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The Conditioned Medium of Calcined Tooth Powder Promotes the Osteogenic and Odontogenic Differentiation of Human Dental Pulp Stem Cells via MAPK Signaling Pathways.

ABSTRACT: The calcined tooth powder (CTP), a type of allogeneic biomimetic mineralized material, has been confirmed that can promote new bone formation when obtained at high temperature. The aim of this study was to investigate effects of the conditioned medium of calcined tooth powder (CTP-CM) on the osteogenic and odontogenic differentiation of human dental pulp stem cells (hDPSCs) and the underlying mechanisms involved. First, ALP activity assay determined that 200??g/mL was the optimal concentration of CTP-CM for the following experiments. CTP-CM had no significant effect on the proliferation of hDPSCs as indicated by CCK-8 and FCM analysis. Both the gene and protein (DSPP/DSPP, RUNX2/RUNX2, OCN/OCN, OSX/OSX, OPN/OPN, ALP/ALP, and COL-1/COL-1) expression levels increased in the CTP-CM-induced hDPSC group as compared with those in the control group at day 3 or 7, showing the positive regulation of CTP-CM on the osteo/odontogenic differentiation of hDPSCs. Mechanistically, MAPK signaling pathways were activated after the CTP-CM treatment, and the inhibitors targeting MAPK were identified which weakened the effects of CTM-CM on the committed differentiation of hDPSCs. These findings could lead to the creation of stem cell therapies for dental regeneration.

SUBMITTER: Wu J

PROVIDER: S-EPMC6444228 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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The Conditioned Medium of Calcined Tooth Powder Promotes the Osteogenic and Odontogenic Differentiation of Human Dental Pulp Stem Cells via MAPK Signaling Pathways.

Wu Jintao J Li Na N Fan Yuan Y Wang Yanqiu Y Gu Yongchun Y Li Zehan Z Pan Yin Y Romila Gobin G Zhou Zuomin Z Yu Jinhua J

Stem cells international 20190319

The calcined tooth powder (CTP), a type of allogeneic biomimetic mineralized material, has been confirmed that can promote new bone formation when obtained at high temperature. The aim of this study was to investigate effects of the conditioned medium of calcined tooth powder (CTP-CM) on the osteogenic and odontogenic differentiation of human dental pulp stem cells (hDPSCs) and the underlying mechanisms involved. First, ALP activity assay determined that 200 <i>μ</i>g/mL was the optimal concentr ...[more]

PMID: 31015840

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Project description:BackgroundChemotherapy-induced alopecia (CIA) is a distressing adverse effect of chemotherapy, with an estimated incidence of 65% and limited treatment options. Cyclophosphamide (CYP) is a common alopecia-inducing chemotherapy agent. Human dental pulp stem cells (DPSCs) secrete several paracrine factors that up-regulate hair growth. Conditioned medium (CM) collected from DPSCs (DPSC-CM) promotes hair growth; culturing mesenchymal stem cells under hypoxic conditions can enhance this effect.MethodsThe effect of DPSC-CM cultured under normoxic (N-) and hypoxic (H-) conditions against CYP-mediated cytotoxicity in keratinocytes was examined using cell viability assay, lactate dehydrogenase (LDH) cytotoxicity assay, and apoptosis detection. The damage-response pathway was determined in a well-established CIA mouse model by analyzing macroscopic effects, histology, and apoptosis. Reverse transcription-quantitative PCR and Caspase-3/7 activity assay were used to investigate the impact of DPSC-CM on the molecular damage-response pathways in CYP-treated mice. The effect of post-CIA DPSC-CM application on post-CIA hair regrowth was analyzed by macroscopic effects and microstructure observation of the hair surface. Furthermore, to investigate the safety of DPSC-CM as a viable treatment option, the effect of DPSC-CM on carcinoma cell lines was examined by cell viability assay and a subcutaneous tumor model.ResultsIn the cell viability assay, DPSC-CM was observed to increase the number of keratinocytes over varying CYP concentrations. Furthermore, it reduced the LDH activity level and suppressed apoptosis in CYP-treated keratinocytes. DPSC-CM exhibited the cytoprotective role in vivo via the dystrophic anagen damage-response pathway. While both N-CM and H-CM downregulated the Caspase-3/7 activity level, H-CM downregulated Caspase-3 mRNA expression. The proportion of post-CIA H-CM-treated mice with > 90% normal hair was nearly twice that of vehicle- or N-CM-treated mice between days 50 and 59 post-depilation, suggesting that post-CIA H-CM application may accelerate hair regrowth and improve hair quality. Furthermore, DPSC-CM suppressed proliferation in vitro in certain carcinoma cell lines and did not promote the squamous cell carcinoma (SCC-VII) tumor growth rate in mice.ConclusionsThe potentiality of DPSC-CM and H-CM as a promising cytoprotective agent and hair regrowth stimulant, respectively, for CIA needs in-depth exploration.

Project description:IntroductionOdontogenic differentiation of human dental pulp stem cells (hDPSCs) is a key step of pulp regeneration. Recent studies showed that circular RNAs (circRNAs) have many biological functions and that competing endogenous RNA (ceRNA) is their most common mechanism of action. However, the role of circRNAs in hDPSCs during odontogenesis is still unclear.MethodsIsolated hDPSCs were cultured in essential and odontogenic medium. Total RNA was extracted after 14 days of culture, and then, microarray analysis was performed to measure the differential expressions of circRNAs. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was then performed to validate the microarray results. Based on microarray data from this study and available in the database, a ceRNA network was constructed to investigate the potential function of circRNAs during odontogenesis. In addition, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to investigate the potential correlation between signaling pathways and circRNAs. In addition, qRT-PCR and Western blot analysis were used to explore the function of hsa_circRNA_104101.ResultsWe found 43 upregulated circRNAs and 144 downregulated circRNAs during the odontogenic differentiation process (fold change > 1.5 and <-1.5, respectively; P < 0.05). qRT-PCR results were in agreement with the microarray results. Bioinformatic analysis revealed that the Wnt signaling pathway and the TGF-β signaling pathway, as well as the other pathways associated with odontogenic differentiation, were correlated to the differentially expressed circRNAs. hsa_circRNA_104101 was proved to promote the odontogenic differentiation of hDPSCs.ConclusionThis study reported 187 circRNAs that were differentially expressed in hDPSCs during odontogenic differentiation. Bioinformatic analysis of the expression data suggested that circRNA-miRNA-mRNA networks might act as a crucial mechanism for hDPSC odontogenic differentiation, providing a theoretical foundation for the study of pulp regeneration regulation by circRNAs.

Dataset Information

The Conditioned Medium of Calcined Tooth Powder Promotes the Osteogenic and Odontogenic Differentiation of Human Dental Pulp Stem Cells via MAPK Signaling Pathways.

Publications

The Conditioned Medium of Calcined Tooth Powder Promotes the Osteogenic and Odontogenic Differentiation of Human Dental Pulp Stem Cells via MAPK Signaling Pathways.

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