Characterization of human bone marrow fibroblast colony-forming cells (CFU-F) and their progeny

Abstract

The bone marrow stromal cell population is comprised of fibroblasts, endothelial cells, fat cells and “reticular cells.” The characteristics and the regulatory role in hematopoiesis of each of these cell types are unknown. A liquid culture system has been used to clone and to characterize human bone marrow fibroblast colony-forming cells (CFU-F). The linear relationship between the number of cells plated and the number of colonies formed suggests that fibroblast colonies originate in a single cell. Bone marrow CFU-F were adherent and nonphagocytic. The majority (90% ± 2%) were less dense than 1.070 g/cu cm. Velocity sedimentation separation demonstrated a heterogeneous CFU-F sedimentation rate, with a modal sedimentation of 4.95 ± 0.15 mm/hr. Analysis of CFU-F proliferative status by the thymidine suicide technique indicated that this cell was noncycling in individuals with undisturbed bone marrow function. Some of the more distinctive products of fibroblasts, other stromal cells, and hematopoietic colony-forming cells were used as positive and negative markers for CFU-F and the cells derived from them in vitro. Complement-mediated cytotoxicity using anti-Ia and anti-factor-VIII antigen antisera did not inhibit fibroblast colony formation. In contrast, a striking reduction of granulocyte-macrophage colony formation (CFU-c) was seen when bone marrow cells were treated with anti-Ia antiserum. Immunofluorescence staining was used to characterize the cells derived from CFU-F in vitro. No staining was observed after incubation of subconfluent cultures with anti-Ia and anti-factor-VIII antigen antisera. A positive immunofluorescent staining was obtained when isolated antibodies against three of the main proteins of bone marrow matrix: type I collagen, type III collagen, and fibronectin were used. Ultrastructural analysis showed that CFU-F progeny, in contrast to endothelial cells, did not contain Weibel-Palade bodies. These data support the conclusion that the colonies described in this study are of fibroblastic nature.