• 한국작물학회지
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• 제56권4호
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• pp.349-360
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• 2011

The objectives of present study were to characterize the peptides which were isolated from Korean fermented soybean paste, chungkukjang, and to determine their antioxidant activities. Four fractions were collected from the methanol extract of chungkukjang by using a recycling preparative HPLC. Among fractions, Fr-2 was identified to be highly potent free radical scavenging activity in the assay of 1,1-diphenyl-2-picryl-hydrazyl (DPPH) and nitroblue tetrazolium(NBT)-reduction inhibition. Base on antioxidant effects, fraction Fr-2 was employed for the refraction with a prep-column and separated into five fractions of which two fractions were identified to have higher antioxidant activity. To confirm the amino acid constituents of antioxidant fractions Fr-2-2 and Fr-2-3 were analyzed, and eight kinds of amino acids such as aspartic acid, threonine, serine, glutamic acid, glycine, lysine, histidine, and arginine were identified as the constituent amino acids. Antioxidant activities of the separated peptides were further assessed cell viability with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl terazolium bromide (MTT), and fluorescence-activated cell sorting (FACS) analysis of H4IIE cells treated with hydrogen peroxide (H2O2). Chungkukjang peptides have shown their ability to protect H4IIE rat hepatoma cells against H2O2- induced oxidative stress by concentration and time-dependent manner. Therefore, These results indicated that fermented soybean paste chungkukjang will be promoted the antioxidant and radical scavenging activities, and beneficial for health. The antioxidant peptide fractions Fr-2-2 and Fr-2-3 were denominated as P-NICS-1 and P-NICS-2, respectively. However, further studies were required to clarify their amino acid sequences and molecular properties, and physiological significances.

• Nutritional Sciences
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• 제9권1호
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• pp.9-13
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• 2006

The differential effects of various fatty acids such as n-3 and n-6 types or degrees of unsaturation on the CYP2E1 induction and the production of lipid peroxidation (LPO) were investigated. The CYP2E1-transduced human hepatoma HepG2 cells (E47) were cultured in RPMI 1640 media containing different concentrations of various fatty acids up to 48 h incubation compared to 04 cells and CYP2E1-null cells. Treated fatty acids were linoleic acid (LA:n-6, C18:2), arachidonic acid (AA:n-6, C20:4) and docosahexaenoic acid (DHA:n-3, C22:6). The cell survival rate was decreased corresponding to the degree of unsaturation (LA>AA $\cong$DHA) and to LPO production in E47 and 04 cells. The four or five unsaturation degree of fatty acids, AA and DHA, caused time- and dose-dependent cell death in E47 cells but not as much as in C34 (without CYP2E1), suggesting an important role of CYP2E1 in the DHA mediated damage. In the levels of lipid peroxides (LPO), AA also elevated LPO by 3- and 5- fold compared to DHA or LA treated E47 cells. However, AA did not increase LPO until 48 h incubation in C34 cells. In conclusion, the polyunsaturated fatty acids induced CYP2E1 induction might be changed by the elevated levels of lipid peroxide (LPO) and oxidative stress through the connection of CYP2E1 and degrees of unsaturated fatty acids.

• Molecular & Cellular Toxicology
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• 제5권4호
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• pp.310-316
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• 2009

Some environmental chemicals have been shown to cause liver-toxicity as the result of bioaccumulation. Particularly, fungicides have been shown to cause varying degrees of hepatictoxicity and to disrupt steroid hormone homeostasis in in vivo models. The principal objective of this study was to evaluate the liver-toxic responses of environmental chemicals-in this case selected fungicides and parasiticides-in order to determine whether or not this agent differentially affected its toxicogenomic activities in hepatic tumor cell lines. To determine the gene expression profiles of 3 fungicides (triadimefon, myclobutanil, vinclozolin) and 1 parasiticide (dibutyl phthalate), we utilized a modified HazChem human array V2. Additionally, in order to observe the differential alterations in its time-dependent activities, we conducted two time (3 hr, 48 hr) exposures to the respective IC20 values of four chemicals. As a result, we analyzed the expression profiles of a total of 1638 genes, and we identified 70 positive significant genes and 144 negative significant genes using four fungicidic and parasiticidic chemicals, using SAM (Significant Analysis of Microarray) methods (q-value<0.5%). These genes were analyzed and identified as being related to apoptosis, stress responses, germ cell development, cofactor metabolism, and lipid metabolism in GO functions and pathways. Additionally, we found 120 genes among those time-dependently differentially expressed genes, using 1-way ANOVA (P-value<0.05). These genes were related to protein metabolism, stress responses, and positive regulation of apoptosis. These data support the conclusion that the four tested chemicals have common toxicogenomic effects and evidence respectively differential expression profiles according to exposure time.

