Mangan K F, D'Alessandro L, Mullaney M T
J Lab Clin Med. 1986 Apr;107(4):353-64.
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.
开展了多项研究，以确定一种马抗胸腺细胞球蛋白制剂（ATGAM）（HATG）及其对照制剂马γ球蛋白（HIgG）对正常人骨髓和血液红系祖细胞（CFU-E、BFU-E）体外增殖的作用机制。在与骨髓单个核细胞和补体的预孵育研究中，HATG并未显著增强CFU-E或BFU-E的生长，其增强程度未超过因该制剂去除骨髓T细胞而预期的增长幅度。然而，将HATG而非HIgG直接添加到骨髓培养物中，可显著刺激CFU-E和BFU-E，其增殖程度高达培养基或HIgG对照的两到四倍（P小于0.05）。在HATG浓度为10至100微克/毫升时观察到峰值效应；HATG在1000微克/毫升时具有毒性。相比之下，OKT3单克隆抗体的刺激作用小于HATG。HATG的体外促红细胞生成刺激作用依赖于辅助细胞的存在，因为去除T细胞或单核细胞（少于2%至5%）或用T细胞或单核细胞吸附HATG会降低其刺激作用，高度纯化的几乎不含辅助细胞的BFU-E需要经辐照的辅助细胞才能显示出HATG的刺激作用，并且在存在HATG而非HIgG的情况下，T细胞或未分离的单个核细胞会释放一种促红细胞爆式集落形成活性物质。在促红细胞生成素存在的情况下，HATG的增强作用在培养的最初96小时内最为显著，并且使用三批不同的HATG均可重复出现该现象。高达16%的受HATG刺激的红系集落表达了非红系谱系细胞。在10微克/毫升时，HATG和HIgG对CFU-E或BFU-E来源集落的血红素中59铁掺入的促进作用相同。促红细胞生成素剂量反应曲线以及抗促红细胞生成素血清的研究表明，HATG还可增加红系祖细胞对极低浓度促红细胞生成素的敏感性。我们得出结论，HATG而非HIgG对照具有强大的剂量依赖性红系祖细胞生长增强作用。除了HATG裂解T抑制细胞的能力外，这些发现表明，HATG还可能通过从T细胞和其他骨髓辅助细胞释放生长增强因子、使红系祖细胞对低浓度促红细胞生成素敏感以及刺激双潜能祖细胞生长来间接刺激红细胞生成。总体而言，这些作用可能解释了HATG在治疗某些造血骨髓衰竭状态患者中的疗效。
