Action of antithymocyte globulin on normal human erythroid progenitor cell proliferation in vitro: erythropoietic growth-enhancing factors are released from marrow accessory cells.
Studies were undertaken to determine the mechanism of action of a horse antithymocyte globulin preparation (ATGAM) (HATG) and its control preparation of horse gamma globulin (HIgG) on the proliferation of normal human marrow and blood erythroid progenitor cells (CFU-E, BFU-E) in vitro. In preincubation studies with marrow mononuclear cells and complement, HATG did not significantly augment CFU-E or BFU-E growth greater than that expected because of removal of marrow T cells by this agent. However, direct addition of HATG but not HIgG to marrow cultures significantly stimulated CFU-E and BFU-E up to two to four times that of media or HIgG controls (P less than 0.05). The peak effect was observed at 10 to 100 micrograms/ml HATG; HATG was toxic at 1000 micrograms/ml. By contrast, the OKT3 monoclonal antibody was less stimulatory than HATG. The in vitro erythropoietic stimulatory effect of HATG was dependent on the presence of accessory cells because removal of T cells or monocytes (less than 2% to 5%) or adsorptions of HATG with T cells or monocytes reduced its stimulatory effect, highly purified BFU-E nearly devoid of accessory cells required irradiated accessory cells for demonstration of the HATG stimulatory effect, and an erythroid burst-promoting activity was released from T cells or unseparated mononuclear cells in the presence of HATG but not HIgG. The HATG enhancing effect was optimal in the first 96 hours of cultures in the presence of erythropoietin, and was reproducible with three separate lots of HATG. Up to 16% of HATG-stimulated erythroid colonies expressed nonerythroid lineage cells. Iron 59 incorporation into heme of CFU-E- or BFU-E-derived colonies was augmented equally by HATG or HIgG at 10 micrograms/ml. Erythropoietin dose-response curves and studies with antierythropoietin sera suggested that HATG also increased the sensitivity of erythroid progenitor cells to very low concentrations of erythropoietin. We conclude that HATG but not HIgG control has potent dose-dependent erythroid progenitor cell growth-enhancing effects. In addition to the ability of HATG to lyse T-suppressor cells, these findings suggest that HATG may also stimulate erythropoiesis indirectly by releasing growth-enhancing factor(s) from T cells and other marrow accessory cells, sensitizing erythroid progenitor cells to low concentrations to erythropoietin, and stimulating growth of bipotential progenitor cells. Collectively, these effects may explain the efficacy of HATG in the treatment of some patients with erythropoietic marrow failure states.
K F
KF
Mangan
L
L
D'Alessandro
M T
MT
Mullaney
D000818:Animals; D000961:Antilymphocyte Serum / Q000494:pharmacology*; D001853:Bone Marrow / Q000378:metabolism / Q000473:pathology; D001854:Bone Marrow Cells; D002455:Cell Division / Q000187:drug effects; D003114:Colony-Forming Units Assay; D003165:Complement System Proteins / Q000276:immunology; D004921:Erythropoietin / Q000502:physiology; D006133:Growth Substances / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006736:Horses; D006801:Humans; D007700:Kinetics; D013601:T-Lymphocytes / Q000276:immunology; D005719:gamma-Globulins / Q000494:pharmacology
D000961:Antilymphocyte Serum; D006133:Growth Substances; D005719:gamma-Globulins; D004921:Erythropoietin; D003165:Complement System Proteins