Association between gender and self-assessment skills amongst Japanese dental students.

Background/purpose: Self-assessment in dental education is considered a fundamental skill for proficient oral healthcare providers. While previous studies looked at self-assessment in education, few have done so at Japanese institutions. This study aimed to assess potential associations between gender and self-assessment skills in Japanese dental students. Materials and methods: Dental students from Niigata University (n = 301) completed preclinical prosthodontic exercises and self-assessed their practical exam performance. Three calibrated faculty evaluated students' work using the same rubric. The difference between student self-assessment score and mean faculty grade was defined as the student-faculty (S-F) gap, indicating how accurately students evaluated their work. The gap scores were stratified by quartiles by faculty grade and by gender. Regression analysis was used to investigate potential associations. Results: Students mean S-F gap was 2.9 ± 10.5 % with a significant negative association to faculty grades (coefficient, -0.32; P < 0.001). There was no significant difference between male and female students for faculty grades, self-assessment scores, and S-F gaps. When stratified by quartile, mean S-F gaps of bottom quartile students (5.7 ± 11.9 %) were significantly higher than the rest of the class (1.9 ± 9.8 %; P = 0.01), while the mean S-F gaps of students in the top quartile (-0.8 ± 10.2 %) were significantly lower than the rest of the class (4.2 ± 10.3 %; P < 0.001). Conclusion: Niigata University students generally overestimated their performance. Students with higher faculty grades self-assessed themselves more accurately than students with lower faculty grades. Gender did not influence S-F gaps, despite dentistry in Japan shifting toward a female-dominated field.

Controlling redox state by edaravone at transplantation site enhances bone regeneration.

In cell-based bone augmentation, transplanted cell dysfunction and apoptosis can occur due to oxidative stress caused by the overproduction of reactive oxygen species (ROS). Edaravone (EDA) is a potent free radical scavenger with potential medical applications. This study aimed to investigate the effect of controlling oxidative stress on bone regeneration using EDA. Bone marrow-derived cells were collected from 4-week-old rats, and EDA effects on cell viability and osteogenic differentiation were evaluated. Collagen gels containing PKH26-prelabeled cells were implanted into the calvarial defects of 12-week-old rats, followed by daily subcutaneous injections of normal saline or 500 µM EDA for 4 d. Bone formation was examined using micro-computed tomography and histological staining. Immunofluorescence staining was performed for markers of oxidative stress, macrophages, osteogenesis, and angiogenesis. EDA suppressed ROS production and hydrogen peroxide-induced apoptosis, recovering cell viability and osteoblast differentiation. EDA treatment in vivo increased new bone formation. EDA induced the transition of the macrophage population toward the M2 phenotype. The EDA group also exhibited stronger immunofluorescence for vascular endothelial growth factor and CD31. In addition, more PKH26-positive and PKH26-osteocalcin-double-positive cells were observed in the EDA group, indicating that transplanted cell survival was prolonged, and they differentiated into bone-forming cells. This could be attributed to oxidative stress suppression at the transplantation site by EDA. Collectively, local administration using EDA facilitates bone regeneration by improving the local environment and angiogenesis, prolonging survival, and enhancing the osteogenic capabilities of transplanted cells.

Multiomics analysis of cultured mouse periodontal ligament cell-derived extracellular matrix.

A comprehensive understanding of the extracellular matrix (ECM) is essential for developing biomimetic ECM scaffolds for tissue regeneration. As the periodontal ligament cell (PDLC)-derived ECM has shown potential for periodontal tissue regeneration, it is vital to gain a deeper understanding of its comprehensive profile. Although the PDLC-derived ECM exhibits extracellular environment similar to that of periodontal ligament (PDL) tissue, details of its molecular composition are lacking. Thus, using a multiomics approach, we systematically analyzed cultured mouse PDLC-derived ECM and compared it to mouse PDL tissue as a reference. Proteomic analysis revealed that, compared to PDL tissue, the cultured PDLC-derived ECM had a lower proportion of fibrillar collagens with increased levels of glycoprotein, corresponding to an immature ECM status. The gene expression signature was maintained in cultured PDLCs and was similar to that in cells from PDL tissues, with additional characteristics representative of naturally occurring progenitor cells. A combination of proteomic and transcriptomic analyses revealed that the cultured mouse PDLC-derived ECM has multiple advantages in tissue regeneration, providing an extracellular environment that closely mimics the environment in the native PDL tissue. These findings provide valuable insights for understanding PDLC-derived ECM and should contribute to the development of biomimetic ECM scaffolds for reliable periodontal tissue regeneration.

Acellular Extrinsic Fiber Cementum Is Invariably Present in the Superficial Layer of Apical Cementum in Mouse Molar.

