• Asian-Australasian Journal of Animal Sciences
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• 제30권9호
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• pp.1245-1252
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• 2017

Objective: Phellodendron amurense (P. amurense) and Humulus japonicus (H. japonicus) are closely involved in anti-oxidative response and increasing antioxidant enzymes activities. However, the effects of their extracts on development of preimplantation bovine embryos have not been investigated. Therefore, we investigated the effects of P. amurense and H. japonicus extracts on developmental competence and quality of preimplantation bovine embryos. Methods: After in vitro fertilization, bovine embryos were cultured for 7 days in Charles Rosenkrans amino acid medium supplemented with P. amurense ($0.01{\mu}g/mL$) and H. japonicus ($0.01{\mu}g/mL$). The effect of this supplementation during in vitro culture on development competence and antioxidant was investigated. Results: We observed that the blastocysts rate was significantly increased (p<0.05) in P. amurense ($28.9%{\pm}2.9%$), H. japonicus ($30.9%{\pm}1.5%$), and a mixture of P. amurense and H. japonicus ($34.8%{\pm}2.1%$) treated groups compared with the control group ($25.4%{\pm}1.6%$). We next confirmed that the intracellular levels of reactive oxygen species (ROS) were significantly decreased (p<0.01) in P. amurense and/or H. japonicus extract treated groups when compared with the control group. Our results also showed that expression of cleaved caspase-3 and apoptotic cells of blastocysts were significantly decreased (p<0.05) in bovine blastocysts derived from both P. amurense and H. japonicus extract treated embryos. Conclusion: These results suggest that proper treatment with P. amurense and H. japonicus extracts in the development of preimplantation bovine embryos improves the quality of blastocysts, which may be related to the reduction of ROS level and apoptosis.

• 대한한의학방제학회지
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• 제25권4호
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• pp.509-526
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• 2017

Background and objectives : Soeumin Bojungykgi-tang (seBYTE) has been used to supplement qi in Korean medicine. It has been demonstrated to possess various biological functions such as anti-cancer, anti-aging and anti-inflammatory effects. The present study evaluated the protective roles of seBYTE in hepatotoxic in vitro and in vivo model. Methods : To investigate cytoprotective effect of seBYTE, HepG2 cells were pretreated with seBYTE and then subsequently exposed to $10{\mu}m$ AA for 12 h, followed by $5{\mu}m$ iron. Cell viability was examined by MTT assay, and expression of apoptosis-related proteins was evaluated by immunoblot analysis. For responsible molecular mechanisms, ROS production, GSH contents, and mitochondrial membrane potential were measured. In addition, hepatoprotective effect of seBYTE in vivo was assessed in $CCl_4$-induced animal model. Results : seBYTE prevented AA + iron-induced cytotoxicity in concentration dependent manner. In addition, ROS production, GSH depletion, and mitochondrial dysfunction induced by AA + iron were significantly reduced by seBYTE pretreatment. Furthermore, seBYTE recovered expression of the pro-apoptotic proteins such as PARP and pro-caspase-3. In animal experiment, plasma ALT and AST levels were significantly elevated in $CCl_4$ treatment, but seBYTE significantly decreased the ALT and AST levels. Moreover, seBYTE alleviated the numbers of histological activity index, percentages of degenerative regions, degenerated hepatocytes, infiltrated inflammatory cells, nitrotyrosine- and 4-hydroxynonenal-positive cells in liver. Conclusions : These results showed that hepatoprotective effect of seBYTE against on $CCl_4$-induced hepatic damages is partly due to antioxidative and anti-apoptotic process.

