• The Korean Journal of Physiology and Pharmacology
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• 제25권2호
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• pp.159-166
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• 2021

Nicotinamide adenine dinucleotide phosphate oxidases (NOXs) are the major enzymatic source of reactive oxygen species (ROS). NOX2 and NOX4 are expressed in the heart but its role in hypoxia-induced atrial natriuretic peptide (ANP) secretion is unclear. This study investigated the effect of NOX on ANP secretion induced by hypoxia in isolated beating rat atria. The results showed that hypoxia significantly upregulated NOX4 but not NOX2 expression, which was completely abolished by endothelin-1 (ET-1) type A and B receptor antagonists BQ123 (0.3 μM) and BQ788 (0.3 μM). ET-1-upregulated NOX4 expression was also blocked by antagonists of secreted phospholipase A2 (sPLA2; varespladib, 5.0 μM) and cytosolic PLA2 (cPLA2; CAY10650, 120.0 nM), and ET-1-induced cPLA2 expression was inhibited by varespladib under normoxia. Moreover, hypoxia-increased ANP secretion was evidently attenuated by the NOX4 antagonist GLX351322 (35.0 μM) and inhibitor of ROS N-Acetyl-D-cysteine (NAC, 15.0 mM), and hypoxia-increased production of ROS was blocked by GLX351322. In addition, hypoxia markedly upregulated Src expression, which was blocked by ET receptors, NOX4, and ROS antagonists. ET-1-increased Src expression was also inhibited by NAC under normoxia. Furthermore, hypoxia-activated extracellular signal-regulated kinase 1/2 (ERK1/2) and protein kinase B (Akt) were completely abolished by Src inhibitor 1 (1.0 μM), and hypoxia-increased GATA4 was inhibited by the ERK1/2 and Akt antagonists PD98059 (10.0 μM) and LY294002 (10.0 μM), respectively. However, hypoxia-induced ANP secretion was substantially inhibited by Src inhibitor. These results indicate that NOX4/Src modulated by ET-1 regulates ANP secretion by activating ERK1/2 and Akt/GATA4 signaling in isolated beating rat hypoxic atria.

• Molecules and Cells
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• 제37권12호
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• pp.907-911
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• 2014

The function of reactive oxygen species (ROS) as second messengers in cell differentiation has been demonstrated only for a limited number of cell types. Here, we used a well-established protocol for BMP2-induced neuronal differentiation of neural crest stem cells (NCSCs) to examine the function of BMP2-induced ROS during the process. We first show that BMP2 indeed induces ROS generation in NCSCs and that blocking ROS generation by pretreatment of cells with diphenyleneiodonium (DPI) as NADPH oxidase (Nox) inhibitor inhibits neuronal differentiation. Among the ROS-generating Nox isozymes, only Nox4 was expressed at a detectable level in NCSCs. Nox4 appears to be critical for survival of NCSCs at least in vitro as down-regulation by RNA interference led to apoptotic response from NCSCs. Interestingly, development of neural crest-derived peripheral neural structures in Nox4-/- mouse appears to be grossly normal, although Nox4-/- embryos were born at a sub-Mendelian ratio and showed delayed over-all development. Specifically, cranial and dorsal root ganglia, derived from NCSCs, were clearly present in Nox4-/- embryo at embryonic days (E) 9.5 and 10.5. These results suggest that Nox4-mediated ROS generation likely plays important role in fate determination and differentiation of NCSCs, but other Nox isozymes play redundant function during embryogenesis.

