Different functions for the interleukin 8 receptors (IL-8R) of human neutrophil leukocytes: NADPH oxidase and phospholipase D are activated through IL-8R1 but not IL-8R2.
Two monoclonal antibodies, anti-IL8R1 and anti-IL8R2, raised against both interleukin 8 receptors (IL-8R) of human neutrophils, IL-8R1 and IL-8R2, were used to study individual receptor functions after stimulation with IL-8, GRO alpha, or NAP-2. Efficacy and selectivity of the antibodies were tested in Jurkat cells transfected with cDNA coding for one or the other receptor. The binding of 125 I labeled IL-8 and IL-8-induced changes of the cytosolic free Ca2+ concentration were inhibited by anti-IL8RI in cells expressing IL-8R1 and by anti-IL8R2 in cells expressing IL-8R2. In human neutrophils, release of elastase was observed after stimulation with IL-8 or GRO alpha. The response to IL-8 was inhibited slightly by anti-IL8R1 and more substantially when both monoclonal antibodies were present, while the response to GRO alpha was inhibited by anti-IL8R2 but was not affected by anti-IL8R1. These results indicate that both IL-8 receptors can signal independently for granule enzyme release. Superoxide production, a measure of the respiratory burst, was obtained with increasing concentrations of IL-8 with maximum effects at 25 to 50 nM, but no response was observed upon challenge with GRO alpha or NAP-2 up to 1000 nM. The superoxide production induced by IL-8 was inhibited by anti-IL8R1, but was not affected by anti-IL8R2. Stimulation of neutrophils with IL-8, in contrast to GRO alpha or NAP-2, also elicited phospholipase D activity. The effect of IL-8 was again inhibited by anti-IL-8R1 but not by anti-IL8R2, indicating that this response, like the respiratory burst, was mediated by IL-8R1. Taken together, our results show that IL-8R1 and IL-8R2 are functionally different. Responses, such as cytosolic free Ca2+ changes and the release of granule enzymes, are mediated through both receptors, whereas the respiratory burst and the activation of phospholipase D depend exclusively on stimulation through IL-8R1.
Jones
S A
SA
Theodor Kocher Institute, University of Bern, Switzerland.
Wolf
M
M
Qin
S
S
Mackay
C R
CR
Baggiolini
M
M
D015703:Antigens, CD / Q000145:classification / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D019272:Leukocyte Elastase; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D019255:NADPH Oxidases; D009504:Neutrophils / Q000378:metabolism*; D010196:Pancreatic Elastase / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D018123:Receptors, Interleukin / Q000145:classification / Q000378:metabolism*; D023062:Receptors, Interleukin-8A; D016897:Respiratory Burst; D014162:Transfection
D015703:Antigens, CD; D018123:Receptors, Interleukin; D023062:Receptors, Interleukin-8A; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D010739:Phospholipase D; D010196:Pancreatic Elastase; D019272:Leukocyte Elastase; D002118:Calcium