Ratajczak M Z, Ratajczak J, Kuczynski W, Light B, Lusk E J, Gewirtz A M
Department of Pathology, University of Pennsylvania School of Medicine, Philadelphia 19104.
Exp Hematol. 1993 Dec;21(13):1663-7.
Protracted engraftment and prolonged thrombocytopenia remain problems when marrow purged with 4-hydroperoxy-cyclophosphamide (4-HC) is employed for autotransplants. Toxic effects of 4-HC on early progenitor and stem cells might play an important causative role. Surprisingly, few investigations have examined the effects of 4-HC on progenitor cells other than colony-forming units-granulocyte/macrophage (CFU-GM) and burst-forming units-erythroid (BFU-E), and only one study could be found where 4-HC exposure was carried out on cells purified beyond the buffy coat stage. Since the cellular milieu, and in particular the red blood cell count, can ameliorate 4-HC toxicity, the suppressive effect of this agent on marrow stem cells might be underestimated. We therefore investigated the relative 4-HC sensitivity of different human bone marrow progenitor cells in vitro using partially purified adherent cell- and T lymphocyte-depleted bone marrow mononuclear cells (A-T-MNC). Cells were exposed to increasing doses (10 to 200 micrograms/mL) of 4-HC using a standard purging protocol established for bone marrow buffy coat cells. Effects on mixed (CFU-Mix), erythroid (CFU-E and BFU-E), granulocyte-macrophage (CFU-GM), and stromal cell (CFU-F) progenitors were determined. In addition, we examined the 4-HC sensitivity of megakaryocyte progenitors (CFU-Meg) since, to our knowledge, this has not been reported before. As expected, increasing doses of 4-HC led to progressive inhibition of hematopoietic colony formation. CFU-F, CFU-Mix, and CFU-Meg appeared most resistant to 4-HC exposure, while CFU-E, BFU-E, and CFU-GM appeared most sensitive. At doses over 100 micrograms/mL, the usual concentration recommended for purging of buffy coat cells, hematopoietic colony formation was completely inhibited. These data suggest that if more highly purified marrow cells are employed for purging, lower 4-HC doses may need to be used. They also suggest that the thrombocytopenia that frequently accompanies 4-HC purging is not likely due to loss of CFU-Meg.
当采用经4-氢过氧环磷酰胺（4-HC）净化的骨髓进行自体移植时，持久植入和长期血小板减少仍然是问题。4-HC对早期祖细胞和干细胞的毒性作用可能起着重要的致病作用。令人惊讶的是，很少有研究考察4-HC对除集落形成单位-粒细胞/巨噬细胞（CFU-GM）和爆式红系集落形成单位（BFU-E）之外的祖细胞的影响，并且仅发现一项研究对超过血沉棕黄层阶段纯化的细胞进行了4-HC处理。由于细胞环境，尤其是红细胞计数，可以减轻4-HC的毒性，该药物对骨髓干细胞的抑制作用可能被低估。因此，我们使用部分纯化的贴壁细胞和T淋巴细胞去除的骨髓单个核细胞（A-T-MNC），在体外研究了不同人类骨髓祖细胞对4-HC的相对敏感性。使用为骨髓血沉棕黄层细胞建立的标准净化方案，将细胞暴露于递增剂量（10至200微克/毫升）的4-HC中。测定对混合祖细胞（CFU-Mix）、红系祖细胞（CFU-E和BFU-E）、粒细胞-巨噬细胞祖细胞（CFU-GM）和基质细胞祖细胞（CFU-F）的影响。此外，我们还检测了巨核细胞祖细胞（CFU-Meg）对4-HC的敏感性，因为据我们所知，此前尚未有相关报道。正如预期的那样，递增剂量的4-HC导致造血集落形成逐渐受到抑制。CFU-F、CFU-Mix和CFU-Meg似乎对4-HC暴露最具抗性，而CFU-E、BFU-E和CFU-GM似乎最敏感。在超过100微克/毫升的剂量下，即通常推荐用于净化血沉棕黄层细胞的浓度，造血集落形成被完全抑制。这些数据表明，如果使用更高纯度的骨髓细胞进行净化，可能需要使用更低剂量的4-HC。它们还表明，4-HC净化时常伴随的血小板减少不太可能是由于CFU-Meg的丢失。
