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In this study, silk fibroin microfiber (mSF) was applied to assist spheroid assemblies of rBMSCs (rabbit bone marrow stem cells) (S/B). Alkaline hydrolysis was induced with different times and conditions to manufacture the various sizes of mSF. The mSF was incorporated in the rBMSC with different amounts to optimize proper content for spheroid assembly. The formation of the S/B was confirmed under optical microscopy and SEM. Proliferation and viability were characterized by CCK-8 and live/dead staining. Osteogenesis was analyzed with ALP (alkaline phosphatase) activity studies and real-time polymerase chain reaction. The S/B was successfully produced and displayed uniformly distributed cells and mSF with the presence of a gap in the structure. Proliferation and viability of the S/B significantly increased when compared to rBMSC spheroids (B), which is potentially due to the enhanced transportation of oxygen and nutrients to the cells located in the core region. Additionally, ALP activity and osteogenic markers were significantly upregulated in the optimized S/B under osteogenic media conditions. Overall, a hybrid-spheroid system with a simple 3D cell culture platform provides a potential approach for engineering therapeutic stem cells.

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Fabrication and Characterization of Silk Fibroin Microfiber-Incorporated Bone Marrow Stem Cell Spheroids to Promote Cell–Cell Interaction and Osteogenesis