[Osteogenesis of human adipose-derived mesenchymal stem cells-biomaterial mixture in vivo after 3D bio-printing].

OBJECTIVE: To construct human adipose-derived mesenchymal stem cells (hASCs)-biomaterial mixture 3D bio-printing body and detect its osteogenesis in vivo, and to establish a guideline of osteogenesis in vivo by use of 3D bio-printing technology preliminarily. METHODS: P4 hASCs were used as seed cells, whose osteogenic potential in vitro was tested by alkaline phosphatase (ALP) staining and alizarin red staining after 14 d of osteogenic induction. The cells were added into 20 g/L sodium alginate and 80 g/L gelatin mixture (cell density was 1 × 10(6)/mL), and the cell-sodium alginate-gelatin mixture was printed by Bioplotter 3D bio-printer (Envision company, Germany), in which the cells'survival rate was detected by live- dead cell double fluorescence staining. Next, the printing body was osteogenically induced for 1 week to gain the experimental group; and the sodium alginate-gelatin mixture without cells was also printed to gain the control group. Both the experimental group and the control group were implanted into the back of the nude mice. After 6 weeks of implantation, the samples were collected, HE staining, Masson staining, immunohistochemical staining and Inveon Micro CT test were preformed to analyze their osteogenic capability. RESULTS: The cells'survival rate was 89%± 2% after printing. Six weeks after implantation, the samples of the control group were mostly degraded, whose shape was irregular and gel-like; the samples of the experimental group kept their original size and their texture was tough. HE staining and Masson staining showed that the bone-like tissue and vessel in-growth could be observed in the experimental group 6 weeks after implantation, immunohistochemical staining showed that the result of osteocalcin was positive, and Micro CT results showed that samples of the experimental group had a higher density and the new bone volume was 18% ± 1%. CONCLUSION: hASCs -biomaterial mixture 3D bio-printing body has capability of ectopic bone formation in nude mice, and it is feasible to apply cells-biomaterial mixture 3D bio-printing technology in the area of bone formation in vivo.

[Short-term effect of CaCl2on human adipose-derived mesenchymal stem cells proliferation and osteogenic differentiation].

OBJECTIVE: To examine the effect of CaCl2, a sodium alginate crosslinker, to stimulate cells for a short time period on human adipose-derived mesenchymal stem cells (hASCs) proliferation and osteogenic differentiation ability, and to determine the appropriate concentration of CaCl2for post three-dimensional biological experiments. METHODS: hASCs stimulated with or without CaCl2at various concentrations were seeded and cultured in control medium and osteogenic medium, respectively. The cell counting kit-8 (CCK8) was used to estimate the cell proliferation level of each group. After 7 days of osteogenic induction, alkaline phosphatase (ALP) staining and activity assays were performed using an ALP kit. After 14 days of osteogenic induction, alizarin red staining and quantitative detection were used to determine the calcium mineral density. The results were analyzed using analysis of variance (ANOVA) and Student-Newman-Keuls (SNK) tests for pairwise comparisons implemented in the SPSS 17.0 software. RESULTS: The CCK-8 assays showed that the differences between the control groups and experimental groups were not statistically significant, so different concentrations of CaCl2had no significant effect on hASCs proliferation. The ALP staining and activity assays showed that ALP activity first increased and then decreased as the CaCl2concentration increased. Furthermore, the differences between all the groups were statistically significant (P<0.05), except the difference between the 50 mmol/L CaCl2group and the 100 mmol/L CaCl2group, and between the osteogenetic medium(OM) group and the 200 mmol/L CaCl2group. Alizarin red staining and quantitative detection showed that the differences between all pairwise combinations of the groups were statistically significant (P<0.05). As the CaCl2 concentration increased, the calcium deposition increased, initially in the form of a scattered sheet and eventually a laminated sheet. CONCLUSION: Stimulation by a high concentration of CaCl2over a short time period can enhance hASCs osteogenic differentiation ability, but has no effect on hASCs proliferation.

