Comparative evaluation of microbial profiles of oral samples obtained at different collection time points and using different methods.

OBJECTIVES: Recently, the oral microbiome has been found to be associated with oral and general health status. Although various oral sample collection protocols are available, the potential differences between the results yielded by these protocols remain unclear. In this study, we aimed to determine the effects of different time points and methods of oral sample collection on the outcomes of microbiome analysis. MATERIALS AND METHODS: Oral samples were collected from eight healthy individuals at four different time points: 2 h after eating, immediately after teeth brushing, immediately after waking up, and 2 h after eating on the subsequent day. Four methods of saliva collection were evaluated: spitting, gum chewing, cotton swab, and oral rinse. Oral microbiomes of these samples were compared by analyzing the bacterial 16S rRNA gene sequence data. RESULTS: The oral microbial composition at the genus level was similar among all sample collection time points and methods. Alpha diversity was not significantly different among the groups, whereas beta diversity was different between the spitting and cotton swab methods. Compared with the between-subject variations, the weighted UniFrac distances between the groups were not minor. CONCLUSIONS: Although the oral microbiome profiles obtained at different collection time points and using different methods were similar, some differences were detected. CLINICAL RELEVANCE: The results of the present study suggest that although all the described protocols are useful, comparisons among microbiomes of samples collected by different methods are not appropriate. Researchers must be aware of the issues regarding the impact of saliva collection methods.

Microsurgical mandibular reconstruction using a resin surgical guide combined with a metal reconstructive plate.

INTRODUCTION: In mandibular reconstruction, repositioning the mandibular position is still challenging and time consuming. We invented a new re-positioning technique using a resin plate combined with a reconstructive plate in reconstructing the mandible with an osteocutaneous free flap. The purpose of this report is to introduce this technique and evaluate the accuracy of mandibular reconstruction using free flaps. We hypothesized that this technique is precise and can reduce intraoperative plate bending at a low cost and short preparation period. METHODS: Mandibular reconstruction was successfully performed in a total of 10 cases without any complications. In this technique, a pre-bent reconstructive plate was prepared in accordance with a three-dimensional model, and then coated with resin. Intraoperatively, the mandibles were secured by fitting these plates snugly and fixing them using a reconstructive plate. Then the resin was removed and free osteocutaneous free flaps were transfer to the defect. Ten patients with a mean age of 68.2 who underwent mandibular resection for aggressive benign (n = 1) or malignant disease (n = 9) were reconstructed using this technique. Seven cases were reconstructed using fibular osteocutaneous free flaps, while scapular osteocutaneous free flaps were used in the remaining cases. The resections entailed: unilateral symphysis and lateral body in four cases, angle to ipsilateral angle in two, ramus to symphysis in two, and lateral body plus angle to symphysis in one case. The deviation of the mandible was evaluated by measuring the preoperative versus postoperative differences in the distances between six bilateral landmarks. RESULTS: There were no complications and flap failure in any of the 10 cases. No further intraoperative plate bending was required. One case underwent additional mucosal resection due to recurrence of cancer. Three cases were referred to postoperative chemoradiotherapy. Two patients expired during follow-up due to recurrence of cancer. Six cases were put back on a normal diet. The other cases who lacked opposing teeth had to remain on a soft diet. The mean follow-up period was 46.2 months. The average of the absolute deviation values was 1.45 mm. This value was 0.94 mm in six cases with mandibular body defects and 2.26 mm in four cases with mandibular defects involving the ramus. CONCLUSIONS: The present novel technique is simple, quick to prepare, and accurate. This technique can be a viable option for microsurgical mandibular reconstruction.

Removal of a Fixation Screw That Was Forced Into the Postsuperior Maxillary Sinus Wall: Case Report.

We have encountered a rare case in which the subject underwent maxillary sinus floor elevation at another hospital, and a screw to fix the grafted bone substitute was forced into the maxillary sinus and intruded into the bone. Various different foreign bodies have been reported as being forced into the maxillary sinus due to dental treatment, and these foreign bodies are often retained on the maxillary sinus mucous membrane. However, no reports have described a screw forced in and intruded into the peculiar position in the bone, as seen in the present case, which we report here with additional discussion.

Reconstruction of the Alveolar Bone Using Bone Augmentation With Selective Laser Melting Titanium Mesh Sheet: A Report of 2 Cases.

