• 대한핵의학회지
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• 제38권4호
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• pp.294-299
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• 2004

목적 : 현재 생체 내로 이식된 세포를 추적하는데 여러 가지 리포터 유전자들이 이용되고 있다. 이 연구에서는 사람 나트륨 옥소 공동 수송체 (hNIS)와 도파민 2 수용체($D_2R$)를 이중 리포터 유전자로 사용하여 각각을 비교하였다. 대상 및 방법: hNIS와 $D_2R$의 발현이 동시에 이루어지도록 하기 위해서 IRES (Internal ribosome entry site)로 연결된 재조합 플라스미드(pIRES-hNIS/D_2R)를 제조하였다. $pIRES-hNIS/D_2R$를 사람의 간암세포주인 SK-Hep1에 lipofactamine을 이용하여 형질을 도입시킨 후, 항생제(G418)를 농도별로 처리하여 2주간 선별하였다(HEP-ND). hNIS와 $D_2R$발현 유무와 발현 정도를 알아보기 위하여 각 유전자에 특이적인 프라이머를 이용하여 RT-PCR을 수행하였다. 각 형질 도입세포군에서, hNIS의 활성은 $^{125}I$ 섭취율을 이용하여 측정하고 $D_2R$의 활성은 $[^3H]spiperone$을 리간드로 이용하여 수용체 결합 정도를 측정하였다. 결과: 선별된 HEP-ND세포에서 hNIS와 $D_2R$의 발현을 RT-PCR로 확인하였을 때 IRES로 연결된 hNIS와 $D_2R$의 발현 정도는 서로 비슷하였다. HEP-ND세포의 $^{125}I$ 섭취율은 대조군인 SK-Hep1세포에 배해 30-40배 증가되었고, $KClO_4$에 의해 $^{125}I$ 섭취가 저해되었다. $D_2R$의 발현 정도를 측정할 수 있는 수용체 결합 분석법을 통해G418 농도별로 나눈 두 종류의 세포주에서, $[^3H]spiperone$을 이용한 해리상수 ($K_d$)와 최대결합 부위농도 ($B_{max}$)는 각각 2.92 nM, 745.25 fmol/mg protein과 8.91nM, 1323 fmole/mg protein이었다. hNIS와 $D_2R$발현의 상관관계에서는 높은 상관관계를 나타내었다. 결론: 이 연구에서 hNIS와 $D_2R$가 이입된 세포주에서 이중 유전자, 감마 영상 리포터 시스템을 개발하였으며, $D_2R$와 HNIS 유전자를 이중 핵 영상 시스템으로서 서로 상호보완적으로 이용할 수 있을 것으로 기대하고 있다.

• 한국정밀공학회지
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• 제31권8호
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• pp.743-747
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• 2014

In this study, drop-on-demand system using piezo-elecrtric inkjet printers was employed for preparation of collagen microspheres, and its application was made to the HepG2 cell-laden microsphere preparation. The collagen microspheres were injected into beaker filled with mineral oil and incubated in a water bath at $37^{\circ}C$ for 45 minutes to induce gelation of the collagen microsphere. The size of collagen microsphere was $100{\mu}m$ in diameter and $80{\mu}m$ in height showing spherical shape. HepG2 cells were encapsulated in the collagen microsphere. The cell-laden microspheres were inspected by the microscopic images. The encapsulation of cells may be beneficial for applications ranging from tissue engineering to cell-based diagnostic assays.

• Journal of Nutrition and Health
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• 제44권6호
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• pp.488-497
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• 2011

