A B S T R A CT To examine the switch from fetal to adult hemoglobin at the cellular level, erythroid progenitor cells from newborn infants and adults were cultured in methyl cellulose with erythropoietin. Individual erythroid colonies were labeled with [3H]-leucine at various times, and globin synthesis patterns examined by gel electrophoresis and fluorography. The percent Ay-or fl-globin synthesis was determined from the total of y + P, and the percent Gy from the total of Gy + Ay. The nonparametric correlation coefficients of percent G-y with percent y or # were obtained. Each group of colonies at each time point was examined separately. In colonies from adult blood, the proportion of Gy-synthesis did not correlate with the proportion of y-synthesis. Colonies from newborn blood fell into two groups. Those that developed from relatively mature progenitor cells, and were seen on day 14, showed a strong negative correlation of Gy with f3-globin synthesis. However, those newborn colonies that developed from immature progenitors, and were seen later in culture (days 17 and 21), showed no correlation of Gy with A-synthesis. These findings are compatible with a clonal model for hemoglobin switching. Fetal progenitors, in which G'y-and #-syntheses are negatively correlated, are gradually replaced during ontogeny by adult progenitors. The adult progenitors produce more If (less y), and the proportions of Goy-and y-or Bi-synthesis are not correlated.
The transient fetal-like erythropoiesis which appears during recovery from bone marrow transplantation has now been examined at the level of erythroid progenitor cells. A 7-year-old boy with beta +-thalassaemia major was studied during engraftment from his beta-thalassaemia trait sister. Hb F and i antigen rose as expected. Macrocytosis never developed, but red cell size distribution became very heterogeneous. Bone marrow CFU-E and BFU-E were detected by 30 d, prior to the appearance of reticulocytes. Marrow erythroid progenitor cell numbers were normal by 146 d, while those in the blood became normal by 360 d. After transplantation globin synthesis ratios in erythroid colonies were diagnostic of thalassaemia trait, indicating engraftment. Individual erythroid colonies derived from both blood and marrow at all times during reconstitution showed no correlation of G gamma and gamma. Thus the fetal-like stress erythropoiesis of marrow expansion following transplantation was derived from adult and not fetal progenitor cells.
Specific globin mRNA accumulation was quantitated in several lines of K562 cells in the absence and the presence of hemin. Using specific cloned DNA probes, the amounts of zeta, alpha, epsilon and gamma mRNAs were shown to be increased 2–3-fold in the presence of 20 microM hemin. No delta- or beta-globin mRNAs were detectable in any of the lines. In one line, Bos, there was a marked decrease in epsilon-globin mRNA, which increased with hemin, although still to much lower levels than in the other lines. The decreased epsilon-globin mRNA accumulation in Bos is shown to be due to decreased epsilon-globin gene transcription.
Specific globin mRNA accumulation was quantitated in several lines of K562 cells in the absence and the presence of hemin. Using specific cloned DNA probes, the amounts of zeta, alpha, epsilon and gamma mRNAs were shown to be increased 2–3-fold in the presence of 20 microM hemin. No delta- or beta-globin mRNAs were detectable in any of the lines. In one line, Bos, there was a marked decrease in epsilon-globin mRNA, which increased with hemin, although still to much lower levels than in the other lines. The decreased epsilon-globin mRNA accumulation in Bos is shown to be due to decreased epsilon-globin gene transcription.
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