• Journal of Life Science
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• v.23 no.10
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• pp.1223-1229
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• 2013

Ursolic acid (UA) a bio-active ingredient found in a variety of fruits and vegetables, and it has potent antioxidant activity. However, the role of UA in mouse embryonic stem (ES) cells is poorly understood. This study investigated the functional role of UA in regulating the development of mouse ES cells under hypoxia. Hypoxia did not exert a significant effect on the undifferentiated state of mouse ES cells. However, it induced reactive oxygen species (ROS) generation and increased the level of lactate dehydrogenase (LDH) production at 48 h of hypoxic exposure. Conversely, oxidative stress induced by hypoxia was significantly inhibited by UA ($30{\mu}M$) pretreatment. Hypoxia significantly decreased cell survival and the level of [$^3H$] thymidine incorporation, both of which recovered following pretreatment of UA. In addition, UA decreased the apoptotic effect of hypoxia by attenuating caspase-3 cleavage or by recovering cellular inhibition of the apoptotic protein (cIAP)-2 and Bcl-2 expression. We further found that UA decreased senescence-associated beta-galactosidase activity. We suggest that UA is a natural antioxidant and one of the functional modulators of hypoxia-induced survival, apoptosis, proliferation, and aging in mouse ES cells.

• Journal of Life Science
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• v.23 no.3
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• pp.462-469
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• 2013

Withaferin A is a steroidal lactone purified from the Indian medicinal plant Withania somnifera. It exhibits a wide variety of activities, including anti-tumor, anti-inflammation, and immunomodulation properties. In this review, we focused on the anti-cancer effects of withaferin A. Withaferin A inhibits cell proliferation, metastasis, invasion, and angiogenesis in cancer cells. Furthermore, it sensitized irradiation, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-, and doxorubicin-mediated apoptosis. The results showed that multiple mechanisms were involved in withaferin A-mediated anti-cancer effects. First, withaferin A increased intracellular reactive oxygen species (ROS) production and induced ER stress- and mitochondria-mediated apoptosis. Second, withaferin A inhibited the signaling pathways (Jak/STAT, Akt, Notch, and c-Met), which are important in cell survival, proliferation, and metastasis. Third, it induced apoptosis and inhibited cancer cell migration through the up-regulation of prostate apoptosis protein-4 (Par-4). Finally, withaferin A up-regulated pro-apoptotic protein expression levels through the inhibition of proteasome activity. Our findings suggested that withaferin A is a potential, potent therapeutic agent.

• Journal of Life Science
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• v.23 no.4
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• pp.479-486
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• 2013

The DNA-binding protein from starved cells (DPS), originally identified as a DNA binding protein in Escherichia coli, is known to play an important role in DNA protection. The aim of this study was to evaluate the functional roles of DPS in E. coli against various kinds of external stresses by comparing the properties of wild-type E. coli cells and dps knockout mutant E. coli (${\Delta}dps$) cells. Under various stress conditions, we measured the cell growth of the wild-type E. coli and the dps knockout mutant E. coli (${\Delta}dps$) cells using a UV spectrophotometer. The growth rate of the cells was compared to investigate the functional roles of the DPS protein in E. coli. In comparison to the properties of the wild-type E. coli cells, the dps knockout mutant E. coli (${\Delta}dps$) cells showed highly sensitive phenotypes under various stress conditions, such as heat shock, acidic pH, nutrient deficiency, and different concentrations of reactive oxygen species (ROS), suggesting that DPS plays key roles in E. coli in combating diverse external stresses. The DPS DNA-binding protein in E. coli plays crucial roles in bacterial cell growth and in the protection of the cells from environmental stresses by tightly binding and preserving their DNA molecules.

