Induction of nuclear factor kappaB but not kappaB-responsive cytokine expression during myocardial reperfusion injury after neutropenia.
Neutrophils may contribute to myocardial ischemia/reperfusion (I/R) injury by generating reactive oxygen intermediates (ROIs). ROIs activate nuclear factor (NF)-kappaB, which regulates genes for cytokines with negative inotropic effects (interleukin [IL]-1beta, IL-6, and tumor necrosis factor [TNF]-alpha). We investigated the impact of neutrophil depletion on NF-kappaB DNA binding activity, and expression of these cytokines during myocardial I/R injury. Male WKY rats (n = 28) received a single dose of antineutrophil antiserum (i/v). Twenty two hours later, when the peripheral blood neutrophil counts were profoundly decreased (94% reduction), the animals underwent 15 min of left anterior descending coronary artery ligation followed by reperfusion for 0.25, 0.5, 1, 2, 3, and 6 h (n = 4/group). Saline-treated animals underwent a similar protocol, and served as controls (n = 28, 4/group). Neutrophil accumulation, defined by myeloperoxidase activity, was present in controls, but not in anti-PMN antisera-treated animals (at least p <0.05 at 1, 2, 3, and 6 h R). Despite this difference, in both saline- and antiserum-treated animals, the GSH levels were very similar and fell significantly (p < 0.0001) at 15 min R; the levels increased gradually over time. In contrast, GSSG levels rose at 15 and 30 min R (p < 0.05), and declined thereafter. NF-kappaB DNA binding activity increased in both groups at 15 min and again at 3 h of R. Both NF-kappaBp50 and p65 subunits were detected by supershift assay. In saline-injected controls both mRNA and protein for IL-1beta, IL-6, and TNF-alpha were detected at 1 h R; levels remained high until 3 h, then fell (except IL-6, which was elevated at 6 h). In neutropenic animals, however, a significant decrease in mRNA (at least 1.7-fold, p < 0.05) as well as protein levels (at least 2. 3-fold, p < 0.01) for all three cytokines was observed. Thus, while neutrophils had minimal effects on oxidative stress (GSH/GSSG) and oxidative stress-responsive NF-kappaB activity, they contributed significantly to myocardial cytokine expression.
Department of Medicine, University of Texas Health Science Center and South Texas Veterans Health Care System, Audie L. Murphy Division, San Antonio, TX 78229-3900, USA.
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B
Chandrasekar
J T
JT
Colston
J
J
Geimer
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D
Cortez
G L
GL
Freeman
D000818:Animals; D016207:Cytokines / Q000235:genetics* / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D015870:Gene Expression; D005978:Glutathione / Q000378:metabolism; D018836:Inflammation Mediators / Q000378:metabolism; D007375:Interleukin-1 / Q000235:genetics / Q000378:metabolism; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism; D008297:Male; D015428:Myocardial Reperfusion Injury / Q000235:genetics* / Q000378:metabolism*; D016328:NF-kappa B / Q000096:biosynthesis*; D009503:Neutropenia / Q000235:genetics* / Q000378:metabolism*; D009504:Neutrophils / Q000378:metabolism; D009195:Peroxidase / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011921:Rats, Inbred WKY; D017382:Reactive Oxygen Species / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism
D016207:Cytokines; D018836:Inflammation Mediators; D007375:Interleukin-1; D015850:Interleukin-6; D016328:NF-kappa B; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D004247:DNA; D009195:Peroxidase; D005978:Glutathione