MSCs-Derived Decellularised Matrix: Cellular Responses and Regenerative Dentistry.

The decellularised extracellular matrix (dECM) of in vitro cell culture is a naturally derived biomaterial formed by the removal of cellular components. The compositions of molecules in the extracellular matrix (ECM) differ depending on various factors, including the culture conditions. Cell-derived ECM provides a 3-dimensional structure that has a complex influence on cell signalling, which in turn affects cell survival and differentiation. This review describes the effects of dECM derived from mesenchymal stem cells (MSCs) on cell responses, including cell migration, cell proliferation, and cell differentiation in vitro. Published articles were searched in the PubMed databases in 2005 to 2022, with assigned keywords (MSCs and decellularisation and cell culture). The 41 articles were reviewed, with the following criteria. (1) ECM was produced exclusively from MSCs; (2) decellularisation processes were performed; and (3) the dECM production was discussed in terms of culture systems and specific supplementations that are suitable for creating the dECM biomaterials. The dECM derived from MSCs supports cell adhesion, enhances cell proliferation, and promotes cell differentiation. Importantly, dECM derived from dental MSCs shows promise in regenerative dentistry applications. Therefore, the literature strongly supports cell-based dECMs as a promising option for innovative tissue engineering approaches for regenerative medicine.

Simvastatin Induces Apoptosis but Attenuates Migration in SCAPs.

AIM: Simvastatin has emerged as having a promising role in controlling stem cell behaviours. This study aimed to evaluate the effects of simvastatin on the viability, growth, and migration of stem cells isolated from apical papillae (SCAPs) in vitro. METHODS: SCAPs were isolated and characterised. The viability and proliferation were assessed using live/dead and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assays, respectively. Cell migration was evaluated using scratch assays. Cell cycle progression and apoptosis were examined using flow cytometry analysis. RESULTS: Simvastatin at a concentration of 100 to 1000 nM did not exhibit cytotoxicity. Simvastatin reduced cell numbers at days 3 and 7. In addition, simvastatin markedly decreased colony formation in both colony number and cell density in a dose-dependent manner. An increase in apoptosis was observed at day 7. There was statistically significant increased in sub G0 population. An in vitro cell migration was attenuated in a dose-dependent manner. CONCLUSION: Simvastatin affects SCAPs' viability, proliferation, and cell migration. The reduction of cell viability at day 7 could be due to apoptotic induction.

Effects of ethylenediaminetetraacetic acid on stem cells from the apical papilla: In vitro study.

Background/purpose: Ethylenediaminetetraacetic acid (EDTA) is used as an irrigant in regenerative endodontic treatment. The present study aimed to investigate the effects of EDTA on stem cells from apical papilla (SCAPs) in vitro. Materials and methods: Human SCAPs were isolated and characterised. The cells were treated with media supplemented with EDTA at concentrations ranging from 1.25% to 17%. Cell proliferation and apoptosis were examined using MTT assay and annexin V/propidium iodide staining. Cell migration was determined by a scratch assay. Gene expression was evaluated using a real-time polymerase chain reaction. Mineral deposition, a hallmark of osteogenesis in vitro, was determined using alizarin red s staining. Results: Overall, SCAPs exhibited mesenchymal stem cell characteristics. EDTA treatment at 2.50% and 1.25% did not significantly exhibit cytotoxicity and alter cell morphology. However, EDTA attenuated cell proliferation and reduced MKI67 mRNA expression in SCAPs. Further, EDTA significantly induced early cell apoptosis at 48 h. Cell migration was delayed with EDTA treatment. After maintaining SCAPs in an osteogenic induction medium, EDTA diminished mineral deposition by SCAPs on day 14. Conclusion: EDTA treatment exhibits adverse effects on SCAPs in vitro. Hence, EDTA exposure to periapical tissues should be avoided to minimise the negative impacts on SCAPs cells in regenerative processes.

Indirect Immobilised Jagged-1 Enhances Matrisome Proteins Associated with Osteogenic Differentiation of Human Dental Pulp Stem Cells: A Proteomic Study.

