Direct Effect of ECM Component on Multilineage Colony Formation Assay of BM Cells and Nonadherent Cells Produced by LTBMCs
| Cells Examined . | Methylcellulose Medium . | Control . | TN-C DM . | |||
|---|---|---|---|---|---|---|
| ECM (μg/mL) . | GF . | No. Colonies . | No. Erythroid Bursts . | No. Colonies . | No. Erythroid Bursts . | |
| BM cells | ||||||
| FN | 0 | + | 78 ± 4 | 6 ± 1 | 55 ± 3 | 4 ± 0 |
| 2 | + | 78 ± 5 | 6 ± 2 | 57 ± 2 | 4 ± 1 | |
| 10 | + | 75 ± 3 | 6 ± 0 | 52 ± 3 | 4 ± 2 | |
| 0 | − | 1 ± 0 | 0 | 2 ± 1 | 0 | |
| 10 | − | 2 ± 1 | 0 | 1 ± 0 | 0 | |
| HS | 0 | + | 77 ± 4 | 7 ± 1 | 55 ± 3 | 4 ± 0 |
| 5 | + | 78 ± 2 | 7 ± 2 | 55 ± 5 | 4 ± 0 | |
| 50 | + | 76 ± 0 | 7 ± 1 | 54 ± 2 | 5 ± 1 | |
| 0 | − | 2 ± 1 | 0 | 2 ± 1 | 0 | |
| 50 | − | 1 ± 1 | 0 | 2 ± 2 | 0 | |
| Nonadherent cells from LTBMCs (wk 3) | ||||||
| Treated with 10 μg/mL of FN | ||||||
| FN | 0 | + | 143 ± 3 | 11 ± 1 | 118 ± 5 | 8 ± 1 |
| 10 | + | 145 ± 6 | 11 ± 3 | 110 ± 6 | 6 ± 1 | |
| 10 | − | 2 ± 1 | 0 | 2 ± 2 | 0 | |
| Treated without FN | ||||||
| FN | 0 | + | 122 ± 3 | 14 ± 1 | 88 ± 3 | 7 ± 1 |
| 10 | + | 118 ± 7 | 13 ± 2 | 85 ± 5 | 7 ± 2 | |
| Treated with 50 μg/mL of HS | ||||||
| HS | 0 | + | 154 ± 4 | 12 ± 2 | 112 ± 5 | 9 ± 2 |
| 50 | + | 148 ± 6 | 11 ± 2 | 110 ± 5 | 9 ± 1 | |
| 50 | − | 2 ± 0 | 0 | 1 ± 1 | 0 | |
| Treated without HS | ||||||
| HS | 0 | + | 122 ± 2 | 12 ± 2 | 89 ± 3 | 9 ± 2 |
| 50 | + | 125 ± 6 | 11 ± 1 | 92 ± 4 | 8 ± 1 | |
| Cells Examined . | Methylcellulose Medium . | Control . | TN-C DM . | |||
|---|---|---|---|---|---|---|
| ECM (μg/mL) . | GF . | No. Colonies . | No. Erythroid Bursts . | No. Colonies . | No. Erythroid Bursts . | |
| BM cells | ||||||
| FN | 0 | + | 78 ± 4 | 6 ± 1 | 55 ± 3 | 4 ± 0 |
| 2 | + | 78 ± 5 | 6 ± 2 | 57 ± 2 | 4 ± 1 | |
| 10 | + | 75 ± 3 | 6 ± 0 | 52 ± 3 | 4 ± 2 | |
| 0 | − | 1 ± 0 | 0 | 2 ± 1 | 0 | |
| 10 | − | 2 ± 1 | 0 | 1 ± 0 | 0 | |
| HS | 0 | + | 77 ± 4 | 7 ± 1 | 55 ± 3 | 4 ± 0 |
| 5 | + | 78 ± 2 | 7 ± 2 | 55 ± 5 | 4 ± 0 | |
| 50 | + | 76 ± 0 | 7 ± 1 | 54 ± 2 | 5 ± 1 | |
| 0 | − | 2 ± 1 | 0 | 2 ± 1 | 0 | |
| 50 | − | 1 ± 1 | 0 | 2 ± 2 | 0 | |
| Nonadherent cells from LTBMCs (wk 3) | ||||||
| Treated with 10 μg/mL of FN | ||||||
| FN | 0 | + | 143 ± 3 | 11 ± 1 | 118 ± 5 | 8 ± 1 |
| 10 | + | 145 ± 6 | 11 ± 3 | 110 ± 6 | 6 ± 1 | |
| 10 | − | 2 ± 1 | 0 | 2 ± 2 | 0 | |
| Treated without FN | ||||||
| FN | 0 | + | 122 ± 3 | 14 ± 1 | 88 ± 3 | 7 ± 1 |
| 10 | + | 118 ± 7 | 13 ± 2 | 85 ± 5 | 7 ± 2 | |
| Treated with 50 μg/mL of HS | ||||||
| HS | 0 | + | 154 ± 4 | 12 ± 2 | 112 ± 5 | 9 ± 2 |
| 50 | + | 148 ± 6 | 11 ± 2 | 110 ± 5 | 9 ± 1 | |
| 50 | − | 2 ± 0 | 0 | 1 ± 1 | 0 | |
| Treated without HS | ||||||
| HS | 0 | + | 122 ± 2 | 12 ± 2 | 89 ± 3 | 9 ± 2 |
| 50 | + | 125 ± 6 | 11 ± 1 | 92 ± 4 | 8 ± 1 | |
Colonies (>50 cells) were counted at day 14. Visualized hemoglobinized colonies were counted as erythroid bursts (BFU-E). Data represented as mean colony number ± SD per 5 × 104nonadherent cells from triplicate cultures. Progenitor cell assays were performed using BM cells before introduction of LTBMCs and nonadherent cells from LTBMCs treated with or without ECM component. No statistical differences were observed in colony formation of hematopoietic cells of TN-C DM or control mice treated with or without ECM between methylcellurose cultures in the presence of increased concentration of ECM and those without ECM component (P > .05, Student'st-test).
Abbreviations: TN-C DM, tenascin-C–deficient mutant mice; GF, growth factors containing IL-3 (1% SCCM) and erythropoietin (2 U/mL).