The pathogenesis of multiple sclerosis. Cytotoxic antibodies against myelin sheath tissue in multiple sclerosis

Cytotoxic antibodies to myelin can be demonstrated by the method of 51Cr-release from chick erythrocytes coated with myelin basic protein. The cytotoxic antibody is inactivated by heating to 56 degrees C and needs complement in order to exert its action. The antibody was determined as IgM and IgG. It has relative specifity and shows cross-reaction with other basic proteins. The cytotoxic antibody was found in only 8% of healthy persons. Patients with multiple sclerosis were positive in 87% of the cases and in acute cases in 94%. In other neurological diseases cytotoxic antibody was present in 64%. The occurrence of cytotoxic antibody to myelin protein is not specific for a particular neurological disorder, especially not for multiple sclerosis. Cytotoxic antibodies arise as a secondary phenomenon, they are not the cause of the disease involved. They appear to be suitable, however, to determine, in association with cellular immunological reactions against myelin which may be regarded as the "primary" immunological processes, the demyelination process in the multiple sclerosis focus.     

Spontaneous and induced remyelination in multiple sclerosis and the Theiler's virus model of central nervous system demyelination

Remyelination in the central nervous system, originally thought to occur rarely, if ever, is now an established phenomena in multiple sclerosis patients. However, the extent of myelin repair is incomplete and limited. Experimental models of central nervous system demyelination provide an opportunity to study the cellular and molecular events involved in remyelination. These models may provide some clue to why remyelination in multiple sclerosis is incomplete as well as suggest potential methods to stimulate central nervous system repair. In this review we examine the morphological aspects of central nervous system remyelination and discuss both spontaneous and induced remyelination in multiple sclerosis and experimental models of central nervous system demyelination. We give special emphasis to the Theiler's virus model of central nervous system demyelination and its usefulness to identify therapeutic agents to promote remyelination. The role of immunoglobulins in promoting remyelination in both the Theiler's model system and in multiple sclerosis is discussed. Finally, we examine the potential physiological role of demyelination and remyelination and its relationship with clinical manifestations of central nervous system disease.     

The neuregulin, glial growth factor 2, diminishes autoimmune demyelination and enhances remyelination in a chronic relapsing model for multiple sclerosis

Glial growth factor 2 (GGF2) is a neuronal signal that promotes the proliferation and survival of the oligodendrocyte, the myelinating cell of the central nervous system (CNS). The present study examined whether recombinant human GGF2 (rhGGF2) could effect clinical recovery and repair to damaged myelin in chronic relapsing experimental autoimmune encephalomyelitis (EAE) in the mouse, a major animal model for the human demyelinating disease, multiple sclerosis. Mice with EAE were treated with rhGGF2 during both the acute and relapsing phases. Clinically, GGF2 treatment delayed signs, decreased severity, and resulted in statistically significant reductions in relapse rate. rhGGF2-treated groups displayed CNS lesions with more remyelination than in controls. This correlated with increased mRNA expression of myelin basic protein exon 2, a marker for remyelination, and with an increase in the CNS of the regulatory cytokine, interleukin 10, at both the RNA and protein levels. Thus, a beneficial effect of a neurotrophic growth factor has been demonstrated on the clinical, pathologic, and molecular manifestations of autoimmune demyelination, an effect that was associated with increased expression of a T helper 2 cytokine. rhGGF2 treatment may represent a novel approach to the treatment of multiple sclerosis.     

Inflammation and the brain

Inflammation in the brain selectively damages the myelin sheath resulting in a variety of clinical syndromes of which the most common is multiple sclerosis. In these disorders, the areas of inflammation and demyelination can be identified in life by magnetic resonance imaging. Events occurring at the blood-brain barrier depend on T-cell activation, which increases immune surveillance within the central nervous system. T-cells activated against brain antigens persist to establish the conditions needed for inflammatory demyelination and this depends on local release of cytokines, culminating in removal of oligodendrocytes and their myelin lamellae by macrophages or microglia. These interactions involve binding between receptors present on microglia for the Fc portion of antibody and complement components to corresponding ligands on target cells. Taken together, the evidence from clinical and experimental studies provides a rationale for the issue of immunological treatments in patients with multiple sclerosis.     

