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Proliferation, odontogenic/osteogenic differentiation, and cytokine production by human stem cells of the apical papilla induced by biomaterials: a comparative study.

ABSTRACT: Introduction:Clinical applications of bioactive materials are increasing in biomedical tissue engineering. This study sought to assess the effect of calcium enriched mixture (CEM) cement, Biodentine, mineral trioxide aggregate (MTA), octacalcium phosphate (OCP), and Atlantik on proliferation, odontogenic/osteogenic differentiation, and pro-inflammatory cytokine production by human stem cells of the apical papilla (SCAPs). Materials and methods:Proliferation of SCAPs treated with different biomaterials was evaluated using trypan blue exclusion test and flow cytometry. Differentiation of cells was evaluated using ALP activity, alizarin red staining, and RT-PCR. The expression of genes of pro-inflammatory cytokines was also evaluated using RT-PCR. Results:The SCAPs treated with biomaterials showed significantly higher proliferation, increased ALP activity, higher number of calcified nodules, and up-regulation of genes related to odontogenic/osteogenic markers compared to the control group. The expression of pro-inflammatory cytokines increased in all groups compared to the control group. Conclusion:The tested biomaterials could induce odontogenic/osteogenic differentiation in SCAPs. MTA had a greater potential for induction of differentiation of SCAPs to odontoblast-like cells while OCP had higher potential to induce differentiation of SCAPs to osteoblast-like cells (MTA? BD? CEM? Atlantik? OCP).

SUBMITTER: Saberi E

PROVIDER: S-EPMC6636314 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Proliferation, odontogenic/osteogenic differentiation, and cytokine production by human stem cells of the apical papilla induced by biomaterials: a comparative study.

Saberi Eshaghali E Farhad-Mollashahi Narges N Sargolzaei Aval Fereydoon F Saberi Mersad M

Clinical, cosmetic and investigational dentistry 20190712

<h4>Introduction</h4>Clinical applications of bioactive materials are increasing in biomedical tissue engineering. This study sought to assess the effect of calcium enriched mixture (CEM) cement, Biodentine, mineral trioxide aggregate (MTA), octacalcium phosphate (OCP), and Atlantik on proliferation, odontogenic/osteogenic differentiation, and pro-inflammatory cytokine production by human stem cells of the apical papilla (SCAPs).<h4>Materials and methods</h4>Proliferation of SCAPs treated with d ...[more]

PMID: 31372059

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Project description:Stem cells from apical papilla (SCAP) possess clear osteo?/odontogenic differentiation capabilities, and are regarded as the major cellular source for root dentin development. Bone morphogenetic protein 2 (BMP2) and vascular endothelial growth factor (VEGF) serve pivotal roles in the modulation of tooth development and dentin formation. However, the synergistic effects of BMP2 and VEGF on osteo?/odontogenic differentiation of SCAP remain unclear. The current study aimed to investigate the proliferative and osteo?/odontogenic differentiating capabilities of BMP2 and VEGF gene-co-transfected SCAP (SCAP-BMP2-VEGF) in vitro. The basic characteristics of the isolated SCAP were identified by the induction of multipotent differentiation and by flow cytometry. Lentiviral vector?mediated gene transfection was conducted with SCAP in order to construct blank vector?transfected SCAP (SCAP-green fluorescent protein), BMP2 gene-transfected SCAP (SCAP-BMP2), VEGF gene?transfected SCAP (SCAP?VEGF) and SCAP-BMP2-VEGF. The Cell Counting Kit 8 assay was used to analyze the proliferative capacities of the four groups of cells. The expression of osteo-/odontogenic genes and proteins in the cells were evaluated by reverse transcription-quantitative polymerase chain reaction and western blotting. The mineralized nodules formed by the four group cells were visualized by alkaline phosphatase (ALP) staining. Among the four groups of cells, SCAP?VEGF was demonstrated to exhibit increased proliferation, and SCAP?BMP2?VEGF exhibited reduced proliferation during eight days observation. SCAP?BMP2?VEGF exhibited significantly increased expression levels of ALP, osteocalcin, dentin sialophosphoprotein, dentin matrix acidic phosphoprotein gene 1 and dentin sialoprotein than the other three groups at the majority of the time points. Furthermore, the SCAP?BMP2?VEGF group exhibited a significantly greater number of ALP?positive mineralized nodules than the other groups following 16 days culture in vitro. In conclusion, lentiviral vector-mediated BMP2 and VEGF gene co-transfection significantly activated the osteo?/odontogenic differentiation of human SCAP.

