scholarly journals Characterization of Odontogenic Differentiation from Human Dental Pulp Stem Cells Using TMT-Based Proteomic Analysis

2020 ◽  
Vol 2020 ◽  
pp. 1-14
Author(s):  
Xijuan Xiao ◽  
Caihong Xin ◽  
Yuqin Zhang ◽  
Jie Yan ◽  
Zhao Chen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Proteomic Analysis ◽  
Dental Pulp ◽  
Kegg Pathway ◽  
Dental Pulp Stem Cells ◽  
Differentially Expressed ◽  
Tandem Mass ◽  
Differentially Expressed Proteins ◽  
Ppi Networks ◽  
Odontogenic Differentiation

Background. The repair of dental pulp injury relies on the odontogenic differentiation of dental pulp stem cells (DPSCs). To better understand the odontogenic differentiation of DPSCs and identify proteins involved in this process, tandem mass tags (TMTs) coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) were applied to compare the proteomic profiles of induced and control DPSCs. Methods. The proteins expressed during osteogenic differentiation of human DPSCs were profiled using the TMT method combined with LC-MS/MS analysis. The identified proteins were subjected to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Then, a protein-protein interaction (PPI) network was constructed. Two selected proteins were confirmed by western blotting (WB) analysis. Results. A total of 223 proteins that were differentially expressed were identified. Among them, 152 proteins were significantly upregulated and 71 were downregulated in the odontogenic differentiation group compared with the control group. On the basis of biological processes in GO, the identified proteins were mainly involved in cellular processes, metabolic processes, and biological regulation, which are connected with the signaling pathways highlighted by KEGG pathway analysis. PPI networks showed that most of the differentially expressed proteins were implicated in physical or functional interaction. The protein expression levels of FBN1 and TGF-β2 validated by WB were consistent with the proteomic analysis. Conclusions. This is the first proteomic analysis of human DPSC odontogenesis using a TMT method. We identified many new differentially expressed proteins that are potential targets for pulp-dentin complex regeneration and repair.

scholarly journals Magnesium-enriched microenvironment promotes odontogenic differentiation in human dental pulp stem cells by activating ERK/BMP2/Smads signaling

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Yuanyuan Kong ◽  
Xiaoli Hu ◽  
Yingqun Zhong ◽  
Ke Xu ◽  
Buling Wu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Rna Sequencing ◽  
Dental Pulp ◽  
Kegg Pathway ◽  
Dental Pulp Stem Cells ◽  
Kegg Pathway Analysis ◽  
Odontogenic Differentiation ◽  
New Strategy ◽  
Human Dental Pulp ◽  
Positive Effect

Abstract Background Magnesium (Mg2+)-enriched microenvironment promotes odontogenic differentiation in human dental pulp stem cells (DPSCs), but the regulatory mechanisms remain undefined. The aim of this work was to assess magnesium’s function in the above process and to explore the associated signaling pathway. Methods DPSCs underwent culture in odontogenic medium with the addition of 0, 1, 5, or 10 mM MgCl2. Intracellular Mg2+ levels in DPSCs were evaluated flow cytometrically using Mag-Fluo-4-AM. Mg2+-entry was inhibited by TRPM7 inhibitor 2-aminoethoxydiphenyl borate (2-APB). RNA-Sequencing was carried out for assessing transcriptome alterations in DPSCs during odontogenic differentiation associated with high extracellular Mg2+. KEGG pathway analysis was performed to determine pathways related to the retrieved differentially expressed genes (DEGs). Immunoblot was performed for assessing magnesium’s role and exploring ERK/BMP2/Smads signaling. Results Mg2+-enriched microenvironment promoted odontogenic differentiation in DPSCs via intracellular Mg2+ increase. Consistently, the positive effect of high extracellular Mg2+ on odontogenic differentiation in DPSCs was blocked by 2-APB, which reduced Mg2+ entry. RNA-sequencing identified 734 DEGs related to odontogenic differentiation in DPSCs in the presence of high extracellular Mg2+. These DEGs participated in many cascades such as MAPK and TGF-β pathways. Consistently, ERK and BMP2/Smads pathways were activated in DPSCs treated with high extracellular Mg2+. In agreement, ERK signaling inhibition by U0126 blunted the effect of high extracellular Mg2+ on mineralization and odontogenic differentiation in DPSCs. Interestingly, BMP2, BMPR1, and phosphorylated Smad1/5/9 were significantly decreased by U0126, indicating that BMP2/Smads acted as downstream of ERK. Conclusions Mg2+-enriched microenvironment promotes odontogenic differentiation in DPSCs by activating ERK/BMP2/Smads signaling via intracellular Mg2+ increase. This study revealed that Mg2+-enriched microenvironment could be used as a new strategy for dental pulp regeneration.

scholarly journals circRNA Expression Profile in Dental Pulp Stem Cells during Odontogenic Differentiation

2020 ◽  
Vol 2020 ◽  
pp. 1-19
Author(s):  
Ming Chen ◽  
Yeqing Yang ◽  
Junkai Zeng ◽  
Zilong Deng ◽  
Buling Wu
Keyword(s):  
Stem Cells ◽  
Signaling Pathway ◽  
Dental Pulp ◽  
Bioinformatic Analysis ◽  
Dental Pulp Stem Cells ◽  
Differentially Expressed ◽  
Common Mechanism ◽  
Pulp Regeneration ◽  
Qrt Pcr ◽  
Odontogenic Differentiation

Introduction. Odontogenic 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. Methods. Isolated 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. Results. We 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. Conclusion. This 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.

