| Cardiac patches are biomaterials that can be used for transplantation and repair of damaged myocardium by combining seed cells with the ability to form cardiomyocytes and suitable scaffold materials.Cardiac patches provide temporary support to the infarcted area on the one hand,and repair the damaged myocardium by delivering cells or bioactive factors to integrate with the host on the other hand,and their preparation has become a hot research topic in recent years.The polymer melt electrowriting technology can orderly control the location of fiber deposition and has excellent mechanical properties.The melt differential electrospinning technology has the characteristics of small diameter of prepared fibers and high porosity of fiber membrane.Both techniques have great potential for tissue engineering applications.Based on the review of cardiac patch preparation technology at home and abroad,combined with the advantages of melt electrowriting and melt differential electrospinning technology,a new cardiac patch preparation technology which mimics the anisotropic mechanical/electrical properties of natural myocardium was proposed and studied in this paper.The research is mainly conducted from the following aspects:（1）Rhomboid polycaprolactone（PCL）scaffold with anisotropic regulation was prepared by melt electrospinning direct writing technique.The effects of structural parameters such as the number of layers,fiber spacing and mesh Angle on the mechanical properties of the scaffolds were studied experimentally.Post-treatment combined with external conductive material carbon nanotubes（CNTs）gave the scaffolds conductive properties.The results show that the change of the number of layers has little effect on the mechanical properties of the scaffold,and the increase of the fiber spacing decreases the overall mechanical properties of the scaffold.The tensile strength and Young’s modulus in the longitudinal direction increased,the tensile strength and Young’s modulus in the transverse direction decreased,and the anisotropic stiffness ratio increased with the decrease of the mesh angle.The longitudinal tensile strength and Young’s modulus of the 15-layer,300μm,70°scaffold were 0.45 MPa and 1.67MPa,respectively,and the transverse tensile strength and Young’s modulus were 0.3 MPa and 0.47 MPa,respectively,with an anisotropic stiffness ratio of 3.55.The longitudinal conductivity of the scaffolds after treatment with ultrasonically dispersed CNTs solution was 1.15×10-3 S/cm and the lateral conductivity was 8.67×10-4 S/cm.The scaffolds showed no significant changes in conductivity during the regular cell culture cycle（3-30 d）and could be used stably.（2）To prevent problems such as cell leakage due to large pore size of scaffolds during in vitro cell culture,PCL fiber membranes were prepared by melt differential electrospinning technology to support and seal the bottom of scaffolds.The spinnability was first investigated experimentally,and then the effects of process parameters such as spinning temperature and spinning voltage on the diameter of PCL fibers and the porosity,pore size distribution and mechanical properties of the fiber membrane were studied.The study showed that with the increase of spinning temperature,the average diameter of PCL fibers first became finer and then coarser.The higher the spinning voltage,the smaller the average diameter of fibers,and the smallest fiber diameter is 1.65μm at 200°C and 35 k V.The porosity of fiber membrane increases with the increase of spinning temperature,and the highest porosity of fiber membrane is 72.1%when the spinning temperature reaches 220°C.The higher the spinning voltage,the greater the degree of inter-fiber entanglement,the smaller the pore size distribution and the higher the mechanical properties of the fiber membrane.The average pore size at 35 k V was 9.3581μm,which was lower than the average diameter of rat H9c2 cardiomyocytes,and the tensile strength was up to 8.5 MPa and the elongation at break was up to 57%.（3）The cytotoxicity of the scaffolds was assessed by CCK-8 assay.The cell survival rates of all groups of scaffolds on day 1 and day 3 of co-culture were above 80%with no significant differences,and the scaffolds were biocompatible.PCL/CNTs diamond scaffolds and PCL/CNTs square scaffolds were designed as experimental groups,and pure PCL scaffolds were designed as control groups,and the scaffolds were grown in vitro with rat H9c2 cardiomyocytes.The cytoskeletal F-actin was fluorescently stained after 1 and 3 days of culture.The results showed that the cells in the control group were mostly round after 1 day of culture,and the cells in the experimental group showed a natural cardiac muscle-like long shuttle shape.The adhesion and spreading of the cells on the scaffold surface in the experimental group were better than those in the control group after 3days of culture,and the scaffold in the PCL/CNTs diamond-shaped group showed a certain degree of orientation.In summary,in this paper,precisely tunable conductive anisotropic PCL/CNTs scaffolds were prepared by melt electrowriting,and the mechanical/conductive properties of the scaffolds were close to those of natural myocardium.Secondly,PCL fiber membranes were prepared by melt differential electrospinning to support and seal the bottom of the scaffolds.Finally,in vitro cell culture showed that the scaffold promoted the growth and adhesion of rat H9c2 cardiomyocytes,providing an important basis for promoting further applications of cardiac patches. |
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