Silencing of p53 and CDKN1A establishes sustainable immortalized megakaryocyte progenitor cells from human iPSCs

• DOI: 10.1016/j.stemcr.2021.11.001
• Publication Date: 2021-12-02T15:30:19Z
• Authors: Masamitsu Sone; Sou Nakamura; Sachiko Umeda; Harumi Ginya; Motohiko Oshima; Maria Alejandra K...; Sudip Kumar Paul; Kanae Hashimoto; Emiri Nakamura; Yasuo Harada; Kyoko Tsujimura; Atsunori Saraya; Tomoyuki Yamaguchi; Naoshi Sugimoto; Akira Sawaguchi; Atsushi Iwama; Koji Eto; Naoya Takayama
• Keywords: p53; Blood Platelets; Cyclin-Dependent Kinase Inhibitor p21; Induced Pluripotent Stem Cells; bcl-X Protein; regenerative medicine; immortalization; Cell Line; megakaryocyte; Proto-Oncogene Proteins c-myc; 03 medical and health sciences; Report; Humans; Gene Silencing; Cell Proliferation; Megakaryocyte Progenitor Cells; platelet; Polycomb Repressive Complex 1; 460; 0303 health sciences; iPSC; cyclin-dependent kinase inhibitor; Clone Cells; Up-Regulation; CDKN1A; HEK293 Cells; Gene Knockdown Techniques; Tumor Suppressor Protein p53

Platelet transfusions are critical for severe thrombocytopenia but depend on blood donors. The shortage of donors and the potential of universal HLA-null platelet products have stimulated research on the ex vivo differentiation of human pluripotent stem cells (hPSCs) to platelets. We recently established expandable immortalized megakaryocyte cell lines (imMKCLs) from hPSCs by transducing MYC, BMI1, and BCL-XL (MBX). imMKCLs can act as cryopreservable master cells to supply platelet concentrates. However, the proliferation rates of the imMKCLs vary with the starting hPSC clone. In this study, we reveal from the gene expression profiles of several MKCL clones that the proliferation arrest is correlated with the expression levels of specific cyclin-dependent kinase inhibitors. Silencing CDKN1A and p53 with the overexpression of MBX was effective at stably inducing imMKCLs that generate functional platelets irrespective of the hPSC clone. Collectively, this improvement in generating imMKCLs should contribute to platelet industrialization and platelet biology.