한수지 47(6), , 2014 Original Article Kor J Fish Aquat Sci 47(6), ,2014 RAW 세포에서의미역 (Undaria pinnatifida) 뿌리에탄올추출물의항염증활성 강보경 안나경 최연욱 김

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1 한수지 47(6), , 214 Originl Article Kor J Fish Aqut Sci 47(6), ,214 RAW 세포에서의미역 (Unri pinntifi) 뿌리에탄올추출물의항염증활성 강보경 안나경 최연욱 김민지 1 박시우 박원민 김보람 김꽃봉우리 1 안동현 * 부경대학교식품공학과 / 식품연구소, 1 부경대학교수산과학연구소 Anti-inflmmtory Activity of Ethnol Extrct of Unri pinntifi Root in RAW Cells Bo-Kyeong Kng, N-Kyung Ahn, Yeon-Uk Choi, Min-Ji Kim 1, Si-Woo Brk, Won-Min Pk, Bo-Rm Kim, Koth-Bong-Woo-Ri Kim 1 n Dong-Hyun Ahn* Deprtment of Foo Science n Technology/Institute of Foo Science, Pukyong Ntionl University, Busn , Kore 1 Institute of Fisheries Sciences, Pukyong Ntionl University, Busn , Kore The nti-inflmmtory effects of the ethnol extrct of Unri pinntifi root (UPREE) were investigte using the lipopolyscchrie (LPS)-inuce inflmmtory response in RAW cells y mesuring the prouction of nitric oxie (NO), interleukin (IL)-6, tumor necrosis fctor (TNF)-α, n IL-1β, n cell prolifertion. We foun tht NO levels were reuce y 34% t 1 μg/ml. Moreover, the prouction of IL-6 n TNF-α ws suppresse y the UPREE tretment. In prticulr, the IL-6 prouction ws inhiite y more thn 3% t 1 μg/ml UPREE. The prolifertion of RAW cells ws mesure y MTT ssy, n we foun no cytotoxicity in those cells trete with UPREE compre to the control. Our results suggest tht UPREE shows promise s therpeutic nti-inflmmtory tretment. Key wors: Unri pinntifi, Anti-inflmmtory Activity, IL-6, TNF-α 서론 (Unri pinntifi) (Choi et l., 1992; Kim n Choi, 1994).,. 4-6% (Ahn et l., 24).,. (Ahn et l., 24), (Jung, 213), (Cho et l., 24). (Jeong et l., 212; Choi n Kim, 213). (Olefsky n Glss, 21)., (Alin n Reichner, 1995). lipopolyscchrie (LPS) Kor J Fish Aqut Sci 47(6) , Decemer 214 This is n Open Access rticle istriute uner the terms of the Cretive Commons Attriution Non-Commercil Licens ( permits unrestricte non-commercil use, istriution, n reprouction in ny meium, provie the originl work is properly cite. Receive 14 August 214; Revise 22 Octoer 214; Accepte 17 Novemer 214 *Corresponing uthor: Tel: Fx: E-mil ress: hhn@pknu.c.kr Copyright 214 The Koren Society of Fisheries n Aqutic Science 751 pissn: , eissn:

2 752 강보경ㆍ안나경ㆍ최연욱ㆍ김민지ㆍ박시우ㆍ박원민ㆍ김보람ㆍ김꽃봉우리ㆍ안동현, nitric oxie (NO), tumor necrosis fctor (TNF)-, interleukin (IL)-1, IL-6 (Wu et l., 24; Vlleor et l., 21). (Dinrello, 1998),,,,,, (Willoughy, 1975).,,. (Kwk n Lee, 214).,., LPS RAW cells NO, IL-6, TNF- IL-1. 실험재료 재료및방법 ( ), (EyelFDU-21, Tokyo Rikkiki Co., Tokyo, Jpn) -2. 추출 1 95% (UNION 32R, Hnil Co., Incheon, Kore) 3, rpm filter pper (Avntec5A, Toyo Roshi Kish, Tokyo, Jpn). 35 wter th rotry evportor (RE2, Ymto Co., Tokyo, Jpn) 세포배양 RAW (KCLB471), DMEM (GIBCO, Grn Isln, NY, USA) 1% inctivte fetl ovine serum (FBS) 1% penicillin-streptomycin 37, 5% CO 2 incutor (MCO-15AC, Snyo, Osk, Jpn). 8-9%, 2 pssges. 세포독성측정 3-(4,5-imethylthizol- 2-yl)-2,5-iphenyltetrzolium romie (MTT) ssy. RAW cell (1 1 6 cells/ml) 96-well plte 2, 2 g/ml LPS (Sigm, St. Louis, MO, USA) (.1, 1, 1, 5, 1 g/ml) 37, 5% CO 2 22., 5 mg/ml MTT 2 4, 2, rpm 1., well imethyl sulfoxie (DMSO) microplte reer (Mo el 55, Bio-R, Richmon, CA, USA) 54 nm.. Prolifertion Inex (%)=smple /control 1 NO 분비량측정 NO nitrite griess. RAW DMEM cells/ml 24-well plte 37, 5% CO 2 incutor 2., RAW g/ml LPS (.1, 1, 1, 5, 1 g/ml) 24., griess (1% sulfnilmie.1% nphthylen imine ihyrochlorie, 1 1) 1, microplte reer 54 nm. NO soium nitrite (NNO 2 ). Pro-inflmmtory cytokine 분비량측정 IL-6, TNF- IL-1 cytokine ELISA-kit (BD Bioscience, Sn Jose, CA, USA). RAW DMEM cells/ml 24-well plte 37, 5% CO 2 incutor g/ml LPS ethnol 12., ELISA microplte nti-mouse IL-6, TNF- IL-1 mouse ntioy (ma) coting..5% tween 2 phosphte uffere sline (PBST), 1% FBS locking. PBST, microplte 2. PBST

3 미역뿌리에탄올추출물의항염증활성 Prolifertion inex Nitric oxie (µm) c 1 Smple (µg/ml) e (µg/ml) Fig. 1. Inhiitory effect of ethnol extrct of Unri pinntifi root on the prouction of prolifertion in RAW cells. - Mens with ifferent superscripts re significntly ifferent (P<.5). iotinylte nti-mouse IL-6, TNF- ma streptviinhorserish peroxise conjugte 1. IL-1, iotinylte nti-mouse IL-1 etection ntioy 1, streptviinhorserish peroxise conjugte 3., PBST, -phenyleneimine (OPD) microplte reer 49 nm. 통계처리 SAS progrm (Sttisticl nlyticl system V8.2, SAS Institute Inc., Cry, NC, USA) ANOVA, Duncn P<.5. 결과및고찰 미역뿌리에탄올추출물에의한세포독성평가 MTT ssy RAW ,.1, 1, 1, 5, 1 g/ml (Fig. 1). NO cytokine. 미역뿌리에탄올추출물에의한 NO 생성억제효과 NO NO synthse (NOS) L- Fig. 2. Inhiitory effect of ethnol extrct of Unri pinntifi root on the prouction of NO in RAW cells. -e Mens with ifferent superscripts re significntly ifferent (P<.5). rginine, (Weisz et l., 1996; Kim et l., 212). LPS pro-inflmmtory cytokine NO, NO (Stuehr et l., 1991). NO RAW LPS (.1, 1, 1, 5, 1 g/ml) NO., LPS 21 NO LPS 2%, 4%, 22%, 32%, 34% NO (P<.5), IC g/ml (Fig. 2). Choi et l. (28) 2 g/ml NO 21% NO. NO. 미역뿌리에탄올추출물에의한 RAW 세포의 IL-6 분비감소효과 RAW LPS IL-6, TNF- IL-1 proinflmmtory cytokine (Guh n Mckmn, 21). IL-6 T, (Akir et l.,199; Shn

