Characterization of a colony-stimulating factor produced by the human monocytic leukemia cell line, THP-1

Abstract

The human monocytic leukemia cell line, THP-1, acquires macrophage-like characteristics following exposure to mezerein. Serum-free medium conditioned by mezerein-activated cells was observed to contain colony-stimulating factor (CSF) activity in assays with murine bone marrow cultures. Isoelectrofocusing revealed that CSF activity displayed charge heterogeneity and migrated in a pl range of 4.4-5.3. Treatment with neuraminidase did not affect biological activity but did reduce charge heterogeneity. Reisofocusing of neuraminidase-treated CSF revealed a peak of activity at pl 4.9. The active component was shown to be an acidic sialoglycoprotein, resistant to proteolytic cleavage but completely inactivated by 2-mercaptoethanol. This CSF has been purified from THP-1-conditioned serum-free medium by preparative isoelectrofocusing, gel filtration through Sephacryl S-200, ion exchange chromatography on DEAE-Sephacel, neuraminidase treatment, and tris-glycinate polyacrylamide gel electrophoresis (PAGE). Elution from SDS-PAGE revealed a single peak of activity corresponding to an apparent molecular weight of 70,000 daltons. Preliminary characterization of the bone marrow cells in colonies showed that THP-1 cells produced macrophage-specific CSF.