S2 Peptide-Conjugated SARS-CoV-2 Virus-like Particles Provide Broad Protection against SARS-CoV-2 Variants of Concern.

Approved COVID-19 vaccines primarily induce neutralizing antibodies targeting the receptor-binding domain (RBD) of the SARS-CoV-2 spike (S) protein. However, the emergence of variants of concern with RBD mutations poses challenges to vaccine efficacy. This study aimed to design a next-generation vaccine that provides broader protection against diverse coronaviruses, focusing on glycan-free S2 peptides as vaccine candidates to overcome the low immunogenicity of the S2 domain due to the N-linked glycans on the S antigen stalk, which can mask S2 antibody responses. Glycan-free S2 peptides were synthesized and attached to SARS-CoV-2 virus-like particles (VLPs) lacking the S antigen. Humoral and cellular immune responses were analyzed after the second booster immunization in BALB/c mice. Enzyme-linked immunosorbent assay revealed the reactivity of sera against SARS-CoV-2 variants, and pseudovirus neutralization assay confirmed neutralizing activities. Among the S2 peptide-conjugated VLPs, the S2.3 (N1135-K1157) and S2.5 (A1174-L1193) peptide-VLP conjugates effectively induced S2-specific serum immunoglobulins. These antisera showed high reactivity against SARS-CoV-2 variant S proteins and effectively inhibited pseudoviral infections. S2 peptide-conjugated VLPs activated SARS-CoV-2 VLP-specific T-cells. The SARS-CoV-2 vaccine incorporating conserved S2 peptides and CoV-2 VLPs shows promise as a universal vaccine capable of generating neutralizing antibodies and T-cell responses against SARS-CoV-2 variants.

Runx2 heterozygosity alters homeostasis of the periodontal complex.

BACKGROUND AND OBJECTIVE: Haploinsufficiency of Runx2 (Runx2+/- ) causes dental anomalies. However, little is known about the involvement of Runx2 in the maintenance of dentin, cementum, and the periodontal ligament (PDL) during adulthood. This study aimed to observe the effects of Runx2+/- on homeostasis of the periodontal complex. MATERIALS AND METHODS: A total of 14 three-month-old Runx2+/- mice and their wild-type littermates were examined using micro-computed tomography, histology, and immunohistochemistry. Phenotypic alterations in the dentin, cementum, and PDL were characterized and quantified. RESULTS: Haploinsufficiency of Runx2 caused cellular changes in the PDL space including reduction of cell proliferation and apoptosis, and irregular attachment of the collagen fibers in the PDL space into the cementum. Absence of continuous thickness of cementum was also observed in Runx2+/- mice. CONCLUSION: Runx2 is critical for cementum integrity and attachment of periodontal fibers. Because of its importance to cementum homeostasis, Runx2 is essential for homeostasis of periodontal complex.

Novel S2 subunit-specific antibody with broad neutralizing activity against SARS-CoV-2 variants of concern.

Introduction: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), had a major impact on both the global health and economy. Numerous virus-neutralizing antibodies were developed against the S1 subunit of SARS-CoV-2 spike (S) protein to block viral binding to host cells and were authorized for control of the COVID-19 pandemic. However, frequent mutations in the S1 subunit of SARS-CoV-2 enabled the emergence of immune evasive variants. To address these challenges, broadly neutralizing antibodies targeting the relatively conserved S2 subunit and its epitopes have been investigated as antibody therapeutics and universal vaccines. Methods: We initiated this study by immunizing BALB/c mice with ß-propiolactone-inactivated SARS-CoV-2 (IAV) to generate B-cell hybridomas. These hybridomas were subsequently screened using HEK293T cells expressing the S2-ECD domain. Hybridomas that produced anti-S2 antibodies were selected, and we conducted a comprehensive evaluation of the potential of these anti-S2 antibodies as antiviral agents and versatile tools for research and diagnostics. Results: In this study, we present a novel S2-specific antibody, 4A5, isolated from BALB/c mice immunized with inactivated SARS-CoV-2. 4A5 exhibited specific affinity to SARS-CoV-2 S2 subunits compared with those of other ß-CoVs. 4A5 bound to epitope segment F1109-V1133 between the heptad-repeat1 (HR1) and the stem-helix (SH) region. The 4A5 epitope is highly conserved in SARS-CoV-2 variants, with a significant conformational feature in both pre- and postfusion S proteins. Notably, 4A5 exhibited broad neutralizing activity against variants and triggered Fc-enhanced antibody-dependent cellular phagocytosis. Discussion: These findings offer a promising avenue for novel antibody therapeutics and insights for next-generation vaccine design. The identification of 4A5, with its unique binding properties and broad neutralizing capacity, offers a potential solution to the challenge posed by SARS-CoV-2 variants and highlights the importance of targeting the conserved S2 subunit in combating the COVID-19.

