• The Korean Journal of Physiology and Pharmacology
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• 제13권5호
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• pp.401-408
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• 2009

$K^+$-$Cl^-$-cotransport (KCC) has been reported to have various cellular functions, including proliferation and apoptosis of human cancer cells. However, the signal transduction pathways that control the activity of KCC are currently not well understood. In this study we investigated the possible role of phospholipase $A_2$ ($PLA_2$)-arachidonic acid (AA) signal in the regulatory mechanism of KCC activity. Exogenous application of AA significantly induced $K^+$ efflux in a dose-dependent manner, which was completely blocked by R-(+)-[2-n-butyl-6,7 -dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1Hinden-5-yl]oxy]acetic acid (DIOA), a specific KCC inhibitor. N-Ethylmaleimide (NEM), a KCC activatorinduced $K^+$ efflux was significantly suppressed by bromoenol lactone (BEL), an inhibitor of the calciumindependent $PLA_2$ ($iPLA_2$), whereas it was not significantly altered by arachidonyl trifluoromethylketone ($AACOCF_3$) and p-bromophenacyl bromide (BPB), inhibitors of the calcium-dependent cytosolic $PLA_2$ ($cPLA_2$) and the secretory $PLA_2$ ($sPLA_2$), respectively. NEM increased AA liberation in a doseand time-dependent manner, which was markedly prevented only by BEL. In addition, the NEM-induced ROS generation was significantly reduced by DPI and BEL, whereas $AACOCF_3$ and BPB did not have an influence. The NEM-induced KCC activation and ROS production was not significantly affected by treatment with indomethacin (Indo) and nordihydroguaiaretic acid (NDGA), selective inhibitors of cyclooxygenase (COX) and lipoxygenase (LOX), respectively. Treatment with 5,8,11,14-eicosatetraynoic acid (ETYA), a non-metabolizable analogue of AA, markedly produced ROS and activated the KCC. Collectively, these results suggest that $iPLA_2$-AA signal may be essentially involved in the mechanism of ROS-mediated KCC activation in HepG2 cells.

• Biomolecules & Therapeutics
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• 제20권6호
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• pp.526-531
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• 2012

During our on-going studies to identify bioactive compounds in medicinal herbs, we found that saucerneol F (SF), a naturally occurring sesquilignan isolated from Saururus chinensis (S. chinensis), showed in vitro anti-inflammatory activity. In this study, we examined the effects of SF on the generation of 5-lipoxygenase (5-LO) dependent leukotriene $C_4$ ($LTC_4$), cyclooxygenase-2 (COX-2) dependent prostaglandin $D_2$ ($PGD_2$), and on phospholipase $C{\gamma}1$ ($PLC{\gamma}1$)-mediated degranulation in SCF-induced mouse bone marrow-derived mast cells (BMMCs). SF inhibited eicosanoid ($PGD_2$ and $LTC_4$) generation and degranulation dose-dependently. To identify the molecular mechanisms underlying the inhibition of eicosanoid generation and degranulation by SF, we examined the effects of SF on the phosphorylation of $PLC{\gamma}1$, intracellular $Ca^{2+}$ influx, the translocation of cytosolic phospholipase $A_2$ ($cPLA_2$) and 5-LO, and on the phosphorylation of MAP kinases (MAPKs). SF was found to reduce intracellular $Ca^{2+}$ influx by inhibiting $PLC{\gamma}1$ phosphorylation and suppressing the nuclear translocations of $cPLA_2$ and 5-LO via the phosphorylations of MAPKs, including extracellular signal-regulated protein kinase-1/2 (ERK1/2), c-Jun N-terminal kinase (JNK), and p38. Taken together, these results suggest that SF may be useful for regulating mast cell-mediated inflammatory responses by inhibiting degranulation and eicosanoid generation.