• Molecules and Cells
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• v.20 no.3
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• pp.401-408
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• 2005

Reactive oxygen species (ROS) contribute to the development of various human diseases. Cu,Zn-superoxide dismutase (SOD) is one of the major means by which cells counteract the deleterious effects of ROS. SOD activity is dependent upon bound copper ions supplied by its partner metallochaperone protein, copper chaperone for SOD (CCS). In the present study, we investigated the protective effects of PEP-1-CCS against neuronal cell death and ischemic insults. When PEP-1-CCS was added to the culture medium of neuronal cells, it rapidly entered the cells and protected them against paraquat-induced cell death. Moreover, transduced PEP-1-CCS markedly increased endogenous SOD activity in the cells. Immunohistochemical analysis revealed that it prevented neuronal cell death in the hippocampus in response to transient forebrain ischemia. These results suggest that CCS is essential to activate SOD, and that transduction of PEP-1-CCS provides a potential strategy for therapeutic delivery in various human diseases including stroke related to SOD or ROS.

• Herbal Formula Science
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• v.23 no.2
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• pp.189-198
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• 2015

Objectives : Sparganii Rhizoma is frequently used in traditional herbal medicine for treatment of blood stasis, amenorrhea and functional dyspepsia and has been reported to exhibit anti-oxidant, anti-proliferation and anti-angiogenesis peoperties. In this study, we investigated the cytoprotective effect and underlying mechanism of Sparganii Rhizoma water extract (SRE) against oxidative stress-induced mitochondrial dysfunction and apoptosis in hepatocyte. Methods : To determine the effects of SRE on oxidative stress, we induced synergistic cytotoxicity by co-treatment of arachidonic acid (AA) and iron in the HepG2 cell, a human derived hepatocyte cell line. Results : Treatment of SRE increased relative cell viability and altered the expression levels of apoptosis-related proteins such as Bcl-xL, Bcl-2 and procaspase-3. And SRE also inhibited the mitochondrial dysfunction and excessive reactive oxygen species production induced by AA+iron. In addition, SRE activated of AMP-activated protein kinase (AMPK), a potential target for cytoprotection, by increasing the phosphorylation of AMPKα at Thr-172. Morever, SRE increased phosphorylation of acetyl-CoA carboxylase, a direct downstream target of AMPK. Conclusion : These results indicated that SRE has the ability to protect against oxidative stress-induced hepatocyte damage, which may be mediated with AMPK pathway.

• The Journal of Korean Medicine
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• v.30 no.1
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• pp.51-63
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• 2009

Objectives: Peroxynitrite ($ONOO^-$), superoxide anion radical (${\cdot}{O_2}^-$ and nitric oxide (NO) are cytotoxic because they can oxidize several cellular components such as proteins, lipids and DNA. They have been implicated in the aging processes, and age-related diseases such as Alzheimer's disease, rheumatoid arthritis, cancer, diabetes, obesity and atherosclerosis. The aim of this study was to investigate the $ONOO^-$, NO, ${\cdot}{O_2}^-$ scavenging and NF-${\kappa}B$ related anti-inflammatory activities of Sotosaja-hwan in ob/ob mice. Methods: Mice were grouped and treated for 5 weeks as follows. Both the normal lean (C57/BL6J black mice) and control obese (ob/ob mice) groups have received standard chow. The experimental groups were fed with a diet of chow supplemented with 30 and 90 mg Sotosaja-hwan per 1 kg of body weight for 14 days. For this study, the fluorescent probes, namely 2',7'-dichlorodihydrofluorescein diacetate (DCFDA), 4,5-diaminofluorescein (DAF-2) and dihydrorhodamine 123 (DHR 123) were used. Western blotting was performed using anti-phospho-$I{\kappa}B$-${\alpha}$, anti-IKK-${\alpha}$, anti-NF-${\kappa}B$ (p50, p65), anti-COX-2, anti-iNOS, anti-YCAM-1 and anti-MMP-9 antibodies, respectively. Results: Sotosaja-hwan inhibited the generation of $ONOO^-$, NO and ${\cdot}{O_2}^-$ in the lipopolysaccharide (LPS)-treated mouse kidney postmitochondrial fraction in vitro. The generation of $ONOO^-$, NO, ${\cdot}{O_2}^-$ and PGE2 were inhibited in the Sotosaja-hwan-administered ob/ob mice groups. The GSH/GSSG ratio was decreased in the ob/ob mice, whereas the ratio was improved in the Sotosaja-hwan-administered groups. Sotosaja-hwan inhibited the protein expression levels of phospho-$I{\kappa}B$-${\alpha}$, IKK-${\alpha}$, NF-${\kappa}B$ (p50, p65), COX-2, iNOS, YCAM-1 and MMP-9 genes. Conclusions: These results suggest that Sotosaja-hwan is an effective $ONOO^-$, ${\cdot}{O_2}^-$ and NO scavenger and has NF-kB related anti-inflammatory activity in ob/ob mice. Therefore, Sotosaja-hwan might be a potential therapeutic drug against the inflammation process and inflammation-related diseases.

