Wirkungen der natürlich vorkommenden α-Synuclein-Autoantikörper auf toxische α-Synuclein-Fragmente

Das idiopathische Parkinson-Syndrom (IPS) gehört zu den häufigsten neurodegenerativen Erkrankungen und trotz intensiver Bemühungen stehen bislang noch keine kausalen Therapiemöglichkeiten zur Verfügung. Lewy-Körperchen, eine der typischen Veränderungen in überlebenden nigralen Neuronen, gelten als p...

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Bibliographic Details
Main Author: Rabenstein, Monika
Contributors: Dodel, R. (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Language:German
Published: Philipps-Universität Marburg 2014
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Table of Contents: The idiopathic Parkinson’s Disease (PD) is a very common neurodegenerative disorder. There is no causal treatment available so far. Lewy bodies, the characteristic feature in surviving nigral neurons, are one of the pathologic hallmarks in PD. Aggregates of fibrillated α-Synuclein (α-Syn), a 140 amino-acids-long protein that is mainly expressed at presynaptic terminals in the CNS, are the major components of Lewy bodies. Recently, natural occurring autoantibodies against α-Syn (nAB α-Syn) were de- tected in the peripheral blood of PD patients and controls (Papachroni et al., 2007). In our group, we affinity-purified nABs α-Syn from intravenous immuno- globulins (IVIG) for further characterization. Since the physiological function of the nAB α-Syn in the human organism is still poorly known, the aim of this project was to specify the interaction of the nABs α-Syn with α-Syn-fragments for providing better understanding and possible treatment application of the nABs α-Syn. This was tested with truncated α-Syn-fragments representing the functional regions of the protein: residues 1-60 (N-terminal domain), residues 1-95 (N- terminal domain + NAC-region), residues 61-140 (NAC-region + C-terminal domain), residues 96-140 (C-terminal domain) and NACP112 (alternative splicing-induced deletion variant lacking amino acids 103-130). The fragments were either freshly prepared or aged for 7 days. First, the epitop(s) on α-Syn for the nAB α-Syn should to be determined. Therefore, binding properties of nAB α-Syn and α-Syn-fragments were analysed in antigen-antibody-binding-assays such as ELISA and Dot Blot. However, neither epitopes for the nAB α-Syn nor clear binding characteristics of the nAB α-Syn to the α-Syn-fragment regions could be identified. The differing binding properties in the assays points to the coexistence of linear and conformational epitopes. Second, the influence of the α-Syn-fragments on cell integrity and viability was tested on primary murine cortical neurons and primary murine microglia using LDH- and MTT-assays. The fragments 1-95, 61-140 and α-Syn 112 decreased cell viability in the primary neuronal culture as measured by MTT-assay. These cytotoxic α-Syn-fragments were subsequently preincubated with the nAB α-Syn-fragments and then added to the primary neuronal culture. There was a rescue effect on the α-Syn-fragments 1-95 and α-Syn 112 as shown by increased cell viabilty. Though, the same rescue effect was attained by preincubation with IVIG. Third, α-Syn-fragments added extracellularly to a primary murine micoglia culture were compared in their ability to induce the release of the proin- flammatory cytokines (TNFα and IL-6). Nearly all α-Syn-fragments (except for 1-60 for IL-6) induced the release of these two cytokines. Subsequently, the impact of the nAB α-Syn on the cytokine realease was determined. There was a highly significant reduction of the TNFα- and IL-6 release by preincubation with the nAB α-Syn (except for 61-140 for TNFα). These results underline for the first time the possible anti-inflammatory effect of the nAB α-Syn on α-Syn-induced microglia activation. Furthermore the findings point to a possible neuroprotective potential for the nAB α-Syn and similarly for IVIG. The overactivation of microglia is an important pathomechanism in PD. Modulating the microglia activity, for example by blocking the release of proinflammatory cytokines with the nAB α-Syn, may therefore provide an important therapeutic target. Further research has to be carried out for the exploration of the nAB α-Syn anti-inflammatory potential.