Development of nucleocapsid-specific monoclonal antibodies for SARS-CoV-2 and their ELISA diagnostics on an automatic microfluidic device.

Coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has threatened public health globally, and the emergence of viral variants has exacerbated an already precarious situation. To prevent further spread of the virus and determine government action required for virus control, accurate and rapid immunoassays for SARS-CoV-2 diagnosis are urgently needed. In this study, we generated monoclonal antibodies (mAbs) against the SARS-CoV-2 nucleocapsid protein (NP), compared their reactivity using an enzyme-linked immunosorbent assay (ELISA), and selected four mAbs designated 1G6, 3E10, 3F10, and 5B6 which have higher reactivity to NP and viral lysates of SARS-CoV-2 than other mAbs. Using an epitope mapping assay, we identified that 1G6 detected the C-terminal domain of SARS-CoV-2 NP (residues 248-364), while 3E10 and 3F10 bound to the N-terminal domain (residues 47-174) and 3F10 detected the N-arm region (residues 1-46) of SARS-CoV-2 NP. Based on the epitope study and sandwich ELISA, we selected the 1G6 and 3E10 Abs as an optimal Ab pair and applied them for a microfluidics-based point-of-care (POC) ELISA assay to detect the NPs of SARS-CoV-2 and its variants. The integrated and automatic microfluidic system could operate the serial injection of the sample, the washing solution, the HRP-conjugate antibody, and the TMB substrate solution simply by controlling air purge via a single syringe. The proposed Ab pair-equipped microsystem effectively detected the NPs of SARS-CoV-2 variants as well as in clinical samples. Collectively, our proposed platform provides an advanced protein-based diagnostic tool for detecting SARS-CoV-2.

Serum BRD2 autoantibody in hepatocellular carcinoma and its detection using mimotope peptide­conjugated BSA.

Tumor­associated (TA) autoantibodies are considered to be promising biomarkers for the early detection of cancer, prior to the development of clinical symptoms. In the present study, a novel TA autoantibody was detected, which may prove to be useful as a diagnostic marker of human HCC using an HBx­transgenic (HBx­tg) hepatocellular carcinoma (HCC) mouse model. Its target antigen was identified as the bromodomain­containing protein 2 (BRD2), a transcriptional regulator that plays a pivotal role in the transcriptional control of diverse genes. BRD2 was upregulated in HCC tissues of the H­ras12V­tg mouse and human subjects, as demonstrated using western blotting or immunohistochemical analysis, with the BRD2 autoantibody. In addition, the truncated BRD2 reactive to the BRD2 autoantibody was detected in tumor cell­derived exosomes, which possibly activated TA immune responses and the generation of autoantibodies. For the detection of the serum BRD2 autoantibody, epitope mimicries of autoantigenic BRD2 were screened from a random cyclic peptide CX7C library with the BRD2 autoantibody. A mimotope with the sequence of CTSVFLPHC, which was cyclized by one pair of cysteine residues, exhibited high affinity to the BRD2 autoantibody and competitively inhibited the binding of the autoantibody to the cellular BRD2 antigen. The use of this cyclic peptide as a capture antigen in human serum enzyme­linked immunosorbent assay allowed the distinction of patients with HCC from healthy subjects with 64.41% sensitivity and 82.42% specificity (area under the ROC curve, 0.7761), which is superior to serum alpha­fetoprotein (AFP; 35.83% sensitivity; 100% specificity; area under the ROC curve, 0.5337) for the diagnosis of HCC. In addition, the detection of the BRD2 autoantibody combined with other autoantibody biomarkers or AFP has increased the accuracy of HCC diagnosis, suggesting that the combinational detection of cancer biomarkers, including the BRD2 autoantibody, is a promising assay for HCC diagnosis.

