SUMMARY: Nuclear factor-B (NF-B) is a transcriptional regulator of inducible expression of genes includingcyclooxygenase-2 (COX-2), regulating cell proliferation. NF-B is kept silent in the cytoplasm via interaction with the inhibitory protein IB␣ and transmigrated into the nucleus upon activation. However, constitutive NF-B has been found in the nucleus of some cancer cells. We investigated the role of NF-B in COX-2 expression and cell proliferation in human gastric cancer AGS cells. AGS cells were treated with antisense oligodeoxynucleotide (AS ODN) or sense oligodeoxynucleotide (S ODN) for the NF-B subunit p50, or they were transfected with a mutated IB␣ gene (MAD-3 mutant) or a control vector, pcDNA-3. AGS cells were treated with COX-2 inhibitors such as indomethacine and NS-398 or prostaglandin E 2 . mRNA expression for COX-2, and protein levels for p50, IB␣, and COX-2 were determined by reverse transcription polymerase chain reaction and Western blot analysis. The NF-B levels were examined by electrophoretic mobility shift assay. Thromboxane B 2 (TXB 2 ) and 6-ketoprostaglandin F 1␣ (6-keto-PGF 1␣ ) levels were determined by enzyme-linked immunosorbent assay. Cell proliferation was assessed by viable cell counting, [ Cell proliferation, mRNA expression and protein level of COX-2, and production of TXB 2 and 6-keto-PGF 1␣ were inhibited in cells treated with AS ODN or transfected with the mutated IB␣ gene, which had lower NF-B levels than cells treated with S ODN or transfected with control vector. COX-2 inhibitors suppressed cell proliferation and production of TXB 2 and 6-keto-PGF 1␣ , in a dose-dependant manner. Prostaglandin E 2 prevented the inhibition of proliferation in cells treated with AS ODN or transfected with the mutated IB␣ gene. In conclusion, NF-B mediates COX-2 expression, which may be related to cell proliferation, in human gastric cancer cells. (Lab Invest 2001, 81:349 -360).

Cell death linked to oxidative DNA damage has been implicated in acute pancreatitis. The severe DNA damage, which is beyond the capacity of the DNA repair proteins, triggers apoptosis. It has been hypothesized that oxidative stress may induce a decrease in the Ku70 and Ku80 levels and apoptosis in pancreatic acinar cells. In this study, it was found that oxidative stress caused by glucose oxidase (

SUMMARY:Reactive oxygen species (ROS) has been considered to be an important regulator in the development and pathogenesis of pancreatitis and an activator of the transcription factor, nuclear factor-B (NF-B), regulating inflammatory cytokine gene expression. NF-B activation was demonstrated in cerulein pancreatitis, which rapidly induces an acute, edematous form of pancreatitis. This study aimed to investigate whether cerulein induced ROS generation, lipid peroxide and hydrogen peroxide production, NF-B activation, and expression of cytokines (IL-1␤, IL-6) in pancreatic acinar cells. An additional aim was to establish whether these alterations were inhibited by antioxidants such as glutathione, superoxide dismutase, and catalase and an inhibitor of NF-B activation, pyrrolidine dithiocarbamate (PDTC). To determine the possible interactions of the antioxidants and PDTC with cerulein-induced signaling, Ca 2ϩ signal and amylase release were monitored in the pancreatic acinar cells treated with cerulein in the presence or absence of either the antioxidants or PDTC. The results showed that cerulein generated ROS and increased lipid peroxide and hydrogen peroxide production in the acinar cells, as determined by dichlorofluorescein diacetate dye. This resulted in NF-B activation and the induction of cytokine gene expression in the cells. The cerulein-induced NF-B activation was in parallel to IB␣ degradation. Cerulein also induced Ca 2ϩ signals and amylase release in acinar cells. Both antioxidants (glutathione, superoxide dismutase, catalase) and PDTC inhibited the cerulein-induced, oxidant-mediated alterations but did not affect the cerulein-evoked Ca 2ϩ signals and amylase release in acinar cells. In conclusion, ROS, generated by cerulein, activates NF-B, resulting in the up-regulation of inflammatory cytokine gene expression in acinar cells. NF-B inhibition by scavenging ROS might alleviate the inflammatory response in pancreatic acinar cells by suppressing cytokine gene expression. (Lab Invest 2002, 82:1359 -1368.

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Helicobacter pylori (H. pylori) infection leads to gastric inflammation, peptic ulcer and gastric carcinoma. H. pylori activates NADPH oxidase and increases reactive oxygen species (ROS), which induce NF-κB activation and IL-8 expression in gastric epithelial cells. Dysfunctional mitochondria trigger inflammatory cytokine production. Peroxisome proliferator-activated receptors-γ (PPAR-γ) regulate inflammatory response. Astaxanthin is a powerful antioxidant that protects cells against oxidative stress. The present study was aimed at determining whether astaxanthin inhibits H. pylori-induced mitochondrial dysfunction, NF-κB activation, and IL-8 expression via PPAR-γ activation in gastric epithelial cells. Gastric epithelial AGS cells were treated with astaxanthin, NADPH oxidase inhibitor apocynin and PPAR-γ antagonist GW9662, and infected with H. pylori. As a result, H. pylori caused an increase in intracellular and mitochondrial ROS, NF-κB activation and IL-8 expression, but decreased mitochondrial membrane potential and ATP level. Astaxanthin inhibited H. pylori-induced alterations (increased ROS, mitochondrial dysfunction, NF-κB activation, and IL-8 expression). Astaxanthin activated PPAR-γ and its target gene catalase in H. pylori-infected cells. Apocynin reduced ROS and inhibited IL-8 expression while astaxanthin did not affect NADPH oxidase activity. Inhibitory effects of astaxanthin on ROS levels and IL-8 expression were suppressed by addition of GW9662. In conclusion, astaxanthin inhibits H. pylori-induced mitochondrial dysfunction and ROS-mediated IL-8 expression by activating PPAR-γ and catalase in gastric epithelial cells. Astaxanthin may be beneficial for preventing oxidative stress-mediated gastric inflammation-associated H. pylori infection.