Generation of directly converted fuman osteoblasts that are free of exogenous gene and xenogenic protein.
Generation of directly converted fuman osteoblasts that are free of exogenous gene and xenogenic protein.
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产生不含外源基因和异种蛋白的直接转化的福曼成骨细胞。
DOI：
10.1002/jcb.25546
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发表时间：
2016
期刊：
影响因子：
4
通讯作者：
Mazda O.
中科院分区：
文献类型：
--
作者：
Yamamoto K;Sato Y;Honjo K;Ichioka H;Oseko F;Sowa Y;Yamamoto T;Kanamura N;Kishida T;Mazda O.
从人体细胞中培养成骨细胞可能是一种有效的骨移植治疗方法。最近，我们建立了一种通过逆转录病毒载体转导某些转录因子基因，将人成纤维细胞直接转化为成骨细胞的方法。然而，逆转录病毒载体介导的转导可能会导致受感染细胞形成肿瘤，因此非病毒基因转染方法可能更适合用于制备用于移植治疗的成骨细胞。在这里，我们构建了编码Oct 4、Osterix和L-Myc的质粒载体，它们是用于此目的的转录因子的适当组合。在用质粒转染的正常人成纤维细胞中诱导成骨细胞样表型，包括高碱性磷酸酶（ALP）活性、骨基质生成和成骨细胞特异性基因表达，然后在成骨培养基中培养。即使在不存在异种蛋白的情况下，也获得了质粒驱动的直接转化的成骨细胞（p-dOB）。质粒载体序列已从p-dOB中脱落。细胞移植到小鼠体内后原位形成钙化小体沉积。这些结果有力地表明，p-dOB可以实际用于针对骨疾病的新型基于细胞的疗法。J.细胞。117：2538-2545，2016.© 2016 Wiley Periodicals，Inc.
Generation of osteoblasts from human somatic cells may be applicable in an effective transplantation therapy against bone diseases. Recently we established a procedure to directly convert human fibroblasts into osteoblasts by transducing some transcription factor genes via retroviral vectors. However, retroviral vector‐mediated transduction may potentially cause tumor formation from the infected cells, thus a non‐viral gene transfection method may be more preferable for preparation of osteoblasts to be used for transplantation therapy. Here, we constructed a plasmid vector encoding Oct4, Osterix, and L‐Myc that were an appropriate combination of transcription factors for this purpose. Osteoblast‐like phenotypes including high alkaline phosphatase (ALP) activity, bone matrix production and osteoblast‐specific gene expression were induced in normal human fibroblasts that were transfected with the plasmid followed by culturing in osteogenic medium. The plasmid‐driven directly converted osteoblasts (p‐dOBs) were obtained even in the absence of a xenogenic protein. The plasmid vector sequence had fallen out of the p‐dOBs. The cells formed deposition of calcified bodies in situ after transplantation into mice. These results strongly suggest that p‐dOBs can be put into practical use for a novel cell‐based therapy against bone diseases. J. Cell. Biochem. 117: 2538–2545, 2016. © 2016 Wiley Periodicals, Inc.