| Pulp tissue can be affacted by deep caries and accidental trauma or restorative procedure,which influence the root development and apex closure and clinicians often preserve the part or whole life of the tooth pulp using vital pulp therapy.The formation of tertiary dentin contributes to successful cure of pulp injury,in which the newly formed dentine protects the pulp from external stimuli and microbiological challenges,leading to long-term survival of the dental pulp tissue.In this regard,the use of biomaterials as a pulp-capping material to promote the formation of the tertiary dentin and improve the success rate of vital pulp therapy has been widely studied and applied in clinical practice.Bioceramic materials such as MTA and Biodentine have significant advantages in properties such as sealing,adhesion,biocompatibility,promotion of biomineralization and induction of odontogenic differentiation,but they also have some shortages like long setting-time,insufficient ability to promote pulp-dentin regeneration.However,in recent years,drug-loaded microspheres loaded with osteogenic active bioactive substances for vital pulp therapy have been promising.Dexamethasone,a synthetic glucocorticoid,is used as an anti-inflammatory drug and considered as a long-acting steroid.It has been proved that DEX has osteoinductive effect on bone marrow stromal cells,dental pulp cells and dental follicle cells.However,large doses of DEX results in tissue necrosis because it could inhibit osteogenic activity and significantly enhance the formation of lipid.Compared with transforming growth factor beta 1(TGF-β1),bone morphogenetic protein 2(BMP-2),dexamethasone is a small molecular bioactive substance with longer half-life,lower cost.However,large doses of DEX results in tissue necrosis because it could inhibit osteogenic activity and significantly enhance the formation of lipid.The use of drug delivery systems can make up for this shortcoming.The purpose of the drug delivery system is to achieve a sustained therapeutic effect in a local microenvironment at a lower concentration through its slow release action.Hollow hydroxyapatite micropheres,with hollow core and porous shell structure and good biocompatibility,which can efficiently increase the drug loading rate of hydroxyapatite,are regarded as good carriers for drug molecules and other bioactive factors.HHAM are widely used for sustained release,which can deliver active substances in local and slow-release manner overtime.In the previous experiments of our group,dexamethasone was loaded into hollow hydroxyapatite microspheres to form a sustained release system.The system has an good drug loading rate and encapsulation efficiency and a sustained release effect in vitro.And the experimental method is simple,feasible,and repeatable.In this study,the effects of dexamethasone hollow hydroxyapatite microspheres on the proliferation and odontogenic differentiation of hDPCs were examined.Our study will provide an evidence for the clinical application prospect of vital pulp therapy.Methods:The dental pulp cells were extracted from the pulp tissue of healthy human permanent teeth.After subculture,the second to fourth generations were selected for subsequent experiments.The viability of hDPCs was quantitatively assessed in the presence of DHHAM using the method of Cell Counting Kit-8 assay At the same concentration of DHHAM,HHAM and DEX,the Quantitative Polymerase Chain Reaction,alkaline phosphatase staining and Alizarin Red S staining results tested the effect of DHHAM on biomineralization of hDPCs in the absence of osteogenic medium and observed the gene expression of alkaline phosphatase,runt-related transcription factor 2,osteocalcin,dentin sialophosphoprotein and dentin matrix protein 1 in this study.Results:In this experiment the drug loading efficiency of DHHAM is 16.7%.The drug-loading efficiency of the sustainereach d-release system is good,and the sustained-release time in vitro can 30 days,which can significantly prolong the action time of the drug,has excellent performance,and can exert a stable and local slow release effect.The effect of DHHAM on the proliferation of hDPCs was dose-dependent and inversely correlated.With the increase of DHHAM concentration,the proliferation of hDPCs was inhibited,and 1 μg/ml of DHHAM was the proper concentration for proliferation of hDPCs.At the same concentration of DHHAM,HHAM and DEX,the secretion of alkaline phosphatase increased gradually from day 7 to day 14,and the expression of alkaline phosphatase in DHHAM group was higher than that in other groups on day 7 or 14(P<0.05).The results of Alizarin Red S staining showed that DHHAM has a stronger ability to promote extracellular matrix mineralization of hDPCs.qPCR assay showed that DHHAM significantly promoted the expression of ALP,Runx2,OCN,DSPP and DMP-1 in human dental pulp cells during odontogenic differentiation than DEX and HHAM(P<0.05).Conclusion:Compared with dexamethasone or hollow hydroxyapatite alone,the sustained release system significantly promotes the mineralization of human dental pulp cells in vitro,and increases alkaline phosphatase,Runx2,dentin sialophosphoprotein,osteocalcin,and dental matrix protein-1.And it provides a basis for the pulp capping agent for animal experiments in pulp preservation. |
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