• 한국식품과학회지
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• 제46권5호
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• pp.609-615
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• 2014

국내에 충분히 보고되지 못한 소재로서 drumstick-tree (Moringa Oleifera Lam.)를 고부가가치 식품 자원으로서 그 활용가능성을 알아보기 위해 주요 영양성분 분식 및 in vitro 신경세포 보호효과에 대해서 연구하였다. Drumstick-tree의 주요 무기성분으로는 칼슘으로 2658.67 mg/100 g이 함유되어 있었고, 다음으로 칼륨, 마그네슘 및 인 등이 함유되어 있었다. 주요 지방산으로서 포화 지방산으로는 palmitic acid (16.33%)와 불포화 지방산으로서 gadoleic acid (66.34%)가 상대적으로 많이 함유되어 있었고, 지용성 비타민인 vitamin E가 94.78 mg/100 g 그리고 niacin이 112.61 mg/100 g 함유되어 있는 것을 알 수 있었다. 80% methanol에 추출한 drum-stick-tree 추출물을 활용하여 $H_2O_2$ 처리한 PC12 cell 내의 활성산소 생성억제효과를 DCF-DA assay를 통해 측정한 결과 drumstick-tree 추출물은 농도의존적인 활성산소 생성 억제효과를 보였다. MTT assay를 이용하여 $H_2O_2$로 유도된 PC12 신경세포에 대한 보호효과를 측정한 결과 vitamin C group 대비 효과적인 신경세포 보호효과를 확인하였고, LDH release assay를 통해 일정 수준의 세포막 보호효과를 역시 확인하였다. 또한 PC12 cell의 oxidative stress-induced apoptosis에 대한 세포 보호효과를 측정하기 위한 caspase assay 실험 결과, 세포 내 caspase activity가 추출물의 의해 효과적으로 감소됨을 알 수 있었다. 결국 본 연구결과를 종합해 볼 때, 우수한 영양 구성 성분과 함께 신경세포 내 oxidative stress의 저감화 등을 통한 drumstick-tree 추출물의 신경 세포 보호 효과는 고부가가치 천연 소재로서의 다양한 산업적 활용 가능성을 암시하는 것으로 판단된다.

• Applied Biological Chemistry
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• 제50권3호
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• pp.228-232
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• 2007

본 연구는 만형자(Vitex rotundifolia)의 세포독성을 확인하기 위한 목적으로 methanol, ethanol, acetone으로 추출한 뒤, 인간 유래의 대장암 세포주인 HT-29에 처리하여 MTT reduction assay를 이용하여 실험을 하였다. 그 결과, 모든 만형자 추출물에서 농도 의존적으로 성장 저해 효과가 나타났으며, 그 중에서도 acetone 추출물을 50 ${\mu}g/ml$ 농도로 처리 시 18%로 가장 높은 항암 효과를 보였다. 또한 각 용매별 추출물을 50 ${\mu}g/ml$를 처리하여 HT-29 세포주의 성장을 관찰한 결과, acetone 추출물은 대조구에 비하여 암세포의 수축 등 형태변화가 두드러지게 나타났으며, Hoechst 33342 staining을 통하여 apoptotic body가 형성된 것을 확인 하였다. 그리고 가장 활성이 좋은 acetone 추출물을 용매 분획하여 얻은 분획물 중, n-hexane 분획물에서 농도 의존적으로 세포독성을 나타냄을 알 수 있었다. 또한 만형자의 n-hexane 분획물은 다른 대장암 세포주 SW620에 대해서도 50 ${\mu}g/ml$ 농도에서 대조군에 비하여 7배 정도의 높은 세포독성을 나타내고 있음을 MTT reduction assay와 LDH release assay를 통하여 확인 할 수 있었다.