Influence of the Depth of the Convolutional Neural Networks on an Artificial Intelligence Model for Diagnosis of Orthognathic Surgery.

The aim of this study was to investigate the relationship between image patterns in cephalometric radiographs and the diagnosis of orthognathic surgery and propose a method to improve the accuracy of predictive models according to the depth of the neural networks. The study included 640 and 320 patients requiring non-surgical and surgical orthodontic treatments, respectively. The data of 150 patients were exclusively classified as a test set. The data of the remaining 810 patients were split into five groups and a five-fold cross-validation was performed. The convolutional neural network models used were ResNet-18, 34, 50, and 101. The number in the model name represents the difference in the depth of the blocks that constitute the model. The accuracy, sensitivity, and specificity of each model were estimated and compared. The average success rate in the test set for the ResNet-18, 34, 50, and 101 was 93.80%, 93.60%, 91.13%, and 91.33%, respectively. In screening, ResNet-18 had the best performance with an area under the curve of 0.979, followed by ResNets-34, 50, and 101 at 0.974, 0.945, and 0.944, respectively. This study suggests the required characteristics of the structure of an artificial intelligence model for decision-making based on medical images.

Comparative Analysis of the Differences in Dentofacial Morphology According to the Tongue and Lip Pressure.

The aim of this study was to evaluate the effects of the tongue and lip pressure on dentofacial morphology. The subjects comprised 194 patients with malocclusion. Anterior and posterior tongue elevation and lip pressures were evaluated using the Iowa Oral Performance Instrument (IOPI) device. The lateral cephalograms of each subject were traced and digitized to perform the analysis. Statistical analysis was used to investigate the relationship between perioral muscle force and the cephalometric variables. Anterior and posterior tongue pressure was both higher in males than in females. No sex difference in lip pressure was observed. The group with a low posterior tongue pressure showed a short ramus height, short posterior facial height, and clockwise-rotated mandible. On the other hand, lip pressure had a significant influence on maxillary incisor angulation. Skeletal pattern was not found to be significantly related with lip pressure. The anterior tongue pressure appeared as a mixed pattern of the two results. Tongue pressure was related to skeletal measurements, such as short posterior facial height, and lip pressure was related to the angulation of the anterior teeth. This study suggests that there may be differences in dentofacial morphology according to the differences in perioral muscle force.

Cyclic Peptide Mimotopes for the Detection of Serum Anti-ATIC Autoantibody Biomarker in Hepato-Cellular Carcinoma.

Tumor-associated (TA) autoantibodies have been identified at the early tumor stage before developing clinical symptoms, which holds hope for early cancer diagnosis. We identified a TA autoantibody from HBx-transgenic (HBx-tg) hepatocellular carcinoma (HCC) model mouse, characterized its target antigen, and examined its relationship to human HCC. The mimotopes corresponding to the antigenic epitope of TA autoantibody were screened from a random cyclic peptide library and used for the detection of serum TA autoantibody. The target antigen of the TA autoantibody was identified as an oncogenic bi-functional purine biosynthesis protein, ATIC. It was upregulated in liver cancer tissues of HBx-tg mouse as well as human HCC tissues. Over-expressed ATIC was also secreted extracellularly via the cancer-derived exosomes, which might cause auto-immune responses. The cyclic peptide mimotope with a high affinity to anti-ATIC autoantibody, CLPSWFHRC, distinguishes between serum samples from HCC patients and healthy subjects with 70.83% sensitivity, 90.68% specificity (AUC = 0.87). However, the recombinant human ATIC protein showed a low affinity to anti-ATIC autoantibody, which may be incompatible as a capture antigen for serum TA autoantibody. This study indicates that anti-ATIC autoantibody can be a potential HCC-associated serum biomarker and suggests that autoantibody biomarker's efficiency can be improved by using antigenic mimicry to native antigens present in vivo.

