MAINTENANCE AND EXPANSION OF ERYTHROPOIESIS IN HUMAN LONG-TERM BONE-MARROW CULTURES IN PRESENCE OF ERYTHROPOIETIN PLUS STEM-CELL FACTOR ANDINTERLEUKIN-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 freed om from relapse. Long-term human bone marrow cultures (LTHBMC) represe nt the best available approximation of in vivo hematopoiesis. In the t raditional 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 Erythropoi etin (Epo), SCF, IL-3 and/or IL-11 by the stromal and hematopoietic ce lls ex vivo and/or suboptimal addition of these cytokines. Therefore, we have investigated the optimal presence of erythropoietin pills SCF, IL-3 and/or IL-11 as requirements for the maintenance and expansion o f erythropoiesis in LTHBMC in an effort to overcome the defective eryt hropoiesis of the traditional LTHBMC. In LTHBMC containing Epo alone a nd its combinations with SCF, IL-3 and/or IL-11, the nonadherent cells consisted mainly of erythroblast and normoblast cells which became th e majority in the third and fourth weeks whereas granulocytes and macr ophages declined steadily from the second week. A significant increase in the number of erythroblast and normoblast cells was produced throu ghout 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 clonoge nic cells were successfully maintained in cultures containing Epo + SC F + IL-3 or Epo + SCF + IL-ll or Epo + SCF + IL-3 + IL-11 for the 4 we eks 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 f ourth weeks, respectively, p < 0.01. Our optimum cultures, including E po + SCF + IL-3 or Epo + SCF + IL-11, maintained the production of non adherent erythroid clonogenic cells for 4 weeks in culture, representi ng 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 p roduction and the supplementation with either of the two combinations: Epo + SCF + IL-3 or Epo + SCF + IL-11 is sufficient for maintaining e rythropoiesis in LTHBMC. The present LTHBMC system should have applica tions to the analysis and manipulation of human erythropoiesis.