Stimulation of adult human bone marrow by factors secreted by fetal liver hematopoietic cells: In vitro evaluation using semisolid clonal assay system

Abstract

Fetal liver infusion (FLI) therapy has been used in various disorders, such as aplastic anemia, leukemia, metabolic disorders, etc., and has been shown to result in stimulation of autologous hematopoiesis in many cases. The aim of the present study was to elucidate the mechanism of stimulation of adult hematopoiesis by fetal liver hematopoietic cells (FLHC) and to identify the factors involved in the process using a clonal assay system in vitro. The effect of FLHC on the clonal growth of bone marrow cells was studied using a co-culture system consisting of mitomycin C-treated FLHC with 2 x 105 bone marrow (BM) mononuclear cells. It was observed that FLHC induced a two- to four-fold increase in the BM colony formation. A further increase in the number of FLHC did not, however, result in an equivalent fold increase in the colony formation, indicating that the number of cells in the BM population responsive to FLHC was perhaps the limiting factor. When the effect of fetal liver cell conditioned medium (FLCM) was examined in a similar fashion, it was observed that the FLCM showed a 1.5- to 4-fold increase in the colony formation when used at 1%-5% along with limiting amounts of growth factors. Higher concentrations of conditioned medium resulted in inhibitory responses. One of the principal factors responsible for the stimulatory activity of FLCM was shown to be transforming growth factor-β1 (TGF-β1), by a variety of experiments such as its quantitation in FLCM by enzyme-linked immunosorbent assay, antibody neutralization, and reconstruction experiments using purified TGF-β1 and normal medium. In these reconstitution experiments, TGF-β1 stimulated the colony formation when it was applied at 1-50 pg/ml, but at higher concentration it induced an inhibitory effect, mimicking the behavior earlier seen with FLCM. Our data strongly suggest that one of the mechanisms in stimulation of a recipient's hematopoiesis could be mediated by the action of TGF-β1 secreted by infused FLHC and could provide a rational framework on which FLI therapy can be further evaluated.