Defining the cellular origin of seminoma by transcriptional and epigenetic mapping to the normal human germline.

Aberrant male germline development can lead to the formation of seminoma, a testicular germ cell tumor. Seminomas are biologically similar to primordial germ cells (PGCs) and many bear an isochromosome 12p [i(12p)] with two additional copies of the short arm of chromosome 12. By mapping seminoma transcriptomes and open chromatin landscape onto a normal human male germline trajectory, we find that seminoma resembles premigratory/migratory PGCs; however, it exhibits enhanced germline and pluripotency programs and upregulation of genes involved in apoptosis, angiogenesis, and MAPK/ERK pathways. Using pluripotent stem cell-derived PGCs from Pallister-Killian syndrome patients mosaic for i(12p), we model seminoma and identify gene dosage effects that may contribute to transformation. As murine seminoma models do not exist, our analyses provide critical insights into genetic, cellular, and signaling programs driving seminoma transformation, and the in vitro platform developed herein permits evaluation of additional signals required for seminoma tumorigenesis.

Evidence Supporting Oral Hygiene Management by Owners through a Genetic Analysis of Dental Plaque Bacteria in Dogs.

With the increase in the number of households raising dogs and the reports of human-to-dog transmission of oral bacteria, concerns about dogs' oral health and the need for oral hygiene management are increasing. In this study, the owners' perceptions about their dogs' oral health and the frequency of oral hygiene were determined along with the analysis of dog dental plaque bacteria through metagenomic amplicon sequencing so as to support the need for oral hygiene management for dogs. Although the perception of 63.2% of the owners about their dogs' oral health was consistent with the veterinarian's diagnosis, the owners' oral hygiene practices regarding their dogs were very poor. The calculi index (CI) and gingiva index (GI) were lower in dogs who had their teeth brushed more than once a week (57.89%) than in dogs brushed less than once a month (42.10%); however, the difference was nonsignificant (CI: p = 0.479, GI: p = 0.840). Genomic DNA was extracted from dental plaque bacteria removed during dog teeth scaling, and metagenomic amplicons were sequenced. The 16S amplicons of 73 species were identified from among the plaque bacteria of the dogs. These amplicons were of oral disease-causing bacteria in humans and dogs. The 16S amplicon of Streptococcus mutans matched that of the human S. mutans, with type c identified as the main serotype. This result suggests that human oral bacteria can be transmitted to dogs. Therefore, considering the high frequency of contact between dogs and humans because of communal living and the current poor oral health of dogs, owners must improve the oral hygiene management of their dogs.

CRISPR loss of function screening to identify genes involved in human primordial germ cell-like cell development.

Despite our increasing knowledge of molecular mechanisms guiding various aspects of human reproduction, those underlying human primordial germ cell (PGC) development remain largely unknown. Here, we conducted custom CRISPR screening in an in vitro system of human PGC-like cells (hPGCLCs) to identify genes required for acquisition and maintenance of PGC fate. Amongst our candidates, we identified TCL1A, an AKT coactivator. Functional assessment in our in vitro hPGCLCs system revealed that TCL1A played a critical role in later stages of hPGCLC development. Moreover, we found that TCL1A loss reduced AKT-mTOR signaling, downregulated expression of genes related to translational control, and subsequently led to a reduction in global protein synthesis and proliferation. Together, our study highlights the utility of CRISPR screening for human in vitro-derived germ cells and identifies novel translational regulators critical for hPGCLC development.

Metabolic memory of Δ9-tetrahydrocannabinol exposure in pluripotent stem cells and primordial germ cells-like cells.

Cannabis, the most consumed illicit psychoactive drug in the world, is increasingly used by pregnant women. However, while cannabinoid receptors are expressed in the early embryo, the impact of phytocannabinoids exposure on early embryonic processes is lacking. Here, we leverage a stepwise in vitro differentiation system that captures the early embryonic developmental cascade to investigate the impact of exposure to the most abundant phytocannabinoid, Δ9-tetrahydrocannabinol (Δ9-THC). We demonstrate that Δ9-THC increases the proliferation of naive mouse embryonic stem cells (ESCs) but not of their primed counterpart. Surprisingly, this increased proliferation, dependent on the CB1 receptor binding, is only associated with moderate transcriptomic changes. Instead, Δ9-THC capitalizes on ESCs' metabolic bivalence by increasing their glycolytic rates and anabolic capabilities. A memory of this metabolic rewiring is retained throughout differentiation to Primordial Germ Cell-Like Cells in the absence of direct exposure and is associated with an alteration of their transcriptional profile. These results represent the first in-depth molecular characterization of the impact of Δ9-THC exposure on early stages of germline development.

