Study Of The Repairing Effects Of Newly Constructed Tissue-engineered Nerve Conduits On Spinal Cord Injury

BackgroundTissue-engineered scaffold material has been used for the repairment of spinal cord injury,but it has limitations, and the effects are not satisfied.ObjectivesTo newly construct tissue-engineered composite scaffolds of nerve conduits based on poly-lactic-co-glycolic acid(PLGA) and salidroside microsphere, and observe their repairing effects on spinal cord injury in rats so as to provide basic data for their application in the repairment of clinical spinal cord injury.MethodsEmulsification-solvent evaporation was used to prepare dexamethasone, salidroside, and mitomycin C microspheres. The influence of drug dosage, the amount of PLGA and the mass fraction of polyvinyl alcohol(PVA) on microsphere preparation process was evaluated by orthogonal test according to the total score of the entrapment efficiency, drug loading and yield.The characterization of microspheres was observed under scanning electron microscope.Electrospinning technique was used to prepare the shell layer of scaffold with collagen and polycaprolactone(PCL) as raw materials. Microspheres were mixed with collagen to prepare the core layer of scaffold with special mould. The composite scaffolds were divided into no-microsphere group, dexamethasone and salidroside microsphere group, and mitomycin C and salidroside microsphere group, and co-cultured with rat bone marrow mesenchymal stem cells,respectively. Wistar rats were used to establish the animal model of spinal cord injury, and then randomly divided into no-microsphere group, dexamethasone and salidroside microsphere group,and mitomycin and salidroside microsphere group with 20 rats in each group. The histological specimens were taken on weeks 4 and 16 after operation respectively, routinely fixed and stained with HE. Immunofluorescence staining was used to detect the expressions of S100, neurofilament,glial fibrillary acidic protein, and macrophage surface-specific antibody ED-1 to comprehensively evaluate the repairing effects of scaffolds on spinal cord injury.ResultsThe best preparation process for salidroside microsphere was 6 mg salidroside, 120 mgPLGA and 1% mass fraction of PVA, and the drug loading, encapsulation efficiency and yield of salidroside microsphere were(3.86 + 0.03)%,(83.62 + 0.31)% and(90.67 + 2.35)% respectively.The best preparation process for dexamethasone microsphere was 10 mg dexamethasone, 80 mg PLGA and 0.5% mass fraction of PVA, and the drug loading, encapsulation efficiency and yield of dexamethasone microsphere were(2.26 + 0.03)%,(83.62 + 0.21)% and(90.87 + 2.45)%respectively. The best preparation process for mitomycin C microsphere was 200 μg mitomycin C,60 mg PLGA, and 0.5% mass fraction of PVA, and drug loading, encapsulation efficiency and yield of mitomycin microsphere were(4.86 + 0.04)%,(76.62 + 0.52)% and(90.23 + 1.75)%respectively. The microspheres were round with smooth surface. The rat bone marrow mesenchymal stem cells grew well on the composite scaffolds with microspheres. The results of HE staining showed that on weeks 4 and 16 after operation the composite scaffolds were obviously or completely degraded and grown with a large number of cells in microsphere groups while there was a small number of cells grown in no-microsphere group. Immunofluorescence staining manifested that the expressions of S100 and NF were gradually increased while the expressions of glial fibrillary acidic protein and ED-1 were gradually decreased in no-microsphere group, dexamethasone and salidroside microsphere group, and mitomycin C and salidroside microsphere group. The repairing effects in microsphere groups were significantly better than in no-microsphere group.ConclusionsThe newly constructed tissue-engineered nerve conduits in this experiment can effectively promote the repairment of spinal cord injury, and has potential clinical application prospects.