The effects of collagen cross-link deficiency on osseointegration process of pure titanium implants.

PURPOSE: This study aimed to investigate the effect of collagen cross-link deficiency on collagen fiber formation around an implant and its effect on the osseointegration process. METHODS: Wistar rats were fed 0.1% beta-aminopropionitrile (BAPN) dissolved in water to induce collagen cross-link deficiency. Custom-made mini-implants with machined surfaces were placed proximal to the tibia. At 1, 2, and 4 weeks postoperatively, the bone area around the implant, bone-implant contact ratio, osteoclast/osteocyte activity, and osseointegration strength were evaluated using histological and immunohistochemical analyses and biomechanical tests. RESULTS: Long-term disturbance of collagen cross-link formation in the BAPN group resulted in faster collagen fiber maturation than that in controls, with a defective collagen structure, low bone formation quantity, and low bone-implant contact values. Deficiency of collagen cross-links resulted in increased bone resorption and decreased osteocyte activity. CONCLUSIONS: Collagen cross-linking is important for the formation of the collagen matrix, and their deficiency may impair bone activity around implants, affecting the osseointegration process.

The relationship between dental metal allergy, periodontitis, and palmoplantar pustulosis: An observational study.

PURPOSE: This study aimed to investigate the relationship between dental metal allergy, periodontitis, and palmoplantar pustulosis among patients from a dental metal allergy clinic over a period of 8 years. METHODS: This study included 436 patients who visited our dental metal allergy clinic between April 1, 2009 and March 31, 2016. Diagnoses of skin diseases, periodontal records, dental metal series patch test results, and electron probe microanalysis (EPMA) data were obtained from medical records. Relative risk (RR) values were estimated from these data. RESULTS: Of the 359 patients who underwent the patch test, 241 showed a positive reaction. Of the 187 patients who underwent EPMA, 113 had allergenic metals in their dental prostheses. These patients were suspected to have a dental metal allergy. Furthermore, 150 of the 436 patients were diagnosed with palmoplantar pustulosis (PPP). The RR of metal allergy between patients with PPP and healthy subjects was 3.88. The RR of periodontal disease between patients with PPP and PPP-negative patients in the national average was 2.54. CONCLUSION: In this study, both dental metal allergy and periodontitis showed a high RR for PPP.

Comparing dental student preclinical self-assessment in the United States and Japan.

PURPOSE/OBJECTIVES: Self-assessment is an essential skill for dental professionals. Understanding global trends in self-assessment can highlight the learning needs of students across a diversity of cultural backgrounds. The aim of this study is to compare the self-assessment ability of dental students in the United States and Japan, where cultural backgrounds may differ. METHODS: Students in the United States (n = 176) completed a typodont premolar and anterior Class II and Class III preparation and restoration. Students in Japan (n = 175) completed a typodont premolar crown preparation. Students and faculty then evaluated the student performance using rubrics for each respective procedure. The difference between the student's self-assessment score and the average faculty score (S-F gap) was calculated and the data were analyzed. RESULTS: The mean S-F gap was 2.8% in Japan and 7.6% in the United States. This indicates that Japanese students tended to assess themselves closer to their faculty graders than students in the United States. On average, students in both countries scored themselves higher than their faculty graders. Students in the United States more frequently overestimated their performance and students in Japan more frequently underestimated their performance. For students in the lower quartile, the mean S-F gap was 5.1% in Japan and 14.6% in the United States, indicating a large cultural discrepancy in the lower quartile groups. CONCLUSIONS: Although different preclinical procedures were compared, our findings demonstrated that Japanese students may score themselves more closely to their faculty assessors than students in the United States. Further investigation with more students completing the same preclinical activity will be needed.

Technical skill training and assessment in dental education.

Highly competent clinical practice requires cognitive, psychomotor and affective skills. Therefore, the ultimate goal of dental education is for practitioners to be competent in all of these domains. While many methods have been introduced to assess knowledge and non-technical skills, it is still very difficult for educators to assess technical skill. Assessment methods for technical skills are still not well established because it is very difficult to assure objectivity, validity and fairness. Nonetheless, technical skill is especially important in dental treatments, along with knowledge and attitude. The aim of this review was to summarize the methods of technical skill training in dental education and how they are assessed. This is a literature review. We searched PubMed MEDLINE using terms related to technical skill training and those assessment as of June 2020 and reviewed them. There have been many reports introducing methods of technical skill training and assessment, including the use of digital technology. However, no single assessment method had demonstrated validity of it. Technical skill training is very important in dental education and there are various ways of learning. The validity of current assessment methods is limited; therefore, a combination of several methods may achieve the best results.