• Journal of Applied Biological Chemistry
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• v.59 no.4
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• pp.365-372
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• 2016

The objective of the present study was to evaluate the protective effect of red ginseng (RG) according to steaming time on 150 mM HCl/60 % ethanol induced gastric ulcer models in mice. The sample was divided into 3 groups-G (dried ginseng), RG 4 (steamed 4 h and dried ginseng), RG 6 (steamed 6 h and dried ginseng), and determined through in vitro experiments, such as 1,1-diphenyl-2-picrylhydrazyl and 2,2'-azinobis-3-ethyl-benzothiazoline-6-sulfonic acid radical scavenging activity, HPLC analysis, total polyphenol, and flavonoid contents. In vitro experiment results were depended on steaming hours. Based on the results, we chose two samples (G, RG 6) and conducted in vivo experiments. Mice were divided into 5 groups: Nor (normal group), Con (acute gastritis mice treated with distilled water), G (gastris induced by HCl/Ethanol treated with 100 mg/kg G), RG 6 (gastris induced by HCl/ethanol treated with 100 mg/kg RG 6), and SC (gastris induced by HCl/Ethanol treated with 10 mg/kg sucralfate). In our results revealed that RG 6 suppressed elevated reactive oxygen species, and inflammatory related makers, such as cyclooxygenase-2, inducible nitric oxide synthase, tumor necrosis factor alpha, and interleukin-1 beta. In addition, gastric lesion area was improved. These results suggest that RG 6 protects the stomach by attenuating oxidative stress and inflammatory response under gastric ulcer conditions. Therefore, RG 6 should be a potential therapeutic agent for the treatment of acute gastric ulcer.

• The Korean Journal of Physiology and Pharmacology
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• v.19 no.3
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• pp.219-228
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• 2015

Excessive microglial activation and subsequent neuroinflammation lead to synaptic loss and dysfunction as well as neuronal cell death, which are involved in the pathogenesis and progression of several neurodegenerative diseases. Thus, the regulation of microglial activation has been evaluated as effective therapeutic strategies. Although dieckol (DEK), one of the phlorotannins isolated from marine brown alga Ecklonia cava, has been previously reported to inhibit microglial activation, the molecular mechanism is still unclear. Therefore, we investigated here molecular mechanism of DEK via extracellular signal-regulated kinase (ERK), Akt and nicotinamide adenine dinuclelotide phosphate (NADPH) oxidase-mediated pathways. In addition, the neuroprotective mechanism of DEK was investigated in microglia-mediated neurotoxicity models such as neuron-microglia co-culture and microglial conditioned media system. Our results demonstrated that treatment of anti-oxidant DEK potently suppressed phosphorylation of ERK in lipopolysaccharide (LPS, $1{\mu}g/ml$)-stimulated BV-2 microglia. In addition, DEK markedly attenuated Akt phosphorylation and increased expression of $gp91^{phox}$, which is the catalytic component of NADPH oxidase complex responsible for microglial reactive oxygen species (ROS) generation. Finally, DEK significantly attenuated neuronal cell death that is induced by treatment of microglial conditioned media containing neurotoxic secretary molecules. These neuroprotective effects of DEK were also confirmed in a neuron-microglia co-culture system using enhanced green fluorescent protein (EGFP)-transfected B35 neuroblastoma cell line. Taken together, these results suggest that DEK suppresses excessive microglial activation and microglia-mediated neuronal cell death via downregulation of ERK, Akt and NADPH oxidase-mediated pathways.