• 대한한방내과학회지
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• 제21권2호
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• pp.259-266
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• 2000

Objectives : The effects of aqueous extracts of Cortex ulmi pumilae (a traditional medicine for cancer treatment in oriental medicine) on the induction of apoptotic cell death were investigated in human liver origm hepatoma cell lines, HepG2. Methods : The death of HepG2 cells was markedly induced by the addition of extracts of Cortex ulmi pumilae in a dose-dependent manner. The apoptotic characteristic ladder pattern of DNA strand break was not observed in cell death of HepG2. In addition, it was not shown nucleus chromatin condensation and fragmentation under hoechst staining. However, by the using annexin V staining assay, externalizations of phosphatidylserine in HepG2 cell which were treated with Cortex ulmi pumilae extracts were detected in the early time (at 9 hr after extract treatment). Furthermore, LDH release was not detected in this early stage. Therefore, Cortex ulmi pumilae extracts-induced cell death of HepG2 cells is mediated by apoptotic death signal processes. Result : The activity of caspase 3-like proteases remained in a basal level in HepG2 cells which treated with the extract of Cordyceps sinensis. However, it was markedly increased in HepG2 cells which treated with two extracts of Cortex ulmi pumilae (C.U.P.-C, C.U.P.-K) which were differently extracted (respectively, 2.3 and 3.3 fold). On a while, the phosphotransferase activities of JNK1 was markedly induced in HepG2 cells which were treated with two extracts of Cortex ulmi pumilae. On the contrary, the activation of transcriptional activator, activating protein1(AP-1) and NF-kB were severely decreased by these two extracts of Cortex ulmi pumilae (C.U.P.-C, C.U.P.-K). In addition, antioxidants (GSH and NAC) and intracellular $Ca2^+$ level regulator (Bapta/AM and Thapsigargin) did not affect Cortex ulmi pumilae extracts-induced apoptotic death of HepG2 cells. Conclusions : In conclusion, our results suggest that two extracts of Cortex ulmi pumilae (C.U.P.-C, C.U.P.-K) induces the apoptotic death of human liver origin hepatoma HepG2 cells via activation of caspase 3-like proteases as well as JNK1, and inhibition of transcriptional activators, AP-1 and $NK-{\kappa}B$.

• 대한한방내과학회지
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• 제21권3호
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• pp.467-475
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• 2000

The water extracts of Jagamcho-tang has been used for treatment of arrhythmia and palpitation in oriental traditional medicine. Brain is provided with blood flow by heart. Jagamcho-tang has been studied on ischemia and infarction in heart. However, little is known about the mechanism by which the water extracts of Jagamcho-tang rescues brain cells from ischemic damages. To elucidate the protective mechanism on ischemic induced cytotoxicity, the effects of Jagamcho-tang on ischemia induced cytotoxicity and generation of nitric oxide(NO) are investigated in C6 glioma cells. Jagamcho-tang induce NO in a dose dependent manner up to 2.5mg/ml in C6 glioma cells. The pretreatment of Jagamcho-tang protect sodium nitroprusside(SNP) (2mM) induced cytotoxicity. This effect of Jagamcho-tang is mimicked by treatment by pretreatment of SNP($100{\mu}M$), an exogenous NO donor. NG-monomethyl-L-arginine($N^{G}MMA$), a specific inhibitor of nitric oxide synthase (NOS), significantly blocks the protective effects of Jagamcho-tang on cell toxicity by ischemia. In addition, lipopolysaccharide(LPS) and phorhol 12 myristate 13-acetate(PMA) treatment for 72h in C6 glial cells markedly induce NO, but treatment of the cells with the water extracts of Jagamcho-tang decrease nitrite formation in a dose dependent manner. In addition, LPS and PMA treatment for 72h induce severe cell death and LDH release into medium in C6 glial cells. However treatment of the cells with the water extracts of Jagamcho-tang dose not induce significant changes compare to control cells. Furthermore, the protective effects of the water extracts of Jagamcho-tang is mimicked by treatment of $N^{G}MMA$. Taken together, I suggest that the protective effects of the water extracts of Jagamcho-tang against ischemic brain damages may be mediated by regulation of iNOS during ischemic condition.