Cannabis is the most widely used illicit drug in the world, with 4.3% of the global adult population estimated to have used it in the previous year. In particular, the consumption of cannabis by pregnant women has almost doubled in recent years and is particularly increased in those aged under 18. The main psychoactive component in cannabis, known as Δ9-THC, activates cannabinoid receptors in the brain, including the receptor CB1. Recent research has shown that CB1 is also active in the mouse embryo, but it remained unclear if Δ9-THC could also affect the development of an embryo. To better understand the potential effects of this exposure, scientists can study stem cells that develop into germ cells (which go on to form egg and sperm), which have been grown in the laboratory. Emerging research has shown that germ cells are particularly sensitive to changes in their environment and due to their role in reproduction, changes can have knock-on effects for embryos. Verdikt et al. studied the effects of Δ9-THC on mouse embryonic stem cells, finding that it caused them to multiply more quickly. This was dependent on both Δ9-THC binding to the CB1 receptor that causes the psychoactive effects of cannabis in the brain and an increased energy metabolism. Blocking an important metabolic pathway called glycolysis caused the Δ9-THC-treated cells to return to a normal multiplication rate. The exposed stem cells also gave rise to germ cells with abnormal metabolism and altered gene expression, suggesting that this metabolic 'memory' can be passed on to cells in the next developmental stage. Overall, the findings indicate that exposure to Δ9-THC alters the metabolism in early embryonic cells of mice and that these effects can be lasting. This emphasises the need for further research on the impact of cannabis use during pregnancy, particularly as the drug's availability is expected to increase significantly with changes in regulation. The work also contributes to research highlighting the inheritance of metabolism.

Alpha-Defensin 1: An Emerging Periodontitis Biomarker.

Background: Research on the development of reliable diagnostic targets is being conducted to overcome the high prevalence and difficulty in managing periodontitis. However, despite the development of various periodontitis target markers, their practical application has been limited due to poor diagnostic accuracy. In this study, we present an improved periodontitis diagnostic target and explore its role in periodontitis. Methods: Gingival crevicular fluid (GCF) was collected from healthy individuals and periodontitis patients, and proteomic analysis was performed. The target marker levels for periodontitis were quantified in GCF samples by enzyme-linked immunosorbent assay (ELISA). Mouse bone marrow-derived macrophages (BMMs) were used for the osteoclast formation assay. Results: LC-MS/MS analysis of whole GCF showed that the level of alpha-defensin 1 (DEFA-1) was higher in periodontitis GCF than in healthy GCF. The comparison of periodontitis target proteins galactin-10, ODAM, and azurocidin proposed in other studies found that the difference in DEFA-1 levels was the largest between healthy and periodontitis GCF, and periodontitis was more effectively distinguished. The differentiation of RANKL-induced BMMs into osteoclasts was significantly reduced by recombinant DEFA-1 (rDEFA-1). Conclusions: These results suggest the regulatory role of DEFA-1 in the periodontitis process and the relevance of DEFA-1 as a diagnostic target for periodontitis.

Reconstitution of Human Prospermatogonial Development from Human-Induced Pluripotent Stem Cells.

There is a scarcity of information regarding the molecular mechanisms underlying human germ cell development due to limitations in obtaining the relevant materials. Reconstitution of human germ cell development from pluripotent stem cells in vitro would provide critical insight into the etiology of various reproductive conditions and disorders, including infertility.Recently, we reported the in vitro reconstitution of human prospermatogonial development from human-induced pluripotent stem cells through human primordial germ cell (PGC)-like cells (hPGCLCs) using long-term cultured xenogeneic reconstituted testes. Here, we describe a method to generate M-prospermatogonia-like cells (MLCs) and T1-prospermatogonia-like cells (T1LCs), which closely resemble M- and T1-prospermatogonia present in second-trimester human fetal testes in vivo.