Differential expression of long noncoding RNAs in SDF-1α-induced dental pulp stem cells

2021 ◽  
Author(s):  
min xiao ◽  
Bo Yao ◽  
Xiaohan Mei ◽  
yu bai ◽  
Jueyu Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dental Pulp ◽  
Rho Gtpase ◽  
Noncoding Rnas ◽  
Dental Pulp Stem Cells ◽  
Long Noncoding Rnas ◽  
Nodule Formation ◽  
Related Factor ◽  
Altered Expression ◽  
Odontogenic Differentiation

Abstract Background SDF-1α cotreatment was shown to have synergistic effects on BMP-2-induced odontogenic differentiation of human apical dental papillary stem cells (SCAP) both in vitro and in vivo. Long noncoding RNAs (lncRNAs) have an important role in the odontogenic differentiation of dental pulp stem cells (DPSCs). Methods We examined the altered expression of lncRNAs in SDF-1α-induced odontogenic differentiation of DPSCs by lncRNA microarray and quantitative reverse transcription polymerase chain reaction (qRT-PCR) analyses. Alterations in lncRNA expression during odontogenic differentiation of DPSCs were identified. Moreover, bioinformatic analysis [Gene Ontology (GO) analysis and coding-noncoding gene coexpression (CNC) analysis] was conducted to predict the interactions of lncRNAs and identify core regulatory factors in SDF-1α-induced odontogenic differentiation of DPSCs. Results The microarray analysis identified 206 differentially expressed lncRNAs (134 lncRNAs with upregulated expression and 72 with downregulated expression) at 7 days post‑treatment. The data demonstrated that one lncRNA, AC080037.1, regulates SDF-1α-induced odontogenic differentiation of DPSCs. Our data showed that lncRNA AC080037.1 siRNA suppresses DPSCs migration and the expression of Rho GTPase induced by SDF-1α. Moreover, AC080037.1 knockdown significantly affected SDF-1α- and BMP-2-induced mineralized nodule formation and strongly suppressed Runt-related factor-2 (RUNX-2), DMP-1 and DSPP expression in DPSCs. Conclusions Our

scholarly journals METTL3-mediated M6a Modification Regulates Cell Cycle Progression of Dental Pulp Stem Cells

2021 ◽  
Author(s):  
Haiyun Luo ◽  
Wenjing Liu ◽  
Yanli Zhang ◽  
Xiao Jiang ◽  
Shiqing Wu ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Tissue Engineering ◽  
Stem Cells ◽  
Flow Cytometry ◽  
Differentially Expressed Genes ◽  
Dental Pulp ◽  
Cell Cycle Control ◽  
Dental Pulp Stem Cells ◽  
Differentially Expressed ◽  
Expression Level

Abstract Background: Dental pulp stem cells (DPSCs) exhibited self-renewal, pluripotency capacity and served as promising cells source in endodontic regeneration and tissue engineering. Meanwhile, the regenerative capacity of DPSCs is limited and reduced in long lifespan. N6-methyladenosine (m6A) is the most prevalent, reversible internal modification in RNAs. The methyltransferases complex and demethylases mediated m6A methylation and cooperated to impact various biological processes associated with stem cell fate determination. However, the biological effect of m6A methylation in DPSCs remained unclear. Methods: Cell surface markers and differentiation potential of primary DPSCs were identified and m6A immunoprecipitation with deep sequencing (m6A RIP-seq) was used to uncover characteristics of m6A modifications in DPSCs transcriptome. Expression level of m6A-related genes were evaluated in immature/mature pulp tissues and cells. Lentiviral vectors were constructed to knockdown or overexpress methyltransferase like 3 (METTL3). Cell morphology, viability, senescence and apoptosis were further analyzed by β-galactosidase, TUNEL staining and flow cytometry. Bioinformatic analysis combing m6A RIP and shMETTL3 RNA-seq was used to functionally enrich overlapped genes and screen target of METTL3. Cell cycle distributions were assayed by flow cytometry and m6A RIP-qPCR was used to confirm METTL3 mediated m6A methylation in DPSCs. Results: Here, m6A peaks distribution, binding area and motif in DPSCs were first revealed by m6A RIP-seq. We also found a relative high expression level of METTL3 in immature DPSCs with superior regenerative potential and METTL3 knockdown induced cell apoptosis and senescence. Furthermore, Conjoint analysis of m6A RIP and RNA-sequencing showed differentially expressed genes affected by METTL3 depletion was mainly enriched in cell cycle, mitosis and alteration of METTL3 expression resulted in cell cycle arrest which indicated METTL3 make essential effect in cell cycle control. To further investigate underlying mechanisms, we explored proteins interaction network of differentially expressed genes and Polo-like Kinase 1 (PLK1), a critical cycle modulator was identified as target of METTL3-mediated m6A methylation in DPSCs. Conclusions: These results revealed m6A methylated hallmarks in DPSCs and a regulatory role of METTL3 in cell cycle control. Our study shed light on therapeutic approaches in vital pulp therapy and serve new insight in stem cells based tissue engineering.

scholarly journals The role of osteomodulin on osteo/odontogenic differentiation in human dental pulp stem cells

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Wenzhen Lin ◽  
Li Gao ◽  
Wenxin Jiang ◽  
Chenguang Niu ◽  
Keyong Yuan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells ◽  
Odontogenic Differentiation ◽  
Human Dental Pulp

scholarly journals Endogenous GDF11 regulates odontogenic differentiation of dental pulp stem cells

2020 ◽  
Vol 24 (19) ◽  
pp. 11457-11464
Author(s):  
Xingying Qi ◽  
Qingyue Xiao ◽  
Rui Sheng ◽  
Shuang Jiang ◽  
Quan Yuan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dental Pulp ◽  
Dental Pulp Stem Cells ◽  
Odontogenic Differentiation
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