Bone morphogenetic protein&amp;nbsp;2‑enhanced osteogenic differentiation of stem cell spheres by regulation of Runx2 expression

Background and Objectives: Vitamin D is a bone modulator widely used in regenerative medicine. This study aimed to analyze the effects of vitamin D on the osteogenic differentiation and mineralization of human mesenchymal stem cells. Materials and Methods: Spheroids were fabricated using human bone marrow-derived stem cells, and were cultured in the presence of vitamin D at concentrations of 0, 0.1, 1, 10, and 100 nM. Stem cell spheroids were fabricated and the morphological evaluation was conducted on days 1, 3, 7 and 14. Determination of qualitative cellular viability was performed with Live/Dead Kit assay on days 1 and 7. Quantitative cellular viability was evaluated with Cell Counting Kit-8 on days 1, 3, 7, and 14. To analyze the osteogenic differentiation of cell spheroids, alkaline phosphatase activity assays were performed with commercially available kit on days 7 and 14. Real-time polymerase chain reaction was used to determine the expression levels of RUNX2, BSP, OCN, and COL1A1 on days 7 and 14. Results: The stem cells produced well-formed spheroids, and addition of vitamin D did not result in any noticeable changes in the shape. The addition of vitamin D did not significantly change the diameter of the spheroids at 0, 0.1, 1, 10, or 100 nM concentrations. Quantitative cell viability results from days 1, 3, 7 and 14 showed no significant difference between groups (p > 0.05). There was significantly higher alkaline phosphatase activity in the 0.1 nM group when compared with the control group on day 14 (p < 0.05). Real-time polymerase chain reaction results demonstrated that the mRNA expression levels of RUNX2, OCN, and COL1A1 were significantly increased when vitamin D was added to the culture. Conclusions: Based on these findings, we concluded that vitamin D could be applied to the increased osteogenicity of stem cell spheroids.

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“…No. : NM 001101: forward: 5 -AATGCTTCTAGGCGGACTATGA-3 , reverse: 5 -TTTCTGCGCAAGTTAGGTTTT-3 ) [24,25].…”

Section: Evaluation Of Alkaline Phosphatase Activitymentioning

confidence: 99%

Vitamin D Enhanced the Osteogenic Differentiation of Cell Spheroids Composed of Bone Marrow Stem Cells

et al. 2021

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“…For example, spheroids composed of MSCs or BMSCs cultured in an osteogenic medium containing BMP‐2 showed minimal enhancement of Runx2 expression compared to a spheroid group without inductive factors. [ 33 , 34 ] Additionally, the SOX9 expression of MSCs within the spheroid cultured in a chondrogenic medium containing TGF‐ β was similar to that of the control, indicating that the diffusional limitation should be addressed for effective differentiation of stem cells within spheroids. [ 35 ] As shown in Figure 1 , the gene expression in previous studies was similar to our results (PS/B and PS/T), where the spheroids were stimulated externally during culture in vitro while the growth factors immobilized on the fibers upregulated osteogenic and chondrogenic differentiation of hADSCs more effectively from the inside of BS and TS.…”

Section: Discussionmentioning

confidence: 98%

Directed Regeneration of Osteochondral Tissue by Hierarchical Assembly of Spatially Organized Composite Spheroids

Lee

Huh

et al. 2021

Advanced Science

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The use of engineered scaffolds or stem cells is investigated widely in the repair of injured musculoskeletal tissue. However, the combined regeneration of hierarchical osteochondral tissue remains a challenge due to delamination between cartilage and subchondral bone or difficulty in spatial control over differentiation of transplanted stem cells. Here, two types of composite spheroids are prepared using adipose‐derived stem cells (hADSCs) and nanofibers coated with either transforming growth factor‐β3 or bone morphogenetic growth factor‐2 for chondrogenesis or osteogenesis, respectively. Each type of spheroid is then cultured within a 3D‐printed microchamber in a spatially arranged manner to recapitulate the bilayer structure of osteochondral tissue. The presence of inductive factors regionally modulates in vitro chondrogenic or osteogenic differentiation of hADSCs within the biphasic construct without dedifferentiation. Furthermore, hADSCs from each spheroid proliferate and sprout and successfully connect the two layers mimicking the osteochondral interface without apertures. In vivo transplantation of the biphasic construct onto a femoral trochlear groove defect in rabbit knee joint results in 21.2 ± 2.8% subchondral bone volume/total volume and a cartilage score of 25.0 ± 3.7. The present approach can be an effective therapeutic platform to engineer complex tissue.

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“…Expression levels of various genes were tested to evaluate osteogenic capability including those of RUNX2, BSP, OCN and COL2A1 [ 23 , 28 ]. RUNX2 is known to be one of the key transcription factors related with osteogenic differentiation [ 29 ]. The secretion of BSP is considered to be the marker of the start of the osteogenic differentiation and BSP knockout led to little or no expression of osteogenic markers without mineralized colonization [ 30 , 31 ].…”

Section: Discussionmentioning

confidence: 99%

Evaluation of the Effects of Cuminum cyminum on Cellular Viability, Osteogenic Differentiation and Mineralization of Human Bone Marrow-Derived Stem Cells

Lee

Song

Park³

et al. 2021

Medicina

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Background and Objectives: Cuminum cyminum L. has long been used in the treatment of various diseases in multiple geographical regions. This study was performed to determine the effects of C. cyminum methanolic extract (CCT) on the cellular viability, alkaline phosphatase activity and mineralization of human mesenchymal stem cells. Materials and Methods: Bone marrow-derived stem cells were cultured in the presence of CCT at concentrations of 0, 0.001, 0.01, 0.1 and 1 μg/mL. Evaluations of cell morphology were performed on days 1, 3, 7 and 14. Cellular viability was evaluated on days 1, 3, 5 and 7. On the 7th and 14th day, alkaline phosphatase activity measurements and Alizarin red S staining were conducted to assess the osteogenic differentiation of stem cells. A real-time polymerase chain reaction was used to determine the expression levels of RUNX2, BSP, OCN, COL2A1 and β-catenin mRNAs. Results: Stem cells in the control group showed fibroblast-like morphology and the addition of CCT at 0.001, 0.01, 0.1 and 1 μg/mL did not generate noticeable changes in morphology compared with the untreated control group. The application of CCT did not produce significant changes in cellular viability or alkaline phosphatase activity compared with controls. Alizarin Red S staining was significantly increased with the application of CCT. Treatment with CCT increased the expressions of RUNX2, BSP and OCN. Conclusions: These results indicate that CCT enhanced the osteogenic differentiation of stem cells derived from bone marrow by regulating the expressions of RUNX2, BSP and OCN. Thus, the use of CCT may be applied to achieve beneficial effects on the mineralization of stem cells.

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Bone morphogenetic protein 2‑enhanced osteogenic differentiation of stem cell spheres by regulation of Runx2 expression

Cited by 12 publications

References 35 publications

Vitamin D Enhanced the Osteogenic Differentiation of Cell Spheroids Composed of Bone Marrow Stem Cells

Vitamin D Enhanced the Osteogenic Differentiation of Cell Spheroids Composed of Bone Marrow Stem Cells

Directed Regeneration of Osteochondral Tissue by Hierarchical Assembly of Spatially Organized Composite Spheroids

Evaluation of the Effects of Cuminum cyminum on Cellular Viability, Osteogenic Differentiation and Mineralization of Human Bone Marrow-Derived Stem Cells

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