Heterogeneity of buoyant density and proliferative state of circulating erythropoietic progenitor cells (BFU-E) in man.
Normal human blood BFU-E are believed to be in a quiescent state with respect to DNA synthesis, since few or none of these progenitors can be killed by cycle-active agents. Using Percoll discontinuous density gradient centrifugation of normal human blood mononuclear cells, we have separated two subpopulations of BFU-E with different proportions in DNA synthesis. Mononuclear cells were obtained with Ficoll-Isopaque from 24 samples of normal blood. BFU-E were assayed with the methylcellulose technique, and their proliferative state was studied with the hydroxyurea (HU) suicide method. The results obtained with a five-step gradient of density range 1.060-1.068 g/ml showed that the vast majority of BFU-E were distributed approximately equally between pooled fractions with rho = 1.060 + 1.062 g/ml and those with rho = 1.064 + 1.066 g/ml. Among nonseparated cells or cells in the fraction of rho = 1.060 + 1.062 g/ml, we failed to detect a significant proportion of DNA-synthesizing BFU-E in the great majority of samples. In contrast, in the pooled fractions with rho = 1.064 + 1.066 g/ml, 14 of 24 samples showed significant kill, and among these 14, a highly significant proportion of BFU-E were killed by HU (39.3% +/- 3.4%). Therefore, the separation of mononuclear cells on the basis of their different buoyant densities revealed the presence of DNA-synthesizing BFU-E in normal human blood. Either DNA-synthesizing BFU-E have a higher buoyant density than non-DNA-synthesizing BFU-E, or else cells of lower buoyant density normally inhibit DNA synthesis in BFU-E.
D002455:Cell Division; D002499:Centrifugation, Density Gradient; D004920:Erythropoiesis; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D006918:Hydroxyurea
D006918:Hydroxyurea