Conditions for development of chimeric mice and colony formation from muscle of chimeric mice
| Experiment no. . | Chimeric mice . | Cell culture . | |||||||
|---|---|---|---|---|---|---|---|---|---|
| Age of donor mice (week) . | Donor BM cells transplanted3-150 . | No. recipients . | Time of harvest after BMT (month) . | No. muscle cells harvested . | % Donor cells in recipients' PB . | Cells cultured . | Total CFU-Cs . | % Donor cells in pooled colonies3-151 . | |
| 1 | 10 | CD34− | 8 | 7 | 5.0 × 105 | 25 ± 20 | Muscle 4 × 104 | 51 | 6 |
| 2.0 × 105 | BM 4 × 104 | 150 | 26 | ||||||
| 2 | 2 | CD34+ | 3 | 11 | 1.1 × 105 | 52 ± 17 | Muscle 2 × 104 | 46 | 18 |
| 4.0 × 104 | BM 2 × 104 | 43 | 12 | ||||||
| 3 | 10 | CD34− | 10 | 3 | 2.8 × 105 | 38 ± 12 | Muscle 2 × 104 | 67 | 44 |
| 2.0 × 105 | BM 4 × 104 | 106 | 55 | ||||||
| 4 | 19 | CD34− | 5 | 12 | 8.0 × 105 | 37 ± 17 | Muscle 4 × 104 | 21 | 63 |
| 2.9 × 105 | BM 4 × 104 | 180 | 62 | ||||||
| 5 | 10 | Unfractionated | 7 | 3 | 6.1 × 105 | 94 ± 2 | Muscle 4 × 104 | 34 | 100 |
| 2.0 × 1063-152 | BM 6 × 1033-153 | 17 | 100 | ||||||
| 6 | 5 | CD34+ | 6 | 10 | 4.5 × 105 | 47 ± 10 | |||
| 5.0 × 104 | |||||||||
| 7 | 9 | Unfractionated | 10 | 6 | 1.7 × 106 | 99 ± 0 | |||
| 2.0 × 1063-152 | |||||||||
| 8 | 10 | Unfractionated | 8 | 10 | 1.2 × 106 | 99 ± 1 | |||
| 2.0 × 1063-152 | |||||||||
| Experiment no. . | Chimeric mice . | Cell culture . | |||||||
|---|---|---|---|---|---|---|---|---|---|
| Age of donor mice (week) . | Donor BM cells transplanted3-150 . | No. recipients . | Time of harvest after BMT (month) . | No. muscle cells harvested . | % Donor cells in recipients' PB . | Cells cultured . | Total CFU-Cs . | % Donor cells in pooled colonies3-151 . | |
| 1 | 10 | CD34− | 8 | 7 | 5.0 × 105 | 25 ± 20 | Muscle 4 × 104 | 51 | 6 |
| 2.0 × 105 | BM 4 × 104 | 150 | 26 | ||||||
| 2 | 2 | CD34+ | 3 | 11 | 1.1 × 105 | 52 ± 17 | Muscle 2 × 104 | 46 | 18 |
| 4.0 × 104 | BM 2 × 104 | 43 | 12 | ||||||
| 3 | 10 | CD34− | 10 | 3 | 2.8 × 105 | 38 ± 12 | Muscle 2 × 104 | 67 | 44 |
| 2.0 × 105 | BM 4 × 104 | 106 | 55 | ||||||
| 4 | 19 | CD34− | 5 | 12 | 8.0 × 105 | 37 ± 17 | Muscle 4 × 104 | 21 | 63 |
| 2.9 × 105 | BM 4 × 104 | 180 | 62 | ||||||
| 5 | 10 | Unfractionated | 7 | 3 | 6.1 × 105 | 94 ± 2 | Muscle 4 × 104 | 34 | 100 |
| 2.0 × 1063-152 | BM 6 × 1033-153 | 17 | 100 | ||||||
| 6 | 5 | CD34+ | 6 | 10 | 4.5 × 105 | 47 ± 10 | |||
| 5.0 × 104 | |||||||||
| 7 | 9 | Unfractionated | 10 | 6 | 1.7 × 106 | 99 ± 0 | |||
| 2.0 × 1063-152 | |||||||||
| 8 | 10 | Unfractionated | 8 | 10 | 1.2 × 106 | 99 ± 1 | |||
| 2.0 × 1063-152 | |||||||||
After lethal irradiation of the recipients, designated numbers of donor (Ly-5.1) BM cells were injected into recipient (Ly-5.2) mice along with 2 × 105 unfractionated marrow cells from Ly-5.2 mice as radioprotective cells. In experiments 1 through 5, cells were harvested from the muscle and BM of chimeric mice 3 to 12 months after transplantation, and the indicated numbers of unfractionated cells were cultured in the presence of thrombopoietin, steel factor, erythropoietin, IL-11, and IL-3. On day 8 of culture, all hematopoietic colonies consisting of 50 or more cells were scored, pooled, and analyzed for Ly-5 expression. In experiments 6 through 8, the chimeric mice were used only for the secondary transplantation studies. In experiments 1, 2, 4, and 6, the chimeric mice were survivors of studies described previously.14
BM indicates bone marrow; BMT, bone marrow transplantation; PB, peripheral blood; CFU-Cs, colony-forming units in culture.
CD34+ and CD34− populations of BM mononuclear cells were prepared by fluorescence-activated cell sorting with fluorescein isothiocyanate–conjugated anti-CD34.14
Significant correlation between muscle and BM chimerism was observed (Pearson correlation coefficient, 0.961; P < .01).
Ly-5.1 cells were transplanted without Ly-5.2 radioprotective cells.
Mononuclear cells were used for culture.