• 동의생리병리학회지
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• 제19권3호
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• pp.640-646
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• 2005

This study was performed for the investigation of anticancer effects of Siegesbeckia glabrescens(SG) on HepG2 cells, a human hepatoma cell line. In the previous study, we examined the involvement of nitric oxide (NO) on anti-proliferative and apoptotic efficacy of SG in vascular smooth muscle cells. The possible mechanism of the apoptotic effects of SG was investigated in HepG2 cells. SG showed potent cytotoxic activity in HepG2 but not chang cells, liver normal cells. SG treatment caused morphological change such as cell shrinkage, nuclei condensation and cell blebbing in HepG2 cells. SG also increased the nitrite production of HepG2 cells in a dose-dependent manner. Furthermore, L-NNA treatment inhibited the anti-proliferative effect of SG. From RT-PCR, SG decreased Bcl-2 but no affected on Bax. Western blot for procaspase-3 and COX-2 showed that degradation of procaspase-3 protein level or inhibition of COX-2 protein expression by SG treatment. In addition, the apoptotic effect of SG was also demonstrated by DNA laddering. In conclusion, SG-induced HepG2 cells death can occur via apoptosis which was dose-dependent, and associated with apoptosis-related Bcl-2/Bax gene expressions, COX-2 inhibition, caspase-3 activation and NO pathway. These results suggest that SG is potentially useful as a chemotherapeutic/chemopreventive agent in hepatocellular carcinoma.

• 한국식품영양학회지
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• 제27권3호
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• pp.400-405
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• 2014

최근 알로에 베라 섭취에 의한 독성 간염이 보고되고 있으나, 아직까지 알로에 베라의 간에 대한 염증 효과가 명확히 밝혀지지 않았다. 본 연구는 알로에 베라 에탄올 추출물이 간세포의 염증 발현에 미치는 영향과 그 기전을 밝히기 위해 수행되었다. $0.001{\sim}100{\mu}g/mL$의 알로에 베라 추출물을 HepG2 세포에 처리하여 MTT assay를 시행한 결과, 모든 농도에서 세포사멸이 유도되지 않았다. 그러나 모든 농도에서 알로에 베라 추출물은 염증성 사이토카인인 IL-8의 분비를 15.7~25.8%까지 유의적으로 증가시켰다(p<0.05). 또 다른 사이토카인인 M-CSF도 알로에 베라 추출물에 의해 36.3~61.5%까지 유의적으로 분비가 증가되었다(p<0.05). 본 연구 결과는 알로에 베라 추출물이 IL-8과 M-CSF와 같은 염증성 사이토카인 분비기전에 의해 간에 염증을 유발할 수 있음을 보여준다. 또한 알로에 추출물에 의해 유발될 수 있는 간염기전을 제시함으로써, 향후 수행될 추가 실험을 위한 기초 자료로 그 가치가 높을 것으로 사료된다.

