Study On Topological Bionic Microenvironment And Heparin Loading Of PCL Scaffolds For Vascular Tissue Engineering

Large-diameter artificial blood vessels have been extensively used in clinical treatment,but small-diameter artificial blood vessels,such as coronary artery(diameter < 6mm),are prone to cause thrombosis and secondary obstruction after transplantation.Therefore,small-diameter tissue engineered artificial blood vessels with good service performance are urgently needed.Tissue engineered vascular scaffold is a necessary condition for vascular regeneration,and its quality determines the performance of blood vessels.To construct tissue-engineered blood vessels with good biocompatibility and mechanical properties,it is necessary to investigate how vascular scaffolds can provide bionic microenvironment and necessary mechanical properties for tissue-engineered blood vessels.Based on the study of extracellular matrix(ECM)-mimicking bionic microenvirenment created by the surface topography of nanofiber of vascular tissue engineering scaffold,the relationship between the bionic microenvirenment and the topology and mechanical properties of the scaffold has been developed.The single and synergistic effect of the physical clue and chemical factor provided by surface topography of the scaffold and the chemical clue provide by heparin loading on cell behaviors has been explored,and the effective approach for further improving the properties and functionality of the scaffold has been investigated.The study provides theoretical and expterimental basis for developing small-diamter vascular grafts with good performance.(1)Cell behaviors influenced by the morphology of the polycaprolactone fibersThe adhesion,migration,and proliferation of cells on tissue-engineered scaffolds are mechanical responses to the surrounding microenvironment.The research in Chapter 2 focuses on the effect of fiber morphology on endothelial cell behavior.Electrospun fibrous membranes with different fiber diameters and arrangements were prepared by adjusting the concentration of spinning solution and the receiving device.Human umbilical vein endothelial cells(HUVECs)were cultured on fiber membrane to investigate the effects of fiber diameter and orientation on cell adhesion,cell proliferation and cell morphology.The results suggested that due to the contact guidance,nano-scale fibers are more conducive to cell proliferation than micro-fibers,and aligned fibers can induce oriented cell alignment.Cell alignment on micron-aligned fibers is more significant,probably because the size of cell surface proteins is closer to that of microfibers,which is easier to be sensed by cells.(2)Cell behaviors influenced by the surface topographical structure of PCL electrospun fibersThe smooth surface of traditional electrospun fiber scaffolds which is researched in Chapter 2 is different from the topological structure of collagen fibers in extracellular matrix(ECM).To creat a more simulated biomimetic ECM microenvironment,in Chapter 3,the electrospun fibers were modified with shish-kebab structure by self-induced crystallization technology.The shish-kebab on the fiber surface can well simulate the periodic topographical structure of collagen fiber surface in extracellular matrix(ECM).The effect of crystal size on HUVECs adhesion and proliferation were studied by adjusting the shish-kebab size,surface roughness and hydrophilicity of fiber scaffold.The results showed that the specific surface area of material and surface roughness increased with the increase of kebab size,which provided more adhesion sites for cells and promoted cell adhesion.Moreover,increased hydrophilicity promoted cell proliferation.(3)Mechanical properties and anticoagulation of PCL tissue-engineered scaffoldsVascular scaffolds should have not only good mechanical properties to comply with blood flow and support blood vessel regeneration,but also good biocompatibility and blood compatibility.When vascular scaffolds are implanted,platelets will adhere to the surface of scaffolds.With the deposition of platelets,the inner wall of scaffolds will be narrowed or even cause thrombosis,which will eventually lead to the failure of replacement of tissue-engineered blood vessels.Therefore,the anticoagulant function of fiber scaffolds is particularly important.In Chapter 4,hydrophilic heparin was evenly dispersed in PCL solution.Due to the addition of heparin,the tensile properties,compliance and bursting strength of PCL/heparin hybrid fiber scaffolds meet the requirements of natural blood vessels.The loaded heparin in PCL/heparin blended fiber had better sustained release effect,which was beneficial to long-term anticoagulant activity of scaffolds.PCL/heparin blended fiber scaffolds can promote cell proliferation through enhanced biocompatibility.(4)Coupling effect of surface topography of fiber scaffolds with anticoagulant loadingBased on the results of the first three chapters,in Chapter 5,the shish-kebab structure and heparin were simultaneously loaded onto PCL fiber by means of blending and surface coating.The changes of scaffold properties such as morphology and chemical composition were studied by adjusting the loading mode of heparin.The coupling effects of heparin loading and topographical structure on HUVECs behavior,anticoagulant activity,inflammatory response,and vascular remodeling in vivo were further studied by cell culture and in vivo animal experiments.The results showed that the synergistic effect of heparin loading,and topographical structure could promote the proliferation of HUVECs and reduce platelet adhesion.Animal study results showed that the blended heparin group with shish-kebab structure could effectively reduce the inflammatory response,and intimal hyperplasia,which ultimately promote the reconstruction of vascular tissue.