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Inhibition of FOXD3 O-GlcNAc Modification Ameliorates Spinal Cord Injury by Promoting STUB1-Mediated Ubiquitination
1Department of Neurosurgery, the 1st Affiliated Hospital, Jiangxi Medical College, Nanchang University, 17 Yongwai Street, Nanchang, 330006, Jiangxi, China.
Abstract:
Spinal cord injury (SCI) is a serious complication of spinal fractures and/or dislocations, characterized by sensory and motor dysfunction in the trunk and limbs. The pathogenesis of SCI is highly complex and remains poorly understood. The role of O-GlcNAc modification and FOXD3 in SCI was studied in this study. The cell and animal models of SCI were established by H2O2 stimulation and heavy object impact method, respectively. HE and Nissl staining were used to analyze pathological changes and neuronal loss in the spinal cord tissues. The motor ability of rats was assessed by BBB score, ladder climbing, and grid climbing tests. Cell viability and apoptosis were assessed by CCK8, flow cytometry, and TUNEL staining, respectively. Co-IP assay detected O-GlcNAc modification level of FOXD3 protein. The interaction between FOXD3 and STUB1 promoter was analyzed by dual luciferase reporter gene and ChIP assays. O-GlcNAc modification level was significantly elevated in the cell and animal models of SCI. O-GlcNAc modification increased both the protein stability and expression of FOXD3. O-GlcNAc modification inhibition or FOXD3 knockdown reduced oxidative stress damage and apoptosis in H2O2-treated PC12 cells. Moreover, FOXD3 mediated transcriptional inhibition of STUB1, and STUB1 induced HMGB1 ubiquitination and degradation in PC12 cells. STUB1 knockdown or HMGB1 overexpression negated the protective effects of FOXD3 knockdown on H2O2-mediated oxidative stress damage and apoptosis in PC12 cells. Inhibiting the O-GlcNAc modification of FOXD3 alleviated oxidative stress damage and apoptosis in nerve cells to mitigate SCI by enhancing STUB1-induced HMGB1 ubiquitination degradation.
Insights
O-GlcNAc modification of FOXD3 protein exacerbates spinal cord injury (SCI) by increasing oxidative stress and apoptosis. Inhibiting this modification mitigates SCI by promoting HMGB1 degradation.
Area of Science:
- Neuroscience
- Molecular Biology
- Biochemistry
Background:
- Spinal cord injury (SCI) leads to severe sensory and motor dysfunction.
- The complex pathogenesis of SCI, particularly the roles of O-GlcNAc modification and FOXD3, is poorly understood.
- Understanding molecular mechanisms is crucial for developing effective SCI treatments.
Purpose of the Study:
- To investigate the role of O-GlcNAc modification and FOXD3 in spinal cord injury.
- To elucidate the molecular pathway involving FOXD3, STUB1, and HMGB1 in SCI pathogenesis.
- To explore therapeutic potential of targeting O-GlcNAc modification in SCI.
Main Methods:
- Established cell and animal models of SCI using H2O2 stimulation and impact methods.
- Assessed pathological changes, neuronal loss, and motor function using HE staining, Nissl staining, and behavioral tests (BBB score, ladder/grid climbing).
- Evaluated cell viability, apoptosis, protein modification, gene expression, and protein interactions using CCK8, flow cytometry, TUNEL, Co-IP, dual luciferase reporter, and ChIP assays.
Main Results:
- O-GlcNAc modification levels were significantly elevated in SCI models.
- O-GlcNAc modification enhanced FOXD3 protein stability and expression, contributing to oxidative stress and apoptosis.
- FOXD3 inhibited STUB1 transcription, which in turn induced HMGB1 ubiquitination and degradation.
- Inhibition of O-GlcNAc modification or FOXD3 knockdown protected against SCI-induced damage.
Conclusions:
- O-GlcNAc modification of FOXD3 plays a critical role in exacerbating SCI by promoting oxidative stress and apoptosis.
- The FOXD3-STUB1-HMGB1 pathway is a key mediator of SCI pathogenesis.
- Targeting O-GlcNAc modification of FOXD3 offers a potential therapeutic strategy for mitigating SCI by enhancing HMGB1 degradation.
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