Abstract
Ischemia/reperfusion (I/R) injury is characterized by the induction of oxidative stress and proinflammatory cytokine expression. Recently demonstrating that oxidative stress and TNF-alpha each stimulate interleukin (IL)-18 expression in cardiomyocytes, we hypothesized that I/R also induces IL-18 expression and thus exacerbates inflammation and tissue damage. Neutralization of IL-18 signaling should therefore diminish tissue injury following I/R. I/R studies were performed using a chronically instrumented closed chest mouse model. Male C57BL/6 mice underwent 30 min of ischemia by LAD coronary artery ligation followed by various periods of reperfusion. Sham-operated or ischemia-only mice served as controls. A subset of animals was treated with IL-18-neutralizing antibodies 1 h prior to LAD ligation. Ischemic LV tissue was used for analysis. Our results demonstrate that, compared with sham operation and ischemia alone, I/R significantly increased (i) oxidative stress (increased MDA/4-HNE levels), (ii) neutrophil infiltration (increased MPO activity), (iii) NF-kappaB DNA binding activity (p50, p65), and (iv) increased expression of IL-18Rbeta, but not IL-18Ralpha or IL-18BP transcripts. Administration of IL-18-neutralizing antibodies significantly reduced I/R injury measured by reduced infarct size (versus control IgG). In isolated adult mouse cardiomyocytes, simulated ischemia/reperfusion enhanced oxidative stress and biologically active IL-18 expression via IKK-dependent NF-kappaB activation. These results indicate that IL-18 plays a critical role in I/R injury and thus represents a promising therapeutic target.
Highlights
Ischemia/reperfusion (I/R) injury is characterized by the induction of oxidative stress and proinflammatory cytokine expression
Circulating neutrophils recruited to the myocardium can exacerbate tissue injury by generating oxygen free radicals via the NADPH oxidase enzyme, releasing proteases and proinflammatory cytokines, and plugging microcapillaries, IL-18 has been shown to directly influence neutrophil function, since IL-18 activates p38 MAP kinase, primes NADPH oxidase, enhances chemotaxis, and increases azurophilic granule release [30]
During the reperfusion phase of the I/R injury, there was a significant increase in MPO activity within the ischemic myocardium (*, p Ͻ 0.0001; Fig. 1C); no increase in activity was seen during the ischemic phase
Summary
Ischemia/reperfusion (I/R) injury is characterized by the induction of oxidative stress and proinflammatory cytokine expression. In isolated adult mouse cardiomyocytes, simulated ischemia/reperfusion enhanced oxidative stress and biologically active IL-18 expression via IKK-dependent NF-B activation. These results indicate that IL-18 plays a critical role in I/R injury and represents a promising therapeutic target. The precise role of IL-18 in I/R injury and its regulation in vivo are incompletely understood, and the signal transduction pathshift assay; ELISA, enzyme-linked immunosorbent assay; GFP, green fluorescent protein; IFN, interferon; IB, inhibitory B; IKK, IB kinase; I/R, ischemia/reperfusion; sI/R, simulated I/R; LAD, left anterior descending; LPS, lipopolysaccharide; LV, left ventricle; MDA, malondialdehyde; 4-HNE, 4-hydroxyalkenals; MOI, multiplicity of infection; MPO, myeloperoxidase; NF-B, nuclear factor B; PDTC, pyrrolidine dithiocarbamate; RT, reverse transcription; qPCR, quantitative polymerase chain reaction; TNF, tumor necrosis factor; TTC, 2,3,5-triphenyltetrazolium chloride; PBS, phosphatebuffered saline; nt, nucleotides; DCFH-DA, 2Ј,7Ј-dichlorofluorescein diacetate; rRNA, ribosomal RNA; JNK, c-Jun N-terminal kinase; ROS, reactive oxygen species. Interleukin-18 and Reperfusion Injury ways involved in its induction in myocardial constituent cells in vitro have not been identified
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