An increased number of follicles containing activated CD69+ helper T cells and proliferating CD71+ B cells are found in H. pylori-infected gastric mucosa

Multiple sclerosis is characterized by myelin destruction and oligodendrocyte loss. The neuropathological hallmark of the disease is the presence of demyelinated plaques in the central nervous system. We have recently found a gliotoxic factor in MS cerebrospinal fluid which induces programmed cell death in vitro, in glial cells. Here we show DNA fragmentation and glial cell death in biopsy samples, obtained from a patient who underwent surgery with suspicion of tumor, and whose disease record, including brain autopsy, demonstrated an active multiple sclerosis. We used the in situ TUNEL technique, a method which sensitively detects the DNA fragmentation accompanying programmed cell death in tissue sections, and compatible with classical fixation techniques. We found intense DNA fragmentation in nuclei of glial cells at-or very near-to the site of demyelination. A double labeling technique showed that glial fibrillary associated protein positive astrocytes may undergo programmed cell death in multiple sclerosis.     

Autoimmunity to myelin oligodendrocyte glycoprotein in rats mimics the spectrum of multiple sclerosis pathology

Multiple sclerosis is a chronic inflammatory disease characterized by perivenous inflammation and focal destruction of myelin. Many attempts have been undertaken previously to create animal models of chronic inflammatory demyelinating diseases through autoimmunity or virus infection. Recently, however, a new model of myelin oligodendrocyte glycoprotein (MOG) induced autoimmune encephalomyelitis became available, which, in a very standardized and predictable way, leads to chronic (relapsing or progressive) disease and widespread CNS demyelination. In the present study we actively induced MOG-experimental autoimmune encephalomyelitis (EAE) in different inbred rat strains using different immunization protocols. The pathology found in our models closely reflects the spectrum of multiple sclerosis (MS) pathology: Classical MS as well as variants such as optic neuritis, Devic's disease and Marburg's type of acute MS are mimicked in rats immunized with MOG antigen. Furthermore we demonstrate, that by using the proper strain/sensitization regime, subforms of MS such as for instance neuromyelitis optica can be reproducibly induced. Our study further supports the notion, that incidence and expression of the disease in this model, alike the situation in multiple sclerosis, is determined by genetic and environmental factors.     

Generation of autonomously pathogenic neo-autoreactive Th1 cells during the development of the determinant spreading cascade in murine autoimmune encephalomyelitis

Chronic progression of autoimmune disease is accompanied by the acquisition of autoreactivity to new self-determinants. Recent evidence indicates that this process, commonly referred to as determinant spreading, may be pathogenic for chronicity. Our studies on experimental autoimmune encephalomyelitis (EAE), a murine model widely used in multiple sclerosis (MS) studies, have shown that determinant spreading develops as a predictable sequential cascade of neo-autoimmunity during progression to chronic disease. By 7-8 weeks after immunization of (SWR x SJL)F1 mice with the immunodominant myelin proteolipid protein determinant (PLP 139-151), splenocytes consistently respond to the immunodominant myelin basic protein determinant (MBP 87-99). In the present study, we directly address the pathogenicity of neo-autoimmunity resulting from endogenous self-priming during the course of disease. Our results indicate that T cells responding to the spreading MBP 87-99 determinant produce a proinflammatory cytokine profile consistent with type 1 helper T cells (Th1) cells. In addition, splenocytes activated to the spreading MBP 87-99 determinant consistently transfer acute EAE in naive recipients even when T cells reactive to the priming PLP 139-151 immunogen are eliminated by bromodeoxyuridine (BUdR)-mediated photolysis. Our data indicate that endogenous neo-autoantigen priming during chronic autoimmune disease generates type 1 helper T cells (Th1) cells that are autonomously pathogenic. These results provide further evidence supporting the view that determinant spreading is a pathogenic process that leads to chronic progression of autoimmune disease.     

A journey from blame to empathy in a family assessment of a mother and her sons

Ultrastructural analysis of myelin from 8-month-old mice deficient in the myelin-associated glycoprotein revealed pronounced and characteristic alterations of the periaxonal oligodendrocyte processes, consisting of intracytoplasmic deposition of vesicular material, multivesicular bodies, mitochondria, and lipofuscin granules, as well as granular or paracrystalline inclusions. These alterations are similar to those described before as "dying-back oligodendrogliopathy" in diseases of toxic or immune-mediated demyelination including multiple sclerosis.     