Bone heating and implant removal using a high-frequency electrosurgical device: An in vivo experimental study.

OBJECTIVES: Failed implant removal using a high-frequency electrosurgical device (HFED) has been reported to be less invasive than other surgical techniques. We sought to clarify the mechanism of removal torque reduction in an implant by heating with HFED. MATERIALS AND METHODS: Sixty-eight Wistar rats received titanium implants on the maxillary bone 4 weeks after extraction of the first and second molars. The control group was sacrificed 6 weeks after implant installation. In the experimental group, the implant was heated by HFED for 10 s using three different power outputs, and samples were collected at 3, 7, and 14 days after heating. Removal torque measurement and histological analysis were performed in the control and experimental groups. Implant surfaces were observed using an electron-probe microanalyzer (EPMA). Data were analyzed using Mann-Whitney U test at a significance level of 5%. RESULTS: The removal torque could not be measured in the control group due to fracture of the implant. After heating, the removal torque was measurable without fracture and decreased significantly at 14 days as compared with that at 3 days (p < .05). Heating with "min" power output resulted in a significantly smaller blank lacunae area and fewer osteoclasts at 14 days after heating (p < .05). EPMA revealed bone matrix adherence to outer surface of heated implant. CONCLUSIONS: After heating, an enlarged area of blank lacunae around the implant and an increased number of osteoclasts into the bone marrow cavity were observed, which may have contributed to the reduction in removal torque.

Peri-implant bone alterations under the influence of abutment screw preload stress. A preclinical vivo study.

OBJECTIVES: The study purpose was to examine peri-implant bone alternations around osseointegrated implants caused solely by abutment screw preload stress using different tightening torque values. MATERIALS AND METHODS: Twenty 20- to 22-week-old Japanese white rabbits received two implants each in right and left femurs. Implants were randomly assigned to one of three tightening torque groups or the control (Cont) group. After 8 weeks, 35 Ncm torque was delivered to abutment screws in the recommended torque (RT) group (n = 16). Other screws received 70 Ncm torque as the high torque (HT) group (n = 16). Temporary tightening (TT) groups (n = 8) received only 70 Ncm torque without preload stress as screws were untightened immediately. Cont group (n = 40) remained in situ. Animals were euthanized at 4, 6, 8, and 10 weeks after torque application. Micro-CT images were then taken, and undecalcified ground sections were stained with toluidine blue. RESULTS: Cross-sections of cortical bone showed remodeling activities adjacent to the implant in all groups. While bone marrow spaces appearance was relatively small in Cont and TT groups, RT and HT groups showed large bone marrow spaces and extensive remodeling activity. Bone-to-implant contact was significantly less in RT and HT groups compared with Cont and TT groups at different time points (p Ë‚ .05). Furthermore, RT and HT groups showed significantly less bone volume and area (p Ë‚ .05). CONCLUSION: Results suggested that preload stress without any occlusal loading might negatively affect peri-implant bone stability and initiate bone remodeling. This could alter bone mechanical properties, subsequently influencing long-term implant success.

Biological reaction control using topography regulation of nanostructured titanium.

The micro- and nanosize surface topography of dental implants has been shown to affect the growth of surrounding cells. In this study, standardized and controlled periodic nanopatterns were fabricated with nanosized surface roughness on titanium substrates, and their influence on bone marrow stromal cells investigated. Cell proliferation assays revealed that the bare substrate with a 1.7 nm surface roughness has lower hydrophilicity but higher proliferation ability than that with a 0.6 nm surface roughness. Further, with the latter substrate, directional cell growth was observed for line and groove patterns with a width of 100 nm and a height of 50 or 100 nm, but not for those with a height of 10 or 25 nm. With the smooth substrate, time-lapse microscopic analyses showed that more than 80% of the bone marrow cells on the line and groove pattern with a height of 100 nm grew and divided along the lines. As the nanosized grain structure controls the cell proliferation rate and the nanosized line and groove structure (50-100 nm) controls cell migration, division, and growth orientation, these standardized nanosized titanium structures can be used to elucidate the mechanisms by which surface topography regulates tissue responses to biomaterials.

Cyclophilin B control of lysine post-translational modifications of skin type I collagen.