[Chemical constituents of Sapium sebiferum leaves].

Chemical constituents of ethyl acetate extract of Sapium sebiferum leaves were isolated and purified by various chromatographic methods, including column chromatographies over silica gel, macroporous adsorption resin, and Sephadex LH-20, as well as preparative TLC and semi preparative HPLC. As a results, 15 compounds were separated from Sapium sebiferum leaves and their structures were examined by spectral analysis including NMR and MS data and identified as( + )-(7R,7'R,7"S,7'"S,8S,8'S,8"S,8'"S)-4", 4"'-dihydroxy-3,3',3",3',5,5'-hexamethoxy-7,9';7',9-diepoxy-4,8";4',8'"-bisoxy-8,8'-dineo-lignan-7",7"',9",9"'-tetraol(1) ,1-(4'- hydroxy-3'-methoxyphenyl)-2-[4"-(3-hydroxypropyl) -2", 6"-dimethoxyphenoxy] propane-1, 3-diol (2), Thero-2, 3-bis-(4-hydroxy-3- methoxypheyl)-3-methoxy-propanol(3) , threo-5-hydroxy-3,7-dimethoxyphenyl propane-8,9-diol (4), boropinol B (5), threo-8S-7-methoxysyringylglycerol(6), 5-hydroxymethylfurfural(7), 5-( methoxy-methyl)-1H-pyrrole-2-carbaldehyde (8), quercetin (9) , kaempferol (10), ethyl gallate(11), coniferaldehyde(12), vanillin(13), 7-hydroxy-6-methoxy-2H-1-henzopyran-2-one(14),and 1-heptacosanol (15). All compounds except for compounds 9-11,14 were separated from this plant for the first time.

[Three dimensional bioprinting technology of human dental pulp cells mixtures].

OBJECTIVE: To explore the three dimensional(3D)bioprinting technology, using human dental pulp cells (hDPCs) mixture as bioink and to lay initial foundations for the application of the 3D bioprinting technology in tooth regeneration. METHODS: Imageware 11.0 computer software was used to aid the design of the 3D biological printing blueprint. Sodium alginate-gelatin hydrosol was prepared and mixed with in vitro isolated hDPCs. The mixture contained 20 g/L sodium alginate and 80 g/L gelatin with cell density of 1×10(6)/mL. The bioprinting of hDPCs mixture was carried out according to certain parameters; the 3D constructs obtained by printing were examined; the viability of hDPCs after printing by staining the constructs with calcein-AM and propidium iodide dye and scanning of laser scanning confocal microscope was evaluated. The in vitro constructs obtained by the bioprinting were cultured, and the proliferation of hDPCs in the constructs detected. RESULTS: By using Imageware 11.0 software, the 3D constructs with the grid structure composed of the accumulation of staggered cylindrical microfilament layers were obtained. According to certain parameters, the hDPCs-sodium alginate-gelatin blends were printed by the 3D bioprinting technology. The self-defined shape and dimension of 3D constructs with the cell survival rate of 87%± 2% were constructed. The hDPCs could proliferate in 3D constructs after printing. CONCLUSION: In this study, the 3D bioprinting of hDPCs mixtures was realized, thus laying initial foundations for the application of the 3D bioprinting technology in tooth regeneration.

[Quantitative evaluation for the accuracy of dental model three-dimensional scanner].

OBJECTIVE: To establish a method to evaluate dental model three-dimensional scanner quantitatively, and to evaluate the accuracy which is a core indicator of 3Shape D700 scanner. METHODS: A standard geometric model similar to the dental arch was designed by three-dimensional reverse software and processed by high precision CNC (computer numerical control) processing technology. Core indicators of dental model three-dimensional scanner including single scanning accuracy, space consistency and rescan accuracy were evaluated. RESULTS: The result of single scanning accuracy of 3Shape D700 scanner was (15.00±10.84) µm, and there was no statistics difference between the accuracy given by manufacturer's instructions which is 20 µm (P=0.053), and same as the results of space consistency (compare the accuracy in vertical direction and horizontal direction, P=0.524) and rescan accuracy (compare the rescan accuracy in vertical direction, P=0.633, and in horizontal direction P=0.221). CONCLUSION: It is feasible to evaluate accuracy of dental model three-dimensional scanner by this method, which can avoid observer error caused by selecting points manually.

