Role of interleukin-15 and interleukin-18 in the secretion of sIL-6R and sgp130 by human neutrophils.
BACKGROUND
Available data indicate that neutrophils (PMN) produce a wide range of cytokines with the potential to modulate immune response. Recent investigation have shown that interleukin (IL)-15 and IL-18 potentiated several functions of normal neutrophils. It has been reported that IL-18-induced cytokine production may be significantly enhanced by coincident addition of IL-15.


OBJECTIVE
In the present study we compared the effect of recombinant human (rh)IL-15 and rhIL-18 as well as effect of a rhIL-15 and rhIL-18 combination on the induction secretion of sIL-6Ralpha and sgp130 by human neutrophils.


METHODS
PMN were isolated from heparinized whole blood of healthy persons. The PMN were cultured for 18 h at 37 degrees C in a humidified incubator with 5% CO(2). rhIL-15 and/or rhIL-18 and lipopolysaccharide were tested to PMN stimulation. The culture supernatants of PMN were removed and examined for the presence of sIL-6R and sgp130 by human enzyme-linked immunosorbent assay kits. Cytoplasmic protein fractions of PMN were analysed for the presence of sIL-6R and sgp130 by western blotting using monoclonal antibodies capable of detecting these proteins. Cells were lysed and cytoplasmic proteins were electrophoresed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The resolved proteins were transferred onto nitrocellulose and incubated with the primary monoclonal antibodies anti-sIL-6R and anti-sgp130. The membranes were incubated at room temperature with alkaline phosphatase anti-mouse immunoglobulin G. Immunoreactive protein bans were visualized by an AP Conjugate Substrate Kit.


CONCLUSIONS
The results of our investigation revealed that IL-15 alone, similarly to IL-18, has no significant ability for the regulation of both soluble IL-6 receptors, sIL-6R and sgp130, released by human neutrophils. It is interesting to note that the secretion of sgp130 was changed after PMN stimulation with rhIL-15 in the presence of rhIL-18. The combination of rhIL-15 and rhIL-18 was shown to induce PMN to secretion relatively higher amounts of sgp130 compared with the stimulation of PMN with rhIL-15 alone and rhIL-18 alone. The results obtained suggest that IL-15 and IL-18, belonging to the inflammatory cytokines, through the regulation of sgp130 secretion must be also considered as anti-inflammatory mediators that may influence the balance reactions mediated by the IL-6 cytokine family.
Jablonska
E
E
Department of Immunology, Medical University of Bialystok, Waszyngtona 15A 15-274 Bialystok, Poland. ewaj@amb.edu.pl
Marcinczyk
M
M
D015153:Blotting, Western; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D019409:Interleukin-15 / Q000494:pharmacology / Q000502:physiology*; D020382:Interleukin-18 / Q000494:pharmacology / Q000502:physiology*; D009025:Morpholines / Q000378:metabolism*; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D009994:Osmolar Concentration; D019947:Receptors, Interleukin-6 / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology; D012995:Solubility
D019409:Interleukin-15; D020382:Interleukin-18; D009025:Morpholines; D019947:Receptors, Interleukin-6; D011994:Recombinant Proteins; C005759:2-phenyl-5,5-dimethyltetrahydro-1,4-oxazine