Radio Frequency Identification Gate System to Identify Misused Personal Dosimeters.

The use of two personal dosimeters, one worn over and one worn under a protective apron, provides the best estimate of effective dose. However, inappropriate positioning of dosimeters is a common occurrence, resulting in abnormally high or low radiation exposure records. Although such incorrect positioning can be identified by radiation exposure records, doing so is time-consuming and labor-intensive for administrators. Therefore, a system that can identify incorrect locations of dosimeters without burdening administrators must be developed. In this study, we developed a radio frequency identification (RFID) gate system that can differentiate between two RFID-tagged dosimeters placed over and under a metal apron and identify misused dosimeters. To simulate the position of the RFID-tagged dosimeters, we designed four dosimeter-wearing classes, including "proper use" and three types of "misuse" (i.e., "reversed," "both under," and "both over"). When the system predicts "misuse" based on the tag reading, the worker is alerted with lights and alarms. The system performance was evaluated using a confusion matrix, with an overall accuracy of 97.75%, demonstrating high classification performance. The safety of the system against life support devices was also investigated, demonstrating that they were not affected by the electric field at 0.3 m or more from the antenna of the system under any transmit powers tested. This RFID gate system is highly capable of identifying incorrectly positioned dosimeters, enabling real-time monitoring of dosimeters to manage their positioning.

Evaluation of dental hygiene learning outcomes through smartphone-based educational slides targeting dental hygienists: A pilot study.

OBJECTIVE: Education reflecting fundamental knowledge is required for competent health care providers, but often lectures are not available for this purpose. This study aimed to evaluate the dental hygiene learning outcomes following the presentation of web-based slides on a smartphone to dental hygienists. METHODS: A two-group nonblinded quasi-experimental design was used for this study. Forty-six dental hygienists were assigned to a study (n = 31) or control group (n = 15). The study group viewed 22 slides on fundamental oral health knowledge using smartphones. Pre and postviewing tests (score range: 1-13) and a questionnaire were conducted to evaluate knowledge acquisition and to receive feedback from participants. Differences between the study and control group and intrastudy group differences were statistically evaluated. RESULTS: The fundamental knowledge of dental hygienists improved after viewing the slides: the study group had a significantly higher mean score than the control group (10.87 vs. 6.60; p < 0.001). Study group participants also had substantially higher post-test than pretest knowledge scores (mean 10.87 vs. 6.26, p < 0.001). In the questionnaire, more than 85% of the participants answered that the content of the slides would be useful in their clinical practice. CONCLUSION: Smartphone-based educational slides were beneficial for conveying fundamental and recent oral health knowledge to dental hygienists.

The Identification of Marker Genes for Predicting the Osteogenic Differentiation Potential of Mesenchymal Stromal Cells.

Mesenchymal stromal cells (MSCs) have the potential to differentiate into a variety of mature cell types and are a promising source of regenerative medicine. The success of regenerative medicine using MSCs strongly depends on their differentiation potential. In this study, we sought to identify marker genes for predicting the osteogenic differentiation potential by comparing ilium MSC and fibroblast samples. We measured the mRNA levels of 95 candidate genes in nine ilium MSC and four fibroblast samples before osteogenic induction, and compared them with alkaline phosphatase (ALP) activity as a marker of osteogenic differentiation after induction. We identified 17 genes whose mRNA expression levels positively correlated with ALP activity. The chondrogenic and adipogenic differentiation potentials of jaw MSCs are much lower than those of ilium MSCs, although the osteogenic differentiation potential of jaw MSCs is comparable with that of ilium MSCs. To select markers suitable for predicting the osteogenic differentiation potential, we compared the mRNA levels of the 17 genes in ilium MSCs with those in jaw MSCs. The levels of 7 out of the 17 genes were not substantially different between the jaw and ilium MSCs, while the remaining 10 genes were expressed at significantly lower levels in jaw MSCs than in ilium MSCs. The mRNA levels of the seven similarly expressed genes were also compared with those in fibroblasts, which have little or no osteogenic differentiation potential. Among the seven genes, the mRNA levels of IGF1 and SRGN in all MSCs examined were higher than those in any of the fibroblasts. These results suggest that measuring the mRNA levels of IGF1 and SRGN before osteogenic induction will provide useful information for selecting competent MSCs for regenerative medicine, although the effectiveness of the markers is needed to be confirmed using a large number of MSCs, which have various levels of osteogenic differentiation potential.