[Quantitative assessment on artifacts of dental restorative materials in cone beam computed tomography].

OBJECTIVE: To quantitatively evaluate the artifacts appearance of eight kinds of common dental restorative materials, such as zirconia. METHODS: For the full-crown tooth preparation of mandibular first molar, eight kinds of full-crowns, such as zirconia all-ceramic crown, glass ceramic crown, ceramage crown, Au-Pt based porcelain-fused-metal (PFM) crown, Pure Titanium PFM crown, Co-Cr PFM crown, Ni-Cr PFM crown, and Au-Pd metal crown were fabricated. And natural teeth in vitro were used as controls. These full-crown and natural teeth in vitro were mounted an ultraviolet-curable resin fixed plate. High resolution cone beam computed tomography (CBCT) was used to scan all of the crowns and natural teeth in vitro, and their DICOM data were imported into software MIMICS 10.0. Then, the number of stripes and the maximum diameters of artifacts around the full-crowns were evaluated quantitatively in two-dimensional tomography images. RESULTS: In the two-dimensional tomography images,the artifacts did not appear around the natural teeth in vitro, glass ceramic crown, and ceramage crown. But thr artifacts appeared around the zirconia all-ceramic and metal crown. The number of stripes of artifacts was five to nine per one crown. The maximum diameters of the artifacts were 2.4 to 2.6 cm and 2.2 to 2.7 cm. CONCLUSION: In the two-dimensional tomography images of CBCT, stripe-like and radical artifacts were caused around the zirconia all-ceramic crown and metal based porcelain-fused-metal crowns. These artifacts could lower the imaging quality of the full crown shape greatly. The artifact was not caused around the natural teeth in vitro, glass ceramic crown, and ceramage crown.

[Study on low temperature degradation of dental zirconia ceramic].

OBJECTIVE: To evaluate the properties of low temperature degradation (LTD) of dental Y-TZP by experiments. METHODS: According to ISO 13356-2008 standards, 72 blanks of Y-TZP were sintered at different temperature (1 400 °C,1 450 °Cor 1 530 °C) and in different heat preserving time(2 h or 5 h), and then the specimens were grounded and polished. The blanks were divided into 12 groups according to the different sintering temperature, heat preserving time and aging time. The surface of specimens was observed by SEM to evaluate the grain size of specimens; Each group of specimens was exposed to 140 °C hydrothermal aging environment for 0 h, 5 h or 24 h. After aging, XRD analysis was undertaken to determine the amount of phase transformation from tetragonal to monoclinic for each groups. At last, the biaxial flexural strengths of all specimens were measured and calculated. RESULTS: The SEM showed that the grain size of each group of Y-TZP increased with the increase of sintering temperature and heat preserving time, however, all of them were below 1 µm. The XRD analysis showed the contents of monoclinic of each group varied with different sintering temperatures, heat preserving times and aging time. The flexural strength of 24 h aging groups declined significantly; but that of 5 h aging group showed increase or decrease according to the different sintering temperatures and heat preserving times. CONCLUSION: Different sintering procedures to some extent may affect the Y-TZP aging properties.

[Three-dimensional model analysis of the gingival sulcus width from different retraction time].

