Physical Properties and Antimicrobial Release Ability of Gentamicin-Loaded Apatite Cement/α-TCP Composites: An In Vitro Study.

Apatite cement (AC), which has excellent osteoconductive ability, and alpha-tricalcium phosphate (α-TCP), which can be used for bone replacement, are useful bone substitute materials. The objective of this study was to clarify the physical properties and antimicrobial release ability of antibiotic-loaded AC/α-TCP composites in vitro. Gentamicin-loaded, rapid setting AC/α-TCP composites were prepared in 2 mixing ratios (10:3 and 10:6). The cement paste of AC/α-TCP composites was prepared in a plastic mold and dried in a thermostatic chamber at 37 °C and 100% relative humidity for 24 h. A diametral tensile strength test, powder X-ray diffraction analysis, and gentamicin release test were performed. The diametral tensile strengths of the AC/α-TCP composites were significantly less than that of AC alone. Powder X-ray diffraction patterns exhibited the characteristic peaks of hydroxyapatite in the AC/α-TCP composites and gentamicin-loaded AC/α-TCP composites. The concentration of the released gentamicin was maintained above the minimum inhibitory concentration of Staphylococcus aureus until Day 30 in both the gentamicin-loaded AC/α-TCP composites (10:3 and 10:6). Our results suggest that a gentamicin-loaded AC/α-TCP composite has potential as a drug delivery system. Further study is essential to investigate the antimicrobial activity and safety of the gentamicin-loaded AC/α-TCP composites in animal models.

LL-37-dsRNA Complexes Modulate Immune Response via RIG-I in Oral Keratinocytes.

Recognition of nucleic acids as pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) promotes an inflammatory response. On the other hand, LL-37, an antimicrobial peptide, is a multifunctional modulator of immune response, though whether it modulates inflammatory responses induced by nucleic acids in oral keratinocytes is unknown. In this study, we firstly investigated the effect of LL-37 on CXCL10 induced by DAMPs and PAMPs in immortalized oral keratinocytes, RT7. Furthermore, the effects of LL-37 on translocation of exogenous nucleic acids into cytoplasm as well as cytosolic receptor, RIG-I on immune responses mediated by LL-37-nucleic acid complexes were examined. From these results, LL-37 enhanced necrotic cell supernatant (NCS)-induced CXCL10 expression in RT7, while the response was decreased by RNase. Complexes of LL-37 and double-stranded (ds) RNA, Poly(I:C) enhanced CXCL10 expression in comparison with each alone, which were associated with NF-κB activation. Furthermore, LL-37 was shown to bind with ds nucleotides and translocate into cytoplasm. Knockdown of RIG-I decreased expression of CXCL10 induced by LL-37-Poly(I:C) complexes, and RIG-I were co-localized with Poly(I:C) entered by LL-37 in cytoplasm. LL-37 modulates dsRNA-mediated inflammatory response via RIG-I in oral keratinocytes, which may play an important role in the pathogenesis of oral inflammatory diseases.

Correction of Severe Skeletal Class II High Angle with Mandibular Retrusion and Gummy Smile by Double-Jaw Surgery.

This report describes the treatment of severe skeletal Class II malocclusion in a young woman with a gummy smile and pronounced lower anterior facial height. Overjet and overbite were +12.0 mm and -1.0 mm, respectively. Cephalometric analysis revealed inferior positioning of the maxilla and severe mandibular retrusion with clockwise rotation. Both the upper and lower anterior teeth showed labial inclination. Based on a diagnosis of a skeletal Class II high angle with mandibular retrusion and a gummy smile, double-jaw orthognathic surgeries for upper and lower premolar extraction were chosen to gain ideal occlusion and an improvement in the esthetic facial profile. Le Fort I osteotomy was performed to move the anterior and posterior teeth upward by 4.0 mm and achieve mandibular counterclockwise rotation. Short lingual sagittal split ramus osteotomy was performed to move the mandible forward by 3.0 mm. As a result, normal overjet and overbite were achieved together with a straight profile and a good smile. After surgery, electromyographic evaluation of anterior temporal muscle activity showed an improvement in the percentage overlapping coefficient value (a symmetric index of bilateral muscle activity) from 28.1% to 63.2% compared to at pre-treatment. The pattern of jaw movement also showed an improvement. These results suggest that orthognathic surgery in skeletal Class II cases can improve not only malocclusion and the skeletal relationship of the jaws, but also masticatory function and jaw movement.