Oxygen is necessary to sustain life, yet cellular oxygen metabolism creates destructive elements called free radicals. Free radicals are chemically unbalanced and carrying free electrons that can damage molecules, potentially damaging the cell itself. For this reason, many antioxidant products, including supplements and functional foods, are being developed. In particular, natural products are rich sources of pharmacologically active compounds. The purpose of this study was to investigate the antioxidant effects of target biomaterials in Korean traditional spices such as diallyl sulfide (DAS), capsaicin (CAP), and gingerol (GGR), and to investigate the response of the antioxidant defense system to oxidative stress by hydrogen peroxide ($H_2O_2$) compared to sulforaphane (SFN) in HepG2 cells. After the analysis of the cell viability using Cell Counting kit-8 (CCK-8) assay, we determined that the optimum levels were $200{\mu}M$ DAS, $25{\mu}M$ CAP, $50{\mu}M$ GGR, and $12.5{\mu}M$ SFN. Antioxidant enzymes were measured and protein expression was detected by Western blotting. All treatments showed a significant decrease in antioxidant enzyme activity such as superoxide dismutase, catalse, and glutathione peroxidase in HepG2 cells. Additionally, DAS, CAP, GGR and SFN increased the antioxidant system-related transcription factor Nrf2 which was found to be regulated by the activation of MAPK-JNK in this study. In conclusion, these results indicate the protective effects of DAS CAP, GGR, and SFN against $H_2O_2$-induced oxidative stress.

• Archives of Pharmacal Research
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• 제27권4호
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• pp.402-406
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• 2004

This research team found in previous studies, that the ginseng saponin metabolite IH901 induces apoptosis in HepG2 cells via a mitochondrial-mediated pathway, which resulted in the activation of caspase-9 and subsequently of caspase-3 and -8. Based on these results, the involvement of the Fas/Fas ligand (FasL) death-receptor pathway, in IH901-induced apoptosis in HepG2 cells, was investigated. Levels of Fas and the Fas ligand (FasL) mRNA or protein were not increased by IH901, rather they were decreased significantly at 18 h post treatment. Soluble FasL (sFasL) was detectable by immunoprecipitation analysis En the medium of HepG2 cells treated with IH901. Increased levels of sFasL were inversely correlated with the levels of FasL. Preincubation of HepG2 cells with antagonistic anti-Fas antibody showed little protective effect, if any, on IH901-induced cell death. At a $30{\mu}M$ (24 and 48 h) and $40{\mu}M$ (24 h) concentration of IH901, the cytotoxic effect of IH901 was less then $50\%$, anti-Fas antibody prevented IH901-induced cell death. However, at a $60{\mu}M$ (24 and 48 h) and $40{\mu}M$ (48 h) concentration of IH901, cell death rates were about $80\%$ or more and most of the chemopreventive and chemotherapeutic effects of IH901 were manifested. Blocking the Fas receptor did not influence IH901-induced cell death. These results indicate that the Fas/FasL system is engaged, but not required for IH901-induced cell death, at pharmacologically significant concentrations.

• Journal of Microbiology and Biotechnology
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• 제21권9호
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• pp.903-913
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• 2011

Vibrio parahaemolyticus, which causes gastroenteritis, wound infection, and septicemia, has two sets of type III secretion systems (TTSS), TTSS1 and TTSS2. A TTSS1-deficient vcrD1 mutant of V. parahaemolyticus showed an attenuated cytotoxicity against HEp-2 cells, and a significant reduction in mouse lethality, which were both restored by complementation with the intact vcrD1 gene. V. parahaemolyticus also triggered phosphorylation of mitogen-activated protein kinases (MAPKs) including p38 and ERK1/2 in HEp-2 cells. The ability to activate p38 and ERK1/2 was significantly affected in a TTSS1-deficient vcrD1 mutant. Experiments using MAPK inhibitors showed that p38 and ERK1/2 MAPKs are involved in V. parahaemolyticus-induced death of HEp-2 cells. In addition, caspase-3 and caspase-9 were processed into active forms in V. parahaemolyticus-exposed HEp-2 cells, but activation of caspases was not essential for V. parahaemolyticus-induced death of HEp-2 cells, as shown by both annexin V staining and lactate dehydrogenase release assays. We conclude that secreted protein(s) of TTSS1 play an important role in activation of p38 and ERK1/2 in HEp-2 cells that eventually leads to cell death via a caspase-independent mechanism.