OBJECTIVE: To explore the effect of gingival retraction time on the gingival sulcus width in different gingival areas, using three-dimensional (3D) model analysis. METHODS: Forty-six premolars from 10 volunteers were included, gingival retraction cords were applied on the buccal sulcus for three times, and the retraction time was 5, 10 and 15 minutes respectively. The least interval between two retraction procedures was 2 weeks. Impressions and stone models were made before and after gingival retraction. 3D data of the models were acquired by a 3D line laser scanner. Models were superimposed and tooth coordinate was set up with special software. Sulcus widths from different retraction time and different gingival areas were measured and compared. RESULTS: Sulcus widths between retraction of 5 and 10 minutes were significantly different at midbuccal and transitional line angle (TLA) gingival areas. At TLA areas, sulcus widths between retraction of 10 and 15 minutes were also statistically different, but the difference was not significant when compared with that of 5 and 10 minutes. Sulcus widths at midbuccal areas were significantly greater than that at TLA areas for all three retraction time. CONCLUSION: Before making impressions, gingival retraction time of single-cord mechanical technique should be at least 10 minutes. For different gingival areas, the enlargement of gingival sulcus from gingival retraction and the optimal retraction time were different. 3D model analysis could be used as a new objective method to evaluate gingival retraction effect from different retraction time.

[Digital inlaid design based on anatomical characteristics and preliminary implementation of its program].

OBJECTIVE: To explore the technical guideline of inlays computer aided design (CAD) for further development, to test the feasibility of processing for the design results of the technical guideline, and to assist co-operation with automatic inlay CAD software development. METHODS: On the basis of point cloud data of right mandibular first molar of standard crown, we established the coordinate system,the landmarks and lines and the characteristic curves. A dental cast of right mandibular first molar MOD-inlay of an individual normal occlusion was scanned. The following objects were digitized: prepared tooth, neighbor teeth and intercuspal bite record. On the basis of parametric standard crown established in the research, using Surfacer 11.0, we achieved the reconstruction of MOD-inlay by extracting margin line, constructing tissue surface, constructing outer surface and occlusal grinding, then the design result was processed and try-in on the cast. We also developed automatic inlay CAD software based on the guideline of inlay CAD. RESULTS: The guideline of inlay CAD was established, the design results were processed and an automatic inlay CAD software was developed on basis of Surfacer 11.0. CONCLUSION: It is a feasible method to establish technical guideline of inlay based on reverse engineering technology and process its design result, and a automatic inlay CAD software could be developed based on the technical guideline.

[Computer-aided design for occlusal rest of the removable partial denture].

OBJECTIVE: To explore the route of automatic computer-aided design for occlusal rest of the removable partial denture. METHODS: A new module for the design of occlusal rest was developed. The three-dimensional data of a partially dentate dental cast was obtained and imported into the new module. The contour of the occlusal's tissue surface was identified. Then, the tissue surface and polished surface were constructed respectively and connected to each other to form an integrated entity. RESULTS: The surface of the occlusal rest was smooth and the curvature matched that of the surrounding area of the abutment tooth. The accuracy was satisfying. CONCLUSION: Design of occlusal rest was much more convenient and time-saving using the newly developed module. The future development of special software package for the design of removable partial denture will benefit from it.

[Methodology of computer-aided design of automatic artificial tooth selection for complete denture].

OBJECTIVE: To study the method of automatic tooth selection for computer-aided design in complete denture, evaluate the results by comparison with traditional denture tooth selection. METHODS: 3D data of occlusal rim made by the dentist were obtained with a 3D laser scanner and specific points on the occlusal rim were measured on Imageware11 platform. Based on Matlab 7.1 software platform, an automatic tooth selection software was programmed with adequate consideration of complete denture tooth selection factors, for example: face form of the patient, mesiodistal diameter of the artificial anterior teeth and posterior teeth, etc. Oclusal rim and edentulous models were selected from 20 patients. The artifical teeth were selected by both technician and software. The result of automatic tooth selection was evaluated by comparison with traditional method. RESULTS: In 20 cases the concordant rate of automatic and traditional method is 70%. CONCLUSION: The method of automatic tooth selection for computer-aided design in complete denture was realized in accordance with traditional method of denture tooth selection, the relationship between the curve of artificial teeth arranged on the occlusal rim and cuspid line--which was calculated accurately; automatic denture tooth selection; the denture tooth selected by software can be directly used in CAD complete denture tooth arrangement.