Amino acids within residues 181-200 of the nicotinic acetylcholine receptor alpha1 subunit involved in nicotine binding

Hematogenous macrophages and resident brain microglia are agents of demyelination in multiple sclerosis (MS) and paradoxically may also participate in remyelination. In vitro studies have shown that macrophage enrichment of aggregate brain cultures promotes myelination per se and enhances the capacity to remyelinate following a demyelinating episode. It has been hypothesized that remyelination in MS is implemented by surviving dedifferentiated oligodendrocytes or by newly recruited progenitors that migrate, proliferate and synthesize myelin in response to signalling molecules in the local environment. We postulate that macrophage-derived cytokines or growth factors may directly or indirectly promote oligodendroglial proliferation and differentiation, contributing to myelin repair in inflammatory demyelinating disease.     

Pathogenesis of multiple sclerosis. Antibody dependent cytotoxicity of lymphocytes against myelin basic proteins in multiple sclerosis

In 79 of 100 patients with multiple sclerosis, serum antibodies of the IgG type were demonstrated which render normal lymphocytes cytotoxic against basic protein of myelin. In 11 cerebrospinal fluids which were tested the antibody was also found. The method used was the release of 51Cr from chicken erythrocytes coated with antigen. The antibody-dependent lymphocyte cytotoxicity is related to the degree of activity of the disease and not to the evolution of the disease or its course. As the disease worsens, the frequency of positive reactions is higher than in inactive stages. The immunological reaction is very specific for multiple sclerosis, in patients with other neurological disorders only 5% had positive findings. The antibody-dependent cytotoxicity of lymphocytes against basic protein of myelin may be attributed with a certain diagnostic significance. The reaction seems suitable for the supervision of the course of multiple sclerosis and to check the effect of therapeutic measures. The antibody-dependent lymphocyte cytotoxicity against basic protein of myelin is considered to be significant in the pathogenesis of multiple sclerosis.     

Chromosome 14 contains determinants that regulate susceptibility to Theiler's virus-induced demyelination in the mouse

Theiler's murine encephalomyelitis virus causes a chronic demyelinating disease in susceptible strains of mice that is similar to human multiple sclerosis. Several nonmajor histocompatibility complex-linked genes have been implicated as determinants of susceptibility or resistance to either demyelination or virus persistence. In this study, we used linkage analysis of major histocompatibility complex identical H-2d (DBA/2J x B10.D2) F2 intercross mice to identify loci associated with susceptibility to virus-induced demyelinating disease. In a 20-cM region on chromosome 14, we identified four markers, D14Mit54, D14Mit60, D14Mit61, and D14Mit90 that are significantly associated with demyelination. Because two peaks were identified, one near D14Mit54 and one near D14Mit90, it is possible that two loci in this region are involved in controlling demyelination.     

Perspectives in multiple sclerosis research]

There is little doubt that multiple sclerosis (MS) is an immune mediated disease, yet the exact immunological mechanisms, that are responsible for inflammation and demyelination in this disease are controversial. Recent evidence is summarized here, which suggests that heterogeneous pathogenetic mechanisms may lead to the inflammatory demyelinating plaques in different MS patients. This heterogeneity apparently involves the antigen specificity of the immune response as well as the mechanisms, responsible for the destruction of myelin sheaths. Since such a pathogenetic heterogeneity may have consequences for the design of therapeutic studies, strategies are discussed, which should allow a more accurate categorization of patient sub-groups in the future.     

Failure of treatment with Linomide or oral myelin tolerization to ameliorate demyelination in a viral model of multiple sclerosis

Both Linomide (quinoline-3-carboxamide) and tolerization with self-antigens have been demonstrated to successfully ameliorate demyelinating disease in experimental autoimmune encephalomyelitis (EAE). Based on the autoimmune hypothesis of multiple sclerosis (MS), both agents have been tested in clinical trials but have been found to be toxic or not efficacious. We investigated the efficacy of these immunomodulators in an alternative experimental model of MS, a virus-induced demyelinating disease. Oral administration of Linomide to Theiler's virus-infected mice beginning either at time of infection or at day 15 post-infection (p.i.) resulted in an increased percentage of spinal cord quadrants with demyelination. Administration of Linomide beginning at day 15 p.i. increased lesion size as compared to infected control-treated mice. Treatment with 80 mg kg(-1) day(-1) of Linomide beginning at the time of infection significantly increased the number of Theiler's murine encephalomyelitis virus (TMEV)-positive cells mm(-2) of spinal cord white matter. There were no differences in the amount of remyelination between mice treated with Linomide or water. However, chronically infected mice treated with Linomide had severely reduced spontaneous vertical activity as measured using a activity wheel. Oral tolerization of mice with mouse or bovine myelin had no effect on virus-induced demyelination or virus antigen expression. The contrasting results obtained between the TMEV model and the autoimmune model of demyelination do not support recent reports suggesting that the underlying mechanism of demyelination in the Theiler's model is autoimmune.     

