Expandable progenitors from induced pluripotent stem cells

We read with interest the Review by Wu et al. (Induced pluripotent stem cells: at the heart of cardiovascular precision medicine. Nat. Rev. Cardiol. 13, 333–349; 2016)¹, which provided an insight into precision medicine aimed at successful drug screening and cardiovascular disease modelling. Regarding the differentiation process, we raise two limitations: heterogeneity, and the time-consuming and costly process. Variations exist not only among established clones of human induced pluripotent stem cells (hiPSCs)², but also among hiPSC-derived cardiomyocytes (hiPSC-CMs)³ as final products, suggesting large lot-to-lot variations during differentiation. Given the long duration of the differentiation process, the cytokines required during this process are costly.

Here, we propose that expandable progenitors could be meaningful for resolving these issues (Tables 1,2). When expandable progenitors are established, these cell lines will be self-renewable and suitable for cryopreservation. Furthermore, differentiation from these cell lines (not from iPSCs) will be possible, indicating that time and cost can be reduced, and less heterogeneous products can be obtained.

Table 1 Expandable progenitors without immortalization

Table 2 Expandable progenitors with immortalization

Several studies have demonstrated that expandable cardiac progenitor cells (CPCs) have been successfully captured (Table 1) without the procedure of immortalization. Lalit et al. have shown that five cardiac factors (mesoderm posterior protein 1, T-box transcription factor TBX5, transcription factor GATA-4, homeobox protein Nkx-2.5, and BRG1-associated factor 60C) transdifferentiate mouse fibroblasts into induced CPCs (iCPCs), which can be expanded >10¹⁵-fold by BIO (6-bromoindirubin-3′- oxime, a canonical Wnt activator) and leukaemia inhibitory factor⁴. iCPCs can differentiate into cardiomyocytes, smooth muscle cells, and endothelial cells. Zhang et al. demonstrated that short-term expression of Yamanaka factors with tyrosine-protein kinase JAK inhibitor JI1 and BACS (bone morphogenetic protein 4, activin, CHIR99021, and SU5402) transdifferentiate mouse fibroblasts into induced expandable CPCs (ieCPCs), which can be expanded >10¹⁰-fold by BACS, and can differentiate into cardiomyocytes, smooth muscle cells, and endothelial cells⁵. A study by Cao et al. showed that homogeneous cardiovascular progenitor cells (CVPCs) can be obtained from human pluripotent stem cells (hPSCs) with combined cytokines, and they can self-renew and expand >10⁷-fold⁶.

Another potential strategy for obtaining expandable progenitors involves transducing various genes (for example, MYC) for immortalization (Table 2). Birket et al. applied a doxycycline (Dox)-inducible MYC expression system on hiPSCs, which enabled robust expansion (>40 population doublings) of CPCs⁷. To induce differentiation, Dox was removed to turn off MYC transgene expression. Similar strategies have been reported for production of platelets^8,9 or erythrocytes¹⁰. Eto and colleagues established immortalized megakaryocyte progenitor cell lines from hPSC-derived haematopoietic progenitors through overexpression of MYC, BMI1, and BCL2L1 (Ref. 8). Collectively, without immortalization, more effective methods of progenitors expansion will be needed. Alternatively, we can apply the Dox-inducible system using transgenes for immortalization.