• The Korean Journal of Physiology and Pharmacology
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• v.26 no.1
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• pp.25-36
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• 2022

To identify the effect and mechanism of carbon monoxide (CO) on delayed rectifier K⁺ currents (I_K) of human cardiac fibroblasts (HCFs), we used the wholecell mode patch-clamp technique. Application of CO delivered by carbon monoxidereleasing molecule-3 (CORM3) increased the amplitude of outward K⁺ currents, and diphenyl phosphine oxide-1 (a specific I_K blocker) inhibited the currents. CORM3-induced augmentation was blocked by pretreatment with nitric oxide synthase blockers (L-NG-monomethyl arginine citrate and L-NG-nitro arginine methyl ester). Pretreatment with KT5823 (a protein kinas G blocker), 1H-[1,-2,-4] oxadiazolo-[4,-3-a] quinoxalin-1-on (ODQ, a soluble guanylate cyclase blocker), KT5720 (a protein kinase A blocker), and SQ22536 (an adenylate cyclase blocker) blocked the CORM3 stimulating effect on I_K. In addition, pretreatment with SB239063 (a p38 mitogen-activated protein kinase [MAPK] blocker) and PD98059 (a p44/42 MAPK blocker) also blocked the CORM3's effect on the currents. When testing the involvement of S-nitrosylation, pretreatment of N-ethylmaleimide (a thiol-alkylating reagent) blocked CO-induced I_K activation and DL-dithiothreitol (a reducing agent) reversed this effect. Pretreatment with 5,10,15,20-tetrakis(1-methylpyridinium-4-yl)-21H,23H porphyrin manganese (III) pentachloride and manganese (III) tetrakis (4-benzoic acid) porphyrin chloride (superoxide dismutase mimetics), diphenyleneiodonium chloride (an NADPH oxidase blocker), or allopurinol (a xanthine oxidase blocker) also inhibited CO-induced I_K activation. These results suggest that CO enhances I_K in HCFs through the nitric oxide, phosphorylation by protein kinase G, protein kinase A, and MAPK, S-nitrosylation and reduction/oxidation (redox) signaling pathways.

• Journal of Applied Biological Chemistry
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• v.59 no.1
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• pp.25-29
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• 2016

Hepatitis B virus infection can cause hepatic fibrosis leading to cirrhosis and hepatocellular carcinoma. However the mechanism remains poorly understood. In this study, we found that Hepatitis B virus X-protein (HBx) increases vimentin, fibronectin, slug, snail and NOX4 expression. Because NOX4-mediated reactive oxygen species can increase slug and snail, which can induce fibrosis, HBx may be a key regulator of hepatic fibrosis development via NOX4 induction.

• The Korean Journal of Pharmacology
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• v.26 no.1
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• pp.55-66
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• 1990

In both resting and opsonized zymosan activated neutrophils, ATP stimulated superoxide generation, whereas adenosine inhibited it slightly. The superoxide generation in activated neutrophils to ATP was greater than that of resting neutrophils. In $Ca^{++}$ free medium, inhibitory effect of adenosine on superoxide generation was detectable, whereas ATP did not have any effect. The stimulatory effect of ATP on superoxide generation was inhibited by adenosine in a dose dependent manner. Neither ATP nor adenosine had any effect on NADPH oxidase acitivity. Effects of ATP or adenosine on superoxide generation were more prominent than that by other triphosphate nucleotides or nucleosides. ATP and ADP further stimulated $Ca^{++}$ uptake and increased cytosolic free $Ca^{++}$ level in neutrophils activated by opsonized zymosan, but adenosine inhibited a $Ca^{++}$ mobilization. Verapamil effectively and tetrodotoxin slightly inhibited an increase of cytosolic free $Ca^{++}$ level induced by ATP. Inhibitory effect of either verapamil or tetrodotoxin on superoxide generation in the ATP plus opsonized zymosan-activated neutrophils was greater than in the cells activated by opsonized zymosan alone. Tetraethylammonium chloride had no apparent effect on superoxide generation. CCCP, 2,4-dinitrophenol, diphenylhydantoin and procaine all inhibited superoxide generation in neutrophils activated by opsonized zymosan. Among these, CCCP only inhibited a stimulatory effect of ATP. ATP further stimulated a loss of sulfhydryl groups in activated neutrophils, whereas adenosine had no effect on it. These results suggest that functional responses of neutrophils may be regulated at least partly by purines. ATP and adenosine may further after functional responses of activated neutrophils through their effect on $Ca^{++}$ uptake, membrane phosphorylation and oxidation of soluble sulfhydryl groups.