Prevalence of immune-related adverse events and anti-tumor efficacy following immune checkpoint inhibitor therapy in Japanese patients with various solid tumors.

BACKGROUND: While immune checkpoint inhibitors (ICIs) occasionally cause immune-related adverse events (irAEs) in various organs, the prevalence of irAEs and potential risk factors have not been clarified. We identified irAE predictive factors and examined the relationship between the effect of ICIs and irAEs for patients with malignancies. METHODS: A total of 533 cases treated with ICIs, including programmed death 1 (PD-1), PD-ligand 1 (PD-L1), and cytotoxic T-lymphocyte antigen 4 (CTLA-4), for various malignancies were included retrospectively. We recorded irAEs from medical records and graded them using the Common Terminology Criteria for Adverse Events version 5. Prevalence and predictive factors associated with immune-related liver injury and the relationship between irAE and treatment response were analyzed. RESULTS: During a median of 10 (1-103) cycles with a median follow-up after several ICI initiations of 384 (21-1715) days, irAEs with all grades and with grade ≥ 3 developed in 144 (27.0%) and 57 (10.7%) cases. Cumulative irAE development rates were 21.9, 33.5, and 43.0% in all grades and 8.8, 14.9, and 20.7% in grade ≥ 3 at 5, 10, and 20 cycles, respectively. Patients who received anti-CTLA4 therapy were more likely to develop irAEs compared to those who received anti-PD-1 or anti-PD-L1 monotherapy. Liver injury was the most common irAE. Multivariate analysis identified the combination of PD-1 and anti-CTL-4 antibodies (hazard ratio [HR], 17.04; P < 0.0001) and baseline eosinophil count ≥130/µL (HR, 3.01 for < 130; P = 0.012) as independent risk factors for the incidence of immune-related liver injury with grade ≥ 2. Patients who developed irAEs had a higher disease control rate (P < 0.0001) and an increased overall survival rate compared to those without irAEs (P < 0.0001). CONCLUSION: Combination therapy with anti-PD-1 and anti-CTL-4 antibodies resulted in higher a frequency of irAEs. Baseline absolute eosinophil count was found to be a predictive factor for immune-related liver injury. Occurrence of irAEs may be associated with higher efficacy of ICI treatment and longer survival among patients who receive ICI therapy.

Sunitinib promotes apoptosis via p38 MAPK activation and STAT3 downregulation in oral keratinocytes.

OBJECTIVE: Sunitinib, a targeted cancer drug, inhibits tyrosine kinases receptors and is widely used as first-line treatment for metastatic renal cell carcinoma. Patients undergoing chemotherapy with sunitinib frequently have oral mucosal complications, such as oral stomatitis, though cytotoxic effects of the drug on oral keratinocytes remain unknown. METHODS: The effects of sunitinib on immortalized oral keratinocytes, RT7 cells, in regard to cell injury and apoptosis, as well as apoptosis-mediated signaling pathways were investigated. RESULTS: Sunitinib treatment caused a significant increase in lactate dehydrogenase (LDH) in RT7 cells and primary oral keratinocytes. Additionally, the drug induced apoptosis-related events, such as DNA fragmentation, decreased anti-apoptotic Bcl-2 protein expression, and induction of cleaved PARP and caspase 3/9 in RT7 cells. Furthermore, phosphorylation of p38 MAPK, but not of ERK or JNK, was increased. On the contrary, constitutive phosphorylated STAT3 was decreased by sunitinib treatment, which was recovered by exposure to SB203580, a p38 MAPK inhibitor. Finally, SB203580 was found to reduce sunitinib-induced cell injury and apoptosis. CONCLUSION: The present results indicate that sunitinib promotes cell injury and apoptosis in oral keratinocytes via p38 activation and STAT3 downregulation. Sunitinib-mediated oral complications may be associated with cytotoxic effects of the drug on oral keratinocytes.

