Quantitative analysis of the role of accessory cells in the development of human blood BFU-E-derived erythroid colonies.

A simplified method for the purification of human peripheral blood erythroid progenitor cells (BFU-E) using standard immunological techniques is described. Following removal of platelets, erythrocytes, nylon-wool-adherent cells, and sheep erythrocyte rosette-forming cells (RFC), BFU-E are routinely concentrated tenfold in the null cell fraction. Null cells plated at low density in erythroid cell cultures containing optimal amounts of methylcellulose, erythropoietin, and fetal calf serum did not give rise to spontaneous erythroid colonies. Coculture of null cells with highly purified, autologous RFC at a ratio of 1:25 yielded well-hemoglobinized erythroid colonies which were noticeably smaller than those found in cultures containing unfractionated peripheral blood mononuclear cells. However, further addition of very low numbers of purified adherent cells to null plus RFC dramatically increased the total hemoglobin content as well as the size and number of BFU-E-derived erythroid colonies. Addition of adherent cells alone to null cells had virtually no effect. Under conditions of optimal stimulation by adherent cells and RFC, the number of erythroid bursts was linearly related to null cells plated over an eightfold range. The synergism exhibited between adherent cells and RFC was not restricted by mismatched histocompatibility antigens. This system should be generally useful in quantitating the roles of more highly purified cellular and molecular populations in human erythropoiesis.