Table 3.
Derivation of natural killer cells from human pluripotent stem cells.
| Author | Stem cell type | Cell type derived | Time line in days | Culture conditions | Remarks |
|---|---|---|---|---|---|
| Hermanson et al.[73] | PSCs/iPSCs | NK cells | 32+ | Cells were cultured in 96-well plate format. EBs were derived prior to differentiation to NK cells. D0–D10: EBs inducted with SCF medium + VEGF + BMP4, etc., for hematopoietic differentiation to generate CD34+/CD43 NK cells. D11–D32+: EBs further differentiated into NK cells using IL-3 + IL-15 + IL-7 + SCF + FLT3 ligand. Medium change was done weekly for 28–32 days and harvested for APC expansion. |
KIR, CD16, NKp46, NKG2D and NKG2A analyzed with flow cytometry. |
| Woll et al.[69] | ESC | NK cells | 35+ | D0–D17: hESCs co-cultured with murine bone marrow stromal cell line M210-B4 along with medium containing RPMI 1640 + 15% FBS + glutamine + NEAA + penicillin/streptomycin + β-mercaptoethanol for next 17 days. D17–D20: CD34+CD45+ double-positive cells were isolated and single-cell suspension prepared. Used aforementioned medium (refer to article for more details). D20–D35+: Separated cells were co-cultured with murine fetal liver-derived stromal cell line AFT024 in medium containing DMEM/Ham's F12 + human serum AB (heat-inactivated) + L-glutamine + penicillin/streptomycin + sodium selenite + ethanolamine + β-mercaptoethanol + ascorbic acid + IL-3 (for 1 week) + IL-15 + SCF + IL-7 + tyrosine kinase 3 ligand for next 30–35 days. Medium was replenished every 5–7 days. |
M210-B4 cells found more efficiently compared with S17 cells. CD56/CD45 NK cells were found mostly when analyzed with flow cytometry |
| Zeng et al,[74] | iPSCs | NK cells | 40–47 | D0–D11: hPSCs were seeded on stromal feeder cells (OP9) in presence of α-MEM and 20% FBS for hematopoietic differentiation. D12–D47: Early hematopoietic progenitors were harvested and co-cultured with modified OP9 cell line expressing Notch ligand DLL1 first to increase hematopoietic progenitors and then driven toward lymphoid lineages. |
Clonality assays to detect rearranged TCRβ and TCRγ chain genes in PBC/iPSC lines were performed. Phenotypic characterization was carried out by flow cytometry and immunocytochemistry. |
| Zhu and Kaufman [75] | iPSCs | NK cells | D0–D7; D8–D14; D15–D43 |
Feeder-free adapted ESCs/iPSCs with ROCKi for EB formation. Hematopoietic cell (CD34+) derivation and NK cell differentiation under feeder-free conditions in the presence of IL-3, IL-6, IL-16, SCF and FLT3. NK cell expansion was carried out under feeder-free conditions in the presence of IL-2 and aAPCs. |
Phenotypic and functional characterization of hESC/iPSC-derived NK cells was done by flow cytometry. |
aAPCs, artificial antigen-presenting cells; APC, antigen-presenting cell; DMEM, Dulbecco's Modified Eagle's Medium; EB, embryoid body; ESCs, embryonic stem cells; FBS, fetal bovine serum; hESCs, human embryonic stem cells; MEM, Minimum Essential Medium; NEAA, non-essential amino acid; PBC, primary biliary cirrhosis; ROCKi, Rho kinase inhibitor; SCF, stem cell factor; VEGF, vascular endothelial growth factor.