• ALGAE
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• v.27 no.1
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• pp.21-32
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• 2012

We report an oxidative burst triggered by prostaglandin $A_2(PGA_2)$ in the brown algal kelp Laminaria digitata, constituting the first such discovery in an alga and the second finding of an oxidative burst triggered by a prostaglandin in a living organism. The response is more powerful than the oxidative burst triggered by most other chemical elicitors in Laminaria. Also, it is dose-dependent and cannot be inhibited by diphenylene iodonium, suggesting that another source than NAD(P)H oxidase is operational in the production of reactive oxygen species. Despite the very strong oxidative response, rather few effects at other levels of signal transduction pathways could be identified. $PGA_2$ does not increase lipolysis (free fatty acids) in Laminaria, and only one oxylipin (15-hydroxyeicosatetraenoic acid; 15-HETE) was found to be upregulated in Laminaria. In a subsequent set of experiments in the genome model Ectocarpus siliculosus, none of 5 selected candidate genes, all established participants in various stress responses, showed any significant differences in their expression profiles.

• Archives of Pharmacal Research
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• v.28 no.10
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• pp.1183-1189
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• 2005

Although plant-derived phenolic acids have been reported to have anti-cancer activity, the exact mechanism is not completely understood. In this study, we investigated the role for reactive oxygen species (ROS) as a mediator of the apoptosis induced by caffeic acid (CA) and ferulic acid (FA), common phenolic acids in plants in HepG2 human hepatoma cells. CA and FA reduced cell viability, and induced apoptotic cell death in a dose-dependent manner. In addition, they evoked a dose-related elevation of intracellular ROS. Treatment with various inhibitors of NADPH oxidase (diphenylene iodonium, apocynin, neopterine) significantly blunted both the generation of ROS and the induction of apoptosis induced CA and FA. These results suggest that ROS generated through activation of NADPH oxidase may play an essential role in the apoptosis induced by CA and FA in HepG2 cells. These results further suggest that CA and FA may be valuable for the therapeutic management of human hepatomas.

• YAKHAK HOEJI
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• v.56 no.2
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• pp.80-85
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• 2012

Previously, we have reported that arachidonic acid (AA) appears to be involved in the induction of apoptosis in HepG2 human hepatoblastoma cells. In this study we investigated the possible role of the NADPH oxidase, a membranebound enzyme generating reactive oxygen species (ROS), in the mechanism of AA-induced apoptosis in HepG2 cells. Apoptotic cell death induced by AA was significantly suppressed by various inhibitors of the NADPH oxidase, diphenylene iodonium (DPI), apocynin (Apo) and neopterine (NP). In addition, these inhibitors of the NADPH oxidase completely blunted the AA-induced ROS elevation. Next, we investigated the implication of metabolic pathway of AA in these AA actions. Both apoptosis and ROS production induced by AA were not significantly altered by treatment with indomethacin (Indo) or nordihydroguaiaretic acid (NDGA), selective inhibitors of cyclooxygenase (COX) and lipoxygenase (LOX), respectively, suggesting that AA metabolites produced by COX or LOX may not have an essential role in the AA-induced apoptosis and ROS generation. Collectively, these results suggest that the NADPH oxidase may be a key player in the mechanism of AA-induced apoptosis in HepG2 cells. These results further suggest that NADPH oxidase may be a good target for the management of human hepatomas.

• YAKHAK HOEJI
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• v.55 no.6
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• pp.485-491
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• 2011

Previously, we have reported that apigenin, a natural flavonoid found in a variety of vegetables and fruits, stimulated melanogenesis through the activation of $K^+-Cl^-$-cotransport (KCC) in B16 melanoma cells. In this study we investigated the possible involvement of reactive oxygen species (ROS) in the mechanism of apigenin-induced melanogenesis in B16 cells. Apigenin elevated intracellular ROS level in a dose-dependent manner. Treatment with various inhibitors of NADPH oxidase, diphenylene iodonium (DPI), apocynin (Apo) and neopterine (NP) significantly inhibited both the generation of ROS and melanogenesis induced by apigenin. In addition these inhibitors profoundly inhibited apigenin-induced $Cl^-$-dependent $K^+$ efflux, a hallmark of KCC activity. However, the apigenin-induced ROS generation was not significantly affected by treatment with a specific KCC inhibitor R-(+)-[(2-n-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy]acetic acid (DIOA). These results indicate that the ROS production may be a upstream regulator of the apigenin-induced KCC stimulation, and in turn, melanogenesis in the B16 cells. Taken together, these results suggest that the NADPH oxidase-mediated ROS production may play an important role in the apigenin-induced melanogenesis in B16 cells. These results further suggest that NADPH oxidase may be a good target for the management of hyperpigmentation disorders.