Microbial changes in biofilms on composite resins with different surface roughness: An in vitro study with a multispecies biofilm model.

STATEMENT OF PROBLEM: The single-species biofilm method cannot represent the interaction and complex functions of microorganisms associated with oral biofilms. PURPOSE: The purpose of this in vitro study was to investigate microbial changes in biofilms on composite resins of varying surface roughness by using a multispecies biofilm model with early-colonizing streptococci, middle colonizer, and late-colonizing gram-negative anaerobes. MATERIAL AND METHODS: Composite resin disks were prepared with different roughness: SR180, SR400, SR1500, and SRGlass roughened with 180-, 400-, and 1500-grit silicon carbide paper and glass (control surface without surface roughening). Surface roughness was analyzed by confocal laser scanning and scanning electron microscopy. After multispecies biofilms had been grown on the composite resin surfaces, the adhesion of Streptococcus mutans, Streptococcus sobrinus, Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, and of total bacteria was determined after 1 (T1) and 4 (T2) days. Differences in surface roughness among the 4 groups were tested with 1-way ANOVA. Multifactorial analysis of variance was used to determine the time-related differences in the bacterial composition with respect to surface roughness (α=.05). RESULTS: The order of SR, from highest to lowest, was SR180 (1.45 ±0.11 µm), SR400 (0.62 ±0.05 µm), SR1500 (0.35 ±0.02 µm), and SRGlass (0.15 ±0.01 µm) (SR180>SR400>SR1500>SRGlass, P<.001). Increased surface roughness was not proportional to bacterial adhesion. Significant differences in the adhesion of total bacteria was only found between SRGlass and SR180 (SR180>SRGlass, P=.029). The adhesion of S. mutans and S. sobrinus to SR180 and SR400 was higher than that to SRGlass (SR180=SR400>SRGlass; S. mutans, P=.003; S. sobrinus, P=.002). However, the adhesion of A. actinomycetemcomitans and P. gingivalis to composite resin was not significantly influenced by surface roughness. Adhesion of total bacteria, S. mutans, and S. sobrinus increased from T1 to T2 (T1T2; A. actinomycetemcomitans, P<.001; P. gingivalis, P=.013). CONCLUSIONS: Decreased adhesion of cariogenic streptococci and total bacteria was observed at surface roughness values of around 0.15 µm. Periodic finishing of surface roughness should be considered to minimize the adhesion of cariogenic streptococci to composite resin surfaces.

Artificial Intelligent Model With Neural Network Machine Learning for the Diagnosis of Orthognathic Surgery.

Diagnosis and treatment planning are the most important steps in the orthognathic surgery for the successful treatment. The purpose of this study was to develop a new artificial intelligent model for surgery/non-surgery decision and extraction determination, and to evaluate the performance of this model. The sample used in this study consisted of 316 patients in total. Of the total sample, 160 were planned with surgical treatment and 156 were planned with non-surgical treatment. The input values of artificial neural network were obtained from 12 measurement values of the lateral cephalogram and 6 additional indexes. The artificial intelligent model of machine learning consisted of 2-layer neural network with one hidden layer. The learning was carried out in 3 stages, and 4 best performing models were adopted. Using these models, decision-making success rates of surgery/non-surgery, surgery type, and extraction/non-extraction were calculated. The final diagnosis success rate was calculated by comparing the actual diagnosis with the diagnosis obtained by the artificial intelligent model. The success rate of the model showed 96% for the diagnosis of surgery/non-surgery decision, and showed 91% for the detailed diagnosis of surgery type and extraction decision. This study suggests the artificial intelligent model using neural network machine learning could be applied for the diagnosis of orthognathic surgery cases.