• 대한임상독성학회지
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• 제15권2호
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• pp.107-115
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• 2017

Purpose: Glehnia littoralis has been used to treat ischemic stroke, phlegm, cough, systemic paralysis, antipyretics and neuralgia. The pharmacological mechanisms of Glehnia littoralis include calcium channel block, coumarin derivatives, anticoagulation, anti-convulsive effect, as well as anti-oxidant and anti-inflammatory effects. Alpha-amanitin (${\alpha}$-amanitin) is a major toxin from extremely poisonous Amanita fungi. Oxidative stress, which may contribute to severe hepatotoxicity was induced by ${\alpha}$-amanitin. The aim of this study was to investigate whether Glehnia littoralis ethyl acetate extract (GLEA) has the protective antioxidant effects on ${\alpha}$-amanitin -induced hepatotoxicity. Methods: Human hepatoma cell line HepG2 cells were pretreated in the presence or absence of GLEA (50, 100 and $200{\mu}g/ml$) for 4 hours, then exposed to $60{\mu}mol/L$ of${\alpha}$-amanitin for an additional 4 hours. Cell viability was evaluated using the MTT method. AST, ALT, and LDH production in a culture medium and intracellular MDA, GSH, and SOD levels were determined. Results: GLEA (50, 100 and $200{\mu}g/ml$) significantly increased the relative cell viability by 7.11, 9.87, and 14.39%, respectively, and reduced the level of ALT by 10.39%, 34.27%, and 52.14%, AST by 9.89%, 15.16%, and 32.84%, as well as LDH by 15.86%, 22.98%, and 24.32% in culture medium, respectively. GLEA could also remarkably decrease the level of MDA and increase the content of GSH and SOD in the HepG2 cells. Conclusion: In the in vitro model, Glehnia littoralis was effective in limiting hepatic injury after ${\alpha}$-amanitin poisoning. Its antioxidant effect is attenuated by antidotal therapy.

• 한국식품영양과학회지
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• 제35권9호
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• pp.1121-1126
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• 2006

In the present study, the potential hepatoprotective effects of poly herbal formulation, Hepa-1000, against oxidative damages induced by t-BHP were evaluated in HepG2 cells in order to relate in vitro antioxidant activity with cytoprotective effects. The t-BHP induced considerable cell damage in HepG2 cells was shown by significant glutamic oxaloacetic transaminase (GOT) and lactate dehydrogenase (LDH) leakage, and increased lipid peroxidation. Hepa-1000-treated cells showed an increased resistance to oxidative challenge, as revealed by higher survival capacity than the one of control cells against t-BHP induced oxidative stress and hepatotoxicity. In addition, the Hepa-1000 had hepatoprotective effects lowering the activity of GOT and LDH, simultaneously. That is, it could inhibit the cell membrane damages resulting in the increased activities of GOT and LDH in the cell culture media. Furthermore, the Hepa-1000 could reduce t-BHP enhanced lipid peroxidation, which was evaluated by measuring the production of malonedialdehyde. Based on the data described above, it could be suggested that the Hepa-1000 has significant hepatoprotective effects and plays a protective role against lipid peroxidation by free radicals.

• 한국약용작물학회지
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• 제27권3호
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• pp.208-217
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• 2019

Background: In the present study, we identified two cytotoxic compounds from the root of Rumex crispus L. using a bioassay-based method. Methods and Results: Compared with the other fractions, the diethyl ether ($Et_2O$) fraction of R. crispus root extract exhibited the strongest of 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging effect [scavenging concentration 50% $(SC_{50})=63.8{\pm}1.47{\mu}g/m{\ell}$], nitric oxide (NO) production inhibitory effect on the mouse macrophage cell line RAW264.7 [inhibitory concentration 50% $(IC_{50})=60.9{\pm}7.52{\mu}g/m{\ell}$] and cytotoxicity effect on the human hepatoma cell line, HepG2 [lethal concentration 50% $(LC_{50})=115.4{\pm}1.86{\mu}g/m{\ell}$]. According to the bioassay-based method, two cytotoxic compounds were purified from the $Et_2O$ fraction by using column chromatography and preparative high performance liquid chromatography (prep-HPLC). These two compounds were identified as parietin and chrysophanol by using nuclear magnetic resonance (NMR) and liquid chromatography quadruple time of flight mass spectrometry (LC-QTOF-MS). In addition, both parietin and chrysophanol exhibited a cytotoxicity effect on HepG2 cells, their $LC_{50}$ values were $169.1{\pm}17.67{\mu}M$ and $111.5{\pm}6.62{\mu}M$, respectively. Conclusions: Parietin and chrysophanol isolated from the $Et_2O$ fraction of the R. crispus root extract showed cytotoxicity in HepG2 cell.

