Although induced pluripotent stem cells hold promise as a potential source of osteoblasts for skeletal regeneration, the induction of pluripotency followed by directed differentiation into osteoblasts is time consuming and low yield. In contrast, direct lineage reprogramming without an intervening stem/progenitor cell stage would be a more efficient approach to generate osteoblasts. We screened combinations of osteogenic transcription factors and identified four factors, Runx2, Osx, Dlx5, and ATF4, that rapidly and efficiently reprogram mouse fibroblasts derived from 2.3 kb type I collagen promoter-driven green fluorescent protein (Col2.3GFP) transgenic mice into induced osteoblast cells (iOBs). iOBs exhibit osteoblast morphology, form mineralized nodules, and express Col2.3GFP and gene markers of osteoblast differentiation. The global transcriptome profiles validated that iOBs resemble primary osteoblasts. Genomewide DNA methylation analysis demonstrates that within differentially methylated loci, the methylation status of iOBs more closely resembles primary osteoblasts than mouse fibroblasts. We further demonstrate that Col2.3GFP+ iOBs have transcriptome profiles similar to GFP+ cells harvested from Col2.3GFP mouse bone chips. Functionally, Col2.3GFP+ iOBs form mineralized bone structures after subcutaneous implantation in immunodeficient mice and contribute to bone healing in a tibia bone fracture model. These findings provide an approach to derive and study osteoblasts for skeletal regeneration. © 2019 American Society for Bone and Mineral Research.

中文翻译：

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将小鼠成纤维细胞直接重编程为功能性成骨细胞。


尽管诱导多能干细胞有望作为骨骼再生成骨细胞的潜在来源，但诱导多能性随后定向分化为成骨细胞非常耗时且产量低。相比之下，无需干预干细胞/祖细胞阶段的直接谱系重编程将是生成成骨细胞的更有效方法。我们筛选了成骨转录因子的组合，并确定了四种因子：Runx2、Osx、Dlx5 和 ATF4，它们可以快速有效地将源自 2.3 kb I 型胶原启动子驱动的绿色荧光蛋白 (Col2.3GFP) 转基因小鼠的小鼠成纤维细胞重新编程为诱导的成骨转录因子。成骨细胞（iOB）。 iOB 表现出成骨细胞形态，形成矿化结节，并表达 Col2.3GFP 和成骨细胞分化的基因标记。全局转录组图谱证实 iOB 与原代成骨细胞相似。全基因组 DNA 甲基化分析表明，在差异甲基化位点内，iOB 的甲基化状态比小鼠成纤维细胞更类似于原代成骨细胞。我们进一步证明 Col2.3GFP+ iOB 的转录组谱与从 Col2.3GFP 小鼠骨芯片中收获的 GFP+ 细胞相似。从功能上讲，Col2.3GFP+ iOB 在皮下植入免疫缺陷小鼠体内后形成矿化骨结构，并有助于胫骨骨折模型中的骨愈合。这些发现提供了一种衍生和研究用于骨骼再生的成骨细胞的方法。 © 2019 美国骨与矿物质研究学会。