The Korea Journal of Herbology (대한본초학회지)

The Korea Association of Herbology (대한본초학회)
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The study of anti-inflammatory effect of Hyeonto-dan extract in RAW 264.7 macrophage

현토단(玄兎丹)의 RAW 264.7 대식 세포에서의 항염증 효과에 관한 연구

  • Kim, Ma-Ryong (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University) ;
  • Kang, Ok-Hua (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University) ;
  • Kong, Ryong (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University) ;
  • Seo, Yun-Soo (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University) ;
  • Zhou, Tian (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University) ;
  • Kim, Sang-A (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University) ;
  • Kim, Eun-Su (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University) ;
  • Sin, Min-A (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University) ;
  • Lee, Young-Seob (Department of Herbal Crop Research, National Institute of Horticultural & Herbal Science, RDA) ;
  • Kwon, Dong-Yeul (Department of Oriental Pharmacy, College of Pharmacy, Wonkwang Oriental Medicines Research Institute, Wonkwang University)
  • 김마룡 (원광대학교 약학대학 한약학과, 원광한약연구소) ;
  • 강옥화 (원광대학교 약학대학 한약학과, 원광한약연구소) ;
  • 공룡 (원광대학교 약학대학 한약학과, 원광한약연구소) ;
  • 서윤수 (원광대학교 약학대학 한약학과, 원광한약연구소) ;
  • 주전 (원광대학교 약학대학 한약학과, 원광한약연구소) ;
  • 김상아 (원광대학교 약학대학 한약학과, 원광한약연구소) ;
  • 김은수 (원광대학교 약학대학 한약학과, 원광한약연구소) ;
  • 신민아 (원광대학교 약학대학 한약학과, 원광한약연구소) ;
  • 이영섭 (농촌진흥청 국립원예특작과학원 인삼특작부 인삼특작이용팀) ;
  • 권동렬 (원광대학교 약학대학 한약학과, 원광한약연구소)
  • Received : 2017.01.19
  • Accepted : 2017.03.15
  • Published : 2017.03.30
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Abstract

Objectives : This study aimed to investigate the unknown mechanisms behind the anti- inflammatory activity of Hyeonto-dan(HT) 70% ethanol extract on LPS-stimulated RAW 264.7 cells. Methods : Cells were treated with Hyeonto-dan 1 h prior to addition of 200 ng/mL of LPS. Cell viability was measured by the MTS assay. Nitric oxide levels were determined by the Griess assay. $PGE_2$ were measured using EIA kit. Pro-inflammatory cytokine production was measured by the enzyme-linked immunosorbent assay (ELISA). The expression of COX-2, iNOS, and MAPKs was investigated by Western blot, qRT-PCR. $NF-{\kappa}B$/p65 localization and interaction of the TLR-4 receptor with LPS was examined by immunofluorescence assays. Results : Hyeonto-dan had no cytotoxicity at the measured concentration. Hyeonto-dan inhibited NO production and pro-inflammatory cytokines such as IL-6, $TNF-{\alpha}$, and PGE2 as well as the protein and mRNA expression of iNOS and COX-2. Moreover, Hyeonto-dan inhibited the interaction between LPS and TLR-4 in murine macrophages. It suppressed phosphorylation of extracellular signal-regulated kinase (ERK 1/2), c-jun N-terminal kinase (JNK 1/2) and p38. Finally, it inhibited translocation of $NF-{\kappa}B$ in response to competitive LPS. Conclusions : Based on the results of this study, Hyeonto-dan inhibited the binding of TLR-4 receptor to LPS and inhibited the phosphorylation of extracellular signaling pathway MAPKs. These inhibitory effects are thought that the amount of $NF-{\kappa}B$ delivered to the nucleus was decreased and the inflammatory reaction was prevented by decreasing the production of LPS-induced $PGE_2$, NO, IL-6 and $TNF-{\alpha}$.

