Cedric S. Raine

Identification of autoantibodies associated with myelin damage in multiple sclerosis

The molecular mechanisms underlying myelin sheath destruction in multiple sclerosis lesions remain unresolved. With immunogold–labeled peptides of myelin antigens and high–resolution microscopy, techniques that can detect antigen–specific antibodies in situ, we have identified autoantibodies specific for the central nervous system myelin antigen myelin/oligodendrocyte glycoprotein. These autoantibodies were specifically bound to disintegrating myelin around axons in lesions of acute multiple sclerosis and the marmoset model of allergic encephalomyelitis. These findings represent direct evidence that autoantibodies against a specific myelin protein mediate target membrane damage in central nervous system demyelinating disease.

Glutamate excitotoxicity in a model of multiple sclerosis

Glutamate excitotoxicity mediated by the AMPA/kainate type of glutamate receptors damages not only neurons but also the myelin-producing cell of the central nervous system, the oligodendrocyte. In multiple sclerosis, myelin, oligodendrocytes and some axons are lost as a result of an inflammatory attack on the central nervous system. Because glutamate is released in large quantities by activated immune cells, we expected that during inflammation in MS, glutamate excitotoxicity might contribute to the lesion. We addressed this by using the AMPA/kainate antagonist NBQX to treat mice sensitized for experimental autoimmune encephalomyelitis, a demyelinating model that mimics many of the clinical and pathologic features of multiple sclerosis. Treatment resulted in substantial amelioration of disease, increased oligodendrocyte survival and reduced dephosphorylation of neurofilament H, an indicator of axonal damage. Despite the clinical differences, treatment with NBQX had no effect on lesion size and did not reduce the degree of central nervous system inflammation. In addition, NBQX did not alter the proliferative activity of antigen-primed T cells in vitro, further indicating a lack of effect on the immune system. Thus, glutamate excitotoxicity seems to be an important mechanism in autoimmune demyelination, and its prevention with AMPA/kainate antagonists may prove to be an effective therapy for multiple sclerosis.

Multiple sclerosis: Re-expression of a developmental pathway that restricts oligodendrocyte maturation

During mammalian central nervous system (CNS) development, contact-mediated activation of Notch1 receptors on oligodendrocyte precursors by the ligand Jagged1 induces Hes5, which inhibits maturation of these cells. Here we tested whether the Notch pathway is re-expressed in the adult CNS in multiple sclerosis (MS), an inflammatory demyelinating disease in which remyelination is typically limited. We found that transforming growth factor-β1 (TGF-β1), a cytokine upregulated in MS, specifically re-induced Jagged1 in primary cultures of human astrocytes. Within and around active MS plaques lacking remyelination, Jagged1 was expressed at high levels by hypertrophic astrocytes, whereas Notch1 and Hes5 localized to cells with an immature oligodendrocyte phenotype, and TGF-β1 was associated with perivascular extracellular matrix in the same areas. In contrast, there was negligible Jagged1 expression in remyelinated lesions. Experiments in vitro showed that Jagged1 signaling inhibited process outgrowth from primary human oligodendrocytes. These data are the first to implicate the Notch pathway in the limited remyelination in MS. Thus, Notch may represent a potential target for therapeutic intervention in this disease.

GFAP Is Necessary for the Integrity of CNS White Matter Architecture and Long-Term Maintenance of Myelination

To investigate the structural role of glial fibrillary acidic protein (GFAP) in vivo, mice carrying a null mutation in GFAP were generated. In 7/14 mutant animals older than 18 months of age, hydrocephalus associated with white matter loss was detected. Mutant mice displayed abnormal myelination including the presence of actively myelinating oligodendrocytes in adults, nonmyelinated axons in optic nerve, and reduced myelin thickness in spinal cord. White matter was poorly vascularized and the blood-brain barrier was structurally and functionally impaired. Astrocytic structure and function were abnormal, consisting of shortened astrocytic cell processes, decreased septation of white matter, and increased CNS extracellular space. Thus, GFAP expression is essential for normal white matter architecture and blood-brain barrier integrity, and its absence leads to late-onset CNS dysmyelination.

Myelin antigen-specific CD8+ T cells are encephalitogenic and produce severe disease in C57BL/6 mice

Encephalitogenic T cells that mediate experimental autoimmune encephalomyelitis (EAE) are commonly assumed to be exclusively CD4+, but formal proof is still lacking. In this study, we report that synthetic peptides 35–55 from myelin oligodendrocyte glycoprotein (pMOG35–55) consistently activate a high proportion of CD8+ αβTCR+ T cells that are encephalitogenic in C57BL/6 (B6) mice. The encephalitogenic potential of CD8+ MOG-specific T cells was established by adoptive transfer of CD8-enriched MOG-specific T cells. These cells induced a much more severe and permanent disease than disease actively induced by immunization with pMOG35–55. CNS lesions in pMOG35–55 CD8+ T cell-induced EAE were progressive and more destructive. The CD8+ T cells were strongly pathogenic in syngeneic B6 and RAG-1−/− mice, but not in isogeneic β2-microglobulin-deficient mice. MOG-specific CD8+ T cells could be repeatedly reisolated for up to 287 days from recipient B6 or RAG-1−/− mice in which disease was induced adoptively with <1 × 106 T cells sensitized to pMOG35–55. It is postulated that MOG induces a relapsing and/or progressive pattern of EAE by eliciting a T cell response dominated by CD8+ autoreactive T cells. Such cells appear to have an enhanced tissue-damaging effect and persist in the animal for long periods.

