Characterisation of Immune Response to Neurofilament Light in Experimental Autoimmune Encephalomyelitis

Autoimmunity to neuronal proteins occurs in several neurological syndromes where cellular and humoral responses are directed to surface, as well as intracellular antigens. Similar to myelin autoimmunity, pathogenic immune responses to neuroaxonal components such as neurofilaments may contribute to neurodegeneration in multiple sclerosis. We studied the immune response to the axonal protein neurofilament light (NF-L) in the experimental autoimmune encephalomyelitis (EAE) animal model of multiple sclerosis. To examine the association between T cells and axonal damage, pathology studies were performed on NF-L immunised mice. The interaction of T cells and axons was analysed by confocal microscopy of central nervous system tissues, and T-cell and antibody responses to immunodominant epitopes identified in Biozzi ABH (H2-A) and SJL/J (H2-A) mice. These epitopes, algorithm-predicted peptides and encephalitogenic motifs within NF-L were screened for encephalitogenicity. Confocal microscopy revealed both CD4 and CD8 T cells alongside damaged axons in the lesions of NF-L immunised mice. CD4 T cells dominated the areas of axonal injury in the dorsal column of spastic mice in which the expression of granzyme B and perforin was detected. Identified NF-L epitopes induced mild neurological signs similar to the observed with the NF-L protein, yet distinct from those characteristic of neurological disease induced with myelin oligodendrocyte glycoprotein. Our data suggest that CD4 T cells are associated with spasticity, axonal damage and neurodegeneration in NF-L immunised mice. In addition, defined T-cell epitopes in the NF-L protein might be involved in the pathogenesis of the disease. Introduction Multiple sclerosis (MS) is a chronic demyelinating and neurodegenerative disease of the central nervous system (CNS) widely considered to be due to aggressive, autoreactive T cells and antibodies to myelin (1-3). However, accumulating evidence shows that immune responses to neuronal and axonal proteins are also present in a wide range of neurodegenerative disorders including MS (4-6). That these responses may contribute to axonal and neuronal damage, pathological hallmarks of MS, is supported by observations that immunisation with neuronal antigens and transfer of antibodies directed to neuronal and axonal proteins induce neuronal damage in animals (7-10). The lack of expression of molecules of the major histocompatibility complex (MHC) class II on neurons indicates that neurons cannot activate CD4 cells in an antigen specific manner. However, neurons constitutively express or readily up regulate expression of MHC class I during inflammation, indicating that neurons may become targets for CD8 T cells (11). Conceptually, both CD4 and CD8 T cells could mediate attack on axons and neurons, either by direct contact via antigenindependent interactions or as a result of collateral damage (12). Activated T cells in the CNS are reported to produce cytotoxic molecules as well as glutamate, nitric oxide and reactive oxygen species that could contribute to the damage and progressive neurodegeneration observed in MS and other neurodegenerative diseases in which inflammation has been described (13-15). In addition, activation of B cells might lead to the production of specific antibodies to neuronal antigens that could also contribute to the damage and progressive neurodegeneration (16). To examine the mechanisms of autoimmunity to neurons we have developed a model of autoimmune induced axonal and neuronal damage following immunisation of mice with the neuronal cytoskeletal protein neurofilament light (NF-L) (10). 4