Identification of anti-SF3B1 autoantibody as a diagnostic marker in patients with hepatocellular carcinoma.

BACKGROUND: Tumor-associated (TA) autoantibodies, which are generated by the immune system upon the recognition of abnormal TA antigens, are promising biomarkers for the early detection of tumors. In order to detect autoantibody biomarkers effectively, antibody-specific epitopes in the diagnostic test should maintain the specific conformations that are as close as possible to those presenting in the body. However, when using patients' serum as a source of TA autoantibodies the characterization of the autoantibody-specific epitope is not easy due to the limited amount of patient-derived serum. METHODS: To overcome these limits, we constructed a B cell hybridoma pool derived from a hepatocellular carcinoma (HCC) model HBx-transgenic mouse and characterized autoantibodies derived from them as tumor biomarkers. Their target antigens were identified by mass spectrometry and the correlations with HCC were examined. With the assumption that TA autoantibodies generated in the tumor mouse model are induced in human cancer patients, the enzyme-linked immunosorbent assays (ELISA) based on the characteristics of mouse TA autoantibodies were developed for the detection of autoantibody biomarkers in human serum. To mimic natural antigenic structures, the specific epitopes against autoantibodies were screened from the phage display cyclic random heptapeptide library, and the streptavidin antigens fused with the specific epitopes were used as coating antigens. RESULTS: In this study, one of HCC-associated autoantibodies derived from HBx-transgenic mouse, XC24, was characterized. Its target antigen was identified as splicing factor 3b subunit 1 (SF3B1) and the high expression of SF3B1 was confirmed in HCC tissues. The specific peptide epitopes against XC24 were selected and, among them, XC24p11 cyclic peptide (-CDATPPRLC-) was used as an epitope of anti-SF3B1 autoantibody ELISA. With this epitope, we could effectively distinguish between serum samples from HCC patients (n = 102) and healthy subjects (n = 85) with 73.53% sensitivity and 91.76% specificity (AUC = 0.8731). Moreover, the simultaneous detection of anti-XC24p11 epitope autoantibody and AFP enhanced the efficiency of HCC diagnosis with 87.25% sensitivity and 90.59% specificity (AUC = 0.9081). CONCLUSIONS: ELISA using XC24p11 peptide epitope that reacts against anti-SF3B1 autoantibody can be used as a novel test to enhance the diagnostic efficiency of HCC.

Alveolar Molding Effect in Infants With Unilateral Cleft Lip and Palate: Comparison of Two- and Three-Dimensional Measurements.

The purpose of this study was to compare the results of two- (2D) and three-dimensional (3D) measurements for the alveolar molding effect in patients with unilateral cleft lip and palate. The sample consisted of 23 unilateral cleft lip and palate infants treated with nasoalveolar molding (NAM) appliance. Dental models were fabricated at initial visit (T0; mean age, 23.5 days after birth) and after alveolar molding therapy (T1; mean duration, 83 days). For 3D measurement, virtual models were constructed using a laser scanner and 3D software. For 2D measurement, 1:1 ratio photograph images of dental models were scanned by a scanner. After setting of common reference points and lines for 2D and 3D measurements, 7 linear and 5 angular variables were measured at the T0 and T1 stages, respectively. Wilcoxon signed rank test and Bland-Altman analysis were performed for statistical analysis. The alveolar molding effect of the maxilla following NAM treatment was inward bending of the anterior part of greater segment, forward growth of the lesser segment, and decrease in the cleft gap in the greater segment and lesser segment. Two angular variables showed difference in statistical interpretation of the change by NAM treatment between 2D and 3D measurements (ΔACG-BG-PG and ΔACL-BL-PL). However, Bland-Altman analysis did not exhibit significant difference in the amounts of change in these variables between the 2 measurements. These results suggest that the data from 2D measurement could be reliably used in conjunction with that from 3D measurement.

Induction of S100A4 in periodontal ligament cells enhances osteoclast formation.

