Electrospun Beaded Fibers Carries Controlled Release NGF In The Application Of Nerve Conduit

Porous nanostructured fibers prepared by electrostatic spinning have similar structure with extra cellular matrix(ECM).The nerve growth factor(NGF)in the beaded structure of fibers can promote the growth of nerve growth and provide nourishment,and together,provide a micro environment for nerve to regenerate.However,currently there are two main problems about nerve conduits.The first problem is the carrying biological macromolecules technology is not mature enough,so under the harsh conditions of fiber preparing(strong shear force or organic solvent),the biologically active molecules is easily inactivated.The second problem is the material hydrophilic,biocompatibility and mechanical properties of electrospinning preparation are not good enough.The purpose of this thesis is to prepare the nerve conduits with good biocompatibility and mechanical properties,and achieve the long-term sustained release of bioactive factorsIn this study,we firstly using aqueous phase –aqueous phase Freezing-induced phase separationto prepare NGF-Dextran particles and then wrapped them to PLLA beaded fibers.Then explored the best preparing conditions for PLLA beaded fibers and PVA fibers.The best preparing conditions for PLLA are 9.5% concentration,10 k V voltage,1m L/h flow rate,15 cm receiving distance.The best preparing conditions for PVA are 12.5% concentration,15 k V voltage,0.5 m L/h flow rate and 10 cm receiving distance.Following the PLLA-PVA composite fibers,PLLA beaded fibers and emulsion PLLA fibers are prepared.The morphology,mechanical properties and contact angle are characterized.The results showed that PVA had the best hydrophilic and mechanical properties.Then Elisa kit was used to detect the quantities of NGF release in vitro.The results indicated the presence of burst release of emulsion PLLA fibers and in the 13 th day,there was no NGF detected.The control release of PLLA beaded fibers and PLLA-PVA fibers are better.During the first 15 days,the amount of release is more than 200 pg.Later,the releasing amount declined in certain degree but still remained in the range of 50-200 pg.In 30 days,the cumulative releasing amount of PLLA-PVA reached to 8965±367 pg(n=3),PLLA double fibers is 8512±326 pg(n=3),emulsion PLLA fiber is4076±181 pg(n=3).The results indicated that both PLLA-PVA composite fiber and PLLA beaded fiber can protect the activity of NGF protein and ensure the continuous and controlled release.The cell proliferation was used to investigate the cell responsiveness of these three fibers.The results showed that the cell responsiveness of PLLA-PVA composite fibers and PLLA beaded fibers are similar,both are better than emulsion PLLA fibers.In the later stage,the cell responsiveness of PLLA-PVA fibers is slightly better than PLLA beaded fibers.Finally,three kinds of electrospun fibers were fabricated into nerve conduits,then transplanted to damage place of rat sciatic nerve.Autologous nerve graft was conducted for positive comparison.The effect of nerve regeneration was tested by muscle atrophy,nerve conduction velocity and myelin sheath regeneration.The results showed that the effect of PLLA-PVA composite fibers and PLLA beaded fibers are similar,slightly worse than the autologous nerve graft,better than emulsion PLLA fibers.These results laid a good foundation for further studies of long-term sustained nerve conduits.