Modulation Of NLRP1-dependent Neuronal Proptosis For Alzheimer’s Disease Therapy

Background:Increasing evidence has shown the aberrant expression of inflammasome related proteins in Alzheimer’s disease (AD) brain; these proteins, including NLRPI inflammasome, are implicated in the execution of inflammatory response and pyroptotic death. Although current data are associated NLRP1 genetic variants with AD, the involvement of NLRP1 inflammasome in AD pathogenesis is still unknown. Objective: To investigate the hypothesis that amyloid-β (Aβ) induces NLRP1-dependent neuronal pyroptosis in models of AD, and targeting NLRPI inflammasome as a valid therapeutic approach for the neuroprotection in AD. Methods:1. The involvement of NLRP1 in Aβ induced neuronal pyroptosis was confirmed by silencing of NLRPI in primary cortical neurons. Cell viability and pyroptosis cells were determined by MTT assay and TUNEL assay. Cytotoxicity was determined by measuring the release of LDH. Levels of inflammatory cytokine IL-1β in cell culture supernatants were measured by ELISA kit. Levels of NLRP1 and caspase-1 activity were measured by immunoblotting. And the inflammasome activation was examined by ASC (apoptosis associated speck-like protein containing a CARD) speck visualization.2. NLRP1 in the NeuN-positive neurons of APPswe/PS1dE9 transgenic mice was detected by double immunofluorescence staining. Using in-vivo non-viral RNA interference methodology, cerebral NLRPI or caspase-1 expression in APPswe/PS 1 dE9 mice was knockdown. The spatial learning and memory performance was detected by Morris water maze. The distribution of AP was assessed by immunohistochemical analysis in brain sections. The neuronal pyroptosis was observed by TUNEL assay. And the neuronal morphology was evaluated by cresyl violet staining of nissl bodies in brain sections. Levels of caspase-1 activity and IL-1β were measured by immunoblotting. Results:1. Using APPswe/PS1dE9 transgenic mice and age- and background-matched wild-type mice, we screened the cerebral expression of NLRP1 in 3-6-. and 9-month-old mice and found that cerebral NLRP1 levels were upregulated in the NeuN-positive neurons of APPswe/PS1dE9 mice during aging.2. Our in-vitro studies further showed that increased NLRP1-mediated caspase-1-dependent ’pyroptosis’ in cultured cortical neurons in response to Aβ.Changing the levels of NLRP1 may protect against Aβ neurotoxicity through the pyroptosis mechanism in cultured neurons.3. We employed direct in-vivo infusion of non-viral small-interfering RNA (siRNA) to knockdown NLRP1 or caspase-1 in APPswe/PS1dE9 brain, and discovered that these NLRP1 or caspase-1 deficiency mice resulted in significantly reduced neuronal pyroptosis and reversed cognitive impairments. Conclusion:Our findings indicate an important role for NLRP1/caspase-1 signaling in AD progression, and point to the modulation of NLRP1 inflammasome as a promising strategy for AD therapy.