• 한국식품영양과학회지
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• 제42권3호
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• pp.369-377
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• 2013

고구마뿌리에 비하여 많은 양이 폐기되는 고구마 줄기와 잎의 이용을 활성화시키고 기능성식품 소재로의 가능성을 알아보기 위하여 잎이 달려 있는 보라색 고구마줄기를 재래시장에서 구입하여 고구마잎(SL), 고구마줄기(ST), 껍질 제거한 고구마줄기(PST)의 총 폴리페놀과 총 플라보노이드 함량을 측정하고, in vitro 시스템에서의 항산화효과, 항알레르기 및 항염증효과를 검색하고자 하였다. 고구마잎은 데치고, 고구마줄기는 생으로 동결건조한 후 에탄올 추출물을 얻어 실험하였다. 총 폴리페놀 함량은 SL(11.03 mg tannic acid/g)> ST(0.87 mg tannic acid/g)> PST(0.37 mg tannic acid/g)이었고, 총 플라보노이드 함량은 SL(9.01 mg rutin/g)> ST(0.50 mg rutin/g)> PST(0.25 mg rutin/g)이었다(p<0.001). DPPH 라디칼을 50% 제거시키는 에탄올 추출물의 농도($IC_{50}$)는 SL($43.6{\mu}g/mL$)< ST($308.4{\mu}g/mL$)$1,631.3{\mu}g/mL$)로 고구마잎이 가장 우수한 효과를 나타내었다. 양성대조시약으로 사용한 BHA의 $IC_{50}$값은 $8.30{\mu}g/mL$이었으며, 처리 농도 $1,000{\mu}g/mL$에서의 환원력은 SL($59.72{\mu}g$ ascorbic acid eq./mL)> ST($12.56{\mu}g$ ascorbic acid eq./mL)> PST($2.18{\mu}g$ ascorbic acid eq./mL)로 고구마잎이 가장 좋았다(p<0.001). 한편 염증반응에 관여하는 xanthine oxidase(XO) 활성저해율을 측정한 결과 처리 농도 $250{\mu}g/mL$에서 SL(13.06%)> ST(5.05%)> PST(0.0%)로 고구마잎이 줄기에 비하여 우수하였으며(p<0.01), 처리 농도 $50{\mu}g/mL$에서의 5-lipoxygenase(LOX) 활성저해율은 SL(91.16%)> ST(33.38%)> PST(14.93%)로(p<0.001) 고구마잎의 효과는 양성대조시약인 EGCG의 저해율(94.42%)과 비슷한 정도로 매우 우수하였다. 또한 $250{\mu}g/mL$ 농도로 처리 시 cyclooxygenase(COX)-2 활성저해율은 SL(55.34%)> ST(2.18%)> PST(0.0%)로 XO 활성저해율과 비슷한 패턴을 보였다(p<0.001). 측정변수들 간의 상관관계를 분석해 본 결과 총 폴리페놀 함량은 플라보노이드 함량($r^2$=0.9988, p<0.001), 환원력($r^2$=0.9982, p<0.001), XO 활성저해율($r^2$=0.8322, p<0.05), COX-2 활성저해율($r^2$=0.9950, p<0.001), 5-LOX 활성저해율과 양의 상관관계($r^2$=0.9823, p<0.001)를 나타내었으며, 플라보노이드 함량은 환원력($r^2$=0.9946, p<0.001), XO 활성저해율($r^2$=0.8392, p<0.05), 5-LOX 활성저해율($r^2$=0.9749, p<0.01), COX-2 활성저해율과 유의한 양의 상관관계($r^2$=0.9937, p<0.001)를 보였다. 또한, 환원력은 XO 활성 저해율($r^2$=0.8384, p<0.05), 5-LOX 활성저해율($r^2$=0.9883, p<0.001) 및 COX-2 활성저해율과 유의한 양의 상관관계($r^2$=0.9954, p<0.001)를 나타내었으며, XO 활성저해율은 5-LOX 활성저해율과 유의한 양의 상관관계를 보였으나($r^2$=0.8786, p<0.05) COX-2 활성저해율과는 상관성을 보이지 않았다. 5-LOX 활성저해율은 COX-2 활성저해율과 유의한 양의 상관관계($r^2$=0.9815, p<0.01)를 나타내었다. 이상의 결과들로부터 고구마잎은 폴리페놀 및 플라보노이드 함량이 매우 높았고 우수한 항산화효과를 보였으며, 알레르기 및 염증반응과 관련이 있는 효소인 XO, 5-LOX 및 COX-2의 활성도 모두 억제하는 효과도 우수하였지만 특히 5-LOX 활성 억제효과는 EGCG와 비슷한 정도로 매우 우수하였다. 따라서 건강을 위하여 고구마잎의 섭취를 증대시킬 수 있는 다양한 방안을 강구할 필요가 있으며, 고구마 줄기를 섭취할 경우에는 가능한 껍질을 모두 섭취하는 것이 좋겠다.