DEC1 regulates the rhythmic expression of PPARγ target genes involved in lipid metabolism in white adipose tissue.

Previously, we found that the basic helix-loop-helix transcriptional repressor DEC1 interacts with the PPARγ:RXRα heterodimer, a master transcription factor for adipogenesis and lipogenesis, to suppress transcription from PPARγ target genes (Noshiro et al., Genes to Cells, 2018, 23:658-669). Because the expression of PPARγ and several of its target genes exhibits circadian rhythmicity in white adipose tissue (WAT), we examined the expression profiles of PPARγ target genes in wild-type and Dec1-/- mice. We found that the expression of PPARγ target genes responsible for lipid metabolism, including the synthesis of triacylglycerol from free fatty acids (FFAs), lipid storage and the lipolysis of triacylglycerol to FFAs, oscillates in a circadian manner in WAT. Moreover, DEC1 deficiency led to a marked increase in the expression of these genes at night (Zeitgeber times 16 and 22), resulting in disruption of circadian rhythms. Serum FFA levels in wild-type mice also showed circadian oscillations, but these were disrupted by DEC1 deficiency, leading to reduced FFA levels. These results suggest that PPARγ:RXRα and DEC1 cooperatively generate the circadian expression of PPARγ target genes through PPAR-responsive elements in WAT.

Deficiency of the basic helix-loop-helix transcription factor DEC1 prevents obesity induced by a high-fat diet in mice.

Obesity is a major public health problem in developed countries resulting from increased food intake and decreased energy consumption and usually associated with abnormal lipid metabolism. Here, we show that DEC1, a basic helix-loop-helix transcription factor, plays an important role in the regulation of lipid consumption in mouse brown adipose tissue (BAT), which is the major site of thermogenesis. Homozygous Dec1 deletion attenuated high-fat-diet-induced obesity, adipocyte hypertrophy, fat volume and hepatic steatosis. Furthermore, DEC1 deficiency increased body temperature during daytime and enhanced the expression of uncoupler protein 1, a key factor of thermogenesis, and various lipolysis-related genes in interscapular BAT. In vitro experiments suggested that DEC1 suppresses the expression of various lipolysis-related genes induced by the heterodimer of peroxisome proliferator-activated receptor γ and retinoid X receptor α (RXRα) through direct binding to RXRα. These observations suggest that enhanced lipolysis in BAT caused by DEC1 deficiency leads to an increase in lipid consumption, thereby decreasing lipid accumulation in adipose tissues and the liver. Thus, DEC1 may serve as an energy-saving factor that suppresses lipid consumption, which may be relevant to managing obesity.

Mesenchymal stem cells ameliorate experimental peritoneal fibrosis by suppressing inflammation and inhibiting TGF-ß1 signaling.

