Nanogels Of Carboxymethyl Chitosan And Lysozyme Encapsulated Amorphous Calcium Phosphate Occlude Dentinal Tubules

ObjectiveThis study aimed to develop of a rapid and effective method to occlude dentinal tubules.First,carboxymethyl chitosan and lysozyme（CMC/LYZ）are used to encapsulate amorphous calcium phosphate（ACP）to form carboxymethyl chitosan/lysozyme-amorphous calcium phosphate nanogels（CMC/LYZ-ACP）.The effective closure of the dentinal tubules based on the transformation of amorphous calcium phosphate into hydroxyapatite（HAP）.MethodsIn this work,CMC/LYZ was used to stabilize ACP and form CMC/LYZ-ACP nanogels.The morphology of the nanogels was examined by transmission electron microscopy（TEM）.Quantitative analysis of particle size was measured by dynamic light scattering.Distribution and quantity of elements in CMC/LYZ-ACP nanogels were determined by element mapping and energy dispersive X-Ray spectroscopy（EDX）.X ray diffraction（XRD）and Fourier Transform Infrared Spectroscopy Analyzer（FTIR）was used for the analysis of the phase and component of nanogels.At the same time,the CMC/LYZ-ACP nanogel was compared with the CMC-ACP nanoparticles which were studied by the previous research group.And then 32 fresh teeth in vitro were collected for periodontal disease,orthodontic meiosis or wisdom tooth extraction without caries,cracks,and obvious wear.Divided into 4 groups:normal dentin group,dentin tubule exposed group,CMC/LYZ-ACP nanogels treatment group,Green Or^TM positive control group,8 samples in each group.The nanogel-encapsulated ACP was applied to artificial dentin sensitive model,exposed dentinal tubule surfaces.After ultrasonic cleaning,scanning electron microscopy（SEM）images,XRD measurements and nanoindentation were applied to check the efficacy of tubular occlusion.And contrast with Green Or^TMM groups.ResultsTEM revealed that CMC/LYZ-ACP nanogels were spherical dense gel particles with size approximately 50-500nm.DLS analysis of particle diameter was about354.8 nanometers.Element mapping and EDX indicated that C,N,O,Ca,P,and S in the microspheres are thoroughly represented.The phase analysis of nano-gel by XRD indicates that there is no crystal phase in CMC/LYZ-ACP nanogels.FTIR results suggest that there is a strong interaction between carboxymethyl chitosan and lysozyme in the composition of nanogels.CMC/LYZ-ACP nanogels were compared with CMC-ACP nanoparticles.It was found that the particles were loose without lysozyme.The addition of lysozyme could form a more boundary CMC/LYZ-ACP nanomaterials;this gel can be applied in clinical.SEM images show that the thickness of the coating layer was approximately 1–2μm after ultrasonic cleaning.The depth to which the mineralized substance enters the dentinal tubule was approximately 4-8μm.While,the Green Or^TM group only formed the sediment on the surface of the dentinal tubule to block the tube,did not enter the inner tube.The results of XRD measurement indicate that both the CMC/LYZ-ACP nanogels group and the Green Or^TM group have obvious crystallization.And nanoindentation indicated that the occluding mineralized substance observed were similar to nature dentin in hardness and modulus of elasticity.ConclusionCMC can form spherical dense nanogels loaded with ACP under the participation of lysozyme.The CMC/LYZ-ACP nanogels could increase the dentinal tubule occluding effectiveness compared with Green Or^TM group.These results indicated that finding and developing novel nanomaterials of CMC/LYZ-ACP would be an effective strategy for the treatment of dentin hypersensitivity.