• The Korean Journal of Physiology and Pharmacology
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• 제20권1호
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• pp.101-109
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• 2016

Reducing $[Mg^{2+}]_o$ to 0.1 mM can evoke repetitive $[Ca^{2+}]_i$ spikes and seizure activity, which induces neuronal cell death in a process called excitotoxicity. We examined the issue of whether cultured rat hippocampal neurons preconditioned by a brief exposure to 0.1 mM $[Mg^{2+}]_o$ are rendered resistant to excitotoxicity induced by a subsequent prolonged exposure and whether $Ca^{2+}$ spikes are involved in this process. Preconditioning by an exposure to 0.1 mM $[Mg^{2+}]_o$ for 5 min inhibited significantly subsequent 24 h exposure-induced cell death 24 h later (tolerance). Such tolerance was prevented by both the NMDA receptor antagonist D-AP5 and the L-type $Ca^{2+}$ channel antagonist nimodipine, which blocked 0.1 mM $[Mg^{2+}]_o$-induced $[Ca^{2+}]_i$ spikes. The AMPA receptor antagonist NBQX significantly inhibited both the tolerance and the $[Ca^{2+}]_i$ spikes. The intracellular $Ca^{2+}$ chelator BAPTA-AM significantly prevented the tolerance. The nonspecific PKC inhibitor staurosporin inhibited the tolerance without affecting the $[Ca^{2+}]_i$ spikes. While $G{\ddot{o}}6976$, a specific inhibitor of $PKC{\alpha}$ had no effect on the tolerance, both the $PKC{\varepsilon}$ translocation inhibitor and the $PKC{\zeta}$ pseudosubstrate inhibitor significantly inhibited the tolerance without affecting the $[Ca^{2+}]_i$ spikes. Furthermore, JAK-2 inhibitor AG490, MAPK kinase inhibitor PD98059, and CaMKII inhibitor KN-62 inhibited the tolerance, but PI-3 kinase inhibitor LY294,002 did not. The protein synthesis inhibitor cycloheximide significantly inhibited the tolerance. Collectively, these results suggest that low $[Mg^{2+}]_o$ preconditioning induced excitotoxic tolerance was directly or indirectly mediated through the $[Ca^{2+}]_i$ spike-induced activation of $PKC{\varepsilon}$ and $PKC{\xi}$, JAK-2, MAPK kinase, CaMKII and the de novo synthesis of proteins.

• 대한한방내과학회지
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• 제29권1호
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• pp.130-148
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• 2008

Objective : This study was designed to investigate the effect of the water extract of Gamisamgibopae-tang(GMSGBPT), an oriental herbal formulation, on the growth of NCI-H460 and A549 human non-small-cell lung cancer cell lines. Methods : Cytotoxicity and cell morphology were evaluated by MTT assay and inverted microscope, respectively. Apoptosis was detected using agarose gel electrophoresis and flow cytometer. The expression levels of mRNAs and proteins of target genes were determined by RT-PCR and western blot analyses, respectively Result and Conclusion : We found that exposure of A549 cells to GMSGBPT resulted in the growth inhibition in a dose-dependent manner as measured by MTT assay, but GMSGBPTdid not affect the growth of NCI-H460 cells. The anti-proliferative effect of GMSGBPT treatment in A549 cells was associated with morphological changes, formation of apoptotic bodies and DNA fragmentation, and flow cytometry analysis confirmed that GMSGBPT treatment increased the populations of apoptotic-sub G1 phase. Growth inhibition and apoptotic cell death by GMSGBPT were connected with a up-regulation of cyclin-dependent kinase inhibitor p21 (WAF1/CIP1) mRNA and protein in a tumor suppressor p53-independent fashion. However GMSGBPT treatment did not affect other growth regulation-related genes such as early growth response-1 (Egr-1), nonsteroidal anti-inflammatory drug (NSAID)-activated gene-1 (NAG-1), inducible nitric oxide synthase (iNOS), cyclooxygenases (COXs), telomere-regulatory factors in A549 orNCI-H460 cells. Taken together, these findings partially provide novel insights into the possible molecular mechanism of the anti-cancer activity of GMSGBPT.