Mesenchymal stem cells (MSCs) are multipotent adult stem cells that have regenerative capability and exert paracrine actions on damaged tissues. Since peritoneal fibrosis is a serious complication of peritoneal dialysis, we tested whether MSCs suppress this using a chlorhexidine gluconate model in rats. Although MSCs isolated from green fluorescent protein-positive rats were detected for only 3 days following their injection, immunohistochemical staining showed that MSCs suppressed the expression of mesenchymal cells, their effects on the deposition of extracellular matrix proteins, and the infiltration of macrophages for 14 days. Moreover, MSCs reduced the functional impairment of the peritoneal membrane. Cocultures of MSCs and human peritoneal mesothelial cells using a Transwell system indicated that the beneficial effects of MSCs on the glucose-induced upregulation of transforming growth factor-ß1(TGF-ß1) and fibronectin mRNA expression in the human cells were likely due to paracrine actions. Preincubation in MSC-conditioned medium suppressed TGF-ß1-induced epithelial-to-mesenchymal transition, α-smooth muscle actin, and the decrease in zonula occludens-1 in cultured human peritoneal mesothelial cells. Although bone morphogenic protein 7 was not detected, MSCs secreted hepatocyte growth factor and a neutralizing antibody to this inhibited TGF-ß1 signaling. Thus, our findings imply that MSCs ameliorate experimental peritoneal fibrosis by suppressing inflammation and TGF-ß1 signaling in a paracrine manner.

Regulation of basic helix-loop-helix transcription factors Dec1 and Dec2 by RORα and their roles in adipogenesis.

DEC1 and DEC2, members of the basic helix-loop-helix superfamily, are involved in various biological phenomena including clock systems, cell differentiation and metabolism. In clock systems, Dec1 and Dec2 expression are up-regulated by the CLOCK:BMAL1 heterodimer via E-box (CACGTG), exhibiting a circadian rhythm in the suprachiasmatic nucleus (SCN), the central circadian pacemaker and other peripheral tissues. In this study, using assays of luciferase reporters, electrophoretic mobility shift and chromatin immunoprecipitation, we identified novel nuclear receptor response elements, ROR response elements (RORE), in Dec1 and Dec2 promoters. These ROREs responded to the transcriptional activator RORα, but not to the repressor REVERBα, although the Bmal1 promoter responded to both RORα and REVERBα. Therefore, RORα, but not REVERBα, is involved in the regulation of Dec1 and Dec2 expression without significantly affecting their rhythmicity. Since RORα, DEC1 and DEC2 reportedly suppressed adipogenic differentiation, we examined expression of Rorα, Dec1, Dec2 and other clock-controlled genes in differentiating 3T3-L1 adipocytes. The results suggested that RORα suppresses adipogenic differentiation at a later stage of differentiation by RORE-mediated stimulation of Dec1 and Dec2 expression.

Use of an educational video with mobile technology for the informed consent process: A randomised controlled trial on patients undergoing surgery for an impacted lower third molar.

The purpose of this study was to determine whether audiovisual presentation of consent information would significantly improve patients' postoperative recall of risks and complications regarding surgical removal of impacted lower third molars compared to the presentation of traditional written consent information. A randomised controlled study on 59 patients undergoing third molar removal was conducted. Patients in the intervention group (n = 30) viewed an educational video on risks and complications related to surgery using mobile tablets. Control-group patients (n = 29) received written information of the risks and complications. Patients' postoperative recall of potential risks for dysesthesia of the lower lip and tongue, infection, and bleeding along with surgical complications of facial oedema, trismus, and pain were assessed using true-false tests. The effect of audiovisual information on postoperative recall of the risks and complications was determined by comparing accuracy scores between the intervention group and control group using the independent t-test. The intervention group was found to have significantly better recall scores of the potential risks and complications, due to much higher accuracy in their recall of bleeding and dysesthesia of the lower lip and/or tongue, compared to the control group [mean (SD) 4.70 (0.94) vs 3.76 (1.50), p = 0.003]. The use of an educational video played on mobile tablets rather than a written pamphlet may lead to better understanding of the informed consent process in patients.

Evaluation of Inappropriate Positioning of Dosimeters in Medical Workers Based on Dose Equivalent Hp(10).

