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논문 기본 정보

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학술저널
저자정보
Seung Hyun L. Kim (Seoul National University) Sungwoo Cho (Seoul National University) Seoyeon Kim (Seoul National University) 권정후 (서울대학교) Jaeyoung Lee (Kangwon National University) Rachel H. Koh (Seoul National University) Ju Hyun Park (Kangwon National University) 이화진 (서울대학교) Tai Hyun Park (Seoul National University) Nathaniel S. Hwang (Seoul National University)
저널정보
한국생체재료학회 생체재료학회지 생체재료학회지 제26권 제3호
발행연도
2022.9
수록면
562 - 579 (18page)
DOI
https://doi.org/10.1186/s40824-022-00280-8

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Background: The number of patients suffering from osteoporosis is increasing as the elderly population increases. The demand for investigating bone regeneration strategies naturally arises. One of the approaches to induce bone regeneration is somatic cell transdifferentiation. Among the transcriptional regulators for transdifferentiation, octamer-binding transcription factor 4 (OCT4) is famous for its role in the regulation of pluripotency of stem cells. Bone morphogenetic protein 4 (BMP4) is another factor that is known to have a significant role in osteogenic differ entiation. Previous studies have achieved transdifferentiation of cells into osteoblasts using viral and plasmid deliver ies of these factors. Although these methods are efficient, viral and plasmid transfection have safety issues such as permanent gene incorporations and bacterial DNA insertions. Herein, we developed a cell penetrating protein-based strategy to induce transdifferentiation of endothelial cells into osteoblasts via nuclear delivery of OCT4 recombinant protein combined with the BMP4 treatment. For the nuclear delivery of OCT4 protein, we fused the protein with 30Kc19, a cell-penetrating and protein stabilizing protein derived from a silkworm hemolymph of Bombyx mori with low cytotoxic properties. This study proposes a promising cell-based therapy without any safety issues that existing transdifferentiation approaches had. Methods: OCT4-30Kc19 protein with high penetrating activities and stability was synthesized for a protein-based osteogenic transdifferentiation system. Cells were treated with OCT4-30Kc19 and BMP4 to evaluate their cellular penetrating activity, cytotoxicity, osteogenic and angiogenic potentials in vitro. The osteogenic potential of 3D cell spheroids was also analyzed. In addition, in vivo cell delivery into subcutaneous tissue and cranial defect model was performed. Results: OCT4-30Kc19 protein was produced in a soluble and stable form. OCT4-30Kc19 efficiently penetrated cells and were localized in intracellular compartments and the nucleus. Cells delivered with OCT4-30Kc19 protein com bined with BMP4 showed increased osteogenesis, both in 2D and 3D culture, and showed increased angiogenesis capacity in vitro. Results from in vivo subcutaneous tissue delivery of cell-seeded scaffolds confirmed enhanced osteogenic properties of transdifferentiated HUVECs via treatment with both OCT4-30Kc19 and BMP4. In addition, in vivo mouse cranial defect experiment demonstrated successful bone regeneration of HUVECs pretreated with both OCT4-30Kc19 and BMP4. Conclusions: Using a protein-based transdifferentiation method allows an alternative approach without utilizing any genetic modification strategies, thus providing a possibility for safer use of cell-based therapies in clinical applications.
#Transdifferentiation #Osteogenesis #Angiogenesis #HUVECs #Cell penetrating protein

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