Metabolic memory of Δ9-tetrahydrocannabinol exposure in pluripotent stem cells and primordial germ cells-like cells.

Cannabis, the most consumed illicit psychoactive drug in the world, is increasingly used by pregnant women. However, while cannabinoid receptors are expressed in the early embryo, the impact of phytocannabinoids exposure on early embryonic processes is lacking. Here, we leverage a stepwise in vitro differentiation system that captures early embryonic developmental cascade to investigate the impact of exposure to the most abundant phytocannabinoid, Δ9-tetrahydrocannabinol (Δ9-THC). We demonstrate that Δ9-THC increases the proliferation of naïve mouse embryonic stem cells (ESCs) but not of their primed counterpart. Surprisingly, this increased proliferation, dependent on the CB1 receptor binding, is only associated with moderate transcriptomic changes. Instead, Δ9-THC capitalizes on ESCs' metabolic bivalence by increasing their glycolytic rates and anabolic capabilities. A memory of this metabolic rewiring is retained throughout differentiation to Primordial Germ Cell-Like Cells in the absence of direct exposure and is associated with an alteration of their transcriptional profile. These results represent the first in-depth molecular characterization of the impact of Δ9-THC exposure on early stages of germline development.

Role of cathepsin D induced by Porphyromonas gingivalis lipopolysaccharide in periodontitis.

Periodontitis is an inflammatory disease of tooth-supporting tissues caused by oral bacteria. Periodontal ligament loss and alveolar bone destruction occur in progressive periodontitis. Since gingival crevicular fluids (GCF) reflects the inflammatory environment of the periodontal pocket, it is a very important specimen for developing targets for periodontitis diagnosis. An antibody array was performed using GCF collected from healthy participants and patients with periodontitis to identify the proteolytic enzymes involved in periodontitis. Of 21 targets on the antibody array membrane, kallikrein 6 (KLK6), kallikrein 10 (KLK10), cathepsin A (CathA), and cathepsin D (CathD) showed higher levels in periodontitis GCF than in GCF from healthy participants. Lipopolysaccharide stimulation of Porphyromonas gingivalis (PG-LPS) in immortalized gingival fibroblasts only increased CathD protein levels among the four targets. The substrate cleavage activity of CathD was increased in PG-LPS-treated immortalized gingival fibroblast extract. The PG-LPS-induced substrate cleavage effect was abolished by the CathD inhibitor pepstatin A. Osteoclast formation was promoted by treatment with conditioned media from PG-LPS- treated immortalized gingival fibroblasts but inhibited by the CathD inhibitor pepstatin A. These results suggest that PG-LPS affected the osteoclast formation process by increasing CathD expression in cells around the alveolar bone, thereby participating in periodontitis progression.

Effect of self-learning media based on 360° virtual reality for learning periodontal instrument skills.

INTRODUCTION: Of 360° Virtual Reality (VR) is possibly produced and sufficiently effective as a consumer-friendly VR learning medium. Therefore, it is also expected to be useful in the dental practice field, as a self-learning medium for non-face-to-face skill training during the ongoing pandemic (COVID-19). Accordingly, this study was conducted to assess 360° VR self-learning media for a periodontal instrument operation. MATERIALS AND METHODS: We recruited 30 participants who had never experienced instrument training. We offered basic education and initial assessment (IA), then divided them into three groups: 1) PAPER: trained only with paper handouts; 2) 2D: trained with 2D video; 3) VR: trained with 360° VR. Each group performed self-learning and mid-term assessment (MA). Subjects then implemented home self-learning with the same media for one week, which was then followed by a final assessment (FA). RESULT: Analysis of IA-to-FA improvement scores showed that VR and 2D video were significantly higher than the PAPER groups. Meanwhile, analysis of MA-to-FA improvement scores showed that only VR was substantially higher than the PAPER group. Although VR and 2D video groups were not considerably different, VR scores were numerically higher than 2D video in all improvement score analyses. DISCUSSION: Both 2D video and 360° VR training were helpful to participants for an effective self-learning and also had good portability and accessibility as online-based learning methods. 360° VR showed higher learning efficiency than regular 2D video, possibly due to its autonomy, 360° visual information and physical and immersive characteristics, which positively affected self-training. CONCLUSION: Our findings showed the potential of 360° VR learning media and, further, suggest its usefulness as a novel self-learning method in future dental education.