ABSTRACT: This report presents a new method to characterize the inappropriate positioning of dosimeters based on the dose equivalent Hp(10). The Hp(10) values of medical workers were measured monthly for 12 mo using two personal dosimeters. Using the ratio between the values of Hp(10) recorded from dosimeters worn over and under protective aprons [Hp(10) over and Hp(10) under , respectively], 670 pairs of dosimeter readings were categorized into a proper use group [Hp(10) over /Hp(10) under ≥ 5] and a misuse group [Hp(10) over /Hp(10) under < 5]. Following personal interviews, the readings in the misuse group were classified into the following six subgroups: "reversed," "sometimes reversed," "both under," "both over," "without apron," and "not specified." Ultimately, the scatter plot of "Hp(10) over - Hp(10) under " vs. Hp(10) over was identified as the most promising tool for clarifying the misuse patterns of dosimeters, as individual readings were mapped to the locations of the corresponding subgroups in the obtained graphs. Our results are expected to facilitate efficient and accurate usage of dosimeters by medical workers.

IL-1ß-mediated up-regulation of DEC1 in human gingiva cells via the Akt pathway.

Growing evidence indicates that inflammation is a contributing factor leading to cancer development. However, pathways involved in this progression are not well understood. The involvement of DEC1 in cancer prompted us to examine whether pro-inflammatory cytokine interleukin-1ß (IL-1ß) induces the expression of DEC1 in oral inflammation. We found that IL-1ß up-regulated DEC1 and hypoxia-inducible factor-1α (HIF-1α) protein and elevated the HIF-1α-responsive gene vascular endothelial growth factor (VEGF) expression in human primary gingival cells. HIF-1α and DEC1 immunoreactivity were significantly higher in the cases of gingival inflammation. We demonstrate that IL-1ß up-regulates DEC1 and HIF-1α protein through a classical inflammatory signaling pathway involving Akt. Our data strongly suggest that PI-3K-Akt is an upstream participant in IL-1ß-mediated DEC1 and HIF-1α induction. This is supported by the following data: (1) IL-1ß induces 473 serine phosphorylation of Akt; (2) IL-1ß-mediated Akt activation occurs in a PI-3K-dependent manner, and specific inhibition of PI-3K prevents Akt phosphorylation; and (3) inhibition of Akt prevents IL-1ß-mediated DEC1 and HIF-1α induction. Taken together, these results suggest that DEC1 is one of the important transcription factors in inflammation.

Smad3 and Snail show circadian expression in human gingival fibroblasts, human mesenchymal stem cell, and in mouse liver.

Smads are intracellular signaling mediators. Complexes of Smad2 and Smad3 with Smad4 transmit transforming growth factor-beta (TGF-ß) receptor-induced signaling. Snail plays important roles in mesoderm formation, gastrulation, neural crest development, and epithelial mesenchymal transition. However, it remains unknown whether Smad3 and Snail expression is circadian rhythm-dependent. Here, we showed for the first time that Smad3 and Snail show circadian expression in human gingival fibroblasts (HGF-1) and human mesenchymal stem cells (MSC) after serum shock. They also showed circadian expression in the mouse liver. We confirmed that BMAL1/2, DEC1/2, VEGF, and PER1/2/3 also show circadian expression in both HGF-1 and MSC. The mRNA peaks and phases in circadian expression of these genes differed between HGF-1 and MSC. In a luciferase assay, Smad3 promoter activity was upregulated by CLOCK/BMAL1. These findings suggest that Smad3 and Snail have circadian rhythm in vitro and vivo, and that circadian expression of Smad3 depends on CLOCK/BMAL1.

Basic helix-loop-helix transcription factor DEC1 negatively regulates cyclin D1.

