• Biomolecules & Therapeutics
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• 제26권2호
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• pp.175-181
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• 2018

Carbon monoxide (CO) is well-known as toxic gas and intrinsic signaling molecule such as neurotransmitter and blood vessel relaxant. Recently, it has been reported that low concentration of CO exerts therapeutic actions under various pathological conditions including liver failure, heart failure, gastric cancer, and cardiac arrest. However, little has been known about the effect of CO in neurodegenerative diseases like Parkinson's disease (PD). To test whether CO could exert a beneficial action during oxidative cell death in PD, we examined the effects of CO on 6-hydroxydopamine (6-OHDA)-induced cell death in C6 glioma cells. Treatment of CO-releasing molecule-2 (CORM-2) significantly attenuated 6-OHDA-induced apoptotic cell death in a dose-dependent manner. CORM-2 treatment decreased Bax/Bcl2 ratio and caspase-3 activity, which had been increased by 6-OHDA. CORM-2 increased phosphorylation of NF-E2-related factor 2 (Nrf2) which is a transcription factor regulating antioxidant proteins. Subsequently, CORM-2 also increased the expression of heme oxygenase-1 and superoxide dismutases (CuZnSOD and MnSOD), which were antioxidant enzymes regulated by Nrf2. These results suggest that CO released by CORM-2 treatment may have protective effects against oxidative cell death in PD through the potentiation of cellular adaptive survival responses via activation of Nrf2 and upregulation of heme oxygenase-1, leading to increasing antioxidant defense capacity.

• 한방비만학회지
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• 제15권2호
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• pp.93-103
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• 2015

Objectives: It was investigated the cytoprotective efficacies of isorhamnetin, a flavonoid originally derived from Hippophae rhamnoides L., against oxidative stress-induced apoptosis in C2C12 myoblasts. Methods: The effects of isorhamnetin on cell growth, apoptosis and reactive oxygen species (ROS) generation were evaluated by trypan blue dye exclusion assay, 4',6-diamidino-2-phenylindole staining and flow cytometry. The levels of apoptosis-regulatory and nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway-related proteins, and caspase activities (caspase-3 and -9) were determined by Western blot analysis and colorimetric assay, respectively. Results: Our results revealed that treatment with isorhamnetin prior to hydrogen peroxide ($H_2O_2$) exposure significantly increased the C2C12 cell viability and, indicating that the exposure of C2C12 cells to isorhamnetin conferred a protective effect against oxidative stress. Isorhamnetin also effectively attenuated $H_2O_2$-induced apoptosis and ROS generation, which was associated with the restoration of the upregulation of Bax and downregulation of Bcl-2 induced by $H_2O_2$. In addition, $H_2O_2$ enhanced the activation of caspase-9 and -3, and degradation of poly (ADP-ribose)-polymerase, a typical substrate protein of activated caspase-3; however, these events were almost totally reversed by pretreatment with isorhamnetin. Moreover, isorhamnetin increased the levels of heme oxygenase-1, a potent antioxidant enzyme, associated with the induction of Nrf2. Conclusions: Our data indicated that isorhamnetin may potentially serve as an agent for the treatment and prevention of muscle disorders caused by oxidative stress.

• 한방안이비인후피부과학회지
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• 제34권4호
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• pp.24-36
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• 2021

Objectives : This study was conducted to confirm the anti-oxidative effect of Chungpyesagan-tang(CPSGT) extract. Methods : In this study, MTT assay was performed to confirm cell viability, and DPPH and ABTS were performed to confirm radical scavenging ability. The ROS scavenging ability and the protective effect against DNA damage were confirmed by 2,7-dichlorofluorescin diacetate(DCF-DA) and 4',6-diamidino-2-phenylindole(DAPI) staining and comet assay. mRNA expression of Heme oxygenase-1(HO-1) was measured by real-time PCR, and expression of HO-1 and Kelch-like ECH-associated protein 1(Keap1) proteins was measured by western blot. Results : CPSGT was not cytotoxic at 50-400㎍/㎖. The radical scavenging activity was increased, and the ROS scavenging activity and the protective effect against DNA damage were increased compared to the LPS-treated group. The mRNA expression and protein expression of HO-1 were increased in a concentration-dependent manner. The protein expression level of Keap1 was decreased in a concentration-dependent manner. Conclusion : This suggests that CPSGT has an antioxidant effect and can be used as a potential material for skin diseases.

