Perilla-derived chalcone inhibits α-synuclein fibrillization and prevents aggregate-induced disruption of intracellular α-synuclein conformation in neuronal cells.

The pathological aggregation of α-synuclein (α-syn) is a hallmark of Parkinson's disease, where extracellular α-syn aggregates can disrupt intracellular α-synuclein conformation in neurons. In this study, we investigated the impact of 2',3'-dihydroxy-4',6'-dimethoxychalcone (DDC), isolated from green perilla, on α-syn fibrillization and subsequently on intracellular α-synuclein conformation. DDC concentration-dependently inhibited α-syn fibrillization in vitro, as determined by thioflavin S fluorescence and blue native polyacrylamide gel electrophoresis. DDC also disaggregated preformed α-syn fibrils, as indicated by decreased thioflavin S fluorescence. Structure-activity relationship analysis revealed that both the 2',3'-dihydroxy and 4',6'-dimethoxy substituents on the B-ring contributed to the inhibitory effect of DDC on α-syn fibrillization. Exposure of SH-SY5Y cells overexpressing α-synuclein to extracellular aged α-syn (i.e., fibrillized α-syn prepared by shaking) significantly reduced diffuse C-terminal α-syn immunoreactivity. Co-incubation of DDC during the fibrillization process preserved this diffuse staining, whereas adding it after fibrillization to preformed α-syn fibrils further diminished it. However, the direct addition of 30 μM DDC to the culture medium significantly attenuated the loss of diffuse C-terminal α-syn immunoreactivity caused by aged α-syn, likely due to a higher DDC-α-syn ratio. These findings demonstrate that DDC not only inhibited α-syn aggregation but also prevented the aggregate-induced conformational disruption of intracellular α-synuclein, highlighting its potential as a therapeutic agent against synucleinopathies.