CDKN2B upregulation prevents teratoma formation in multipotent fibromodulin-reprogrammed cells
Zhong Zheng, … , Kang Ting, Chia Soo
Zhong Zheng, … , Kang Ting, Chia Soo
Published July 15, 2019
Citation Information: J Clin Invest. 2019;129(8):3236-3251. https://doi.org/10.1172/JCI125015.
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Research Article Cell biology

CDKN2B upregulation prevents teratoma formation in multipotent fibromodulin-reprogrammed cells

Abstract

Tumorigenicity is a well-documented risk to overcome for pluripotent or multipotent cell applications in regenerative medicine. To address the emerging demand for safe cell sources in tissue regeneration, we established a novel, protein-based reprogramming method that does not require genome integration or oncogene activation to yield multipotent fibromodulin (FMOD)-reprogrammed (FReP) cells from dermal fibroblasts. When compared with induced pluripotent stem cells (iPSCs), FReP cells exhibited a superior capability for bone and skeletal muscle regeneration with markedly less tumorigenic risk. Moreover, we showed that the decreased tumorigenicity of FReP cells was directly related to an upregulation of cyclin-dependent kinase inhibitor 2B (CDKN2B) expression during the FMOD reprogramming process. Indeed, sustained suppression of CDKN2B resulted in tumorigenic, pluripotent FReP cells that formed teratomas in vivo that were indistinguishable from iPSC-derived teratomas. These results highlight the pivotal role of CDKN2B in cell fate determination and tumorigenic regulation and reveal an alternative pluripotent/multipotent cell reprogramming strategy that solely uses FMOD protein.

Authors

Zhong Zheng, Chenshuang Li, Pin Ha, Grace X. Chang, Pu Yang, Xinli Zhang, Jong Kil Kim, Wenlu Jiang, Xiaoxiao Pang, Emily A. Berthiaume, Zane Mills, Christos S. Haveles, Eric Chen, Kang Ting, Chia Soo

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Figure 1

FReP cells display myogenic differentiation potential in vitro.

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FReP cells display myogenic differentiation potential in vitro. (A) Myog...
(A) Myogenic markers ACTN, MYOD, ACTA1, myosin, and desmin were found in FReP cells after cultivation using the established in vitro 2-stage skeletal myogenic differentiation protocol. Under the same differentiation conditions, parental unreprogrammed BJ fibroblasts and FReP-basal cells were used as negative controls, while retrovirus-mediated BJ-iPSCs and differentiated human satellite cells were used as positive controls. White arrowheads indicate the fusing myogenically differentiated cells; scale bar: 100 μm. (B) A PCR array revealed significantly upregulated myogenesis-related genes in FReP cells during the 3-week in vitro myogenic differentiation period. Unprocessed original Ct data are shown in Supplemental Table 1. n = 3 independent experiments. (C) The creatine kinase activity assay suggests that, after undergoing myogenic differentiation, FReP cells have biological activities similar to those of human satellite cells in vitro. **P < 0.005 (analyzed by 1-tailed Mann-Whitney and Kruskal-Wallis ANOVA tests); n = 6.