Keywords

References

  1. Kim Yang-Ho. Pathology. Korean Society of Pathlogists : Gomunsa. 1991 : 71-92.
  2. Laroux FS. Mechanisms of inflammation : the good, the bad and the ugly. Frontiers in Bioscience. 2004. 9 : 3156-62. https://doi.org/10.2741/1468
  3. Barnes P.J. and M. Karin. Nuclear factor-kB : a pivotal transcription factor in chronic inflammatory diseases. New England Jounal of Medicine. 1997 ; 336(15) : 1066-71. https://doi.org/10.1056/NEJM199704103361506
  4. Lee J.I. and G.J. Burckart. Nuclear factor kappa B : important transcription factor and therapeutic target. The Journal of Clinical Pharmacology. 1998 ; 38(11) : 981. https://doi.org/10.1177/009127009803801101
  5. Grover A, Shandilya A, Punetha A, Bisaria VS, Sundar D. Inhibition of the $NEMO/IKK{\beta}$ association complex formation, a novel mechanism associated with the $NF-{\kappa}B$ activation suppression by Withania somnifera's key metabolite withaferin A. BMC Genomics. 2010 ; 11(4) : 25. https://doi.org/10.1186/1471-2164-11-25
  6. Tak P.P. and G.S. Firestein. NF-kappaB : a key role in inflammatory diseases. Journal of Clinical Investigation. 2001 ; 107(1) : 7-12. https://doi.org/10.1172/JCI11830
  7. Sun SC, Chang JH, Jin J. Regulation of nuclear $factor-{\kappa}B$ in autoimmunity. Trends Immunol. 2013 ; 34(6) : 282-9. https://doi.org/10.1016/j.it.2013.01.004
  8. Kim Seung-Hyun, Kong Gu. Role of Immune-Inflammatory Mechanisms in Alzheimer's Disease and Other Degenerative Diseases. Journal of the Korean Neurological Association. 2002 ; 20(6) : 575-84 .
  9. Leon CG, Tory R, Jia J, Sivak O, Wasan KM. Discovery and Development of Toll-Like Receptor-4 (TLR4) Antagonists. A New Paradigm for Treating Sepsis and Other Diseases : Pharmaceutical Research. 2008 ; 25(8) :1751-61. https://doi.org/10.1007/s11095-008-9571-x
  10. Ui Yeok-lim. Seuideukhyobang. Uiseongdang. 1990 : 220.
  11. Heo Joon. Donguibogam 3rd ed. Yeogang Publishing Co. 2005 : 2010.
  12. Oky Maeng, Yong Chan Kim, Young Sang Kim, Sang-Gi Paik, Hayyoung Lee. Identification of a gene that is up-regulated in RAW264.7 macrophage cells by nitric oxide. Bulletin of Biotechnology Chungnam National University. 2002 ; 8(2) : 1-9.
  13. Mu MM, Chakravortty D, Sugiyama T, Koide N, Takahashi K, Mori I, Yoshida T, Yokochi T. The inhibitory action of quercetin on lipopolysaccharide-induced nitric oxide production in RAW 264.7 macrophage cells : J. Endotoxin Res. 2001 ; 7(6) : 431-8. https://doi.org/10.1177/09680519010070060601
  14. Dubois R.N et al. Cyclooxygenase in biology and disease. The Federation of American Societies for Experimental Biology Journal. 1998 ; 12(12) : 1063. https://doi.org/10.1096/fasebj.12.12.1063
  15. Bosca L, Zeini M, Traves PG, Hortelano S. Nitric oxide and cell viability in inflammatory cells : a role for NO in macrophage function and fate. Toxicology. 2005 ; 208(2) : 249-58. https://doi.org/10.1016/j.tox.2004.11.035
  16. Qi M, Elion EA. MAP kinase pathways. J Cell Sci. 2005 ; 118 : 3569-72. https://doi.org/10.1242/jcs.02470
  17. Gray Pearson, Fred Robinson, Tara Beers Gibson, Bing-e Xu, Mahesh Karandikar, Kevin Berman, and Melanie H. Cobb. Mitogen-Activated Protein (MAP) Kinase Pathways : Regulation and Physiological Functions. Endocrine reviews. 2001 ; 22(2) : 153-83. https://doi.org/10.1210/er.22.2.153
  18. Guo AK, Hou YY, Hirata H, Yamauchi S, Yip AK, Chiam KH et al. Loss of p53 enhances NF-B-dependent lamellipodia formation. J Cell Physiol. 2014 ; 229(6) : 696-704. https://doi.org/10.1002/jcp.24505
  19. Kaisho T, Akira S. Toll-like receptor function and signaling. J Allergy Clin Immunol. 2006 ; 117(5) : 979-87. https://doi.org/10.1016/j.jaci.2006.02.023
  20. Miller SI, Ernst RK, Bader MW. LPS, TLR4 and infectious disease diversity. Nat Rev Microbiol. 2005 ; 3(1) : 36-46. https://doi.org/10.1038/nrmicro1068