Covalent intermolecular cross-linking of collagen is essential for tissue stability. Recent studies have demonstrated that cyclophilin B (CypB), an endoplasmic reticulum (ER)-resident peptidyl-prolyl cis-trans isomerase, modulates lysine (Lys) hydroxylation of type I collagen impacting cross-linking chemistry. However, the extent of modulation, the molecular mechanism and the functional outcome in tissues are not well understood. Here, we report that, in CypB null (KO) mouse skin, two unusual collagen cross-links lacking Lys hydroxylation are formed while neither was detected in wild type (WT) or heterozygous (Het) mice. Mass spectrometric analysis of type I collagen showed that none of the telopeptidyl Lys was hydroxylated in KO or WT/Het mice. Hydroxylation of the helical cross-linking Lys residues was almost complete in WT/Het but was markedly diminished in KO. Lys hydroxylation at other sites was also lower in KO but to a lesser extent. A key glycosylation site, α1(I) Lys-87, was underglycosylated while other sites were mostly overglycosylated in KO. Despite these findings, lysyl hydroxylases and glycosyltransferase 25 domain 1 levels were significantly higher in KO than WT/Het. However, the components of ER chaperone complex that positively or negatively regulates lysyl hydroxylase activities were severely reduced or slightly increased, respectively, in KO. The atomic force microscopy-based nanoindentation modulus were significantly lower in KO skin than WT. These data demonstrate that CypB deficiency profoundly affects Lys post-translational modifications of collagen likely by modulating LH chaperone complexes. Together, our study underscores the critical role of CypB in Lys modifications of collagen, cross-linking and mechanical properties of skin.

Clinical guidelines for dental cone-beam computed tomography.

Dental cone-beam computed tomography (CBCT) received regulatory approval in Japan in 2000 and has been widely used since being approved for coverage by the National Health Insurance system in 2012. This imaging technique allows dental practitioners to observe and diagnose lesions in the dental hard tissue in three dimensions (3D). When performing routine radiography, the examination must be justified, and optimal protection should be provided according to the ALARA (as low as reasonably achievable) principles laid down by the International Commission on Radiological Protection. Dental CBCT should be performed in such a way that the radiation exposure is minimized and the benefits to the patient are maximized. There is a growing demand for widespread access to cutting-edge health care through Japan's universal health insurance system. However, at the same time, people want our limited human, material, and financial resources to be used efficiently while providing safe health care at the least possible cost to society. Japan's aging population is expected to reach a peak in 2025, when most of the baby boomer generation will be aged 75 years or older. Comprehensive health care networks are needed to overcome these challenges. Against this background, we hope that this text will contribute to the nation's oral health by encouraging efficient use of dental CBCT.

Insulin-like growth factor binding Protein-3 suppresses osteoblast differentiation via bone morphogenetic protein-2.

Bone augmentation therapy is used in dental implantation. While techniques to induce bone formation are generally successful, the maintenance of bone mass is more difficult. Therefore, it is important to understand the mechanisms that regulate this process. Insulin-like growth factor-1 (IGF-1) is one of the most abundant growth factors that regulate bone mass, promote osteoblast differentiation, and accelerate bone formation. The activity of IGF-1 is regulated by IGF-binding proteins (IGFBPs). IGFBP-3 forms a ternary complex with IGF-1, extending its half-life in the circulating system. Therefore, IGFBP-3 acts as a stabilizer and transporter of IGF-1. Recent studies reported new IGF-1-independent functions of IGFBP-3 related with bone metabolism. In this study, we investigated the function of IGFBP-3 in osteoblast differentiation. Our results showed that IGFBP-3 decreases the expression of osteoblast differentiation markers, whose expression is enhanced by bone morphogenetic protein-2 (BMP-2). IGFBP-3 also reduced BMP-2 effect on ALP activity and mineral nodule formation. In addition, IGFBP-3 suppresses the activity of the Smad Binding Element (SBE) reporter, induced by BMP-2 signaling. These results suggest that IGFBP-3 inhibits osteoblast differentiation through the BMP-2 signal pathway, and that IGFBP-3 might play a role in bone mass maintenance in an IGF-1-dependent and -independent manner.

Extracellular matrix with defective collagen cross-linking affects the differentiation of bone cells.

