Research On The Construction Of 3D Printed New Drug Delivery System

Spinal cord injury?SCI?has a high rate of incidence and a low cure rate,and the functional distinction of motor sensory neurons in the gray matter and white matter area of the spinal cord is very obvious.Traditional medical treatment and surgical treatment cannot fundamentally solve the problem,and it is difficult to locate and restore the function.The use of tissue engineering to transplant cells and nutrient scaffold materials can provide exogenous cells to help restore the physiological function of the injury,and can also promote the repair of endogenous neurons,which is conducive to tissue reconstruction.Although there are many reports of scaffold transplantation for the treatment of spinal cord injury,traditional scaffold transplantation is also limited to the filling of defects in the spinal cord injury.Based on this problem,due to 3D printing has the characteristics of precise customization,this study intends to use 3D printing to build a new slow release drug delivery system:nanoparticle composite scaffolds,prepare new printable hydrogels and drug loaded nanoparticles,and carry out cells to study the differentiation of neurons in vivo and in vitro,so as to achieve the treatment and repair of spinal cord injury.Chapter one:ReviewsThis chapter reviews three aspects of spinal cord injury treatment,tissue engineering scaffold materials and 3d printing construction of sustained-release scaffolds.The treatment of spinal cord injury includes drug treatment and surgical treatment,among which drug treatment is mainly anti-inflammatory and provides nutritional factors.Tissue engineering scaffold materials,including natural sources and synthetic materials,are introduced.Finally,the research on 3D printing construction of slow-release scaffold is reviewed.Chapter two:Preparation and characterization of PLGA-loaded nanoparticlesIn this chapter,BDNF-loaded poly?lactic-glycolic acid?nanoparticles?BDNF@PLGA?were prepared by the double emulsion method.The detection methods of bovine serum albumin?BSA?and brain-derived neurotrophic factor?BDNF?were verified.The precision and recovery rate meet the methodological requirements.Using BSA as a model drug,orthogonal design was used to optimize the preparation process.The factor level was determined by single factor experiment,and the optimal process was determined by the encapsulation efficiency and drug-loading rate.The optimal conditions for preparation of BDNF-PLGA-NPs included a volume ratio of primary emulsion to external water phase of 1:15,the mass ratio of dosage to PLGA dosage of 6:100,a concentration of polyvinyl alcohol of 1%?w/v?and an ultrasonic time of 5 minutes.The BDNF nanoparticles obtained had a diameter of?215±0.28?nm,zata potential-23.31 mV and an EE of?78.75±2.61?%.The nanoparticles could release the drug in vitro for 14 days without burst effect,and the cumulative release rate was?84.65±3.35?%at equilibrium.Chapter three:Preparation and characterization of 3D printed nano-composite scaffoldsIn this chapter,3D printing was used to prepare calcium alginate-Matrigel composite scaffold loaded with BDNF nanoparticles.The printing material is composed of sodium alginate and Matrigel.Adjusting the proportion of mixed hydrogel has a great effect on the viscosity.The viscosity of 0.5%sodium alginate-30%Matrigel?SA-MA?mixed solution is suitable for printing.It is cross-linked with 3%CaCl₂ to form calcium alginate-Matrigel scaffold.The scaffold can meet the test of gel inversion experiment.BDNF@PLGA is added to SA-MA,and the BDNF@PLGA-SA-MA composition scaffold can be prepared by 3D printer dual-channel printing.Scanning electron microscopy showed that the scaffolds had abundant pore structure,nanoparticles were evenly distributed in the scaffolds,and the spherical shape remained intact.Compared with SA-MA scaffold?BDNF-SA-MA?loaded with free drugs,BDNF@PLGA-SA-MA composite scaffolds have stronger drug release ability.The results of Alamar Blue staining showed that the nanocomposite scaffold could promote cell proliferation.The results of Calcein-AM/PI staining showed that the nanocomposite scaffolds had high biocompatibility.The cell survival rate was?88.04±3.10?%after three days of culture.Chapter four:The 3D printed nano-composite scaffold induced differentiation of ectodermal mesenchymal stem cells into neurons in vitroIn this chapter,the application of 3D printed BDNF@PLGA-SA-MA composition scaffolds in vitro was investigated.The ability to promote the differentiation of EMSCs into neurons was investigated,and a BDNF-SA-MA scaffold with two-dimensional culture and free drug loading was used as a control group.The results of immunofluorescence showed that EMSCs could express specific neuronal markers in above culture environments.After induction,the neurons on BDNF@PLGA-SA-MA composite scaffold were more mature and could form neural network.qRT-PCR was used to detect the relative mRNA expression of the target gene,and the mRNA expression of EMSCs in the nanoparticle composite scaffold was higher.Flow cytometry was used to detect the expression of TUJ1,and the positive rate of EMSCs in the nanoparticle composite scaffold was higher.The three-dimensional environment can simulate the role of extracellular matrix,and the inclusion of nanoparticles can reduce the loss of BDNF,slowly release drugs to induce cell differentiation,and EMSCs differentiate into neurons more efficiently.Chapter five:Research on the repair of spinal cord injury with 3D printed nano-composite scaffoldStudy on BDNF@PLGA-SA-MA composite scaffold with EMSCs to repair spinal cord injury in rats in vivo were carried out.Spinal cord injury modeling by truncating T9-T10 spinal cord,transplantation of carrier cell composite scaffolds as a treatment strategy.The recovery of motor function was evaluated by BBB score and open field test.BDNF-SA-MA scaffold with free drug in the first four weeks after injury had better recovery ability,As the scaffold degrades BDNF@PLGA-SA-MA,it can slowly release the drug encapsulated by the nanoparticles,so it shows a higher effect on the recovery of motor function.The pathological section results showed that after BDNF@PLGA-SA-MA scaffold grafting,the damage cavity was reduced,the intercellular space was significantly reduced,the arrangement was orderly,and the tissues were favorably reconstructed.