• The Korean Journal of Physiology and Pharmacology
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• 제15권2호
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• pp.83-88
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• 2011

A large body of evidence has indicated that induction of endogenous antioxidative proteins seems to be a reasonable strategy for delaying the progression of cell injury. In our previous study, cilostazol was found to increase the expression of the antioxidant enzyme heme oxygenase-1 (HO-1) in synovial cells. Thus, the present study was undertaken to examine whether cilostazol is able to counteract tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$)-induced cell death in endothelial cells via the induction of HO-1 expression. We exposed human umbilical vein endothelial cells (HUVECs) to TNF-${\alpha}$ (50 ng/ml), with or without cilostazol ($10{\mu}M$). Pretreatment with cilostazol markedly reduced TNF-${\alpha}$-induced viability loss in the HUVECs, which was reversed by zinc protoporphyrine IX (ZnPP), an inhibitor of HO-1. Moreover, cilostazol increased HO-1 protein and mRNA expression. Cilostazol-induced HO-1 induction was markedly attenuated not only by ZnPP but also by copper-protoporphyrin IX (CuPP). In an assay measuring peroxisome proliferator-activated receptor-${\gamma}$ (PPAR-${\gamma}$) transcription activity, cilostazol directly increased PPAR-${\gamma}$ transcriptional activity which was completely abolished by HO-1 inhibitor. Furthermore, increased PPAR-${\gamma}$ activity by cilostazol and rosiglitazone was completely abolished in cells transfected with HO-1 siRNA. Taken together, these results indicate that cilostazol up-regulates HO-1 and protects cells against TNF-${\alpha}$-induced endothelial cytotoxicity via a PPAR-${\gamma}$-dependent pathway.

• Biomolecules & Therapeutics
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• 제21권1호
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• pp.60-65
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• 2013

3,4,5-Trihydroxycinnamic acid (THC) is a derivative of hydroxycinnamic acids, which have been reported to possess a variety of biological properties such as anti-inflammatory, anti-tumor, and neuroprotective activities. However, biological activity of THC has not been extensively examined. Recently, we reported that THC possesses anti-inflammatory activity in LPS-stimulated BV2 microglial cells. However, its precise mechanism by which THC exerts anti-inflammatory action has not been clearly identified. Therefore, the present study was carried out to understand the anti-inflammatory mechanism of THC in BV2 microglial cells. THC effectively suppressed the LPS-induced induction of pro-inflammatory mediators such as NO, TNF-${\alpha}$, and IL-$1{\beta}$. THC also suppressed expression of MCP-1, which plays a key role in the migration of activated microglia. To understand the underlying mechanism by which THC exerts these anti-inflammatory properties, involvement of Nrf2, which is a cytoprotective transcription factor, was examined. THC resulted in increased phosphorylation of Nrf2 with consequent expression of HO-1 in a concentration-dependent manner. THC-induced phosphorylation of Nrf2 was blocked with SB203580, a p38 MAPK inhibitor, indicating that p38 MAPK is the responsible kinase for the phosphorylation of Nrf2. Taken together, the present study for the first time demonstrates that THC exerts anti-inflammatory properties through the activation of Nrf2 in BV2 microglial cells, suggesting that THC might be a valuable therapeutic adjuvant for the treatment of inflammation-related disorders in the CNS.

• Natural Product Sciences
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• 제24권1호
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• pp.28-35
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• 2018

