Introduction/Objectives: Post-traumatic edema mediated by aquaporin-4 (AQP4) dysregulation is a critical determinant of secondary neuronal injury after spinal cord injury (SCI). This study aimed to investigate the therapeutic potential of miR-29a-3p in modulating AQP4 to alleviate edema and improve neurological recovery following SCI.
Methods: A mouse contusion SCI model was established (n=3/group, total n=30). Pathological validation was performed via H&E staining. AQP4 spatiotemporal expression was analyzed by immunofluorescence and Western blot (days 1, 7, 14 post-injury). Edema was assessed by spinal cord wet weight and in vivo MRI. In vitro, an IL-1α-induced astrocyte injury model was used. Bioinformatics screening and dual-luciferase assays identified miR-29a-3p as a direct AQP4 regulator. GFAP-Cre-driven Tg-29a transgenic mice (n=5/group, total n=60) enabled astrocyte-specific miR-29a-3p overexpression. Functional recovery was evaluated using the Basso Mouse Scale (BMS) and footprint analysis over 4 weeks.
Results: AQP4 expression was significantly upregulated (>6-fold vs. sham, p<0.01) and redistributed by day 7 post-SCI, correlating with increased tissue water content. In the injured spinal cord and in IL-1α-stimulated astrocytes, miR-29a-3p was downregulated while AQP4 was upregulated. miR-29a-3p directly targeted the 3’UTR of AQP4. Astrocyte-specific overexpression of miR-29a-3p in Tg-29a mice significantly suppressed AQP4 protein levels, reduced edema (MRI and tissue water content, p<0.01), attenuated glial scar formation (lowered GFAP and vimentin), and improved functional recovery (higher BMS scores, p<0.05) compared to wild-type SCI controls. Increased neuronal marker NF-200 expression was also observed.
Conclusions: AQP4 dysregulation is a pivotal mechanism in SCI-induced edema. miR-29a-3p serves as a key post-transcriptional regulator of AQP4. Astrocyte-targeted overexpression of miR-29a-3p effectively mitigates edema, modulates astrogliosis, and promotes functional recovery, highlighting its potential as a novel therapeutic strategy for SCI.
Ethical Statement: All animal procedures were approved by the Institutional Animal Care and Use Committee and conducted in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals.