Bernhard Hemmer

A point mutation in PTPRC is associated with the development of multiple sclerosis

Multiple sclerosis (MS) is the most common demyelinating disease of the central nervous system. It is widely accepted that a dysregulated immune response against brain resident antigens is central to its yet unknown pathogenesis. Although there is evidence that the development of MS has a genetic component, specific genetic factors are largely unknown. Here we investigated the role of a point mutation in the gene (PTPRC) encoding protein-tyrosine phosphatase, receptor-type C (also known as CD45) in the heterozygous state in the development of MS. The nucleotide transition in exon 4 of the gene locus interferes with mRNA splicing and results in altered expression of CD45 isoforms on immune cells. In three of four independent case-control studies, we demonstrated an association of the mutation with MS. We found the PTPRC mutation to be linked to and associated with the disease in three MS nuclear families. In one additional family, we found the same variant CD45 phenotype, with an as-yet-unknown origin, among the members affected with MS. Our findings suggest an association of the mutation in PTPRC with the development of MS in some families.

Pathogenesis of multiple sclerosis: an update on immunology

&NA; Multiple sclerosis is characterized by demyelination and chronic inflammation of the central nervous system. Extensive studies in the animal model experimental autoimmune encephalomyelitis have suggested that multiple sclerosis is an autoimmune disorder mediated by myelin‐specific CD4 T cells secreting T helper type 1 cytokines and tumor necrosis factor alpha. This concept has been widely used to develop new experimental therapies. However, recent findings in both experimental autoimmune encephalomyelitis and multiple sclerosis question a simple CD4 T helper type 1 T cell paradigm and provide evidence for the role of various immune cells in the pathogenesis of experimental autoimmune encephalomyelitis and multiple sclerosis. In this paper we review recent progress and discuss the implications for new therapeutic strategies. Curr Opin Neurol 15:227‐231.

CD45 isoform expression in autoimmune myasthenia gravis.

In myasthenia gravis (MG), humoral and cellular immune mechanisms are involved in the autoimmune pathogenesis. In this study, we investigated the role of the CD45 molecule in MG, having recently reported an association in multiple sclerosis. CD45, a protein-tyrosine phophatase receptor type C (PTPRC), is essential for both thymic selection and peripheral activation of T and B cells. Our aims were to determine (a) the prevalence of a functional mutation in the CD45 gene (exon 4 77C M G; prevalence analysis), and (b) the distribution of memory (CD45RO+) and naïve (CD45RA+) T cells in the peripheral blood (subset analysis). T cells from 78 patients with generalised MG were stained with monoclonal antibodies against CD45RO, CD45RA, CD4 and CD8 and quantified by four-colour flow cytometry. The control panel for the prevalence analysis (a) consisted of 303 healthy individuals. (b) From those, 67 age- and sex-matched probands were randomly selected as controls for the subset analysis. Patients were stratified according to their MG onset age, thymic pathology and immunosuppressive treatment. Statistical analysis was performed by Fishers exact test, asymptotic h 2 test, the two-sided Mann-Whitney test and Spearmans correlation coefficient. As a result, the 77C M G mutation in exon 4 of the CD45 gene was found in 1 of 78 patients versus none of the 303 controls. Thus, no association was detected with this single nucleotide polymorphism in MG patients overall. Surprisingly, however, ratios of CD45RO+ to CD45RA+ T cells were lower among CD8+ T cells from patients with late-onset MG ( P =0.023). Thymoma patients also showed a similar trend among CD4+ and CD8+ T-cells, as expected. These differences were not related to immunosuppressive drug treatment or thymectomy (in the 67 informative patients). Since there is no other evidence for increased thymopoiesis in late-onset MG, we propose an altered subset balance in the circulation.

Dipeptidyl Peptidase IV (CD26): Role in T Cell Activation and Autoimmune Disease

The ectoenzyme dipeptidyl peptidase IV (DP IV; EC 3.4.14.5; CD26) has been shown to play a crucial role in T cell activation. In the present study, we show by flow cytometry and by enzymatic DP IV assay that myelin basic protein (MBP)-specific, CD4+ T cell clones (TCC) derived from patients with multiple sclerosis (MS) express high levels of DP IV/CD26. The enzymatic activity of resting TCC was found to be three to fourfold higher than on resting peripheral blood T cells and close to that of T cells 48 hours after PHA stimulation. The DP IV inhibitors Lys[Z(NO2)]-thiazolidide and Lys[Z(NO2)]-pyrrolidide suppress in a dose-dependent manner DNA synthesis and IFN-gamma, IL-4, and TNF-alpha production of the antigen-stimulated TCC. These data suggest that CD26 plays a role in regulating activation of autoreactive TCC. Further in vivo investigations will clarify, whether the inhibition of the enzymatic activity of DP IV could be a useful tool for therapeutic interventions in MS and/or other autoimmune diseases.

Identification of Target Antigens in CNS Inflammation by Protein Array Technique

Multiple Sclerosis (MS) is the most prevalent chronic inflammatory disease of the CNS, causing severe disability in a significant proportion of patients. Although many findings suggest that MS is caused by an immune response to proteins expressed in the CNS, the target antigens are still unknown. Among the candidates are self- and foreign proteins expressed in the CNS compartment. Here, we describe a new approach to dissect immune responses in the CNS. We applied a protein array based on a human brain cDNA library to decrypt the specificity of the local antibody response in MS. The macroarray, containing 37,000 proteins, enabled us to perform a large-scale screening for disease-associated antigens. Target proteins were further mapped to identify highaffinity ligands and possible mimics. Using this approach, we found MS-specific high-affinity antibody responses to two peptide sequences derived from Epstein-Barr virus (EBV) proteins. Several mimics with lower affinity also were identified. Subsequent analysis revealed an elevated and specific immune response in MS patients against both EBV proteins, suggesting a putative role of EBV in the pathogenesis of MS. The study demonstrates that protein arrays can be successfully applied to identify disease-associated antibody responses in neuroinflammatory diseases.