Preparation Of TNTs/PLGA/PEI Sustained-release Insulin Implant And Its Promotion Of Implant Osseointegration

Objective:The Fourth National Oral Epidemiological Survey shows that nearly 67% of people aged 55～64 in our country suffer from dentition defects of varying degrees,and 82% of patients with dentition defects in the 65～74 age group.Implants have the characteristics of no damage to adjacent teeth and high chewing efficiency and have become the first choice for repairing dentition defects.The osseointegration rate of patients with osteoporosis is reduced,which affects the early stability of the implant and osteoporosis has become a relative contraindication for implants.In recent years,the effect of insulin on bone regeneration has attracted attention,but the in vivo activity time of insulin is only 5-10 minutes.Studies have found that insulin-loaded polylactic-co-glycolic-acid(PLGA)microspheres can effectively control the release of insulin,but the drug loading is small;titanium dioxide nanotubes(TNTs)have strong loading and adsorption capacity,but the simple construction of TNTs drug-loaded coating will cause early burst release of the drug;polyethyleneimine(PEI)as an excitation layer can enhance the effect of titanium implant osseointegration.The purpose of this study was to prepare insulin-loaded TNTs/PLGA/PEI surface structures to slowly release insulin,and to investigate the effect of this surface structure on implant osseointegration under osteoporotic conditions.Methods:The PLGA nanoparticles were prepared by ultrasonic re-emulsification and then incubated with PEI solution to adsorb PEI on the surface of the PLGA nanoparticles.TNTs structures were prepared on pure titanium-based surfaces,and PLGA/PEI insulin-loaded nanoparticle complexes were loaded into the TNTs structures by vacuum perfusion method to monitor insulin release.This study was divided into four groups:smooth surface titanium sheet group,TNTs group,non-insulin-loaded TNTs/PLGA/PEI group,and insulin-loaded TNTs/PLGA/PEI group.For the effect on cell proliferation,real-time quantitative polymerase chain reaction(Real-time q PCR)was used to detect the effect of insulin-loaded TNTs/PLGA/PEI surface structure on osteogenic ability.A rat model of osteoporosis was constructed,implants were implanted,and the experimental rats were sacrificed 4 weeks later.Using methylene blue/acid fuchsin staining,the formation of new bone around implants in all groups was observed and expressed by the bone-to-implant contact ratio(BIC%).A portable digital torque tester was used to perform the torque test to detect the removal torque value(Removal Torque,RTQ).Results:Narrow size distribution,uniform insulin-loaded PLGA/PEI nanoparticles were successfully fabricated,with spherical and smooth surface morphology,which exhibited an initial burst release(about 40% within 24 hours),but the remaining insulin was released continuously for more than 15 days.At the same time,the PLGA/PEI insulin-loaded nanoparticle complexes were successfully loaded into the TNTs structure.The comparison with the cells cultured in the smooth titanium sheet group,TNTs group,and non-insulinloaded TNTs/PLGA/PEI group showed that the number of cells in the insulin-loaded TNTs/PLGA/PEI group increased significantly at 1,4,and 7 days.Real-time q PCR results showed that on the 3rd,7th,and 14 th days,the alkaline phosphatase(Alkaline Phosphatase,ALP),osteogenesis-related transcription factor antibody(Osterix,OSX),Runt-related transcription factor 2(Runt-related transcription factor 2,RUNX2)in the insulin-loaded TNTs/PLGA/PEI group concentration was significantly higher than those other three groups.In a rat model of osteoporosis,in histomorphometry analysis,the bone implants in the smooth-surfaced titanium sheet group,the TNTs group,non-insulin-loaded TNTs/PLGA/PEI group,and the insulin-loaded TNTs/PLGA/PEI group were 40.3±3.5%,45.3±3.5%,50.3±3.5% and 65.3±5.0%,respectively.The BIC% of the insulin-loaded TNTs/PLGA/PEI group was significantly higher than those other three groups.The mean RTQ values of the non-insulin-loaded TNTs/PLGA/PEI group and the insulin-loaded TNTs/PLGA/PEI group were significantly higher than those of other groups,which were12.3±3.1N·cm and 16.3±3.5N·cm,respectively.Conclusion:Insulin can be continuously released around the implants,promoting cell proliferation and osteogenic differentiation,by preparing insulin-loaded implants with TNTs/PLGA/PEI surface structure,and improving the osseointegration rate of implants under osteoporosis conditions.