4 754 강보경ㆍ안나경ㆍ최연욱ㆍ김민지ㆍ박시우ㆍ박원민ㆍ김보람ㆍ김꽃봉우리ㆍ안동현 et l., 29). IL-6 (Hur et l., 213). LPS RAW IL-6, LPS pg/ml, , 29.77, 2.61, 167.9, pg/ml IL-6 (Fig. 3), 5 1 g/ml 35% 38% RAW IL-6. Choi et l. (213) ethyl cette 5 g/ml 1 g/ml 63%,. 미역뿌리에탄올추출물에의한 RAW 세포의 TNF-α 분비감소효과 TNF- cytokine,, T,,, (Kim et l., 29). cytokine TNF- LPS, TNF- ELISA. RAW TNF- LPS 2,31 pg/ml..1-1 g/ml LPS, (Fig. 4). 1 g/ml 22%. ethyl cette 1 g/ml TNF- 21% (Choi et l., 213). NO IL-6 RAW TNF-. 미역뿌리에탄올추출물에의한 RAW 세포의 IL-1β 분비감소효과 IL-6, TNF- cytokine IL-1 NO,, B., T, B NK cell cytokine (Lee, 211). TNF-α (pg/ml) 2,5 2, 1,5 1, 5 f Fig. 4. Inhiitory effect of ethnol extrct of Unri pinntifi root on the prouction of TNF-α in RAW cells. -f Mens with ifferent superscripts re significntly ifferent (P<.5)..1 c (µg/ml) e 8 IL-6 (pg/ml) c c IL-1β (pg/ml) e (µg/ml) (µg/ml) Fig. 3. Inhiitory effect of ethnol extrct of Unri pinntifi root on the prouction of IL-6 in RAW cells. -e Mens with ifferent superscripts re significntly ifferent (P<.5). Fig. 5. Inhiitory effect of ethnol extrct of Unri pinntifi root on the prouction of IL-1β in RAW cells. - Mens with ifferent superscripts re significntly ifferent (P<.5).