• 생명과학회지
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• 제26권7호
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• pp.764-771
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• 2016

개똥쑥 추출물은 항박테리아, 항바이러스 그리고 항산화효과를 포함한 다양한 기능을 가지고 있는 것으로 잘 알려져 있다. 그러나, 개똥쑥 항증식 작용기전은 알려지지 않았다. 따라서, 우리는 Hep3B 간암 세포에서 AAE추출물의 apoptotic 효과를 알아보고자 한다. 본 연구의 목적은 AAE가 인체 간암 세포주(Hep3B)의 증식에 미치는 효과를 분석하고 이에 대한 apoptosis의 효과를 조사하는데 있다. 인산화에 의해 활성화된 Akt는 TSC2, mTOR 그리고 GSK-3β의 인산화를 유도하여 세포증식을 유도한다. 본 연구에서, 우리는 AAE가Akt-mTOR-GSK3β 신호 경로와 mitochondria를 매개하는 apoptotic 단백질을 통한 암세포의 apoptosis 유도할 것이라고 추측하였다. 이를 위해, 먼저 AAE가 처리농도에 따라 세포증식에 미치는 효과를 분석하였다. AAE처리는 세포증식을 억제시켰을 뿐만 아니라 젖산 탈수소 효소의 방출을 유도하였다. 이러한 결과는 MTT assay, LDH assay로 확인하였다. 또한 Hoechst 33342 staining, Annexin Ⅴ- PI staining, JC-1 staining 그리고 Western blotting을 통해 apoptosis 효과를 확인하였다. 본 연구에서는 간암세포에 AAE의 처리가 Akt, TSC2, GSK-3β-phosphorylated, Bim, Bcl-2, pro-caspase 3의 억제와 Bak, Bax 활성을 유도한다는 것을 확인하였다. 이러한 결과는 AAE가 Akt-mTOR-GSK-3β 신호 경로를 통해 intrinsic apoptosis를 유도한다는 것을 나타낸다.

• 생명과학회지
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• 제17권5호
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• pp.671-677
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• 2007

발효 및 비발효 차가버섯 수용성 추출물이 정상 세포주 NIH3T3 mouse normal fibroblast cell 및 인체 종양 세포주 AGS human gastric cancer cell(위암), HCT-15 human colon cancer cell(대장암), Hep3B human hepatoma cancer cell(간암), MCF-7 human breast cancer cell(유방암), HeLa human cervical cancer cell(자궁경부암)에서 MTT(3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay 방법에 의한 세포 증식 억제와 암세포 증식억제의 기전 연구의 일환으로 apoptosis가 일어날 때 나타나는 DNA fragmentation을 agarose gel electrophoresis 방법으로 검토하였다. 인체 종양 세포주의 생육저해 효과가 발효 차가버섯 추출물이 비발효 차가버섯 추출물보다 강한 것으로 나타났다. 그러나 동일한 실험조건하에서 마우스 정상 세포주 NIH3T3은 80% 이상의 생존율을 나타내어 정상 세포주에는 큰 영향을 미치지 않는 것으로 나타났다. 특히, 발효 및 비발효 차가버섯 추출물에서 본 실험에 사용한 세포주 중에서 대장암 세포주 HCT-15에 대해 가장 세포 증식 억제효과가 뛰어났으며, 이러한 효과는 첨가 농도 의존적 이였다. 발효 및 비발효 차가버섯 추출물에 의한 암세포 증식억제가 기전 연구로 apoptosis가 일어날 때 나타나는 DNA fragmentation을 세포로부터 genomic DNA를 분리하여 agarose gel electrophoresis 방법으로 조사한 결과, 정상세포인 NIH3T3 세포는 DNA fragmentation이 거의 일어나지 않아 세포 생존율 결과와 유사한 경향을 보였으나, 특히 대장암 세포주인 HCT-15에서는 발효 차가버섯뿐만 아니라 비발효 차가버섯 추출물에서도 DNA fragmentation이 많이 일어나는 것이 관찰되어 암세포 증식억제 효과가 높다는 결과를 뒷받침 해주고 있다.