• 대한화장품학회지
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• 제46권3호
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• pp.253-263
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• 2020

활성산소종(reactive oxygen species, ROS)은 우리 몸의 항상성 유지에 있어 중요한 역할을 한다. 그러나 과도한 ROS의 생성은 단백질, 지질, 핵산과 같은 세포 구성성분을 손상시키고 피부노화를 촉진시킨다. 이에 본 연구에서는 과도한 산화 스트레스를 예방하기 위해 Chungsimyeonja-um (CSYJE)의 항산화 효과를 확인하였다. 먼저 DPPH 및 ABTS assay를 실시하여 CSYJE의 항산화 효과를 확인한 결과 농도 의존적으로 radical 소거 활성을 확인하였다. 세포생존율 확인을 위해 MTT assay를 실시한 결과 1,000 ㎍/mL 농도에서 세포 독성이 없음을 확인하였다. 항산화 관련 단백질인 nuclear-E2-related factor 2 (Nrf2), Heme oxygenase-1 (HO-1)의 발현 수준을 확인하기 위해 western blotting을 실시한 결과 농도 의존적으로 발현이 증가하는 것을 확인하였다. 세포 내 ROS유발 물질인 lipopolysaccharide (LPS)로 ROS를 유도한 후, ROS생성 억제효과를 확인하기 위해 DCF-DA 염색법을 실시한 결과 농도 의존적으로 ROS 생성 억제효과를 확인하였으며 ROS의 생성으로 인한 염증성 사이토카인과 염증인자의 mRNA발현 수준을 확인하기 위해 real-time RT-PCR을 실시한 결과 농도 의존적으로 염증성 사이토카인과 염증인자의 mRNA 발현을 억제시켰다. 따라서, 본 연구는 Nrf2/HO-1 신호 전달 경로 활성을 통해 CSYJE의 항산화효과를 확인했으며 이는 CSYJE가 활성산소를 억제하여 항산화 화장품의 재료로서 사용될 수 있음을 시사한다.

• 생명과학회지
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• 제23권8호
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• pp.1057-1063
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• 2013

진피(Citri Pericarpium)의 항암작용 기전 해석을 위하여 U937 백혈병 세포의 apoptosis 유발에 미치는 메탄올 추출물(EMCP)의 영향을 조사하였으며, apoptosis 조절에 중요한 몇 가지 유전자들의 발현 및 활성 변화, ROS의 생성 변화를 조사하였다. EMCP 처리에 의한 U937 세포의 증식 억제는 apoptosis 유도와 연관성이 있음을 DAPI 염색을 통한 apoptotic body 출현의 증가 및 Flow cytometry 분석에 의한 Sub-G1기 세포 빈도의 증가로 확인하였다. EMCP 처리에 의한 apoptosis 유도에서 Bax 발현 증가, caspases의 활성 및 PARP의 단편화 등이 동반되었으며, ROS 생성의 증가와 연관성이 있었다. 산화적 손상에 대해 세포나 조직을 보호하는 역할을 하는 것으로 알려진 세포 내 항산화 효소인 HO-1의 발현이 EMCP의 처리에 의해 증가되었으나 ROS 생성 억제제인 NAC의 전처리에 의해 감소된 HO-1의 발현은 전사인자인 Nrf2의 핵으로의 이동 억제와 관련되어 있었다. 이상의 결과에서 EMCP 처리에 의한 U937 세포의 apoptosis 유발에는 ROS 생성의 증가와 pNrf2에 의해 조절되는 HO-1의 발현 증가가 중요한 기전으로 작용한다는 것을 알 수 있었다. 이러한 자료는 진피의 항암기전 해석을 이해하는데 중요한 기초자료로서 활용될 수 있을 것으로 생각된다.

