Human marrow erythropoiesis in culture. I. Characterization of methylcellulose colony assay

Abstract

The morphologic and functional characteristics were examined of human marrow erythrocytes cultured with a recently developed methylcellulose colony assay technique. Erythrocytic cells in various stages of development were observed, and a significant degree of maturational synchrony within individual colonies was noted. By light microscopy, colonies consisting of late normoblasts appeared compact, had an orange hue attributable to their hemoglobin, and demonstrated pseudoperoxidase activity, whereas colonies composed of early erythroblasts grew less compact or in clusters of smaller cell aggregates and showed no reddish tinge. Colonies possessing intermediate features were also observed. Maturational synchrony of individual colonies was confirmed using transmission and scanning electron microscopy. The ultrastructure and cytochemistry of most immature cells were normal. The mature erythrocytes, however, were severely microcytic and hypochromic and contained one to several Heinz bodies. These defects in the cytoplasmic maturation of erythrocytes correspond with impaired granulocytic maturation in culture, which was observed previously and suggest environmental or nutritional defects in culture. Linearity of the method was confirmed using 5 normal bone marrows. Erythropoietin dose responses observed in 10 normal marrows were comparable to the previously reported results and revealed significant variation in individual plating efficiencies.