• 대한한방내과학회지
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• 제26권1호
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• pp.182-198
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• 2005

Object : This study was designed to research whether the protection and inhibitory effects of cardiovascular diseases in L-NAME induced rat or ECV 304 cell lines through the Cell morphological pattern, Tunel assay, LDH activity, heart rate, blood pressure and immunohistochemistric analysis by Boonsimgieum water extract Methods : Nitric oxide(NO) play an important role in normal and pathophysiological cells including as a messenger molecule, neurotransmitter, microbiocidal agent, or dilator of blood vessels and artheriosclerosis, hypertension, myocardial infarction, respectively. Endothelial cell products can modulate the magnitude of a response to a vasoconstrictor, as evinced by the greater constriction after endothelium removal or NO synthesis blockade. To investigate that Boonsimgieum in the potential contribution of the levels of nitric oxide generated by endothelial nitric oxide synthase (eNOS) and the mechanisms of protection against NG-nitro-L-arginine methyl ester (L-NAME), human ECV 304 cells, which normally do not express eNOS, were expressed by L-NAME. L-NAME stimulated rat or cells were found to be resistant to injury and delayed death following the Boonsimgieum. Inhibition of nitric oxide synthesis abolished the protective effect against L-NAME, thrombin and collagen exposure. Interestingly, such effects have been observed during stimulation with agents such as phenylephrine and KCl on L-NAME mediate rats, were damaged by the NOS inhibitor L-NAME. Result : As the result of this study, In group, the anti-apoptosis and necrosis in the cardiovascular system have a potential capacity for prevented, protected and treating the diseases of cardiovascular system, against the necrosis of rat and ECV 304 cells with Caspase 3 and calpain expression by L-NAME is promoted. Conclusion : these results demonstrate neuroprotective and memory enhancing effects of ZIBU, suggesting its beneficial actions for the treatment of AD.

• Journal of Ginseng Research
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• 제44권4호
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• pp.664-671
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• 2020

Background: Ginsenoside compound-Mc1 (Mc1) is a member of the deglycosylated ginsenosides obtained from ginseng extract. Although several ginsenosides have a cardioprotective effect, this has not been demonstrated in ginsenoside Mc1. Methods: We treated H9c2 cells with hydrogen peroxide (H₂O₂) and ginsenoside Mc1 to evaluate the antioxidant effects of Mc1. The levels of antioxidant molecules, catalase, and superoxide dismutase 2 (SOD2) were measured, and cell viability was determined using the Bcl2-associated X protein (Bax):B-cell lymphoma-extra large ratio, a cytotoxicity assay, and flow cytometry. We generated mice with high-fat diet (HFD)-induced obesity using ginsenoside Mc1 and assessed their heart tissues to evaluate the antioxidant effect and the fibrosis-reducing capability of ginsenoside Mc1. Results: Ginsenoside Mc1 significantly increased the level of phosphorylated AMP-activated protein kinase (AMPK) in the H9c2 cells. The expression levels of catalase and SOD2 increased significantly after treatment with ginsenoside Mc1, resulting in a decrease in the production of H₂O₂-mediated reactive oxygen species. Treatment with ginsenoside Mc1 also significantly reduced the H₂O₂-mediated elevation of the Bax:Bcl2 ratio and the number of DNA-damaged cells, which was significantly attenuated by treatment with an AMPK inhibitor. Consistent with the in vitro data, ginsenoside Mc1 upregulated the levels of catalase and SOD2 and decreased the Bax:B-cell lymphoma-extra large ratio and caspase-3 activity in the heart tissues of HFD-induced obese mice, resulting in reduced collagen deposition. Conclusion: Ginsenoside Mc1 decreases oxidative stress and increases cell viability in H9c2 cells and the heart tissue isolated from HFD-fed mice via an AMPK-dependent mechanism, suggesting its potential as a novel therapeutic agent for oxidative stress-related cardiac diseases.