• 생명과학회지
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• 제27권11호
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• pp.1340-1348
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• 2017

Sanguinarine은 다양한 목적으로 사용되고 있는 Sanguinaria canadensis L.의 뿌리에서 유래된 benzophenanthridine alkaloid 계열 물질중의 하나이다. 그동안 sanguinarine의 다양한 약리학적인 효능이 알려져 왔고, 항암활성에 대한 연구도 여러 암세포들을 대상으로 수행되어 왔다. 그러나 sanguinarine에 의한 암세포의 apoptosis 유도에 대한 현상은 여전히 많은 부분에서 연구의 대상으로 남아 있다. 본 연구는 Hep3B 인체 간암세포를 대상으로 sanguinarine의 항암활성에 대한 추가적인 자료를 제시하기 위하여 수행되었다. 본 논문의 결과에 의하면, sanguinarine은 처리 농도 의존적으로 Hep3B 세포의 증식을 억제하였으며, 이는 apoptosis 유도와 연관성이 있었다. Sanguinarine은 두 가지 apoptosis 경로인 extrinsic 및 intrinsic 경로의 개시 initiator caspase인 caspase-8 및 caspase-9 뿐만 아니라 대표적인 effector caspase인 caspase-3의 활성을 증가시켰고, caspase-3의 기질인 PARP의 분절을 유발하였다. 아울러 sanguinarine은 DR-related 유전자들의 발현을 부분적으로 증가시켰으며, Bcl-2 family에 속하는 pro-apoptotic Bax의 발현을 증가시킨 반면, anti-apoptotic Bcl-2의 발현은 억제시켰다. 또한 sanguinarine은 Bid의 truncation을 촉진하였고, MMP의 소실에 따른 cytochrome c를 미토콘드리아에서 세포질로의 이동을 증가시켰다. 그리고 sanguinarine에 의한 apoptosis 유도 및 세포 증식율 억제 현상이 caspase의 활성을 인위적으로 억제하였을 경우, 모두 사라졌다. 따라서 sanguinarine에 의하여 유도하는 Hep3B 세포의 apoptosis 유발에는 caspase 의존적으로 extrinsic 및 intrinsic 경로가 모두 관여하고 있음을 알 수 있었다.

• 약학회지
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• 제59권5호
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• pp.201-206
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• 2015

The aim of the present study was to optimize in vitro method for the prediction of drug-induced liver injury using human liver microsomes (HLM). Cytotoxicity test of cyclophosphamide and acetaminophen in HepG2 cells cultured with HLM showed that the newly established condition using 0.375 mg/ml HLM for 24 hr incubation was comparable or more sensitive than the previously established condition using 0.75 mg/ml HLM for 12 hr incubation. Although the cytotoxic effect of troglitazone was completely attenuated by 0.75 mg/ml HLM, it was augmented by 0.375 mg/ml HLM in the presence of the NADPH-generating system. The cytotoxic effect of chlormezanone, a withdrawn drug due to hepatotoxicity in human, was increased by HLM in the presence of the NADPH-generating system. In contrast, the cytotoxic effect of methapyrilene, a withdrawn drug due to hepatotoxicity in rats, was decreased by HLM in the presence of the NADPH-generating system. The present study suggests that the optimized in vitro method using HLM can be useful for the prediction of drug-induced hepatotoxicity.

• 한국철도학회논문집
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• 제13권6호
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• pp.559-564
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• 2010

본 논문은 동력 집중식 철도차량에서 전력변환장치로서 객차용 전원공급장치의 전력품질을 향상시키기 위해 보완된 필터설계를 제안한다. 먼저, 필터설계를 위해 객차용 전원공급장치와 연결된 변압기의 출력전력 파형을 측정하였다. 실험 및 시뮬레이션 결과를 통해 스위칭 기법은 개선된 특정고조파 소거 펄스폭 변조이고 스위칭 주파수는 300Hz임을 추정하였다. 따라서 추정된 파라미터를 바탕으로 전력품질을 향상시키기 위해 필터를 설계하였다. 시뮬레이션 결과 전체적으로 고조파의 크기가 감소함을 확인하였다.

