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BioMed Research International 2020This study is aimed at evaluating the effects of platelet-rich plasma (PRP) on proliferation, viability, and odontogenic differentiation of neural crest stem-like cells...
OBJECTIVE
This study is aimed at evaluating the effects of platelet-rich plasma (PRP) on proliferation, viability, and odontogenic differentiation of neural crest stem-like cells (NCSCs) derived from human dental apical papilla.
MATERIALS AND METHODS
Cells from apical papillae were obtained and then induced to form neural spheres. The expression of NCSC markers p75NTR and HNK-1 in neural sphere cells was detected by immunofluorescence staining. Human PRP was prepared by a 2-step centrifugation method and activated by CaCl and thrombin. The concentrations of PDGF-BB and TGF-1 in whole blood and PRP were measured by an ELISA kit. PRP in five different concentrations (0%, 2.5%, 5%, 10%, and 25%) was applied to culture NCSCs. On the 1, 3, 5, and 7 days, cell proliferation was evaluated by CCK8. Cell viability was tested by a live/dead staining kit. mRNA and protein expression of DSPP and BMP4 were analyzed by RT-qPCR and western blot, respectively. Statistical analysis was performed by a one-way analysis of variance (ANOVA) test or -test.
RESULTS
Dental apical papilla cells formed neural spheres, from which cells displayed positive expression of p75NTR and HNK-1. The concentrations of PDGF-BB and TGF-1 in PRP were about 3.5-fold higher than those in whole blood. 5% and 10% PRP significantly promoted proliferation of NCSCs, while 25% and 50% PRP inhibited cell proliferation from Day 3 to Day 7. Low-concentration (2.5%, 5%, and 10%) PRP slightly improved viability of NCSCs on Day 7. On the other hand, high-concentration (25% and 50%) PRP significantly inhibited viability of NCSCs from Day 3 to Day 7. RT-qPCR and western blot results indicated that 10% PRP could promote odontogenic differentiation of NCSCs on Day 7. mRNA and protein expression of DSPP and BMP4 were significantly upregulated in the 10% PRP group compared to those in the control group ( < 0.05).
CONCLUSIONS
PRP is a simply acquirable blood derivative which contains high concentration of growth factors like PDGF-BB and TGF-1. PRP in a proper concentration could promote proliferation, viability, and odontogenic differentiation of NCSCs derived from human dental apical papilla.
Topics: Biological Products; Cell Proliferation; Cell Survival; Cells, Cultured; Dental Papilla; Humans; Neural Crest; Neural Stem Cells; Odontogenesis; Platelet-Rich Plasma
PubMed: 32461990
DOI: 10.1155/2020/4671989 -
Tissue & Cell Aug 2015Stem cells from the human dental apical papilla (SCAP) can be obtained from almost all extracted wisdom teeth with an immature tooth root. Although different stem cell...
Stem cells from the human dental apical papilla (SCAP) can be obtained from almost all extracted wisdom teeth with an immature tooth root. Although different stem cell lines are used for studies, it remains elusive whether specific characteristics of the dental stem cell cultures such as proliferation rates or the cell differentiation potential are related to the cell source, e.g. the donor tissue of the dental apical papilla. To answer this question, we compared two independent SCAP cell lines from the same donor and compared them with a third cell line from another donor. We investigated the expression of stem cell markers, the efficiency of colony forming units, cell proliferation and the differentiation potential. Results showed particular differences for typical stem cell attributes such as stem cell marker expression, cell proliferation and the adipogenic differentiation. These differences were regardless of the donor of the cell lines. In conclusion, we suppose that stem cell characteristics of SCAP cell cultures are independent from the donor.
