Project description:miRNAs are not well known their expression and function in tooth development. To identify the miRNAs expression during tooth development, tooth germs were dissected from the initiation bud, cap and bell stages. miRNA-chip expression analysis was performed with RNAs of the molar tooth germs from embryos of pregnant mice at emrbryonic day 11, 12, 14, and 16, using Agilent's miRNA microarray.
Project description:To identify genes heretofore undiscovered as critical players in the biogenesis of teeth, we have used microarray gene expression analysis of the developing mouse molar tooth (DMT) between 1 and 10 days postnatal to identify genes differentially expressed when compared to 16 control tissues (GEO accession # GSE1986). Of the top 100 genes exhibiting increased expression in the DMT, 29 were found to have been previously associated with tooth development. Differential expression of the remaining 71 genes not previously associated with tooth development was confirmed by qRT-PCR analysis. Further analysis of seven of the latter genes by mRNA in situ hybridization found that five were specific to the developing tooth in the craniofacial region (Rspo4, Papln, Amtn, Gja1, Maf). Of the remaining two, one was found to be more widely expressed (Sp7) and the other was found to be specific to the nasal serous gland, which is close to, but distinct from, the developing tooth (Vrm). Experiment Overall Design: mRNA from molar teeth extracted from Swiss Webster mouse pups between 1 and 10 days post-natal was pooled, labeled, and hybridized in quadruplicate to Affymetrix Mouse Genome Expression 430 2.0 microarrays. This data was compared to that of 16 control tissues (GEO accession # GSE1986) to identify genes differentially expressed in the DMT mRNA.
Project description:Organoid models provide powerful tools to study tissue biology and development in a dish. Here, we established first-time organoid models from early-postnatal (postnatal day 7) mouse molar and incisor, capable of differentiation toward ameloblast-like cells in vitro. To more in detail characterise organoids from mouse molar and incisor, bulk RNA-sequencing was performed on the following (1) early passage (passage 0) organoids from both tooth types grown in basal tooth organoid medium (TOM) with or without addition of exogenous epidermal growth factor (EGF); and (2) late passage (passage 5) organoids grown in TOM+EGF or differentiation medium (DM).
Project description:Dioxins are ubiquitous environmental poisons that are developmentally toxic. Exposure of mouse E14 tooth germs to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) leads to reduced tooth size and deformation of cuspal morphology implying the induction of a variety of biological responses on both cellular and molecular levels. To verify such responses at the gene level, mouse embryonic cap staged tooth germs were cultured for 24 h with/without 1 µM TCDD. Keywords: tissue culture, exposure to TCDD, tooth development
Project description:Throughout the various stages of tooth development, reciprocal epithelial-mesenchymal interactions are the driving force, for instance crucially involved in the differentiation of mature enamel-forming ameloblasts and dentin-producing odontoblasts. Here we established mouse tooth ‘assembloids’, comprised of tooth organoid-derived dental epithelial cells (from mouse molars and incisors) cultured together with molar dental pulp stem cells (DPSCs), to mimic these developmental interactions. Assembloids from both tooth types were grown both in basal- and differentiation-inducing conditions. Single cell transcriptomics analysis was applied to in detail characterize and validate the newly developed mouse tooth assembloid model and evaluate the induced differentiation processes.
Project description:Foxi3 is a transcription factor expressed in the epithelium during tooth development. In this study we used a microarray analysis to indentify differentially expressed genes in bud stage (E13.5 ) mandibular molar epithelium of conditional Foxi3 knock-out embryos compared to control littermates.
