Extracellular Vesicle‐Conjugated Functional Matrix Hydrogels Prevent Senescence by Exosomal miR‐3594‐5p‐Targeted HIPK2/p53 Pathway for Disc Regeneration

Abstract Nucleus pulposus stem cells (NPSCs) senescence plays a critical role in the progression of intervertebral disc degeneration (IDD). Stem cell‐derived extracellular vesicles (EV) alleviate cellular senescence. Whereas, the underlying mechanism remains unclear. Low stability largely limited the administration of EV in vivo. RGD, an arginine‐glycine‐aspartic acid tripeptide, strongly binds integrins expressed on the EV membranes, allowing RGD to anchor EV and prolong their bioavailability. An RGD‐complexed nucleus pulposus matrix hydrogel (RGD‐DNP) is developed to enhance the therapeutic effects of small EV (sEV). RGD‐DNP prolonged sEV retention in vitro and ex vivo. sEV‐RGD‐DNP promoted NPSCs migration, decreased the number of SA‐β‐gal‐positive cells, alleviated cell cycle arrest, and reduced p16, p21, and p53 activation. Small RNA‐seq showed that miR‐3594‐5p is enriched in sEV, and targets the homeodomain‐interacting protein kinase 2 (HIPK2)/p53 pathway. The HIPK2 knockdown rescues the impaired therapeutic effects of sEV with downregulated miR‐3594‐5p. RGD‐DNP conjugate with lower amounts of sEV achieved similar disc regeneration with free sEV of higher concentrations in DNP. In conclusion, sEV‐RGD‐DNP increases sEV bioavailability and relieves NPSCs senescence by targeting the HIPK2/p53 pathway, thereby alleviating IDD. This work achieves better regenerative effects with fewer sEV and consolidates the theoretical basis for sEV application for IDD treatment.