Multiple sclerosis: altered glutamate homeostasis in lesions correlates with oligodendrocyte and axonal damage.

Glutamate excitotoxicity, recently demonstrated in an animal model of multiple sclerosis (MS), is evoked by altered glutamate homeostasis. In the present study, we investigated the major regulating factors in glutamate excitotoxicity by immunohistochemistry in MS and control white matter with markers for glutamate production (glutaminase), glutamate transport (GLAST, GLT‐1 and EAAT‐1), glutamate metabolism (glutamate dehydrogenase [GDH] and glutamine synthetase [GS]), axonal damage (SMI 32) and CNS cell types. Active MS lesions showed high‐level glutaminase expression in macrophages and microglia in close proximity to dystrophic axons. Correlation between glutaminase expression and axonal damage was confirmed experimentally in animals. White matter from other inflammatory neurologic diseases displayed glutaminase reactivity, whereas normals and noninflammatory conditions showed none. All three glutamate transporters were expressed robustly, mainly on oligodendrocytes, in normal, control and MS white matter, except for GLT‐1, which showed low‐level expression around active MS lesions. GS and GDH were present in oligodendrocytes in normal and non‐MS white matter but were absent from both active and chronic silent MS lesions, suggesting lasting metabolic impediments. Thus, imbalanced glutamate homeostasis contributes to axonal and oligodendroglial pathology in MS. Manipulation of this imbalance may have therapeutic import.

Proteomic analysis of active multiple sclerosis lesions reveals therapeutic targets

Understanding the neuropathology of multiple sclerosis (MS) is essential for improved therapies. Therefore, identification of targets specific to pathological types of MS may have therapeutic benefits. Here we identify, by laser-capture microdissection and proteomics, proteins unique to three major types of MS lesions: acute plaque, chronic active plaque and chronic plaque. Comparative proteomic profiles identified tissue factor and protein C inhibitor within chronic active plaque samples, suggesting dysregulation of molecules associated with coagulation. In vivo administration of hirudin or recombinant activated protein C reduced disease severity in experimental autoimmune encephalomyelitis and suppressed Th1 and Th17 cytokines in astrocytes and immune cells. Administration of mutant forms of recombinant activated protein C showed that both its anticoagulant and its signalling functions were essential for optimal amelioration of experimental autoimmune encephalomyelitis. A proteomic approach illuminated potential therapeutic targets selective for specific pathological stages of MS and implicated participation of the coagulation cascade.

Nervous tissue as an immune compartment: the dialect of the immune response in the CNS

Here, Zsuzsa Fabry and colleagues address the question of whether the unique cellular environment of the central nervous system (CNS) contributes to the observed differences in immunological functions between the CNS and other organs. In particular, they discuss the significance within the CNS of the blood-brain barrier, the nonconstitutive expression of major histocompatibility complex (MHC) molecules, the unusual set of potential antigen-presenting and effector cells, and the production of immune or neuromediators from various cellular sources.

The endocannabinoid anandamide protects neurons during CNS inflammation by induction of MKP-1 in microglial cells.

Endocannabinoids are released after brain injury and believed to attenuate neuronal damage by binding to CB(1) receptors and protecting against excitotoxicity. Such excitotoxic brain lesions initially result in primary destruction of brain parenchyma, which attracts macrophages and microglia. These inflammatory cells release toxic cytokines and free radicals, resulting in secondary neuronal damage. In this study, we show that the endocannabinoid system is highly activated during CNS inflammation and that the endocannabinoid anandamide (AEA) protects neurons from inflammatory damage by CB(1/2) receptor-mediated rapid induction of mitogen-activated protein kinase phosphatase-1 (MKP-1) in microglial cells associated with histone H3 phoshorylation of the mkp-1 gene sequence. As a result, AEA-induced rapid MKP-1 expression switches off MAPK signal transduction in microglial cells activated by stimulation of pattern recognition receptors. The release of AEA in injured CNS tissue might therefore represent a new mechanism of neuro-immune communication during CNS injury, which controls and limits immune response after primary CNS damage.

Late complications of immune deviation therapy in a nonhuman primate.

The administration of antigens in soluble form can induce antigen-specific immune tolerance and suppress experimental autoimmune diseases. In a marmoset model of multiple sclerosis induced by myelin oligodendrocyte glycoprotein (MOG), marmosets tolerized to MOG were protected against acute disease, but after tolerization treatment a lethal demyelinating disorder emerged. In these animals, MOG-specific T cell proliferative responses were transiently suppressed, cytokine production was shifted from a T helper type 1 (TH1) to a TH2 pattern, and titers of autoantibodies to MOG were enhanced. Thus, immune deviation can increase concentrations of pathogenic autoantibodies and in some circumstances exacerbate autoimmune disease.