• 한국식품영양과학회지
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• 제38권12호
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• pp.1691-1698
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• 2009

본 연구에서는 현재까지 연구되어진 감잎 추출물(PLE)의 추출방법을 달리하고 그 추출물을 비 특이적 흡착 resin을 사용하여 부분 정제함으로서 추출물의 활성을 높이고자 하였으며 PLE와 PLE의 부문 정제물(PPLE)의 페놀성 화합물 및 플라보노이드 함량을 측정하였다. 또한 이들의 항산화제와 항알레르기 개선제로써의 유효성을 알아보기 위해 항산화 효과에 관여하는 DPPH 라디칼 포착효능, superoxide 음이온 라디칼 포착효능과 제Ⅰ형 알레르기 반응 억제 효능에 관여하는 5-LO억제 효능, COX억제효능, NO소거효능 및 수동피부아나필락시스 억제 효능(PCA)을 측정하였다. 그 결과 PLE와 PPLE의 페놀성 화합물은 각각 $230.0{\pm}19.6$ mg/g, $475{\pm}38.7$ mg/g이 함유되어 있었고, 플라보노이드는 각각 $34.8{\pm}6.5$ mg/g, $78.8{\pm}3.6$ mg/g 함유되어 있는 것으로 나타났다. 그리고 PLE와 PPLE이 각각 $23.8{\pm}63.2$ mg/g, $10.0{\pm}1.3$ mg/g의 농도에서 DPPH 라디칼을, $47.6{\pm}3.4$ ppm, $22.4{\pm}3.3$ ppm의 농도에서 superoxide 음이온 라디칼을 50% 소거하는 것으로 나타났다. PLE와 PPLE의 항알레르기 효능에 관하여 5-LO의 $IC_{50}$값이 각각 $77.1{\pm}11.7$ ppm, $38.6{\pm}7.0$ ppm으로 양성대조군인 EGCG의 $IC_{50}$($18.9{\pm}5.5ppm$)보다는 높았지만 PLE와 PPLE가 EGCG와 같은 단일물질이 아니라 천연혼합물임을 감안하였을 때 5-LO를 비교 적 낮은 농도에서 억제하고 있다고 볼 수 있다. 또한, COX-2의 선택적 저해율을 COX-1과 COX-2의 비율 비교를 통해 나타낸 결과 양성대조군으로 사용한 EGCG보다는 떨어지는 결과를 나타냈으나 다른 천연물과 비교하였을 때 COX-2를 상당히 선택적으로 저해하고 있음을 확인하였다. 마우스의 대식세포인 RAW 264.7 세포주를 사용하여 낮은 세포독성과 NO소거에도 우수한 효과가 있음을 확인하였고, 마우스를 이용하여 수동피부아나필락시스 반응 억제 효과도 확인하였다. 이상의 결과에 따르면, 감잎 추출물이 항산화 및 항알레르기 효능을 갖고 있어 알레르기성 비염이나 아토피와 같은 알레르기관련 질병 예방 및 개선 또는 건강식품 원료로서 유효하게 사용될 것이라 생각된다.

• 대한한의학회지
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• 제39권4호
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• pp.86-113
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• 2018

