Electrospun Membranes Loading MiRNA-126/miRNA-145 And Modification For Vascular Regeneration

Small-diameter(<6mm)vascular grafts are difficult to be widely used because of thrombosis,intimal hyperplasia during in vivo transplantation.It is currently improved by the modification of the surface morphology and hydrophilicity of the material,and the addition of growth factors in materials.However,due to the lack of design in the structure and function on vascular grafts,it cannot meet clinical requirements.Micro RNAs(mi RNAs)are important regulators of gene expression.In this thesis,the electrospun membranes with the target carriers of mi RNAs were prepared and delivered to vascular smooth muscle cells(SMCs)or vascular endothelial cells(ECs)to study the effects on vascular regeneration,inflammation and calcification.The main contents consists of four parts.Short peptide Val-Ala-Pro-Gly(VAPG)was linked to PEG-g-trimethylated chitosan polymer to prepare TMC-g-PEG-VAPG(TPV)with different molecular weights(5 k Da,20 k Da and 50 k Da)for specially targeted delivery of mi RNA-145 to SMCs.TMC-g-PEG-VAPG with relatively higher molecular weight of chitosan(50k Da)could significantly enhance cellular uptake in SMCs over that in ECs.Moreover,loading with TMC-g-PEG-VAPG/mi RNA-145 complexes,the poly(ethylene glycol)-b-poly(L-lactide-co-(?)-caprolactone)(PELCL)electrospun membranes were capable of modulating the contractile phenotype of SMCs in the prolonged duration.Arg-Glu-Asp-Val(REDV)-modified electrospun membranes was developed specifically consisting of the PELCL and REDV-linked PELCL with TMC-g-PEGREDV(TPR)/ mi RNA-126 to regulate ECs growth under shear stress.PELCL-REDV was located on the fiber surfaces,which could accelerate the release of mi RNA-126 and facilitate ECs adhesion and proliferation.And ECs showed elongated cell shape and parallel to the flow direction under shear stress(1 Pa).An efficient bioactive trilayered tissue-engineered vascular graft was prepared specifically consisting of PELCL/REDV-modified PELCL as inner layer loading with TPR/mi RNA-126,PELCL as middle layer loading with TPV/mi RNA-145,and PCL as the outer layer.In vitro cell culture results demonstrated that the accelerated release of mi RNA-126(71% in 56 days)had significant biological advantages in enhanced proliferation and intracellular nitric oxide production in ECs.Phenotypes and restraint of calcium deposition of SMCs were modulated by slow-releasing mi RNA-145(34%in 56 days).In vivo implantation in a rat abdominal aorta interposition model suggested that the local delivery of mi RNA-126 and mi RNA-145 exerted a positive effect on accelerating endothelialization(nearly 100% at 4 weeks),improving contractile SMCs regeneration,promoting normal extracellular matrix formation,regulating inflammation and depressing calcification possibly by facilitating transformation of macrophages into the anti-inflammatory M2 phenotype,which could improve the patency rate of small-diameter vascular grafts.REDV was reacted with polyurethane(PU)and blended with PCL to prepare electrospun membranes.The tensile properties and compliance suggested that the mechanical properties of PU-REDV/PCL electrospun membranes matched that of natural blood vessels.