Role of IL-12 and IFN-gamma in Pseudomonas aeruginosa corneal infection.
OBJECTIVE
In Pseudomonas aeruginosa ocular infection, T-helper cell 1-responsive mouse strains are susceptible (the cornea perforates), and neutralization of IFN-gamma before infection has been shown to delay the onset of perforation. IFN-gamma is the predominant cytokine induced by IL-12, and positive regulation of IL-12 by IFN-gamma, if unchecked, leads to excessive cytokine production and toxicity. Despite its potential importance, the role of IL-12 in ocular infection with P. aeruginosa remains unexplored and was the purpose of this study.


METHODS
IL-12 knockout mice, histopathology, RT/PCR and ELISA analyses, immunocytochemistry, and quantitation of viable bacteria in cornea were used to examine the role of IL-12 in IFN-gamma production and the susceptibility phenotype.


RESULTS
To directly test the effect of IL-12 on IFN-gamma production, IL-12 knockout and wild-type C57BL/6 mice were used. Both groups of mice were susceptible to infection, with corneal perforation seen at 5 to 7 days after infection. RT-PCR and ELISA analyses confirmed that IL-12 message and protein levels were elevated after infection only in the wild-type mouse cornea. Other differences between the two groups were detected. Knockout versus wild-type mice showed a significant decrease in IFN-gamma mRNA levels in the cornea and cervical lymph nodes and decreased TNF-alpha protein levels in cornea. Corneas of knockout mice also had a significant increase in bacterial load at 5 days after infection when compared with wild-type mice.


CONCLUSIONS
These data provide evidence that IL-12 is important in IFN-gamma production and in the absence of the cytokine, both IFN-gamma and TNF-alpha levels in cornea are significantly decreased, resulting in unchecked bacterial growth and perforation.
Hazlett
Linda D
LD
Department of Anatomy and Cell Biology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA. lhazlett@med.wayne.edu
Rudner
Xiaowen L
XL
McClellan
Sharon A
SA
Barrett
Ronald P
RP
Lighvani
Shahrzad
S
D000818:Animals; D015169:Colony Count, Microbial; D003315:Cornea / Q000378:metabolism / Q000473:pathology; D003320:Corneal Ulcer / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology; D004797:Enzyme-Linked Immunosorbent Assay; D015818:Eye Infections, Bacterial / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology; D005260:Female; D007124:Immunoenzyme Techniques; D007371:Interferon-gamma / Q000235:genetics / Q000378:metabolism*; D018664:Interleukin-12 / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D011552:Pseudomonas Infections / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology; D011550:Pseudomonas aeruginosa / Q000302:isolation & purification; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D018664:Interleukin-12; D007371:Interferon-gamma