Heparin crosslinked chitosan microspheres for the delivery of neural stem cells and growth factors for central nervous system repair

An effective paradigm for transplanting large numbers of neural stem cells after central nervous system (CNS) injury has yet to be established. Biomaterial scaffolds have shown promise in cell transplantation and in regenerative medicine, but improved scaffolds are needed. In this study we designed and optimized multifunctional and biocompatible chitosan-based films and microspheres for the delivery of neural stem cells and growth factors for CNS injuries. The chitosan microspheres were fabricated by coaxial airflow techniques, with the sphere size controlled by varying the syringe needle gauge and the airflow rate. When applying a coaxial airflow at 30 standard cubic feet per hour, ∼300 μm diameter spheres were reproducibly generated that were physically stable yet susceptible to enzymatic degradation. Heparin was covalently crosslinked to the chitosan scaffolds using genipin, which bound fibroblast growth factor-2 (FGF-2) with high affinity while retaining its biological activity. At 1 μg ml−1 approximately 80% of the FGF-2 bound to the scaffold. A neural stem cell line, GFP + RG3.6 derived from embryonic rat cortex, was used to evaluate cytocompatibility, attachment and survival on the crosslinked chitosan-heparin complex surfaces. The MTT assay and microscopic analysis revealed that the scaffold containing tethered FGF-2 was superior in sustaining survival and growth of neural stem cells compared to standard culture conditions. Altogether, our results demonstrate that this multifunctional scaffold possesses good cytocompatibility and can be used as a growth factor delivery vehicle while supporting neural stem cell attachment and survival.