3D Spheroid Cultivation Alters the Extent and Progression of Osteogenic Differentiation of Mesenchymal Stem/Stromal Cells Compared to 2D Cultivation

Abstract

Mesenchymal stem/stromal cells (MSC) are capable of progenitor cell fraction renewal or tissue-specific differentiation. These properties are maintained during in vitro cultivation, making them an interesting model system for testing biological and pharmacological compounds. Cell cultivation in 2D is commonly used to study cellular responses, but the 2D environment does not reflect the structural situation of most cell types. Therefore, 3D culture systems have been developed to provide a more accurate physiological environment in terms of cell–cell interactions. Since knowledge about the effects of 3D culture on specific differentiation processes is limited, we studied the effects on osteogenic differentiation and the release of factors affecting bone metabolism for up to 35 days and compared them with the effects in 2D culture. We demonstrated that the selected 3D model allowed the rapid and reliable formation of spheroids that were stable over several weeks and both accelerated and enhanced osteogenic differentiation compared with the 2D culture. Thus, our experiments provide new insights into the effects of cell arrangement of MSC in 2D and 3D. However, due to the different culture dimensions, various detection methods had to be chosen, which in principle limits the explanatory power of the comparison between 2D and 3D cultures.

Engineering three-dimensional bone macro-tissues by guided fusion of cell spheroids

“…This is associated with a reduction in cell viability over time confirming cell death in the core region (Figure 2D). The presence of a necrotic core has not been reported in other osteogenic spheroids (26,29,30) which may be due to cell type and size differences. These studies produced spheroids of approximately 600 µm; however, dRObs formed spheroids of >1 mm.…”

Co-culture of STRO1 + human gingival mesenchymal stem cells and human umbilical vein endothelial cells in 3D spheroids: enhanced in vitro osteogenic and angiogenic capacities

“…3D culture of MSCs allows them to form cell spheroids through self-assembly, better simulating cell-cell and cell-ECM interactions and forming more complex cellular morphologies and structures to mimic the in vivo environment ( Ryu et al, 2019 ). Compared to the 2D monolayer culture method, MSC spheroids cultivated in 3D demonstrate superior potential regarding cell vitality, multidirectional differentiation, and transplantability ( Duval et al, 2017 ; Wolff et al, 2023 ). This increased potential is a key reason behind this study’s adoption of a 3D MSC spheroids system for bone regeneration.…”

Advantages of Using 3D Spheroid Culture Systems in Toxicological and Pharmacological Assessment for Osteogenesis Research

“…Moreover, the interaction between calcium and phosphate was upregulated in bone 3D spheroids [ 63 ]. Calcium and phosphate are essential minerals involved in various cellular processes, including the mineralization and deposition of minerals such as hydroxyapatite [ 64 ].…”