The Preventive Effect of Melatonin on Radiation-Induced Oral Mucositis.

Melatonin exerts various physiological effects through melatonin receptors and their ability to scavenge free radicals. Radiotherapy is a common treatment for head and neck tumors, but stomatitis, a side effect affecting irradiated oral mucosa, can impact treatment outcomes. This study investigated the preventive effect of melatonin, a potent free radical scavenger, on radiation-induced oral mucositis. Mice were irradiated with 15 Gy of X-ray radiation to the head and neck, and the oral mucosa was histologically compared between a melatonin-administered group and a control group. The results showed that radiation-induced oral mucositis was suppressed in mice administered melatonin before and after irradiation. It was suggested that the mechanism involved the inhibition of apoptosis and the inhibition of DNA damage. From these findings, we confirmed that melatonin has a protective effect against radiation-induced oral mucositis.

Dosage compensation of Z sex chromosome genes in avian fibroblast cells.

In birds, sex is genetically determined; however, the molecular mechanism is not well-understood. The avian Z sex chromosome (chrZ) lacks whole chromosome inactivation, in contrast to the mammalian chrX. To investigate chrZ dosage compensation and its role in sex specification, we use a highly quantitative method and analyze transcriptional activities of male and female fibroblast cells from seven bird species. Our data indicate that three fourths of chrZ genes are strictly compensated across Aves, similar to mammalian chrX. We also present a complete list of non-compensated chrZ genes and identify Ribosomal Protein S6 (RPS6) as a conserved sex-dimorphic gene in birds.

A New Implantation Method for Orthodontic Anchor Screws: Basic Research for Clinical Applications.

This study aimed to determine whether the positional relationship between the underside of the screw head and the surface of the alveolar bone could alter the stress on the two surfaces and affect the stability of implanted anchor screws. First, in order to confirm the extent of the gap between the mini-screw and the bone surface, a mini-screw was placed in the palate of rabbits and examined histologically. As a result, in the conventional screw implantation procedure, oral mucosa between the base of the screw head and the bone creates a spatial gap. Removal of the oral mucosa eliminates this gap. Then, we compared the positional difference of the screw in a contact and gap group by analyzing stress distribution on the bone and screw. Analysis using the finite element method showed that more stress was loaded on both the bone and screw in the gap group than in the contact group. Cortical bone thickness did not affect stress in either group. The effects of different load strengths were similar between groups. A surgical procedure in which mucosal coverings are removed so that implanted anchor mini-screws are in contact with the bone surface was found to reduce the stress load on both the bone and screw. This procedure can be used to prevent undesirable dislodgement of implanted mini-screws.

Inhibitory effect of 405-nm blue LED light on the growth of Candida albicans and Streptococcus mutans dual-species biofilms on denture base resin.

We investigated whether irradiation with 405-nm blue LED light could inhibit the growth of not only single- but dual-species biofilms formed by Candida albicans and Streptococcus mutans on denture base resin and cause the alteration in gene expression related to adhesion and biofilm formation. C. albicans and S. mutans single-/dual-species biofilms were formed on the denture base specimens. The biofilms were irradiated with 405-nm blue LED light (power density output: 280 mW/cm2) for 0 (control) and 40 min. Dual-species biofilms were analyzed using CFU assay and fluorescence microscopy, and single-/dual-species biofilms were analyzed using alamarBlue assays and gene expression analysis. To assess the inhibitory effect of irradiation on dual-species biofilms, specimens after irradiation were aerobically incubated for 12 h. After incubation, the inhibition of growth was assessed using CFU assays and fluorescence microscopy. Data were analyzed using the Mann-Whitney U or Student's t test (p < 0.05). Irradiation produced a significant inhibitory effect on biofilms. Fluorescence microscopy revealed that almost all C. albicans and S. mutans cells were killed by irradiation, and there was no notable difference in biofilm thickness immediately after irradiation and after irradiation and incubation for 12 h. alamarBlue assays indicated the growth of the biofilms was inhibited for 12-13 h. The expression of genes associated with adhesion and biofilm formation-als1 in C. albicans and ftf, gtfC, and gtfB in S. mutans-significantly reduced by irradiation. Irradiation with 405-nm blue LED light effectively inhibited the growth of C. albicans and S. mutans dual-species biofilms for 12 h.