Project description:In this study, using mouse molar as the model, we developed a dual fluorescence reporter mouse to precisely track and analyze dental epithelium and mesenchyme at single-cell resolution from early embryonic to postnatal stages. Moreover, we constructed the virtual molar explorer (VMEx) to spatially map 15,967 molar-expressed genes and identified that Msx1+ Sdc1+ marked the developing dental papilla while surrounded by Msx1+ Sdc1- molar niche. Through tooth germ reconstitution and organoid culture in vitro and kidney capsule transplantation in vivo, we provided evidence that the Msx1+ Sdc1- dental follicle cells might function as the tooth organizers that promoted epithelium survival and tooth germ organization. Furthermore, the appearance of Msx1+ Sdc1+ dental papilla cells relied on the interaction between dental epithelium and Msx1+ Sdc1- dental follicle cells. Together, our results revealed the cellular dynamics of tooth development in mice and identified that the dental follicle might be the key driver of epithelial-mesenchymal interaction and tooth morphogenesis.
Project description:Sequenced samples are cultured posterior parts of the first mouse molar tooth primordia. RNA sequencing was performed based on explants after 0, 16 or 24 hours of in vitro culture respectively, with aim to define candidate genes playing a role in the tooth germ development.
Project description:The tooth is derived from both the ectoderm and the neural crest (ectomesenchyme). It is often used as a model for studies of the basic mechanisms of organ development, including differentiation, cellular interaction, morphogenesis, and mineralization of extracellular matrices. In mouse, tooth development begins at embryonic day 11.5 (E11.5) by thickening of the dental epithelium, whereas mineralization of enamel and dentin in first molars starts at postnatal day 0 (P0). Tooth development entails a multistep and complex process of gene expression. Studies with genetically modified mice have contributed substantially to our understanding of genes that regulate tooth development and morphology. MicroRNAs (miRNA) are a class of non-coding RNAs that regulate gene expression at a post-transcriptional level, and are considered as important regulatory molecules during foetal development. By binding to target mRNAs, miRNA induce mRNA decay or translation repression. Recent bioinformatic predictions of miRNA targets in vertebrates indicate that hundreds of miRNAs are responsible for regulation of expression of up to 30% of the human protein-coding genes. Recent experiments has verified that miRNAs can regulate expression of hundreds of mRNAs in cultured cells. Previous study has also shown that microarrays can be used to detect physiologically relevant effects of miRNAs on expression of mRNAs in vertebrates; miRNAs have been shown to alter expression of a greater number of transcripts than previously appreciated, primarily through interaction with 3M-4UTRs. Understanding the biological function of miRNAs require knowledge of their mRNA targets. Bioinformatic approaches have been used to predict mRNA targets, among which both transcription factors and pro-apoptopic genes were prominent candidates. The precise molecular function of miRNAs remains largely unknown and a better understanding may require loss-of-function studies in vivo. It has been shown that intravenous administration of a novel class of chemically engineered oligonucleotides termed M-^SantagomirsM-^T resulted in a marked reduction of corresponding miRNA levels in most tissues. This resulted in upregulated expression of hundreds of genes predicted to be repressed by miR-122 because these genes had a miR-122 recognition motif in their 3M-4UTR region. Paradoxically, antagomir treatment also revealed a significant number of downregulated genes that may be activated by miR-122. The mechanism by which miRNAs may activate gene expression in vivo is unknown. However, it may involve an indirect effect (the suppression of a transcriptional repressor), or alternatively, miRNA may have a direct effect on gene activation (e.g. chromatin remodeling). Studies of microRNA expression profiles of the developing murine molar tooth germ have shown these to be highly dynamic, some miRNAs being abundantly expressed during the early development of the molar tooth germ, while others are highly expressed only at later developmental stages. A recent study also suggested that miRNAs modulate tooth morphogenesis and ameloblast differentiation, perhaps largely by fine tuning conserved signaling networks. The function of microRNA-214 (miR-214) is unknown, however, it has been related to normal growth and skeletal development in mice. In tooth germs miR-214 is more highly expressed during later developmental stages. It was selected as a target for silencing using anti-miR-214. Here we report effects from injections of 1-100 pmol of anti-miR-214 close to developing first mandibular molar in newborn mice. At 24 hrs, or more, after injection significant changes in gene expression, and in levels of several correspondingly encoded proteins, were found in the tooth germ.