A Study Of Electrospun PHBV Nanofibers For Bone Regeneration And Nerve Regeneration

Tissue engineering comprised three elements:cells,growth factors and scaffolds.The PHBV(poly(3-hydroxybutyrate-co-3-hydroxyvalerate))was a bio-absorbable polymeric material that was synthesized by microorganisms.The biomaterial was recognized for its biodegradability,biocompatibility and non-immunogenicity.To mimic the natural structure of extracellular matrices,electrospinning was applied to fabricate biomaterials into ultrafine fibers.Based on the electrospun PHBV nanofibers,four kinds of nanofibers were fabricated for bone regeneration including aligned PHBV nanofibers,aligned PHBV/HA nanofibers,random-oriented PHBV nanofibers and random-oriented PHBV/HA nanofibers,another four kinds of nanofibers were fabricated for nerve regeneration including aligned PHBV nanofibers,.aligned PHBV/Laminin nanofibers,random-oriented PHBV nanofibers and random-oriented PHBV/Laminin nanofibers.In the vitro study,four kinds of nanofibers were fabricated for bone regeneration were investigated.The BMSCs cultured on the flat films were tested by CCK-8 kits and osteogenic markers.Cell proliferation tested with CCK-8 assay indicated that the BMSCs attached and proliferated more favorably on random-oriented PHBV nanofibrous meshes without HA.After one,two and four weeks of cell seeding,osteogenic markers including alkaline phosphate(ALP),osteocalcin(OCN),and mineralized matrix deposits were detected,respectively.The results indicated that the introduction of HA could induce the BMSCs to differentiate into osteoblasts.As similar,the Schwann cells seeded on four kinds of nanofibers fabricated for nerve regeneration were investigated through CCK-8 kits test and specific markers test.The results showed that the attachment and proliferation of Schwann cells on the PHBV meshes was good.SEM showed the BMSCs and Schwann cells migrated and elongated along the same diection on the aligned meshes,BMSCs and Schwann cells migrated and elongated randomly on the random-oriented meshes.In vivo study,the PHBV nanofibers were fabricated into 3D scaffolds and then the scaffolds were implanted into the rabbit 1.5cm radius defect for bone regeneration.The outcome of repairing the defect were tested through gross observation,X-ray,histological examination,SEM,CT three dimensional reconstruction and biomechanical test.The results showed that the four kinds nanoibrous scaffolds could heal the bone defect 16 weeks after the operation,the random-oriented PHBV/HA nanofibrous scaffolds had the best osteogenic capacity among the four kinds nanoibrous scaffolds.The results in vivo and vitro study indicated that the HA loaded on the PHBV nanofibers could improve the bone regeneration capacity of the nanofibers.The random-oriented nanofibers may affect bone regeneration in a certain extent.The blood tests were applied in the rabbits before the operation and after the operation.The results showed that the PHBV nanofibers had good biocompatibility.The PHBV nanofibers were fabricated into nerve guide conduits and then the conduits were implanted into the SD rats sciactic nerve defect for repairing the 12mm nerve gap.The outcome of nerve regeneration were tested through gross observation,sciatic functional test,histological examination,and electrophysiological test.The results showed that the four kinds nanoibrous scaffolds could repair the sciatic nerve gap 8 weeks after the operation,the aligned PHBV/Laminin nanofibrous conduits and the aligned PHBV nanofibrous conduits could repair the nerve defect better than the random-oriented nanofibrous conduits,the Laminin did not affect the nerve regeneration capacity of the nanofibers both in vivo and vitro.The PHBV nanofibers could repair the nerve injury due to its capability of affecting endogenous neurotrophic factors secrection.