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.
Chandrasekar
B
B
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.
Colston
J T
JT
Geimer
J
J
Cortez
D
D
Freeman
G L
GL
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