• 대한한방내과학회지
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• 제31권3호
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• pp.650-666
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• 2010

Objectives : In this study, the author tried to investigate whether wood vinegar produced from Morus alba (MA) significantly affects the increase in airway epithelial mucosubstances and hyperplasia of tracheal goblet cells of rats, and in vitro airway mucin secretion and PMA- or EGF- or TNF-alpha-induced MUC5AC mucin production / gene expression from human airway epithelial cells. Materials and Methods : For the in vivo experiment, the author induced hypersecretion of airway mucus and goblet cell hyperplasia by exposure of rats to SO2 over 3 weeks. Effect of orally-administered MA over 2 weeks on increase in airway epithelial mucosubstances from tracheal goblet cells of rats and hyperplasia of goblet cells were assessed using histopathological analysis after staining the epithelial tissue with alcian blue. For the in vitro experiment, confluent RTSE cells were chased for 30 min in the presence of MA to assess the effect of MA on mucin secretion by enzyme-linked immunosorbent assay (ELISA). Also, effects of MA on PMA- or EGF- or TNF-alpha-induced MUC5AC mucin production and gene expression from human airway epithelial cells (NCI-H292) were investigated. Confluent NCI-H292 cells were pretreated for 30 min in the presence of MA and treated with PMA (10 ng/ml), EGF (25 ng/ml) or TNF-alpha (0.2 nm) for 24 hrs, to assess both effects of MA on PMA- or EGF- or TNF-alpha-induced MUC5AC mucin production by enzyme-linked immunosorbent assay (ELISA) and gene expression by reverse transcription-polymerase chain reaction (RT-PCR). Possible cytotoxicities of MA in vitro were assessed by examining LDH release from RTSE cells and the rate of survival and proliferation of NCI-H292 cells. In vivo liver and kidney toxicities of MA were evaluated by measuring serum GOT/GPT activities and serum BUN/creatinine concentrations of rats after administering MA orally. Results : 1. MA decreased the amount of intraepithelial mucosubstances of rats exposed to sulfur dioxide inhalationally. 2. MA decreased in vitro mucin secretion from cultured RTSE cells. 3. MA significantly inhibited PMA-, EGF-, and TNF-alpha-induced MUC5AC mucin productions and the expression levels of MUC5AC mRNA from NCI-H292 cells. 4. MA did not show either in vitro or in vivo hepatic or renal toxicities. Conclusion : The results from this study suggests that MA can regulate the secretion, production and gene expression of airway mucin observed in diverse respiratory diseases accompanied by mucus hypersecretion and does not show in vivo toxicity to liver and kidney functions after oral administration. Effects of MA should be further studied using animal experimental models that simulate the diverse pathophysiology of respiratory diseases via future research.

• Journal of Microbiology and Biotechnology
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• 제26권4호
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• pp.790-798
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• 2016

Chlamydiae, obligate intracellular bacteria, are associated with a variety of human diseases. The chlamydial life cycle undergoes a biphasic development: replicative reticulate bodies (RBs) phase and infectious elementary bodies (EBs) phase. At the end of the chlamydial intracellular life cycle, EBs have to be released to the surrounded cells. Therefore, the interactions between Chlamydiae and cell death pathways could greatly influence the outcomes of Chlamydia infection. However, the underlying molecular mechanisms remain elusive. Here, we investigated host cell death after Chlamydia infection in vitro, in L929 cells, and showed that Chlamydia infection induces cell necrosis, as detected by the propidium iodide (PI)-Annexin V double-staining flow-cytometric assay and Lactate dehydrogenase (LDH) release assay. The production of reactive oxygen species (ROS), an important factor in induction of necrosis, was increased after Chlamydia infection, and inhibition of ROS with specific pharmacological inhibitors, diphenylene iodonium (DPI) or butylated hydroxyanisole (BHA), led to significant suppression of necrosis. Interestingly, live-cell imaging revealed that Chlamydia infection induced lysosome membrane permeabilization (LMP). When an inhibitor upstream of LMP, CA-074-Me, was added to cells, the production of ROS was reduced with concomitant inhibition of necrosis. Taken together, our results indicate that Chlamydia infection elicits the production of ROS, which is dependent on LMP at least partially, followed by induction of host-cell necrosis. To our best knowledge, this is the first live-cell-imaging observation of LMP post Chlamydia infection and report on the link of LMP to ROS to necrosis during Chlamydia infection.

