Abstract
Alzheimer’s disease (AD) is thought to be caused by amyloid-β (Aβ) accumulation in the central nervous system due to deficient clearance. The aim of the present study was to investigate the effect of ganoderic acid A (GAA) on Aβ clearance in microglia and its anti-AD activity. Aβ degradation in BV2 microglial cells was determined using an intracellular Aβ clearance assay. GAA stimulated autophagosome formation via the Axl receptor tyrosine kinase (Axl)/RAC/CDC42-activated kinase 1 (Pak1) pathway was determined by Western blot analyses, and fluorescence-labeled Aβ42 was localized in lysosomes in confocal laser microscopy images. The in vivo anti-AD activity of GAA was evaluated by object recognition and Morris water maze (MWM) tests in an AD mouse model following intracerebroventricular injection of aggregated Aβ42. The autophagy level in the hippocampus was assayed by immunohistochemical assessment against microtubule-associated proteins 1A/1B light-chain 3B (LC3B). Intracellular Aβ42 levels were significantly reduced by GAA treatment in microglial cells. Additionally, GAA activated autophagy according to increased LC3B-II levels, with this increased autophagy stimulated by upregulating Axl and Pak1 phosphorylation. The effect of eliminating Aβ by GAA through autophagy was reversed by R428, an Axl inhibitor, or IPA-3, a Pak1 inhibitor. Consistent with the cell-based assay, GAA ameliorated cognitive deficiency and reduced Aβ42 levels in an AD mouse model. Furthermore, LC3B expression in the hippocampus was up-regulated by GAA treatment, with these GAA-specific effects abolished by R428. GAA promoted Aβ clearance by enhancing autophagy via the Axl/Pak1 signaling pathway in microglial cells and ameliorated cognitive deficiency in an AD mouse model.
Highlights
Introduction published maps and institutional affilAlzheimer’s disease (AD) is a progressive and nonreversible disorder characterized by impairment of memory and cognition
FITC-labeled Aβ42 is taken up and degraded by microglia, and after Aβ42 degradation, the fluorophore remains in the cells; total cellular fluorescence correlates to total Aβ42 uptake and can be monitored by flow cytometry [31]
The effect of ganoderic acid A (GAA) in Aβ42-treated mice was blocked by the Axl-specific inhibitor, R428 (Figure 4B–J and Figure S3A), consistent with the in vitro study. These results indicated that GAA ameliorated cognitive deficits and decreased the accumulation of Aβ42 accumulation by promoting autophagy in an Axl-dependent manner
Summary
Alzheimer’s disease (AD) is a progressive and nonreversible disorder characterized by impairment of memory and cognition. Aggregation of amyloid-β (Aβ), the primary component of senile plaques, is the critical factor of AD pathology. The amyloid hypothesis assumes that the Aβ peptide is the causative agent in AD and is strongly supported by data from rare autosomal dominant forms of AD [2]. Numerous studies demonstrate that AD might be caused by mutations in three genes (APP, PSEN1, and PSEN2), involved in Aβ production [3]. The development of late-onset AD appears to arise from the failure of Aβ-clearance mechanisms rather than overproduction of the peptide [6]. The clearance of intracellular Aβ occurs through proteolysis by a family of amyloid-degrading enzymes (ADEs) or through other iations
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