Pulegone is a naturally occurring organic compound obtained from essential oils from a variety of plants. The aim of this study was to investigate the anti-inflammatory effects through the inhibitory mechanism of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX-2), nuclear factor kappa B ($NF-{\kappa}B$), mitogen-activated protein kinases (MAPK) pathways and the activation of nuclear factor erythroid 2-related factor 2 (Nrf2)/ heme oxygenase (HO)-1 pathways in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. Results revealed that pulegone significantly inhibited NO production as well as iNOS and COX-2 expressions. Meanwhile, western blot analysis showed that pulegone down-regulated LPS-induced $NF-{\kappa}B$ and MAPKs activation in RAW 264.7 cells. Furthermore, the selected compound suppressed LPS-induced intracellular ROS production in RAW 264.7 cells, while the expression of stress response gene, HO-1, and its transcriptional activator, Nrf-2 was upregulated upon pulegone treatment. Taking together, these findings provided that pulegone inhibited the LPS-induced expression of inflammatory mediators via the down-regulation iNOS, COX-2, $NF-{\kappa}B$, and MAPKs signaling pathways as well as up-regulation of Nrf-2/HO-1 indicating that pulegone has a potential therapeutic and preventive application in various inflammatory diseases.

• Natural Product Sciences
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• 제24권1호
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• pp.13-20
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• 2018

Estragole is a naturally occurring phenylpropanoid obtained from essential oils found in a broad diversity of plants. Although the phenylpropanoids show many biological activities, clear regulation of the inflammatory signaling pathways has not yet been determined. Here, we scrutinized the anti-inflammatory effect of estragole. The anti-inflammatory effect of estragole was determined through the inhibitory mechanisms of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX-2), nuclear factor kappa B ($NF-{\kappa}B$), and mitogen-activated protein kinases (MAPK) pathways and the activation of nuclear factor erythroid 2-related factor 2 (Nrf-2)/heme oxygenase (HO)-1 pathways in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. Estragole significantly inhibited NO production, iNOS and COX-2 expression as well as LPS-induced $NF-{\kappa}B$ and MAPK activation. Furthermore, estragole suppressed LPS-induced intracellular ROS production but up-regulated the stress response gene HO-1 via the activation of transcription factor Nrf-2. These findings demonstrate that estragole inhibits the LPS-induced expression of inflammatory mediators via the down-regulation of iNOS, COX-2, $NF-{\kappa}B$, and MAPK pathways, as well as the up-regulation of the Nrf-2/HO-1 pathway, indicating that this phenylpropanoid has potential therapeutic and preventive applications in various inflammatory diseases.

• 생약학회지
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• 제51권3호
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• pp.163-170
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• 2020

Mahaenggamsuk-tang (MHGS) has been widely used in Korea and China for the treatment of various diseases. MHGS was constituted the Ephedrea Herba, Armenicae Semen, Glycyrrhizae Radix and Gypsum Fibrosum. In this study, we have made three different solvents extract as MHGS water extract (MHGS-W), MHGS 50% EtOH extract (MHGS-50E), and MHGS 100% EtOH extract (MHGS-100E). The MHGS-W, MHGS-50E and MHGS-100E showed the discernible difference patterns on HPLC analysis. Furthermore, MHGS-50E and MHGS-100E significantly increased the DPPH and ABTS radical scavenging effects than MHGS-W. In addition, the MHGS-50E and MHGS-100E also inhibited significantly nitric oxide (NO) and prostaglandin E2 (PGE₂) production, and inducible nitric oxide synthase (iNOS) cyclooxygenase-2 (COX-2) protein expression in RAW264.7. On the other hand, MHGS-50E and MHGS-W showed remarkable protection on the HT22 cell via heme oxygenase (HO)-1, but MHGS-100E did not show. The results of this study proved that MHGS-50E has greater potential therapeutic uses by exerting antioxidant, anti-inflammatory and neuroprotective effects compared to MHGS-100E, MHGS-W. Our study suggests that the different solvent might be affected the biological activities when make the traditional herbal medicines including MHGS.