• Animal Bioscience
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• 제34권4호
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• pp.546-557
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• 2021

Objective: If fertilization does not occur within a specific period, the quality of unfertilized oocytes in the oviduct (in vivo aging) or in culture (in vitro aging) will deteriorate over time. Icariin (ICA), found in all species of Epimedium herbs, has strong antioxidant activity, and is thought to exert anti-aging effects in vitro. We asked whether ICA protects oocytes against age-related changes in vitro. Methods: We analyzed the reactive oxygen species (ROS) levels and expression of antioxidant, maternal, and estrogen receptor genes, and along with spindle morphology, and the developmental competence and quality of embryos in the presence and absence of ICA. Results: Treatment with 5 μM ICA (ICA-5) led to a significant reduction in ROS activity, but increased mRNA expression of glutathione and antioxidant genes (superoxide dismutase 1 [SOD1], SOD2, peroxiredoxin 5, and nuclear factor erythroid 2-like 2), during aging in vitro. In addition, ICA-5 prevented defects in spindle formation and chromosomal alignment, and increased mRNA expression of cytoplasmic maturation factor genes (bone morphogenetic protein 15, cyclin B1, MOS proto-oncogene, serine/threonine kinase, and growth differentiation factor-9). It also prevented apoptosis, increased mRNA expression of antiapoptotic genes (BCL2-like 1 and baculoviral IAP repeat-containing 5), and reduced mRNA expression of pro-apoptotic genes (BCL2 antagonist/killer 1 and activation of caspase-3). Although the maturation and cleavage rates were similar in all groups, the total cell number per blastocyst and the percentage of apoptotic cells at the blastocyst stage were higher and lower, respectively, in the control and ICA-5 groups than in the aging group. Conclusion: ICA protects oocytes against damage during aging in vitro; therefore, it can be used to improve assisted reproductive technologies.

• 대한한의학방제학회지
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• 제31권4호
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• pp.231-243
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• 2023

Objectives : Jasminum officinale L. var. grandiflorum is used as a traditional or folk remedy in China to treat arthritis, hepatitis, duodenitis, conjunctivitis, gastritis, and diarrhea. In this study, we aimed to study the hepatocyte protective activity and molecular mechanism of Jasminum officinale L. var. grandiflorum aqueous extract (JGW) using HepG2 hepatocyte cell lines. Methods : HepG2 cells were pretreated with diverse concentrations of JGW, and then the cells were exposed to tert-butyl hydroperoxide (tBHP) for inducing oxidative stress. Hydrogen peroxide (H₂O₂) production, glutathione (GSH) concentration, mitochondrial membrane potential (MMP) and cell viability were measured to investigate hepato-protective effects of JGW. Phosphorylation of AMP-activated protein kinases (AMPK), acetyl coenzyme A carboxylase (ACC) and effects of compound C on cell viability were examined to observe the role of AMPK on JGW-mediated cytoprotection. Results : Pretreatment with JGW (10-300 ㎍/mL) significantly suppressed cytotoxicity induced by tBHP in a concentration dependent manner and reduced the expression of cleaved PARP and cleaved caspase-3 proteins related to apoptosis in HepG2 cells. In addition, pretreatment with JGW significantly prevented the increase in H₂O₂ production, GSH depletion, and lower MMP induced by tBHP. Treatment with JGW (30 minutes of incubation and concentrations of 100 and 300 ㎍/mL) increased the phosphorylation of AMPK and ACC and treatment with compound C, a chemical inhibitor of AMPK, inhibited the cytoprotective effect of JGW. Conclusions : Our results demonstrated that JGW may protect hepatocytes from oxidative stress via activation of AMPK.

• 한국식품과학회지
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• 제53권4호
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• pp.423-433
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• 2021

