IL-2R+ CD4+ T cells and CD45RB− CD4+ T cells in the central nervous system in experimental allergic encephalomyelitis

We examined lymphocytes isolated from the spinal cord (SC), peripheral blood (PB) and lymph nodes (LN) draining the immunization site of Lewis rats with acute experimental allergic encephalomyelitis (EAE). Cells were analysed for T cell subset markers CD4 (mAb W3/25) and CD8 (mAb OX8), for IL-2R (mAb OX39), and for high molecular mass leukocyte common antigen (LCA, CD45RB) expression (mAb OX22). T cells expressing high (CD45RB+) or low (CD45RB-) molecular mass LCA are of different maturational stages and/or separate lineages. CD4+ T cells were more predominant in SC than in PB and LN; CD8+ T cells were scarce in SC but common in PB and LN. Activated CD4+ T cells (IL-2R+) were common in the SC and LN but infrequent in blood. CD4+ T cells that were CD45RB+ were scarce in the SC. In contrast, the majority of CD4+ T cells in the PB and LN were CD45RB+. The preferential accumulation of IL-2R+ CD4+ T cells and of CD45RB- CD4+ T cells in the central nervous system (CNS) indicates that a selective mechanism directs cell egress into CNS lesions in EAE.

Neutralizing antibodies to interferon

The impact of neutralizing antibodies (NAbs) to interferon-beta (IFN-β) on clinical and imaging parameters in multiple sclerosis (MS) is reviewed. An effect on relapse rates and imaging parameters was noted in patients who tested positive for NAbs, but disability measures were unaffected or showed a trend toward improvement. Patients who developed NAbs during treatment with IFN-β1a tended to remain NAb+, whereas those who developed NAbs during IFN-β1b treatment tended to revert to NAb− over time. NAbs were more persistent in patients with high titers. The prevalence of NAbs was lower when a higher-than-standard dose of IFN-β1b was given in a dose-comparison study. The prevalence of NAbs in an observational study of MS patients who exhibited suboptimal treatment responses to IFN-β1b was significantly less than the reported prevalence in clinical trials. An immunoregulatory effect of immune complexes of cytokine and anticytokine antibodies is proposed to account for the variability of clinical responses seen in patients who develop NAbs to IFN-β1b.

Role of interferons in demyelinating diseases

IFN beta-1b reduces the frequency of major multiple sclerosis attacks by 50 percent. Serial MRI scanning over the course of the clinical trial that led to approval of the agent revealed a significant lessening both in disease activity and in accumulating burden of disease in IFN beta-1b-treated patients compared to placebo-treated controls. The mechanism by which IFN beta-1b exerts its beneficial effect in multiple sclerosis is unknown. T suppressor cell function fails during MS attacks and is persistently subnormal in multiple sclerosis patients with progressive disease. IFN beta-1b partially restores suppressor function in multiple sclerosis patients. IFN beta-1b also inhibits release of lymphotoxin, tumor necrosis factor, and interferon gamma, at least in vitro. All three cytokines are toxic to oligodendrocytes. In contrast; production of transforming growth factor beta-1 (TGF beta 1) is increased by IFN beta-1b. TGF beta 1 is an immunosuppressive cytokine. All of the above listed actions of IFN beta-1b could contribute to its beneficial effect. Perhaps all do.