Effects of CEACAM1 in oral keratinocytes on HO-1 expression induced by Candida ß-glucan particles.

OBJECTIVE: Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is a member of the carcinoembryonic antigen family. Although its expression has been found in chronic oral inflammatory epithelium, this study aimed to know whether CEACAM1 in oral keratinocytes participates in host immune response against Candida albicans . METHODOLOGY: We investigated CEACAM1 expression in oral keratinocytes induced by C. albicans as well as by Candida cell wall component ß-glucan particles (ß-GPs). Furthermore, the effects of CEACAM1 on ß-GPs-induced heme oxygenase-1 (HO-1) expression and its related signals were examined. RESULTS: Fluorescence staining showed CEACAM1 expression in oral keratinocytes (RT7) cells, whereas quantitative reverse transcription (RT)-PCR indicated that both live and heat-killed C. albicans increased CEACAM1 mRNA expression in RT7 cells. Examinations using quantitative RT-PCR and western blotting indicated that CEACAM1 expression was also increased by ß-GPs derived from C. albicans . Specific siRNA for CEACAM1 decreased HO-1 expression induced by ß-GPs from C. albicans as well as the budding yeast microorganism Saccharomyces cerevisiae . Moreover, knockdown of CEACAM1 decreased ß-GPs-induced ROS activity in the early phase and translocation of Nrf2 into the nucleus. CONCLUSION: CEACAM1 in oral keratinocytes may have a critical role in regulation of HO-1 for host immune defense during Candida infection.

Long-standing temporomandibular joint dislocation treated by intraoral condylectomy: a case report and review of the literature.

BACKGROUND: Noninvasive management by closed reduction is a desirable treatment for temporomandibular joint dislocation. However, reduction of long-standing temporomandibular joint dislocation is often difficult. Various conservative treatments have been attempted, but these often render poor outcomes. This article reports the case of long-standing temporomandibular joint dislocation that was successfully closed using intraoral condylectomy. CASE PRESENTATION: A 69-year-old Japanese man who sustained an injury in a car collision was unable to close his mouth. Owing to the diagnosis of long-standing temporomandibular joint dislocation, intraoral condylectomy was performed. In the case of temporomandibular joint dislocation, it is convenient to reach the condyle from the oral cavity because sufficient opening is maintained. The condyle can be clearly visualized using an approach similar to sagittal split ramus osteotomy, and the operation using surgical instruments can be facilitated by resecting the coronoid process. By separating the surrounding soft tissue and pulling the cut condyle with sufficient visual field, the condyle can be resected while addressing the hemostasis. During the 12-month postoperative follow-up period, no temporomandibular joint dislocation recurred and the occlusion remained stable. CONCLUSIONS: The limited intraoral incision of this surgical technique provides sufficient access for condylectomy. The results of this case report suggest that condylectomy by intraoral approach could become the treatment of choice for long-standing temporomandibular joint dislocation.

Expression of anti-fungal peptide, ß-defensin 118 in oral fibroblasts induced by C. albicans ß-glucan-containing particles.