CD8+ myelin peptide-specific T cells can chemoattract CD4+ myelin peptide-specific T cells: importance of IFN-inducible protein 10

The demyelination process that occurs in the central nervous system (CNS) of patients with multiple sclerosis (MS) is due, in part, to an inflammatory response in which CD4+ and CD8+ T cells and macrophages infiltrate white matter. While it is thought that the inflammatory and demyelination process in MS is the product of Th1-associated cytokines secreted by CD4+ myelin protein-specific T cells present in the CNS, the mechanisms that are responsible for the recruitment and maintenance of these myelin-reactive CD4+ T cells in the CNS have not been elucidated. We have shown previously that CD8+ CTL that recognize peptides derived from sequences of the myelin proteolipid protein (PLP) presented by HLA class I molecules can be generated in vitro, and that these PLP-specific CD8+ CTL secrete the proinflammatory chemokines macrophage-inflammatory protein-1alpha and -1beta, IL-16, and IP-10. In this study, we demonstrate that soluble products of these PLP-specific CD8+ CTL can chemoattract CD4+ T cells that are specific for a myelin basic protein peptide and a PLP peptide, and that the majority of this chemotactic activity is mediated by IFN-inducible protein 10. These results demonstrate that PLP-specific CD8+ T cells can play a role in the recruitment and retention of myelin-derived peptide-specific CD4+ T cells, and indicate that they may play a proinflammatory role in the pathogenesis of MS.     

Primary demyelination in transgenic mice expressing interferon-gamma

Ever since the use of interferon-gamma to treat patients with multiple sclerosis resulted in enhanced disease, the role of IFN-gamma in demyelination has been under question. To address this issue directly, transgenic mice were generated that expressed the cDNA of murine IFN-gamma in the central nervous system by using an oligodendrocyte-specific promoter. Expression of the transgene occurred after 8 weeks of age, at which time the murine immune and central nervous systems are both fully developed. Directly associated with transgene expression, primary demyelination occurred and was accompanied by clinical abnormalities consistent with CNS disorders. Additionally, multiple hallmarks of immune-mediated CNS disease were observed including upregulation of MHC molecules, gliosis and lymphocytic infiltration. These results demonstrate a direct role for IFN-gamma as an inducer of CNS demyelination leading to disease and provide new opportunities for dissecting the mechanism of demyelination.     

Treatment of experimental encephalomyelitis with a novel chimeric fusion protein of myelin basic protein and proteolipid protein

It has been shown that peripheral T cell tolerance can be induced by systemic antigen administration. We have been interested in using this phenomenon to develop antigen-specific immunotherapies for T cell-mediated autoimmune diseases. In patients with the demyelinating disease multiple sclerosis (MS), multiple potentially autoantigenic epitopes have been identified on the two major proteins of the myelin sheath, myelin basic protein (MBP) and proteolipid protein (PLP). To generate a tolerogenic protein for the therapy of patients with MS, we have produced a protein fusion between the 21.5-kD isoform of MBP (MBP21.5) and a genetically engineered form of PLP (deltaPLP4). In this report, we describe the effects of treatment with this agent (MP4) on clinical disease in a murine model of demyelinating disease, experimental autoimmune encephalomyelitis (EAE). Treatment of SJL/J mice with MP4 after induction of EAE either by active immunization or by adoptive transfer of activated T cells completely prevented subsequent clinical paralysis. Importantly, the administration of MP4 completely suppressed the development of EAE initiated by the cotransfer of both MBP- and PLP-activated T cells. Prevention of clinical disease after the intravenous injection of MP4 was paralleled by the formation of long-lived functional peptide-MHC complexes in vivo, as well as by a significant reduction in both MBP- and PLP-specific T cell proliferative responses. Mice treated with MP4 were resistant to disease when rechallenged with an encephalitogenic PLP peptide emulsified in CFA, indicating that MP4 administration had a prolonged effect in vivo. Administration of MP4 was also found to markedly ameliorate the course of established clinical disease. Finally, MP4 therapy was equally efficacious in mice defective in Fas expression. These results support the conclusion that MP4 protein is highly effective in suppressing disease caused by multiple neuroantigen epitopes in experimentally induced demyelinating disease.     

