Food Science and Preservation (한국식품저장유통학회지)

The Korean Society of Food Preservation (한국식품저장유통학회)
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Effect of garlic (Allium sativum L.) stems on inflammatory cytokines, iNOS and COX-2 expressions in Raw 264.7 cells induced by lipopolysaccharide

Lipopolysaccharide로 유도된 Raw 264.7 세포에서 마늘대 추출물(Allium sativum L. Stems)의 염증성 사이토카인 및 iNOS, COX-2 발현에 대한 효과 검증

  • Cho, Yong Hun (Department of Cosmeceutical Science, Daegu Hanny University) ;
  • Kim, Hyeon Jeong (Department of Cosmeceutical Science, Daegu Hanny University) ;
  • Kim, Dong In (Department of Cosmeceutical Science, Daegu Hanny University) ;
  • Jang, Jae Yoon (Department of Cosmeceutical Science, Daegu Hanny University) ;
  • Kwak, Jae Hoon (Department of Cosmeceutical Science, Daegu Hanny University) ;
  • Shin, Yu Hyeon (Department of Cosmeceutical Science, Daegu Hanny University) ;
  • Cho, Yeon Gje (School of Food Science and Biotechnology / Food and Bio-Industry Research Institute, Kyungpook National University) ;
  • An, Bong Jeon (Department of Cosmeceutical Science, Daegu Hanny University)
  • 조용훈 (대구한의대학교 화장품약리학과) ;
  • 김현정 (대구한의대학교 화장품약리학과) ;
  • 김동인 (대구한의대학교 화장품약리학과) ;
  • 장재윤 (대구한의대학교 화장품약리학과) ;
  • 곽재훈 (대구한의대학교 화장품약리학과) ;
  • 신유현 (대구한의대학교 화장품약리학과) ;
  • 조영제 (경북대학교 식품공학부/식품생물산업연구소) ;
  • 안봉전 (대구한의대학교 화장품약리학과)
  • Received : 2015.03.12
  • Accepted : 2015.07.15
  • Published : 2015.08.30
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Abstract

In this study, the anti-oxidant and anti-inflammatory activities of water extract (ASSW) and 70% ethanol extract (ASSE) of Allium sativum L. stems were investigated using Raw 264.7 cells induced by lipopolysaccharide (LPS). ABTS radical scavenging activities of ASSW and ASSE at $1000{\mu}g/mL$ concentration were 96.9% and 97.8%, respectively. In order to investigate the potential anti-inflammatory effects of ASSW and ASSE, nitric oxide (NO), pro-inflammatory cytokines, interleukin-6 (IL-6), and tumor necrosis factor including ${\alpha}$ (TNF-${\alpha}$), interleukin-$1{\beta}$ (IL-$1{\beta}$), and prostaglandin-E2 (PGE2) were measured. ASSW and ASSE at $100{\mu}g/mL$ concentration showed inhibitory effects against NO production by 18% and 23%, respectively. Production of IL-$1{\beta}$ and IL-6 after treatment with ASSW and ASSE at $100{\mu}g/mL$ decreased by approximately 28% and 15% for ASSW and 17% and 12% for ASSE, respectively. In addition, production of TNF-${\alpha}$ after treatment of $100{\mu}g/mL$ of ASSW and ASSE decreased by 24% and 23%, respectively. In addition, the treatment of $100{\mu}g/mL$ of ASSW and ASSE showed inhibitory expressions against PGE2 by 45.47% and 33.87%, respectively. These results suggested that ASSE showed greater inhibitory activity than that of the ASSW by the suppression of inflammatory mediators, including NO, IL-6, TNF-${\alpha}$ and PGE2 production, and the expressions of iNOS and COX-2 in macrophages. In conclusion, ASSW and ASSE may have some ancillary effects on inflammatory factors as potential anti-inflammatory agents.

