Ninjurin-1 (NINJ-1) is a protein that plays an important role in plasma membrane rupture during lytic cell death, during which DAMP (damage-associated molecular patterns) molecules are released from damaged cells and trigger the inflammatory process. In patients with chronic inflammatory diseases, complications may arise due to the release of inflammatory DAMP molecules resulting from plasma membrane disruption by NINJ-1, which may contribute to the progression of inflammatory diseases. In this thesis, we aimed to develop a novel approach to inhibit NINJ-1 activity. We used antisense oligonucleotides (ASO), as they enable precise and specific targeting and have also beenproven to be a safe and effective approach for regulation of gene expression. In the master’s thesis, we tested the effect of different ASO on the expression of NINJ-1 and developed various systems to evaluate ASO activity. Using the propidium iodide viability assay, we demonstrated that NINJ-1 overexpression triggers cell death in the HEK293T cell line. The activity of five different ASOs was tested using two approaches – first, we developed a luciferase reporter system, and second, we used a system based on the quantification of cell death with propidium iodide. The tested ASOs did not reduce NINJ-1 expression. In scope of this project, we also developed three additional approaches for indirect evaluation of ASO activity. While the indirect effects of ASOs on NINJ-1 expression cannot be monitored through the activation of inflammation-responsive promoters changes in NINJ-1 expression can be indirectly assessed via inflammatory cytokines secreted by immune cells that come into contact with NINJ-1-expressing cells.
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