Apigenin enhances Nrf2-induced chaperone-mediated autophagy and mitigates α-synuclein pathology: Implications for Parkinson's disease therapy.

The aggregation of α-synuclein (SNCA) in dopaminergic neurons of the substantia nigra is a key factor in the pathogenesis of Parkinson's disease (PD). Despite years of drug discovery efforts targeting SNCA aggregation, no disease-modifying drugs have been approved to date. The failure of numerous clinical trials can be attributed, at least in part, to the difficulty in identifying potent compounds during preclinical investigations. Establish a screening approach based on molecular docking and autophagic flux detection to identify natural compounds from new perspectives of SNCA clearance and to explore its mechanism. Molecular docking technique combined with autophagic flux detection was performed for preliminary screening of flavonoids in PubChem and CHEBI databases. Western blotting was utilized to detect the levels of SNCA, chaperone-mediated autophagy (CMA)-associated proteins, apoptosis-related proteins, and neuroinflammatory biomarkers, alongside the assessment of phosphorylation status of proteins implicated in signaling cascades. JC-1 staining was used to measure the mitochondrial transmembrane potential (MMP). RNA-sequencing and Kyoto encyclopedia of genes and genomes/gene ontology (KEGG/GO) analysis were optimized to detect gene expression. PD mouse motor function was assessed using rotarod, pole, open field, footprint, and gait analyses. Immunofluorescence staining was employed to detect the expression of Nuclear factor erythroid 2-related factor 2 (Nrf2), dopaminergic neuronal deficits, microglia activation, and production of inflammatory factors. LysoTracker Red staining and pSIN-PAmCherry-KFERQ-NE plasmid were used to evaluate the lysosomal activity. pHrodo™ Green E.coli BioParticles™ were employed to measure phagocytosis activity. By molecular docking and autophagic flux detection, we evaluated the efficacy of flavone derivatives and identified apigenin (AP) as a candidate that activates CMA to promote SNCA clearance and thereby inhibits SNCA-induced neurotoxicity. AP inhibited apoptosis by promoting SNCA degradation through Nrf2-mediated CMA activation. Moreover, AP could also inhibit apoptosis via the Nrf2/extracellular regulated protein kinases (ERK) feedback loop that operates independently of CMA activation. Additionally, AP enhanced the phagocytosis capabilities of BV2 cells and inhibited SNCA-induced neuroinflammation, both in vitro and in vivo. AP activates CMA to promote the clearance of SNCA, thereby inhibiting SNCA-induced neurotoxicity. Nrf2 and its role in AP-mediated neuroprotection may provide new insights that target degradation pathways to counteract SNCA pathology in PD.