TY - JOUR
T1 - Metalloproteinases are involved in lipopolysaccharide- and tumor necrosis factor-α-mediated regulation of CXCR1 and CXCR2 chemokine receptor expression
AU - Khandaker, Masud H.
AU - Mitchell, Gordon
AU - Xu, Luoling
AU - Andrews, Joseph D.
AU - Singh, Rajkumari
AU - Leung, Harry
AU - Madrenas, Joaquín
AU - Ferguson, Stephen S.G.
AU - Feldman, Ross D.
AU - Kelvin, David J.
PY - 1999/4/1
Y1 - 1999/4/1
N2 - The neutrophil-specific G-protein-coupled chemokine receptors, CXCR1 and CXCR2, bind with high affinity to the potent chemoattractant interleukin-8 (IL-8). The mechanisms of IL-8 receptor regulation are not well defined, although previous studies have suggested a process of ligand-promoted internalization as a putative regulatory pathway. Herein, we provide evidence for two distinct processes of CXCR1 and CXCR2 regulation. Confocal microscopy data showed a redistribution of CXCR1 expression from the cell surface of neutrophils to internal compartments after stimulation with IL-8, whereas stimulation with bacterial lipopolysaccharide (LPS) or tumor necrosis factor- α (TNF-α) did not induce CXCR1 internalization but instead mediated a significant loss of membrane-proximal CXCR1 staining intensity. To investigate whether proteolytic cleavage was the mechanism responsible for LPS- and TNF-α-induced downmodulation of IL-8 receptors, we tested a panel of proteinase inhibitors. The downmodulation of CXCR1 and CXCR2 by LPS and TNF-α was most dramatically inhibited by metalloproteinase inhibitors; 1,10- phenanthroline and EDTA significantly attenuated LPS- and TNF-α-induced loss of CXCR1 and CXCR2 cell surface expression. Metalloproteinase inhibitors also blocked the release of CXCR1 cleavage fragments into the cell supernatants of LPS- and TNF-α-stimulated neutrophils. In addition, while treatment of neutrophils with LPS and TNF-α inhibited IL-8 receptor-mediated calcium mobilization and IL-8-directed neutrophil chemotaxis, both 1,10- phenanthroline and EDTA blocked these inhibitory processes. In contrast, metalloproteinase inhibitors did not affect IL-8-mediated downmodulation of CXCR1 and CXCR2 cell surface expression or receptor signaling. Thus, these findings may provide further insight into the mechanisms of leukocyte regulation during immunologic and inflammatory responses.
AB - The neutrophil-specific G-protein-coupled chemokine receptors, CXCR1 and CXCR2, bind with high affinity to the potent chemoattractant interleukin-8 (IL-8). The mechanisms of IL-8 receptor regulation are not well defined, although previous studies have suggested a process of ligand-promoted internalization as a putative regulatory pathway. Herein, we provide evidence for two distinct processes of CXCR1 and CXCR2 regulation. Confocal microscopy data showed a redistribution of CXCR1 expression from the cell surface of neutrophils to internal compartments after stimulation with IL-8, whereas stimulation with bacterial lipopolysaccharide (LPS) or tumor necrosis factor- α (TNF-α) did not induce CXCR1 internalization but instead mediated a significant loss of membrane-proximal CXCR1 staining intensity. To investigate whether proteolytic cleavage was the mechanism responsible for LPS- and TNF-α-induced downmodulation of IL-8 receptors, we tested a panel of proteinase inhibitors. The downmodulation of CXCR1 and CXCR2 by LPS and TNF-α was most dramatically inhibited by metalloproteinase inhibitors; 1,10- phenanthroline and EDTA significantly attenuated LPS- and TNF-α-induced loss of CXCR1 and CXCR2 cell surface expression. Metalloproteinase inhibitors also blocked the release of CXCR1 cleavage fragments into the cell supernatants of LPS- and TNF-α-stimulated neutrophils. In addition, while treatment of neutrophils with LPS and TNF-α inhibited IL-8 receptor-mediated calcium mobilization and IL-8-directed neutrophil chemotaxis, both 1,10- phenanthroline and EDTA blocked these inhibitory processes. In contrast, metalloproteinase inhibitors did not affect IL-8-mediated downmodulation of CXCR1 and CXCR2 cell surface expression or receptor signaling. Thus, these findings may provide further insight into the mechanisms of leukocyte regulation during immunologic and inflammatory responses.
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U2 - 10.1182/blood.v93.7.2173
DO - 10.1182/blood.v93.7.2173
M3 - Article
C2 - 10090924
AN - SCOPUS:0033120707
SN - 0006-4971
VL - 93
SP - 2173
EP - 2185
JO - Blood
JF - Blood
IS - 7
ER -