CMPs and GMPs can give rise to NK cells under the influence of stroma and HDC
| . | 1 cell/well . | 10 cells/well . | 1 cell/well in methylcellulose . | |||||||
|---|---|---|---|---|---|---|---|---|---|---|
| Cytokines only . | Stroma + cytokines . | HDC + stroma + cytokines . | Cytokines only . | Stroma + cytokines . | HDC + stroma + cytokines . | Erythroid colonies . | Myeloid colonies . | Mixed colonies . | Cloning efficiency . | |
| CMP | ||||||||||
| BG118 | 0 | 1.67 | 3.33 | 0 | 30 | 51.67 | 15 | 40 | 8.3 | 63.3 |
| BG119 | 3.33 | 35 | 27.5 | 8.33 | 100 | 100 | 35 | 46.7 | 10 | 91.7 |
| BG120 | 0 | NE | 17.5 | NE | 86.67 | 100 | 30 | 43.3 | 11.7 | 85 |
| BG121 | 0 | 3.33 | 8.33 | 1.67 | 68.33 | 55 | 26.7 | 23.3 | 11.7 | 61.7 |
| BG122 | 0 | 23.33 | 34.17 | 5 | 98.15 | 100 | 18.3 | 56.7 | 6.7 | 81.7 |
| Median | 0 | 13.3 | 17.5 | 3.3 | 86.7 | 100 | 26.7 | 43.3 | 10 | 81.7 |
| Average | 0.7 | 15.8 | 18.2 | 3.7 | 76.6 | 81.3 | 25 | 42 | 9.7 | 76.7 |
| GMP | ||||||||||
| BG118 | 0 | 8.33 | 15 | 0 | 50 | 36.7 | 0 | 55 | 0 | 55 |
| BG119 | 1.7 | 21.7 | 25 | 5 | 100 | 100 | 0 | 83.3 | 0 | 83.3 |
| BG120 | 0 | 31.7 | 29.2 | ND | 75 | 88 (5/well) | 0 | 61.7 | 0 | 61.7 |
| BG121 | 0 | 25 | 5 | 5 | 73.3 | 30 | 0 | 43.3 | 0 | 43.3 |
| BG122 | 1.7 | 15 | 22.3 | 16.7 | 88.1 | 95 | 0 | 70 | 0 | 70 |
| Median | 0 | 21.7 | 22.3 | 5 | 75 | 88 | 0 | 61.7 | 0 | 61.7 |
| Average | 0.7 | 20.3 | 19.3 | 6.7 | 77.3 | 69.9 | 0 | 62.7 | 0 | 62.7 |
| . | 1 cell/well . | 10 cells/well . | 1 cell/well in methylcellulose . | |||||||
|---|---|---|---|---|---|---|---|---|---|---|
| Cytokines only . | Stroma + cytokines . | HDC + stroma + cytokines . | Cytokines only . | Stroma + cytokines . | HDC + stroma + cytokines . | Erythroid colonies . | Myeloid colonies . | Mixed colonies . | Cloning efficiency . | |
| CMP | ||||||||||
| BG118 | 0 | 1.67 | 3.33 | 0 | 30 | 51.67 | 15 | 40 | 8.3 | 63.3 |
| BG119 | 3.33 | 35 | 27.5 | 8.33 | 100 | 100 | 35 | 46.7 | 10 | 91.7 |
| BG120 | 0 | NE | 17.5 | NE | 86.67 | 100 | 30 | 43.3 | 11.7 | 85 |
| BG121 | 0 | 3.33 | 8.33 | 1.67 | 68.33 | 55 | 26.7 | 23.3 | 11.7 | 61.7 |
| BG122 | 0 | 23.33 | 34.17 | 5 | 98.15 | 100 | 18.3 | 56.7 | 6.7 | 81.7 |
| Median | 0 | 13.3 | 17.5 | 3.3 | 86.7 | 100 | 26.7 | 43.3 | 10 | 81.7 |
| Average | 0.7 | 15.8 | 18.2 | 3.7 | 76.6 | 81.3 | 25 | 42 | 9.7 | 76.7 |
| GMP | ||||||||||
| BG118 | 0 | 8.33 | 15 | 0 | 50 | 36.7 | 0 | 55 | 0 | 55 |
| BG119 | 1.7 | 21.7 | 25 | 5 | 100 | 100 | 0 | 83.3 | 0 | 83.3 |
| BG120 | 0 | 31.7 | 29.2 | ND | 75 | 88 (5/well) | 0 | 61.7 | 0 | 61.7 |
| BG121 | 0 | 25 | 5 | 5 | 73.3 | 30 | 0 | 43.3 | 0 | 43.3 |
| BG122 | 1.7 | 15 | 22.3 | 16.7 | 88.1 | 95 | 0 | 70 | 0 | 70 |
| Median | 0 | 21.7 | 22.3 | 5 | 75 | 88 | 0 | 61.7 | 0 | 61.7 |
| Average | 0.7 | 20.3 | 19.3 | 6.7 | 77.3 | 69.9 | 0 | 62.7 | 0 | 62.7 |
CMP and GMP were double-sorted from UCB and deposited in 96-well plates at 1 cell/well or 10 cells/well (see “CD34+ cells isolation” for details). Culture conditions included cytokines alone (IL-3, IL-7, IL-15, SCF, and FLT-3L), cytokines and stroma, cytokines and stroma and HDC, or methylcellulose enriched with growth factors. Sixty replicates were performed for each donor (n = 5). At the end of the culture period each well was assayed for NK cells with the use of FACS. Shown is the percentage of wells that contained NK cells. Cells cultured in methylcellulose were examined for outgrowth of erythroid, myeloid, and mixed colonies. The median and average for all the experiments (n = 300 wells with the use of 5 separate donors for each condition) are shown.
NE indicates not evaluable; ND, not done.