Topics: Biomarkers; Cell Differentiation; Cell Proliferation; Dental Papilla; Humans; Mesenchymal Stem Cells; Molar, Third; Osteocalcin; Osteogenesis; Stem Cells; Tissue Donors
PubMed: 26014130
DOI: 10.1016/j.tice.2015.05.002 -
Journal of Cellular Physiology Nov 2015Bmp2 is essential for dentin formation. Bmp2 cKO mice exhibited similar phenotype to dentinogenesis imperfecta, showing dental pulp exposure, hypomineralized dentin, and...
Bmp2 is essential for dentin formation. Bmp2 cKO mice exhibited similar phenotype to dentinogenesis imperfecta, showing dental pulp exposure, hypomineralized dentin, and delayed odontoblast differentiation. As it is relatively difficult to obtain lot of primary Bmp2 cKO dental papilla mesenchymal cells and to maintain a long-term culture of these primary cells, availability of immortalized deleted Bmp2 dental papilla mesenchymal cells is critical for studying the underlying mechanism of Bmp2 signal in odontogenesis. In this study, our goal was to generate an immortalized deleted Bmp2 dental papilla mesenchymal (iBmp2(ko/ko)dp) cell line by introducing Cre recombinase and green fluorescent protein (GFP) into the immortalized mouse floxed Bmp2 dental papilla mesenchymal (iBmp2(fx/fx)dp) cells. iBmp2(ko/ko)dp cells were confirmed by GFP and PCR. The deleted Bmp2 cells exhibited slow cell proliferation rate and cell growth was arrested in G2 phase. Expression of tooth-related marker genes and cell differentiation were decreased in the deleted cells. Importantly, extracellular matrix remodeling was impaired in the iBmp2(ko/ko)dp cells as reflected by the decreased Mmp-9 expression. In addition, with exogenous Bmp2 induction, these cell differentiation and mineralization were rescued as well as extracellular matrix remodeling was enhanced. Therefore, we for the first time described establishment of iBmp(ko/ko) cells that are useful for study of mechanisms in regulating dental papilla mesenchymal cell lineages.
Topics: Animals; Bone Morphogenetic Protein 2; Cell Differentiation; Cell Line; Cell Lineage; Cell Proliferation; Dental Papilla; Gene Expression Regulation, Developmental; Gene Knockout Techniques; Mesenchymal Stem Cells; Mice; Odontoblasts; Odontogenesis; Tooth
PubMed: 26037045
DOI: 10.1002/jcp.25061 -
Connective Tissue Research Jan 2024The important role of non-coding RNAs in odontoblastic differentiation of dental tissue-derived stem cells has been widely demonstrated; however, whether piRNA (a...
PURPOSE
The important role of non-coding RNAs in odontoblastic differentiation of dental tissue-derived stem cells has been widely demonstrated; however, whether piRNA (a subclass of non-coding RNA) involved in the course of odontoblastic differentiation is not yet available. This study aimed to investigate the expression profile of piRNA during odontogenic differentiation of mDPCs and the potential molecular mechanism in vitro.
MATERIALS AND METHODS
The primary mouse dental papilla cells (mDPCs) were isolated from the first molars of 1-day postnatal Kunming mice. Then, they were cultured in odontogenic medium for 9 days. The expression profile of piRNA was detected by Small RNA sequencing. RT-qPCR was used to verify the elevation of piR-368. The mRNA and protein levels of mineralization markers were examined by qRT-PCR and Western blot analysis. Alkaline phosphatase (ALP) activity and alizarin red S staining were conducted to assess the odontoblastic differentiation ability.
RESULTS
We validated piR-368 was significantly upregulated and interference with piR-368 markedly inhibited the odontogenic differentiation of mDPCs. In addition, the relationship between Smad1/5 signaling pathway and piR-368-induced odontoblastic differentiation has been discovered. Finally, we demonstrated Smurf1 as a target gene of piR-368 using dual-luciferase assays.
CONCLUSION
This study was the first to illustrate the participation of piRNA in odontoblastic differentiation. We proved that piR-368 promoted odontoblastic differentiation of mouse dental papilla cells via the Smad1/5 signaling pathway by targeting Smurf1.