• 동의생리병리학회지
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• 제18권6호
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• pp.1666-1685
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• 2004

This present study was designed to screen medicinal plants for the treatment of brain diseases such as Parkinson's disease or aging. We tested the effects of the water extracts from 38 species medicinal plants on antioxidant capacity, monoamine oxidase B (MAO-B) inhibitory activity, acetylcholinesterase (AChE) inhibition and antiperoxidation activity in vitro. The water extracts from 38 species were tested on their antioxidant activity using radical scavenging effects against ABTS+. The water extract of C. sappan was showed the highest antioxidant capacity, the antioxidant activity at 1 Jig of herbal extract being 0.38mM TE. Lipid peroxidation in brain homogenates induced by NADPH and ADP-Fe/sup 2+/ was strong inhibited by C. sappan and R. palmatum extracts. Among the 38 medicinal plants investigated, R. palmatum showed significant biological activity (antioxidant capacity, MAO-B inhibiory activity, and AChE inhibitory activity). The protective efficacy of R. palmatum water extract on 1-methyl-4­phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced parkinsonism and its possible mechanism were studied in C57BL/6 mice. Treatment of R. palmatum water extract protected biomacromolecules such as lipids from oxidative damage induced by MPTP. The content of MDA in brain tissue was decreased significantly by R. palmatum extract. These results suggest that R. palmatum water extract plays on effective role in attenuating MPTP-induced neurotoxicity in mice. This protective effect of R. palmatum might be estimated the result from the inhibitory activity on monoamine oxidase B and the enhancement of antioxidant activity.

• 한국식품영양과학회지
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• 제35권8호
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• pp.979-984
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• 2006

과잉 철은 폐경기 여성 및 핀란드 남성에서의 심혈관계질환 증가와 예방통계학적으로 밀접한 관련이 있다고 보고되고 있다. 허혈성 심장질환, 뇌 심혈관계 질환, 암 및 노화의 원인으로 산화적 스트레스가 자유기 반응을 자극하고 지질 과산화 반응 등을 연쇄적으로 촉진시키는데 철이 위험 인자로 인식되고 있다. 그러나 뇌심혈관계 질환 유발의 중요인자인 혈소판 활성화와 관련하여 철로 인한 산화적 스트레스와 항산화작용의 연구는 부족하다고 한다. 산화적 스트레스에서 철 및 과산화수소의 자유기 형성과 관련하여 토끼 혈액에서 분리한 세정 혈소판을 사용하여 연구하였다. 산화적 스트레스를 통해 혈소판 응집을 유도하고 이에 미치는 영향을 연구한 결과에서 $H_2O_2$ 단독 투여시 혈소판 응집작용은 나타나지 않았다. $FeSO_4$ 단독 투여시 농도 의존적으로 혈소판 응집작용이 증가하여 나타내지만, $H_2O_2$ 존재 하에 $FeSO_4$ 투여시 농도 의존적으로 혈소판 응집작용이 증가되어 나타났다. 혈소판 응집을 유도하는 collagen 최적의 농도$(2\;{\mu}g/mL)$보다 낮은 1/10 농도$(2\;{\mu}g/mL)$)에서 $H_2O_2$ 및 $FeSO_4$의 영향은 농도 의존적으로 혈소판 응집작용이 증가되었다. 철 단독 투여시보다 과산화수소와 함께 투여시 농도 의존적으로 혈소판 활성화가 증대되었고 이러한 혈소판 활성화는 NAD/NADP, catalase, glutathione, mannitol, tiron 등에 의해 농도 의존적으로 억제되었고, NADH/NADPH, SOD, aspirin 등에 의해서는 영향이 없었다. 그러므로, 이러한 NAD(H)/NADP(H) cofactor는 혈소판 응집작용을 일으키는 radical을 직접 억제하기보다 radical 생성에 관련하는 것으로 사료된다. 이상의 결과에서 과잉철은 혈소판 활성화에 직접적으로 관여하고 $H_2O_2$ 존재하에 2가 철을 촉매로 하여 Fenton 반응으로 생성된 OH. 자유기가 혈소판 활성화에 중요한 역할을 한다. 그러나 혈소판에서 자유기가 arachidonic acid 대사의 활성화와 인산화 단백질로 인한 세포내 정보전달에 관한 연구가 더 이루어져야 한다고 사료된다.