Fibrillar type I collagen, the predominant organic component in bone, is stabilized by lysyl oxidase (LOX)-initiated covalent intermolecular cross-linking, an important determinant of bone quality. However, the impact of collagen cross-linking on the activity of bone cells and subsequent tissue remodeling is not well understood. In this study, we investigated the effect of collagen cross-linking on bone cellular activities employing a loss-of-function approach, using a potent LOX inhibitor, ß-aminopropionitrile (BAPN). Osteoblastic cells (MC3T3-E1) were cultured for 2 weeks in the presence of 0-2 mM BAPN to obtain low cross-linked collagen matrices. The addition of BAPN to the cultures diminished collagen cross-links in a dose-dependent manner and, at 1 mM level, none of the major cross-links were detected without affecting collagen production. After the removal of cellular components from these cultures, MC3T3-E1, osteoclasts (RAW264.7), or mouse primary bone marrow-derived stromal cells (BMSCs) were seeded. MC3T3-E1 cells grown on low cross-link matrices showed increased alkaline phosphatase (ALP) activity. The number of multinucleate tartrate-resistant acid phosphatase (TRAP)-positive cells increased in RAW264.7 cells. Initial adhesion, proliferation, and ALP activity of BMSCs also increased. In the animal experiments, 4-week-old C57BL/6 mice were fed with BAPN-containing diet for 8 weeks. At this point, biochemical analysis of bone demonstrated that collagen cross-links decreased without affecting collagen content. Then, the diet was changed to a control diet to minimize the direct effect of BAPN. At 2 and 4 weeks after the change, histological samples were prepared. Histological examination of femur samples at 4 weeks showed a significant increase in the number of bone surface osteoblasts, while the bone volume and surface osteoclast numbers were not significantly affected. These results clearly demonstrated that the extent of collagen cross-linking of bone matrix affected the differentiation of bone cells, underscoring the importance of collagen cross-linking in the regulation of cell behaviors and tissue remodeling in bone. Characterization of collagen cross-linking in bone may be beneficial to obtain insight into not only bone mechanical property, but also bone cellular activities.

Cyclophilin B Deficiency Causes Abnormal Dentin Collagen Matrix.

Cyclophilin B (CypB) is an endoplasmic reticulum-resident protein that regulates collagen folding, and also contributes to prolyl 3-hydroxylation (P3H) and lysine (Lys) hydroxylation of collagen. In this study, we characterized dentin type I collagen in CypB null (KO) mice, a model of recessive osteogenesis imperfecta type IX, and compared to those of wild-type (WT) and heterozygous (Het) mice. Mass spectrometric analysis demonstrated that the extent of P3H in KO collagen was significantly diminished compared to WT/Het. Lys hydroxylation in KO was significantly diminished at the helical cross-linking sites, α1/α2(I) Lys-87 and α1(I) Lys-930, leading to a significant increase in the under-hydroxylated cross-links and a decrease in fully hydroxylated cross-links. The extent of glycosylation of hydroxylysine residues was, except α1(I) Lys-87, generally higher in KO than WT/Het. Some of these molecular phenotypes were distinct from other KO tissues reported previously, indicating the dentin-specific control mechanism through CypB. Histological analysis revealed that the width of predentin was greater and irregular, and collagen fibrils were sparse and significantly smaller in KO than WT/Het. These results indicate a critical role of CypB in dentin matrix formation, suggesting a possible association between recessive osteogenesis imperfecta and dentin defects that have not been clinically detected.

Cyclophilin-B Modulates Collagen Cross-linking by Differentially Affecting Lysine Hydroxylation in the Helical and Telopeptidyl Domains of Tendon Type I Collagen.

Covalent intermolecular cross-linking provides collagen fibrils with stability. The cross-linking chemistry is tissue-specific and determined primarily by the state of lysine hydroxylation at specific sites. A recent study on cyclophilin B (CypB) null mice, a model of recessive osteogenesis imperfecta, demonstrated that lysine hydroxylation at the helical cross-linking site of bone type I collagen was diminished in these animals (Cabral, W. A., Perdivara, I., Weis, M., Terajima, M., Blissett, A. R., Chang, W., Perosky, J. E., Makareeva, E. N., Mertz, E. L., Leikin, S., Tomer, K. B., Kozloff, K. M., Eyre, D. R., Yamauchi, M., and Marini, J. C. (2014) PLoS Genet 10, e1004465). However, the extent of decrease appears to be tissue- and molecular site-specific, the mechanism of which is unknown. Here we report that although CypB deficiency resulted in lower lysine hydroxylation in the helical cross-linking sites, it was increased in the telopeptide cross-linking sites in tendon type I collagen. This resulted in a decrease in the lysine aldehyde-derived cross-links but generation of hydroxylysine aldehyde-derived cross-links. The latter were absent from the wild type and heterozygous mice. Glycosylation of hydroxylysine residues was moderately increased in the CypB null tendon. We found that CypB interacted with all lysyl hydroxylase isoforms (isoforms 1-3) and a putative lysyl hydroxylase-2 chaperone, 65-kDa FK506-binding protein. Tendon collagen in CypB null mice showed severe size and organizational abnormalities. The data indicate that CypB modulates collagen cross-linking by differentially affecting lysine hydroxylation in a site-specific manner, possibly via its interaction with lysyl hydroxylases and associated molecules. This study underscores the critical importance of collagen post-translational modifications in connective tissue formation.