• 대한약리학회지
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• 제27권2호
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• pp.197-205
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• 1991

Adriamycin (Doxorubicin HCl)의 혈관밖 유출에 따른 조직의 손상, 특히 피부괴양 및 괴사 기전을 규명하기 위한 연구의 일환으로 흰쥐 피부세포를 이용한 in vitro 실험에서 adriamycin에 의한 산소라디칼 생성 및 그와 관련된 세포독성 기전으로 지질과산화를 검토하였다. Adriamycin은 흰쥐 태자 피부의 배양세포에서 lactic dehydrogenase(LDH) 유리를 용량 및 시간 의존적으로 증가 시켰으며, NADPH 및 NADH 첨가 조건에서 $superoxide\;anion(O^-\;_2{\cdot})$ 생성을 현저히 증가시켰다. Adriamycin은 지질과산화 반응의 척도인 malondialdehyde(MDA) 생성을 역시 NADPH, NADH 존재하에서 용량의존적으로 증가시켰고, 산소라디칼 제거물질들인 superoxide dismutase (SOD), catalase 및 thiourea와 항산화물질인 butylated hydroxytoluene(BHT), ${\alpha}-tocopherol$은 MDA 생성증가를 현저히 억제하였다. 1, 3,-bis(2-chloroethyl)-1-nitrosourea(BCNU)를 처리하여 산화성 공격에 대한 방어기전의 하나인 glutahione 체계를 억제할 경우 adriamycin에 의한 MDA 생성은 더욱 현저히 증가하였고, 이는 역시 항산화 물질들에 의하여 억제되었다. 이상의 연구성적에서 adriamycin은 산소라디칼 생성의 증가와 그에 따른 지질과산화를 촉진하므로서 피부세포에 손상을 줄 것으로 사료되었다.

• 대한화장품학회:학술대회논문집
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• 대한화장품학회 2003년도 IFSCC Conference Proceeding Book I
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• pp.13-34
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• 2003

Fructose-1, 6-diphosphate (FOP), a glycolytic metabolite is reported to ameliorate inflammation and inhibit the nitric oxide production in murine macrophages stimulated with endotoxin. It is also reported that FOP has cytoprotective effects against hypoxia or ischemia/reperfusion injury in brain and heart. In this study, we examined whether FDP has protective effects on UV-induced oxidative damage in skin cell culture system and human skin in vivo. FDP had a protective role in UVB-induced LDH release and ROS accumulation in HaCaT although it did not show direct radical scavenging effect in the experiment using 1, 1-diphenyl-2-picrylhydrazyl (DPPH). FDP also preserved cellular GSH content after UV irradiation in HaCaT and normal human fibroblast culture system. Cellular oxidative stress induces multiple downstream signaling pathways that regulate expression of multiple gene including MMP-1 and collagen, we examined the effects of FDP on UV-induced alteration of these protein expression in fibroblast culture and human skin in vivo. The increased MMP-1 expression in fibroblast and human skin by UV irradiation was significantly decreased by FDP. FDP also prevented the UV-induced decrease of collagen expression in fibroblast and human skin. Moreover, the decreasing the intracellular levels of reducing equivalents in human fibroblast by glutathione (GSH) depletion lowered the UVA dose threshold for reduction of procollagen expression, indicating that the differences of glutathione contents define the susceptibility of fibroblasts towards UV-induced reduction of procollagen expression. FDP also preserved cellular GSH content after UV irradiation, indicating that FDP has protective effects on UV-induced reduction of procollagen expression, which are possibly through maintaining intracellular reducing equivalent. Based on these premises, we examined the effect of daily use of a moisturizer containing FDP on facial wrinkle in comparison with vehicle moisturizer lacking FDP. In the clinical study, FDP significantly decreased facial wrinkle compared with vehicle alone after 6 months of use. Our results suggest that FDP has anti-aging effects in skin by increasing cellular antioxidant system and preventing oxidative signal and inflammatory reaction. Therefore FDP may be useful anti-aging agent for cosmetic purpose.