본 연구에서는 두충 잎의 분획물을 이용하여 미세먼지(PM_2.5)로 유도한 in vitro 뇌 신경세포 독성에 대한 항산화 활성 및 세포보호 효과를 확인하였다. EFEL은 우수한 ABTS 및 DPPH 라디칼 소거능을 나타냈으며, 지질과산화물 억제활성에서 양성대조군인 catechin과 동일한 농도에서 유사한 활성을 나타냈다. 또한, EFEL은 미세먼지로 유도된 해마세포(HT22), 뇌신경세포(MC-IXC) 및 미세아교세포(BV-2) 내의 ROS 수준을 효과적으로 감소시켜주었으며, 세포 생존능을 증가시킴으로써 세포보호 효과를 나타냈다. 미세먼지로 활성화된 BV-2에서 EFEL은 세포 활성화와 관련된 세포막 수용체인 TLR4와 p-JNK의 발현을 감소시켰다. 또한, 세포 사멸에 관여하는 caspase-3 활성화 전 단계인 procaspase-3와 세포 생존 경로에 관여하는 p-Akt 발현의 증가된 수준을 나타냈며, 염증성 사이토카인으로써 증가된 TNF-α의 수준을 감소시켰다. EFEL의 생리활성 물질을 HPLC로 분석한 결과, rutin과 chlorogenic acid가 확인되었다. 결과적으로, 두충 잎의 아세트산에틸 분획물은 우수한 페놀성 화합물 및 플라보노이드 함량을 나타냄으로써 높은 항산화능을 보였으며, 미세먼지로 유도된 산화적 스트레스에서 사멸 및 염증반응을 조절하여 뇌 신경세포를 보호하였다. 이를 고려할 때, EFEL은 미세먼지로 유도된 염증성 뇌신경세포 관련 질환 예방에 도움을 줄 수 있는 건강기능식품 소재로 활용될 수 있을 것으로 판단된다.

• 한국식품과학회지
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• 제51권4호
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• pp.379-392
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• 2019

본 연구에서는 갑주백목 에탄올 추출물(ethanolic extraction from Diospyros kaki (Gabjubaekmok))을 이용하여 in vitro 항산화 활성과 더불어 $A{\beta}$로 유도된 인지기능 저하를 갖는 마우스 모델에서 인지기능 및 뇌 신경세포 보호 효과를 검증하였다. 갑주백목 추출물은 양성대조군과 비교하여 우수한 ABTS 및 DPPH 라디칼 소거활성($IC_{50}=83.30$, $403.88{\mu}g/mL$) 및 MDA 생성 억제 활성($IC_{50}=62.10{\mu}g/mL$)을 보여주었고, in vitro 아세틸콜린 분해효소 억제 활성은 $312.82{\mu}g/mL$의 $IC_{50}$ 값을 나타내었다. 또한, MC-IXC 뇌 신경세포에 과산화수소를 처리하여 산화적 스트레스를 유발시킨 뇌 신경세포 사멸에 대해 보호 효과를 나타냈다. 갑주 백목 추출물의 인지기능 개선 효과를 확인하기 위하여 $A{\beta}$를 사용하여 인지기능 장애 마우스 모델을 수립하였으며, Y-미로, 수동 회피 및 Morris 수중 미로 실험과 같은 행동실험을 통해 인지 및 기억능력에 대한 개선 효과를 나타냈다. 이후 마우스 뇌조직에서의 아세틸콜린 함량의 증가 및 아세틸콜린 분해효소의 활성을 억제함으로써 cholinergic 시스템을 보호하였고, SOD, 환원형 GSH 및 MDA 함량 측정을 통해 항산화 시스템을 개선시켜줌을 확인하였다. 더불어 뇌 조직의 미토콘드리아에서 ROS의 생성 억제, MMP 보호 및 ATP 함량을 회복시켜주었으며, western blot 분석을 통해 $TNF-{\alpha}$와 같은 염증성 사이토카인을 억제시켜 줌으로써 면역반응에 관여하는 JNK의 인산화를 감소시키고 Akt 신호전달을 활성화시켜 세포자동 사멸화를 억제시키는 것으로 확인되었다. 마지막으로 HPLC 분석을 통해서 갑주백목의 주요 생리활성물질이 갈산으로 확인되었다. 이러한 결과를 종합하였을 때, 갑주백목 에탄올 추출물은 뇌 조직에서의 cholinergic 및 항산화 시스템 보호효과를 통해 $A{\beta}$ 처리에 대하여 학습 및 기억 능력을 개선 시킬 수 있는 천연 소재로서의 가능성뿐만 아니라 $A{\beta}$ 및 과산화수소로부터 유발된 산화적 스트레스의 환경에서 뇌신경세포를 보호함으로써 알츠하이머성 질환과 같은 퇴행성 뇌신경질환을 예방할 수 있는 고부가가치 건강기능식품 소재로서의 활용 가능성을 확인하였다.

