Skeletal reconstruction is necessary in cases of bone defects created by tumors, trauma, and abnormalities. Regeneration of bone defects remains a critical problem, and current approaches are based on biocompatible scaffolds. Spheroids represent a simple 3D system since no supporting material is required for cell growth. Different techniques are used to generate spheroids, such as hanging drop, low-attachment plates, and magnetic nanoparticles. The idea of using magnetic nanoparticles is to cross-link through cell membrane overnight to create complex 3D cellular spheroid by using magnets to guide the cellular response. Herein, the current study aimed to achieve 3D human fetal osteoblast (hFOB) spheroid under magnetic levitation. Formation of 3D spheroid culture under magnetic levitation was evaluated by cell viability at 3, 7, and 14 days. Morphology of the 3D hFOB spheroid was analyzed by SEM and fluorescence microscopy and the differentiation towards mineralized lineage by ALP assay, qPCR, and alizarin red staining. The cell viability indicated that the 3D hFOB spheroid still viable after 14 days of culture. ALP assay, qPCR analysis expression of Col1, ALP, and Itg-β1 molecules, and calcium deposition with alizarin red showed a high level of bioactivity of the 3D hFOB spheroid. SEM images allowed the morphological analysis of the 3D microtissue-like spheroid with the presence of matrix deposition. These results indicate that magnetic levitation culture enables 3D stable osteoblast spheroids and could be a promising strategy for engineering application in the 3D construct in surgery regeneration of mineralized tissue.
Bone defects lead to the structural loss of normal architecture, and those in the field of bone tissue engineering are searching for new alternatives to aid bone regeneration. Dental pulp-mesenchymal stem cells (DP-MSC) could provide a promising alternative to repair bone defects, principally due to their multipotency and capacity to fabricate three-dimensional (3D) spheroids. The present study aimed to characterize the 3D DP-MSC microsphere and the osteogenic differentiation capacity potential cultured by a magnetic levitation system. To achieve this, the 3D DP-MSC microsphere was grown for 7, 14, and 21 days in an osteoinductive medium and compared to 3D human fetal osteoblast (hFOB) microspheres by examining the morphology, proliferation, osteogenesis, and colonization onto PLA fiber spun membrane. Our results showed good cell viability for both 3D microspheres with an average diameter of 350 μm. The osteogenesis examination of the 3D DP-MSC microsphere revealed the lineage commitment, such as the hFOB microsphere, as evidenced by ALP activity, the calcium content, and the expression of osteoblastic markers. Finally, the evaluation of the surface colonization exhibited similar patterns of cell-spreading over the fibrillar membrane. Our study demonstrated the feasibility of forming a 3D DP-MSC microsphere structure and the cell-behavior response as a strategy for the applications of bone tissue guiding.
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