Regulation of interleukin-8 binding and function by heparin and alpha2-macroglobulin.
BACKGROUND
Increased expression of interleukin-8 (IL-8), a potent neutrophil chemoattractant, is associated with a number of inflammatory diseases. Interleukin-8 binds to the glycosaminoglycan (GAG) heparin and the protease inhibitor alpha2-macroglobulin, molecules which regulate the function of a number of cytokines. Heparan sulphate was previously shown to enhance neutrophil chemotactic responses to IL-8.


OBJECTIVE
The purpose of this study was to investigate the effect of heparin, heparan sulphate and alpha2-macroglobulin on IL-8 binding to neutrophils and subsequent functional effects in vitro.


METHODS
The binding of 125I-IL-8 to normal neutrophils at 4 degrees C was studied and the IL-8 induced neutrophil chemotactic response was investigated using micro-Boyden chambers. Complexation of IL-8 with alpha2-macroglobulin was confirmed using gel filtration chromatography.


RESULTS
Heparin, but not heparan sulphate, inhibited the binding of 125I-IL-8 to neutrophils (IC50=26 microg/mL) and IL-8 induced neutrophil chemotactic responses (IC50=4 microg/mL). The specific inhibitory effect of heparin was apparently due to an interaction with IL-8 which was charge-dependent, since dextran sulphate had a greater inhibitory effect on chemotactic responses (IC50=2 microg/mL) and FITC-heparin did not bind to neutrophils. The heparin-induced inhibition of IL-8 binding and chemotactic responses was reversed in a dose-dependent manner in the presence of alpha2-macroglobulin. The binding of 125I-IL-8 to neutrophils in the presence of alpha2-macroglobulin appears to be, in part, through the specific IL-8 receptor.


CONCLUSIONS
These results point to an anti-inflammatory role for heparin and a novel, potentially, pro-inflammatory role for alpha2-macroglobulin which together indicate the importance of cytokine-binding macromolecules in determining net cytokine function.
Ramdin
L
L
University Medicine, Southampton General Hospital, UK.
Perks
B
B
Sheron
N
N
Shute
J K
JK
D015703:Antigens, CD / Q000378:metabolism; D002634:Chemotaxis, Leukocyte / Q000187:drug effects; D002850:Chromatography, Gel; D006493:Heparin / Q000378:metabolism / Q000494:pharmacology*; D006497:Heparan Sulfate / Q000494:pharmacology; D006801:Humans; D015151:Immunoblotting; D016209:Interleukin-8 / Q000378:metabolism / Q000502:physiology*; D009504:Neutrophils / Q000378:metabolism; D011485:Protein Binding; D018123:Receptors, Interleukin / Q000378:metabolism; D023062:Receptors, Interleukin-8A; D000511:alpha-Macroglobulins / Q000494:pharmacology*
D015703:Antigens, CD; D006493:Heparin; D006497:Heparan Sulfate; D016209:Interleukin-8; D018123:Receptors, Interleukin; D023062:Receptors, Interleukin-8A; D000511:alpha-Macroglobulins