• BMB Reports
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• 제31권2호
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• pp.161-169
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• 1998

TThe reactive oxygen species oxidatively modify the biological macromolecules, including proteins, lipids, and nucleic acids. Iron- and heme-mediated Fenton-like reactions produce different pro-oxidants. However, these reactive products have not been clearly characterized. We examined the nature of the oxidizing species from the different iron sources by measuring oxidative protein modification and spectroscopic study. Hemoglobin (Hb) and methemoglobin (metHb) were oxidatively modified in $O{\array-\\\dot{2}}$ and $H_{2}O_{2}$ generating systems. Globin and bovine serum albumin (BSA) were also modified by iron, iron-EDTA, hematin, and Hb in an $O{\array-\\\dot{2}}$ generating system. In a $H_{2}O_{2}$ generating system, the iron- and iron-EDTA-mediated protein modifications were markedly reduced while the Hb-and hematin-mediated modifications were slightly increased. In the $O{\array-\\\dot{2}}$ generating system, the iron- and iron-EDTA-mediated protein modifications were strongly inhibited by superoxide dismutase (SOD) or catalase, but heme- and Hb-mediated protein modifications were inhibited only by catalase and slightly increased by SOD. Mannitol, 5,5-dimethyl-l-pyrroline-N-oxide (DMPO), deoxyribose, and thiourea inhibited the iron-EDTA-mediated protein modification. Mannitol and DMPO, however, did not exhibit significant inhibition in the hematin-mediated modification. Desferrioxamine (DFO) inhibited protein modification mediated by iron, but cyanide and azide did not, while the hematin-mediated protein modification was inhibited by cyanide and azide, but not significantly by DFO. The protein-modified products by iron and heme were different. ESR and UV-visible spectroscopy detected the DMPO spin adduct of the hydroxyl radical and ferryl ion generated from iron-EDTA and metHb, respectively. These results led us to conclude that the main oxidizing species are hydroxyl radical in the iron-EDTA type and the ferry I ion in the hematin type, the latter being more effective for protein modification.

• 분석과학
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• 제27권5호
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• pp.223-227
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• 2014

${\delta}$-Aminolevulinic acid (ALA)는 생물권내에 폭넓게 존재하는 화합물이고, 포유류에서 헴(heme)과 식물에서 엽록소 생성을 하게하는 경로에서 만들어진 테트라피롤의 중간물질로써 생체내 중요한 역할을 한다. ALA는 생분해성 매개자, 성장 조절자, 헴단백질의 전구체 그리고 암 치료에서 사용되는 효과적인 물질로써 관심이 있다. 최근에는 ALA가 피부 치료의 좋은 효과를 가지고 있어 피부과학에서 빈번히 사용된다고 보고되어 있는데, 하지만 지난 몇 십 년 동안, 많은 연구가 ALA 메커니즘의 설명과 치료적 활성 개선에 초점을 맞춰왔지만, 세포 기능과 세포생장에 대한 ALA의 효과는 아직 불분명하다. 본 연구에서는 ALA의 약물학적 효과가 HEK293T 뿐만 아니라, HaCaT와 HeLa 세포에서 세포분열을 억제하는 것으로 확인을 하였다. 또한, ALA가 처리된 세포에서는 세포예정사가 유도되는 것을 확인하였다. 이러한 결과들은 특정 세포에 ALA가 처리되면 세포증식을 억제하며, 특히 인간의 암세포의 죽임을 유발하는 효과적인 약물 중 하나로써 ALA를 개선된 치료 전략으로 가능성을 제시한다.

