Interleukin-1β stimulates IL-8 expression through MAP kinase and ROS signaling in human gastric carcinoma cells

Abstract

Recent studies have suggested that the expression of interleukin-8 (IL-8) directly correlates with the vascularity of human gastric carcinomas. In this study, the effect of IL-1b on IL-8 expression in human gastric cancer TMK-1 cells and the underlying signal transduction pathways were investigated. IL-1b induced the IL-8 expression in a time-and concentration-dependent manner. IL-1b induced the activation of extracellular signalregulated kinases-1/2 and P38 mitogen-activated protein kinase (MAPK), but not the activation of c-jun aminoterminal kinse and Akt. Specific inhibitors of MEK-1 (PD980590) and P38 MAPK (SB203580) were found to suppress the IL-8 expression and the IL-8 promoter activity. Expression of vectors encoding a mutated-type MEK-1 and P38 MAPK resulted in decrease in the IL-8 promoter activity. IL-1b also induced the production of reactive oxygen species (ROS). N-acetyl cysteine (NAC) prevented the IL-1b-induced ROS production and IL-8 expression. In addition, exogenous H 2 O 2 could induce the IL-8 expression. Deletional and site-directed mutagenesis studies on the IL-8 promoter revealed that activator protein-1 (AP-1) and nuclear factor (NF)-jB sites were required for the IL-1b-induced IL-8 transcription. Electrophoretic mobility shift assay confirmed that IL-1b increased the DNA-binding activity of AP-1 and NF-jB. Inhibitor (PD980590, SB203580) and ROS scavenger (NAC) studies revealed that the upstream signalings for the transcription factors AP-1 and NF-jB were MAPK and ROS, respectively. Conditioned media from the TMK-1 cells pretreated with IL-1b could remarkably stimulate the in vitro growth of HUVEC and this effect was partially abrogated by IL-8-neutralizing antibodies. The above results suggest that MAPK-AP-1 and ROS-NF-jB signaling pathways are involved in the IL-1b-induced IL-8 expression and that these paracrine signaling pathways induce endothelial cell proliferation.

Pro-inflammatory cytokines increase reactive oxygen species through mitochondria and NADPH oxidase in cultured RPE cells

“…Of note, the concentration of IL-1β (0.02 ng/ml) capable of inducing maximal intracellular RPE ROS levels is below the concentration (20 ng/ml) that maximally induces extracellular ROS; it is also lower than the concentrations found to maximally induced intracellular ROS in other cell types (Hwang et al, 2004), suggesting the potential importance of ROS in IL-1β-mediated signal transduction in the RPE.…”

“…Intracellular ROS Production-Intracellular ROS production by human RPE cells in response to TNF-α, IL-1β, or IFN-γ treatment was measured by a cell-based fluorometric assay based on deacetylation and oxidation of non-fluorescent reduced CM-H 2 DCFDA (Molecular Probes) into fluorescent CM-DCF. The reliability and specificity of the probe have been established in various cells including cultured human RPE cells (Deshpande et al, 2000;Nishikawa et al, 2000;Touyz et al, 2001;Yamagishi et al, 2001;Higgins et al, 2003;Li et al, 2003;Fukami et al, 2004;Hermann et al, 2004;Hwang et al, 2004;Kannan et al, 2004;Shanker et al, 2004;Kim et al, 2005). RPE cells were seeded into 96-well plates pre-coated with poly-D-lysine (Sigma-Aldrich) and cultured for 1 day or 7days.…”

“…TNF-α increases mitochondrial ROS production in tumor cells, endothelial cells, and hepatocytes (Schulze-Osthoff, 1993;Corda et al, 2001). IL-1β stimulates ROS production in various cell types (Mendes et al, 2003;Brigelius-Flohe et al, 2004;Hwang et al, 2004;Kaur et al, 2004). IFN-γ induces an immediate and marked augmentation of intracellular ROS in transformed lymphoblast cell lines (HSC536/N and PD149L) (Pearl-Yafe et al, 2003, 2004.…”

ATR-dependent radiation-induced γH2AX foci in bystander primary human astrocytes and glioma cells

“…a protein) produced by targeted cells and passed on through the medium or the induction of a (secondary) bystander signal in the Mechanisms of bystander cH2AX foci induction S Burdak-Rothkamm et al bystander cells themselves. As some cytokines can be induced by increased ROS levels (Bellocq et al, 1999;Ayache et al, 2002;Hsu and Wen, 2002;Kosmidou et al, 2002;Hwang et al, 2004;Ryan et al, 2004), they are likely candidates for a secondary bystander signal. Furthermore, some of these cytokines (e.g.…”

“…The involvement of soluble factors like reactive oxygen species (ROS) (Azzam et al, 2002;Shao et al, 2003aShao et al, , 2005, nitric oxide (NO) (Shao et al, 2003b(Shao et al, , 2005Sokolov et al, 2005) and cytokines released from irradiated cells as well as gap junction intercellular communication (Azzam et al, 2001;Shao et al, 2003a;Mitchell et al, 2004;Sokolov et al, 2005) have recently been reported. Most interestingly, several cytokines can be induced by ROS and NO/NOS (Ayache et al, 2002;Hsu and Wen, 2002;Kosmidou et al, 2002;Hwang et al, 2004;Ryan et al, 2004) providing a possible link between different factors potentially involved in bystander signalling. End points used for the study of bystander effects in vitro have included micronuclei formation (Azzam et al, 2002;Kashino et al, 2004;Shao et al, 2004), gene mutations and genomic instability (Zhou et al, 2000), gene expression changes (Azzam et al, 2002;Yang et al, 2005), transformation (Sawant et al, 2001), proliferation (Gerashchenko and Howell, 2003), cell survival, apoptosis (Belyakov et al, 2002;Lyng et al, 2002), cell cycle arrest (Azzam et al, 2000) and most recently the induction of gH2AX foci in bystander cells (Hu et al, 2005;Sokolov et al, 2005;Yang et al, 2005).…”