5 미역뿌리에탄올추출물의항염증활성 755 IL-1 TNF-, (Lee, 29). LPS RAW IL-1, LPS 62.6 pg/ml, (.1, 1, 1, 5, 1 g/ml) (Fig. 5). LPS RAW NO, IL-6 TNF-. 사사 214 ( ). References Ahn SJ, Kim YS n Kwon PP. 24. Storge of wste-rown sewee n egrtion of lginte using microorgnism. J Environ Sci 13, Akir S, Hirno T, Tg T n Kishimoto T Biology of multifunctionl cytokines: IL 6 n relte molecules (IL 1 n TNF). FASEB J 11, Alin JE n Reichner JS Nitric oxie in inflmmtion n immunity. New Horiz 3, Cho JS, Prk IN, Heo JS n Lee YS. 24. Biosorption n esorption of hevy metls using Unri sp. Koren J Environ Agric 23, Choi HS, Kim SS, Kim JG n Kim WJ Effect of temperture on some qulity chrcteristics of queous extrcts of se mustr. Kore J Foo Sci Technol 26, Choi JS, Shin SH, H YM, Kim YC, Kim TB, Prk SM, Choi IS, Song HJ n Choi YJ. 28. Minerl contents n physiologicl ctivities of rie se tngle (Lminri jponic) collecte from Gijng n Wno in Kore. J Life Sci 18, Choi MW n Kim JI Anti-inflmmtory of ethyl cette frction isolte from Unri pinntifi on lipopolyscchries-stimulte RAW cells. Kor J Fish Aqut Sci 46, Dinrello CA Interleukin-1, Interleukin-1 receptors n interleukin-1 receptor ntgonist. Int Rev Immunol 16, Guh M n Mckmn N. 21. LPS inuction of gene expression in humn monocytes. Cell Signl 21, Hur JW, Cho EH, Lee BK, Lee UY n Yoon TJ The enhnce effect of Oplopnx eltus Nki on the immune system n ntitumor ctivity. Koren J Foo Nutr 26, Kw CS n Lee JH In vitro ntioxint n nti-inflmmtory effects of ethnol extrcts from sprout of evening primrose (Oenother lcinit) n gooseerry (Actinii rgut). J Koren Soc Foo Sci Nutr 43, x.oi.org/1.3746/jkfn Jung HD Effect of ietry supplementtion of fermente se mustr n sewee fusiforme y-proucts on prouctivity in poultry. Ph. D. Thesis, Konkuk University, Seoul, Kore. Jeong DH, Kim KBWR, Kng BK, Jung SA, Kim HJ, Jeong HY, Brk SW n Ahn DH Anti-inflmmtory ctivity of the Unri pinntifi wter extrct. J Appl Biol Chem 55, Kim SH, Yun JW, Lee YH n Cheon EJ. 29. Polymorphisms of tumor necrosis fctor-lph promoter gene in Kwski isese n reltion to the risk of coronry rtery lesion. Koren J Peitr 52, Kim WJ n Choi HS Development of comine methos for effective extrction of se mustr. Kore J Foo Sci Techonol 26, Kim YS, Lee SJ, Hwng JW, Kim EH, Prk PJ n Jeong JH Anti-inflmmtory effects of extrcts from Ligustrum ovlifolium H. leves on RAW mcrophges. J Koren Soc Foo Sci Nutr 41, org/1.3746/jkfn Lee E. 29. Effect of Ephere her on immunomoultory ctivity in lipopolyscchrie-expose rts n Rw cells. Koren J Plnt Res 22, Lee E Effects of Ixeris entt extrct on the prouction of pro inflmmtory cytokines in the LPS stimulte rt n Rw cells. Koren J Plnt Res 24, Olefsky JM n Glss CK. 21. Mcrophges, inflmmtion n insulin resistnce. Annu Rev Plnt Physiol 72, Shn J, Fu J, Zho Z, Kong X, Hung H, Luo L n Yin Z. 29. Chlorogenic ci inhiits lipopolyscchrie-inuce cyclooxygense-2 expression in RAW cells through suppressing NF-κB n JNK/AP-1 ctivtion. Int Immunophrmcol 9, Stuehr DJ, Cho HJ, Kwon NS, Weise MF n Nthn CF Purifiction n chrcteriztion of the cytokine-inuce mcrophge nitric oxie synthse: n FAD- n FMN- contining flvoprotein. Proc Ntl Ac Sci USA 88, Vlleor AF, Coml M, Sntmrí-Bi LF, Lloers J n Cel A. 21. Mcrophge proinflmmtory ctivtion n ectivtion: question of lnce. Av Immunol 18, 1-2. Weisz A, Cictiello L n Esumi H Regultion of the mouse inucile-type nitric oxie synthse gene promoter y interferon-γ, cteril lipopolyscchrie n N G -

6 756 강보경ㆍ안나경ㆍ최연욱ㆍ김민지ㆍ박시우ㆍ박원민ㆍ김보람ㆍ김꽃봉우리ㆍ안동현 monomethyl-ʟ-rginine. Biochem J 316, Willoughy DA Humn rthritis pplie to niml moels. Towrs etter therpy. Ann Rheum Dis 34, Wu MJ, Wng L, Ding HY, Weng CY n Yen JH. 24. Glossogyne tenuifoli cts to inhiit inflmmtory meitor. J Biome Sci 11,

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untitled 3. 농업환경연구과 과제구분 기본연구 수행시기 전반기 연구과제 및 세부과제 수행 기간 소 속 책임자 농가에 적합한 부식성곤충 대량 사육기술 개발 12~ 13 농업환경연구과 곤충팀 이영혜 1) 부식성 곤충 먹이 제조 기술 개발 12~ 13 농업환경연구과 곤충팀 이영혜 색인용어 부식성곤충, 장수풍뎅이, 계통, 먹이제조 ABSTRACT In first check,