• Journal of Acupuncture Research
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• 제19권2호
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• pp.211-220
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• 2002

Objective : This study was undertaken to determine whether Scutellaria baicalensis Georgi extract (SbG) exerts the protective effect against oxidant-induced alterations in organic cation transport in the renal proximal tubule. Methods : Organic cation transport was estimated by examining alterations in tetraethylammonium (TEA) uptake in rabbit renal cortical slices. The slices were treated with 0.2 mM tBHP for 60 min at $37^{\circ}C$. tBl-IP caused an inhibition in TEA uptake by renal cortical slices. Such an effect was accompanied by depressed Na+-K+-ATPase activity and ATP depletion. Result : SbG prevented tBHP-induced inhibition of TEA uptake in a dose-dependent manner at the concentration ranges of 0.05-0.1%. SbG also prevented H2O2-induced reduction in TEA uptake. tBHP-induced inhibition of Na+-K+-ATPase activity and ATP depletion were significantly prevented by 0.05% SbG. Oxidants increased LDH release, which was blocked by SbG. Oxidants caused a significant increase in lipid peroxidation and its effect was prevented by SbG. Conclusion : These results suggest that SbG prevents oxidant-induced alterations in organic cation transport in rabbit renal cortical slices. Such protective effects of SbG may be attributed to inhibition of peroxidation of membrane lipid.

• 대한약리학회지
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• 제32권1호
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• pp.31-37
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• 1996

허혈전처치(IP)의 히혈-재관류손상에 대한 심근 보호작용의 기전을 규명하기 위한 일환으로 denosine에 의한 PKC자극이 허혈전처치의 주요 기전으로 작용할 가능성을 조사하였다. 흰쥐 적출심장의 Langendorff 관류 표본에서 실험적인 허혈(30분)-재관류(20분)손상을 유도하였고, 허혈전처치는 허혈-재관류 손상 유도 전에 5분 허혈-5분 재관류를 3회 반복하여 시행하였다. 심근 손상의 지표로 심수축기능, 세포질효소 유출을 측정하였다. Adenosine이 허혈전처치의 심보호 효과에 관여하는지를 관찰하기 위하여 adenosine수용체 억제제인 8-(p-sulfophenyl)-theophylline(SPT), Xanthine amine congener(XAC) 및 8-cyclopentyl-1,3-dipropylxanthine (DPCPX)을 허혈전처치 유도 전에 투여하였다. 또한 PKC가 허혈전처치의 세포내 매개인자로 관여 할 가능성을 관찰하기 위하여 PKC활성 억제제인 polymyxin B 및 chelerythrine과 PKC translocation 억제제인 colchicine을 허혈전처치 유도 전에 투여하였다. 연구성적은 다음과 같다. 1) 허혈전처치는 허혈재관류 심장의 심기능의 저하를 현저히 회복시켜 심기능 회복률은 75%에 달하였다. 2) 허혈-재관류 심장에서 lactate dehydrogenase유출증가는 허혈전처치에 의해 현저히 저하되었다. 3) Adenosine 비선택적 차단제인 SPT와 Al 선택적 차단제인 DPCPX 및 XAC의 투여가 허혈전처치에 의한 심기능회복 및 LDH 유출 감소에 영향을 미치지 않았다. 4) PKC활성 억제제인 polymyxin B 와 chelerythrine을 처치시 히혈전처치 심장의 심기능 회복률이 현저히 감소되었으며 LHD 유출 역시 대조군 심장의 수준으로 증가하였다. 5) PKC translocation을 방해하는 colchicine도 허혈전처치의 심보호 효과를 억제시켰다. 이상의 결과들로부터 adenosine은 흰쥐 심장에서 허혈전처치의 심보호효과에 중요한 세포외 매개물질로 작용할 가능성이 희박하며, PKC는 흰쥐 심장에서 허혈전처치시 세포내 매개 인자로 관여하여 허혈전처치에 의한 심보호효과에 중요한 역할을 할 수 있으리라 사료된다.