Bone augmentation is used to supplement bone defects during dental implant treatment. In this technique, the area filled with bone prosthetic material is covered with an artificial space-making device or titanium mesh sheet, which must be manually adapted to the bone defect during the procedure before being fixed in place. Selective laser melting (SLM) method can be used to preadapt the titanium mesh sheet based on preoperative CT data. This method enables shorter surgery times compared with conventional titanium mesh sheet methods, as well as regeneration of an ideal alveolar bone shape. Here, we present 2 cases of bone augmentation using the SLM titanium mesh sheet method. The postoperative course was without complications in both cases; neither patient experienced mesh exposure or infection during healing. The SLM titanium mesh sheet method should be considered as a new and effective bone augmentation method.

Rapid and Easy Histological Evaluation of Alveolar Human Bone Quality at Dental Implant Sites Using a Nondecalcified Frozen Cryofilm Section Technique: A Technical Report.

INTRODUCTION: The evaluation of bone quality at the site of the alveolar bone for a dental implant is very important. This study presents an easy technique for direct evaluation of alveolar bone quality using nondecalcified cryofilm frozen sections on human alveolar bone core samples. MATERIALS AND METHODS: Core samples harvested from alveolar bone were immediately frozen in cooled hexanen and slowly cut using a disposable tungsten carbide blade; the sliced sections were collected with adhesive cryofilms. Staining was performed using von toluidine blue and von Kossa for microscopic observations. RESULTS: All core samples clearly showed bone structure components of cortical bone, trabecular bone, bone marrow, blood vessels, and bone-related cells. CONCLUSION: These results suggest the efficacy of a nondecalcified cryofilm frozen section technique for histological observation of surgical implant sites.

Clinical Study of 14 Cases of Bone Augmentation with Selective Laser Melting Titanium Mesh Plates.

Additive manufacturing techniques are being used in the medical field. Orthopedic hip prostheses and denture bases are designed and fabricated based on the patient's computer-aided design (CAD) data. We attempted to incorporate this technique into dental implant bone augmentation. Surgical simulation was performed using patient data. Fourteen patients underwent bone augmentation using a selective laser melting (SLM) titanium mesh plate. The results showed no evidence of infection in any of the 14 patients. In 12 patients, only one fixation screw was used, and good results were obtained. The SLM titanium mesh plate was good adaptation in all cases, with bone occupancy greater than 90%. The average bone resorption of the marginal alveolar bone from the time of dental implant placement to the time of the superstructure placement was 0.69 ± 0.25 mm. Implant superstructures were placed in all cases, and bone augmentation with SLM titanium mesh plates was considered a useful technique.

Application of the Homologous Modeling Technique for Precision Medicine in the Field of Oral and Maxillofacial Surgery.

In the field of oral and maxillofacial surgery, establishment of a new method for predicting morphology is desirable. Therefore, the purpose of the present study was to establish a new method for predicting the original shape of a mandibular defect site using the homologous modeling technique. This study used data from 44 patients who underwent computed tomography in the Department of Oral Surgery at Osaka Medical College. Two types of homologous models were constructed: total mandible (TM) and half mandible (HM). Principal component analysis (PCA) was performed using point cloud data of the homologous model M and homologous model HM, and a multiple regression equation was created using the PC value of TM as the object variable and PC value of HM as the explanatory variable. The predicted PC (M) was created from PC (HM) using a regression formula, back-calculated from point cloud data from PC (M), to create the predicted mandible model. Finally, the original image (TC-M) and estimated mandible were superposed and examined. The mean absolute error between the predicted mandible and actual mandible was 1.04 ± 1.35 mm. We believe that this method will be applicable in actual clinical practice.

The effect of simple heat treatment on apatite formation on grit-blasted/acid-etched dental Ti implants already in clinical use.

Grit-blasted/acid-etched titanium dental implants have a moderately roughened surface that is suitable for cell adhesion and exhibits faster osseointegration. However, the roughened surface does not always maintain stable fixation over a long period. In this study, a simple heat treatment at 600°C was performed on a commercially available dental Ti implant with grit-blasting/acid-etching, and its effect on mineralization capacity was assessed by examining apatite formation in a simulated body fluid (SBF). The as-purchased implant displayed a moderately roughened surface at the micrometer scale. Its surface was composed of titanium hydride accompanied by a small amount of alumina particles derived from the grit-blasting. Heat treatment transformed the titanium hydride into rutile without evidently changing the surface morphology. The immersion in SBF revealed that apatite formed on the heated implant at 7 days. Furthermore, apatite formed on the Ti rod surface within 1 day when the metal was subjected to acid and heat treatment without blasting. These indicate that apatite formation was conferred on the commercially available dental implant by simple heat treatment, although its induction period was slightly affected by alumina particles remaining on the implant surface. The heat-treated implant should achieve stronger and more stable bone bonding due to its apatite formation.