Topics: Animals; Mice; Cell Differentiation; Cells, Cultured; Dental Papilla; Dental Pulp; Extracellular Matrix Proteins; Odontoblasts; Piwi-Interacting RNA; Signal Transduction; Smad1 Protein
PubMed: 37978579
DOI: 10.1080/03008207.2023.2281319 -
The Anatomical Record Dec 1992The localization of different cytoskeletal proteins (keratin, vimentin, desmin, actin, and alpha-smooth muscle actin) was examined by immunohistochemistry in normal... (Comparative Study)
Comparative Study
The localization of different cytoskeletal proteins (keratin, vimentin, desmin, actin, and alpha-smooth muscle actin) was examined by immunohistochemistry in normal human adult dental pulp and compared with dental papilla of tooth germs. Keratin and actin were localized in enamel organ. Vimentin and actin were observed in the dental papilla and in the adult dental pulp. Desmin and alpha-smooth muscle actin were present only in the vessel walls. These data are discussed paying particular attention to the origin and the peculiar functional characters of the dental papilla and pulp.
Topics: Actins; Dental Papilla; Dental Pulp; Desmin; Humans; Immunohistochemistry; Keratins; Tooth Germ; Vimentin
PubMed: 1280923
DOI: 10.1002/ar.1092340414 -
International Journal of Oral Science Mar 2009Tooth loss compromises human oral health. Although several prosthetic methods, such as artificial denture and dental implants, are clinical therapies to tooth loss... (Review)
Review
Tooth loss compromises human oral health. Although several prosthetic methods, such as artificial denture and dental implants, are clinical therapies to tooth loss problems, they are thought to have safety and usage time issues. Recently, tooth tissue engineering has attracted more and more attention. Stem cell based tissue engineering is thought to be a promising way to replace the missing tooth. Mesenchymal stem cells (MSCs) are multipotent stem cells which can differentiate into a variety of cell types. The potential MSCs for tooth regeneration mainly include stem cells from human exfoliated deciduous teeth (SHEDs), adult dental pulp stem cells (DPSCs), stem cells from the apical part of the papilla (SCAPs), stem cells from the dental follicle (DFSCs), periodontal ligament stem cells (PDLSCs) and bone marrow derived mesenchymal stem cells (BMSCs). This review outlines the recent progress in the mesenchymal stem cells used in tooth regeneration.
Topics: Adult Stem Cells; Bone Marrow Cells; Dental Papilla; Dental Pulp; Dental Sac; Humans; Mesenchymal Stem Cells; Multipotent Stem Cells; Periodontal Ligament; Regeneration; Tissue Engineering; Tooth; Tooth, Deciduous
PubMed: 20690498
DOI: 10.4248/ijos.08032 -
Scientific Reports Dec 2015Dentinogenesis is the formation of dentin, a substance that forms the majority of teeth, and this process is performed by odontoblasts. Dental papilla cells (DPCs), as...
Dentinogenesis is the formation of dentin, a substance that forms the majority of teeth, and this process is performed by odontoblasts. Dental papilla cells (DPCs), as the progenitor cells of odontoblasts, undergo the odontogenic differentiation regulated by multiple cytokines and paracrine signal molecules. Ape1 is a perfect paradigm of the function complexity of a biological macromolecule with two major functional regions for DNA repair and redox regulation, respectively. To date, it remains unclear whether Ape1 can regulate the dentinogenesis in DPCs. In the present study, we firstly examed the spatio-temporal expression of Ape1 during tooth germ developmental process, and found the Ape1 expression was initially high and then gradually reduced along with the tooth development. Secondly, the osteo/odontogenic differentiation capacity of DPCs was up-regulated when treated with either Ape1-shRNA or E3330 (a specific inhibitor of the Ape1 redox function), respectively. Moreover, we found that the canonical Wnt signaling pathway was activated in this process, and E3330 reinforced-osteo/odontogenic differentiation capacity was suppressed by Dickkopf1 (DKK1), a potent antagonist of canonical Wnt signaling pathway. Taken together, we for the first time showed that inhibition of Ape1 redox regulation could promote the osteo/odontogenic differentiation capacity of DPCs via canonical Wnt signaling pathway.