The cementum is a highly mineralized tissue that covers the tooth root. The regional differences among the types of cementum, especially in the extrinsic fibers that contribute to tooth support, remain controversial. Therefore, this study used second harmonic generation imaging in conjunction with automated collagen extraction and image analysis algorithms to facilitate the quantitative examination of the fiber characteristics and the changes occurring in these fibers over time. Acellular extrinsic fiber cementum (AEFC) was invariably observed in the superficial layer of the apical cementum in mouse molars, indicating that this region of the cementum plays a crucial role in supporting the tooth. The apical AEFC exhibited continuity and fiber characteristics comparable with the cervical AEFC, suggesting a common cellular origin for their formation. The cellular intrinsic fiber cementum present in the inner layer of the apical cementum showed consistent growth in the apical direction without layering. This study highlights the dynamic nature of the cementum in mouse molars and underscores the requirement for re-examining its structure and roles. The findings of the present study elucidate the morphophysiological features of cementum and have broader implications for the maintenance of periodontal tissue health.

Effect of Sparc knockout on the extracellular matrix of mouse periodontal ligament cells.

The periodontal ligament (PDL) is a critical component in maintaining tooth stability. It is composed of cells and an extracellular matrix (ECM), each with unique roles in tissue function and homeostasis. Secreted protein acidic and rich in cysteine (SPARC), a calcium-binding matricellular glycoprotein, plays a crucial role in regulating ECM assembly and turnover, alongside facilitating cellular-ECM interactions. In the present study, mass spectrometry-based proteomics was used to assess the impacts of Sparc-knockout (KO) on PDL-derived cells. Results demonstrated that Sparc-KO significantly reduces ECM production and alters its composition with increased levels of type I collagen. Despite this increase in Sparc-KO, type I collagen was not likely to be effectively integrated into the fibrils due to collagen cross-linking impairment. Furthermore, the pathway and process enrichment analyses suggested that SPARC plays a protective role against ECM degradation by antagonistically interacting with cell-surface collagen receptors. These findings provide detailed insights into the multifaceted role of SPARC in ECM organization, including its impact on ECM production, collagen regulation, and interactions with various cellular compartments. A better understanding of these complex mechanisms is crucial for comprehending the causes of periodontal disease and tissue regeneration, where precise control of ECM organization is necessary.

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.

Extracellular Matrix-Oriented Proteomic Analysis of Periodontal Ligament Under Mechanical Stress.

The periodontal ligament (PDL) is a specialized connective tissue that provides structural support to the tooth and is crucial for oral functions. The mechanical properties of the PDL are mainly derived from the tissue-specific composition and structural characteristics of the extracellular matrix (ECM). The ECM also plays key roles in determining cell fate in the cellular microenvironment thus crucial in the PDL tissue homeostasis. In the present study, we determined the comprehensive ECM profile of mouse molar PDL using laser microdissection and mass spectrometry-based proteomic analysis with ECM-oriented data curation. Additionally, we evaluated changes in the ECM proteome under mechanical loading using a mouse orthodontic tooth movement (OTM) model and analyzed potential regulatory networks using a bioinformatics approach. Proteomic changes were evaluated in reference to the novel second harmonic generation (SHG)-based fiber characterization. Our ECM-oriented proteomics approach succeeded in illustrating the comprehensive ECM profile of the mouse molar PDL. We revealed the presence of type II collagen in PDL, possibly associated with the load-bearing function upon occlusal force. Mechanical loading induced unique architectural changes in collagen fibers along with dynamic compositional changes in the matrisome profile, particularly involving ECM glycoproteins and matrisome-associated proteins. We identified several unique matrisome proteins which responded to the different modes of mechanical loading in PDL. Notably, the proportion of type VI collagen significantly increased at the mesial side, contributing to collagen fibrogenesis. On the other hand, type XII collagen increased at the PDL-cementum boundary of the distal side. Furthermore, a multifaceted bioinformatics approach illustrated the potential molecular cues, including PDGF signaling, that maintain ECM homeostasis under mechanical loading. Our findings provide fundamental insights into the molecular network underlying ECM homeostasis in PDL, which is vital for clinical diagnosis and development of biomimetic tissue-regeneration strategies.

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.

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.

Metallization by Sputtering to Improve the Bond Strength between Zirconia Ceramics and Resin Cements.

Zirconia has been used as a prosthesis material for over a decade because of its excellent mechanical properties and esthetics. The surface treatment for zirconia generally involves sandblasting and the application of primers for favorable bond strength between the surface and resin. However, sandblasting causes the microcracking and chipping of the zirconia surface. To overcome these challenges, the metallization of the zirconia surface was performed. Ti and Au were sputtered on yttria stabilized zirconia (YSZ) disks and heated to 800 °C for 15 min in air. These disks were bonded to stainless-steel rods using resin cement. Then, shear bond strength tests were performed using an Instron-type testing machine. The shear bond strength of the Ti sputtering group was significantly higher than that of the other groups. According to the results of X-ray photoelectron spectroscopy and electron probe microanalysis, the Ti-sputtered YSZ surface contained both sub-titanium oxide and titanium oxide before heating. Sub-titanium oxide was converted to titanium oxide by heating. These results suggest that metallization using Ti is effective for zirconia surface treatment to improve the shear bond strength between YSZ and resin cement. This metallization technique for YSZ has potential in clinical applications.