[Measurement and estimation of the root surface of single-root teeth].

OBJECTIVE: To evaluate the availability of the estimation method of the single-root surface area on periapical radiographs, and to compare with the measured method by 3-dimension laser scanning and reconstructed images. METHODS: Ninety single-root teeth with severe periodontitis were selected, periapcal radiographs were taken before teeth extracted. Root lengths and crown lengths were measured on periapical radiographs. The ratio of crown to root of each tooth was calculated. Then the root surface areas were also estimated according to mathematics limit principle on the periapical radiographs. The image of the roots were reconstructed by using a 3D laser scanning, the root surface areas were measured with a CAD/CAM software. RESULTS: No significant difference was found between the measured values of the ratio of crown to root by 3D laser scanning and the estimated values on periapical radiographs (t=0.58, P>0.05). For the lower incisors, no significant difference was found between the measured root surface areas with the 3D laser scanning and the estimated ones on periapical radiographs(t=0.46, P>0.05). The surface area of the root in each tooth was divided into three parts: the coronal 1/3, the middle 1/3 and the apical 1/3. Each part of root surface area was measured and estimated. And then the percentage of each part in total root surface area was calculated. The measured percentage was similar to the estimated one except the middle 1/3 part of the upper premolars(t=0.77, P>0.05). CONCLUSION: No significant difference was found between the measured ratio of crown to root with 3D laser scanning and the estimated one on periapical radiographs. The evaluated method on periapical radiographs to estimate root surface areas was relatively reliable for the lower incisor.

[A method for three-dimensional analysis of occlusal contacts from digital dental casts].

OBJECTIVE: The study was to investigate the occlusal contacts with a set of dental casts based on their three-dimensional models in the intercuspal position. METHODS: 3D point-clouds of the dental casts in the intercuspal position were acquired by D.02-L-3D scanner with a special casts-locating device, and reconstructed . The areas of occlusal contacts were calculated, and were further compared with the results from the methods of articulating paper and photocclusion technique. RESULTS: There was no significant difference in the result of occlusal contacts analysis between the 3D reconstructive measurement and articulating paper or photocclusion technique. The virtual articulating paper was also preliminarily carried out in this research. CONCLUSION: The study verified the reliability of the measurement for occlusal contacts by the 3D digital model, and may lead to a new way for occlusal contacts research.

[Construction of inlay axial surface using B-Spline curve network].

OBJECTIVE: To realize the requirement of adjacent area accuracy, axial surface physical contour and smoothness in the process of inlay computer-aided design (CAD). METHODS: A compromised method was brought forward that some assistant 3-D constraint lines from occlusion to gingiva were used to lead the inlay contour on axial corner and other characterized areas, and two perpendicular 3-D constructing lines at adjacent area were used to construct accurate 3-D configuration of this area. The axial surface was completely defined by the network of specified curves which were two constructing lines and four borders. RESULTS: Inlay axial surface was smooth, the contour was consistent with the residual tooth body and the 3-D configuration in adjacent area was accurate. CONCLUSION: It is feasible to realize the inlay axial surface accuracy, smoothness, and consistency using the method of special distributed 3-D B-Spline curve network.

[A scanning method for the dental casts of intercuspal position].

OBJECTIVE: To acquire the 3D point-cloud of the dental casts of intercuspal position and rebuild it in computer. METHODS: A special scanning method was used to acquire the point-cloud of dental cast by using the D.02-L-3D SCANNER and rebuild the model by computer. RESULTS: The method could acquire a 3D model with high resolving power and clear-cut dental anatomic configuration for the dental casts of intercuspal position. CONCLUSION: This is a viable method of scanning with many advantages, such as precision, simplicity, high efficiency. It could satisfy the requirements of clinical practice, teaching and scientific research.

Coordinated control and experimentation of the dental arch generator of the tooth-arrangement robot.

