Mian Wan

microRNA miR-34a Regulates Cytodifferentiation and Targets Multi-signaling Pathways in Human Dental Papilla Cells

Odontogenesis relies on the reciprocal signaling interactions between dental epithelium and neural crest-derived mesenchyme, which is regulated by several signaling pathways. Subtle changes in the activity of these major signaling pathways can have dramatic effects on tooth development. An important regulator of such subtle changes is the fine tuning function of microRNAs (miRNAs). However, the underlying mechanism by which miRNAs regulate tooth development remains elusive. This study determined the expression of miRNAs during cytodifferentiation in the human tooth germ and studied miR-34a as a regulator of dental papilla cell differentiation. Using microarrays, miRNA expression profiles were established at selected times during development (early bell stage or late bell stage) of the human fetal tooth germ. We identified 29 differentially expressed miRNAs from early bell stage/late bell stage comparisons. Out of 6 miRNAs selected for validation by qPCR, all transcripts were confirmed to be differentially expressed. miR-34a was selected for further investigation because it has been previously reported to regulate organogenesis. miR-34a mimics and inhibitors were transfected into human fetal dental papilla cells, mRNA levels of predicted target genes were detected by quantitative real-time PCR, and levels of putative target proteins were examined by western blotting. ALP and DSPP expression were also tested by qPCR, western blotting, and immunofluorescence. Findings from these studies suggested that miR-34a may play important roles in dental papilla cell differentiation during human tooth development by targeting NOTCH and TGF-beta signaling.

MicroRNA 224 Regulates Ion Transporter Expression in Ameloblasts To Coordinate Enamel Mineralization.

ABSTRACT Enamel mineralization is accompanied by the release of protons into the extracellular matrix, which is buffered to regulate the pH value in the local microenvironment. The present study aimed to investigate the role of microRNA 224 (miR-224) as a regulator of SLC4A4 and CFTR, encoding the key buffering ion transporters, in modulating enamel mineralization. miR-224 was significantly downregulated as ameloblasts differentiated, in parallel with upregulation of SLC4A4 and CFTR. Overexpression of miR-224 downregulated SLC4A4 and CFTR expression in cultured human epithelial cells. A microRNA luciferase assay confirmed the specific binding of miR-224 to the 3′ untranslated regions (UTRs) of SLC4A4 and CFTR mRNAs, thereby inhibiting protein translation. miR-224 agomir injection in mouse neonatal incisors resulted in normal enamel length and thickness, but with disturbed organization of the prism structure and deficient crystal growth. Moreover, the enamel Ca/P ratio and microhardness were markedly reduced after miR-224 agomir administration. These results demonstrate that miR-224 plays a pivotal role in fine tuning enamel mineralization by modulating SLC4A4 and CFTR to maintain pH homeostasis and support enamel mineralization.

BMP7 and EREG Contribute to the Inductive Potential of Dental Mesenchyme.

Odontogenesis is accomplished by reciprocal signaling between the epithelial and mesenchymal compartments. It is generally accepted that the inductive mesenchyme is capable of inducing the odontogenic commitment of both dental and non-dental epithelial cells. However, the duration of this signal in the developing dental mesenchyme and whether adult dental pulp tissue maintains its inductive capability remain unclear. This study investigated the contribution of growth factors to regulating the inductive potential of the dental mesenchyme. Human oral epithelial cells (OEs) were co-cultured with either human dental mesenchymal/papilla cells (FDPCs) or human dental pulp cells (ADPCs) under 2-dimensional or 3-dimensional conditions. Odontogenic-associated genes and proteins were detected by qPCR and immunofluorescence, respectively, and significant differences were observed between the two co-culture systems. The BMP7 and EREG expression levels in FDPCs were significantly higher than in ADPCs, as indicated by human growth factor PCR arrays and immunofluorescence analyses. OEs co-cultured with ADPCs supplemented with BMP7 and EREG expressed ameloblastic differentiation genes. Our study suggests that BMP7 and EREG expression in late bell-stage human dental papilla contributes to the inductive potential of dental mesenchyme. Furthermore, adult dental pulp cells supplemented with these two growth factors re-established the inductive potential of postnatal dental pulp tissue.

Spatial signalling mediated by the transforming growth factor-β signalling pathway during tooth formation

Tooth development relies on sequential and reciprocal interactions between the epithelial and mesenchymal tissues, and it is continuously regulated by a variety of conserved and specific temporal-spatial signalling pathways. It is well known that suspensions of tooth germ cells can form tooth-like structures after losing the positional information provided by the epithelial and mesenchymal tissues. However, the particular stage in which the tooth germ cells start to form tooth-like structures after losing their positional information remains unclear. In this study, we investigated the reassociation of tooth germ cells suspension from different morphological stages during tooth development and the phosphorylation of Smad2/3 in this process. Four tooth morphological stages were designed in this study. The results showed that tooth germ cells formed odontogenic tissue at embryonic day (E) 14.5, which is referred to as the cap stage, and they formed tooth-like structures at E16.5, which is referred to as the early bell stage, and E18.5, which is referred to as the late bell stage. Moreover, the transforming growth factor-β signalling pathway might play a role in this process.