Microbicidal effect of 405-nm blue LED light on Candida albicans and Streptococcus mutans dual-species biofilms on denture base resin.

This study investigated: (1) the microbicidal effect of 405-nm blue LED light irradiation on biofilm formed by Candida albicans hyphae and Streptococcus mutans under dual-species condition on denture base resin, (2) the generation of intracellular reactive oxygen species (ROS) induced by irradiation, and (3) the existence of intracellular porphyrins, which act as a photosensitizer. Denture base resin specimens were prepared and C. albicans and S. mutans dual-species biofilms were allowed to form on the specimens. The biofilms were irradiated with 405-nm blue LED light and analyzed using the colony-forming unit assay, fluorescence microscopy, and scanning electron microscopy (SEM). Single-species biofilms of C. albicans and S. mutans formed on the specimens were irradiated with 405-nm blue LED light. After the irradiation, the intracellular ROS levels in C. albicans and S. mutans cells were measured. In addition, the level of intracellular porphyrins in C. albicans and S. mutans were measured. Irradiation for more than 30 min significantly inhibited the colony formation ability of C. albicans and S. mutans. Fluorescence microscopy revealed that almost all C. albicans and S. mutans cells were killed by irradiation. SEM images showed various cell damage patterns. Irradiation led to the generation of intracellular ROS and porphyrins were present in both C. albicans and S. mutans cells. In conclusion, irradiation with 405-nm blue light-emitting diode light for 40 min effectively disinfect C. albicans hyphae and S. mutans dual-species biofilms and possibly react with intracellular porphyrins resulting in generation of ROS in each microorganism.

A New Induction Method for the Controlled Differentiation of Human-Induced Pluripotent Stem Cells Using Frozen Sections.

Considering that every tissue/organ has the most suitable microenvironment for its functional cells, controlling induced pluripotent stem cell (iPSC) differentiation by culture on frozen sections having a suitable microenvironment is possible. Induced PSCs were cultured on frozen sections of the liver, the brain, the spinal cord, and cover glasses (control) for 9 days. The iPSCs cultured on the sections of the liver resembled hepatocytes, whereas those on sections of the brain and the spinal cord resembled neuronal cells. The percentage of hepatocytic marker-positive cells in the iPSCs cultured on the sections of the liver was statistically higher than that of those in the iPSCs cultured on the sections of the brain and the spinal cord or on cover glasses. In contrast, the iPSCs cultured on the sections of the brain and the spinal cord revealed a high percentage of neural marker-positive cells. Thus, iPSCs can be differentiated into a specific cell lineage in response to specific factors within frozen sections of tissues/organs. Differentiation efficacy of the frozen sections markedly differed between the iPSC clones. Therefore, our induction method could be simple and effective for evaluating the iPSC quality.

Feasibility of Rapid Diagnostic Technology for SARS-CoV-2 Virus Using a Trace Amount of Saliva.

Containment of SARS-CoV-2 has become an urgent global issue. To overcome the problems of conventional quantitative polymerase chain reaction (qPCR) tests, we verified the usefulness of a mobile qPCR device that utilizes mouthwash to obtain a saliva sample with the aim of developing a rapid diagnostic method for SARS-CoV-2. First, we examined whether anyone could easily operate this device. Then, we examined whether RNA in the mouthwash could be detected in a short time. In addition, we investigated whether it was possible to diagnose SARS-CoV-2 infection using mouthwash obtained from COVID-19 patients undergoing hospitalization. The results revealed that all subjects were able to complete the operation properly without error. In addition, RNase P was detected in the mouthwash without pretreatment. The average detection time was 18 min, which is significantly shorter than conventional qPCR devices. Furthermore, this device detected SARS-CoV-2 in the mouthwash of a COVID-19 patient undergoing hospitalization. The above findings verified the efficacy of this diagnostic method, which had a low risk of infection, was technically simple, and provided stable results. Therefore, this method is useful for the rapid detection of SARS-CoV-2.

