A real-time pluripotency reporter for the long-term and real-time monitoring of pluripotency changes in induced pluripotent stem cells

作者：Shen, Hong-Fen^*; Li, Yong-Long; Huang, Shi-Hao; Xia, Jia-Wei; Yao, Zhi-Fang; Xiao, Gao-Fang; Zhou, Ying; Li, Ying-Chun; Shi, Jun-Wen; Lin, Xiao-Lin; Zhao, Wen-Tao; Sun, Yan; Tian, Yu-Guang^*; Jia, Jun-Shuang^*; Xiao, Dong^*

来源：Aging-US, 2022, 14(10): 4445-4458.

• induced pluripotent stem (iPS) cells
• cancer cell reprogramming
• cancer therapy
• Oct4-EGFP transgene
• pluripotency
• reporter system

To master the technology of reprogramming mouse somatic cells to induced pluripotent stem cells (iPSCs), which will lay a good foundation for setting up a technology platform on reprogramming human cancer cells into iPSCs. Mouse iPSCs (i.e., Oct4-GFP miPSCs) was successfully generated from mouse embryonic fibroblasts (MEFs) harboring Oct4-EGFP transgene by introducing four factors, Oct4, Sox2, c-Myc and Klf4, under mESC (Murine embryonic stem cells) culture conditions. Oct4-GFP miPSCs were similar to mESCs in morphology, proliferation, mESC-specific surface antigens and gene expression. Additionally, Oct4-GFP miPSCs could be cultured in suspension to form embryoid bodies (EBs) and differentiate into cell types of the three germ layers in vitro. Moreover, Oct4-GFP miPSCs could develop to teratoma and chimera in vivo. Unlike cell cycle distribution of MEFs, Oct4-GFP miPSCs are similar to mESCs in the cell cycle structure which consists of higher S phase and lower G1 phase. More importantly, our data demonstrated that MEFs harboring Oct4-EGFP transgene did not express GFP, until they were reprogrammed to the pluripotent stage (iPSCs), while the GFP expression was progressively lost when these pluripotent Oct4-GFP miPSCs exposed to EB-mediated differentiation conditions, suggesting the pluripotency of Oct4-GFP miPSCs can be real-time monitored over long periods of time via GFP assay. Altogether, our findings demonstrate that Oct4-GFP miPSC line is successfully established, which will lay a solid foundation for setting up a technology platform on reprogramming cancer cells into iPSCs. Furthermore, this pluripotency reporter system permits the long-term real-time monitoring of pluripotency changes in a live single-cell, and its progeny.

• 单位
  南方医科大学; 6; 中山大学