• 한국식품과학회지
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• 제17권4호
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• pp.248-252
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• 1985

비이용 자원의 효율적인 이용의 가능성을 검토하기 위하여 앵두씨의 지방질 조성 및 단백질 패턴을 분석한 결과는 다음과 같다. 일반성분 중 조지방질과 조단백질의 함량은 각각 40.38% 및 26.59%였다 앵두씨의 지방질은 중성지방질이 95.4%인 반면에 복합지방질은 4.51%에 불과하였으며, 중성지방질의 성분으로는 트리-글리세리드가 89.86%로서 대부분이었고. 스테롤 4.14%. 모노-글리세리드 2.98%, 스테롤 에스테르 1.77%. 유리지방산 1.07% 및 디 -글리세리드 0.18% 였다. 지방산 조성은 총 지방질, 중성지방절 및 트리 글리세리드에서 올레산(65.06$\sim$66.05%)와 리놀레산(26.56$\sim$28.40%)이 주된 구성 지방산이었고, 당지방질파 인지방질의 경우에는 올FP산(40.55$\sim$51.46%)의 함량이 다른 획분의 함량에 비해 적은 반면, 팔미트산(17.64$\sim$21.43%)과 스테아르산(4.14$\sim$8.07%) 의 함량이 높았으며, 소량의 라오르산도 함유되어 있었다. 염용해성 단백질의 추출율은 약 60%였으며, 분획된 주단백질의 수득율은 약 46.5% 였고 전기영동 결과 7개의 밴드가 확인되었다.

• 한국식품저장유통학회지
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• 제9권1호
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• pp.67-73
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• 2002

수입 참깨가루로 제조한 참기름의 품질과 위생적 안정성에 대한 기초자료를 얻고자 통참깨와 참깨가루를 원료로한 참기름의 일반성분, 화학적 특성, 지방산, sesamol, sesamolin, sesamin 함량 및 AOM에 의한 유도기간 등을 비교 조사하였다. 일반성분 중 수분은 통참깨에 비해 참깨가루가 월등히 낮게 나타났으며, 단백질은 거의 차이가 없었다. 참기름의 화학적 특성으로 산가와 검화가는 참깨가루가 통참깨에 비해 높게 나타났으며, 요오드가는 통참깨군보다 낮았다. 참기름의 지방산 조성에서 linolenic acid과 linoleic acid의 함량은 각각 222.44~144.14, 2713~1776.46 $\mu\textrm{g}$/mL이었고, linoleic acid와 ${\gamma}$-linoleic acid의 함량이 통참깨 참기름에 비해 참깨가루 참기름의 함량이 낮았다. Lignan 물질 중 sesamol 함량은 통참깨 참기름이 참깨가루 참기름보다 함량이 높았고, 인도산 통참깨 참기름의 sesamol 함량이 가장 높았다. Sesamin, sesamolin 함량은 한국만 통참깨 참기름이 가장 높았다. AOM에 의한 산화유도시간은 북한산, 중국산 참깨가루가 각각 6.67, 13.35 시간으로 나타나 통참깨군보다 산화안정성이 낮은 것으로 나타났다.

• 대한한의학방제학회지
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• 제23권2호
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• pp.189-198
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• 2015

Objectives : Sparganii Rhizoma is frequently used in traditional herbal medicine for treatment of blood stasis, amenorrhea and functional dyspepsia and has been reported to exhibit anti-oxidant, anti-proliferation and anti-angiogenesis peoperties. In this study, we investigated the cytoprotective effect and underlying mechanism of Sparganii Rhizoma water extract (SRE) against oxidative stress-induced mitochondrial dysfunction and apoptosis in hepatocyte. Methods : To determine the effects of SRE on oxidative stress, we induced synergistic cytotoxicity by co-treatment of arachidonic acid (AA) and iron in the HepG2 cell, a human derived hepatocyte cell line. Results : Treatment of SRE increased relative cell viability and altered the expression levels of apoptosis-related proteins such as Bcl-xL, Bcl-2 and procaspase-3. And SRE also inhibited the mitochondrial dysfunction and excessive reactive oxygen species production induced by AA+iron. In addition, SRE activated of AMP-activated protein kinase (AMPK), a potential target for cytoprotection, by increasing the phosphorylation of AMPKα at Thr-172. Morever, SRE increased phosphorylation of acetyl-CoA carboxylase, a direct downstream target of AMPK. Conclusion : These results indicated that SRE has the ability to protect against oxidative stress-induced hepatocyte damage, which may be mediated with AMPK pathway.