• The Korean Journal of Physiology and Pharmacology
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• v.24 no.3
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• pp.267-276
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• 2020

T In the present study, we investigated the effect of oncogenic H-Ras on rat mdr1b expression in NIH3T3 cells. The constitutive expression of H-Ras^V12 was found to downregulate the mdr1b promoter activity and mdr1b mRNA expression. The doxorubicin-induced mdr1b promoter activity of the H-Ras^V12 expressing NIH3T3 cells was markedly lower than that of control NIH3T3 cells. Additionally, there is a positive correlation between the level of H-Ras^V12 expression and a sensitivity to doxorubicin toxicity. To examine the detailed mechanism of H-Ras^V12-mediated down-regulation of mdr1b expression, antioxidant N-acetylcysteine (NAC) and NADPH oxidase inhibitor diphenylene iodonium (DPI) were used. Pretreating cells with either NAC or DPI significantly enhanced the oncogenic H-Ras-mediated down-regulation of mdr1b expression and markedly prevented doxorubicin-induced cell death. Moreover, NAC and DPI treatment led to a decrease in ERK activity, and the ERK inhibitors PD98059 or U0126 enhanced the mdr1b-Luc activity of H-Ras^V12-NIH3T3 and reduced doxorubicin-induced apoptosis. These data suggest that Ras^V12 expression could downregulate mdr1b expression through intracellular reactive oxygen species (ROS) production, and ERK activation induced by ROS, is at least in part, contributed to the downregulation of mdr1b expression.

• The Korean Journal of Physiology and Pharmacology
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• v.15 no.6
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• pp.319-326
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• 2011

Quercetin-3-O-${\beta}$-D-glucuronopyranoside (QGC) is a flavonoid glucoside extracted from Rumex Aquaticus Herba. We aimed to explore its protective effect against ethanol-induced cell damage and the mechanism involved in the effect in feline esophageal epithelial cells (EEC). Cell viability was tested and 2',7'-dichlorofluorescin diacetate assay was used to detect intracellular $H_2O_2$ production. Western blotting analysis was performed to investigate MAPK activation and interleukin 6 (IL-6) expression. Exposure of cells to 10% ethanol time-dependently decreased cell viability. Notably, exposure to ethanol for 30 min decreased cell viability to 43.4%. When cells were incubated with $50{\mu}M$ QGC for 12 h prior to and during ethanol treatment, cell viability was increased to 65%. QGC also inhibited the $H_2O_2$ production and activation of ERK 1/2 induced by ethanol. Pretreatment of cells with the NADPH oxidase inhibitor, diphenylene iodonium, also inhibited the ethanol-induced ERK 1/2 activation. Treatment of cells with ethanol for 30 or 60 min in the absence or presence of QGC exhibited no changes in the IL-6 expression or release compared to control. Taken together, the data indicate that the cytoprotective effect of QGC against ethanol-induced cell damage may involve inhibition of ROS generation and downstream activation of the ERK 1/2 in feline EEC.

• Preventive Nutrition and Food Science
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• v.12 no.2
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• pp.74-78
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• 2007

Fucoidan, that is high-molecular-weight sulfated polysaccharides extracted from brown seaweeds has been shown to elicit various biological activities. Here, we investigated the effects of fucoidan on cell proliferation and nitric oxide (NO) production in neuronal blastoma cell (SH-SY5Y). In the present study, we demonstrated that fucoidan treatment resulted in increase of cell proliferation and NO production. When cells were treated with amyloid-${\beta}$ (A${\beta}$) in the absence or presence of fucoidan, fucoidan recovered the cell viability decreased by A${\beta}$ peptides. To further determine whether nitric oxide synthase (NOS) is involved in proliferative effect of fucoidan, cells were treated with NOS inhibitors in the absence or presence of fucoidan. Selective constitutive nitric oxide synthase (cNOS) inhibitor, diphenylene iodonium chloride (DPI), caused a decrease of cell viability, whereas cell viability was increased by specific inducible nitric oxide synthase (iNOS) inhibitor, S-methylisothiourea (SMT), in the fucoidan-untreated cells. Treatment with fucoidan inhibited the cell viability decreased in DPI-exposed cells. In contrast, fucoidan had no effect on cell growth in SMT-treated cells, indicating that cNOS may not play a role in the proliferation of fucoidan-treated cells. The present data suggest that fucoidan has proliferative and neuroprotective effects and these effects may be associated with iNOS.

• Journal of Microbiology and Biotechnology
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• v.26 no.4
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• pp.790-798
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• 2016

Chlamydiae, obligate intracellular bacteria, are associated with a variety of human diseases. The chlamydial life cycle undergoes a biphasic development: replicative reticulate bodies (RBs) phase and infectious elementary bodies (EBs) phase. At the end of the chlamydial intracellular life cycle, EBs have to be released to the surrounded cells. Therefore, the interactions between Chlamydiae and cell death pathways could greatly influence the outcomes of Chlamydia infection. However, the underlying molecular mechanisms remain elusive. Here, we investigated host cell death after Chlamydia infection in vitro, in L929 cells, and showed that Chlamydia infection induces cell necrosis, as detected by the propidium iodide (PI)-Annexin V double-staining flow-cytometric assay and Lactate dehydrogenase (LDH) release assay. The production of reactive oxygen species (ROS), an important factor in induction of necrosis, was increased after Chlamydia infection, and inhibition of ROS with specific pharmacological inhibitors, diphenylene iodonium (DPI) or butylated hydroxyanisole (BHA), led to significant suppression of necrosis. Interestingly, live-cell imaging revealed that Chlamydia infection induced lysosome membrane permeabilization (LMP). When an inhibitor upstream of LMP, CA-074-Me, was added to cells, the production of ROS was reduced with concomitant inhibition of necrosis. Taken together, our results indicate that Chlamydia infection elicits the production of ROS, which is dependent on LMP at least partially, followed by induction of host-cell necrosis. To our best knowledge, this is the first live-cell-imaging observation of LMP post Chlamydia infection and report on the link of LMP to ROS to necrosis during Chlamydia infection.