Peroxiredoxin 5 is involved in cancer cell invasion and tumor growth of oral squamous cell carcinoma.

OBJECTIVES: Peroxiredoxins (Prxs) are antioxidant enzymes that can coordinate cell signal transduction via reactive species scavenging or by acting as redox sensors. The mechanism by which Prxs promote cancer invasion and progression is not yet fully understood. This study aims to elucidate the precise mechanism through which Prx type 5 (Prx5) promotes cancer invasion and tumor growth. MATERIALS AND METHODS: We analyzed the Prx5 expression in oral squamous cell carcinoma (OSCC) by using microarray analysis for gene expression profiling. To identify Prx5 function in cancer, lentiviral short hairpin RNA was used for Prx5 depletion, and invasion assay and mouse xenograft were performed. RESULTS: In microarray data obtained from OSCC patients, Prx5 showed higher expression at the tumor margin (TM) compared to the tumor center (TC) of the collective invasion. The depletion of Prx5 in OSCC cells (Prx5dep ) led to decreased invasion activity. In orthotopic xenograft models, Prx5dep cells harbored delimited tumorigenicity compared to wild-type cells as well as the suppression of lymph node metastasis. Prx5dep cells showed growth retardation and increased cellular reactive oxygen species (ROS) levels. The growth retardation of Prx5dep cells resulted in G1 phase arrest. CONCLUSIONS: This study provides evidence that Prx5 removes excess ROS, especially in the TM, contributing to cancer invasion and tumor progression.

Evaluation of the suitability of Gardenia blue pigment derived from Gardenia jasminoides Ellis (Rubiaceae) as a dental plaque disclosant.

OBJECTIVE: Dental disclosants are used to distinguish the amount and location of dental plaque. Therefore, dental disclosants are useful for dental plaque management and effective in motivating oral care. After reports on the cytotoxicity and carcinogenesis of dental disclosants containing erythrosine, many natural pigments for dental disclosants have been suggested. However, there are insufficient ingredients with proven biocompatibility for human subjects. The purpose of this study was to explore the suitability of Gardenia blue pigment as a dental disclosant. MATERIALS AND METHODS: Natural Gardenia blue pigment was used as the dental disclosant experimental group and 2Tone was used as the control group. The homogeneity of the panelists in the groups was identified by measuring the gingivitis index and dental plaque index of the subjects before the experiments. The degree of pigmentation on the tooth surface was observed immediately after coloring and after 1 h. The remaining pigment on the dental surface was also monitored after brushing the teeth. In the panelist test, the taste and sensation of the pigment were examined, and the overall preference for the pigment as a dental disclosant was examined. RESULTS: After coloration of the tooth surface, neither the natural Gardenia blue pigment nor 2Tone imparted any special taste or sensation. The coloration of dental plaque with Gardenia blue pigment was similar to that of 2Tone, and the difference in the degree of coloration between Gardenia blue pigment and 2Tone was not statistically significant. The residual degree of pigmentation after 1 h of coloring was similar in both groups, but most of it was removed by brushing. There was no statistically significant difference in the overall preference of Gardenia blue pigment over 2Tone. CONCLUSIONS: The results of this study prove that natural Gardenia blue pigment could be a suitable dental disclosant in terms of pigmentation and preference.

The developmental origin and the specification of the adrenal cortex in humans and cynomolgus monkeys.

Development of the adrenal cortex, a vital endocrine organ, originates in the adrenogonadal primordium, a common progenitor for both the adrenocortical and gonadal lineages in rodents. In contrast, we find that in humans and cynomolgus monkeys, the adrenocortical lineage originates in a temporally and spatially distinct fashion from the gonadal lineage, arising earlier and more anteriorly within the coelomic epithelium. The adrenal primordium arises from adrenogenic coelomic epithelium via an epithelial-to-mesenchymal transition, which then progresses into the steroidogenic fetal zone via both direct and indirect routes. Notably, we find that adrenocortical and gonadal lineages exhibit distinct HOX codes, suggesting distinct anterior-posterior regionalization. Together, our assessment of the early divergence of these lineages provides a molecular framework for understanding human adrenal and gonadal disorders.