OBJECTIVE: Although oral fibroblasts are thought to have the potential to enhance host defenses against Candida albicans , it is unknown whether they are able to recognize Candida cell components to increase the expression of antifungal peptides, such as defensin factors, against Candida infection. METHODOLOGY: We performed expression profiles of defensin genes induced by heat-killed C. albicans in oral immortalized fibroblasts (GT1) using cDNA microarray analysis. From those results, quantitative RT-PCR was used to examine the effects of Candida ß-glucan-containing particles (ß-GPs) on ß-Defensin 118 (DEFB 118) expression in oral mucosal cells. Furthermore, the antifungal activities of recombinant DEFB 118 against C. albicans and C. glabrata were investigated using fungicidal assays. RESULTS: Microarray analysis showed that DEFB118, ß-Defensin 129 (DEFB129), and α-Defensin 1 (DEFA1) genes were induced by heat-killed C. albicans and that their mRNA expressions were also significantly increased by live as well as heat-killed C. albicans . Next, we focused on DEFB118, and found that GT1, primary fibroblasts, and RT7 (oral immortalized keratinocytes) constitutively expressed DEFB118 mRNA expression in RT-PCR. Furthermore, C. albicans ß-GPs significantly increased the expression of DEFB118 mRNA in GT1 and primary fibroblasts. Although DEFB118 mRNA expression in RT7 was significantly induced by both live and heat-killed C. albicans, C. albicans ß-GPs failed to have an effect on that expression. Finally, recombinant DEFB118 significantly decreased the survival of both strains of C. albicans in a dose-dependent manner, whereas no effects were seen for both C. glabrata strains. CONCLUSION: DEFB118, induced by C. albicans ß-GPs from oral fibroblasts, may play an important role in oral immune responses against C. albicans infection.

Two PARP13 isoforms are associated with induction of antiviral factors in oral mucosal cells.

Innate immune systems in the oral cavity have important roles in the host defense against viral invasion of oral mucosa. Poly(ADP­ribose) polymerase 13 (PARP13), which has a strong antiviral ability, has been reported to possess two isoforms; a full­length protein, zinc­finger antiviral protein long (ZAPL), and a shorter protein (ZAPS). However, the expression and function of these two isoforms in oral mucosa remain unknown. In the present study, the expression levels of ZAPL and ZAPS induced by transfected double­stranded (ds) RNA, Poly(I:C), and dsDNA, Poly(dA:dT), in immortalized oral keratinocytes and fibroblasts (RT7 and GT1 cell lines, respectively) were investigated. Subsequently, the effects of the knockdown of ZAPL and ZAPS on transfected nucleotide­induced antiviral factors were examined. The results demonstrated constitutive expression of ZAPL and ZAPS in RT7 and GT1 cells, and their expression in both cell types was notably increased by transfection of Poly(I:C) and Poly(dA:dT) when compared with no transfection. Specific knockdown of ZAPL and ZAPS in RT7 cells decreased IFN­ß and C­X­C motif chemokine ligand 10 (CXCL10) expression induced by transfected Poly(I:C) and Poly(dA:dT). On the other hand, knockdown of ZAPL and ZAPS in GT1 cells decreased the expression of CXCL10 induced by the transfected nucleotides, whereas that had no effect on IFN­ß expression induced by Poly(dA:dT). Their knockdown was also associated with transfected nucleotides­induced IFN regulatory factor 3 phosphorylation in both cell types. Taken together, these results indicate that ZAPL and ZAPS, isoforms of PARP13, in oral mucosal cells participate in host defense against viral infection of oral mucosa.

Melatonin enhances cisplatin-induced cell death through inhibition of DERL1 in mesenchymal-like CD44high OSCC cells.

BACKGROUND: Melatonin is a hormone that is primarily produced in the pineal gland and is involved in wide range of biological functions. However, the impact of melatonin on chemotherapy-induced cell death remains to be elucidated in oral squamous cell carcinoma (OSCC) cells. The objective of this study was to clarify the role of melatonin in cisplatin-induced cytotoxicity in CD44high OSCC cells. METHODS: CD44high OSCC cells were cultured on fibronectin-coated hydrogel. A lactate dehydrogenase cytotoxicity assay was performed to evaluate cisplatin-induced cell death. The effect of melatonin on cisplatin-induced cell death and Derlin-1 (DERL1) endoplasmic reticulum membrane protein expression was investigated. RESULTS: CD44high OSCC cells exhibited mesenchymal-like features when cultured on fibronectin-coated hydrogel. Mesenchymal-like CD44high OSCC cells demonstrated strong resistance to cisplatin-induced cell death compared with epithelial-like CD44high OSCC cells. DERL1 mRNA and DERL1 protein expression levels were significantly higher in mesenchymal-like CD44high cells compared with epithelial-like CD44high cells. Cisplatin-induced cell death was significantly enhanced after DERL1 siRNA knockdown, suggesting that DERL1 is involved in resistance to cisplatin-induced cell death. Melatonin significantly inhibited DERL1 expression and enhanced cisplatin-induced cell death in mesenchymal-like CD44high cells. miR-181c-5p expression was significantly upregulated in the presence of melatonin. Furthermore, melatonin-inhibited DERL1 expression was significantly recovered by miR-181c-5p inhibitor. In addition, melatoninenhanced cisplatin-induced cell death was attenuated by miR-181c-5p inhibitor. These results suggest that melatonin-induced miR-181c-5p enhances cisplatin-induced cell death through inhibition of DERL1 in mesenchymal-like CD44high cells. CONCLUSIONS: Melatonin plays a vital role in promoting cisplatin-induced cytotoxicity in mesenchymal-like CD44high OSCC cells.

