Interleukin (IL)-8 (CXCL8) induces cytokine expression and superoxide formation by guinea pig neutrophils infected with Mycobacterium tuberculosis.
METHODS
Interleukin (IL)-8, a neutrophil attracting chemokine, is known to be made by a variety of leukocyte populations following stimulation by Mycobacterium tuberculosis.


OBJECTIVE
The effect of recombinant guinea pig IL-8 on the ability of neutrophils to generate a cytokine response after infection with M. tuberculosis H37Ra was examined.


METHODS
Recombinant gpIL-8 was produced by subcloning the gene into Escherichia coli and purification over a nickel column. The identity of the rgpIL-8 was confirmed by sequencing. Neutrophils were harvested from the blood of non-vaccinated or M. bovis BCG-vaccinated guinea pigs and tested for their ability to migrate toward media alone, 10 microg/ml PPD, f-Met-Leu-Phe (f-MLP), or rgpIL-8 in 96-well chemotactic chambers. Neutrophils were also pre-stimulated with rgpIL-8 then restimulated with LPS (10 microg/ml) or infected in vitro with M. tuberculosis H37Ra (MOI 1:1).


RESULTS
Recombinant gpIL-8 and f-MLP induced significant chemotaxis in neutrophils from both non-vaccinated and BCG-vaccinated guinea pigs, with the best chemotaxis occurring at a concentration of 10(-7)M. Real-time PCR analysis revealed that pre-treatment of neutrophils induced elevated levels of IL-8 and TNF-alpha mRNA and protein as well as superoxide, but not mRNA for MCP-1, IFN-gamma, or TGF-beta when compared to neutrophils pre-stimulated with media alone.


CONCLUSIONS
The presence of IL-8 early in the host response to M. tuberculosis infection may be an important contributor to a successful immune response. How essential a role IL-8 plays remains unknown and merits further study.
Lyons
Mark J
MJ
Department of Medical Microbiology and Immunology, Texas A&M University, System Health Science Center, Reynolds Medical Building Room 463, University Drive, College Station, TX 77843-1114, USA.
Yoshimura
Teizo
T
McMurray
David N
DN
D000818:Animals; D001500:BCG Vaccine / Q000008:administration & dosage; D002478:Cells, Cultured; D002634:Chemotaxis, Leukocyte; D016207:Cytokines / Q000378:metabolism*; D006168:Guinea Pigs; D016209:Interleukin-8 / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D018375:Neutrophil Activation; D009504:Neutrophils / Q000187:drug effects* / Q000276:immunology / Q000382:microbiology*; D011994:Recombinant Proteins / Q000494:pharmacology; D013481:Superoxides / Q000032:analysis / Q000378:metabolism*; D014376:Tuberculosis / Q000276:immunology*
D001500:BCG Vaccine; D016207:Cytokines; D016209:Interleukin-8; D008070:Lipopolysaccharides; D011994:Recombinant Proteins; D013481:Superoxides