Grapefruit seed extract effectively inhibits the Candida albicans biofilms development on polymethyl methacrylate denture-base resin.

This study aimed to investigate the cleansing effects of grapefruit seed extract (GSE) on biofilms of Candida albicans (C. albicans) formed on denture-base resin and the influence of GSE on the mechanical and surface characteristics of the resin. GSE solution diluted with distilled water to 0.1% (0.1% GSE) and 1% (1% GSE) and solutions with Polident® denture cleansing tablet dissolved in distilled water (Polident) or in 0.1% GSE solution (0.1% G+P) were prepared as cleansing solutions. Discs of acrylic resin were prepared, and the biofilm of C. albicans was formed on the discs. The discs with the biofilm were treated with each solution for 5 min at 25°C. After the treatment, the biofilm on the discs was analyzed using a colony forming unit (CFU) assay, fluorescence microscopy, and scanning electron microscopy (SEM). In order to assess the persistent cleansing effect, the discs treated with each solution for 5 min were aerobically incubated in Yeast Nitrogen Base medium for another 24 h. After incubation, the persistent effect was assessed by CFU assay. Some specimens of acrylic resin were immersed in each solution for 7 days, and changes in surface roughness (Ra), Vickers hardness (VH), flexural strength (FS), and flexural modulus (FM) were evaluated. As a result, the treatment with 1% GSE for 5 min almost completely eliminated the biofilm formed on the resin; whereas, the treatment with 0.1% GSE, Polident, and 0.1% G+P for 5 min showed a statistically significant inhibitory effect on biofilms. In addition, 0.1% GSE and 0.1% G+P exerted a persistent inhibitory effect on biofilms. Fluorescence microscopy indicated that Polident mainly induced the death of yeast, while the cleansing solutions containing at least 0.1% GSE induced the death of hyphae as well as yeast. SEM also revealed that Polident caused wrinkles, shrinkage, and some deep craters predominantly on the cell surfaces of yeast, while the solutions containing at least 0.1% GSE induced wrinkles, shrinkage, and some damage on cell surfaces of not only yeasts but also hyphae. No significant changes in Ra, VH, FS, or FM were observed after immersion in any of the solutions. Taken together, GSE solution is capable of cleansing C. albicans biofilms on denture-base resin and has a persistent inhibitory effect on biofilm development, without any deteriorations of resin surface.

Effectiveness of 405-nm blue LED light for degradation of Candida biofilms formed on PMMA denture base resin.

This study investigated (i) the degradation effect of 405-nm blue light-emitting diode (LED) light irradiation on Candida albicans and C. glabrata biofilms formed on denture base resin and (ii) the effects of 405-nm blue LED light irradiation on the mechanical and surface characteristics of the resin. Polymethyl methacrylate denture base resin discs were prepared, and C. albicans or C. glabrata biofilms formed on the denture base resin discs. Each biofilm was irradiated with 405-nm blue LED light under a constant output power (280 mW/cm2) for different times in a moisture chamber with 100% relative humidity. Postirradiation, each biofilm was analyzed using a colony-forming unit assay, fluorescence microscopy, and scanning electron microscopy (SEM). Parallelepiped specimens of acrylic resin were prepared, and changes in their flexural strength (FS), flexural modulus (FM), and surface roughness (Ra) preirradiation and postirradiation with 405-nm blue LED light were evaluated. Irradiation for 30 min completely inhibited colony formation in both Candida species. Fluorescence microscopy showed that almost all Candida cells were killed because of irradiation. SEM images showed various cell damage patterns, such as wrinkles, shrinkage, and cell surface damage. An increase in FS was noted postirradiation, but no significant changes were observed in FM and Ra preirradiation and postirradiation. In conclusion, irradiation with 405-nm blue LED light induces degradation of C. albicans and C. glabrata biofilms on denture base resin, even in the absence of photosensitizers, without resin surface deterioration.