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.

In vivo cell proliferation analysis and cell-tracing reveal the global cellular dynamics of periodontal ligament cells under mechanical-loading.

Periodontal ligament (PDL) is a uniquely differentiated tissue that anchors the tooth to the alveolar bone socket and plays key roles in oral function. PDL cells can respond rapidly to mechanical stimuli, resulting in accelerated tissue remodeling. Cell proliferation is an initial event in tissue remodeling and participates in maintaining the cell supply; therefore, analyzing cell-proliferative activity might provide a comprehensive view of cellular dynamics at the tissue level. In this study, we investigated proliferating cells in mouse molar PDL during orthodontic tooth movement (OTM)-induced tissue remodeling. Our results demonstrated that the mechanical stimuli evoked a dynamic change in the proliferative-cell profile at the entire PDL. Additionally, cell-tracing analysis revealed that the proliferated cells underwent further division and subsequently contributed to tissue remodeling. Moreover, OTM-induced proliferating cells expressed various molecular markers that most likely arise from a wide range of cell types, indicating the lineage plasticity of PDL cells in vivo. Although further studies are required, these findings partially elucidated the global views of the cell trajectory in mouse molar PDL under mechanical-loading conditions, which is vital for understanding the cellular dynamics of the PDL and beneficial for dental treatment in humans.

Reducing bacterial counts around the abutment following professional mechanical plaque removal at the implant bridge: A randomized crossover comparison of removing or not removing the superstructure.

PURPOSE: The purpose of this study was to determine whether removing the superstructure of the implant bridge in cases of full-arch implant restorations for edentulous atrophic arches at the abutment level during professional mechanical plaque removal (PMPR) affects bacterial counts. METHODS: This crossover clinical trial included 20 patients who received screw-retained prostheses at the abutment level. Patients were randomly assigned to two groups and received PMPR with or without removal of the superstructure. After a three-month washout period, the type of treatment was reversed between the groups. Bacterial counts around the cylinder and abutment were measured and compared before and after PMPR. RESULTS: Bacterial numbers around the cylinder and abutment were significantly reduced after PMPR as compared with before PMPR regardless of whether the superstructure was removed (p <0.05). However the ratio of subjects with bacteria at 1.0 × 105 colony forming unit/ml (cfu/ml) or more after PMPR was significantly higher when the superstructure was not removed (p < 0.05). Among patients with bacterial counts of less than 10 × 105 cfu/ml, bacterial loads were reduced to less than 1.0 × 105 cfu/ml even when superstructures were not removed. Among patients with bacterial load of >10 × 105 cfu/ml, bacterial numbers were not reduced to <1.0 × 105 cfu/ml when PMPR was performed without removing the superstructure. CONCLUSIONS: Removal of the superstructure in cases of full-arch implant restorations for edentulous atrophic arches during PMPR reduces bacterial numbers around the implant bridge at the abutment level.

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.

A multi-factorial analysis of bone morphology and fracture strength of rat femur in response to ovariectomy.

BACKGROUND: Postmenopausal osteoporosis develops due to a deficiency of estrogen that causes a decrease in bone mass and changes in the macro- and micro-architectural structure of the bone, leading to the loss of mechanical strength and an increased risk of fracture. Although the assessment of bone mineral density (BMD) has been widely used as a gold standard for diagnostic screening of bone fracture risks, it accounts for only a part of the variation in bone fragility; thus, it is necessary to consider other determinants of bone strength. Therefore, we aimed to comprehensively evaluate the architectural changes of the bone that influence bone fracture strength, together with the different sensitivities of cortical and trabecular bone in response to ovariectomy (OVX). METHODS: Bone morphology parameters were separately analyzed both in cortical and in trabecular bones, at distal-metaphysis, and mid-diaphysis of OVX rat femurs. Three-point bending test was performed at mid-diaphysis of the femurs. Correlation of OVX-induced changes of morphological parameters with breaking force was analyzed using Pearson's correlation coefficient. RESULTS: OVX resulted in a decline in the bone volume of distal-metaphysis trabecular bone, but an increase in distal-metaphysis and mid-diaphysis cortical bone volume. Tissue mineral density (TMD) remained unchanged in both the trabecular and cortical bone of the distal metaphysis but decreased in cortical bone of the mid-diaphysis. The OVX significantly increased the breaking force at mid-diaphysis of the femurs. CONCLUSIONS: OVX decreased the trabecular bone volume of the distal-metaphysis and increased the cortical bone volume of the distal-metaphysis and mid-diaphysis. Despite the reduction in TMD and increased cortical porosity, bone fracture strength increased in the mid-diaphysis after OVX. These results indicate that analyzing a single factor, i.e., BMD, is not sufficient to predict the absolute fracture risk of the bone, as OVX-induced bone response vary, depending on the bone type and location. Our results strongly support the necessity of analyzing bone micro-architecture and site specificity to clarify the true etiology of osteoporosis in a clinical setting.

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.