In the present study, we examined the bone healing capacity of
, a homeobox gene that plays essential roles in the differentiation of a range of developing tissues, and identified its putative ...function in palatogenesis. We applied the knocking down of
in human periodontal ligament fibroblasts to examine the osteogenic potential of
. Additionally, we applied in vivo periodontitis induced experiment to reveal the possible application of
knockdown for 1 and 2 weeks in bone healing processes. We examined the detailed histomorphological changes using Masson's trichrome staining and micro-computed tomography evaluation. Moreover, we observed the localization patterns of various signaling molecules, including α-SMA, CK14, IL-1β, and MPO to examine the altered bone healing processes. Furthermore, we investigated the process of bone formation using immunohistochemistry of Osteocalcin and Runx2. On the basis of the results, we suggest that the knocking down of
via the activation of osteoblast and modulation of inflammation would be a plausible answer for bone regeneration as a gene therapy. Additionally, we propose that the purpose-dependent selection and application of developmental regulation genes are important for the functional regeneration of specific tissues and organs, where the pathological condition of tooth loss lesion would be.
Lipid biosynthesis is recently studied its functions in a range of cellular physiology including differentiation and regeneration. However, it still remains to be elucidated in its precise function. ...To reveal this, we evaluated the roles of lysophosphatidic acid (LPA) signaling in alveolar bone formation using the LPA type 2 receptor (LPAR2) antagonist AMG‐35 (Amgen Compound 35) using tooth loss without periodontal disease model which would be caused by trauma and usually requires a dental implant to restore masticatory function. In this study, in vitro cell culture experiments in osteoblasts and periodontal ligament fibroblasts revealed cell type‐specific responses, with AMG‐35 modulating osteogenic differentiation in osteoblasts in vitro. To confirm the in vivo results, we employed a mouse model of tooth loss without periodontal disease. Five to 10 days after tooth extraction, AMG‐35 facilitated bone formation in the tooth root socket as measured by immunohistochemistry for differentiation markers KI67, Osteocalcin, Periostin, RUNX2, transforming growth factor beta 1 (TGF‐β1) and SMAD2/3. The increased expression and the localization of these proteins suggest that AMG‐35 elicits osteoblast differentiation through TGF‐β1 and SMAD2/3 signaling. These results indicate that LPAR2/TGF‐β1/SMAD2/3 represents a new signaling pathway in alveolar bone formation and that local application of AMG‐35 in traumatic tooth loss can be used to facilitate bone regeneration and healing for further clinical treatment.
Periodontitis is an excessive inflammatory event in tooth-supporting tissues and can cause tooth loss. We used erythropoietin (EPO), which has been reported to play an important role in bone healing ...and modulation of angiogenesis, as a therapeutic agent
in vivo
and
in vitro
experimental models to analyze its effect on periodontitis. First
,
EPO was applied to
in vitro
MC3T3-E1 cells and human periodontal ligament fibroblast (hPDLF) cells to examine its function in altered cellular events and gene expression patterns.
In vitro
cultivation of MC3T3-E1 and hPDLF cells with 10 IU/ml EPO at 24 and 48 h showed an obvious increase in cell proliferation. Interestingly, EPO treatment altered the expression of osteogenesis-related molecules, including alkaline phosphatase (ALP), bone morphogenetic protein-2 (BMP-2), and osteocalcin (OC) in MC3T3-E1 cells but not in hPDLF cells. In particular, MC3T3-E1 cells showed increased expression of ALP, BMP-2, and OC on day 5, while hPDLF cells showed increased expression of BMP-2, and OC on day 14. Based on the
in vitro
examination, we evaluated the effect of EPO on bone formation using an experimentally-induced animal periodontitis model. After the induction of periodontitis in the maxillary left second M, 10 IU/ml of EPO was locally applied to the extraction tooth sockets. Histomorphological examination using Masson’s trichrome (MTC) staining showed facilitated bone formation in the EPO-treated groups after 14 days. Similarly, stronger positive reactions against vascular endothelial growth factor (VEGF), cluster of differentiation 31 (CD31), runt-related transcription factor 2 (RUNX2), and osteocalcin (OC) were detected in the EPO-treated group compared to the control. Meanwhile, myeloperoxidase, an inflammatory marker, was decreased in the EPO-treated group on days 1 and 5. Overall, EPO facilitates bone healing and regeneration through altered signaling regulation and modulation of inflammation in the osteoblast cell lineage and to a lesser extent in hPDLF cells.
