Alzheimer's disease (AD) is the most common neurodegenerative disease, characterized by disturbances in memory, thinking, and behaviour. The pathology of AD involves a complex interplay of multiple mechanisms, limiting the efficacy of existing therapies that target a single mechanism. Compounds that act simultaneously on multiple pathological mechanisms therefore represent a promising approach for the treatment of AD. In our master's thesis, we investigated the efficacy of two dual butyrylcolinesterase (BChE) and p38α mitogen-activated protein kinase (p38α MAPK) inhibitors, namely KES-29 and GDK-767, in a cellular model of the inflammatory response, along with selective p38α MAPK and BChE inhibitors, KES-19 and GUK-1329, respectively. The toxicity of the inhibitors was assessed by evaluating their effects on the metabolic activity of BV2, SH-SY5Y, and HepG2 cells and by determining the percentage of dead BV2 cells after exposure to the inhibitors. We found that the inhibitor GUK-1329 was highly toxic even in the concentration range of 1–10 μM. In contrast, the inhibitors KES-29, GDK-767, and KES-19 only slightly decreased the metabolic activity of BV2 cells at 10 μM, without affecting the metabolic activity of SH-SY5Y and HepG2 cells at this concentration. Moreover, these inhibitors had no significant effect on the percentage of dead BV2 cells within the concentration range of 1–10 μM. Therefore, the protective effects of KES-29, GDK-767, and KES-19 were further tested within this concentration range. To investigate the potential protective effect of the inhibitors, we established and evaluated a cellular model of the inflammatory response by activating the microglia-like BV2 cell line with lipopolysaccharide (LPS). Using Western blotting (WB) and enzyme-linked immunosorbent assay (ELISA), we found that KES-29, GDK-767, and KES-19 effectively reduced the release of the inflammatory mediators interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α) from activated microglia. Notably, the dual inhibitors KES-29 and GDK-767 reduced cytokine release more than the selective inhibitor KES-19. Additionally, all three inhibitors attenuated the morphological changes in cells induced by LPS stimulation. Moreover, KES-29, GDK-767, and KES-19 were found to decrease LPS-induced caspase-3 activity in a concentration range of 1–10 μM, suggesting a protective role against apoptosis. However, it is important to note that only GDK-767 and KES-19 at a concentration of 5 μM significantly reduced overall cell survival. Our results suggest that the dual inhibitors KES-29 and GDK-767 and the selective inhibitor KES-19 effectively reduce LPS-induced inflammation in microglial cells, indicating their potential as novel agents for the treatment of inflammation-related neurodegenerative diseases such as AD.
|
