A burst-promoting activity derived from the human bone marrow stromal cell line KM-102 is identical to the...

Recently, several human bone marrow stromal cell lines have established and produced hematopoietic growth factors. One of these factors, a burst-promoting activity (BPA), was purified from 6 liters of serum-free conditioned medium cultured from stromal cell line KM-102, which was stimulated by phorbol myristate acetate (PMA) and calcium ionophore A23187. This stimulation induced 60 times more production of BPA than the unstimulated control culture. BPA was purified 4000-fold by sequential fractionation using ammonium sulfate precipitation, anion-exchange and lentil lectin affinity chromatographies, high performance gel filtration chromatography, and reversed phase high performance liquid chromatography. Purified BPA gave a single broad band of protein with a molecular weight of approximately 18 kd, as assessed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The concentration required for half maximal growth of early erythroid colonies was estimated as 10 pg/ml or 0.6 pM. At a higher concentration (125 pg/ml) this factor also stimulates the growth of granulocyte, macrophage, and eosinophil colonies in agar culture. The profile of amino acid composition is very similar to that of the human granulocyte-macrophage colony-stimulating factor (GM-CSF) deduced from its complementary DNA sequence. The result of amino-terminal sequence analysis strongly suggests that the purified material consists of GM-CSF and tetrapeptide-deleted GM-CSF. Moreover, antibody against GM-CSF completely neutralized the biological activities of this factor. These results indicate that the human bone marrow stromal cell line secretes GM-CSF as a burst-promoting activity and GM-CSF may play a significant role in the interaction between stem cells and stromal cells in the hematopoietic microenvironment.