Immune response to cytosolic DNA via intercellular receptor modulation in oral keratinocytes and fibroblasts.

OBJECTIVE: Double-strand (ds) DNA-enveloped viruses can cause oral infection. Our aim is to investigate whether oral mucosal cells participate in immune response against cytosolic dsDNA invasion. METHODS: We examined the response to transfected herpes simplex virus (HSV) dsDNA via intracellular receptors in oral keratinocytes (RT7) and fibroblasts (GT1), and the effect of TNF-α on those responses. RESULTS: Transfected dsDNA increased CXCL10 expression via NF-κB activation in both cell types, while those responses were inhibited by knockdown of RIG-I, an RNA sensor. Although IFI16, a DNA sensor, was expressed in the nuclei of both types, its knockdown decreased transfected dsDNA-induced CXCL10 expression in GT1 but not RT7 cells. IFI16 in GT1 cells was translocated into cytoplasm from nuclei, which was attributed to immune response to cytosolic dsDNA. TNF-α enhanced transfected dsDNA-induced CXCL10, and knockdown of IFI16 decreased TNF-α and dsDNA-driven CXCL10 expression in both RT7 and GT1 cells. Finally, the combination of TNF-α and transfected dsDNA resulted in translocation of IFI16 from nuclei to cytoplasm in RT7 cells. CONCLUSION: RIG-I and IFI16 in oral mucosal cells may play important roles in host immune response against DNA viral infection, while TNF-α contributes to development of an antiviral system via those intracellular receptors.

Effects of CEACAM1 in oral keratinocytes on HO-1 expression induced by Candida β-glucan particles

Abstract Objective Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is a member of the carcinoembryonic antigen family. Although its expression has been found in chronic oral inflammatory epithelium, this study aimed to know whether CEACAM1 in oral keratinocytes participates in host immune response against Candida albicans . Methodology We investigated CEACAM1 expression in oral keratinocytes induced by C. albicans as well as by Candida cell wall component β-glucan particles (β-GPs). Furthermore, the effects of CEACAM1 on β-GPs-induced heme oxygenase-1 (HO-1) expression and its related signals were examined. Results Fluorescence staining showed CEACAM1 expression in oral keratinocytes (RT7) cells, whereas quantitative reverse transcription (RT)-PCR indicated that both live and heat-killed C. albicans increased CEACAM1 mRNA expression in RT7 cells. Examinations using quantitative RT-PCR and western blotting indicated that CEACAM1 expression was also increased by β-GPs derived from C. albicans . Specific siRNA for CEACAM1 decreased HO-1 expression induced by β-GPs from C. albicans as well as the budding yeast microorganism Saccharomyces cerevisiae . Moreover, knockdown of CEACAM1 decreased β-GPs-induced ROS activity in the early phase and translocation of Nrf2 into the nucleus. Conclusion CEACAM1 in oral keratinocytes may have a critical role in regulation of HO-1 for host immune defense during Candida infection.

