| Objectives: The purpose of this study was:(1)to evaluate the drug loading capability of Poly(D,L-lactic-co-glycolide)(PLGA)polymers to chlorhexidine,calcium,phosphorus elements as well as the physico-chemical properties,antibacterial ability,biocompatibility,effects on dentin microhardness of the product;(2)to evaluate the drug loading capability of PLGA polymers to a new quaternary ammonium compound K21,calcium,phosphorus elements as well as the physico-chemical properties,antibacterial ability,biocompatibility,in vitro mineralization ability of the product;(3)to use PLGA submicron particles containing K21,calcium,phosphorus to prevent Enterococcus faecalis(E.faecalis)infection in beagle dogs’ root canals.Methods: Experiment 1: PLGA submicron particles carrying chlorhexidine,calcium and phosphorus were prepared according to a modified emulsion-solvent evaporation method.FE-SEM was used to observe the morphology of submicron particles.EDS was conducted to analyse element composition.The diameter of particles and zeta potential were measured by dynamic light scattering.FTIR was used to comfirm the incorporation of chlorhexidine.The release of calcium and phosphorus was measured in Tris-HCl while the release of chlorhexidine and p H values were measured in doubledistilled water.A CFUs count method was applied for the assessment of antibacterial ability against planktonic E.faecalis.To evaluate the biocompatibility of the submicron particles,a CCK-8 kit was used.Dentin slices were medicated by ultrasonic vibration in submicron particles suspensions to assess the infiltration ability into dentinal tubules.Medicated dentin slices were incubated with E.faecalis suspensions for 7 days.Then dentin slices were transferred to fresh culture medium and the optical density values of the medium were measured at time intervals to investigate the inhibition of E.faecalis on dentin.Besides,dentin slices were cultured with submicron particles paste to evaluate the effects on dentin microhardness.Experiment 2: PLGA submicron particles containing K21,calcium and phosphorus elements were prepared according to the modified emulsion-solvent evaporation method in experiment 1.FE-SEM,EDS,dynamic light scattering and FTIR was used to analyse the morphology and physicochemical properties of submicron particles.The p H value change and the release of K21 were measured in double-distilled water while the release of calcium and phosphorus was measured in Tris-HCl.A CFUs count method and a CCK-8 kit were applied respectively for the assessment of antibacterial ability against planktonic E.faecalis and cytotoxicity.The infiltration and inhibition of E.faecalis on dentin were evaluated by the method in experiment 1.To investigate the in vitro mineralization ability,submicron particles compacted into root canals were immersed in simulated body fluid and the formation of crystals on the surface was observed.Experiment 3:Five beagle dogs were used.Bilateral mandibular second and third premolars were opened and instrumented.Then suspensions of PLGA submicron particles containing K21,calcium and phosphorus,chlorhexidine solution,normal saline,calcium hydroxide paste were added into root canals randomly.Root canals were subjected to ultrasonic vibration and pulp chambers were sealed.One week later,temporary restorations were removed.E.faecalis suspensions were added into root canals and pulp chambers were sealed again.Radiographic images were taken every two weeks to observe periapical lesions.8 weeks after bacteria infection,dogs were sacrificed.Experimental teeth with surrounding alveolar bone were dissected and subjected to fixing,decalcifying,dehydration,embedding.Finally tissue sections were obtained.Hematoxylin-eosin staining was used to evaluate periapical lesions.Brown&Brenn staining was conducted to observe bacteria distribution.TRAP staining was performed to observe osteoclasts.Results: Experiment 1: PLGA submicron particles carrying chlorhexidine,calcium and phosphorus were synthesized successfully.The submicron particles had a diameter of approximately 500 nm while the zeta potential was-7 m V,with an acidic p H value.Encapsulated chemicals could be released in a sustained manner from the particles.The particles also exhibited excellent biocompatibility on MC3T3-E1 cells and significant antimicrobial property against planktonic E.faecalis.On dentin slices,the particles could be driven into dentinal tubules by ultrasonic activiation and inhibit E.faecalis colonization.Besides,dentin slices medicated with the particles displayed an increase in microhardness.Experiment 2: PLGA submicron particles containing K21,calcium and phosphorus were synthesized successfully.Submicron particles had a diameter of about 250 nm while the zeta potential was 34 m V.Particles were acidic in double-distilled water.K21,calcium and phosphorus could all be released in a sustained manner from submicron particles.The particles were noncytotoxic,with strong antibacterial ability against E.faecalis.On dentin slices,the particles could be driven into dentinal tubules by ultrasonic vibration and lingered on dentin surface to inhibit E.faecalis colonization.Besides,after immersion in simulated body fluid,massive apatite crystals could be induced on the surface.Experiment 3: In beagle dogs’ root canals,compared with calcium hydroxide and normal saline,PLGA submicron particles containing K21,calcium and phosphorus could decrease inflammatory infiltrate and the number of osteoclasts while inhibited bacteria invasion into dentinal tubules.Cunclusions:(1)PLGA submicron particles carrying chlorhexidine,calcium and phosphorus exhibited favorable physico-chemical properties,biocompatibility,antibacterial ability,infiltration ability into dentinal tubules and dentin microhardness improvement capability.(2)PLGA submicron particles containing K21,calcium and phosphorus showed satisfactory physico-chemical properties with no cytotoxicity and excellent antibacterial ability.The particles also possessed infiltration ability on dentin slices and in vitro mineralization ability.(3)PLGA submicron particles containing K21,calcium and phosphorus demonstrated better prevention to E.faecalis infection in beagle dogs’ root canals compared with calcium hydroxide.The particles might be developed into a new intra-canal disinfection. |
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