본 연구에서는 마늘의 부산물로 발생하는 마늘대의 항산화 및 항염증 효과를 알아보기 위하여 LPS로 염증을 유도한 Raw 264.7 세포에 대한 열수추출물(ASSW)과 70% 에탄올 추출물(ASSE)의 효과를 살펴보았다. ASSW의 폴리페놀 함량은 $37.08{\pm}1.51mg(TAE)/g$, ASSE의 폴리페놀 함량은 $44.7{\pm}1.32mg(TAE)/g$ 이 함유되어있음을 확인하였다. DPPH 실험과 $ABTS^+$ 실험에서 ASSW, ASSE 모두 농도 의존적으로 증가하였으며, DPPH의 경우 $1,000{\mu}g/mL$에서 대조군인 Vit.C의 $50{\mu}g/mL$의 항산화능이 있다는 것이 확인되었고, $ABTS^+$의 경우 $500{\mu}g/mL$ 이상부터 대조군인 Vit.C와 비슷한 효과를 나타냄으로서 ASSW, ASSE 모두 항산화능이 있다는 것을 확인하였다. MTT측정으로 인해 세포 독성을 가지지 않았던 농도대(5, 10, 25, 50, $100{\mu}g/mL$)에서 염증 억제 효과를 살펴보기 위해 NO를 측정한 결과 ASSW, ASSE 모두 $25{\mu}g/mL$에서부터 유의적으로 분비량이 감소함을 확인하였으며 특히 $100{\mu}g/mL$의 농도에서 약 18%, 23%의 억제 효과를 보였다. 또한 대식세포의 염증성 cytokine인 IL-6, TNF-${\alpha}$, IL-$1{\beta}$ 및 $PGE_2$의 분비량을 첨가 농도 의존적으로 억제함을 확인하였다. 특히 $PGE_2$에 대해 ASSW, ASSE $100{\mu}g/mL$의 농도에서 약 55%, 60%의 감소효과를 보였다. ASSW의 iNOS, COX-2의 발현 저해효과는 나타내지 못하였지만, ASSE는 $100{\mu}g/mL$의 농도에서 iNOS의 발현량이 현저하게 억제됨을 확인하였고, COX-2의 경우 농도 의존적으로 저하되어 특히 $50{\mu}g/mL$, $100{\mu}g/mL$의 구간에서 단백질 발현 저해효과가 있음을 확인하였다. 이를 통해 ASSW, ASSE 모두 항산화 효과와 항염증 효과가 있음을 확인하였으며, ASSW 보다 ASSE에서 활성산소종(reactive oxygen species, ROS) 및 ROS에 의해 유발되는 염증을 억제시켜주는 소재가 될수있을 것이라 예상된다.