Leptin promotes wound healing in the skin.

INTRODUCTION: Leptin, a 16 kDa anti-obesity hormone, exhibits various physiological properties. Interestingly, skin wound healing was proven to delay in leptin-deficient ob/ob mice. However, little is known on the mechanisms of this phenomenon. In this study, we attempted to elucidate a role of leptin in wound healing of skin. METHODS: Immunohistochemical analysis was performed to confirm the expression of the leptin receptor (Ob-R) in human and mouse skin. Leptin was topically administered to chemical wounds created in mouse back skin along with sustained-release absorbable hydrogel. The process of wound repair was histologically observed and the area of ulceration was measured over time. The effect of leptin on the proliferation, differentiation and migration of human epidermal keratinocytes was investigated. RESULTS: Ob-R was expressed in epidermal cells of human and mouse skin. Topical administration of leptin significantly promoted wound healing. Histological analysis showed more blood vessels in the dermal connective tissues in the leptin-treated group. The proliferation, differentiation/function and migration of human epidermal keratinocytes were enhanced by exogenous leptin. CONCLUSION: Topically administered leptin was proven to promote wound healing in the skin by accelerating proliferation, differentiation/function and migration of epidermal keratinocytes and enhancing angiogenesis around the wounded area. These results strongly suggest that topical administration of leptin may be useful as a treatment to promote wound healing in the skin.

Leptin promotes wound healing in the oral mucosa.

INTRODUCTION: Leptin, a 16 kDa circulating anti-obesity hormone, exhibits many physiological properties. Recently, leptin was isolated from saliva; however, its function in the oral cavity is still unclear. In this study, we investigated the physiological role of leptin in the oral cavity by focusing on its effect on wound healing in the oral mucosa. METHODS: Immunohistochemical analysis was used to examine the expression of the leptin receptor (Ob-R) in human/rabbit oral mucosa. To investigate the effect of leptin on wound healing in the oral mucosa, chemical wounds were created in rabbit oral mucosa, and leptin was topically administered to the wound. The process of wound repair was histologically observed and quantitatively analyzed by measuring the area of ulceration and the duration required for complete healing. The effect of leptin on the proliferation, differentiation and migration of human oral mucosal epithelial cells (RT7 cells) was investigated using crystal violet staining, reverse transcription polymerase chain reaction (RT-PCR) and a wound healing assay, respectively. RESULTS: Ob-R was expressed in spinous/granular cells in the epithelial tissue and vascular endothelial cells in the subepithelial connective tissue of the oral mucosa. Topical administration of leptin significantly promoted wound healing and shortened the duration required for complete healing. Histological analysis of gingival tissue beneath the ulceration showed a denser distribution of blood vessels in the leptin-treated group. Although the proliferation and differentiation of RT7 cells were not affected by leptin, the migration of these cells was accelerated in the presence of leptin. CONCLUSION: Topically administered leptin was shown to promote wound healing in the oral mucosa by accelerating epithelial cell migration and enhancing angiogenesis around the wounded area. These results strongly suggest that topical administration of leptin may be useful as a treatment to promote wound healing in the oral mucosa.

Expression and cellular localizaion of melatonin-synthesizing enzymes in rat and human salivary glands.