BACKGROUND: The traditional way of acquiring the dental arch curve form is based on manual operation, which will randomly bring numerous errors caused by human factors. The purpose of this paper is to automatically acquire the dental arch curve and implement the coordinated control of the dental arch generator of the multi-manipulator tooth-arrangement robot, which can be used in full denture manufacturing. METHODS: Based on the arc length constant theory, kinematics on dental arch generator is analyzed. The control pulse realization methods of high-resolution timing and CPU time-stamp timing are studied, and testing and comparative analysis of the control precision and stability of the two methods is carried out. Control experimentation of the dental arch generator and preliminary tooth-arrangement experimentation are performed using the multi-manipulator tooth-arrangement robot system. RESULTS: The dental arch generator can automatically generate a dental arch curve that fits a patient according to the patient's jaw arch parameters. Repeated positioning accuracy is 0.12 mm for the slipways which drive the dental arch generator. The maximum value of single point error is 1.64 mm when the arc width direction (x-axis) is 37.25 mm. The experimental results indicate that the method of control pulse realized by high-resolution timing to achieve high precision coordinated motion control of dental arch generator of tooth-arrangement robot is feasible. The error analysis results indicate that the control strategy and technical route can fulfill the requirements for motion speed and location precision. CONCLUSION: A novel system to generate the tooth arch curve has been developed. The traditional method of manually determining the dental arch may soon become obsolete in favour of the use of a robot to assist in generating a more standard tooth arch curve. The system can be used to manufacture a full denture. It will lay an important theoretical foundation for quantitative research of oral restoration, and also provide a way to standardize the manufacturing process of full denture.

[3D-data acquisition of partially edentulous cast using grating projection method].

OBJECTIVE: A method for 3D-data acquisition of partially edentulous cast was introduced to lay the foundation for reconstructing 3D digital model of a partially edentulous cast and studying computer aided design (CAD) to a removable partial denture framework. METHODS: Using projection system controlled by a computer, four frames of sinusoidal fringe pattern at pi/2 intervals were projected onto the surface of partially edentulous cast consecutively within 0.4 second. Deformed fringe patterns modulated by the surface height of the cast were recorded by a digital camera. The phase value with the height information was demodulated with phase-shifting method. The transition from the unwrapped phase to the height of the points on the cast was performed with triangulation method. The multiview 3D data was integrated automatically to obtain the complete 3D profile data and the 3D digital model of the partially edentulous cast was reconstructed. RESULTS: Dense and complete points-cloud data was achieved without scanning blind zone basically. The anatomic structure of reconstructed 3D digital model was sharp and clear. CONCLUSION: As an alternative for acquiring 3D data of partially edentulous cast, this method presented here with high precision and high speed meets the need for the subsequent 3D design to removable partial denture framework.

[Design of geometric models of the removable partial denture framework].

PURPOSE: The aim of this study was to introduce a method on computer aided design (CAD) of removable partial denture(RPD) framework based on domestic CAD/CAM software system, which contributes to the further research to develop the domestic software system applied in restorative dentistry. METHODS: Point cloud data of a partially edentulous cast, a mandibular Kennedy Class II modification 2 arch, was captured by an optical scanning system with projective grating and high-resolution digital camera. Using domestic CAD/CAM software system, the above point cloud data was reduced, digital surveying line and inserting path were determined. In terms of the principle of clinical design, the tissue surfaces and polishing surfaces of every component of RPD were built such as clasp, rest, lingual bar, and mesh construction, et al. As well as the characteristic structures of the framework, including the half-pear shaped cross section of lingual bar, the internal and external finish lines, the tissue stop at the tissue surface of mesh construction and so on, were fabricated on the base of reduced data model. 3-D surface model of the RPD framework was created. RESULTS: Geometric model of the RPD framework was fabricated successfully, which took on good fitting, high visibility and editing conveniently. And the data converted to STL file format that could be read by any other CAD/CAM software system and was in preparation for subsequent computer aided manufacture (CAM) of RPD framework. CONCLUSIONS: It is suggested that this method based on domestic software in CAD of RPD framework be feasible.