DEC1 (also known as Stra13/Bhlhb2/Sharp2) and DEC2 (also known as Bhlhb3/Sharp1) are two paralogous basic helix-loop-helix (bHLH) transcriptional regulators which exhibit a robust circadian gene expression pattern in the suprachiasmatic nucleus (SCN) and in peripheral organs. DEC1 has been suggested to play key roles in mammalian cell differentiation, the cell cycle and circadian regulation, hypoxia response, and carcinogenesis. Here we show that DEC1 overexpression exhibits delayed wound healing and reduces cell proliferation, migration, and invasion. DEC1 strongly repressed the promoter activity of cyclin D1. We further identify a possible DEC-response element in the cyclin D1 promoter region, and confirmed the direct binding of DEC1 to that element. Forced expression of DEC1 efficiently repressed the cyclin D1 promoter and expression. Our clinical data provide the first evidence that there is a strong inverse correlation between DEC1 and cyclin D1 expression in oral cancer, and DEC1 expression significantly correlated with clinicopathological parameters. We suggest that radiation-induced DEC1 overexpression and Akt phosphorylation in cancer cells are mediated via PI-3K signalling. Overexpression of DEC1 activates the PI-3K/Akt signalling pathway through reactive oxygen species (ROS).

Study of Alveolar Bone Remodeling Using Deciduous Tooth Stem Cells and Hydroxyapatite by Vascular Endothelial Growth Factor Enhancement and Inhibition of Matrix Metalloproteinase-8 Expression in vivo.

Background: Periodontitis progression is characterized by alveolar bone loss, and its prevention is a major clinical problem in periodontal disease management. Matrix metalloproteinase-8 (MMP-8) has been shown to adequately monitor the treatment of chronic periodontitis patients as gingival crevicular fluid MMP-8s were positively associated with the severity of periodontal disease. Moreover, modulating the vascular endothelial growth factor (VEGF) levels in bones could be a good way to improve bone regeneration and cure periodontitis as VEGF promotes endothelial cell proliferation, proteolytic enzyme release, chemotaxis, and migration; all of which are required for angiogenesis. Purpose: The aim of this study was to determine the effect of hydroxyapatite incorporated with stem cells from exfoliated deciduous teeth (SHED) in Wistar rats' initial alveolar bone remodeling based on the findings of MMP-8 and VEGF expressions. Methods: A hydroxyapatite scaffold (HAS) in conjunction with SHED was transplanted into animal models with alveolar mandibular defects. A total of 10 Wistar rats (Rattus norvegicus) were divided into two groups: HAS and HAS + SHED. Immunohistochemistry staining was performed after 7 days to facilitate the examination of MMP-8 and VEGF expressions. Results: The independent t-test found significant downregulation of MMP-8 and upregulation VEGF expressions in groups transplanted with HAS in conjunction with SHED compared with the HAS group (p < 0.05). Conclusion: The combination of SHED with HAS on alveolar bone defects may contribute to initial alveolar bone remodeling as evident through the assessments of MMP-8 and VEGF expressions.

Anti-apoptotic effect of the basic helix-loop-helix (bHLH) transcription factor DEC2 in human breast cancer cells.

DEC1 (BHLHB2/Stra13/Sharp2) and DEC2 (BHLHB3/Sharp1) are basic helix-loop-helix (bHLH) transcription factors that are involved in circadian rhythms, differentiation and the responses to hypoxia. We examined whether DEC1 and DEC2 are involved in apoptosis regulation, in human breast cancer MCF-7 cells. We found that siRNA-mediated knockdown of DEC2 resulted in marked enhancement of apoptosis compared with that in control cells transfected with nonspecific siRNA. However, knockdown of DEC1 by siRNA did not affect cell survival. Knockdown of DEC2 affected the expression of mRNA or proteins related to apoptosis, such as Fas, c-Myc, caspase-8, poly (ADP-ribose) polymerase (PARP) and Bax. We also showed that tumor necrosis factor-alpha (TNF-alpha) up-regulates the expression of DEC1 and DEC2. DEC2 over-expression caused by the transfection of an expression vector reduced the amounts of cleaved PARP and caspase-8 induced by TNF-alpha treatment, whereas DEC1 over-expression increased it. Finally, we revealed that treatment with double knockdown against both DEC1 and DEC2 decreased the amounts of cleaved PARP and caspase-8 induced by DEC2 siRNA with or without TNF-alpha. These data indicate that DEC2 has an anti-apoptotic effect, whereas DEC1 has a pro-apoptotic effect, which are involved in the balance of survival of human breast cancer MCF-7 cells.