Objectives: We examined the effects of a mixed formula consisting of dried pomegranate concentrate powder (PCP) and the aqueous extracts of Eucommiae cortex (EC) and Achyranthis radix (AR) in rats with surgically induced osteoarthritis (OA). Methods: Two weeks after OA-inducing surgery, a PCP:EC:AR 5:4:1 (g/g) combination or single formula was orally administered. Changes in body weight, knee thickness, maximum knee extension angle, bone mineral density of the knee joints, femoral and tibial articular surfaces, and compressive strength of the femoral and tibial articular cartilage (AC) were assessed, along with the prostaglandin E2 level, 5-lipoxygenase, matrix metalloproteinase (MMP)-2 and MMP-9 activity, and chondrogenic gene mRNA expression in the femoral and tibial AC with the synovial membrane (SM). In addition, the number of cleaved poly(ADP-ribose) polymerase, cyclooxygenase and tumor necrosis factor-${\alpha}$-immunoreactive cells in the femoral and tibial AC with SM were monitored, and the rate of cell proliferation was determined with a 5-bromo-2'-deoxyuridine uptake assay. Results : The signs of surgically induced OA in rats were significantly inhibited by both PCP, EC and AR combined and single formulas. In particular, the combination formula-treated OA model rats showed dose-dependent, significantly increased inhibitory activity against all tested criteria compared with single formula-treated rats. Conclusions: Taken together, our results suggest that the combination formula synergistically increased the anti-OA effects of its components through anti-inflammatory and chondrogenic activity in rats with surgically induced OA. In addition, 200, 100 and 50 mg/kg combination formula treatments showed dose-dependent inhibitory activity against all of the tested criteria.

• Biomolecules & Therapeutics
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• 제22권1호
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• pp.27-34
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• 2014

Curcumin is naturally occurring polyphenolic compound found in turmeric and has many pharmacological activities. The present study was undertaken to evaluate anti-allergic inflammatory activity of curcumin, and to investigate its inhibitory mechanisms in immunoglobulin E (IgE)/Ag-induced mouse bone marrow-derived mast cells (BMMCs) and in a mouse model of IgE/Ag-mediated passive systemic anaphylaxis (PSA). Curcumin inhibited cyclooxygenase-2 (COX-2) dependent prostaglandin $D_2$ ($PGD_2$) and 5-lipoxygenase (5-LO) dependent leukotriene $C_4$ ($LTC_4$) generation dose-dependently in BMMCs. To probe the mechanism involved, we assessed the effects of curcumin on the phosphorylation of Syk and its downstream signal molecules. Curcumin inhibited intracellular $Ca^{2+}$ influx via phospholipase $C{\gamma}1$ ($PLC{\gamma}1$) activation and the phosphorylation of mitogen-activated protein kinases (MAPKs) and the nuclear factor-${\kappa}B$ (NF-${\kappa}B$) pathway. Furthermore, the oral administration of curcumin significantly attenuated IgE/Ag-induced PSA, as determined by serum $LTC_4$, $PGD_2$, and histamine levels. Taken together, this study shows that curcumin offers a basis for drug development for the treatment of allergic inflammatory diseases.

• Biomolecules & Therapeutics
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• 제23권5호
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• pp.421-427
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• 2015

Imperatorin has been known to exert many biological functions including anti-inflammatory activity. In this study, we investigated the inhibitory effects of imperatorin on the production of inflammatory mediators in mouse bone marrow-derived mast cells (BMMC). Imperatorin inhibited degranulation and the generation of eicosanoids (leukotriene $C_4$ ($LTC_4$) and prostaglandin $D_2$ ($PGD_2$) in IgE/antigen (Ag)-stimulated BMMC. To elucidate the molecular mechanism involved in this process, we investigated the effect of imperatorin on intracellular signaling in BMMC. Biochemical analyses of the IgE/Ag-mediated signaling pathway demonstrated that imperatorin dramatically attenuated degranulation and the production of 5-lipoxygenase-dependent $LTC_4$ and cyclooxygenase-2-dependent $PGD_2$ through the inhibition of intracellular calcium influx/phospholipase $C{\gamma}1$, cytosolic phospholipase $A_2$/mitogen-activated protein kinases and/or nuclear factor-${\kappa}B$ pathways in BMMC. These results suggest that the effects of imperatorin on inhibition of degranulation and eicosanoid generation through the suppression of multiple steps of IgE/Ag-mediated signaling pathways would be beneficial for the prevention of allergic inflammation.

