Marrow Reticulo-Fibroblastoid Colonies (CFU-RF Derived) Spontaneously Release an Erythroid Colony (BFU-E) Enhancing Factor
The marrow microenvironment is composed of an extracellular matrix as well as a heterogeneous population of cells. Isolation of the various cell types and analysis of their function is necessary for a better understanding of their roles in hemopoiesis. We have recently reported a colony assay for a cellular component of the marrow microenvironment. The assay consists of a plasma clot-methylcellulose marrow culture. The stimulator is PHA-stimulated leukocyte conditioned medium (PHA-LCM) and hydrocortisone (5 x 10-5M). The fibrin strands appear to act as a substrate for the growth of Reticulo-Fibroblastoid colonies derived from the CFU-RF precursor. RF colonies can be subcultured forming adherent layers when transferred to liquid cultures. Confluent adherent layers can be maintained for long periods of time by changing medium every 3 to 5 days. Supernatants derived from unstimulated RF cultures (RF-CM) were tested for growth promotion of hemopoietic precursors. We found: (1) RF-CM by itself does not induce colony formation. (2) In the presence of erythropoietin, RF-CM enhances the growth of BFU-E. (3) Recombinant IL 4 also enhances BFU-E formation, but in our assays IL 4 induced fewer colonies than RF-CM and the colonies were smaller. (4) Because neither IL 4 nor RF-CM, by themselves, can stimulate colony formation, we compared the effect of RF-CM on assays that are known to show other IL 4 functions. RF-CM did not induce proliferation of PHA induced blast T cells, a known property of IL 4. In conclusion, RF cells in culture appear to secrete, spontaneously, a factor that enhances BFU-E formation, and, unlike GM-CSF and 1L3, RF-CM has no effect on other hemopoietic colony forming cells. IL 4 has similar properties, but RF-CM appears not to have the B- and T-cell growth factor activities of Interleukin 4.
KeywordsColony Formation Bone Marrow Culture Hemopoietic Stem Cell Adherent Layer Human Bone Marrow Cell
Unable to display preview. Download preview PDF.
- 4.Schrader, J.W. 1983. Bone marrow differentiation in vitro. CRC Crit. Rev. Immunol. 4 (3): 197–277.Google Scholar
- 7.Izaguirre, C.A., F.E. Katz, and M.F. Greaves. 1982. Origin of marrow microenvironmental stromal cells in hemopoietic pluripotent stem cells. Blood 60, Suppl. 1: 323a.Google Scholar
- 11.Singer, J.W., A. Keating, J. Cuttner, A.M. Gown, R. Jacobson, P.D. Killen, J.W. Moohr, V. Najfeld, J. Powell, J. Sanders, G.E. Striker, and P.J. Fialkow. 1984. Evidence for a stem cell common to hematopoiesis and its in vitro micro-environment: Studies of patients with clonal hematopoietic neoplasia. Leuk. Res. 8: 535–545.PubMedCrossRefGoogle Scholar
- 14.Messner, H.A., J.E. Till, and E.A. McCulloch. 1973. Interacting cell populations affecting granulopoietic colony formation by normal and leukemic human marrow cells. Blood 42: 707–710.Google Scholar
- 24.Aye, M.T. 1987. A novel growth factor produced by cells sub cultured from marrow stromal (CFU-RF) colonies. Blood Suppl. 1: 166a.Google Scholar