Einfluss der immunsuppressiven Therapie auf Reifungsstadien und TCR-Vβ-Gebrauch bei der autoimmunen Frühmyasthenie
Ziel der vorliegenden Arbeit ist die Untersuchung von TCR-Vβ-Gebrauch und Thymozyten-Reifungsstadien bei Patienten mit autoimmuner Frühmyasthenie. Auf diese Weise soll nach Hinweisen dafür gesucht werden, ob bei der autoimmunen Frühmyasthenie Mechanismen der zentralen oder der peripheren Toleranz ge...
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The aim of this work is the investigation of TCR Vβ usage and thymocyte maturation stages in patients with autoimmune “early-onset” myasthenia gravis (MG). Thus, it shall be looked for evidence, whether mechanisms of central or peripheral tolerance are disturbed in autoimmune “early-onset” myasthenia gravis. In the literature there are clues for intrathymic changes as well as peripheral regulatory dysfunctions, which are involved in the pathogenesis of autoimmune “early-onset” myasthenia gravis. In order that the pathognomonic nAChR antibodies can be generated by plasma cells, B cells (which then differentiate to plasma cells) must be stimulated by activated and clonal expanded T cells. Thymus and blood lymphocytes from 40 patients with nAChR antibody positive, “early-onset” myasthenia gravis and 22 immunological healthy probands, who had to undergo cardiac surgery, were examined by fluorescence activated cell sorting (FACS). TCR Vβ expansions in the blood and in the thymus were investigated as well as thymocyte maturation stages. In order to examine the pathophysiological influence of immunosuppressive medication the group of MG patients was divided into two subgroups: the ones who had not been treated immunosuppressively at the time of thymectomy on the one hand and those who had taken corticosteroids or azathioprine at that time on the other hand. Vβ chain analysis showed no significant differences in the thymus concerning the number of Vβ chain expansions when the three study subgroups have been compared with each other. In the blood a significant higher number of Vβ expansions was found on CD4+ positive lymphocytes in both the immunosuppressive treated and the immunosuppressive naïve MG subgroups when compared with the normal controls. Comparing the two MG subgroups with each other showed no significant differences with regard to the number of Vβ expansions on CD4+ positive lymphocytes. Concerning CD8+ lymphocytes no significant differences have been found between the three study groups. The fact that a significant higher number of Vβ expansions was found on CD4+ positive lymphocytes of MG patients when compared with normal controls and that no such differences could be detected in the thymus argues for a disturbation of peripheral tolerance in patients with autoimmune “early-onset” myasthenia gravis. Immunosuppressive medication seems to have no influence on this disturbation of peripheral tolerance, because the number of Vβ expansions on CD4+ positive lymphocytes did not differ significantly between the immunosuppressive treated and the immunosuppressive naïve MG subgroup. Investigation of thymocyte maturation stages showed an increased appearance of the late double positive stage in immunosuppressive naïve MG patients when compared with normal controls (1.38±0.71% vs. 1.06±1.24%, p<0.0447). The early CD4+ single positive stage also occurs more frequently in immunosuppressive naïve MG patients when compared with normal controls (3.49±1.49% vs. 2.21±2.08%, p<0.0413). Comparing the two MG subgroups with each other showed a decreased appearance of the mentioned stages in the immunosuppressively treated MG subgroup (late double positive: 0.74±0.64% vs. 1.38±0.71%, p<0.0024; early CD4+ single positive: 2.51±2.24% vs. 3.49±1.49%, p<0.0313). Because negative selection of autoreactive thymocytes exactly takes place in these stages there is evidence for a disturbation of central tolerance in autoimmune “early-onset” myasthenia gravis and an influence of corticosteroids or azathioprine that supports negative selection in the thymus. Thus, this work argues for a disturbation of the central tolerance in patients with autoimmune “early-onset” MG with succeeded expansion of CD4+ positive lymphocytes. These expansions seem to take place after the lymphocytes have emigrated from the thymus (disturbation of the peripheral tolerance). Hence, therapeutic attempts for encouraging both the peripheral (i. e. facilitation of a peripheral regulatory milieu) and the central tolerance (e. g. agents which interfere with thymocyte intracellular signaling or mechanisms of antigen presentation) come into consideration in patients with autoimmune “early-onset” myasthenia gravis.