Expanding homogeneous culture of human primordial germ cell-like cells maintaining germline features without serum or feeder layers.

In vitro expansion of human primordial germ cell-like cells (hPGCLCs), a pluripotent stem cell-derived PGC model, has proved challenging due to rapid loss of primordial germ cell (PGC)-like identity and limited cell survival/proliferation. Here, we describe long-term culture hPGCLCs (LTC-hPGCLCs), which actively proliferate in a serum-free, feeder-free condition without apparent limit as highly homogeneous diploid cell populations maintaining transcriptomic and epigenomic characteristics of hPGCLCs. Histone proteomics confirmed reduced H3K9me2 and increased H3K27me3 marks in LTC-hPGCLCs compared with induced pluripotent stem cells (iPSCs). LTC-hPGCLCs established from multiple human iPSC clones of both sexes were telomerase positive, senescence-free cells readily passaged with minimal cell death or deviation from the PGC-like identity. LTC-hPGCLCs are capable of differentiating to DAZL-positive M-spermatogonia-like cells in the xenogeneic reconstituted testis (xrTestis) organ culture milieu as well as efficiently producing fully pluripotent embryonic germ cell-like cells in the presence of stem cell factor and fibroblast growth factor 2. Thus, LTC-hPGCLCs provide convenient access to unlimited amounts of high-quality and homogeneous hPGCLCs.

Antibacterial and antioxidant effect of ethanol extracts of Terminalia chebula on Streptococcus mutans.

OBJECTIVE: Dental caries is a high prevalent chronic bacterial infectious disease caused by plaque, a bacterial colony deposited on tooth surfaces and gum tissues. Streptococcus mutans is a primary cariogenic bacterium commonly found in the human oral cavity. Oral hygiene products containing antibacterial ingredients can be helpful in caries management. In this study, we investigated the anticaries mechanism of the ethanol extract of Terminalia chebula (EETC) on S. mutans and suggest its possible application as a functional ingredients for oral hygiene products. MATERIALS AND METHODS: The EETC was prepared from the Terminalia chebula fruit. Disk diffusion, minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and colony forming unit (CFU) were analyzed to observe the antibacterial activity of EETC. The glucan formation was measured using the filtrate of bacterial culture medium and sucrose. Gene expression was analyzed using RT-PCR. Cytotoxicity was analyzed using the MTT assay. The radical-scavenging activities of DPPH and ABTS were also tested to verify the antioxidant activity of EETC. RESULTS: The antibacterial activity of the EETC was explored through a disc diffusion analysis and CFU measurement. EETC treatment decreased insoluble glucan formation and gene expression of glycosyltransferase B (gtf B), glycosyltransferase C (gtf C), glycosyltransferase D (gtf D), and fructosyltransferase (ftf). The MIC and MBC of EETC on S. mutans were not cytotoxic to gingival fibroblasts. In addition, we observed DPPH and ABTS-radical scavenging activities of EETC. CONCLUSIONS: These results indicate that the antibacterial and antioxidant effects of EETC may contribute to oral hygiene products for dental caries management.

Anti-Periodontitis Effect of Ethanol Extracts of Alpinia Katsumadai Seeds.