Short communication: Distribution of phospholipids in parotid cancer by matrix-assisted laser desorption/ionization imaging mass spectrometry.

BACKGROUND: Parotid cancer is relatively rare, and malignancy varies; therefore, novel markers are needed to predict prognosis. Recent advances in matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS), useful for visualization of lipid molecules, have revealed the relationship between cancer and lipid metabolism, indicating the potential of lipids as biomarkers. However, the distribution and importance of phospholipids in parotid cancer remain unclear. OBJECTIVE: This study aimed to use MALDI-IMS to comprehensively investigate the spatial distribution of phospholipids characteristically expressed in human parotid cancer tissues. METHODS: Tissue samples were surgically collected from two patients with parotid cancer (acinic cell carcinoma and mucoepidermoid carcinoma). Frozen sections of the samples were assessed using MALDI-IMS in both positive and negative ion modes, with an m/z range of 600-1000. The mass spectra obtained in the tumor and non-tumor regions were compared and analyzed. Ion images corresponding to the peak characteristics of the tumor regions were visualized. RESULTS: Several candidate phospholipids with significantly different expression levels were detected between the tumor and non-tumor regions. The number of unique lipid peaks with significantly different intensities between the tumor and non-tumor regions was 95 and 85 for Cases 1 and 2, respectively, in positive ion mode, and 99 and 97 for Cases 1 and 2, respectively, in negative ion mode. Imaging differentiated the characteristics that phospholipids were heterogeneously distributed in the tumor regions. CONCLUSION: Phospholipid candidates that are characteristically expressed in human parotid cancer tissues were found, demonstrating the localization of their expression. These findings are notable for further investigation of alterations in lipid metabolism of parotid cancer and may have potential for the development of phospholipids as biomarkers.

Bioactivation Treatment with Mixed Acid and Heat on Titanium Implants Fabricated by Selective Laser Melting Enhances Preosteoblast Cell Differentiation.

Selective laser melting (SLM) is a promising technology capable of producing individual characteristics with a high degree of surface roughness for implants. These surfaces can be modified so as to increase their osseointegration, bone generation and biocompatibility, features which are critical to their clinical success. In this study, we evaluated the effects on preosteoblast proliferation and differentiation of titanium metal (Ti) with a high degree of roughness (Ra = 5.4266 ± 1.282 µm) prepared by SLM (SLM-Ti) that was also subjected to surface bioactive treatment by mixed acid and heat (MAH). The results showed that the MAH treatment further increased the surface roughness, wettability and apatite formation capacity of SLM-Ti, features which are useful for cell attachment and bone bonding. Quantitative measurement of osteogenic-related gene expression by RT-PCR indicated that the MC3T3-E1 cells on the SLM-Ti MAH surface presented a stronger tendency towards osteogenic differentiation at the genetic level through significantly increased expression of Alp, Ocn, Runx2 and Opn. We conclude that bio-activated SLM-Ti enhanced preosteoblast differentiation. These findings suggest that the mixed acid and heat treatment on SLM-Ti is promising method for preparing the next generation of orthopedic and dental implants because of its apatite formation and cell differentiation capability.

Histological Evaluation of Porous Additive-Manufacturing Titanium Artificial Bone in Rat Calvarial Bone Defects.

Jaw reconstruction using an additive-manufacturing titanium artificial bone (AMTAB) has recently attracted considerable attention. The synthesis of a titanium artificial bone is based on three-dimensional computed tomography images acquired before surgery. A histological evaluation of porous AMTAB (pAMTAB) embedded in rat calvarial bone defects was conducted. This study examined three groups: rats implanted with mixed-acid and heat-treated pAMTAB, rats implanted with untreated pAMTAB, and rats with no implant. In both pAMTAB groups, bone defects were created in rat calvarial bones using a 5-mm trephine bar, followed by pAMTAB implantation. The pAMTAB was fixed to the defect using the fitting force of the surrounding bones. The rats were sacrificed at 4, 8, and 16 weeks after implantation, and the skull was dissected. Undecalcified ground slides were prepared and stained with Villanueva Goldner. Compared with the no implant control group, both pAMTAB groups exhibited new bone formation inside the defect, with greater bone formation in the mixed-acid and heat-treated pAMTAB group than in the untreated pAMTAB group, but the difference was not significant. These data suggest that pAMTAB induces bone formation after implantation in bone defects. Bone formation appears to be enhanced by prior mixed-acid and heat-treated pAMTAB.