• 한국식품영양과학회지
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• 제25권6호
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• pp.969-975
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• 1996

본 연구는 streptozotocin 유발 당뇨쥐의 간과 폐조직에서의 MFO계 활성 변화 및 지질과산화에 미치는 vitamin E의 영향을 관찰하고자 본 실험을 행하였다. 체중 140g 정도의 Sparague-Dawley종 숫컷을 대조군(40mg vitamin E/kg diet group)과 당뇨유발 실험군으로 나눈 후 실험군은 다시 식이내 vitamin E 공급수준에 따라 DM-0E군(0mg vitamin E/kg diet group), DM-40E군(40mg vitamin E/kg diet group), DM-400E군(400mg vitamin E/kg diet group) 나누어 4주간 사육한 후 대조군을 제외한 실험군을 STZ로 당뇨를 유발시켰다. 당뇨유발 6일 후 쥐를 희생시켜 간조직 및 폐조직 microsome 중의 cytochrome P$_{450}$ 및 cytochrome b$_{5}$함량과 NADPH-cytochrome P$_{450}$ reductase 활성도를 측정하고 아울러 microsome내의 지질과산화물을 측정하여 다음과 같은 결과를 얻었다. 1. 간조직 중의 cytochrome P$_{450}$ 함량은 대조군에 비해 DM-0E군, DM-40E 군 및 DM-400E군이 각각 162%, 150%, 56% 증가하였으며 또 DM-400E군은 DM-0E, DM-40E군에 비해 낮았다. 폐조직 중의 cytochrome P$_{450}$ 함량도 간조직에 비해 함량은 낮았지만 경향은 비슷하였다. 2. 간조직 중의 cytochrome b$_{5}$은 대조군에 비해 DM-0E군, DM-40E군은 각각 70%, 53%씩 증가하였으나 DM-400E군은 대조군 수준이었다. 폐조직 중의 cytochrome b$_{5}$함량도 대조군에 비 해 DM-0E, DM-40E 및 DM-400E군에서 각각 149%, 116%, 50%씩 증가하였다. 3. 간조직에서의 cytochrome P$_{450}$ reductase 활성은 대조군에 비해 DM-0E군은 58%, DM-40E군에서는 47% 증가하였으며 DM-400E는 대조군 수준이었다. 폐조직에서도 간조직에서와 비슷한 경향이었다. 4. 간조직의 지질과산화물가는 대조군에 비해 DM-0E군, DM-40E군, DM-400E군이 각각 143%, 95%, 31%씩 높았다. 또 DM-400E군은 DM-0E, DM-40E군에 비해 약 50% 정도 낮았다. 폐조직에서도 경향은 비슷했는데 간조직 보다 과산화적 손상이 다소 높았다. 이상의 결과에서 STZ 유발 당뇨쥐에서는 MFO system의 활성이 증가되고 지질과산화가 촉진되었다. 그러나 식이 중의 vitamin E의 적절한 공급은 MFO system 활성이 저하되고 과산화적 손상이 현저하게 완화됨을 볼 수 있었다.

• Journal of Periodontal and Implant Science
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• 제47권2호
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• pp.66-76
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• 2017

Purpose: Oral wound healing requires gingival fibroblasts to respond to local growth factors. Epigenetic silencing through DNA methylation can potentially decrease the responsiveness of gingival fibroblasts to local growth factors. In this study, our aim was to determine whether the inhibition of DNA methylation sensitized gingival fibroblasts to transforming growth factor-${\beta}1$ (TGF-${\beta}1$). Methods: Gingival fibroblasts were exposed to 5-aza-2'-deoxycytidine (5-aza), a clinically approved demethylating agent, before stimulation with TGF-${\beta}1$. Gene expression changes were evaluated using quantitative polymerase chain reaction (PCR) analysis. DNA methylation was detected by methylation-sensitive restriction enzymes and PCR amplification. Results: We found that 5-aza enhanced TGF-${\beta}1$-induced interleukin-11 (IL11) expression in gingival fibroblasts 2.37-fold (P=0.008). 5-aza had no significant effects on the expression of proteoglycan 4 (PRG4) and NADPH oxidase 4 (NOX4). Consistent with this, 5-aza caused demethylation of the IL11 gene commonly next to a guanosine (CpG) island in gingival fibroblasts. The TGF-${\beta}$ type I receptor kinase inhibitor SB431542 impeded the changes in IL11 expression, indicating that the effects of 5-aza require TGF-${\beta}$ signaling. 5-aza moderately increased the expression of TGF-${\beta}$ type II receptor (1.40-fold; P=0.009), possibly enhancing the responsiveness of fibroblasts to TGF-${\beta}1$. As part of the feedback response, 5-aza increased the expression of the DNA methyltransferases 1 (DNMT1) (P=0.005) and DNMT3B (P=0.002), which are enzymes responsible for gene methylation. Conclusions: These in vitro data suggest that the inhibition of DNA methylation by 5-aza supports TGF-${\beta}$-induced IL11 expression in gingival fibroblasts.