• 한국식품영양과학회지
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• 제37권1호
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• pp.117-123
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• 2008

국내에서 생산된 감자 중 세풍, 남서, 수미, 조풍 및 대서의 5가지 품종에 대하여 단백질 profile 및 아미노산 조성을 분석하였다. 총 질소함량은 $1.27{\sim}1.64%$이었으며, 남서가 높았고 수미가 낮게 나타났다. 아미노산 조성은 품종 간에 유의적인 차이가 있었다. 한편, 주요 감자 단백질은 papatin(40 kDa), trypsin inhibitor(20 kDa) 및 protease inhibitor(15 kDa)이었으며, 이들의 함량은 각각 $22.16{\sim}25.81%$, $25.22{\sim}20.91%$ 및 $14.12{\sim}25.23%$이었다. Papatin 함량은 조풍, 세풍, 수미감자가 높은 함량을 보인 반면, trypsin inhibitor는 조풍감자가 5.22%로 가장 낮은 함량을 보였다. Protease inhibitors인 20 kDa와 15 kDa를 합한 값은 $24.7{\sim}35.0%$이었으며, 세풍이 가장 적었고 조풍에 가장 많이 함유되어 있었다.

• International Journal of Oral Biology
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• 제31권4호
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• pp.141-148
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• 2006

The effects of adenosine triphosphate(ATP) on salivary glands have been recognized since 1982. The presence of purinergic recepetors(P2Rs) that mediate the effects of ATP in various tissues, including parotid and submandibular salivary gland, has been supported by the cloning of receptor cDNAs and the expression of the receptor proteins. P2Rs have many subtypes, and the activation of these receptor subtypes increase intracellular $Ca^{2+}$, a key ion in the regulation of the secretion in the salivary gland. The apical pores of taste buds in circumvallate and foliate papillae are surrounded by the saliva from von Ebner salivary gland(vEG). Thus, it is important how the secretion of vEG is controlled. This study was designed to elucidate the roles of P2Rs on salivary secretion of vEG. Male Sprague-Dawley rats (about 200 g) were used for this experiment. vEG-rich tissues were obtained from dissecting $500-1,000\;{\mu}m$ thick posterior tongue slices under stereomicroscope view. P2Rs mRNA in vEG acinar cells were identified with RT-PCR. To observe the change in intracellular $Ca^{2+}$ activity, we employed $Ca^{2+}-ion$ specific fluorescence analysis with fura-2. Single acinar cells and cell clusters were isolated by a sequential trypsin/collagenase treatment and were loaded with $10\;{\mu}M$ fura -2 AM for 60 minutes at room temperature. Several agonists and antagonists were used to test a receptor specificity. RT-PCR revealed that the mRNAs of $P2X_4$, $P2Y_1$, $P2Y_2$ and $P2Y_3$ are expressed in vEG acinar cells. The intracellular calcium activity was increased in response to $10\;{\mu}M$ ATP, a P2Rs agonist, and 2-MeSATP, a $P2Y_1$ and $P2Y_2R$ agonist. However, $300\;{\mu}M\;{\alpha}{\beta}-MeATP$, a $P2X_1$ and $P2X_3R$ agonist, did not elicit the response. The responses elicited by $10\;{\mu}M$ ATP and UTP, a $P2Y_2R$ agonists, were maintained when extracellular calcium was removed. $10\;{\mu}M$ suramin, a P2XR antagonist, and reactive blue 2, a P2YR antagonist, partially blocked ATP-induced response. However, when extracellular calciums were removed, suramin did not abolish the responses elicited by ATP. These results suggest that P2Rs play an important role in salivary secretion of vEG acinar cells and the effects of ATP on vEG salivary secretion may be mediated by $P2X_4$, $P2Y_1$, $P2Y_2$, and/or $P2Y_3$.