MicroRNAs (miRNAs) are a class of naturally occurring small non-coding RNAs that post-transcriptionally regulate gene expression in organisms. Most mammalian miRNAs influence biological processes, ...including developmental changes, tissue morphogenesis and the maintenance of tissue identity, cell growth, differentiation, apoptosis, and metabolism. The
has been correlated with cancer; however, developmental roles of this miRNA are unclear. In this study, we examined the expression pattern and evaluated the developmental regulation of
during tooth morphogenesis using ex-vivo culture method. The expression pattern of
was examined in the epithelium and mesenchyme of developing tooth germ with stage-specific manners. Perturbation of the expression of
clearly altered expression patterns of dental-development-related signaling molecules, including
and
. The gene expression complemented with change in cellular events including, apoptosis and proliferation which caused altered crown and pulp morphogenesis in renal-capsule-calcified teeth. Especially, mislocalization of β-Catenin and SMAD1/5/8 were observed alongside dramatic alterations in the expression patterns of
and
. Overall, our data suggest that the
regulate the cellular physiology during tooth morphogenesis through modulation of the Wnt, Bmp, Fgf, and Shh signaling pathways to form proper tooth pulp and crown.
During tooth development, proper protein folding and trafficking are significant processes as newly synthesized proteins proceed to form designated tissues. Endoplasmic reticulum (ER) stress occurs ...inevitably in tooth development as unfolded and misfolded proteins accumulate in ER. 4-Phenylbutyric acid (4PBA) is a FDA approved drug and known as a chemical chaperone which alleviates the ER stress. Recently, several studies showed that 4PBA performs therapeutic effects in some genetic diseases due to misfolding of proteins, metabolic related-diseases and apoptosis due to ER stress. However, the roles of 4PBA during odontogenesis are not elucidated. This study revealed the effects of 4PBA during molar development in mice.
We employed in vitro organ cultivation and renal transplantation methods which would mimic the permanent tooth development in an infant period of human. The
cultivated tooth germs and renal calcified teeth were examined by histology and immunohistochemical analysis.
Our results revealed that treatment of 4PBA altered expression patterns of enamel knot related signaling molecules, and consequently affected cellular secretion and patterned formation of dental hard tissues including dentin and enamel during tooth morphogenesis. The alteration of ER stress by 4PBA treatment during organogenesis would suggest that proper ER stress is important for pattern formation during tooth development and morphogenesis, and 4PBA as a chemical chaperone would be one of the candidate molecules for dental and hard tissue regeneration.
Apigenin, a natural product belonging to the flavone class, affects various cell physiologies, such as cell signaling, inflammation, proliferation, migration, and protease production. In this study, ...apigenin was applied to mouse molar pulp after mechanically pulpal exposure to examine the detailed function of apigenin in regulating pulpal inflammation and tertiary dentin formation.
cell cultivation using human dental pulp stem cells (hDPSCs) and
mice model experiments were employed to examine the effect of apigenin in the pulp and dentin regeneration.
cultivation of hDPSCs with apigenin treatment upregulated bone morphogenetic protein (BMP)- and osteogenesis-related signaling molecules such as BMP2, BMP4, BMP7, bone sialoprotein (BSP), runt-related transcription factor 2 (RUNX2), and osteocalcin (OCN) after 14 days. After apigenin local delivery in the mice pulpal cavity, histology and cellular physiology, such as the modulation of inflammation and differentiation, were examined using histology and immunostainings. Apigenin-treated specimens showed period-altered immunolocalization patterns of tumor necrosis factor (TNF)-α, myeloperoxidase (MPO), NESTIN, and transforming growth factor (TGF)-β1 at 3 and 5 days. Moreover, the apigenin-treated group showed a facilitated dentin-bridge formation with few irregular tubules after 42 days from pulpal cavity preparation. Micro-CT images confirmed obvious dentin-bridge structures in the apigenin-treated specimens compared with the control. Apigenin facilitated the reparative dentin formation through the modulation of inflammation and the activation of signaling regulations. Therefore, apigenin would be a potential therapeutic agent for regenerating dentin in exposed pulp caused by dental caries and traumatic injury.