Improved high-performance liquid chromatographic method for the separation and quantification of lipid classes: application to fish lipids

The demyelination process that occurs in the central nervous system (CNS) of patients with multiple sclerosis (MS) is in part due to an inflammatory response in which CD4+ and CD8+ T cells and macrophages infiltrate white matter. In this study, we have identified a peptide sequence derived from the CNS-specific myelin protein proteolipid protein (PLP) which could bind to HLA-A3 and induce a HLA-A3-restricted CD8+ CTL response from HLA-A3+ donors. These PLP peptide-specific CTL could lyse HLA-A3+ target cells pulsed with a homologous peptide derived from the CRM1 protein of Saccharomyces cerevisae. These findings demonstrate the immunogenic potential of a PLP-derived peptide for generation of autoreactive HLA-A3-restricted CD8+ CTL, and further show that these CTL can be activated by a peptide derived from a common environmental microorganism.     

Oligodendrocyte apoptosis and primary demyelination induced by local TNF/p55TNF receptor signaling in the central nervous system of transgenic mice: models for multiple sclerosis with primary oligodendrogliopathy

The scientific dogma that multiple sclerosis (MS) is a disease caused by a single pathogenic mechanism has been challenged recently by the heterogeneity observed in MS lesions and the realization that not all patterns of demyelination can be modeled by autoimmune-triggered mechanisms. To evaluate the contribution of local tumor necrosis factor (TNF) ligand/receptor signaling pathways to MS immunopathogenesis we have analyzed disease pathology in central nervous system-expressing TNF transgenic mice, with or without p55 or p75TNF receptors, using combined in situ terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling and cell identification techniques. We demonstrate that local production of TNF by central nervous system glia potently and selectively induces oligodendrocyte apoptosis and myelin vacuolation in the context of an intact blood-brain barrier and absence of immune cell infiltration into the central nervous system parenchyma. Interestingly, primary demyelination then develops in a classical manner in the presence of large numbers of recruited phagocytic macrophages, possibly the result of concomitant pro-inflammatory effects of TNF in the central nervous system, and lesions progress into acute or chronic MS-type plaques with axonal damage, focal blood-brain barrier disruption, and considerable oligodendrocyte loss. Both the cytotoxic and inflammatory effects of TNF were abrogated in mice genetically deficient for the p55TNF receptor demonstrating a dominant role for p55TNF receptor-signaling pathways in TNF-mediated pathology. These results demonstrate that aberrant local TNF/p55TNF receptor signaling in the central nervous system can have a potentially major role in the aetiopathogenesis of MS demyelination, particularly in MS subtypes in which oligodendrocyte death is a primary pathological feature, and provide new models for studying the basic mechanisms underlying oligodendrocyte and myelin loss.     

Myelin composition in acute and chronic multiple sclerosis in relation to cerebral lysosomal activity

The neuropathology of 3 cases of acute multiple sclerosis was correlated with biochemical analyses. Astrocytosis was a characteristic feature of the diffuse demyelinating lesions in one case and lymphocytic cuffing characterized the well-defined plaques present in the white matter of the other two cases. No abnormalities were found in the protein or lipid composition of isolated myelin, despite a wide range of recovery. Nevertheless, the gel electrophoretic protein pattern of white matter adjacent to plaque areas showed selective loss of myelin basic protein. Lysosomal acid proteinase and beta-glucuronidase levels were very significantly increased in all white matter samples in which astrocytosis was a major neuropathological feature. Levels were less markedly raised in samples containing discrete active plaques. Enzyme changes were also found in the apparently normal white matter of 2 of the cases. Acid proteinase activity was in the normal range but the activities of beta-glucuronidase and acetylcholine esterase were elevated. The significance of these results in relation to glial cell activity in the early stages of demyelination is discussed.     

Injuries in the etiopathogenesis of multiple sclerosis

An injury might derange the protective function of the blood-brain barrier, and thus it represents one of the possible pathogenetic factors in the demyelination of the neural axis. However, the effect of injury on the occurrence or deterioration of multiple sclerosis is still controversial. According to most authors, the importance of injury in individual cases of multiple sclerosis is undeniable, as well as the fact that injuries are factors of progression and deterioration of the disease, but never its cause. Consequently, injuries can cause only temporary disability, and not permanent. Nevertheless, the incidence of multiple sclerosis increases proportionally to the severity of injury. The length of the period from the occurrence of injury to possible demyelination is still not established. Studies and clinical reports point to the fact that in the evaluation of injury as a precipitating factor for the vulnerability of the blood-brain barrier, the severity of the injury is of greater importance than its site.