Histone Modification in Osteogenic Differentiation of Skeletal Stem Cells

Osteogenic differentiation of skeletal stem cells is an integral part of bone development and homeostasis, and the perturbation of this process is one of the causes to skeletal disease. Understanding of how epigenetic events regulate skeletal stem cell differentiation is therefore of great importance. While the basic epigenetic modifications leading to bone formation are somewhat under explored, a significant amount of research has defined the regulatory roles of histone modifications in osteogenic differentiation. The orchestration of histone modifications is a requirement to establish the epigenetic status which regulates gene transcription during osteogenic differentiation of skeletal stem cells. Here we focus on the roles of histone modification during osteogenic differentiation and review studies that have advanced our knowledge in the field. Before this summary, a brief description is given regarding the up-to-date understanding of the definition of skeletal stem cells and the main mechanisms responsible for histone modifications.

Insights into the Regulation of Yap/Taz from Cellular Systems and Mouse Models.

BACKGROUND The Hippo signaling pathway serves as a main regulator of tissue growth and organ size through the moderation of cell cycle dynamics across many different species. In mammals, the two downstream transcriptional regulators of this pathway are Yes-associated protein (YAP) and transcriptional co-activator with PDZ binding motif (TAZ). Recent studies found in addition to the core Hippo signaling pathway, other signaling pathways can also regulate YAP/TAZ activity, either directly or through crosstalk with the Hippo pathway. OBJECTIVE In this review, we discuss what is known about the regulation of YAP/TAZ from studies conducted using both cell line and mouse models. CONCLUSION To add to the complexity of YAP/TAZ regulation, the activity of YAP/TAZ is also controlled by mechanical and cytoskeletal cues and by multiple extracellular factors.

The Origin and Identification of Mesenchymal Stem Cells in Teeth: from Odontogenic to Non-odontogenic

BACKGROUND Mesenchymal stem cells (MSCs) in teeth have been exploited as vital seed cells for stem cell-based dental medicine. To date, several mesenchymal stem cell populations originated from odontogenic tissue have been isolated and characterized by their expression of MSC surface markers and capacity of multi-lineage differentiation, including dental pulp stem cells (DPSCs), stem cells from human exfoliated deciduous teeth (SHED), stem cells from apical papilla (SCAP) and so on. However, their identity in vivo remains elusive, which hinders further understanding of their application in stem cell-based tooth regeneration. Label retaining and lineage tracing analyses, which serve as gold standards for identification of stem cells in vivo, provide feasibility for identifying MSCs in teeth. OBJECTIVES In this review, we will discuss the issues of MSCs, including the origin and identification of both odontogenic and non-odontogenic MSCs, and address the role of nerve-derived Sonic hedgehog (Shh) in the regulation of MSCs in the neurovascular bundle (NVB). CONCLUSION Based on label retaining and lineage tracing analyses, latest studies have found new populations of non-odontogenic MSCs in teeth, periarterial-derived and glial-derived, regulated by the Shh derived from nerves in the NVB, which provides a new hope for tooth regeneration.

Epigenetic Regulation of Gene Expression in Epithelial Stem Cells Fate

BACKGROUND Epithelial tissues have the ability to self-renew throughout animals life due to the presence of the epithelial stem cells. Except for complicated genes regulation, the fate of epithelial stem cells is also regulated by the epigenetics, including DNA methylation, histone modification and microRNAs, which are emerging as vital elements of epigenetic regulation for epithelial stem cells self-renewal and differentiation. However, the mechanisms underlying these are still poorly understood. OBJECTIVE In this review, we focus on the epigenetic regulation of gene expression in epithelial stem cells fate, using intestinal and epidermal stem cells as models. Meanwhile, a brief description of recent research about the possible impact of network regulation in epithelial stem cell-based amelogenesis by epigenetic regulation is therefore, being discussed. CONCLUSION Epigenetic modification plays a vital role in the epithelial stem cells fate choice through the gene expression. The interaction between epigenetic modification and molecular signaling in epithelial stem cells fate choice still needs further exploration.

Cell culture-based computer-aided design/computer-aided manufacture bio-enamel as novel treatment for enamel defect

Enamel, one of the tissues of the tooth, is the hardest and most highly mineralized substance of the human body. The interwoven architecture of enamel rods contributes to its strength and resistance to fracture. However, these rods still cannot prevent damage to the enamel caused by injury, dental caries, or developmental defects. It is a wellknown fact that enamel is formed by terminally differentiated ameloblasts that are induced by dental mesenchyme signals. Once the formation of enamel is complete and the tooth erupts, the ameloblasts commit apoptosis; furthermore, the enamel compartment is acellular in nature and has no blood/nerve supply to support self-regeneration. Defects in enamel may expose the underlying dentin with dentinal tubules, leading to sensitivity or pain, as well as allowing bacterial penetration, tooth pulp infection, and inflammation. In order to prevent the problem from advancing as well as to protect the tooth and improve cosmetic appearance, acid-etching techniques and application of artificial materials such as resin, amalgam, and other similar use materials are used by dentists. Resin restoration is the most popular method and acid-etching