Abstract
The present study was designed to investigate the role of β‐amyloid (Aβ1‐42) in inducing neuronal pyroptosis and its mechanism. Mice cortical neurons (MCNs) were used in this study, LPS + Nigericin was used to induce pyroptosis in MCNs (positive control group), and Aβ1‐42 was used to interfere with MCNs. In addition, propidium iodide (PI) staining was used to examine cell permeability, lactate dehydrogenase (LDH) release assay was employed to detect cytotoxicity, immunofluorescence (IF) staining was used to investigate the expression level of the key protein GSDMD, Western blot was performed to detect the expression levels of key proteins, and enzyme‐linked immunosorbent assay (ELISA) was utilized to determine the expression levels of inflammatory factors in culture medium, including IL‐1β, IL‐18 and TNF‐α. Small interfering RNA (siRNA) was used to silence the mRNA expression of caspase‐1 and GSDMD, and Aβ1‐42 was used to induce pyroptosis, followed by investigation of the role of caspase‐1‐mediated GSDMD cleavage in pyroptosis. In addition, necrosulfonamide (NSA), an inhibitor of GSDMD oligomerization, was used for pre‐treatment, and Aβ1‐42 was subsequently used to observe the pyroptosis in MCNs. Finally, AAV9‐siRNA‐caspase‐1 was injected into the tail vein of APP/PS1 double transgenic mice (Alzheimer's disease mice) for caspase‐1 mRNA inhibition, followed by observation of behavioural changes in mice and measurement of the expression of inflammatory factors and pyroptosis‐related protein. As results, Aβ1‐42 could induce pyroptosis in MCNs, increase cell permeability and enhance LDH release, which were similar to the LPS + Nigericin‐induced pyroptosis. Meanwhile, the expression levels of cellular GSDMD and p30‐GSDMD were up‐regulated, the levels of NLRP3 inflammasome and GSDMD‐cleaved protein caspase‐1 were up‐regulated, and the levels of inflammatory factors in the medium were also up‐regulated. siRNA intervention in caspase‐1 or GSDMD inhibited Aβ1‐42‐induced pyroptosis, and NSA pre‐treatment also caused the similar inhibitory effects. The behavioural ability of Alzheimer's disease (AD) mice was relieved after the injection of AAV9‐siRNA‐caspase‐1, and the expression of pyroptosis‐related protein in the cortex and hippocampus was down‐regulated. In conclusion, Aβ1‐42 could induce pyroptosis by GSDMD protein, and NLRP3‐caspase‐1 signalling was an important signal to mediate GSDMD cleavage, which plays an important role in Aβ1‐42‐induced pyroptosis in neurons. Therefore, GSDMD is expected to be a novel therapeutic target for AD.
Highlights
Alzheimer's disease (AD) is a neurodegenerative disease characterized by cognitive decline, which is the leading cause of dementia.[1,2] The pathological features of AD mainly include the senile plaque (SP) formed by the deposition of β-amyloid (Aβ) in the extracellular space,[3] neurofibrillary tangles (NFT) formed by the abnormally phosphorylated Tau protein aggregation, vascular amyloidosis and loss of neurons in the cortex and hippocampus.[4]
Aβ1-42 could induce pyroptosis by GSDMD protein, and NLRP3-caspase-1 signalling was an important signal to mediate GSDMD cleavage, which plays an important role in Aβ1-42-induced pyroptosis in neurons
Mice cortical neurons (MCNs) in the Aβ1-42 group were incubated with 25 μmol/L Aβ1-42, and MCNs were transfected with Small interfering RNA (siRNA)-caspase-1 and siRNA-GSDMD
Summary
Alzheimer's disease (AD) is a neurodegenerative disease characterized by cognitive decline, which is the leading cause of dementia.[1,2] The pathological features of AD mainly include the senile plaque (SP) formed by the deposition of β-amyloid (Aβ) in the extracellular space,[3] neurofibrillary tangles (NFT) formed by the abnormally phosphorylated Tau protein aggregation, vascular amyloidosis and loss of neurons in the cortex and hippocampus.[4]. Studies on AD have revealed the aggregation of glial cells around the SP in the brain tissue of patients and AD rats, subsequently causing inflammation, and the abnormally elevated expression of inflammatory factors, including interleukin-1 (IL-1), IL-6 and tumour necrosis factor-α (TNF-α), which further amplifies the inflammatory cascade.[10,11]. Aβ has been found to induce neuroinflammatory responses, central inflammation is mainly mediated by microglia.[8]. It remains uncovered whether Aβ directly causes inflammatory injury of nerve cells as well as the underlying mechanisms. In view of the role of pyroptosis in inflammatory response, in the present study, we investigated whether Aβ directly caused inflammatory injury of nerve cells as well as the underlying mechanisms, to further reveal the role of Aβ in AD-related inflammatory responses
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