H L Wilson1, K Ni, H C O'Neill. 1. Division of Biochemistry and Molecular Biology, School of Life Sciences, Australian National University, Canberra ACT.
Abstract
UNLABELLED: A unique long term culture (LTC) system has been developed which supports the production of dendritic cells (DC). Cell production is dependent on a stromal cell layer derived from murine spleen. This LTC system produces a high turnover of non-adherent cells that express DC morphology, cell-surface markers, and antigen-presenting capacity. OBJECTIVE: The long term production of these cells suggests that the LTC system supports hemopoiesis. It was of interest to examine the number and nature of hemopoietic progenitors present in LTC. MATERIALS AND METHODS: A combination of approaches, including FACS analysis, spleen colony-forming unit assays, and in vitro colony assays were undertaken. RESULTS: Pluripotent haemopoietic stem cells are not detectable among the non-adherent cell population produced in LTC. Instead, LTC support a replicating c-kit+ progenitor population, which generates only dendritic-like colonies in in vitro colony assays. In addition, this population does not respond to combinations of growth factors thought to stimulate DC proliferation, including granulocyte macrophage-colony stimulating factor (GM-CSF) and Flt3L. Production of DC occurs only in the presence of LTC-derived culture supernatant or a confluent stromal cell layer. CONCLUSIONS: These results suggest that LTC contain a dendritic progenitor that is dependent upon the stromal cell network for proliferation and differentiation. The development of only DC within LTC allows easy collection of cells for experimentation. This, in combination with the fact that DC development occurs in the absence of exogenous growth factors, makes the LTC system a practical model for the study of DC function and development.
UNLABELLED: A unique long term culture (LTC) system has been developed which supports the production of dendritic cells (DC). Cell production is dependent on a stromal cell layer derived from murine spleen. This LTC system produces a high turnover of non-adherent cells that express DC morphology, cell-surface markers, and antigen-presenting capacity. OBJECTIVE: The long term production of these cells suggests that the LTC system supports hemopoiesis. It was of interest to examine the number and nature of hemopoietic progenitors present in LTC. MATERIALS AND METHODS: A combination of approaches, including FACS analysis, spleen colony-forming unit assays, and in vitro colony assays were undertaken. RESULTS: Pluripotent haemopoietic stem cells are not detectable among the non-adherent cell population produced in LTC. Instead, LTC support a replicating c-kit+ progenitor population, which generates only dendritic-like colonies in in vitro colony assays. In addition, this population does not respond to combinations of growth factors thought to stimulate DC proliferation, including granulocyte macrophage-colony stimulating factor (GM-CSF) and Flt3L. Production of DC occurs only in the presence of LTC-derived culture supernatant or a confluent stromal cell layer. CONCLUSIONS: These results suggest that LTC contain a dendritic progenitor that is dependent upon the stromal cell network for proliferation and differentiation. The development of only DC within LTC allows easy collection of cells for experimentation. This, in combination with the fact that DC development occurs in the absence of exogenous growth factors, makes the LTC system a practical model for the study of DC function and development.
Authors: Ellen Weisberg; Renee D Wright; Douglas W McMillin; Constantine Mitsiades; Arghya Ray; Rosemary Barrett; Sophia Adamia; Richard Stone; Ilene Galinsky; Andrew L Kung; James D Griffin Journal: Mol Cancer Ther Date: 2008-04-29 Impact factor: 6.261