• Journal of Life Science
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• v.19 no.6
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• pp.818-824
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• 2009

The mechanisms responsible for ischemia/reperfusion (I/R) injury have direct or indirect relevance to clinical lung injury after severe shock, cardiopulmonary bypass, and transplantation. This study investigated the effects of aspirin on intestinal I/R-induced acute lung injury (ALI) in rats. Lipopolysaccharide (LPS) induced cyclooxygenase-2 (COX-2) expression in A549 and RAW264.7 cells. RAW264.7 macrophages had shown greater expression of COX-2 than A549 cells. In addition, the NADPH oxidase inhibitor apocynin and p38 MAPK inhibitor SB203580 attenuated LPS-stimulated COX-2 expression. To induce ALI, intestinal ischemia was performed for 60 min prior to the 4 hr reperfusion by clamping the superior mesenteric artery in Sprague-Dawley rats. In order to test and compare the effect of non-specific COX inhibitor aspirin with the effect of mepacrine, a well known phospholipase$A_{2}$ inhibitor, rats were divided into 4 groups: Sham, I/R, Mepa+I/R (mepacrine, 60 mg/kg, i.p.), ASA+I/R (aspirin, 10 mg/kg, i.p.). In the present investigation, myeloperoxidase activities in the lung and intestinal tissues were increased by I/R. These changes were reduced by single pretreatment of mepacrine (60 mg/kg, i.p.) or aspirin (10 mg/kg, i.p.) 30 min before I/R. Structural studies demonstrated that the tissue injuries in the lung and intestine after I/R were also attenuated by the pretreatment of mepacrine or aspirin. These results suggest that I/R-induced ALI is mediated, in part, by the activation of COX. In addition, pretreatment of aspirin might be helpful for the prevention of ALI in ARDS-prone patients. In addition, the p38 MAPK inhibitor and apocynin also might be helpful to ALI through the inhibition of COX-2 expression.

• Journal of Nutrition and Health
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• v.54 no.4
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• pp.412-421
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• 2021

Purpose: Thioacetamide (TAA) produces reactive oxygen species (ROS) in the liver, and the generated ROS induces liver injury through inflammatory reactions. The current study was undertaken to investigate the hepatoprotective effect of Artemisiae Capillaris Herba water extract (AC), imparted via its antioxidant activity, in an animal model of TAA-induced liver injury. Methods: Animal experiments were conducted in 5 groups: normal, control (TAA 200 mg/kg), SM (TAA 200 mg/kg + silymarin 100 mg/kg), ACL (TAA 200 mg/kg + AC 100 mg/kg), ACH (TAA 200 mg/kg + AC 200mg/kg). TAA (intraperitoneal) and treatment compounds (per oral) were administered for 3 days. Serum levels of ammonia concentration and myeloperoxidase (MPO) activity were subsequently measured. Liver tissues were subjected to western blot analysis for measuring the oxidative stress (NADPH oxidase), anti-oxidative activity (Nrf2, heme oxygenase-1 [HO-1], superoxide dismutase [SOD], catalase, and GPx-1/2), tissue inhibitors of metalloproteinase (TIMP)-1, and matrix metalloproteinases (MMPs) protein expressions. Results: Serum ammonia levels and MPO activity were significantly increased in the TAA-induced control group, whereas groups administered AC treatment showed markedly reduced levels. Western blot analysis revealed significantly increased NOX2 and p22^phox expressions, (oxidative stress-related factors) in the TAA-induced control group. These levels were determined to be significantly decreased after AC exposure. Moreover, antioxidant-related factors including Nrf2, HO-1, SOD, catalase, and GPx-1/2 were significantly decreased in the control group and increased in the AC treated groups. In addition, MMP expressions were significantly suppressed in the AC treatment group due to increased levels of TIMP-1. Conclusion: Taken together, these data indicate that exposure to AC reduces the oxidative stress by inhibiting the expression of NADPH oxidase (NOX2 and p22^phox) through the Nrf2 signaling pathway. We therefore propose the potential of AC for the prevention and treatment of TAA-induced liver injury.

