Authors:	Beutgen, Vanessa Madeleine
Title:	Immunoproteomic analysis of serological autoantibodies in glaucoma
Online publication date:	9-Apr-2021
Year of first publication:	2021
Language:	english
Abstract:	As a neurodegenerative disease, glaucoma is not curable with currently available treatments. Nowadays, only the prevention of further vision loss by delaying disease progression is possible. The focus for glaucoma management lies in the lowering of the intraocular pressure by eye drops, laser treatment or surgery. A timely start of treatment is crucial to prevent glaucoma-caused blindness. Therefore, early diagnosis plays a particularly important role. The pathomechanisms of glaucoma are not conclusively discovered, but different studies have shown an involvement of immunological processes in the pathogenesis of glaucoma. One aspect of glaucoma pathogenesis is related to processes of humoral immunity, resulting in altered serological levels of autoantibodies. In this scope, altered abundance of several specific autoantibodies to ocular antigens, in retina and optic nerve, were found in serum and aqueous humour of glaucoma patients. The trabecular meshwork (TM) is a critical site in glaucoma pathogenesis, too. It is subjected to major structural changes in the course of the disease. Dysregulation of extracellular matrix deposition, the actin cytoskeleton and cell-cell / cell-matrix connections enhance tissue rigidity and thereby increase aqueous humour outflow resistance. This lead to the hypothesis that these major structural and biomechanic alterations in the glaucomatous TM are accompanied by shifts in the abundance of autoantibodies in natural autoimmunity as well. The aim of this doctoral thesis was the identification of antigens in the TM and the investigation of their possible association with glaucoma, using different immunoproteomic techniques. The potential of these autoantibodies as diagnostic biomarker candidates was also evaluated. The research focused on primary open-angle glaucoma (POAG) as the most common form of the disease, however, altered levels of serological antibodies were also analysed for other forms of open-angle glaucoma. Although, it remains unclear if these changed autoantibody levels are cause or consequence of glaucoma, the identification of antibodies to self-antigens in serum is a promising approach towards the discovery of diagnostic and prognostic biomarkers. These biomarkers could facilitate the decision making upon treatment options of ophthalmologists and promote early diagnosis of the disease. With the application of “serological proteome analysis” (SERPA) and ensuing protein microarray analysis, three new biomarker candidates were identified. Autoantibodies to Caldesmon, Voltage-dependent anion-selective channel protein 2 and Phosphoglycerate mutase 1 showed significantly higher levels in POAG patients (p < 0.05) in comparison to a non-glaucomatous control group. Antibody levels of these markers and to two previously known glaucoma-related autoantigens, mitochondrial 60 kDa heat shock protein and Vimentin, were used to classify POAG patient from controls via artificial neural networks. With the panel of these potential biomarkers it was possible to classify the groups with a sensitivity of 81% and a specificity of 93%. In another approach “mass spectrometry-based antibody-mediated identification of autoantigens” (MS-AMIDA) was used for initial autoantibody profiling. Sixty-six autoantigens were identified as targets of the natural autoantibodies. Twentyone of these showed an association with POAG. Subsequent protein microarray validation revealed a significant (p < 0.05) increase of autoantibodies to Threonine--tRNA ligase 1, Complement component 1 Q subcomponent-binding protein and Paraneoplastic antigen Ma2 in POAG patients compared to a non-glaucomatous control group. Furthermore, an association of the identified glaucoma-related autoantigens with the “platelet-derived growth factor receptor β” pathway that is known to be involved in glaucoma-related fibrosis in the trabecular meshwork was discovered. Data retrieved from an explorative microarray experiment analysing sera of POAG, normal tension glaucoma (NTG) and pseudoexfoliation glaucoma (PEXG) patients versus a control group, revealed altered levels of serological autoantibodies in all three manifestations of OAG. The analysis showed especially low levels of anti-clathrin antibodies in sera of PEXG patients and significantly higher levels of β-2 adrenergic receptor antibodies in POAG and NTG patients. In summary, the results in this doctoral thesis provide novel biomarker candidates and give new insights into the characteristics of autoantibodies in glaucoma., whilst demonstrating these to be part of natural immunity. Furthermore, pathways with an enrichment of immunogenic proteins were identified, which delivers additional findings to our knowledge of the natural autoimmunity. These results form the basis for further analysis of autoantibodies in health and disease.
DDC:	570 Biowissenschaften 570 Life sciences 610 Medizin 610 Medical sciences
Institution:	Johannes Gutenberg-Universität Mainz
Department:	FB 04 Medizin FB 10 Biologie
Place:	Mainz
ROR:	https://ror.org/023b0x485
DOI:	http://doi.org/10.25358/openscience-5718
URN:	urn:nbn:de:hebis:77-openscience-dc646185-e96f-4940-8d83-f4ca27ce0a3e9
Version:	Original work
Publication type:	Dissertation
License:	In Copyright
Information on rights of use:	http://rightsstatements.org/vocab/InC/1.0/
Extent:	X, 127 Seiten, Illustrationen, Diagramme
Appears in collections:	JGU-Publikationen