Eosinophil activation by colony-stimulating factor in man: Metabolic effects and analysis by flow cytometry

Abstract

Substantial increases in the killing capacity of human eosinophils after in vitro incubation with human placental conditioned medium (HPCM), a standard source of colony-stimulating factor (CSF), have recently been described. In this article, the interaction between HPCM and purified human eosinophils is analyzed by flow cytometry and by effects on iodination, superoxide production, and protein synthesis. HPCM increased the intensity of natural eosinophil autofluorescence (aFlu) (460 nm) after the absorption of ultraviolet light (360 nm) in a manner that was both time and dose dependent. Measured in arbitrary units, eosinophil aFlu was 72 ± 7.3 (arithmetic mean ± SEM) and 121 ± 3.2 after 18-hr incubations in the absence or presence of HPCM, respectively. The activity in HPCM responsible for these changes cochromatographed on Ultrogel AcA44 columns with CSF and with the less hydrophilic variant of CSF (CSF-α) on phenyl Sepharose. Mouse spleen, but not mouse lung, conditioned medium was also active on human eosinophils in this assay. Both CSF-α and mouse spleen conditioned medium also contain eosinophil colony-stimulating activity (CSA), whereas inactive CSFs with no effect on mature eosinophils, CSF-β, and mouse lung conditioned medium also lack eosinophil CSA. CSF-α stimulated superoxide production of resting eosinophils (from 0.03 ± 0.03 to 0.47 ± 0.08 nmole cytochrome-c reduced/105 eosinophils) and of eosinophils incubated with preopsonized zymosan (from 0.15 ± 0.06 to 0.73 ± 0.07). It also stimulated iodination by resting eosinophils (from 0.76 ± 0.16 to 2.60 ± 0.72 nmoles l/107 eosinophils/hr) and of eosinophils incubated with preopsonized zymosan (from 7.52 ± 2.08 to 29.8 ± 1.32). In contrast, CSF-β was inactive in these assays. CSF-α also stimulated, between 2- and 15-fold, the new protein synthesis of eosinophils. Thus, substances that stimulate the differentiation of progenitor cells into eosinophils also interact with peripheral mature eosinophils, and the activation of postmitotic cells may be a physiologic role of CSF-like molecules.