Expression of anti-fungal peptide, β-defensin 118 in oral fibroblasts induced by C. albicans β-glucan-containing particles

Abstract Objective: Although oral fibroblasts are thought to have the potential to enhance host defenses against Candida albicans , it is unknown whether they are able to recognize Candida cell components to increase the expression of antifungal peptides, such as defensin factors, against Candida infection. Methodology: We performed expression profiles of defensin genes induced by heat-killed C. albicans in oral immortalized fibroblasts (GT1) using cDNA microarray analysis. From those results, quantitative RT-PCR was used to examine the effects of Candida β-glucan-containing particles (β-GPs) on β-Defensin 118 (DEFB 118) expression in oral mucosal cells. Furthermore, the antifungal activities of recombinant DEFB 118 against C. albicans and C. glabrata were investigated using fungicidal assays. Results: Microarray analysis showed that DEFB118, β-Defensin 129 (DEFB129), and α-Defensin 1 (DEFA1) genes were induced by heat-killed C. albicans and that their mRNA expressions were also significantly increased by live as well as heat-killed C. albicans . Next, we focused on DEFB118, and found that GT1, primary fibroblasts, and RT7 (oral immortalized keratinocytes) constitutively expressed DEFB118 mRNA expression in RT-PCR. Furthermore, C. albicans β-GPs significantly increased the expression of DEFB118 mRNA in GT1 and primary fibroblasts. Although DEFB118 mRNA expression in RT7 was significantly induced by both live and heat-killed C. albicans, C. albicans β-GPs failed to have an effect on that expression. Finally, recombinant DEFB118 significantly decreased the survival of both strains of C. albicans in a dose-dependent manner, whereas no effects were seen for both C. glabrata strains. Conclusion: DEFB118, induced by C. albicans β-GPs from oral fibroblasts, may play an important role in oral immune responses against C. albicans infection.

Effects of miR-224-5p-enhanced downregulation of pannexin-1 on docetaxel-induced apoptosis in amoeboid-like CD44high oral cancer cells.

We previously found that microRNAs play major roles in the maintenance of amoeboid-like oral squamous cell carcinoma (OSCC) cells with high expression of CD44 (CD44high ). However, the roles of microRNAs in chemotherapeutic resistance exhibited by CD44high amoeboid-like OSCC cells are unclear. Here, docetaxel-induced apoptosis was examined in CD44high OSCC cells (CD44high OM-1 cells) cultured on laminin-coated silicone gel. Amoeboid-like CD44high OSCC cells exhibited robust resistance to docetaxel-induced apoptosis and significant upregulation of miR-224-5p expression compared with epithelial-like CD44high OSCC cells and mesenchymal-like CD44high OSCC cells. The expression of pannexin-1 (PANX1), a channel-forming protein that regulates the release of ATP, was significantly upregulated following transfection of amoeboid-like CD44high OSCC cells with an miR-224-5p inhibitor. These results suggest that miR-224-5p inhibits PANX1 expression. Furthermore, miR-224-5p inhibitor-transfected amoeboid-like CD44high OSCC cells exhibited significant enhancement of the proportion of apoptotic cells; however, this effect was significantly inhibited by knockdown of PANX1 with PANX1 small interfering RNA. Additionally, the miR-224-5p inhibitor-enhanced extracellular ATP levels were significantly reduced by PANX1 knockdown. These findings imply that miR-224-5p plays a vital role in the resistance to docetaxel-induced apoptosis by attenuating PANX1-induced ATP discharge. Moreover, amoeboid-like CD44high OSCC cells may be involved in chemotherapeutic resistance of OSCC.

The squamous cell carcinoma cell line OM-1 retains both p75-dependent stratified epithelial progenitor potential and cancer stem cell properties.

The low-affinity nerve growth factor receptor p75 is a stratified epithelial stem/progenitor marker of human epithelia. We found OM-1, a human squamous cell carcinoma (SCC) cell line, showed distinct cells with p75 cluster, especially located at the center of a growing colony in a monolayer culture. A cell with p75 cluster was surrounded by cytokeratin 14- and cytokeratin 13-expressing cells that settled at the outer margin of the colony. OM-1 cells were also capable of forming tumor spheres in a cell suspension culture, an ability which was attenuated by the inhibition of p75-signaling. Intriguingly, we also found a p75-negative cell population from a growing culture of OM-1 that re-committed to become p75-clustering cells. These results indicated the possibility that SCC with epithelial multi-layering capacity can exploit the p75-dependent stratified epithelial progenitor property for the cancer stemness.