• Molecules and Cells
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• 제38권5호
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• pp.416-425
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• 2015

NF-E2-related factor 2 (Nrf2), a basic leucine zipper transcription factor, has recently received a great deal of attention as an important molecule that enhances antioxidative defenses and induces resistance to chemotherapy or radiotherapy. In this study, we investigated the apoptosis-inducing and Nrf2- upregulating effects of quercetin on malignant mesothelioma (MM) MSTO-211H and H2452 cells. Quercetin treatment inhibited cell growth and led to upregulation of Nrf2 at both the mRNA and protein levels without altering the ubiquitination and extending the half-life of the Nrf2 protein. Following treatment with quercetin, analyses of the nuclear level of Nrf2, Nrf2 antioxidant response element-binding assay, Nrf2 promoter-luc assay, and RT-PCR toward the Nrf2-regulated gene, heme oxygenase-1, demonstrated that the induced Nrf2 is transcriptionally active. Knockdown of Nrf2 expression with siRNA enhanced cytotoxicity due to the induction of apoptosis, as evidenced by an increase in the level of proapoptotic Bax, a decrease in the level of antiapoptotic Bcl-2 with enhanced cleavage of caspase-3 and PARP proteins, the appearance of a sub-$G_0/G_1$ peak in the flow cytometric assay, and increased percentage of apoptotic propensities in the annexin V binding assay. Effective reversal of apoptosis was observed following pretreatment with the pan-caspase inhibitor Z-VAD. Moreover, Nrf2 knockdown exhibited increased sensitivity to the anticancer drug, cisplatin, presumably by potentiating the oxidative stress induced by cisplatin. Collectively, our data demonstrate the importance of Nrf2 in cytoprotection, survival, and drug resistance with implications for the potential significance of targeting Nrf2 as a promising strategy for overcoming resistance to chemotherapeutics in MM.

• Biomolecules & Therapeutics
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• 제20권6호
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• pp.499-505
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• 2012

Chemoprevention represents a strategy designed to protect cells or tissues against various carcinogens and carcinogenic metabolites derived from exogenous or endogenous sources. Recent studies indicate that plant-derived triterpenoids, like oleanolic acid, may exert cytoprotective functions via regulation of the activity of different transcription factors. The chemopreventive effects may be mediated through induction of the nuclear factor erythroid 2-related factor 2 (Nrf2) transcription factor. Activation of Nrf2 by triterpenoids induces the expression of phase 2 detoxifying and antioxidant enzymes such as NAD(P)H quinone oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1) - proteins which can protect cells or tissues against various toxic metabolites. On the other hand, inhibition of other transcription factors, like NF-${\kappa}B$ leads to the decrease in the pro-inflammatory gene expression. Moreover, the modulation of microRNAs activity may constitute a new mechanism responsible for valuable effects of triterpenoids. Recently, based on the structure of naturally occurring triterpenoids and with involvement of bioinformatics and computational chemistry, many synthetic analogs with improved biological properties have been obtained. Data from in vitro and in vivo experiments strongly suggest synthetic derivatives as promising candidates in the chemopreventive and chemotherapeutic strategies.

• Biomolecules & Therapeutics
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• 제20권6호
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• pp.562-568
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• 2012

Cytochrome P450 BM3 (CYP102A1) from Bacillus megaterium is a self-sufficient monooxygenase that consists of a heme domain and FAD/FMN-containing reductase domain (BMR). In this report, the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) by BMR was evaluated as a method for monitoring BMR activity. The electron transfer proceeds from NADPH to BMR and then to BMR substrates, MTT and CTC. MTT and CTC are monotetrazolium salts that form formazans upon reduction. The reduction of MTT and CTC followed classical Michaelis-Menten kinetics ($k_{cat}=4120\;min^{-1}$, $K_m=77{\mu}M$ for MTT and $k_{cat}=6580\;min^{-1}$, $K_m=51{\mu}M$ for CTC). Our continuous assay using MTT and CTC allows the simple, rapid measurement of BMR activity. The BMR was able to metabolize mitomycin C and doxorubicin, which are anticancer drug substrates for CPR, producing the same metabolites as those produced by CPR. Moreover, the BMR was able to interact with CYP1A2 and transfer electrons to promote the oxidation reactions of substrates by CYP1A2 and CYP2E1 in humans. The results of this study suggest the possibility of the utilization of BMR as a surrogate for mammalian CPR.

