| Objective:The goal of regenerative endodontics is to reinstate normal pulp function,including innate pulp immunity,pulp nutritional function,pulp sensibility and pulp repair function.In the unique microenvironment of the dental pulp,the triad of tissue engineering would require stem cells,biomaterials and scaffoolds under the premise of controlling infection.In this experiment,BMSCs were selected as stem cells,collagen was used as a scaffold,iRoot BP was used as a biomaterial,and implanted subcutaneously into the rats to observe the formation of blood vessels and nerve fibers in the pulp-like tissue.At the same time,observe the biocompatibility and interaction between iRoot BP and dental pulp-like tissue.In order to provide a certain experimental basis for clinical transformation research of dental pulp tissue engineering.Material and methods:(1)Cell culture,isolation and distribution in collagen scaffolds.The cells were harvested from the bone marrow of wistar rat tibia,cultured in vitro by adherent method,passaged,and stored frozen.Collagen scaffolds engrafted with normal rBMSC and cultured in a 37℃incubator under normal conditions for 12 hours.The distribution of cells in the collagen membrane was observed by scanning electron microscopy(SEM).(2)Preparation of rat incisor root fragments.The incisors of rats were extracted and pulp tissue was removed to prepare 3 mm root fragments.After disinfection,collagen scaffolds with rBMSCs were placed in the root canal cavity and cultured in vitro for 12 hours.Subcutaneous transplantation was appled for subsequent experiments.(3)Subcutaneous transplantation and observeation of the formation of blood vessels and nerve fibers in the pulp-like tissue.Collagen scaffolds combined BMSCs in rat incisor fragments were implanted subcutaneously into the rats,to observe the regeneration of the pulp-like tissue and detect the formation of Blood vessels and nerves.Only collagen scaffolds and/or root fragments served as Control group.(4)Observeation of the biocompatibility of iRoot BP and influence on pulp regeneration.iRoot BP was placed on one side of the root fragments and the thickness is 0.5mm to 1mm.Placement of collagen scaffold material in the pulp cavity after coagulation.Composite rBMSCs were cultured in vitro for 12 hours before subcutaneous transplantation.To observe the regeneration of the pulp-like tissue and the biocompatibility of iRoot BP.Results:(1)The BMSCs from wistar rats were isolated and cultured by whole bone marrow adherence method,and the cells showed a uniform long fusiform spiral growth.Collagen scaffolds engrafted with normal rBMSC were also observed by scanning electron microscopy(SEM),After culture of the rBMSC on the scaffold for about 12 hours,abundant rBMSC attached and spread out onto the surface of the scaffold,as observed under SEM.(2)Dental pulp-like tissues were observed in the root fragments in experimental group,and H&E staining showed odontoblasts and massive angiogenesis in the new tissue.Immunohistochemistry showed positive expression of nerve fiber markers PGP9.5.There were litte dental pulp-like tissue in the control group and the blank group.(3)iRoot BP had good biocompatibility with the new dental pulp-like tissues.No inflammation was observed around iRoot BP.The revascularization around iRoot BP was significantly higher than the other areas of the root fragments.Conclusions:(1)BMSCs have a wide range of sources,high isolation rate,strong proliferative capacity and good combination with collagen scaffold,which can be used as the preferred seed cells for pulp tissue regeneration.(2)The rBMSCs combined with scaffolds promote regeneration of dental pulp-like tissues.(3)The iRoot BP has good biocompatibility,induces angiogenesis,and can be used as an ideal material for pulp revascularization. |
PDF Full Text Request