Jp Maciejewski

Long-lasting decrease of marrow and circulating long-term culture initiating cells after allogeneic bone marrow transplant

We investigated bone marrow (BM) and circulating (PB) hematopoietic progenitor cells in 37 normal donors and in 25 patients 1 to 8 years after successful allogeneic bone marrow transplant. At the time of testing, transplanted patients had normal blood counts and bone marrow cellularity. By flow cytometry, BM CD34+ cells were found to be three- to four-fold decreased in transplanted patients compared to normal donors, while the number of PB CD34+cells was the same as in normal donors. Using a methylcellulose colony assay, primary BM colony-forming cells (CFU-GM) were decreased 2.1-fold, whereas PB CFU-GM were only marginally decreased. In a long-term culture initiating cell (LTC-IC) assay, an eight-fold decrease of early progenitor cells was observed in the marrow of transplanted patients compared to normal donors, and a five-fold decrease was documented in peripheral blood. We found that the BM LTC-IC cell number correlated with concurrently determined BM CD34+ cells and committed progenitor cell number (measured as CFU-GM) and with PB LTC-IC number, but not with PB CFU-GM and CD34+ cells. We conclude that marrow and circulating early stem cell compartments, as measured by the LTC-IC assay, are greatly and permanently depressed following bone marrow transplant. The correlation between BM and PB LTC-IC indicates that the enumeration of circulating LTC-IC can be used as a measure of the stem cell compartment in the bone marrow after transplant. It seems that the deficiency of the most immature progenitor cells persists forever after successful bone marrow transplant; this means that a complete hematopoietic reconstitution can be sustained by a reduced stem cell pool.

Sustained long-term hematologic recovery despite a marked quantitative defect in the stem cell compartment of patients with aplastic anemia after immunosuppressive therapy†

Previously, we reported that patients with aplastic anemia (AA) have profoundly decreased numbers of hematopoietic progenitor and stem cells as measured in the long‐term culture initiating cell (LTC‐IC) assay (Blood 1996;88:1983–1991). We now present results of a long‐term prospective study of LTC‐IC numbers in peripheral blood (PB) and bone marrow (BM) of patients treated with antithymocyte globulin and cyclosporin A. Numbers of secondary colony forming cells (secondary CFC) in long‐term bone marrow culture (LTBMC) were used to quantitate LTC‐IC. BM (N = 35) and PB (N = 41) secondary CFC from both untreated severe AA patients and responders to immunosuppressive therapy who were sampled up to 6 years after initial treatment were compared. Normal controls showed 148 ± 38 (N = 17) and 16 ± 3 (N= 14) secondary CFC per 106 in BM and PB, respectively. In cross‐sectional analysis, prior to therapy, AA patients showed 2.6 ± 1 (mean ± SD) secondary CFC/106 BM MNC; within the first year after initial treatment (N = 14), secondary CFC number rose modestly to 8.2 ± 2.2/106 MNC, and further increased to 15.8 ± 7 (N = 17) at 2 years and 16.2 ± 7/106 MNC (N = 25) 3 years after treatment. There was no further improvement in the secondary CFC numbers at 4, 5, and ≥6 years (N = 37). Thus, while BM secondary CFC increased about 6‐fold at 3 years post‐therapy compared to presentation, they remained about only 10% of normal despite hematologic recovery. Similar data were obtained for PB, with approximately 4‐fold increase in secondary CFC numbers within 2 years of therapy, to about 15% of normal values. We confirmed these observations in patients studied serially over a period of 4 years: initial secondary CFC were 2.35 ± 1/106 BM MNC and 0.11 ± 0.1/106 PB MNC improving to an average of 6 ± 1.2 (BM; N = 12) and 2.4 ± 1/106 MNC (PB; N = 14). In many cases of partial recovery, PB counts improve but do not normalize. When we studied secondary CFC numbers only in patients who achieved complete normalization of PB counts (ANC >1,500/mm3; platelets >105/mm3 and absolute reticulocytes >5 × 104/mm3), BM secondary CFC were significantly higher than in patients with partial recovery; the PB secondary CFC number was modestly increased but remained below the normal values. Within the group of patients with complete recovery, there was no correlation between the secondary CFC and time after initial treatment. In addition, there also was no correlation between the secondary CFC number at presentation and the quality of hematopoietic recovery. Despite a limited expansion potential of a severely reduced stem cell pool, their numbers are sufficient to provide a long‐term supply of mature blood cells. Am. J. Hematol. 65:123–131, 2000. Published 2000 Wiley‐Liss, Inc.