• 대한약리학회지
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• 제23권2호
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• pp.151-157
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• 1987

Steroidal alkaloid인 cyclobuxine D의 carrageenin으로 유발한 늑막염과 croton oil로 유발한 육아종양에 대한 영향을 관찰하고 그 작용을 aspirin, dexamethasone과 hydrocortisone의 작용과 비교하였다. Carrageenin으로 유발한 늑막염에서 혈장 삼출정도는 pontamine sky blue을 정맥으로 투여하여 20분 동안 늑막염 삼출물로 삼출되어 나오는 양으로 측정하였다. Cyclobuxine D는 용량적으로 삼출물의 양과 20분 동안 삼출되는 pontamine sky blue의 양을 감소시켰다. Cyclooxygenase 억제제인 aspirin과 phospholipase $A_2$를 억제하여 결과적으로 cyclooxygenase와 lipoxygenase의 생성물을 차단하는 dexamethasone은 현저하게 carrageenin으로 나타나는 염증현상을 억제하였다. Cyclobuxine D는 croton oil로 유발한 육아종양에서 종양 부위에 직접 투여하거나 근육내로 투여해도 pouch내 염증물 양과 pouch wall의 무게를 현저하게 억제하였다. Dexamethasone과 hydrocortisone은 육아종양의 형성을 현저히 억제하였으며 , dexamethasone은 근육내로 투여 하였을 때 육아종양 형성을 완전히 억제하였다. 이상의 결과에서 나타난 cyclobuxine D의 항염증 작용은 prostaglandins의 생성억제 또는 polymorphonuclear cell (PMN)의 유주 억제와 밀접한 관련이 있을 것으로 사려된다.

• Archives of Pharmacal Research
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• 제24권6호
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• pp.552-556
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• 2001

Histamine and arachidonic acid (AA) release was measured using the P2-purinoceptor antaongists, phospholipase $A_2{\;}(PLA_2)$ and cyclooxygenase (COX)/lipoxygenase (LOX) inhibitors to determine whether or not ATP-induced histamine release is associated with arachidonic acid (AA) release in rat peritoneal mast cells. ATP increased histamine release in a dose dependent manner, whereas adenosine did not. PPADS (a selective P2X-purinoceptor antagonist) and suramin (a nonselective P2X,2Y-purinoceptor antagonist) inhibited ATP-induced histamine release in a dose dependent manner. However, RB-2 (a P2Y-purinoceptor antagonist) did not block ATP-induced histamine release. Manoalide and oleyloxyethyl phosphorylcholine (OPC), secretory PLA$_2$ inhibitors, also inhibited ATP-induced histamine release dose-dependently. Both COX inhibitors (ibuprofen and indomethacin) and LOX inhibitors (baicalein and caffeic acid) inhibited ATP-induced histamine in a dose dependent manner. ATP significantly increased [$^3H$]AA release by 54%. PPADS and suramin significantly inhibited ATP-induced [3H]Ph release by 81% and 39%, respectively. ATP-induced histamine release was significantly inhibited by a variety of protein kinase inhibitors, such as bisindolmaleimide, genistein, methyl 2,5-dihydroxycinnamate, W-7 and trifluoperazine. Overall, the results suggest that ATP-induced histamine release is in part related to the PLA2-mediated AA metabolism and P2X-purinoceptors.

• Biomolecules & Therapeutics
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• 제16권3호
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• pp.190-196
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• 2008

Deoxypodophyllotoxin (DPT) is a medicinal herb product isolated from Anthriscus sylvestris. DPT possesses beneficial activities in regulating immediate-type allergic reaction and anti-inflammatory activity through the dual inhibition of cyclooxygenase-2 and 5-lipoxygenase. In the present study, the metabolism of DPT was further characterized in rat liver microsomes isolated from male Sprague Dawley rats. The metabolism of DPT was NADPH-dependent. In addition, when liver microsomes were incubated with SKF-525A, a well-known CYP inhibitor, in the presence of $\beta$-NADPH, the metabolism of DPT was significantly inhibited. Using enriched rat liver microsomes, the anticipated isoforms of cytochrome P450s (CYPs) in the metabolism of DPT were partially characterized. Phenobarbital-induced microsomes increased in the formation of metabolite M1. The metabolite M3 was only produced in the enriched microsomes isolated from dexamethasone-treated rats. The results indicated that the metabolism of DPT would be CYP-dependent and that CYP2B and CYP3A might be important in the metabolism of DPT in rats.