Keywords

References

  1. Haddad JJ (2002) Antioxidant and prooxidant mechanisms in the regulation of redox (y)-sensitive transcription factor. Cell Signal, 14, 879-897 https://doi.org/10.1016/S0898-6568(02)00053-0
  2. Beckman KB, Ames BN (1998) The free radical theory of aging matures. Physiol Rev, 78, 547-581
  3. Balaban RS, Nemoto S, Finkel T (2005) Mitochondria, oxidants, and aging. Cell, 120, 483-495 https://doi.org/10.1016/j.cell.2005.02.001
  4. Sohal RS, Weindruch R (1996) Oxidative stress, caloric restriction, and aging. Science, 273, 59-63 https://doi.org/10.1126/science.273.5271.59
  5. Davies KJ (2000) Oxidative stress, antioxidant defenses, and damage removal, repair, and replacement systems. IUBMB Life, 50, 279-289 https://doi.org/10.1080/15216540051081010
  6. Medzhitov R (2008) Origin and physiological roles of inflammation. Nature, 454, 428-435 https://doi.org/10.1038/nature07201
  7. Guha M, Mackman N (2001) LPS induction of gene expression in human monocytes. Cell Signal, 13, 85-94 https://doi.org/10.1016/S0898-6568(00)00149-2
  8. Coussens LM, Werb Z (2002) Inflammation and cancer. Nature, 420, 860-867 https://doi.org/10.1038/nature01322
  9. Posadas I, Terencio MC, Guillén I, Ferrándiz ML, Coloma J, Payá M, Alcaraz MJ (2000) Co-regulation between cyclo-oxygenase-2 and inducible nitric oxide synthase expression in the time-course of murine inflammation. N-S Arc Pharmacol, 361, 98-106 https://doi.org/10.1007/s002109900150
  10. Cencioni C, Spallotta F, Martelli F, Valente S, Mai A, Zeiher AM, Gaetano C (2013) Oxidative stress and epigenetic regulation in ageing and age-related diseases. Int J Mol Sci, 14, 17643-17663 https://doi.org/10.3390/ijms140917643
  11. Kundu JK, Surh YJ (2008) Inflammation: gearing the journey to cancer. Mutat Res-Rev Mutat, 659, 15-30 https://doi.org/10.1016/j.mrrev.2008.03.002
  12. Lee TB (1979) Illustrated flora of Korea, Hangmunsa, Seoul, Korea, p 203
  13. Jo JS, Hwang SY (1990) Gijeunyungusa, Seoul, Korea, p 154-155
  14. Bozin B, Mimica-Dukic N, Samojlik I, Goran A, Igic R (2008) Phenolics as antioxidants in garlic (Allium sativum L., alliaceae). J Agric Food Chem, 111, 925-929 https://doi.org/10.1016/j.foodchem.2008.04.071
  15. Lee MK, Park JS, Na HS (2005) Proximate compositions of green garlic powder and microbiological properties of bread with green garlic. Korean J Food Preserv, 12, 95-100
  16. Folin O, Denis W (1912) On phosphotungasticphosphomolybetic compounds as color regents. J Biol Chem, 12, 239-249
  17. Carmichael J, DeGraff WG, Gazdar AF, Minna JD, Mitchell HB (1987) Evaluation of a tetrazolium based semiautomated colorimetric assay : assessment of chemosensitivity testing. Cancer Res, 47, 936-942
  18. Ding AH, Nathan CF, Stueher DJ (1988) Release of reactive nitrogen intermediates and macrophages. Comparison of activation cytokines and evidence for independent production. J Immunol, 141, 2407-2412
  19. Duthie G, Crozier A (2000) Plant-derived phenolic antioxidants. Curr Opin Clin Nutr Metab Care, 3, 447-451 https://doi.org/10.1097/00075197-200011000-00006
  20. Ferreres F, Gomes D, Valentao P, Goncalves R, Pio R, Chagas EA, Seabra RM, Andrade PB (2009) Improved loquat (Eriobotrya japonica Lindl.) cultivars : variation of phenolics and antioxidative potential. Food Chem, 114, 1019-1027 https://doi.org/10.1016/j.foodchem.2008.10.065
  21. Kromhout D (1987) Essential micronutrients in relation to carcinogenesis. Am J Clin Nutr, 45, 1361-1467
  22. Park SJ, Park DH, Kim SS, Gon J, Lee HY (2009) Chemical compositions of fermented Codonopsis lanceolata. J Food Sci Nutr, 38, 396-400
  23. Moon JS, Kim SJ, Par YM, Hwang IS, Kim EY, Park JW, Park IB, Kim SW, Kang WG, Park YK, Jung ST (2004) Antimicrobial effect of methanol extracts from some medicinal herbs and the content of phenolic compounds. Korean J Food Preserv, 11, 207-213