BACKGROUND: An increase in the expression of S100A4 has been reported in various inflammatory diseases. However, little is known about the association between periodontal inflammation and S100A4 expression. The aims of this study were to investigate changes in S100A4 expression in human periodontal ligament (hPDL) cells in response to inflammatory stimuli and to describe a possible mechanism underlying the change. METHOD: Human PDL cells were treated with lipopolysaccharide (LPS) and the level of S100A4 was analyzed by real-time RT-PCR and Western blotting. LPS was added to co-cultures of hPDL and osteoclast progenitor cells under osteoclastogenic condition and the formation of osteoclasts was assessed. Alternatively, progenitor cells were directly treated with recombinant S100A4 for evaluation of osteoclastogenesis. The activity of nuclear factor kappaB (NFκB) was examined by Western blotting for phosphorylated forms of inhibitor kappaB (IκB) and p65. An NFκB inhibitor was added to the culture of hPDL cells with LPS and the level of S100A4 was measured by real-time RT-PCR. RESULTS: We found that LPS stimulation resulted in a significant increase of S100A4 expression in hPDL cells. S100A4 protein secretion from hPDL cells was also increased. The enhanced expression of S100A4 in hPDL cells under inflammatory conditions led to stimulation of the generation of osteoclasts. In addition, direct S100A4 treatment stimulated osteoclastogenesis. The underlying mechanism for the increased S100A4 expression in hPDL cells was activation of the NFκB signaling pathway. CONCLUSION: Our results suggest that bone destruction in periodontitis might be associated with increased S100A4 expression in hPDL cells.

Adhesion of mutans streptococci to self-ligating ceramic brackets: in vivo quantitative analysis with real-time polymerase chain reaction.

OBJECTIVE: To analyze in vivo mutans streptococci (MS) adhesion to self-ligating ceramic brackets [Clarity-SL (CSL) and Clippy-C (CC)] and the relationships between bacterial adhesion and oral hygiene indices. MATERIALS AND METHODS: Four central incisor brackets from the maxilla and mandible were collected from 40 patients (20 patients per each bracket type) at debonding immediately after plaque and gingival indices were measured. Adhesions of Streptococcus mutans, S. sobrinus, and total bacteria were quantitatively determined using real-time polymerase chain reaction after genomic DNA was extracted. Factorial analysis of variance was used to analyze bacterial adhesion to the brackets with respect to the bracket type and jaw position. Correlation coefficients were calculated to determine the relationships of bacterial adhesion to oral hygiene indices. RESULTS: Adhesion of total bacteria and S. mutans to CSL was higher than that to CC (P < 0.001). Adhesion of total bacteria to the mandibular brackets was higher than that to the maxillary ones (P < 0.001), while adhesion of S. mutans to the maxillary brackets were higher than that in the mandibular ones (P < 0.001). In particular, the proportion of S. mutans to total bacteria in CSL was higher than CC (P < 0.05) in the maxillary anterior teeth (P < 0.001). There were no significant differences in adhesion of S. sobrinus between the brackets and jaw positions. Interestingly, no significant relationships were found between bacterial adhesions and oral hygiene indices. LIMITATIONS: Complex bracket configurations may significantly influence bacterial adhesion to orthodontic brackets. Further in vivo study using bracket raw materials will help to define the relationships between bacteria adhesion and enamel demineralization. CONCLUSIONS: Because oral hygiene indices were not significantly correlated with adhesions of MS to self-ligating ceramic brackets, careful examinations around the brackets should be needed to prevent enamel demineralization, regardless of oral hygiene status.

Alveolar bone turnover and periodontal ligament width are controlled by Wnt.