Age-related changes in oral tactile and thermal sensation throughout adulthood.

Although the life expectancy of women is over 80 years in many countries, oral sensation has scarcely been compared between adults ≥ 80 years and younger age groups. The purpose of this study was to clarify age-related changes in oral sensation throughout adulthood. After exclusion of individuals with factors that might have confounded somatosensory performance, 123 female participants were divided into four age groups: 20-39 years, 40-59 years, 60-79 years, and 80-96 years. Perceptions of tactile and thermal sensations were examined at points on the anterior and posterior palate, anterior and posterior tongue, lower labial-attached gingiva, lower lip, and buccal mucosa; two-point discrimination was examined only on the tongue. The tactile and two-point discrimination thresholds for the anterior and posterior tongue were significantly higher in the 80-96-year-old group than in any other age group (p < 0.05). The tactile threshold for the buccal mucosa was significantly higher in the 80-96-year-old group than in the 60-79-year-old group (p < 0.05). The percentage of participants able to perceive a warm stimulus (50 °C) in the buccal mucosa was significantly lower in the 80-96-year-old group than in the 20-39-year-old group (p < 0.05). Only the topography of the warm sensation perception changed with age. This cross-sectional study suggests that oral tactile and thermal sensation for warm stimuli deteriorates with age in a site-specific manner, especially after the age of 80 years, but the same does not occur with cool stimuli.

In vitro investigation of the cell compatibility and antibacterial properties of titanium treated with calcium and ozone.

The purpose of this study was to evaluate the surface modification of calcium ions on roughened titanium as a surface treatment of dental implants for cell attachment, growth, and initial bacterial adhesion. When a surface-roughened, pure titanium disk was immersed in a calcium chloride solution (100 mM) containing 20 ppm ozone for 24 h at 25ºC, calcium was detected on the surface by X-ray photoelectron spectroscopy. The calcium-modified, roughened titanium disk had a significantly greater concentration of the initially adhered cells as well as cells cultured over 7 days compared with titanium disks without surface modification. Furthermore, the initial bacterial adhesion on the calcium-ozone treated titanium disk was statistically less than on a pure titanium disk or titanium disk treated without ozone. Dissolved ozone was useful for modifying the surface of roughened titanium with calcium ions and the surface modification may be applicable for dental implants.

TGF-ß1 induces amoeboid-to-mesenchymal transition of CD44high oral squamous cell carcinoma cells via miR-422a downregulation through ERK activation and Cofilin-1 phosphorylation.

BACKGROUND: The objective of this study was to clarify the molecular mechanism of amoeboid-to-mesenchymal transition (AMT) of CD44high oral squamous cell carcinoma (OSCC) cells. METHODS: Morphology and expression of mesenchymal genes were investigated in CD44high OSCC cells (CD44high OM-1 cells) cultured on laminin-coated soft silicone gel. Additionally, microarray analysis was performed to investigate microRNA (miRNA) expression inhibited by transforming growth factor-ß1 (TGF-ß1) in CD44high OM-1 cells. RESULTS: When CD44high OM-1 cells were cultured on 2.0-kPa laminin-coated silicone gel, the cells exhibited an amoeboid-like round morphology. Cofilin-1 expression was found in the nucleus and cytoplasm of amoeboid-like CD44high OM-1 cells. The invasive capacity was significantly reduced after Cofilin-1 knockdown. Additionally, Cofilin-1 knockdown cells had an irregularly extended shape. Phosphorylated Cofilin-1 was significantly upregulated by TGF-ß1. Additionally, TGF-ß1 enhanced N-cadherin and Snail mRNA expression and induced a spindle-shaped morphology. ERK1/2 phosphorylation was induced by TGF-ß1. Microarray analysis revealed that miR-422a exhibited the greatest downregulation (fold change: 0.22) in the presence of TGF-ß1. Importantly, TGF-ß1-inhibited miR-422a expression was recovered by the ERK inhibitor or ERK1/2 knockdown. Additionally, miR-422a inhibitor-transfected CD44high OM-1 cells exhibited high N-cadherin and Snail mRNA expression. Furthermore, Cofilin-1 knockdown and miR-422a inhibition induced a spindle cell morphology. CONCLUSION: Cofilin-1 is involved in the invasive ability of CD44high OSCC cells. TGF-ß1 contributes to AMT by downregulation of miR-422a via ERK activation and Cofilin-1 phosphorylation. Our findings suggest that miR-422a and Cofilin-1 play major roles in the maintenance of amoeboid-like CD44high cells.