Circumvallate papilla (CVP) is a distinctively structured with dome-shaped apex, and the surrounding trench which contains over two hundred taste buds on the lateral walls. Although CVP was ...extensively studied to determine the regulatory mechanisms during organogenesis, it still remains to be elucidated the principle mechanisms of signaling regulations on morphogenesis including taste buds formation. The key role of Yes-associated protein (YAP) in the regulation of organ size and cell proliferation in vertebrates is well understood, but little is known about the role of this signaling pathway in CVP development. We aimed to determine the putative roles of YAP signaling in the epithelial patterning during CVP morphogenesis. To evaluate the precise localization patterns of YAP and other related signaling molecules, including β-catenin, Ki67, cytokeratins, and PGP9.5, in CVP tissue, histology and immunohistochemistry were employed at E16 and adult mice. Our results suggested that there are specific localization patterns of YAP and Wnt signaling molecules in developing and adult CVP. These concrete localization patterns would provide putative involvements of YAP and Wnt signaling for proper epithelial cell differentiation including the formation and maintenance of taste buds.
For hard tissue formation, cellular mechanisms, involved in protein folding, processing, and secretion play important roles in the endoplasmic reticulum (ER). In pathological and regeneration ...conditions, ER stress hinders proper formation and secretion of proteins, and tissue regeneration by unfolded protein synthesis. 4-Phenylbutyric acid (4PBA) is a chemical chaperone that alleviates ER stress through modulation in proteins folding and protein trafficking. However, previous studies about 4PBA only focused on the metabolic diseases rather than on hard tissue formation and regeneration. Herein, we evaluated the function of 4PBA in dentin regeneration using an exposed pulp animal model system via a local delivery method as a drug repositioning strategy. Our results showed altered morphological changes and cellular physiology with histology and immunohistochemistry. The 4PBA treatment modulated the inflammation reaction and resolved ER stress in the early stage of pulp exposure. In addition, 4PBA treatment activated blood vessel formation and TGF-β1 expression in the dentin-pulp complex. Micro-computed tomography and histological examinations confirmed the facilitated formation of the dentin bridge in the 4PBA-treated specimens. These results suggest that proper modulation of ER stress would be an important factor for secretion and patterned formation in dentin regeneration.
FUSE binding protein 1 (
), a regulator of the c-Myc transcription factor and a DNA/RNA-binding protein, plays important roles in the regulation of gene transcription and cellular physiology. In this ...study, to reveal the precise developmental function of
, we examined the detailed expression pattern and developmental function of
during tooth morphogenesis by RT-qPCR, in situ hybridization, and knock-down study using in vitro organ cultivation methods. In embryogenesis,
is obviously expressed in the enamel organ and condensed mesenchyme, known to be important for proper tooth formation. Knocking down
at E14 for two days, showed the altered expression patterns of tooth development related signalling molecules, including
and
. In addition, transient knock-down of
at E14 revealed changes in the localization patterns of c-Myc and cell proliferation in epithelium and mesenchyme, related with altered tooth morphogenesis. These results also showed the decreased amelogenin and dentin sialophosphoprotein expressions and disrupted enamel rod and interrod formation in one- and three-week renal transplanted teeth respectively. Thus, our results suggested that
plays a modulating role during dentinogenesis and amelogenesis by regulating the expression pattern of signalling molecules to achieve the proper structural formation of hard tissue matrices and crown morphogenesis in mice molar development.
Unlike vertebrates, the number of toothed taxa in invertebrates is very few, with leeches being the only tooth-bearing organisms in the phylum Annelida. Copious studies have been conducted regarding ...vertebrate teeth; however, studies regarding the structure and function of invertebrate teeth are limited. In this study, the tooth structure of leeches, specifically
and
, was revealed, which showed sharp and pointed teeth along the apex of three jaws. Understanding conserved signaling regulations among analogous organs is crucial for uncovering the underlying mechanisms during organogenesis. Therefore, to shed light on the evolutionary perspective of odontogenesis to some extent, we conducted de novo transcriptome analyses using embryonic mouse tooth germs,
teeth, and
proboscises to identify conserved signaling molecules involved in tooth development. The selection criteria were particularly based on the presence of tooth-related genes in mice,
teeth, and
proboscis, wherein 4113 genes were commonly expressed in all three specimens. Furthermore, the chemical nature of leech teeth was also examined via TEM-EDS to compare the chemical composition with vertebrate teeth. The examination of tissue-specific genetic information and chemical nature between leeches and mice revealed chemical similarities between leech and mice teeth, as well as conserved signaling molecules involved in tooth formation, including
,
, and
. Based on our findings, we propose that leech teeth express signaling molecules conserved in mice and these conserved tooth-specific signaling for dental hard tissue formation in mice would corresponds to the structural formation of the toothed jaw in leeches.
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