Correlation Between Stress Distributions and Biological Reactions in Bone Surrounding Implants That Support Cantilevers in Supraocclusal Contact in Rats.

PURPOSE: To investigate the relationship between stress distributions and peri-implant bone reactions around maxillary implants that support cantilevers in supraocclusal contact. MATERIALS AND METHODS: After molar extraction, 16 Wistar rats received a titanium implant unilaterally. After healing, 8 rats (control group) were killed and the others received implant-supported cantilever superstructures in supraocclusion (loaded group). After 5 days, they were killed. The maxillae of all rats were scanned by microcomputed tomography (µ-CT). Based on the µ-CT images, bone volumes were measured. For the loaded group, 3D finite element models were created and analyzed under 20-N vertical and 5-N lateral forces, successively. After µ-CT scanning, sections were prepared and observed histologically. RESULTS: When compared with the controls, the bone volume tended to decrease in the loaded group, but the difference was not statistically significant. On average, marginal bone resorption and stresses tended to be higher in 2 rats that occluded on the cantilever arm than in the others, which occluded right on the implant, nevertheless, calculated stress did not surpass the maximum elastic stress (yielding strength) of rat bone. However, at the implant-bone interface of these samples, partial bone resorption surrounded by signs of active resorption was histologically found. CONCLUSION: These findings suggest that in this occlusally loaded rat model, the stress distributions correlated to some extent with bone volume and morphological changes observed on µ-CT images and histological sections.

RhoA-Mediated Functions in C3H10T1/2 Osteoprogenitors Are Substrate Topography Dependent.

Surface topography broadly influences cellular responses. Adherent cell activities are regulated, in part, by RhoA, a member of the Rho-family of GTPases. In this study, we evaluated the influence of surface topography on RhoA activity and associated cellular functions. The murine mesenchymal stem cell line C3H10T1/2 cells (osteoprogenitor cells) were cultured on titanium substrates with smooth topography (S), microtopography (M), and nanotopography (N) to evaluate the effect of surface topography on RhoA-mediated functions (cell spreading, adhesion, migration, and osteogenic differentiation). The influence of RhoA activity in the context of surface topography was also elucidated using RhoA pharmacologic inhibitor. Following adhesion, M and N adherent cells developed multiple projections, while S adherent cells had flattened and widespread morphology. RhoA inhibitor induced remarkable longer and thinner cytoplasmic projections on all surfaces. Cell adhesion and osteogenic differentiation was topography dependent with S < M and N surfaces. RhoA inhibition increased adhesion on S and M surfaces, but not N surfaces. Cell migration in a wound healing assay was greater on S versus M versus N surfaces and RhoA inhibitor increased S adherent cell migration, but not N adherent cell migration. RhoA inhibitor enhanced osteogenic differentiation in S adherent cells, but not M or N adherent cells. RhoA activity was surface topography roughness dependent (S < M, N). RhoA activity and -mediated functions are influenced by surface topography. Smooth surface adherent cells appear highly sensitive to RhoA function, while nano-scale topography adherent cell may utilize alternative cellular signaling pathway(s) to influence adherent cellular functions regardless of RhoA activity.

Observation of the bone surrounding an overloaded implant in a novel rat model.

PURPOSE: Although the recent success rates of dental implants are quite high, it is still true that loading during the bone healing period may be more likely to discourage osseointegration and that occlusal overloading might result in implant failure, even after osseointegration is established. The purposes of this study were to establish an appropriate experimental animal model and to histologically assess degenerative changes in established osseointegration under early and excessive occlusal loading. MATERIALS AND METHODS: Forty rats were divided into control and experimental groups. The maxillary first and second molars on both sides were extracted, and machined-surface titanium implants were placed. In the experimental group, 2 or 4 weeks after implant placement, abutments that were designed to overload the implants were attached. Control group implants did not receive abutments and remained in situ 2 or 4 weeks. Sections were prepared and observed histologically. RESULTS: Attrition of occluding opposite teeth and shiny spots on the abutments indicated that this model was useful for histologic investigation of the remodeling and bone changes around implants. Specimens showed remarkable bone loss and deterioration of osseointegration when overloading began at 2 weeks. Overloading applied after 4 weeks of healing induced active bone resorption in remote areas of the implants after 15 days of occlusion, while bone resorption at the interface was limited. CONCLUSION: The authors successfully established an implant occlusion model using rats. This model revealed degenerative changes in osseointegration and/or in the bone around implants upon excessive occlusal loading. These results emphasize the risks associated with immediate loading and overloading. This is the first study to reveal the possibility of bone loss around overloaded implants in the absence of infection using a small animal model.