IL-10 synergistically enhances GM-CSF-induced CCR1 expression in myelomonocytic cells
Section snippets
Materials and methods
Materials. The following materials were used in this study: human monoblastic leukemic U937 cell line (American Type Culture Collection, Rockville, MD); TRIzol reagent, CCR1 and CCR5 primers (Life Technologies, Gaithersburg, MD); human recombinant GM-CSF, IL-10, and macrophage inhibitory protein-1α (MIP-1α) (BD PharMingen, San Diego, CA); mouse mAb against CCR1 (R&D Systems, Minneapolis, MN); mouse mAbs against phospho-extracellular signal-regulated kinase (ERK) (Thr202/Tyr204) and pan Erk, and
IL-10 augmented GM-CSF-induced CCR1 protein expression
The unprimed U937 cells did not show any detectable level of CCR1 on the cell surface. When used alone, GM-CSF (20 ng/ml) and IL-10 (10 ng/ml) induced minor increases in CCR1 protein expression in U937 cells, with GM-CSF causing significantly greater up-regulation than IL-10 (Table 1). Combined GM-CSF/IL-10 treatment resulted in a synergistic increase in CCR1 protein expression. The unprimed U937 cells expressed low levels of CCR1 mRNA (102±10 copies/103 copies of β-actin, n=4). Fig. 1 shows that
Discussion
The present study demonstrates that IL-10 can synergistically enhance GM-CSF-induced CCR1 protein expression in myelomonocytic cells, and the synergism occurs primarily at the protein translational level, but not the gene transcription level. These conclusions are based on the observations that (a) GM-CSF, but not IL-10, increased both CCR1 mRNA and protein expression; (b) addition of IL-10 to GM-CSF treatment synergistically enhanced CCR1 protein expression with no effect on mRNA expression;
Acknowledgements
This work was supported by a grant from the National Medical Research Council of Singapore (NMRC/0433/2000) to W.S.F.W.
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