BACKGROUND: The molecular signals responsible for maintaining homeostatic control over the periodontal ligament (PDL) are unknown. The purpose of this study is to investigate the role of Wnt signaling in this process using gain- and loss-of-function approaches. METHODS: The function of endogenous Wnt signal in the PDL was evaluated in Lrp5(ACT) mice in which a mutation in the low-density lipoprotein receptor-related protein 5 Wnt coreceptor causes constitutive activation of Wnt signaling, and in adenovirus Dkk1-treated mice in which overexpression of the Wnt inhibitor Dkk1 causes transient Wnt signal inhibition. PDL in both animal models was examined using histology and immunohistochemical analyses for osteopontin, runt-related transcription factor 2 (Runx2), fibromodulin, osterix, ki67, receptor activator of nuclear factor-κB ligand (RANKL), and alkaline phosphatase activity. RESULTS: Lrp5(ACT) mice exhibited a significant narrowing of the PDL space caused by an increase in osteogenic gene expression, a reduction in RANKL expression and osteoclast activity, and an increase in alveolar bone formation. Conversely, adenovirus Dkk1-treated mice showed decreased expression of osteogenic markers, coupled with an increase in osteoclast activity, which resulted in a slight increase in PDL width. CONCLUSION: The Wnt pathway is involved in the homeostatic control of the PDL, and conditions that elevate or repress Wnt signaling alter the expression of osteogenic genes within the PDL space, which in turn affects its overall width.

Influence of temporomandibular joint disc displacement on craniocervical posture and hyoid bone position.

INTRODUCTION: The purpose of this study was to evaluate craniocervical posture and hyoid bone position in orthodontic patients with temporomandibular joint (TMJ) disc displacement. METHODS: The subjects consisted of 170 female orthodontic patients who consented to bilateral magnetic resonance imaging of their TMJs. They were divided into 3 groups based on the results of magnetic resonance imaging of their TMJs: bilateral normal disc position, bilateral disc displacement with reduction, and bilateral disc displacement without reduction. Twenty-five variables from lateral cephalograms were analyzed with 1-way analysis of variance to investigate differences in craniocervical posture and hyoid bone position with respect to TMJ disc displacement status. Pearson correlation coefficients were calculated to analyze the relationships between craniofacial morphology and craniocervical posture or hyoid bone position. RESULTS: Subjects with TMJ disc displacement were more likely to have an extended craniocervical posture with Class II hyperdivergent patterns. The most significant differences were found between patients with bilateral normal disc position and bilateral disc displacement without reduction. However, hyoid bone position in relation to craniofacial references was not significantly different among the TMJ disc displacement groups, except for variables related to the mandible. Pearson correlation coefficients indicated that extended craniocervical posture was significantly correlated with backward positioning and clockwise rotation of the mandible. CONCLUSIONS: This suggests that craniocervical posture is significantly influenced by TMJ disc displacement, which may be associated with hyperdivergent skeletal patterns with a retrognathic mandible.

Downregulation of Wnt causes root resorption.

INTRODUCTION: There are multiple causes of external root resorption, but absent a disease state, it is most often observed when excessive physical force is used during orthodontic treatment. Even without mechanical stimulation, however, root resorption can still occur. The purpose of this study was to test whether Wnt signaling plays a role in pathologic root resorption, by conditionally deleting Wntless (Wls) from odontoblasts and osteoblasts and then evaluating the phenotypic effects on the maintenance of the root surface. METHODS: Ten (age, 1 month) and 20 (age, 3 months) OCN-Cre;Wls(fl/fl) mice and their wild-type littermates were evaluated using microcomputed tomography, histology, and immunohistochemistry. Phenotypic alterations in the alveolar bone, dentin, and cementum were characterized and quantified. RESULTS: In a genetic model of reduced Wnt signaling, we found that RANKL expression is upregulated, and osteoprotegerin expression is downregulated. This molecular disruption results in an increase in osteoclast activity, a decrease in osteoblast activity, and extensive, spontaneous root resorption. A genetic strain of mice in which Wnt signaling is elevated exhibits thicker cementum, whereas, even in the perinatal period, OCN-Cre;Wls(fl/fl) mice exhibit thinner cementum. CONCLUSIONS: Taken together, these data demonstrate that Wnts regulate cementum homeostasis, and that idiopathic cases of root resorption might have as their etiology a reduction in endogenous Wnt signaling.

The molecular and cellular effects of ageing on the periodontal ligament.

