Separation of normal hemopoietic stem cells from clonogenic leukemic cells in a rat model for human acute myelocytic leukemia—I. Velocity sedimentation

Abstract

In a search for preparative methods for obtaining optimal bone marrow grafts for autologous marrow transplantation in acute leukemia, cell separation by velocity sedimentation was investigated in a rat model for human acute myelocytic leukemia (BNML). Although the sedimentation rate profiles of normal and leukemic marrow cells partly overlap, normal hemopoietic stem cells (CFU-S) and clonogenic leukemic cells (LCFU-S) have distinct sedimentation velocities. CFU-S show a peak at a sedimentation rate of 3.5 mm.h −1 and LCFU-S at 7 mm.h −1. Below a sedimentation rate of 4.5 mm.h −1, 70% of the CFU-S and only 1% of the LCFU-S are recovered. Expressed as a percentage of the original number of CFU-S before separation, 40% are recovered. For leukemic cells, DNA histograms prepared from different fractions revealed a direct correlation between cell size and cell cycle phase. The relationship between the proliferative state of the normal and leukemic cell population and the efficacy of separation is discussed. Separation experiments with various mixtures of normal and leukemic marrow cells showed that all leukemic cells could be effectively removed from a cell suspension containing 0.1–0.5 oo o leukemic cells as determined by bioassays (elimination factor: 120–150). The minimal efficacy of remission-induction chemotherapy is discussed to achieve a suitable marrow cell population. After velocity sedimentation separation, this marrow should yield a sufficient number of normal hemopoietic stem cells to repopulate lethally treated recipients and should be completely free of leukemic cells.