Three-dimensional Electrospun Polycaprolactone Composite Nanofibers And Their Cytobiological Properties

The scaffold material is the central link of tissue engineering technology,because it not only plays a role of supporting structure,but also directly regulates the behaviors of cells and provides templates to construct new tissues.Nanofiber materials prepared by electrospinning,which are widely used as biomedicine materials,including drug carriers,wound dressings and tissue engineering scaffolds for possessing the advantages of large specific surface area,high porosity and diverse functional design,simulates the structure of natural extracellular matrix.While,the traditional electrospinning technology can only produce two-dimensional structure so that it can not meet the practical needs of regeneration of human complex tissue and organ.In addition,the synthetic polymers that are suitable for electrospinning have limited application in special tissue engineering(hard tissues,and muscular tissues which are responsive to electrical stimulation),because of the poor mechanical properties and the lack of surface functional groups.Therefore,how to construct nanofiber scaffolds with functionality(performed mechanical property,conductivity,bioactivity and three-dimensional structure)is the main problem that our project aims at.In this study,graphene oxide(GO)was hybridized with polycaprolactone(PCL)to prepare PCL/GO composite nanofibers with reinforced properties via electrospinning.Compared with the pure PCL scaffold,the overall morphologies of the composite nanofiber scaffolds were more uniform and the surfaces of the composite scaffolds were smoother,while the diameters of the single fibers were smaller and the surfaces were slightly rough with grooves and protuberances.The surface chemistry and thermal properties of the composite nanofibers were improved by blending GO,which was not only embedded in the nanofibers,but also exposed to the fiber surfaces.When the GO content was lower,the elastic modulus and tensile strength of the composite nanofibers were enhanced,finally we obtained composite nanofiber scaffolds with improved mechanical properties by controlling the composite content of GO.The results of cytobiology revealed that GO improved cell adhesion and spreading on the composite nanofiber scaffolds,as a result the mouse marrow mesenchymal stem cells(mMSCs)and low-differentiated rat pheochromocytoma(PC12-L)cells exhibited typical fibroblast-and neuron-like morphologies respectively with mature and plump appearance and obvious pseudopods.The appropriate blending of GO into nanofibers not only was conducive to cell proliferation,but also facilitated the differentiation of mMSCs and PC12-L cells into osteo-and neuro-like cells respectively.The directing role of PCL/GO composite nanofiber scaffolds on cell behaviors may be attributed to the nanotopography(surface roughness),surface oxygen-containing functional groups and the conductivity of the scaffold materials.We also fabricated three-dimensional(3D)nanofiber macrostructure with nanotexture by designing the special collector,which was created by embedding stainless steel needles in a hemispherical polytetrafluoroethylene(PTFE)dish.This 3D nanofiber macrostructure reappeared the structure of the collector at macro level,showing a cap-like morphology and the thickness increased with the increase of the electrospinning time,even reaching a few centimeters.The 3D macrostructure simultaneously possessed the dense fibrous region and the loose fibrous region,among which the former can provide mechanical strength and the latter can promote cell infiltration.The 3D nanofiber macrostructures were integrated with 60% acetone/alcohol,which was used as the binder,and the integrated structure was ellipse-like,which was similar to some tissues/organs in macro morphology,such as lung,liver,kidney and heart.In addition,we blended the conductive polymer,polyaniline(PANi),into the PCL nanofiber to prepare 3D PCL/PANi composite nanofiber macrostructures.It was found that the addition of PANi increased the conductivity of electrospinning solution,decreased the average diameter of the fibers,and improved the surface chemistry of the 3D composite macrostructure.The results of cytobiology demonstrated that the appropriate addition of PANi into the 3D composite macrostructure facilitated cell adhesion,spreading,proliferation,myogenic differentiation and myotube maturation.Mouse myoblasts(C2C12 cells)can not only grew and differentiated on the surfaces of the 3D composite nanofiber macrostructure,but also penetrated into the interior,which revealed that on one hand the 3D PCL/PANi composite nanofiber scaffolds provided templates for the growth and filtration of the myoblast,on the other hand the incorporation of PANi facilitated myoblast differentiation and myotube formation.We obtained 3D nanofiber macrostructures with special shapes by combining the origami with electrospinning.Furthermore,we hybridized the bioactive nano-hydroxyapatite(nHA)into PCL nanofiber to prepare PCL/n HA composite with enhanced mechanical and biological properties.Taking the structure design into consideration,we constructed 3D PCL/nHA composite nanofiber scaffolds with special structures via origami,and cultured human fetal osteoblasts(hFOBs)on the 3D scaffold to construct 3D composite nanofiber scaffold/cells complex in vitro,which was expected to provide a new strategy for the construction of bone tissue engineering scaffold.In summary,this thesis aimed at the functionalization and 3D construction of the electrospinning nanofiber scaffold,and obtained composite nanofiber scaffolds with functionality(performed mechanical property,conductivity,bioactivity and three-dimensional structure).Overall studies demonstrated that electrospun PCL-based composite nanofiber scaffolds with good mechanical applicability and biological compatibility was expected to become a new generation of tissue engineering scaffold materials for the repair and reconstruction of tissues/organs,including bone,nerve,blood vessel and heart.