AIM: Many in vitro studies have investigated age-related biological changes in cells comprising the periodontium but the basic question of whether the periodontium can maintain its integrity with age remains unanswered. Thus, the aim of this study was to understand how, in the absence of disease, advancing age impacts the structure of the periodontium. MATERIALS AND METHODS: Of 4, 10, 25, and 50-week-old mice were examined using histology and immunohistochemical analyses for cell proliferation, cell turnover, collagen quantity and quality, osteogenic markers, bone turnover, and cytokine expression. RESULTS: The periodontal ligament (PDL) space shows a gradual decrease in width over the lifespan of the mice. Cell proliferation as well as the quantity and quality of collagen fibres decreased with age although cell density did not appear to be altered. Osteoprogenitor markers in the PDL maintained their expression with increasing age. Alkaline phosphatase (ALP) activity decreased, but osteoclast activity increased with age. CONCLUSIONS: Ageing is associated with a decline in the quality and quantity of collagen and an increase in bone resorption, all of which can diminish the function of the periodontium even in the absence of disease.

Dentofacial characteristics in orthodontic patients with centric relation-maximum intercuspation discrepancy.

INTRODUCTION: To investigate dentofacial characteristics of orthodontic patients with centric relation (CR)-maximum intercuspation (MI) discrepancy and to analyze changes in dentofacial characteristics between CR and MI positions in these patients using lateral cephalograms. MATERIALS AND METHODS: Adult female patients were classified into two groups: large CR-MI discrepancy (greater than 2.0 mm horizontal or vertical mandibular incisor movements during CR to MI change, n  =  20) and small CR-MI discrepancy (less than 1.0 mm horizontal and vertical mandibular incisor movements during CR to MI change, n  =  22). All subjects underwent temporomandibular joint (TMJ) magnetic resonance imaging prior to treatment. Gnathological stabilizing splints were used to find a reliable CR position in patients with large CR-MI discrepancy. Sixteen variables from lateral cephalograms were analyzed to identify differences in cephalometric variables between CR and MI positions in patients with large discrepancy. Differences in dentofacial cephalometric variables at MI positions between patients with large and small CR-MI discrepancies were also analyzed. RESULTS: Patients with large CR-MI discrepancy had backward positioning and rotation of the mandible at the MI position compared to the norm. In addition, the mandible moved more posteriorly and rotated more in a clockwise direction during MI to CR change. Interestingly, all patients with large CR-MI discrepancy had TMJ disk displacement. There were no significant differences in the cephalometric variables of the MI positions between patients with small and large CR-MI discrepancies. CONCLUSIONS: This study suggests that adult patients with backward positioning and rotation of the mandible should be carefully evaluated as a result of the potential CR-MI discrepancy.

Wnt signaling regulates pulp volume and dentin thickness.

Odontoblasts, cementoblasts, ameloblasts, and osteoblasts all form mineralized tissues in the craniofacial complex, and all these cell types exhibit active Wnt signaling during postnatal life. We set out to understand the functions of this Wnt signaling, by evaluating the phenotypes of mice in which the essential Wnt chaperone protein, Wntless was eliminated. The deletion of Wls was restricted to cells expressing Osteocalcin (OCN), which in addition to osteoblasts includes odontoblasts, cementoblasts, and ameloblasts. Dentin, cementum, enamel, and bone all formed in OCN-Cre;Wls(fl/fl) mice but their homeostasis was dramatically affected. The most notable feature was a significant increase in dentin volume and density. We attribute this gain in dentin volume to a Wnt-mediated misregulation of Runx2. Normally, Wnt signaling stimulates Runx2, which in turn inhibits dentin sialoprotein (DSP); this inhibition must be relieved for odontoblasts to differentiate. In OCN-Cre;Wls(fl/fl) mice, Wnt pathway activation is reduced and Runx2 levels decline. The Runx2-mediated repression of DSP is relieved and odontoblast differentiation is accordingly enhanced. This study demonstrates the importance of Wnt signaling in the homeostasis of mineralized tissues of the craniofacial complex.