• The Korean Journal of Physiology and Pharmacology
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• v.15 no.3
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• pp.149-156
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• 2011

Golgi SNAP receptor complex 1 (GS28) has been implicated in vesicular transport between intra-Golgi networks and between endoplasmic reticulum (ER) and Golgi. Additional role(s) of GS28 within cells have not been well characterized. We observed decreased expression of GS28 in rat ischemic hippocampus. In this study, we examined the role of GS28 and its molecular mechanisms in neuronal (SK-N-SH) cell death induced by hydrogen peroxide ($H_2O_2$). GS28 siRNA-transfected cells treated with $H_2O_2$ showed a significant increase in cytotoxicity under glutathione (GSH)-depleted conditions after pretreatment with buthionine sulfoximine, which corresponded to an increase of intracellular reactive oxygen species (ROS) in the cells. Pretreatment of GS28 siRNA-transfected cells with p38 chemical inhibitor significantly inhibited cytotoxicity; we also observed that p38 was activated in the cells by immunoblot analysis. We confirmed the role of p38 MAPK in cotransfected cells with GS28 siRNA and p38 siRNA in the cell viability assay, flow cytometry, and immunoblot. Involvement of apoptotic or autophagic processes in the cells was not shown in the cell viability, flow cytometry, and immunoblot analyses. However, pretreatment of the cells with necrostatin-1 completely inhibited $H_2O_2$-induced cytotoxicity, ROS generation, and p38 activation, indicating that the cell death is necroptotic. Collectively these data imply that $H_2O_2$ induces necroptotic cell death in the GS28 siRNA-transfected cells and that the necroptotic signals are mediated by sequential activations in RIP1/p38/ROS. Taken together, these results indicate that GS28 has a protective role in $H_2O_2$-induced necroptosis via inhibition of p38 MAPK in GSH-depleted neuronal cells.

• The Korean Journal of Physiology and Pharmacology
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• v.17 no.3
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• pp.181-187
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• 2013

Reactive oxygen species (ROS) are generated in various cells, including vascular smooth muscle and endothelial cells, and regulate ion channel functions. $K_{Ca}3.1$ plays an important role in endothelial functions. However, the effects of superoxide and hydrogen peroxide radicals on the expression of this ion channel in the endothelium remain unclear. In this study, we examined the effects of ROS donors on $K_{Ca}3.1$ expression and the $K^+$ current in primary cultured human umbilical vein endothelial cells (HUVECs). The hydrogen peroxide donor, tert-butyl hydroperoxide (TBHP), upregulated $K_{Ca}3.1$ expression, while the superoxide donors, xanthine/xanthine oxidase mixture (X/XO) and lysophosphatidylcholine (LPC), downregulated its expression, in a concentration-dependent manner. These ROS donor effects were prevented by antioxidants or superoxide dismustase. Phosphorylated extracellular signal-regulated kinase (pERK) was upregulated by TBHP and downregulated by X/XO. In addition, repressor element-1-silencing transcription factor (REST) was downregulated by TBHP, and upregulated by X/XO. Furthermore, $K_{Ca}3.1$ current, which was activated by clamping cells with 1 ${\mu}M$ $Ca^{2+}$ and applying the $K_{Ca}3.1$ activator 1-ethyl-2-benzimidazolinone, was further augmented by TBHP, and inhibited by X/XO. These effects were prevented by antioxidants. The results suggest that hydrogen peroxide increases $K_{Ca}3.1$ expression by upregulating pERK and downregulating REST, and augments the $K^+$ current. On the other hand, superoxide reduces $K_{Ca}3.1$ expression by downregulating pERK and upregulating REST, and inhibits the $K^+$ current. ROS thereby play a key role in both physiological and pathological processes in endothelial cells by regulating $K_{Ca}3.1$ and endothelial function.