The indirect immobilisation of Jagged-1 (Jagged-1) promoted osteogenic differentiation of human dental pulp cells (hDPs). Furthermore, the analysis of the Reactome pathway of RNA sequencing data indicates the upregulated genes involved with the extracellular matrix (ECM). Hence, our objective was to investigate the effects of Jagged-1 on proteomic profiles of human dental pulp stem cells (hDPSC). hDPSCs were cultured on the surface coated with human IgG Fc fragment (hFc) and the surface coated with rhJagged1/Fc recombinant protein-coated surface. Cells were differentiated to the osteogenic lineage using an osteogenic differentiation medium (OM) for 14 days, and cells cultured in a growth medium were used as a control. The protein component of the cultured cells was extracted into the cytosol, membrane, nucleus, and cytoskeletal compartment. Subsequently, the proteomic analysis was performed using liquid chromatography-tandem mass spectrometry (LC-MS). Metascape gene list analysis reported that Jagged-1 stimulated the expression of the membrane trafficking protein (DOP1B), which can indirectly improve osteogenic differentiation. hDPSCs cultured on Jagged-1 surface under OM condition expressed COL27A1, MXRA5, COL7A1, and MMP16, which played an important role in osteogenic differentiation. Furthermore, common matrisome proteins of all cellular components were related to osteogenesis/osteogenic differentiation. Additionally, the gene ontology categorised by the biological process of cytosol, membrane, and cytoskeleton compartments was associated with the biomineralisation process. The gene ontology of different culture conditions in each cellular component showed several unique gene ontologies. Remarkably, the Jagged-1_OM culture condition showed the biological process related to odontogenesis in the membrane compartment. In conclusion, the Jagged-1 induces osteogenic differentiation could, mainly through the regulation of protein in the membrane compartment.

Effects of decellularized extracellular matrix derived from Jagged1-treated human dental pulp stem cells on biological responses of stem cells isolated from apical papilla.

Objective: Indirect Jagged1 immobilization efficiently activates canonical Notch signaling in human dental pulp stem cells (hDPSCs). This study aimed to investigate the characteristics of the Jagged1-treated hDPSC-derived decellularized extracellular matrix (dECM) and its biological activity on odonto/osteogenic differentiation of stem cells isolated from apical papilla (SCAPs). Methods: Bioinformatic database of Jagged1-treated hDPSCs was analyzed using NetworkAnalyst. hDPSCs seeded on the Jagged1 immobilized surface were maintained with normal or osteogenic induction medium (OM) followed by decellularization procedure, dECM-N, or dECM-OM, respectively. SCAPs were reseeded on each dECM with either the normal medium or the OM. Cell viability was determined by MTT assay. Characteristics of dECMs and SCAPs were evaluated by SEM, EDX, immunofluorescent staining, and alcian blue staining. Mineralization was assessed by alizarin red S, Von Kossa, and alkaline phosphatase staining. Statistical significance was considered at p < 0.05. Results: RNA-seq database revealed upregulation of several genes involved in ECM organization, ECM-receptor interaction, and focal adhesion in Jagged1-treated hDPSCs. Immobilized Jagged1 increased the osteogenesis of the hDPSC culture with OM. dECMs showed fibrillar-like network structure and maintained major ECM proteins, fibronectin, type I-collagen, and glycosaminoglycans. A decrease in calcium and phosphate components was observed in dECMs after the decellularized process. Cell viability on dECMs did not alter by 7 days. Cell attachment and f-actin cytoskeletal organization of SCAPs proliferated on Jagged1-treated dECMs were comparable to those of the control dECMs. SCAPs exhibited significantly higher mineralization on dECM-N in OM and markedly enhanced on dECM-OM with normal medium or OM conditions. Conclusion: Jagged1-treated hDPSC-derived dECMs are biocompatible and increase odonto/osteogenic differentiation of SCAPs. The results suggested the potential of Jagged1 dECMs, which could be further developed into ECM scaffolds for application in regenerative medicine.