• 대한본초학회지
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• 제30권6호
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• pp.7-15
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• 2015

Objectives : Rosa laevigata Michx. has been used for the treatment of renal disease in traditional Korean medicine. In this study, we investigated cytoprotective effect of R. laevigata water extract (RLE) against oxidative stress induced by arachidonic acid (AA) + iron.Methods : To evaluate the protective effects of RLE against AA + iron-induced oxidative stress in HepG2 cell, cell viability and changes on apoptosis-related proteins were assessed by MTT and immunoblot analyses. The effects of RLE on reduced glutathione level, production of reactive oxygen species and mitochondrial membrane potential were also monitored. Furthermore, to verify underlying molecular mechanism, NF-E2-related factor 2 (Nrf2) was examined by immunoblot analysis. Additionally, Nrf2 transactivation and its downstream target genes expression were also determined by reporter gene and realtime RT-PCR analyses.Results : RLE pretreatment (30-300 μg/ml) prevented cells from AA + iron-mediated cell death in a concentration dependent manner. In addition, 100 μg/ml RLE inhibited AA + iron-induced glutathione depletion, reactive oxygen species production and mitochondrial dysfunction. RLE accumulated nuclear Nrf2 and also transactivated Nrf2, which was evidenced by antioxidant response element- and glutathione S-transferase A2-driven luciferase activities and mRNA level of glutamate-cysteine ligase catalytic subunit, NAD(P)H:quinone oxidoreductase 1 and sestrin 2. Moreover, protective effect of RLE against AA + iron was abolished in Nrf2 knockout cells.Conclusions : These results indicate that RLE has the ability to protect hepatocyte against oxidative stress through Nrf2 activation.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제32권4호
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• pp.295-307
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• 2006

Styela clava, called non-native tunicate or sea squirt, is habitat which include bays and harbors in Korea and several sites in the sea faced world. We fabricate cellulose membrane nerve conduit (CMNC) from this native sea squirt skin, and evaluate the capacity of promoting peripheral nerve regeneration in the rat sciatic nerve defect model. After processing the pure cellulose membrane from the sea squirt skin as we already published before, CMNC was designed as a non-tubular sheet with 14 mm length and 4 mm width. Total eleven male Spraque-Dawley rats (12 weeks, weighing 250 to 300g) were divided into sham group (n=2), silicone tube grafted control group (n=3) and experimental group (n=6). Each CMNC grafted nerve was evaluated after 4, 8 and 12 weeks in the experimental group, and after 12 weeks, sciatic function was evaluated with sciatic function index (SFI) and gait analysis, and histomorphology of nerve conduit and the innervated tissues of sciatic nerve were all examined using image analyzer and electromicroscopic methods in the all groups. The regenerated axon and nerve sheath were found only in the inner surface of the CMNC after 4 weeks and became more thicker after 8 and 12 weeks. In the TEM study, CMNC grafted group showed more abundant organized myelinated nerve fibers with thickened extracellular matrix than silicone conduit grafted group after 12 weeks. The sciatic function index (SFI) and ankle stance angle (ASA) in the functional evaluation were $-47.2{\pm}3.9$, $35.5^{\circ}{\pm}4.9^{\circ}$ in CMNC grafted group (n=2) and $-80.4{\pm}7.4$, $29.2^{\circ}{\pm}5.3^{\circ}$ in silicone conduit grafted group (n=3), respectively. And the myelinated axon was 41.59% in CMNC group and 9.51% in silicone conduit group to the sham group. The development of a bioactive CMNC to replace autogenous nerve grafts offers a potential and available approach to improved peripheral nerve regeneration. As we already published before, small peptide fragment derived from the basement membrane matrix proteins of squirt skin, which is a kind of anchoring protein composed of glycocalyx, induced the effective axonal regeneration with rapid growth of Schwann cells beneath the inner surface of CMNC. So the possibilities of clinical application as a peripheral nerve regeneration will be able to be suggested.

• 한국식품저장유통학회지
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• 제23권7호
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• pp.1026-1032
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• 2016

시기별 병귤 추출물의 항산화 활성은 시기적으로 미숙과 시기인 9월에 가장 높았으며, 이 시기에 총 폴리페놀 함량이 가장 높았다. 특히 rutin, hesperidin, nobiletin의 함량이 높았으며, 이는 항염 활성에 영향을 미치는 것으로 여겨진다. 항염 활성에서는 NO의 생성을 억제하였으며 염증성 cytokine인 TNF-${\alpha}$의 생성 억제 활성이 가장 높았다. 또한 NO 생성을 억제하는 단백질로 알려진 iNOS 단백질의 발현 역시 억제하는 것을 확인 할 수 있었다. 항염 활성에 영향을 미칠 것으로 여겨지는 nobiletin 함량의 경우 12월에 70%이상 감소하는 것을 확인할 수 있었다. 이러한 시기별 플라보노이드 함량 분석결과는 병귤을 천연 소재로 활용하기 위한 수확시기를 확립할 수 있을 것으로 여겨진다.