Oral microbes are intimately associated with many oral and systemic diseases. Ongoing research is seeking to elucidate drugs that prevent and treat microbial diseases. Various functions of Alpinia Katsumadai seed extracts have been reported such as their anti-viral, anti-oxidant, anti-inflammatory, anti-puritic, anti-emetic, and cytoprotective effects. Here, we investigated the anti-periodontitis effect of an ethanol extract of Alpinia Katsumadai seeds (EEAKSs) on dental plaque bacteria (DPB)-induced inflammation and bone resorption. DPB and Porphyromonas gingivalis (P. gingivalis) were cultured and lipopolysaccharide (LPS) was extracted. Prostaglandin E2 (PGE2) and cyclooxygenase 2 (COX-2) levels were estimated using ELISA. Cytotoxicity was also verified. Proteases were screened using a protease antibody array method. Osteoclastic bone resorption was also investigated. EEAKSs suppressed P. gingivalis growth on agar plates. LPS prepared from dental plaque bacteria (DPB-LPS) and P. gingivalis (PG-LPS) significantly increased PGE2 and COX2 levels in immortalized gingival fibroblasts (IGFs), immortalized human oral keratinocytes (IHOKs), and RAW264.7 macrophage cells. However, DPB-LPS and PG-LPS-induced PGE2 and COX-2 increases were effectively abolished by EEAKS treatment at non-cytotoxic concentrations. In the protease antibody array, matrix metalloproteinase (MMP)-2, MMP-3, MMP-7, kallikrein 10, cathepsin D, and cathepsin V levels were increased by PG-LPS stimulation. However, increases in protease levels except for cathepsin D were suppressed by EEAKS treatment. In addition, RANKL-induced osteoclast differentiation was significantly inhibited by EEAKS treatment, leading to reductions in resorption pit formation. These results suggest that EEAKSs exerted a beneficial oral health effect to help prevent DPB-mediated periodontal disease.

Identification of galectin­10 as a biomarker for periodontitis based on proteomic analysis of gingival crevicular fluid.

Periodontitis is a chronic inflammatory disease caused by the gradual breakdown of tissues surrounding the teeth due to various factors. The disease has been frequently noted in dental outpatients for a number of years. Improvements are required to current diagnostic methods, which have limitations in assessing the condition and progression of periodontitis. The development of diagnostic biomarkers for periodontitis to increase the sensitivity and accuracy of diagnosis is important for the management of periodontitis. In the present study, whole gingival crevicular fluid (GCF) from patients with periodontitis and healthy individuals was characterized via liquid chromatography with tandem mass spectrometry. Label­free quantification was used to identify the differentially abundant protein biomarkers. A total of 1,295 proteins were identified from the whole GCF of patients with periodontitis and healthy individuals via proteomic analysis. When analyzing biological processes, 'metabolic process' and 'cell organization and biogenes' were identified to play important roles in GCF under periodontitis conditions according to Gene Ontology. When analyzing molecular functions, 'catalytic activity' and 'protein binding' were the terms most enriched with differentially abundant proteins under periodontitis conditions. Galectin­10 (Gal­10) was one of the most upregulated proteins in the GCF of patients with periodontitis. The levels of prostaglandin E2 were increased in oral keratinocytes and gingival fibroblasts treated with recombinant (r)Gal­10. The levels of interleukin­8, matrix metalloproteinase 9 and C­reactive protein were increased in the conditioned media (CM) of rGal­10­treated gingival fibroblasts. In addition, the CM of rGal­10­treated gingival fibroblasts induced osteoclast differentiation. These results suggested that Gal­10 expression was increased in the GCF of patients with periodontitis and contributed to the process of osteoclastogenesis. Therefore, Gal­10 may be a candidate biomarker for periodontitis.

Reconstitution of prospermatogonial specification in vitro from human induced pluripotent stem cells.

Establishment of spermatogonia throughout the fetal and postnatal period is essential for production of spermatozoa and male fertility. Here, we establish a protocol for in vitro reconstitution of human prospermatogonial specification whereby human primordial germ cell (PGC)-like cells differentiated from human induced pluripotent stem cells are further induced into M-prospermatogonia-like cells and T1 prospermatogonia-like cells (T1LCs) using long-term cultured xenogeneic reconstituted testes. Single cell RNA-sequencing is used to delineate the lineage trajectory leading to T1LCs, which closely resemble human T1-prospermatogonia in vivo and exhibit gene expression related to spermatogenesis and diminished proliferation, a hallmark of quiescent T1 prospermatogonia. Notably, this system enables us to visualize the dynamic and stage-specific regulation of transposable elements during human prospermatogonial specification. Together, our findings pave the way for understanding and reconstructing human male germline development in vitro.

Cytotoxicity of dental disclosing solution on gingival epithelial cells in vitro.