• 대한본초학회지
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• 제34권2호
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• pp.1-14
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• 2019

Objective : This study was designed to evaluate the protective effect of Rehmanniae Radix Crudus (RC) in 150 mM HCl/ethanol induced acute gastritis mice. Methods : ICR mice were divided into 5 groups (normal group, control group, 10 mg/kg sucralfate treated group, 50 mg/kg RC treated group, 100 mg/kg RC treated group, n=8). Normal group was not take any treatment. Control group induced gastritis 1 hour after ingestion of distilled water. 10 mg/kg sucralfate induced group was induced gastritis 1 hour after ingestion of distilled of sucralfate 10 mg/kg. 50 mg/kg and 100 mg/kg RC treated groups were induced gastritis 1 hour after ingestion of distilled of RC 50 mg/kg and 100 mg/kg. After 1 hour of gastritis induction, removed the stomach tissue. We examined histological observations, oxidative stress biomarkers, antioxidant proteins, inflammatory mediators and cytokines. Results : In this study, the RC treatment group showed gastritis and gastric ulcer inhibition, and the area of injury decreased. The oxidative stress biomarkers such as reactive oxygen species (ROS) and peroxy nitrite ($ONOO^-$) in the serum were reduced in the RC treated group. Inaddition, antioxidant proteins (nuclear factor erythroid 2-related factor 2, Heme oxygenase 1) were increased in RC treated group, and the expression of inflammatory mediators and cytokines induced by nuclear factor-kappa B activation was inhibited. Conclusion : According to the results, RC may have an excellent inhibitory effect on acute gastritis and gastric ulcer.

• Biomolecules & Therapeutics
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• 제30권5호
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• pp.447-454
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• 2022

Few studies have evaluated the role of autophagy in the development of oxaliplatin (OXT) resistance in colon cancer cells. In this study, we compared the role of autophagy between SNU-C5 colon cancer cells and OXT-resistant SNU-C5 (SNU-C5/OXTR) cells. At the same concentration of OXT, the cytotoxicity of OXT or apoptosis was significantly reduced in SNU-C5/OXTR cells compared with that in SNU-C5 cells. Compared with SNU-C5 cells, SNU-C5/OXTR cells exhibited low levels of autophagy. The expression level of important autophagy proteins, such as autophagy-related protein 5 (Atg5), beclin-1, Atg7, microtubule-associated proteins 1A/1B light chain 3B I (LC3-I), and LC3-II, was significantly lower in SNU-C5/OXTR cells than that in SNU-C5 cells. The expression level of the autophagy-essential protein p62 was also lower in SNU-C5/OXTR cells than in SNU-C5 cells. In SNU-C5/OXTR cells, the production of intracellular reactive oxygen species (ROS) was significantly higher than that in SNU-C5 cells, and treatment with the ROS scavenger N-acetylcysteine restored the reduced autophagy levels. Furthermore, the expression of antioxidant-related nuclear factor erythroid 2-related factor 2 transcription factor, heme oxygenase-1, and Cu/Zn superoxide dismutase were also significantly increased in SNU-C5/OXTR cells. These findings suggest that autophagy is significantly reduced in SNU-C5/OXTR cells compared with SNU-C5 cells, which may be related to the production of ROS in OXT-resistant cells.

• Archives of Pharmacal Research
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• 제12권2호
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• pp.79-87
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• 1989

Enzymatic methylation of arginine and lysine residues of several cytochrome c and lysine residue of calmodulin always resulted in lowering of their respective isoelectric points (pI). Employing cytochromes c derived from various sources, we examined a possible relationship between the degree of amino acid sequence degeneracy and the magnitude of change in the pI values by enzymatic methylation, and found that there was no correlation between these two parameters. By constructing space-filling models of oligopeptide fragments adjacent to the potential methylation sites, we have noted that not all the methylatable residues are able to form hydrogen bonds prior to the methylation. Two preparations of yeast apocytochrome c, one chemically prepared by removing heme from holocytochrome c and the other by translating yeast iso-1-cytochrome c mRNA in vitro, exhibited slightly higher Stokes radii than the homologous holocytochrome c, indicating relatively 'relaxed or open' conformation of the protein. However, when the in vitro synthesized methylated apocytochrome c was compared with the unmethylated counter-part, the Stokes radius of the latter was found to be larger.