Charakterisierung der humoralen Immunantwort bei der Multiplen Sklerose

Multiple sclerosis (MS) is a chronic inflammatory and demyelinating disease of the central nervous system (CNS) with an yet unknown etiology. Infiltration of immune cells in the CNS and histopathological changes are key features in MS. Although it is likely that the immune system plays an important role in disease process it is still unknown which immune cells are potentially relevant in disease manifestation and progression. Since inflammatory parameters in the cerebrospinal fluid (CSF) correlate with the extent of CNS lesions, CSF abnormalities could reflect lesion pathology. In this study, the distribution of various immune cells and proteins in the CSF and peripheral blood was compared between MS patients and patients with non-inflammatory neurological diseases (NIND) in order to search for CSF abnormalities in MS. The analysis revealed no significant differences in peripheral blood between MS and control patients. In contrast, significant changes were observed in the CSF. In particular, MS patients showed an activation of the humoral immune response which was reflected by a higher percentage of B cells and plasmacells in patient?s CSF and an increased intrathecal immunoglobulin-G (IgG)-synthesis. Furthermore, the analyses enclosed three distinct CSF patterns, which remained stable during different phases of disease: 1. a B cell dominant pathology with high percentages of B cells, low percentages of monocytes and an increased intrathecal IgG-synthesis, 2. a monocyte-dominant pathology with low numbers of B cells, high numbers of monocytes and low IgG-levels and 3. an intermediate pathology with equal numbers of B cells and monocytes and a moderate IgG-synthesis. Further analyses revealed an association between CSF pathology and disease progression. A predominance of B cells was associated with a faster disease progression, whereas a predominance of monocytes was found in patients with slower progression. This study describes the existence of heterogeneity in CSF cytology in MS patients and support the idea of different pathomechanisms in MS. The different CSF patterns may allow for the first time the stratification of MS patients according to their CSF pathology and a prediction of individual disease progression. A hallmark of the MS is the presence of intrathecal antibody synthesis and the occurence of oligoclonal IgG bands (OCB) in the CSF, which are found in 95% of the MS patients but rarely in healthy controls (<1%). These OCB are also found in infectious CNS diseases. In these disorders the OCB are specific for the disease causing pathogen. In MS, the target antigen is still unknown. The aim of the second part of this study was to identify the antigen specificity of the highly focused antibody response. A novel protein-array technology based on a cDNA library of the human brain enabled us to screen for the antibody response against more than 37000 different proteins. Potential candidate antigens which were identified with CSF of MS patients but not with control CSF were selected for large ELISA-experiments to determine the frequency of positive immunreactivities in large MS and control groups. 10 candidate antigens showed significant higher immunreaktivities in MS patients. Interestingly, the different proteins showed overlappings in immunreactivity assuming that all proteins of each group share the same epitope. The underlying epitopes were defined by peptidescan analysis and amino acid substitutional assays. The analyses identified two proteins of the Epstein-Barr virus (EBV): EBNA-1 and BRRF-2. To investigate the immunreactivity against these EBV-Proteins, the BRRF-2 protein was recombinantly expressed in E.coli using a cloning strategy. The analyses revealed a significant higher immunreactivity against EBNA-1 and BRRF-2 in CSF and serum of MS patients compared to NIND patients and patients with other inflammatory neurological diseases (OIND). Furthermore, a BRRF-2 specific intrathecal IgG synthesis was observed. Of significant importance, we could show that the OCB of MS patients were specific for both EBV proteins. The data strongly support the role of EBV in the pathogenesis of MS.