Direct Effect of TN-C on Multilineage Colony Formation Assay of BM Cells and Nonadherent Cells Produced by LTBMCs
| Cells Examined . | Methylcellulose Medium . | Control . | TN-C DM . | |||
|---|---|---|---|---|---|---|
| TN-C (ng/mL) . | GF . | No. Colonies . | No. Erythroid Bursts . | No. Colonies . | No. Erythroid Bursts . | |
| BM Cells | 0 | + | 90 ± 5 | 6 ± 1 | 65 ± 3 | 4 ± 2 |
| 4 | + | 87 ± 2 | 6 ± 2 | NE | NE | |
| 20 | + | 89 ± 3 | 6 ± 2 | 67 ± 4 | 5 ± 3 | |
| 100 | + | 91 ± 4 | 6 ± 2 | 65 ± 4 | 4 ± 2 | |
| 0 | − | 3 ± 1 | 0 | 2 ± 1 | 0 | |
| 100 | − | 4 ± 2 | 0 | 2 ± 2 | 0 | |
| Nonadherent cells from LTBMCs (wk 3) | ||||||
| Treated with 20 ng/mL of TN-C | 0 | + | 175 ± 8 | 17 ± 1 | 107 ± 4 | 8 ± 0 |
| 20 | + | 181 ± 7 | 17 ± 1 | 109 ± 1 | 6 ± 1 | |
| Treated without TN-C | 0 | + | 134 ± 6 | 14 ± 1 | 83 ± 3 | 7 ± 1 |
| 20 | + | 136 ± 7 | 13 ± 2 | 80 ± 5 | 7 ± 2 | |
| Nonadherent cells from LTBMCs (wk 6) | ||||||
| Treated with 100 ng/mL of TN-C | 0 | − | 0 | 0 | 0 | 0 |
| 20 | − | 0 | 0 | 0 | 0 | |
| 100 | − | 0 | 0 | 0 | 0 | |
| 0 | + | 148 ± 2 | 18 ± 2 | 89 ± 3 | 9 ± 2 | |
| 20 | + | 146 ± 2 | 16 ± 2 | 88 ± 2 | 9 ± 1 | |
| 100 | + | 150 ± 5 | 17 ± 3 | 87 ± 4 | 10 ± 1 | |
| Treated without TN-C | 0 | − | 0 | 0 | 0 | 0 |
| 20 | − | 0 | 0 | 0 | 0 | |
| 100 | − | 0 | 0 | 0 | 0 | |
| 0 | + | 103 ± 7 | 12 ± 2 | 45 ± 1 | 5 ± 1 | |
| 20 | + | 105 ± 6 | 11 ± 1 | 45 ± 2 | 5 ± 1 | |
| 100 | + | 103 ± 2 | 10 ± 2 | 46 ± 4 | 5 ± 2 | |
| Cells Examined . | Methylcellulose Medium . | Control . | TN-C DM . | |||
|---|---|---|---|---|---|---|
| TN-C (ng/mL) . | GF . | No. Colonies . | No. Erythroid Bursts . | No. Colonies . | No. Erythroid Bursts . | |
| BM Cells | 0 | + | 90 ± 5 | 6 ± 1 | 65 ± 3 | 4 ± 2 |
| 4 | + | 87 ± 2 | 6 ± 2 | NE | NE | |
| 20 | + | 89 ± 3 | 6 ± 2 | 67 ± 4 | 5 ± 3 | |
| 100 | + | 91 ± 4 | 6 ± 2 | 65 ± 4 | 4 ± 2 | |
| 0 | − | 3 ± 1 | 0 | 2 ± 1 | 0 | |
| 100 | − | 4 ± 2 | 0 | 2 ± 2 | 0 | |
| Nonadherent cells from LTBMCs (wk 3) | ||||||
| Treated with 20 ng/mL of TN-C | 0 | + | 175 ± 8 | 17 ± 1 | 107 ± 4 | 8 ± 0 |
| 20 | + | 181 ± 7 | 17 ± 1 | 109 ± 1 | 6 ± 1 | |
| Treated without TN-C | 0 | + | 134 ± 6 | 14 ± 1 | 83 ± 3 | 7 ± 1 |
| 20 | + | 136 ± 7 | 13 ± 2 | 80 ± 5 | 7 ± 2 | |
| Nonadherent cells from LTBMCs (wk 6) | ||||||
| Treated with 100 ng/mL of TN-C | 0 | − | 0 | 0 | 0 | 0 |
| 20 | − | 0 | 0 | 0 | 0 | |
| 100 | − | 0 | 0 | 0 | 0 | |
| 0 | + | 148 ± 2 | 18 ± 2 | 89 ± 3 | 9 ± 2 | |
| 20 | + | 146 ± 2 | 16 ± 2 | 88 ± 2 | 9 ± 1 | |
| 100 | + | 150 ± 5 | 17 ± 3 | 87 ± 4 | 10 ± 1 | |
| Treated without TN-C | 0 | − | 0 | 0 | 0 | 0 |
| 20 | − | 0 | 0 | 0 | 0 | |
| 100 | − | 0 | 0 | 0 | 0 | |
| 0 | + | 103 ± 7 | 12 ± 2 | 45 ± 1 | 5 ± 1 | |
| 20 | + | 105 ± 6 | 11 ± 1 | 45 ± 2 | 5 ± 1 | |
| 100 | + | 103 ± 2 | 10 ± 2 | 46 ± 4 | 5 ± 2 | |
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 TN-C. No statistical differences were observed in colony formation of hematopoietic cells of TN-C DM or control mice treated with or without TN-C between methylcellulose cultures in the presence of increased concentration of TN-C and those without TN-C (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); NE, not examined.