OBJECTIVE: Coloring dental biofilm and plaque with a dental disclosing solution is visually effective in dental treatment and oral hygiene education. Despite continuous reports of the risk of the product ingredients, dental disclosing solution are widely used in dentistry. However, the cytotoxic mechanism of dental disclosing solution is not known. Here we elucidated the tissue dyeing range and investigated the cytotoxic mechanism of dental disclosing solution. MATERIALS AND METHODS: Gingival epithelial cells and mouse head and neck tissue were stained with dental disclosing solution. Changes in the cell cycle distribution by the dental disclosing solution treatment were analyzed. A deoxynucleotidyl transferase dUTP nick and labeling (TUNEL) assay was performed to examine the apoptotic features of the gingival epithelial cells. RESULTS: Dental disclosing solution stained the chromosome strongly, as well as both the hard and soft tissue of the mouse head and neck. The results of flow cytometric analysis and TUNEL analyses revealed that the cytotoxicity associated with dental disclosing solution was related to the induction of apoptosis. However, the staining of porcine skin by dental disclosing solution was not easily removed, even with a wide range of pH solutions. CONCLUSIONS: These results suggest that dental disclosing solution had strong cytotoxicity and safer alternatives are needed.

Zygotic genome activation in the chicken: a comparative review.

Maternal RNAs and proteins in the oocyte contribute to early embryonic development. After fertilization, these maternal factors are cleared and embryonic development is determined by an individual's own RNAs and proteins, in a process called the maternal-to-zygotic transition. Zygotic transcription is initially inactive, but is eventually activated by maternal transcription factors. The timing and molecular mechanisms involved in zygotic genome activation (ZGA) have been well-described in many species. Among birds, a transcriptome-based understanding of ZGA has only been explored in chickens by RNA sequencing of intrauterine embryos. RNA sequencing of chicken intrauterine embryos, including oocytes, zygotes, and Eyal-Giladi and Kochav (EGK) stages I-X has enabled the identification of differentially expressed genes between consecutive stages. These studies have revealed that there are two waves of ZGA: a minor wave at the one-cell stage (shortly after fertilization) and a major wave between EGK.III and EGK.VI (during cellularization). In the chicken, the maternal genome is activated during minor ZGA and the paternal genome is quiescent until major ZGA to avoid transcription from supernumerary sperm nuclei. In this review, we provide a detailed overview of events in intrauterine embryonic development in birds (and particularly in chickens), as well as a transcriptome-based analysis of ZGA.

Cancer-associated fibroblast stimulates cancer cell invasion in an interleukin-1 receptor (IL-1R)-dependent manner.

Tumor microenvironment serves an important role in tumor growth and metastasis. Cancer cells can promote growth and malignancy by altering the surrounding stroma. Cancer-associated fibroblast (CAF) are an abundant cell type present within the tumor microenvironment and provide tumorigenic features by secreting cytokines. In the current study, the CAF-mediated invasion of oral squamous cell carcinoma (OSCC) was investigated and the associated mechanisms were elucidated. Cancer invasion was estimated using a Matrigel-coated Transwell chamber and FITC-gelatin matrix. To verify the effect of the tumor microenvironment, conditioned media (CM) from normal fibroblast (NF) and CAFs were prepared. An ELISA was performed to estimate the level of IL-1ß. A proteome profiler human protease array was performed to verify the proteases affected by stimulation with CM, from CAF. Recombinant IL-1ß protein increased the invasion of OSCC cells. IL-1ß expression was higher in CAF than NF. CM from CAF (CM-CAF) increased cancer invasion and FITC-gelatin matrix degradation. The invasive capacity provided by CAF was abrogated by an IL-1 receptor (IL-1R) antagonist. Additionally, CM-CAF increased the secretion of ADAM 9 and Kallikrein 11 from OSCC cells. The invasion activity by CM-CAF was partially abrogated by the neutralization of ADAM 9 or Kallikrein 11. In conclusion, by providing stromal factor, CAFs were a critical inducer of OSCC invasion, and CAF secretes the required amount of IL-1ß to increase cancer invasion activity. The invasive capacity of CAF was identified to be IL-1R-dependent. ADAM 9 and Kallikrein 11 were influencing factors involved in the increase of CAF-mediated cancer invasion.