• Nutrition Research and Practice
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• 제7권6호
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• pp.475-480
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• 2013

The purpose of this study was to determine the antioxidant effect of fucoxanthin. After rats were fed a normal fat diet (NF), high fat diet (HF), and high fat with 0.2% fucoxanthin diet (HF + Fxn) for 4 weeks, the markers of oxidative stress and antioxidant capacity like lipid peroxidation, plasma total antioxidant capacity (TAC), and activities of antioxidant enzymes (catalase, superoxide dismutase (SOD), and gluthathione peroxidase (GSH-Px)) were determined. mRNA expression of transcription factor, nuclear erythroid factor like 2 (Nrf2), and its target genes such as NAD(P)H quinone oxidoreductase1 (NQO1) and heme oxygenase-1 (HO-1) were also determined. Mean weight gain in the HF + Fxn group was lower, without statistical significance, and the total food intake in the HF + Fxn group was lower than that in the HF group (P < 0.05). The activity of GSH-Px (P < 0.05) in plasma was significantly higher in the HF + Fxn group than those in the HF group (P < 0.05). In the liver, the activities of catalase (P < 0.05) and GSH-Px (P < 0.05) in the HF + Fxn group were significantly higher than those in the HF group. Plasma TAC level was significantly higher in the HF + Fxn group than that in the HF group (P < 0.05). Lipid peroxidation in plasma tended to be lower without statistical significance. Fucoxanthin supplements were shown to have higher mRNA expression of Nrf2 and NQO1 than those in the high fat diet only group (P < 0.05). In conclusion, supplementation of fucoxanthin improved the antioxidant capacity, depleted by high fat diet, by activating the Nrf2 pathway and its downstream target gene NQO1. Therefore, supplementation of fucoxanthin, especially for those who consume high fat in their diet, may benefit from reduced risk of oxidative stress.

• 한방재활의학과학회지
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• 제30권1호
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• pp.47-61
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• 2020

Objectives This study is carried out to investigate the effects of Lonicera japonica in wound-induced rats. Methods Rats were divided into 5 groups; normal (Nor), control (Veh), positive comparison (PC), Lonicera japonica 100 mg/kg (LL), Lonicera japonica 200 mg/kg (LH), each n=8. Total polyphenol and flavonoid were quantified. 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3 ethylbenzothiazoline-6-sulfonic acid) (ABTS) free radical scavenging activation were measured. Reactive oxygen species (ROS) was measured in serum. Antioxidant factors and inflammatory factors were measured in skin tissue, and also hydroxyproline content. Skin tissue was analyzed by Hematoxylin & Eosin and Masson's trichrome staining method. Results Total polyphenol and flavonoid were 32.86±0.14 mg/g and 67.17±0.57 mg/g. The IC₅₀ values of DPPH and ABTS free radical scavenging activation were 26.69±1.50 ㎍/mL and 49.33±4.52 ㎍/mL. ROS was significantly lower in LL and LH groups. Nuclear factor-erythroid 2-related factor 2 (Nrf2) was significantly higher in LH group and higher in LL group but not significant. Superoxide dismutase 1 (SOD-1), catalase, and heme oxygenase 1 (HO-1) were significantly higher in LL and LH groups. Nuclear factor kappa-B p65 (NF-κBp65), phosphorylated iκBα (p-iκBα), cyclooxygenase 2 (COX-2), and tumor necrosis factor alpha (TNF-α) were significantly lower in LL and LH groups. Hydroxyproline was significantly higher in LL and LH groups. The histopathologic analysis showed that skin tissue had recovered further more in LL and LH groups than in Veh group. Conclusions These results suggest that Lonicera japonica has the anti-oxidant, anti-inflammatory and healing effects in wound-induced rats.