Melatonin­induced miR­181c­5p enhances osteogenic differentiation and mineralization of human jawbone­derived osteoblastic cells.

Our previous study revealed that treatment with a combination of fibroblast growth factor­2 and melatonin (MEL) synergistically augmented osteogenic activity and mineralization of MC3T3­E1 mouse preosteoblast cells. Thus, the objective of the present study was to assess the effect of MEL on osteogenetic characteristics in human osteoblastic cells. Human jawbone­derived osteoblastic (hOB) cells were isolated from mandibular bone fragments. RUNX family transcription factor 2 (Runx2) expression, alkaline phosphatase (ALP) enzyme activity and the mineralization ability of hOB cells in the presence of MEL were evaluated. Microarray analysis was also performed to assess the expression of MEL­induced microRNAs (miRNAs/miRs) in hOB cells. Treatment with MEL significantly enhanced Runx2 expression, ALP activity and mineralization staining. However, this effect was significantly reduced following transforming growth factor­ß1 treatment. In total, 124 miRNAs were differentially expressed in MEL­treated hOB cells, compared with untreated cells. Of the upregulated miRNAs, miR­181c­5p exhibited the largest fold change. Runx2 mRNA expression and mineralization staining in the presence of MEL were significantly reduced following transfection with a miR­181c­5p inhibitor. In addition, transfection with miR-181c-5p mimics significantly increased Runx2 expression and mineralization staining. These results suggested that MEL­induced miR­181c­5p was involved in osteogenic differentiation and mineralization of hOB cells. Using TargetScan, a putative miR­181c­5p binding site was identified in the Notch2 gene. Moreover, Notch2 mRNA and protein expression levels in hOB cells were significantly reduced following transfection with miR­181c­5p mimics, confirming Notch2 as a target gene for miR­181c­5p. Notch2 siRNA knockdown significantly increased Runx2 expression and mineralization staining, which suggested that Notch2 may negatively regulate osteogenic differentiation of hOB cells by downregulating Runx2. In conclusion, MEL­induced expression of miR­181c­5p enhanced osteogenic differentiation and calcification of hOB cells.

Morphological Evaluation of Bone by CT to Determine Primary Stability-Clinical Study.

BACKGROUND: Primary stability is an important prognostic factor for dental implant therapy. In the present study, we evaluate the relationship between implant stability evaluation findings by the use of an implant stability quotient (ISQ), an index for primary stability, and a morphological evaluation of bone by preoperative computed tomography (CT). SUBJECTS AND METHODS: We analyzed 98 patients who underwent implant placement surgery in this retrospective study. For all 247 implants, the correlations of the ISQ value with cortical bone thickness, cortical bone CT value, cancellous bone CT value, insertion torque value, implant diameter, and implant length were examined. RESULTS: 1. Factors affecting ISQ values in all cases: It was revealed that there were significant associations between the cortical bone thickness and cancellous bone CT values with ISQ by multiple regression analysis. 2. It was revealed that there was a significant correlation between cortical bone thickness and cancellous bone CT values with ISQ by multiple regression analysis in the upper jaw. 3. It was indicated that there was a significant association between cortical bone thickness and implant diameter with ISQ by multiple regression analysis in the lower jaw. CONCLUSION: We concluded that analysis of the correlation of the ISQ value with cortical bone thickness and values obtained in preoperative CT imaging were useful preoperative evaluations for obtaining implant stability.