Construction Of Functional Chitosan-Based Bioactive Microcarrier And Its Application In Tissue Repair

Tissue engineering microcarriers have been widely studied in the fields of nerve,bone,cartilage and other tissue repair fields because of their large surface area,flexible and controllable size,adaptability to most shape defects,injection to avoid large area injury and so on.However,it has some problems,such as poor mechanical properties,small size and easy loss,low biological activity,single function,lack of environmental response and so on.On the basis of the previous research of our group,using the conductivity and bioadhesion of polydopamine,we constructed conductive microcarriers,magnetic microcarriers and bioactive microcarriers respectively,and carried out a series of experiments and systematic characterization in vitro.the physical and chemical properties of three kinds of functional microcarriers were regulated,and the response mechanism and their effects on stem cell behavior were studied.The application prospects of the above microcarriers in nerve repair,cartilage repair and bone repair were evaluated,which provided a basis for follow-up clinical trials.Porous chitosan(CS)microcarriers were prepared by emulsion polymerization and low temperature phase separation.By adjusting the conditions of dopamine self-polymerization,three-dimensional conductive layer structure was constructed by in-situ polymerization on CS microcarriers,and conductive CS/PDA microcarriers were prepared.The pore size,porosity,conductivity and biocompatibility of CS/PDA microcarriers were characterized,focusing on the effect of electrical stimulation on SCs cells.The results show that CS/PDA microcarriers have good pore size,porosity and biocompatibility.With the increase of water content,the electrical conductivity of CS/PDA microcarrier increases accordingly,and when saturated with water,the conductivity can reach 10-3 S/cm.When SCs was cultured on conductive CS/PDA microcarrier,it was found that cells could adhere and proliferate inside the microcarrier,and electrical stimulation promoted the faster proliferation of SCs cells.Based on the study of conductive CS/PDA microcarrier,CS/PDA microcarrier loaded with mesenchymal stem cells(rADSCs)(CS/PDA/rADSCs)was used to repair rat sciatic nerve.PCL catheter was used as the substrate,CS/PDA/rADSCs was injected,and routine feeding was carried out after suture.The movement recovery of rats was characterized by gross observation and gait analysis.The degree of sciatic nerve regeneration was evaluated by immunohistochemical staining and gastrocnemius histological staining.The experimental results show that CS/PDA/rADSC can change the microenvironment of nerve conduit,contribute to the growth and migration of Schwann cells,bridge the injured gap,establish signal transduction,and promote sciatic nerve regeneration,so it has a great application prospect in the field of nerve tissue engineering.The core-shell structure PDA@MS was synthesized by in-situ polymerization of dopamine and magnetic Fe3O4(MS)nanoparticles,and then the magnetic CS/PDA@MS microcarrier was successfully prepared by combining the hydroxy group of PDA with chitosan molecules.The magnetic CS/PDA@MS microcarrier has paramagnetism and can respond when an external magnet is added.The pH of the degradation solution of magnetic CS/PDA@MS microcarrier is alkaline,which is helpful to maintain the stability of tissue microenvironment at the defect site.Further increase the magnetic CS/PDA@MS Microcarrier loaded BMSCs were used for cartilage defects in the knee joint of rats,and cylindrical magnets were implanted in the posterior muscles of the knee joint.The results showed that the implanted CS/PDA@MS+BMSCs+Magnet has a more significant repair effect.Octadecalcium phosphate(OCP)was loaded into CS microcarrier to prepare bioactive CS/PDA/OCP microcarrier.CS/PDA/OCP microcarriers have interconnected void structure,high porosity and good biocompatibility.The addition of OCP improved the mechanical properties and protein adsorption capacity of the microcarrier.The mineralization experiment of simulated body fluid in vitro shows that CS/PDA/OCP microcarrier has high mineralization activity,and the pH of mineralization solution is alkaline,which is beneficial to neutralize the acidic environment of tissue defect and alleviate inflammatory reaction.Osteogenic differentiation in vitro showed that OCP microcarrier could promote the expression of alkaline phosphatase and calcium nodules,induce osteogenic differentiation of BMSCs,and have the potential to be used in the field of bone tissue engineering.