Mechanisms of T-cell mediated inflammatory neurodegeneration

by Lukas Müller-Kirschbaum
Doctoral thesis
Date of Examination:2024-09-17
Date of issue:2025-04-04
Advisor:Prof. Dr. Alexander Flügel
Referee:Prof. Dr. Alexander Flügel
Referee:Prof. Dr. Christine Stadelmann-Nessler
Referee:Prof. Dr. Jürgen Wienands
Referee:Prof. Dr. Martin Weber
Referee:Prof. Dr. Ralf Heinrich
Referee:Prof. Dr. Oliver Wirths
Persistent Address: http://dx.doi.org/10.53846/goediss-11193

 

 

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Abstract

English

Multiple sclerosis (MS) is a T-cell mediated, chronic, demyelinating disease of the central nervous system (CNS). Neurodegeneration and axonal loss are associated with increased disability and disease progression. Animal models, adequately reflecting grey matter inflammation and cortical atrophy are required to dissect the mechanisms of T-cell mediated neuronal damage. In a recently developed Lewis rat model of autoimmune grey matter disease induced by anti-b-synuclein response, CD4+ effector T cell infiltration into the cerebral cortex is accompanied by neurodegeneration. In this model, direct contacts between T cells and neurons have been observed but their significance is unclear. In this study, we aimed to develop a research tool to enhance the affinity interaction between T cells and neurons in the context of neuroinflammation. To this end, we engineered T cells expressing a chimeric antigen receptor (CAR) directed against an epitope tag presented on neurons upon AAV transduction. Flow cytometry, qRT-PCR and time-lapse imaging of co-cultured binding partners were used to characterize the system in vitro. We found that established contacts between CAR T cells and epitope-tag expressing neurons activated CAR T cells and led to drastic motility reduction. Additionally, contact formation was associated with neuronal loss in vitro. Truncation of the CAR signaling domain did not alter binding of T cells to neurons, but abrogated artificial activation of T cells and mitigated neuronal damage. Neonatal adeno-associated virus (AAV) administration and adoptive transfer of pathogenic b- synuclein-reactive T cells and ovalbumin-reactive CAR T cells were used to translate the developed system to neuroinflammation in vivo. Bulk RNA sequencing showed that activation of CAR T cells induced increased disease severity via a direct interaction with epitope-tag-expressing cells. Further, intravital two photon microscopy confirmed motility changes of CAR T cells in the CNS parenchyma of AAV-infected animals. Bulk RNA sequencing in combination with single nuclei transcriptome analysis revealed direct effects on specific neuronal populations in epitope-tag-expressing animals. These neuronal cells upregulated inflammatory damage associated pathways at the expense of homeostatic processes. Taken together, in this study we have developed a research tool allowing us to enhance and investigate the affinity interactions of T cells and neurons in the context of grey matter inflammation. Our data suggest that close interactions of the investigated cell types contribute to the induction of pathological changes in neurons and increase disease severity.
Keywords: Multiple Sclerosis; Neuroimmunology; Experimental Autoimmune Encephalomyelitis; CAR T cell