• 한국미생물·생명공학회지
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• 제29권3호
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• pp.181-185
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• 2001

유방암 세포주에서는 우수한 항암활성을 가진 것으로 알려진 indole-3-carbinol을 HepG2세포주에 시간과 농도별로 처리한 결과 cell growth inhibition을 확인하였으며, $IC_{50}$ 값은 48시간배양에서 $446\mu$M 72시간 배양에서 444$\mu$M로 나타났다. $400\mu$M의 I3C을 투여하고, 24, 48, 72시간에 HePG2 세포주의 cell cycle pattern을 분석한 결과, G1 phase에서 P21의증가와 함께 Cdk 6와 cyclin D의 확연한 감소와 Pb protein의 hypo-phosphorylation을 확인하였다. 반면 G2 phase에서는 I3C의 직접적인 억제로 인해 24시간 후부터 Cdc2와 cyclin B1가 급격히 감소하는것을 확인하였다. Flow cytomery 분석결과 I3C 처리 24시간 뒤 G2 arrest (25%)가 발생하였으며, 72시간이 지난후 G1 arrest (53%)가 발생하였다. 이러한 I3C의 간암세포주인 HePG2 cell의 cell cycle arrest가 apoptosis를 유발하는지를 알고자 caspase 3 Bcl2 Bax protein의 발현양상을 확인한 결과 아무런 변화가 보이지 않았다. 즉 I3C은 간암세포주인 HepG2 cell에서 apoptosis를 유도하지 못한다는것을 확인하였따. 결론적으로 I3C은 HepG2 세포주에서 G1와 G2 phase에서 cell cycle arrest는 발생시키나, 특이적으로 apoptosis 와는 연관되지 않는다는 사실을 확인하였다.

• 생명과학회지
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• 제27권9호
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• pp.1020-1030
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• 2017

대장암은 미국 등 서양 국가뿐만 아니라 국내에서도 2번째로 많이 발병이 되는 암으로 알려져 있다. 역학조사에 의하면 오메가-3를 많이 섭취한 인종에서 대장암 발생빈도가 감소하고 최근 오메가-3는 수종의 암에 대해 항암작용을 나타낸다고 한다. 이에 본 연구에서는 대장암에서 DHA의 항침윤, 항혈관 신생 및 항종양 형성능 억제의 기전을 규명하여 다음과 같은 결과를 얻었다. DHA는 인체 대장암 세포주 HT29 의 증식을 농도 의존적으로 억제하였으나 AA는 거의 영향이 없었다. FACS 분석에서 DHA 처리했을 때 Sub G1 phase의 세포가 DHA의 농도 의존적으로 증가 하였다. DHA 처리 후 cleaved PARP가 증가하고, uncelaved caspase-3가 감소 하였다. HT29 세포의 침윤능은 DHA 처리에 의해 억제 되었다. DHA 처리 후 MMP-9 및 MMP-2 mRNA양이 감소 되었을 뿐만 아니라 그 promoter의 reporter 활성도 억제하였으며 VEGF promoter 활성도 DHA에 의해 억제 되었다. NF-kB promoter 활성 및 핵으로의 이동도 DHA에 의해 억제 되었다. In vivo 동물실험에서 생쥐 대장암 세포주인 MCA38에 대한 Fat-1 transgenic mice에서의 종양 형성능은 현저히 억제 되었다. 면역형광염색법을 이용한 Fat-1 transgenic mice의 종양 조직에서의 TUNEL 양성세포는 wild type mice에 비해 현저히 증가하였으나 CD31의 형광강도는 감소 되었다. 이상의 결과로 오메가-3는 대장암 세포에서 NF-kB 억제에 따른 COX-2, MMP-2 및 MMP-9 등 matrix matalloproteinase의 억제를 통한 침윤능의 억제, VEGF 억제를 통한 혈관신생의 억제등 복합적 기전에 의해 항암작용을 나타내리라 생각되며, 따라서 오메가-3는 대장암의 예방 및 치료에 유용하게 사용될 수 있으리라 생각된다.