• The Korean Journal of Physiology and Pharmacology
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• 제21권6호
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• pp.687-694
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• 2017

Plumbagin, a hydroxy 1,4-naphthoquinone compound from plant metabolites, exhibits anticancer, antibacterial, and antifungal activities via modulating various signaling molecules. However, its effects on vascular functions are rarely studied except in pulmonary and coronary arteries where NADPH oxidase (NOX) inhibition was suggested as a mechanism. Here we investigate the effects of plumbagin on the contractility of skeletal artery (deep femoral artery, DFA), mesenteric artery (MA) and renal artery (RA) in rats. Although plumbagin alone had no effect on the isometric tone of DFA, $1{\mu}M$ phenylephrine (PhE)-induced partial contraction was largely augmented by plumbagin (${\Delta}T_{Plum}$, 125% of 80 mM KCl-induced contraction at $1{\mu}M$). With relatively higher concentrations (>$5{\mu}M$), plumbagin induced a transient contraction followed by tonic relaxation of DFA. Similar biphasic augmentation of the PhE-induced contraction was observed in MA and RA. VAS2870 and GKT137831, specific NOX4 inhibitors, neither mimicked nor inhibited ${\Delta}T_{Plum}$ in DFA. Also, pretreatment with tiron or catalase did not affect ${\Delta}T_{Plum}$ of DFA. Under the inhibition of PhE-contraction with L-type $Ca^{2+}$ channel blocker (nifedipine, $1{\mu}M$), plumbagin still induced tonic contraction, suggesting $Ca^{2+}$-sensitization mechanism of smooth muscle. Although ${\Delta}T_{Plum}$ was consistently observed under pretreatment with Rho A-kinase inhibitor (Y27632, $1{\mu}M$), a PKC inhibitor (GF 109203X, $10{\mu}M$) largely suppressed ${\Delta}T_{Plum}$. Taken together, it is suggested that plumbagin facilitates the PKC activation in the presence of vasoactive agonists in skeletal arteries. The biphasic contractile effects on the systemic arteries should be considered in the pharmacological studies of plumbagin and 1,4-naphthoquinones.

• 한방재활의학과학회지
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• 제32권1호
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• pp.1-19
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• 2022

Objectives This study aims to evaluate the wound healing effects of oral administered hot water extracts of Korean black ginseng (KBG). Methods 40 C57BL/6 mice were divided into five groups; normal, control, vitamin E 200 mg/kg, KBG 100 mg/kg, KBG 200 mg/kg, each n=8. Skin wounds were made in the back of all mice except normal group using biopsy punches. Wounds were observed on days 7 and 14 after injury. The anti-oxidant and inflammatory protein levels were evaluated using western blotting. Skin tissue was analyzed by hematoxylin & eosin and Masson's trichrome staining method. Results KBG significantly accelerated reducing wound area. KBG significantly decreased myeloperoxidase activity. KBG significantly decreased oxidative stress factors such as NADPH oxidase-4 and p22^phox and increased antioxidant enzymes including nuclear factor erythroid 2-related factor2, kelch-like ECH-associated protein-1, heme oxygenase-1, superoxide dismutase, catalase and glutathione peroxidase-1/2. Moreover, KBG significantly decreased inflammation factors including nuclear factor-κB, phosphorylated inhibitor of κBα, cyclooxygenase-2, inducible nitric oxide synthase, tumor necrosis factor-α and interleukin (IL)-6 and increased anti-inflammation cytokine such as IL-4 and IL-10. In addition, KBG significantly increased tight junction proteins including claudin-1, claudin-3, claudin-4. In histopathologic, KBG made the epithelium thin and uniform, and accelerated the remodeling of collagen. Conclusions The results suggest that KBG has healing effects on skin wound in mice by anti-inflammatory and antioxidant activity.