  24. Molyneux P (2004) The use of the stable free radical diphenylpicrylhydrazyl (DPPH) for estimating antioxidant activity. Songklanakarin J Sci Technol, 26, 211-219
  25. Lu Y, Foo LY (2000) Antioxidant and radical scavenging activities of polyphenols from apple pomace. J Agric Food Chem, 68, 81-85 https://doi.org/10.1016/S0308-8146(99)00167-3
  26. Rice-Evans CA, Miller NJ, Paganga G (1996) Structureantioxidant activity relationships of flavonoids and phenolic acids. Free Radic Biol Med, 20, 933-956 https://doi.org/10.1016/0891-5849(95)02227-9
  27. Rice-Evans CA, Miller NJ, Bolwell GP, Bramley PM, Pridham JB (1995) The relative antioxidant activities of plant-derived polyphenolic flavonoids. Free Radic Res, 22, 375-383 https://doi.org/10.3109/10715769509145649
  28. Miller NJ, Sampson J, Candeias LP, Bramley PM, Rice-Evans CA (1996) Antioxidant activities of carotenes and xanthophylls. FEBS Lett, 384, 240-242 https://doi.org/10.1016/0014-5793(96)00323-7
  29. Weisz A, Cicatiello L, Esumi H (1996) Regulation of the mouse inducible-type nitric oxide synthase gene promoter by interferongamma, bacterial lipopolysaccharide and NG-monomethyl-L-arginine. Biochem J, 316, 209-215 https://doi.org/10.1042/bj3160209
  30. An SM, Kim HG, Choi EJ, Hwang HH, Lee ES, Baek JH, Boo YC, Koh JS (2014) Screening for antiinflammatory activities in extracts from Korean herb medicines. J Soc Cosmet Scientists Korea, 40, 95-108 https://doi.org/10.15230/SCSK.2014.40.1.95
  31. Kang CH, Choi YH, Choi IW, Lee JD, Kim GY (2011) Inhibition of lipopolysaccharide-induced iNOS, COX-2, and TNF-${\alpha}$ expression by aqueous extract of Orixa japonica in Raw 264.7 cells via suppression of NF-${\kappa}B$ activity. Trop J Pharm Res, 10, 161-168
  32. Shon MS, Song JH, Kim JS, Jang HD, Kim GN (2013) Anti-oxidant activity of oil extracted from Korean red ginseng and its moisturizing function. Korean J Aesthet Cosmetol, 11, 489-494
  33. Lee HJ, Sim BY, Bak JW, Kim DH (2014) Effect of Gami-sopungsan on inflammation and DNCB-induced dermatitis in NC/Nga in mice. Korean J Orient Physiol Pathol, 28, 146-153
  34. Kuo, Li C, Chi CW, Liu TY (2004) The antiinflammatory potential of berberine in vitro and in vivo. Cancer letters, 203, 127-137 https://doi.org/10.1016/j.canlet.2003.09.002
  35. Nilsson G, Svensson V, Nilsson K (1995) Constitutive and inducible cytokine mRNA expression in the human mast cell line HMC-1. Scand J Immunol, 42, 76-81 https://doi.org/10.1111/j.1365-3083.1995.tb03628.x
  36. Papanicolaou DA, Wilder RL, Manolagas SC, Chrousos GP (1998) The pathophysiologic roles of interleukin-6 in human disease. Ann Intern Med, 128, 127-137 https://doi.org/10.7326/0003-4819-128-2-199801150-00009
  37. Zhang Y, Ramos BF, Jakschik BA (1992) Neutrophil recruitment by tumor necrosis factor from mast cells inimmune complex peritonitis. Science, 258, 1957-1959 https://doi.org/10.1126/science.1470922
  38. Roitt I, Brostoff J, Male D (2002) Immunology, 6th ed, London : Mosby
  39. Hur GM, Ryu YS, Yun HY, Jeon BH, Kim YM, Seok JH, Lee JH (1999) Hepatic ischemia/reperfusion in rats induces iNOS gene transcription by activation of NF-kappaB. Biochem Biophys Res Commun, 261, 917-922 https://doi.org/10.1006/bbrc.1999.1143
  40. Sato T, Nakajima H, Fujio K, Mori Y (1997) Enhancement of prostaglandin E2 production by epidermal growth factor requires the coordinate activation of cytosolic phospholipase A2 and cyclooxygenase 2 in human squamous carcinoma A431 cells. Prostaglandins, 53, 355-369
  41. Huang M, Stolina M, Sharma S, Mao JT, Zhu L, Miller PW, Dubinett SM (1998) Non-small cell lung cancer cyclooxygenase-2-dependent regulation of cytokine balance in lymphocytes and macrophages : up-regulation of interleukin 10 and down-regulation of interleukin 12 production. Cancer Res, 58, 1208-1216

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