Project description:IntroductionPeriapical alveolar bone loss is the common consequence of apical periodontitis (AP) caused by persistent local inflammation around the apical area. Human stem cells from apical papilla (hSCAPs) play a crucial role in the restoration of bone lesions during AP. Studies have recently identified the critical role of microRNAs (miRNAs) involved in AP pathogenesis, but little is known about their function and potential molecular mechanism, especially in the osteogenesis of hSCAPs during AP. Here, we investigated the role of clinical sample-based specific miRNAs in the osteogenesis of hSCAPs.MethodsDifferential expression of miRNAs were detected in the periapical tissues of normal and patients with AP via transcriptomic analysis, and the expression of miR-199a-5p was confirmed by qRT-PCR. Treatment of hSCAPs with miR-199a-5p mimics while loaded onto beta-tricalcium phosphate (β-TCP) ceramic particle scaffold to explore its effect on osteogenesis in vivo. RNA binding protein immunoprecipitation (RIP) and Luciferase reporter assay were conducted to identify the target gene of miR-199a-5p.ResultsThe expression of miR-199a-5p was decreased in the periapical tissues of AP patients, and miR-199a-5p mimics markedly enhanced cell proliferation and osteogenic differentiation of hSCAPs, while miR-199a-5p antagomir dramatically attenuated hSCAPs osteogenesis. Moreover, we identified and confirmed Interferon Induced Protein with Tetratricopeptide Repeats 2 (IFIT2) as a specific target of miR-199a-5p, and silencing endogenous IFIT2 expression alleviated the inhibitory effect of miR-199a-5p antagomir on the osteogenic differentiation of hSCAPs. Furthermore, miR-199a-5p mimics transfected hSCAPs loaded onto beta-tricalcium phosphate (β-TCP) scaffolds induced robust subcutaneous ectopic bone formation in vivo.DiscussionThese results strengthen our understanding of predictors and facilitators of the key AP miRNAs (miR-199a-5p) in bone lesion repair under periapical inflammatory conditions. And the regulatory networks will be instrumental in exploring the underlying mechanisms of AP and lay the foundation for future regenerative medicine based on dental mesenchymal stem cells.

Project description:To understand the functions of secretory proteins in odontogenesis and to further the understanding of the different molecular events during odontogenesis and osteogenesis, we induced the odonto/osteogenic differentiation of stem cells from dental apical papilla (SCAPs) and bone marrow-derived stem cells (BMSCs) in vitro and compared the expression of secretory proteins during early odonto/osteogenic differentiation using high-performance liquid chromatography with tandem mass spectrometry. The results revealed significant changes by at least 50% in 139 SCAP proteins and 203 BMSC proteins during differentiation. Of these, 92 were significantly upregulated and 47 were significantly downregulated during the differentiation of SCAPs. Most of these proteins showed the same trend during the differentiation of BMSCs. Among the proteins that showed significantly changes during the differentiation of SCAPs and BMSCs, we found that transforming growth factor-?2 (TGF?2) is a key protein in the network with powerful mediation ability. TGF?2 was secreted more by SCAPs than BMSCs, was significantly upregulated during the differentiation of SCAPs and was significantly downregulated during the differentiation of BMSCs. Furthermore, the effects of recombinant human TGF?2 and TGF?1 on the odonto/osteogenic differentiation of SCAPs and BMSCs were investigated. Real-time reverse transcription polymerase chain reaction (RT-PCR) and western blotting data revealed that TGF?2 enhanced the odontogenic-related markers [dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP1)] and inhibited the osteogenic-related marker bone sialoprotein (BSP) in SCAPs, whereas TGF?1 enhanced the BSP expression and inhibited the DSPP and DMP1 expression at early odonto/osteogenic differentiation of SCAPs. However, in BMSCs, TGF?2 enhanced the expression of alkaline phosphatase (ALP), runt-related transcription factor 2 (RUNX2), DSPP, and DMP1, whereas TGF?1 enhanced the expression of ALP and RUNX2, with no significant intergroup difference of DSPP at the early odonto/osteogenic differentiation of BMSCs. TGF?2 is a potentially important molecule with a distinct function in the regulation of odontogenesis and osteogenesis.

Dataset Information

Proliferation, odontogenic/osteogenic differentiation, and cytokine production by human stem cells of the apical papilla induced by biomaterials: a comparative study.

Publications

Proliferation, odontogenic/osteogenic differentiation, and cytokine production by human stem cells of the apical papilla induced by biomaterials: a comparative study.

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