• Journal of Nutrition and Health
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• v.57 no.4
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• pp.403-417
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• 2024

Purpose: This study investigated the anti-inflammatory effects of Evodiae Fructus (EF) on hydrochloric acid (HCl)/ethanol-induced gastritis, focusing on its impact on oxidative stress by analyzing inflammatory cytokines and inflammation-related factors. The total polyphenol and flavonoid contents were determined through in vitro experiments, while the radical scavenging activity was confirmed using 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) assays. Methods: In vivo experiments were conducted on rats divided into 5 groups (n = 7/in each group): normal group (Normal), 150 mM HCl/60% ethanol-induced gastritis group (Control), 150 mM HCl/60% ethanol-induced gastritis group administered 10 mg/kg sucralfate (SC), 150 mM HCl/60% ethanol-induced gastritis group administered EF at the doses of 100 mg/kg or 200 mg/kg (EF100 or EF200). The mice were pretreated with the extract (EF) or drug (SC), and after 1 hour, 150 mM HCl/60% ethanol (v/v) mixture was administered orally. Reactive oxygen species (ROS) levels, peroxynitrite (ONOO^-), and pro-inflammatory cytokines including tumor necrosis factor-α and interleukin-1 beta were assessed in serum. Additionally, western blotting of the gastric tissues confirmed the expression of inflammation-related proteins. Results: EF alleviated the gastric mucosal damage caused by 150 mM HCl/60% ethanol. The assessment of oxidative stress in the serum showed that EF significantly reduced ROS and ONOO^- levels and significantly decreased the levels of pro-inflammatory cytokines. Western blot analysis revealed that EF reduced ROS-generating nicotinamide adenine dinucleotide phosphate oxidase subunits, including gp91^phox, p22^phox, and p47^phox. Additionally, EF mitigated the inflammation by inhibiting the mitogen-activated protein kinase signaling pathway. Conclusion: These results indicate that EF is a potential herbal medicine candidate for the treatment of oxidative stress-induced gastritis.

• BMB Reports
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• v.49 no.4
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• pp.232-237
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• 2016

Mulberry tree twigs (Ramulus mori) contain large amounts of oxyresveratrols and have traditionally been used as herbal medicines because of their anti-inflammatory properties. However, the signaling mechanism by which R. mori exerts its anti-inflammatory action remains to be elucidated. In this study, we observed that R. mori ethanol extracts (RME) exerted an inhibitory effect on the lipopolysaccharide (LPS)-induced production of the pro-inflammatory cytokine interleukin-6 (IL-6) in Raw264.7 macrophage cells. Additionally, RME inhibited IL-6 production by blocking the leukotriene B₄ receptor-2 (BLT2)-dependent-NADPH oxidase 1 (NOX1)-reactive oxygen species (ROS) cascade, leading to anti-inflammatory activity. Finally, RME suppressed the production of the BLT2 ligands LTB4 and 12(S)-HETE by inhibiting the p38 kinase-cytosolic phospholipase A₂-5-/12-lipoxygenase cascade in LPS-stimulated Raw264.7 cells. Overall, our results suggest that RME inhibits the 'BLT2 ligand-BLT2'-linked autocrine inflammatory axis, and that this BLT2-linked cascade is one of the targets of the anti-inflammatory action of R. mori.

• The Journal of Internal Korean Medicine
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• v.35 no.1
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• pp.92-105
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• 2014

Objectives : Cisplatin is a widely used cancer therapy drug. However, nephrotoxicity resulting in increased oxidative stress is a major side effect of cisplatin chemotherapy, thereby limiting its chemotherapeutic use. Lycium chinense Miller (LCM) has been used as a traditional herbal medicine in various febrile and inflammatory diseases such as night sweat, cough, nosebleed, bronchitis, pulmonary tuberculosis, etc. In this study we investigated the protective and antioxidative potential of LCM against cisplatin-induced nephrotoxicity in rats. Methods : Twenty-four 8-week-old male Wistar rats were divided into four groups: normal untreated; cisplatin treatment only; LCM 10 mg/kg plus cisplatin treatment; and LCM 30 mg/kg plus cisplatin treatment. Twenty-four hours after the last cisplatin injection, all the rats were sacrificed, and serological changes were evaluated. The levels of NF-${\kappa}B$ activity and NOX-4, $p47^{phox}$, $p22^{phox}$, COX-2, iNOS, SOD, catalase expressions were analyzed in Western blot analysis. Results : Cisplatin injection caused an increase in the BUN level, which is a reliable indicator of renal toxicity. The levels of BUN, renal ROS, and renal TBARS were significantly reduced in the LCM groups compared with the cisplatin-only groups. The levels of $p47^{phox}$ and $p22^{phox}$, which are NADPH oxidase subunits, were increased in the cisplatin-only groups, whereas they were decreased in the LCM groups. The levels of renal NF-${\kappa}B$ activity and COX-2, iNOS expressions were increased significantly in the cisplatin-only groups compared with the normal groups, whereas they were decreased in the LCM groups. Compared with the cisplatin-only groups, renal GSH and GSH/GSSG increased in the LCM groups. Also, the administration of LCM increased levels of SOD and catalase as compared with the cisplatin-only groups. Conclusions : These results suggest that LCM protects cisplatin-induced nephrotoxicity via a mechanism that may involves the inhibition of oxidative stress by the activation of antioxidants.

