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Enhanced differentiation of mesenchymal stromal cells by three-dimensional culture and azacitidine

View Article: PubMed Central - PubMed

ABSTRACT

Background: Mesenchymal stromal cells (MSCs) are useful for cell therapy because of their potential for multilineage differentiation. However, MSCs that are expanded in traditional two-dimensional (2D) culture systems eventually lose their differentiation abilities. Therefore, we investigated whether azacitidine (AZA) supplementation and three-dimensional culture (3D) could improve the differentiation properties of MSCs.

Methods: 2D- or 3D-cultured MSCs which were prepared according to the conventional or hanging-drop culture method respectively, were treated with or without AZA (1 µM for 72 h), and their osteogenic and adipogenic differentiation potential were determined and compared.

Results: AZA treatment did not affect the cell apoptosis or viability in both 2D- and 3D-cultured MSCs. However, compared to conventionally cultured 2D-MSCs, AZA-treated 2D-MSCs showed marginally increased differentiation abilities. In contrast, 3D-MSCs showed significantly increased osteogenic and adipogenic differentiation ability. When 3D culture was performed in the presence of AZA, the osteogenic differentiation ability was further increased, whereas adipogenic differentiation was not affected.

Conclusion: 3D culture efficiently promoted the multilineage differentiation of MSCs, and in combination with AZA, it could help MSCs to acquire greater osteogenic differentiation ability. This optimized culture method can enhance the therapeutic potential of MSCs.

No MeSH data available.


Related in: MedlinePlus

Representative photographs showing senescence-associated β-galactosidase (blue) staining of MSCs before and after exposure to AZA. The scale bar represents 100 µm.
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Figure 5: Representative photographs showing senescence-associated β-galactosidase (blue) staining of MSCs before and after exposure to AZA. The scale bar represents 100 µm.

Mentions: To identify any changes in MSC senescence induced by AZA treatment, an SA-β-gal assay was conducted with 2D-MSCs. A higher number of cells showed β-gal staining in the AZA-treated group than in the untreated group, indicating that AZA could induce MSC senescence (Fig. 5). The number of β-gal positive cells was 4-fold higher in AZA-treated cells than in 2D-MSCs (data not shown).


Enhanced differentiation of mesenchymal stromal cells by three-dimensional culture and azacitidine
Representative photographs showing senescence-associated β-galactosidase (blue) staining of MSCs before and after exposure to AZA. The scale bar represents 100 µm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5383582&req=5

Figure 5: Representative photographs showing senescence-associated β-galactosidase (blue) staining of MSCs before and after exposure to AZA. The scale bar represents 100 µm.
Mentions: To identify any changes in MSC senescence induced by AZA treatment, an SA-β-gal assay was conducted with 2D-MSCs. A higher number of cells showed β-gal staining in the AZA-treated group than in the untreated group, indicating that AZA could induce MSC senescence (Fig. 5). The number of β-gal positive cells was 4-fold higher in AZA-treated cells than in 2D-MSCs (data not shown).

View Article: PubMed Central - PubMed

ABSTRACT

Background: Mesenchymal stromal cells (MSCs) are useful for cell therapy because of their potential for multilineage differentiation. However, MSCs that are expanded in traditional two-dimensional (2D) culture systems eventually lose their differentiation abilities. Therefore, we investigated whether azacitidine (AZA) supplementation and three-dimensional culture (3D) could improve the differentiation properties of MSCs.

Methods: 2D- or 3D-cultured MSCs which were prepared according to the conventional or hanging-drop culture method respectively, were treated with or without AZA (1 µM for 72 h), and their osteogenic and adipogenic differentiation potential were determined and compared.

Results: AZA treatment did not affect the cell apoptosis or viability in both 2D- and 3D-cultured MSCs. However, compared to conventionally cultured 2D-MSCs, AZA-treated 2D-MSCs showed marginally increased differentiation abilities. In contrast, 3D-MSCs showed significantly increased osteogenic and adipogenic differentiation ability. When 3D culture was performed in the presence of AZA, the osteogenic differentiation ability was further increased, whereas adipogenic differentiation was not affected.

Conclusion: 3D culture efficiently promoted the multilineage differentiation of MSCs, and in combination with AZA, it could help MSCs to acquire greater osteogenic differentiation ability. This optimized culture method can enhance the therapeutic potential of MSCs.

No MeSH data available.


Related in: MedlinePlus