Ex vivo expansion of murine marrow cells with interleukin-3 (IL-3), IL-6, IL-11, and stem cell factor leads to impaired engraftment in irradiated hosts

Abstract

In vitro incubation of bone marrow cells with cytokines has been used as an approach to expand stem cells and to facilitate retroviral integration. Expansion of hematopoietic progenitor cells has been monitored by different in vitro assays and in a few instances by in vivo marrow renewal in myeloablated hosts. This is the first report of studies, using two competitive transplant models, in which cytokine-treated cells, obtained from nonpretreated donors (eg, 5-fluorouracil), were competed with normal cells. A basic assumption is that the expansion of progenitors assayed in vitro as high- and low-proliferative potential colony-forming cells (HPP- and LPP-CFCs) indicates an expansion of stem cells which will repopulate in vivo. This study shows that culture of marrow cells with four cytokines (stem cell factor, interleukin-3 [IL-3], IL-6, IL-11) induces significant expansion and proliferation of HPP-CFC and LPP-CFC. Cell-cycle analysis showed that these hematopoietic progenitors were induced to actively cell cycle by culture with these cytokines. In the first competitive transplant model, which uses Ly5.2/Ly5.1 congenic mice, cytokine-cultured Ly5.2 cells competed with noncultured Ly5.1 cells led to 5% +/- 1% engraftment at 12 weeks and to 4% +/- 2% engraftment at 22 weeks posttransplantation for the cytokine exposed cells. Noncultured Ly5.2 cells competed with cultured Ly5.1 cells led to 70% +/- 1% engraftment at 12 weeks and to 93% +/- 2% engraftment at 22 weeks posttransplantation. In the second model, which uses BALB/c marrow of opposite genders, cultured male cells lead to 13% +/- 9% engraftment at 10 weeks and 2% +/- 1% engraftment at 14 weeks posttransplantation; noncultured male cells lead to 70% +/- 2% and 95% +/- 2% engraftment at 10 and 14 weeks posttransplantation, respectively. Data presented here from two different competitive transplant studies show a defect of cytokine expanded marrow related to cell cycle activation which manifests as defective long-term repopulating capability in irradiated host mice. The engraftment defect is more profound at longer time intervals, suggesting that the most striking effect may be on long-term repopulating cells.