• The Korean Journal of Physiology and Pharmacology
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• v.26 no.4
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• pp.229-238
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• 2022

Severe bacterial infections are frequently accompanied by depressed neutrophil functions. Thus, agents that increase the microbicidal activity of neutrophils could add to a direct antimicrobial therapy. Lysophosphatidylcholine augments neutrophil bactericidal activity via the glycine (Gly)/glycine receptor (GlyR) α2/TRPM2/p38 mitogen-activated protein kinase (MAPK) pathway. However, the direct effect of glycine on neutrophil bactericidal activity was not reported. In this study, the effect of glycine on neutrophil bactericidal activity was examined. Glycine augmented bactericidal activity of human neutrophils (EC₅₀ = 238 μM) in a strychnine (a GlyR antagonist)-sensitive manner. Glycine augmented bacterial clearance in mice, which was also blocked by strychnine (0.4 mg/kg, s.c.). Glycine enhanced NADPH oxidase-mediated reactive oxygen species (ROS) production and TRPM2-mediated [Ca²⁺]_i increase in neutrophils that had taken up E. coli. Glycine augmented Lucifer yellow uptake (fluid-phase pinocytosis) and azurophil granule-phagosome fusion in neutrophils that had taken up E. coli in an SB203580 (a p38 MAPK inhibitor)-sensitive manner. These findings indicate that glycine augments neutrophil microbicidal activity by enhancing azurophil granule-phagosome fusion via the GlyRα2/ROS/calcium/p38 MAPK pathway. We suggest that glycine could be a useful agent for increasing neutrophil bacterial clearance.

• Journal of the Korean Society of Food Science and Nutrition
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• v.27 no.2
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• pp.319-325
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• 1998

The purpose of this study was to investigate the effect of green tea catechin on microsomal mixed function oxidase(MFO) system of kidney and brain in streptozotocin(STZ) induced diabetic rats. Sprague-Dawley male rats weighing 140$\pm$10g were randomly assigned to one control and three STZ-diabetic groups. Diabetic groups wer classified to DM-0C(catechin 0%/kg diet), DM-0.5C (catechin 0.5%/kg diet), and DM-1.0C(catechin 1%/kg diet) according to the level of catechin supplementation. Diabetes were experimentally induced by intravenous administration of 55mg/kg body weight of STZ in citrate buffer(pH 4.3) after 4 weeks feeding of three experimental diets. Animals were sacrificed at the sixth day of diabetic state. The contents of cytochrome P450 in kidney were increased by 77, 42, 49% in DM-0C, DM-0.5C and DM-1.0C groups, respectively, than normal group. The contents of cytochrome P450 in brain were increased by 43% in DM-0C group than normal group, but those of DM-0.5C and DM-1.0C groups were similar to that of normal group. The contents of cytochrome b5 in kidney were increased by 78, 38, 49% in DM-0C, DM-0.5C and DM-1.0C groups, respectively, than normal group. Meanwhile, the contents of cytochrome b5 in brain were not significantly different among all groups. The activities of NADPH-cytochrome P450 reductase in kidney of DM-group were increased by 27% than normal group, but those of DM-0.5C and DM-1.0C groups were 13 and 15% lower than that of DM-0C group. The activities in brain were also increased by 31% in DM-0C group, but those of DM-0.5C and DM-1.0C groups were similar to than of normal group. Levels of TBARS (thiobarbituric acid reactive substance) in kidney were increased by 147, 60 and 59% in DM-0C, DM-0.5C, and DM-1.0C groups, respectively, compared with normal group, but those of DM-0.5C and DM-1.0C groups were 36, 35% lower than that of DM-0C group. Meanwhile, the levels of TBARS in brain were not significantly different among four groups. These results indicate that dietary catechins in green tea play a powerful antioxidant role in reducing the lipid peroxidation enhanced by activation of MFO system in STZ-induced diabetes.