Topics: Animals; Benzoquinones; Cell Cycle; Cell Differentiation; Cell Proliferation; DNA-(Apurinic or Apyrimidinic Site) Lyase; Dental Papilla; Models, Biological; Odontogenesis; Osteogenesis; Oxidation-Reduction; Propionates; RNA, Small Interfering; Rats; Time Factors; Tooth; Up-Regulation; Wnt Signaling Pathway
PubMed: 26639148
DOI: 10.1038/srep17483 -
Advances in Dental Research Aug 2001Odontoblasts differentiate from the cells of the dental papilla, and it has been well-established that their differentiation in developing teeth is induced by the dental... (Review)
Review
Odontoblasts differentiate from the cells of the dental papilla, and it has been well-established that their differentiation in developing teeth is induced by the dental epithelium. In experimental studies, no other mesenchymal cells have been shown to have the capacity to differentiate into odontoblasts, indicating that the dental papilla cells have been committed to odontoblast cell lineage during earlier developmental stages. We propose that the advancing differentiation within the odontoblast cell lineage is regulated by sequential epithelial signals. The first epithelial signals from the early oral ectoderm induce the odontogenic potential in the cranial neural crest cells. The next step in the determination of the odontogenic cell lineage is the development of the dental papilla from odontogenic mesenchyme. The formation of the dental papilla starts at the onset of the transition from the bud to the cap stage of tooth morphogenesis, and this is regulated by epithelial signals from the primary enamel knot. The primary enamel knot is a signaling center which forms at the tip of the epithelial tooth bud. It becomes fully developed and morphologically discernible in the cap-stage dental epithelium and expresses at least ten different signaling molecules belonging to the BMP, FGF, Hh, and Wnt families. In molar teeth, secondary enamel knots appear in the enamel epithelium at the sites of the future cusps. They also express several signaling molecules, and their formation precedes the folding and growth of the epithelium. The differentiation of odontoblasts always starts from the tips of the cusps, and therefore, it is conceivable that some of the signals expressed in the enamel knots may act as inducers of odontoblast differentiation. The functions of the different signals in enamel knots are not precisely known. We have shown that FGFs stimulate the proliferation of mesenchymal as well as epithelial cells, and they may also regulate the growth of the cusps. We have proposed that the enamel knot signals also have important roles, together with mesenchymal signals, in regulating the patterning of the cusps and hence the shape of the tooth crown. We suggest that the enamel knots are central regulators of tooth development, since they link cell differentiation to morphogenesis.
Topics: Animals; Cell Differentiation; Cell Division; Cell Lineage; Dental Enamel; Dental Papilla; Ectoderm; Epithelial Cells; Epithelium; Fibroblast Growth Factors; Intercellular Signaling Peptides and Proteins; Mesoderm; Morphogenesis; Neural Crest; Odontoblasts; Odontogenesis; Signal Transduction; Tooth Crown; Tooth Germ
PubMed: 12640732
DOI: 10.1177/08959374010150010401 -
Archives of Oral Biology Feb 1999Hepatocyte growth factor/scatter factor (HGF/SF), a broad-spectrum and multifunctional cytokine, is essential for the development of tissues including tooth. Here it was...
