Background: Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by β-amyloid (Aβ) accumulation, neuroinflammation, and neuronal loss. Interferon gamma-inducible protein 16 (IFI16) regulates inflammatory processes in AD, while ubiquitin-specific peptidase 16 (USP16) has been shown to ameliorate memory deficits in AD models. This study aimed to elucidate the interplay between USP16 and IFI16 in AD pathogenesis.

Methods: Differentially expressed genes associated with AD were identified using dataset GSE129296. Glial cells were exposed to β-amyloid peptide (Aβ)1–42 to establish an in vitro AD model, followed by indirect co-culture with neuronal cells. Neuronal viability and apoptosis were evaluated using the Cell Counting Kit-8 assay and flow cytometry, respectively. Levels of tumor necrosis factor (TNF)-α and interleukin (IL)-1β in model glial cells were quantified by Enzyme-linked immunosorbent assay (ELISA). A potential deubiquitylating enzyme of IFI16 was predicted using Ubibrowser and confirmed by co-immunoprecipitation, while the ubiquitination levels of IFI16 were assessed. The expression of USP16/IFI16 and toll-like receptor 4 (TLR4)/NLR family pyrin domain containing 3 (NLRP3) was determined by reverse transcription-quantitative PCR (RT-PCR) and western blotting.

Results: IFI16 was markedly upregulated in AD. Silencing of IFI16 attenuated Aβ1–42-induced proinflammatory responses in glial cells, reduced neuronal apoptosis, and enhanced neuronal viability, accompanied by decreased inflammatory cytokine levels and downregulation of TLR4/NLRP3 expression. USP16, identified as the deubiquitylating enzyme of IFI16, was significantly elevated in Aβ1–42-stimulated glial cells, and its silencing reduced IFI16 expression. Furthermore, IFI16 overexpression reversed the effects of USP16 knockdown on cell survival, apoptosis, and inflammation.

Conclusion: USP16 depletion alleviates inflammation and apoptosis in AD cellular models through modulation of IFI16, suggesting that USP16 may serve as a promising therapeutic target for AD.