Examination of new parameters for sex determination of mandible using Japanese computer tomography data.

OBJECTIVE: In the field of forensic science, sex discrimination of skeletons is an important identification item for personal identification. The individual sex discrimination method using skeletons includes a determination method using measurement values and a macroscopic form observation method. Both methods have advantage and disadvantage. In this study, we used the homologous model technique and principal component (PC) analysis to determine gender difference from morphology of the mandible. METHODS AND MATERIALS: 45 patients (23 males and 22 females) of CT imaging for tooth extraction from January 2018 to March 2019 at department of oral surgery, Osaka Medical College. The mean age was 43.1 ± 14.6. Patients with less than 14 remaining teeth were excluded because the number of remaining teeth may affect the shape of the mandible. 3D images were constructed, and 20 landmarks plotting on the 3D model surfaces. We generated template models of the mandible consisting of approximately 8434 polygons. The template model automatically fitted into the individually scanned point cloud of the mandible by minimising external and internal energy functions. As described above, the mandibles were constructed for each sample by using the Homologous Body Modeling software (HBM, Digital Human Technology, Inc.) and the mHBM-Rugle (Medic Engineering Corporation). The mandibles were analysed using the PCA. RESULTS: The contribution of the most important PC was found to be 27.2%. 12 PCs explained over 75% of the total variance. That is, it was able to express 75% or more of the mandible expression with 12 PCs. A significant difference between male and female was observed in the first PCs (Wilcoxon test, p < 0.05). Visualising the result of the first PC showed that the mandibular branch of male was larger than that of female, and the mandible angle was overhanging outside. CONCLUSION: This method is a combination of the determination method using the previous measurement values and the determination using macroscopic observation, and is considered to be innovative method.

Mechanical, Histological, and Scanning Electron Microscopy Study of the Effect of Mixed-Acid and Heat Treatment on Additive-Manufactured Titanium Plates on Bonding to the Bone Surface.

The additive manufacturing (AM) technique has attracted attention as one of the fully customizable medical material technologies. In addition, the development of new surface treatments has been investigated to improve the osteogenic ability of the AM titanium (Ti) plate. The purpose of this study was to evaluate the osteogenic activity of the AM Ti with mixed-acid and heat (MAH) treatment. Fully customized AM Ti plates were created with a curvature suitable for rat calvarial bone, and they were examined in a group implanted with the MAH-treated Ti in comparison with the untreated (UN) group. The AM Ti plates were fixed to the surface of rat calvarial bone, followed by extraction of the calvarial bone 1, 4, 8, and 12 weeks after implantation. The bonding between the bone and Ti was evaluated mechanically. In addition, AM Ti plates removed from the bone were examined histologically by electron microscopy and Villanueva-Goldner stain. The mechanical evaluation showed significantly stronger bone-bonding in the MAH group than in the UN group. In addition, active bone formation was seen histologically in the MAH group. Therefore, these findings indicate that MAH resulted in rapid and strong bonding between cortical bone and Ti.

Osteogenic capacity of mixed-acid and heat-treated titanium mesh prepared by a selective laser melting technique.

The practical use of additive manufacturing to create artificial bone as a material for repairing complex bone defects is currently attracting attention. In this study, we compared the osteogenic capacity of materials composited by the method developed by Kokubo et al. of treating 3D-printed titanium (Ti) mesh with a mixture of H2SO4 and HCl and heating (mixed-acid and heat treatment) with that of materials subjected to conventional chemical treatment. Ti plates treated with this method have been found to promote highly active bone formation on their surface when inserted into rabbit tibial bone defects. No previous study has compared this method with other surface treatment methods. In this study, we used histological and other observations to compare the bone formation process in bone defects when Ti meshes prepared by the selective laser melting technique (SLM) and treated either with mixed acids and heat or with conventional chemical Ti surface treatments were implanted in a rat calvarial bone defect model. We found that both micro-computed tomography and observations of undecalcified ground sections showed that the best bone formation was observed in rats implanted with mesh treated with mixed acids and heat. Our results suggest that mixed-acid and heat-treated Ti mesh prepared by SLM may have a high osteogenic capacity in bone defects.