Melatonin, discovered in 1958, is secreted by the pineal gland primarily during the night. Its secretion is controlled by the light/dark cycle of the environment. Melatonin is also produced in and secreted by various extrapineal organs, tissues and cells and its synthesizing enzyme arylalkylamine N-acetyltransferase (AANAT) is expressed in various extrapineal organs, tissues and cells. Recently, it was reported that melatonin is present in saliva, but it is not certain where melatonin was synthesized and whether it was secreted into saliva and what function it may have in saliva. The present study was performed to investigate where melatonin was synthesized and whether it was secreted by salivary glands into saliva. We performed immunohistochemical analysis of the expression of AANAT in rat parotid, submandibular and sublingual glands and the expression of both AANAT and hydroxyindole-O-methyltransferase (HIOMT) in human submandibular glands. We evaluated the expression of AANAT and HIOMT mRNA in rat submandibular glands by quantitative reverse transcription-polymerase chain reaction. As a result, we observed expression of AANAT in epithelial cells of striated ducts in rat salivary glands and expression of AANAT, HIOMT and melatonin in epithelial cells of striated ducts in human submandibular glands. In addition, we also confirmed the expression of the most potent melatonin receptor, melatonin 1a receptor, in rat buccal mucosa. Our findings suggest that melatonin might be produced and secreted by salivary glands directly into saliva and that it might play some physiological role in the oral cavity.

Leptin and vascular endothelial growth factor regulate angiogenesis in tooth germs.

Leptin, a 16 kDa non-glycolated polypeptide of 146 amino acids produced by the ob gene, has a variety of physiological roles not only in lipid metabolism, hematopoiesis, thermogenesis and ovarian function, but also in angiogenesis. This study focuses to investigate the possibility that leptin, as an angiogenic factor, may regulate the angiogenesis during tooth development. We firstly studied the expression of leptin and vascular endothelial growth factor (VEGF) during tooth development immunohistochemically. This investigation revealed that leptin is expressed in ameloblasts, odontoblasts, dental papilla cells and stratum intermedium cells. This expression pattern was similar to that of VEGF, one of the most potent angiogenic factors. Interestingly, more leptin-positive cells were observed in the upper third portion of dental papilla, which is closest to odontoblastic layer, compared to middle and lower thirds. Moreover, in the dental papilla, more CD31 and/or CD34-positive vascular endothelial cells were observed in the vicinity of ameloblasts and odontoblasts expressing leptin and VEGF. These findings strongly suggest that ameloblasts, odontoblasts and dental papilla cells induce the angiogenesis in tooth germs by secretion of leptin as well as VEGF.

Possible involvement of maspin in tooth development.

Maspin is a 42 kDa serine protease inhibitor that possesses tumor suppressive and anti-angiogenic activities. Despite of a huge amount of data concerning the expression pattern of maspin in various tissues and its relevance to the biological properties of a variety of human cancer cells, little is known on the maspin expression in skeletal and tooth tissues. Recently, we reported that maspin may play an important role in extracellular matrix formation in bone by enhancing the accumulation of latent TGF-ß in the extracellular matrix. This study was performed to elucidate the possible role of maspin in tooth development. First, an immunohistochemical analysis for human tooth germs at the late bell stage showed the expression of maspin by active ameloblasts and odontoblasts that were forming enamel and dentin, respectively. During rat tooth development, maspin expression was observed for the first time in inner and outer enamel epithelial cells and dental papilla cells at early bell stage. The neutralizing anti-maspin antibody inhibited the proper dental tissue formation in organ cultures of mandibular first molars obtained from 21-day-old rat embryos. In addition, the proliferation of HAT-7 cells, a rat odontogenic epithelial cell line, and human dental papilla cells were suppressed in a dose-dependent manner with anti-maspin antibody. Moreover, RT-PCR analysis showed that the expression of mRNA for tooth-related genes including dentin matrix protein 1, dentin sialophosphoprotein and osteopontin in human dental papilla cells was inhibited when treated with anti-maspin antibody. These findings suggest that maspin expressed in ameloblasts and odontoblasts plays an important physiological role in tooth development through the regulation of matrix formation in dental tissues.