CHRONO and DEC1/DEC2 compensate for lack of CRY1/CRY2 in expression of coherent circadian rhythm but not in generation of circadian oscillation in the neonatal mouse SCN.

Clock genes Cry1 and Cry2, inhibitory components of core molecular feedback loop, are regarded as critical molecules for the circadian rhythm generation in mammals. A double knockout of Cry1 and Cry2 abolishes the circadian behavioral rhythm in adult mice under constant darkness. However, robust circadian rhythms in PER2::LUC expression are detected in the cultured suprachiasmatic nucleus (SCN) of Cry1/Cry2 deficient neonatal mice and restored in adult SCN by co-culture with wild-type neonatal SCN. These findings led us to postulate the compensatory molecule(s) for Cry1/Cry2 deficiency in circadian rhythm generation. We examined the roles of Chrono and Dec1/Dec2 proteins, the suppressors of Per(s) transcription similar to CRY(s). Unexpectedly, knockout of Chrono or Dec1/Dec2 in the Cry1/Cry2 deficient mice did not abolish but decoupled the coherent circadian rhythm into three different periodicities or significantly shortened the circadian period in neonatal SCN. DNA microarray analysis for the SCN of Cry1/Cry2 deficient mice revealed substantial increases in Per(s), Chrono and Dec(s) expression, indicating disinhibition of the transactivation by BMAL1/CLOCK. Here, we conclude that Chrono and Dec1/Dec2 do not compensate for absence of CRY1/CRY2 in the circadian rhythm generation but contribute to the coherent circadian rhythm expression in the neonatal mouse SCN most likely through integration of cellular circadian rhythms.

Liver X receptors (LXRalpha and LXRbeta) are potent regulators for hepatic Dec1 expression.

DEC1 (BHLHB2/Stra13/Sharp2)-a basic helix-loop-helix transcription factor-is known to be involved in various biological phenomena including clock systems and metabolism. In the clock systems, Dec1 expression is dominantly up-regulated by CLOCK : BMAL1 heterodimer, and it exhibits circadian rhythm in the suprachiasmatic nucleus (SCN)-the central circadian pacemaker-and other peripheral tissues. Recent studies have shown that the strong circadian rhythmicity of Dec1 in the SCN was abolished by Clock mutation, whereas that in the liver was affected, but not abolished, by Clock mutation. Moreover, feeding conditions affected hepatic Dec1 expression, which indicates that Dec1 expression is closely linked with the metabolic functions of the liver. Among ligand-activated nuclear receptors examined, LXRalpha and LXRbeta with T0901317-agonist for LXR-were found to be potent enhancers for Dec1 promoter activity, and a higher expression level of LXRalpha protein was detected in the liver than in the kidney and heart. T0901317 increased the levels of endogenous Dec1 transcript in hepatoma cells. Chromatin immunoprecipitation assay indicated that LXRalpha bound to the Dec1 promoter, and an LXRalpha-binding site was identified. These observations indicate that hepatic DEC1 mediates the ligand-dependent LXR signal to regulate the expression of genes involved in the hepatic clock system and metabolism.

Identification of mesenchymal stem cell (MSC)-transcription factors by microarray and knockdown analyses, and signature molecule-marked MSC in bone marrow by immunohistochemistry.

Although ex vivo expanded mesenchymal stem cells (MSC) have been used in numerous studies, the molecular signature and in vivo distribution status of MSC remain unknown. To address this matter, we identified numerous human MSC-characteristic genes--including nine transcription factor genes--using DNA microarray and real-time RT-PCR analyses: Most of the MSC-characteristic genes were down-regulated 24 h after incubation with osteogenesis-, chondrogenesis- or adipogenesis-induction medium, or 48-72 h after knockdown of the nine transcription factors. Furthermore, knockdowns of ETV1, ETV5, FOXP1, GATA6, HMGA2, SIM2 or SOX11 suppressed the self-renewal capacity of MSC, whereas those of FOXP1, SOX11, ETV1, SIM2 or PRDM16 reduced the osteogenic- and/or adipogenic potential. In addition, immunohistochemistry using antibodies for the MSC characteristic molecules--including GATA6, TRPC4, FLG and TGM2--revealed that MSC-like cells were present near the endosteum and in the interior of bone marrow of adult mice. These findings indicate that MSC synthesize a set of MSC markers in vitro and in vivo, and that MSC-characteristic transcription factors are involved in MSC stemness regulation.