Hepatocyte growth factor/scatter factor (HGF/SF), a broad-spectrum and multifunctional cytokine, is essential for the development of tissues including tooth. Here it was found that the HGF/SF content of human dental papillae obtained from 8 to 16-year-old individuals decreased significantly with age. Cultured fibroblasts prepared from the dental papillae of individuals of different ages produced HGF/SF at almost the same rate, but the sensitivities of the cells to interleukin-1alpha and tumour necrosis factor-alpha for the production of HGF/SF increased with age. Generally, mesenchymal cells such as fibroblasts produce HGF/SF but do not express c-Met, a receptor for HGF/SF, yet fibroblasts in dental papilla and cultured fibroblasts prepared from dental papilla did express c-Met, as determined by immunohistochemistry, in situ hybridization and reverse transcription-polymerase chain reaction. Recombinant human [125I]iodo-HGF/SF specifically bound to cell-surface macromolecules with a mol. wt of 146,000, which is the same as that of the beta-subunit of c-Met. The physiological role of c-Met on fibroblasts in dental papilla is unknown, but the addition of 2 ng of HGF/SF per ml to the culture medium significantly stimulated DNA synthesis in the cells, as determined by pulse labelling with [3H]thymidine. Exogenous HGF/SF also stimulated secretion by the cells of vascular endothelial growth factor, a cytokine that induces blood vessel-formation. These results suggest that HGF/SF may be involved in tooth development via autocrine mechanisms.
Topics: Adolescent; Aging; Autocrine Communication; Cells, Cultured; Child; DNA; Dental Papilla; Endothelial Growth Factors; Fibroblasts; Gene Expression Regulation; Hepatocyte Growth Factor; Humans; Immunohistochemistry; In Situ Hybridization; Interleukin-1; Iodine Radioisotopes; Lymphokines; Mesoderm; Molecular Weight; Neovascularization, Physiologic; Odontogenesis; Polymerase Chain Reaction; Proto-Oncogene Proteins c-met; Radiopharmaceuticals; Thymidine; Tritium; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors
PubMed: 10206332
DOI: 10.1016/s0003-9969(98)00101-0 -
Journal of Cellular Physiology Nov 2015Fam20c is essential for the normal mineralization of dentin and bone. The generation of odontoblast and osteoblast cell lines carrying floxed Fam20c allele can offer...
Fam20c is essential for the normal mineralization of dentin and bone. The generation of odontoblast and osteoblast cell lines carrying floxed Fam20c allele can offer valuable tools for the study of the roles of Fam20c in the mineralization of dentin and bone. The limited capability of the primary odontoblasts and osteoblasts to proliferate necessitates the development of odontoblast and osteoblast cell lines serving as substitutes for the study of differentiation and mineralization of the odontoblasts and osteoblasts. In this study, we established and characterized immortalized mouse floxed Fam20c dental papilla mesenchymal and osteoblast cell lines. The isolated primary mouse floxed Fam20c dental papilla mesenchymal cells and osteoblasts were immortalized by the infection of lentivirus containing Simian Virus 40 T-antigen (SV40 T-Ag). The immortalization of floxed Fam20c dental papilla mesenchymal cells and osteoblasts was verified by the long-term passages and genomic integration of SV40 T-Ag. The immortalized floxed Fam20c dental papilla mesenchymal and osteoblast cell lines not only proliferated at a high rate and retained the morphology of their primary counterparts, but also preserved the dentin and bone specific gene expression as the primary dental papilla mesenchymal cells and osteoblasts did. Consistently, the capability of the primary floxed Fam20c dental papilla mesenchymal cells and osteoblasts to mineralize was also inherited by the immortalized dental papilla mesenchymal and osteoblast cell lines. Thus, we have successfully generated the immortalized mouse floxed Fam20c dental papilla mesenchymal and osteoblast cell lines.
Topics: Animals; Bone Morphogenetic Protein 2; Calcification, Physiologic; Calcium-Binding Proteins; Cell Differentiation; Cell Line; Cell Proliferation; Dental Papilla; Dentin; Extracellular Matrix Proteins; Gene Expression Regulation, Developmental; Mesenchymal Stem Cells; Mice; Osteoblasts
PubMed: 25833681
DOI: 10.1002/jcp.25008