Injectable smart-blended hydrogel cross-linked with Vanillin to accelerate differentiation of intervertebral disc-derived stem cells (IVDSCs) for promoting degenerative nucleolus pulposus in a rat model.

Abstract

The nucleus pulposus (NP) degradation is a primary factor in intervertebral disk degeneration (IVD) and a major contributor to low back pain. Intervertebral disk-derived stem cell (IVDSC) therapy presents a promising solution, yet identifying suitable cell carriers for NP transplantation remains challenging. The present study investigates this issue by developing smart injectable hydrogels incorporating vanillin (V) and hyaluronic acid (HA) encapsulated with IVDSCs to facilitate IVD regeneration. The hydrogel was cross linked by carbodiimide-succinimide (EDC-NHS) method. Enhanced mechanical properties were achieved by integrating collagen and HA into the hydrogel. The rheological analysis revealed the pre-gel viscoelastic and shear-thinning characteristics. In vitro, cell viability was maintained up to 500µg/mL, with a high proliferation rate observed over 14days. The hydrogels supported multilineage differentiation, as confirmed by osteogenic and adipogenic induction. Anti-inflammatory effects were demonstrated by reduced cytokine release (TNF-α, IL-6, IL-1β) after 24h of treatment. Gene expression studies indicated elevated levels of chondrocyte markers (Acan, Sox9, Col2). In vivo, hydrogel injection into the NP was monitored via X-ray imaging, showing a significant increase in disk height index (DHI%) after 8weeks, alongside improved histologic scores. Biomechanical testing revealed that the hydrogel effectively mimicked NP properties, enhancing compressive stiffness and reducing neutral zone stiffness post-denucleation. The results suggest that the synthesized VCHA-NP hydrogel can be used as an alternative to NPs, offering a promising path for IVD regeneration.