The basic helix-loop-helix proteins differentiated embryo chondrocyte (DEC) 1 and DEC2 function as corepressors of retinoid X receptors.

The basic helix-loop-helix proteins differentiated embryo chondrocyte 1 (DEC1) and DEC2 are involved in circadian rhythm control. Because the metabolism of dietary nutrients has been linked to circadian regulation, we examined the effect of DEC1 and DEC2 on the function of the metabolite-sensing nuclear receptors, ligand-dependent transcription factors, including retinoid X receptor (RXR) and liver X receptor (LXR). Transfection assays showed that DEC1 and DEC2 repressed ligand-dependent transactivation by RXR. Knockdown of endogenous DEC1 and DEC2 expression with small interfering RNAs augmented ligand-dependent RXRalpha transactivation. DEC1 and DEC2 interacted directly with RXRalpha, and ligand addition enhanced their association. DEC1 and DEC2 modified interaction of RXRalpha with cofactor proteins. Transfection assays using DEC1 and DEC2 mutants revealed that the C-terminal region of DEC2 is required for repression and that an LXXLL motif in DEC1 and DEC2 is necessary for RXRalpha repression. DEC1 and DEC2 repressed the induction of LXR target genes, associated with the promoter of an LXR target gene, and dissociated from the promoter with ligand treatment. Knockdown of endogenous DEC1 and DEC2 enhanced the LXR target gene expression in hepatocytes. Expression of Dec1, Dec2, and Srebp-1c showed a circadian rhythm in the liver of mice, whereas that of Lxralpha, Lxrbeta, and Rxralpha was not rhythmic. DEC1 and DEC2 also repressed the transactivation of other RXR heterodimers, such as farnesoid X receptor, vitamin D receptor, and retinoic acid receptor. Thus, the repressor function of DEC1 and DEC2 may be extended to other RXR heterodimer nuclear receptors.

Basic-helix-loop-helix (bHLH) transcription factor DEC2 negatively regulates vascular endothelial growth factor expression.

DEC1 (BHLHB2/Sharp2/Stra13) and DEC2 (BHLHB3/Sharp1) are basic-helix-loop-helix (bHLH) transcription factors, involved in cellular differentiation, responses to hypoxia and circadian rhythms. We recently showed that the expression of DEC1 and DEC2 was up-regulated by hypoxia; however, the functions of these two factors under hypoxic conditions have not been elucidated in detail. It is well established that the expression of vascular endothelial growth factor (VEGF) is up-regulated by hypoxia, and the expression of VEGF in response to hypoxia depends on transcriptional activation by a heterodimer comprising hypoxia-inducible factor 1alpha (HIF-1alpha) and arylhydrocarbon receptor nuclear translocator 1 (ARNT1). In the present study, we showed that DEC2, but not DEC1, suppressed VEGF gene expression under hypoxic conditions. DEC2 protein was co-immunoprecipitated with HIF-1alpha but not with ARNT1. The binding of HIF-1alpha to the hypoxia response element (HRE) in the VEGF promoter was decreased by DEC2 over-expression, and increased by DEC2 knockdown. We also showed that the circadian expression of VEGF showed a reciprocal pattern to that of DEC2 in cartilage. DEC2 had a circadian oscillation in implanted Sarcoma 180 cells. We conclude that DEC2 negatively regulates VEGF expression and plays an important role in the pathological conditions in which VEGF is involved.