Transcriptomic Insights into Neuroprotection of Nobiletin and Inhibitory on aβ_1-42 Aggregation

Background: Nobiletin (NOB), a flavone mainly from citrus peels, has drawn increasing attention for its potential protective effect on cognitive function. However, in most studies, either the model was not specifically for Alzheimer's disease (AD), or the action stage of nobiletin-rich extract as an intervention substance was not clearly defined, whether for prevention or treatment.

Method: To assess the prophylactic potential of NOB against AD, the Aβ_1-42 was first characterized in vitro, followed by validation in Caenorhabditis elegans (CL4176) and mice. CL4176 eggs were pretreated with NOB until the L3 stage, with 0.5 mM methylglyoxal induction at L4. Twelve-week-old male C57BL/6J mice were randomly assigned to four groups: (1) Sham (i.p. saline + i.c.v. saline), (2) Model (i.p. saline + i.c.v. Aβ1-42), (3) Positive (i.p. 2 mg/kg donepezil + i.c.v. Aβ1-42), and (4) NOB (i.p. 20 mg/kg NOB + i.c.v. Aβ_1-42). All mice received three weeks of indicated i.p. pretreatment prior to i.c.v. injection (5 μL/mouse).

Result: NOB demonstrated dual efficacy in inhibiting the aggregation of Aβ1-42 peptidesand promoting the disaggregationof Aβ1-42 protofibrils. Thioflavin T fluorescence assays and atomic force microscopy imaging showed that NOB inhibited Aβ_1-42 fibrillation and disaggregated preformed fibrils into amorphous clusters. Consistent with these findings, NOB pretreatment significantly reduced Aβ_1-42 plaque deposition and attenuated reactive oxygen species levels in C. elegans, suggesting a protective role against amyloid-induced proteotoxicity and oxidative stress. For mice, hippocampal transcriptomic analysis via principal component analysis indicating considerable variations among the Sham, Model, Donepezil, and NOB groups. NOB pretreatment modulated 890 differentially expressed genes (DEGs), with 824 up-regulated and 66 down-regulated. Dual volcano plot analysis identified 11 core DEGs. Furthermore, WGCNA highlighted strongly significant correlated with the turquoise module, for which KEGG analysis suggested pathways of neurodegeneration, ADand oxidative phosphorylation. Accordingly, TUNEL and Nissl staining demonstrated that NOB effectively attenuated neuronal injury and apoptosis. Additionally, ultrastructural imaging confirmed that NOB preserved mitochondrial integrity and myelin sheath structure, highlighting its comprehensive neuroprotective effects.

Conclusion: Our findings consistently demonstrated the potential anti-Alzheimer’s disease impact of pretreatment with nobiletin (a citrus flavone).