• 생명과학회지
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• 제16권6호
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• pp.919-924
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• 2006

다양한 독성물질 및 이들의 대사산물의 효율적인 생물독성 평가시스템이 지속적으로 개발되고 있다. 본 연구에서는 대분비계 교란물질인 endosulfan, bisphenol A, vinclozolin 및 3,5-dichloroaniline을 대상으로 인간 간암세포주, 지렁이, 효모를 이용한 세포독성 및 성장억제 효과를 평가하였다. 인간 간암세포주 독성평가에서는 endosulfan, 3,5-dichloroaniline, bisphenol A의 순으로 독성이 나타났으며, 지렁이 독성평가에서는 endosulfan, bisphenol A, 3,5-dichloroaniline의 순으로 독성이 나타났다. 효모를 이용한 독성평가에서는 3,5-dichloroaniline, endosulfan, bisphenol A의 순으로 독성이 나타나 다른 시스템과는 부분적인 차이가 나타났으며, vinclozolin의 경우 3가지 독성평가 시스템에서 모두 독성이 나타나지 않았다. 이러한 결과는, 동일한 물질을 서로 다른 생물 독성평가 시스템을 사용하여 평가하는 경우, 부분적인 오류가 나타날 수 있음을 암시하고 있으며, 독성 유무 판단은 가능하더라도, 독성 정량평가 및 독성 정도를 비교하는 것은 어렵다는 것을 제시하고 있다. 또한 본 결과는, 다양한 물질 및 이들의 대사산물의 일차적 독성평가에는 지렁이 및 효모시스템이 빠르고 경제적임을 암시하고 있으며, 독성이 인정될 경우 인간세포주 및 동물시험에 의한 검증이 효율적이라고 판단된다.

• 대한한의학방제학회지
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• 제16권1호
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• pp.117-130
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• 2008

Korean traditional medicine has been used for the treatment of the various diseases based on both oriental medicinal theory and clinical trials. Thus, the prescriptions of Korean traditional medicine would be useful for the development of new therapeutics. This research focuses on the fundamental study in Korean traditional prescriptions for the development of new hepatoprotective agents. We found two prescriptions. Injinho-Tang and Osumogwa-Tang, showed the significant DPPH free radical scavenging and hepatoprotective effect, respectively. It is well-known that free radical scavenging effect is related to the prevention of various pathological events including liver injury. This paper deals with hepatoprotective effects on tacrine-induced cytotoxicity in Hep G2 cells, free radicals scavenging on both DPPH and superoxide of above two prescriptions. Hot water extract of Injinho-Tang did not show the significant hepatoprotective effect on tacrine-induced cytotoxicity in Hep G2 cells, however, it shows the significant scavenging effects for both DPPH and superoxide radicals. On the other hand, all of the hot water extracts of constituent herbal drugs in Injinho-Tang exhibited the promising protective effect on tacrine-induced cytotoxicity in Hep G2 cells. Of these, water extract of Rhei Rhizoma showed the most prominent effect on tacrine-induced cytotoxicity in Hep G2 cells. Bioassay-guided fractionation of Rhei Rhizoma extract has furnished four compounds, and their chemical structures have been identified by comparison of their spectral data with those of literature as chrysophanol (1), emodin (2), 3,5-dihydroxy-4'- methoxystilbene (3), and rhapontigenin (4), respectively. Among the isolated compounds, compounds 2-4 revealed the significant hepatoprotective effect in vitro when their $EC_{50}$ values compare with that of silybin, as a positive control. It also exhibited that emodin possessed the most hepatoprotective effect among these active compounds. In case of Osumogwa-Tang, its hot water extract showed the moderate protective effect on tacrine-induced cytotoxicity in Hep G2 cells. Hot water extract of Chaenomelis Fructus, one of the constituent herbal drug of this prescription, exhibited the significant hepatoprotective effect with $EC_{50}$ value of $7.8{\pm}0.1\;{\mu}g/ml$, however, it showed strong cytotoxicity in Hep G2 cells above the concentration of $25\;{\mu}g/ml$. It was revealed that both hot water extract of Evodiae Fructus and its butanol soluble fraction showed the moderate hepatoprotective effect but concentration-dependent activity in Hep G2 assay system. Two quinolone alkaloids, evocarpine and dihydroevocarpine, also tested for their hepatoprotective effects on tacrine-induced cytotoxicity in Hep G2 cells, however, these two compounds derived from the Evodiae Fructus as the major constituents did not show in vitro hepatoprotective effect. From these results, it would be necessary to further isolation of its hepatoprotective compounds from the butanol soluble fraction of the hot water extract of Evodiae Fructus.