| Popis: |
The use of a semisolid support like methylcellulose (MC) in a clonogenic assay prevents cell migration and nonspecific aggregation. However, the inhibitory effect of MC on myeloid cell lines has been reported. To assess the effect of MC on human leukemic progenitor cell growth (acute myeloblastic leukemia colony-forming units, AML-CFU), increasing concentrations of MC (0.36%, 0.72%, and 1.44%) were added in a double-feeder culture system. T-lymphocyte-depleted leukemic cells from 12 patients with AML were cultured in the presence of 2.5% phytohemagglutinin (PHA) in a liquid and a semisolid (MC) medium over a leukocyte feeder layer. The leukemic nature of the colonies was confirmed by cytogenetic studies. The median cloning efficiency in the optimal MC assay system was significantly higher (217 leukemic colony-forming units [CFU-L]/5 x 10(4) cells) than the one obtained in the liquid assay system (72.5 CFU-L/5 x 10(4) cells). However, three patterns of growth were observed: 1) colony formation was significantly better in MC than in the liquid assay system (seven of ten cases), 2) there was no difference in growth response (three of ten cases), and 3) colony formation was significantly better in the liquid assay system (one of ten cases). In the semisolid assay system, colony growth was dependent on MC concentration and varied among individual patients. A striking feature was the partial reduction of AML-CFU growth at 1.44% MC, with complete inhibition in 4/11 cases. This phenomenon was not observed for normal progenitors cultured under the same conditions. Cytological evaluation of AML-CFU showed an incomplete maturation to the myelocyte state, accompanied occasionally by macrophagic differentiation. In contrast, maturation of the granulocyte-macrophage colony-forming unit (CFU-GM) clones was harmonious, resulting in greater than 40% polynuclear cells, even from a 7-day culture. Despite a variable clonal response of leukemic progenitors from individual patients, we conclude that 0.72% MC is the optimal concentration of MC in our system, allowing clonal growth of AML-CFU. |
