Zhou Ping, Shi Jia-Min, Song Jing-E, Han Yu, Li Hong-Jiao, Song Ya-Meng, Feng Fang, Wang Jian-Lin, Zhang Rui, Lan Feng
School and Hospital of Stomatology, Lanzhou University, No.222 Tianshui South Road, Chengguan District, Lanzhou, 730000, Gansu Province, People's Republic of China.
College of Life Sciences, Lanzhou University, No.222 Tianshui South Road, Chengguan District, Lanzhou, 730000, Gansu Province, People's Republic of China.
Stem Cell Res Ther. 2021 Jan 7;12(1):41. doi: 10.1186/s13287-020-02085-9.
Derivation of osteoblast-like cells from human pluripotent stem cells (hPSCs) is a popular topic in bone tissue engineering. Although many improvements have been achieved, the low induction efficiency because of spontaneous differentiation hampers their applications. To solve this problem, a detailed understanding of the osteogenic differentiation process of hPSCs is urgently needed.
Monolayer cultured human embryonic stem cells and human-induced pluripotent stem cells were differentiated in commonly applied serum-containing osteogenic medium for 35 days. In addition to traditional assays such as cell viability detection, reverse transcription-polymerase chain reaction, immunofluorescence, and alizarin red staining, we also applied studies of cell counting, cell telomerase activity, and flow cytometry as essential indicators to analyse the cell type changes in each week.
The population of differentiated cells was quite heterogeneous throughout the 35 days of induction. Then, cell telomerase activity and cell cycle analyses have value in evaluating the cell type and tumourigenicity of the obtained cells. Finally, a dynamic map was made to integrate the analysis of these results during osteogenic differentiation of hPSCs, and the cell types at defined stages were concluded.
Our results lay the foundation to improve the in vitro osteogenic differentiation efficiency of hPSCs by supplementing with functional compounds at the desired stage, and then establishing a stepwise induction system in the future.
从人多能干细胞（hPSC）中诱导生成类成骨细胞是骨组织工程领域的一个热门话题。尽管已取得诸多进展，但自发分化导致的低诱导效率阻碍了其应用。为解决这一问题，迫切需要深入了解hPSC的成骨分化过程。
将单层培养的人胚胎干细胞和人诱导多能干细胞在常用的含血清成骨培养基中分化35天。除了细胞活力检测、逆转录聚合酶链反应、免疫荧光和茜素红染色等传统检测方法外，我们还应用细胞计数、细胞端粒酶活性和流式细胞术研究作为重要指标，分析每周的细胞类型变化。
在35天的诱导过程中，分化细胞群体具有高度异质性。此外，细胞端粒酶活性和细胞周期分析对于评估所获细胞的类型和致瘤性具有重要价值。最后，绘制了动态图谱以整合hPSC成骨分化过程中这些结果的分析，并总结了特定阶段的细胞类型。
我们的研究结果为通过在期望阶段补充功能性化合物来提高hPSC体外成骨分化效率奠定了基础，进而为未来建立逐步诱导体系提供了依据。
