Maintenance and expansion of erythropoiesis in human long-term bone marrow cultures in presence of erythropoietin plus stem cell factor and interleukin-3 or interleukin-11.

Recently there has been great interest in the ex vivo expansion and/or purging of human bone marrow cells prior to transplantation in order to obtain a long-lasting restoration of normal hematopoiesis and freedom from relapse. Long-term human bone marrow cultures (LTHBMC) represent the best available approximation of in vivo hematopoiesis. In the traditional LTHBMC, erythropoiesis is short lived (about 2 weeks). The longevity and productivity of erythropoiesis in LTHBMC may be limited by the insufficient production of certain cytokines such as Erythropoietin (Epo), SCF, IL-3 and/or IL-11 by the stromal and hematopoietic cells ex vivo and/or suboptimal addition of these cytokines. Therefore, we have investigated the optimal presence of erythropoietin plus SCF, IL-3, and/or IL-11 as requirements for the maintenance and expansion of erythropoiesis in LTHBMC in an effort to overcome the defective erythropoiesis of the traditional LTHBMC. In LTHBMC containing Epo alone and its combinations with SCF, IL-3, and/or IL-11, the nonadherent cells consisted mainly of erythroblast and normoblast cells which became the majority in the third and fourth weeks whereas granulocytes and macrophages declined steadily from the second week. A significant increase in the number of erythroblast and normoblast cells was produced throughout the whole period of LTHBMC containing Epo + IL-11 or Epo + SCF + IL-11 or Epo + SCF + IL-3 + IL-11. In the presence of Epo alone, BFU-E decreased steadily throughout LTHBMC. However, the erythroid clonogenic cells were successfully maintained in cultures containing Epo + SCF IL-3 or Epo + SCF + IL-11 or Epo + SCF + IL-3 + IL-11 for the 4 weeks and even significantly expanded by 4.5-5.7, 8.1-10 and 5-7-fold more than in the presence of Epo alone in the second, third and fourth weeks, respectively, p < 0.01. Our optimum cultures, including Epo + SCF + IL-3 or Epo + SCF + IL-11, maintained the production of nonadherent erythroid clonogenic cells for 4 weeks in culture, representing a significant improvement over the traditional LTHBMC that exhibits a progressive decline in erythropoiesis during the first 2 weeks. We conclude that Epo alone could not maintain erythroid clonogenic cell production and the supplementation with either of the two combinations: Epo + SCF + IL-3 or Epo + SCF + IL-11 is sufficient for maintaining erythropoiesis in LTHBMC. The present LTHBMC system should have applications to the analysis and manipulation of human erythropoiesis.