• International Journal of Oral Biology
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• 제39권4호
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• pp.207-213
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• 2014

Antimicrobial actions of reactive oxygen/nitrogen species (ROS/RNS) derived from products of NADPH oxidase and inducible nitric oxide (NO) synthase in host phagocytes inactivate various bacterial macromolecules. To cope with these cytotoxic radicals, pathogenic bacteria have evolved to conserve systems necessary for detoxifying ROS/RNS and repairing damages caused by their actions. In response to these stresses, bacteria also induce expression of molecular chaperones to aid in ameliorating protein misfolding. In this study, we explored the function of a newly identified chaperone Spy, that is localized exclusively in the periplasm when bacteria exposed to conditions causing spheroplast formation, in the resistance of Salmonella Typhimurium to ROS/RNS. A spy deletion mutant was constructed in S. Typhimurium by a PCR-mediated method of one-step gene inactivation with ${\lambda}$ Red recombinase, and subjected to ROS/RNS stresses. The spy mutant Salmonella showed a modest decrease in growth rate in NO-producing cultures, and no detectable difference of growth rate in $H_2O_2$ containing cultures, compared with that of wild type Salmonella. Quantitative RT-PCR analysis showed that spy mRNA levels were similar regardless of both stresses, but were increased considerably in Salmonella mutants lacking the flavohemoglobin Hmp, which are incapable of NO detoxification, and lacking an alternative sigma factor RpoS, conferring hypersusceptibility to $H_2O_2$. Results demonstrate that Spy expression can be induced under extreme conditions of both stresses, and suggest that the protein may have supportive roles in maintaining proteostasis in the periplasm where various chaperones may act in concert with Spy, thereby protecting bacteria against toxicities of ROS/RNS.

• Clinical and Experimental Pediatrics
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• 제61권9호
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• pp.271-278
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• 2018

Purpose: Abnormal potassium channels expression affects vessel function, including vascular tone and proliferation rate. Diverse potassium channels, including voltage-gated potassium (Kv) channels, are involved in pathological changes of pulmonary arterial hypertension (PAH). Since the role of the Kv1.7 channel in PAH has not been previously studied, we investigated whether Kv1.7 channel expression changes in the lung tissue of a monocrotaline (MCT)-induced PAH rat model and whether this change is influenced by the endothelin (ET)-1 and reactive oxygen species (ROS) pathways. Methods: Rats were separated into 2 groups: the control (C) group and the MCT (M) group (60 mg/kg MCT). A hemodynamic study was performed by catheterization into the external jugular vein to estimate the right ventricular pressure (RVP), and pathological changes in the lung tissue were investigated. Changes in protein and mRNA levels were confirmed by western blot and polymerase chain reaction analysis, respectively. Results: MCT caused increased RVP, medial wall thickening of the pulmonary arterioles, and increased expression level of ET-1, ET receptor A, and NADPH oxidase (NOX) 4 proteins. Decreased Kv1.7 channel expression was detected in the lung tissue. Inward-rectifier channel 6.1 expression in the lung tissue also increased. We confirmed that ET-1 increased NOX4 level and decreased glutathione peroxidase-1 level in pulmonary artery smooth muscle cells (PASMCs). ET-1 increased ROS level in PASMCs. Conclusion: Decreased Kv1.7 channel expression might be caused by the ET-1 and ROS pathways and contributes to MCT-induced PAH.