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28 July 2022
Kenji Kitajima, Takahiko Hara (Stem Cell Project) and colleagues published an article, “An interferon-γ/FLT3 axis positively regulates hematopoietic progenitor cell expansion from human pluripotent stem cells” in Stem Cells.

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Identification of a key factor for the efficient production of hematopoietic progenitor cells from human induced pluripotent stem cells

Stem cell project team in our institute revealed that in vitro production of hematopoietic progenitor cells from human induced pluripotent stem cells was enhanced by interferon-γ/FLT3 pathway. This study was published as an original research article in US academic journal.

<Title of the paper>
An interferon-γ/FLT3 axis positively regulates hematopoietic progenitor cell expansion from human pluripotent stem cells
<Journal>
Stem Cells
DOI:10.1093/stmcls/sxac052

Background

Human induced pluripotent stem cells (hiPSCs; Figure 1) could be differentiated into various cell types when cultured in specialized conditions. These differentiated cells are useful for regenerative medicine, drug discovery, and pathological studies. It has been known that hematopoietic stem/progenitor cells (HSPCs) could be obtained from hiPSCs by several culture methods. An important feature of HSPCs is bone marrow repopulating activity [1], however, the repopulating activity of hiPSC-derived HSPCs were significantly low. To obtain highly-potent HSPCs from hiPSCs, great efforts have been made in the world.

Key findings

HSPCs are originated from aorta/gonad/mesonephros (AGM) region during mammalian embryogenesis. By mimicking the embryonic HSPC developmental process by hiPSC-derived organoids [2] (Figure 1), we could obtain hematopoietic cells expressing HSPC markers from hiPSCs. However these HSPC-like cells showed low repopulating activity when transplanted into conditioned immune-compromised mice.

Figure1
Figure 1 Human induced pluripotent stem cells (hiPSCs; left) and organoid obtained from hiPSCs (right). When hiPSCs were cultured in a specialized condition, these cells autonomously formed organoid-like structures.

Subsequently these HSPCs were thoroughly investigated to seek their fundamental defects. In bone marrow, HSPCs are differentiated into mature myeloid cells via hematopoietic progenitor cells (HPCs) expressing CD38 on their cell surfaces (Figure 2). These CD38+ HPSCs exhibit highly proliferative potential, hence the small number of HSPCs could give rise to huge number of myeloid cells via multiple cell division steps at the CD38+ HPC stage (Figure 2). Notably, we found that hiPSC-derived HPSCs were differentiated into myeloid cells without or rapidly passing through CD38+ HPC stage (Figure 2). Possibly, this unusual defect would be linked with their low engraftable activity.

Figure2
Figure 2 The difference of hematopoietic differentiation between hematopoietic stem/progenitor cell (HSPC) in bone marrow (BM-HSPC) and that from human induced pluripotent stem cells (iPS-HPSC). BM-HSPC is differentiated into myeloid cells via transient amplification of CD38+ HPCs while iPS-HSPC is rapidly differentiated into myeloid cells.

We next found that the expression of FLT3 was significantly low in the iPS-derived HSPCs when compared with cord blood HSPCs. FLT3 encodes a receptor tyrosine kinase expressed on the cell surface. FLT3 is activated by FLT3L, a cytokine supporting cell proliferation of HSPCs. Thus FLT3 is required for the transduction of mitogenic stimulation from FLT3L. Therefore we examined the effects of enforced FLT3 expression on hematopoietic induction from hiPSCs in the presence of FLT3L. Consequently, FLT3 expression unaffected the number of HSPCs while CD38+ HPCs were significantly increased. Therefore the absence of CD38+ HPCs was caused by the failed expression of FLT3.

By searching several cytokines/chemicals, we found that interferon-γ (IFN-γ) could induce FLT3 expression in hiPSC-derived HSPCs (Figure 3). Furthermore IFN-γ treatment drastically increased CD38+ HPCs from hiPSCs (Figure 3). Therefore it would be plausible that inflammatory signals, such as IFN-γ, is required for establishment of adult-type hematopoietic hierarchy including CD38+ HPCs. However, FLT3 and IFN-γ did not enhance bone marrow populating ativity of hiPSC-derived HSPCs. Thereby other cue(s) would be required to obtain highly-potent HSPCs from hiPSCs. On the other hand, we found that a chemical compound UM171, known to support ex vivo amplification of cord blood HSPCs, was effective for in vitro expansion of hiPSC-derived HSPCs. Thus, UM171-mediated in vitro expansion of hiPSC-derived HSPCs would be useful as a stem cell source for various hematopoietic cells.

Figure3
Figure 3 The expression and fucntion of FLT3 in hematopoietic stem/progenitor cell (HSPC) from human induced pluripotent stem cells (iPS-HPSC). The expression of FLT3 in iPS-HSPCs is induced by interferom-γ (IFN-γ). The FLT3 expression allows FLT3L-induced signal and induces CD38+ hematopoietuc progenitor cells (HPCs).

〈Glossary〉

[1]
The activity is experimentally defined as the continuous supply of circulating multi-lineage mature hematopoietic cells for long-term when transplanted into conditioned immune-compromised mice.
[2]
Three dimensional, mini-organ-like cell aggregates. These aggreagates are formed by culturing stem cells in appropriate and specialized conditions.

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