RIPK3 activation promotes DAXX‐dependent neuronal necroptosis after intracerebral hemorrhage in mice

• Published in: CNS neuroscience & therapeutics 2024-01, Vol.30 (1), p.e14397-n/a
• Main Authors: Bai, Qingqing, Wang, Shuoyang, Rao, Dongmei, Zhou, Zhiming, Wang, Jianfei, Wang, Qi, Qin, Yu, Chu, Zhaohu, Zhao, Shoucai, Yu, Dijing, Xu, Yang
• Format: Article
• Language: English
• Subjects: Animals; Antibodies; Apoptosis-inducing factor; Biotechnology; Brain research; Cell death; Daxx protein; death domain‐associated protein; Enzyme-linked immunosorbent assay; Ethics; Hemorrhage; Immunofluorescence; Immunoprecipitation; intracerebral hemorrhage; Ischemia; Kinases; Localization; Necroptosis; neurobehavioral deficits; Original; Proteins; receptor‐interacting protein kinase 3; Signal transduction; Statistical analysis; Stroke; Traumatic brain injury; Water content; Western blotting
• ISSN: 1755-5930; 1755-5949; 1755-5949
• DOI: 10.1111/cns.14397

Background Necroptosis induced by receptor‐interacting protein kinase 3 (RIPK3) is engaged in intracerebral hemorrhage (ICH) pathology. In this study, we explored the impact of RIPK3 activation on neuronal necroptosis and the mechanism of the death domain‐associated protein (DAXX)‐mediated nuclear necroptosis pathway after ICH. Methods Potential molecules linked to the progression of ICH were discovered using RNA sequencing. The level of DAXX was assessed by quantitative real‐time PCR, ELISA, and western blotting. DAXX localization was determined by immunofluorescence and immunoprecipitation assays. The RIPK3 inhibitor GSK872 and DAXX knockdown with shRNA‐DAXX were used to examine the nuclear necroptosis pathway associated with ICH. Neurobehavioral deficit assessments were performed. Results DAXX was increased in patients and mice after ICH. In an ICH mouse model, shRNA‐DAXX reduced brain water content and alleviated neurologic impairments. GSK872 administration reduced the expression of DAXX. shRNA‐DAXX inhibited the expression of p‐MLKL. Immunofluorescence and immunoprecipitation assays showed that RIPK3 and AIF translocated into the nucleus and then bound with nuclear DAXX. Conclusions RIPK3 revitalization promoted neuronal necroptosis in ICH mice, partially through the DAXX signaling pathway. RIPK3 and AIF interacted with nuclear DAXX to aggravate ICH injury. RIPK3 induces AIF expression and nucleus translocation after ICH. Then, RIPK3 and AIF were combined with DAXX, respectively. DAXX‐RIPK3 and DAXX‐AIF complexes contribute to MLKL phosphorylation. RIPK3 inhibitor GSK872 suppresses the necroptosis cascade and attenuates AIF, DAXX, and MLKL expression. When RIPK3 is inhibited, DAXX‐RIPK3 and DAXX‐AIF binding disappeared.