Intracellular Delivery of Recombinant RUNX2 Facilitated by Cell-Penetrating Protein for the Osteogenic Differentiation of hMSCs

Abstract

Human mesenchymal stem cells (hMSCs) are a commonly used cell source for cell therapy and tissue engineering because of their easy accessibility and multipotency. Runt-related transcription factor 2 (RUNX2) is a master regulator of the osteogenic commitment of hMSCs. Either recombinant plasmid delivery or viral transduction has been utilized to activate RUNX2 gene expression for effective hMSC differentiation. In this study, recombinant RUNX2 fused with cell-penetrating 30Kc19 alpha protein (30Kc19 alpha-RUNX2) was delivered into hMSCs for osteogenic commitment. Fusion of recombinant RUNX2 with 30Kc19 alpha resulted in successful delivery of the protein into cells and enhanced soluble expression of the protein. Intracellular delivery of the 30Kc19 alpha-RUNX2 fusion protein enhanced the osteogenic differentiation of hMSCs in vitro. 30Kc19 alpha-RUNX2 treatment resulted in increased ALP accumulation and elevated calcium deposition. Finally, implantation of hMSCs treated with 30Kc19 alpha-RUNX2 showed osteogenesis via cell delivery into the subcutaneous tissue and bone regeneration in a cranial defect mouse model. Therefore, we suggest that 30Kc19 alpha-RUNX2, an osteoinductive recombinant protein, is an efficient tool for bone tissue engineering.