Journal of Applied Biological Chemistry

The Korean Society for Applied Biological Chemistry (한국응용생명화학회)
권호 표지
DOI QR코드

DOI QR Code

Anti-inflammation effect of rebaudioside A by inhibition of the MAPK and NF-κB signal pathway in RAW264.7 macrophage

RAW264.7 대식세포에서 MAPK 및 NF-κB 신호전달 억제를 통한 rebaudioside A의 항염 효과

  • Choi, Da Hee (School of Cosmetic Science and Beauty Biotechnology, Semyung University) ;
  • Cho, Uk Min (School of Cosmetic Science and Beauty Biotechnology, Semyung University) ;
  • Hwang, Hyung Seo (School of Cosmetic Science and Beauty Biotechnology, Semyung University)
  • Received : 2018.05.11
  • Accepted : 2018.06.11
  • Published : 2018.06.30
Download PDF
⟨ Previous Next ⟩

Abstract

Rebaudioside A is a natural sweetener isolated from Stevia rebaudiana Bertoni, one of the glycosides based on steviol. Recent studies have shown that rebaudioside A inhibits the inflammatory response by inhibiting cytokines secretion such as interleukin-$1{\alpha}/1{\beta}$ in activated RAW264.7 mouse macrophage cells by LPS. However, the inhibitory mechanism of inflammation by rebaudioside A in the presence of LPS has not been fully elucidated. Therefore, in this study, we tried to investigate the anti-inflammatory activity of rebaudioside A at the protein level when RAW264.7 cells were stimulated by LPS. The inducible nitric oxide synthase protein expression level was reduced in the group treated with $250{\mu}M$ rebaudioside A compared to the LPS-treated group. In addition, the mRNA expression level of $NF-{\kappa}B$, which is a representative nuclear transcription factor by inflammatory signal, was also decreased as compared with that of LPS-treated group. In addition, $NF-{\kappa}B$ and inhibitor-${\kappa}B$ ($I-{\kappa}B$) complexes that are known to be dissociated by $I-{\kappa}B$ phosphorylation and ubiquitination were less phosphorylated than LPS treated group in the presence rebaudioside A. Finally, we could find that rebaudioside A was involved in the $NF-{\kappa}B$ pathway through reducing extracellular signal-regulated kinase1/2 phosphorylation in a concentration-dependent manner. These results suggest that rebaudioside A might suppress inflammatory reaction through MAPK and $NF-{\kappa}B$ regulation in LPS-stimulated RAW264.7.

리바우디오사이드 A는 Stevia rebaudiana Bertoni에서 분리된 천연감미료로 널리 알려진 스테비올배당체 중 하나이다. 최근 연구에서 LPS 자극에 의해 활성화된 RAW264.7 마우스 대식 세포에서 리바우디오사이드 A가 인터루킨-$1{\alpha}/1{\beta}$ 같은 염증성 사이토카인 분비를 억제하는 기능이 확인되었다. 그러나 LPS처리 시 리바우디오사이드 A의 항염 활성에 대한염증 억제기작은 정확히 제시하지 못하였다. 따라서 본 연구에서는 리바우디오사이드 A의 LPS 신호전달 메카니즘에서의 항염증 효능을 단백질 수준에서 규명하고자 하였다. NO 생성에 관여하는 iNOS 단백질 발현양을 분석한 결과 리바우디오사이드 A의 $250{\mu}M$ 처리군에서 농도 의존적으로 단백질 발현이 감소하는 것을 확인하였다. 또한 염증 신호에 의한 대표적 핵 전사 인자인 $NF{\kappa}B$의 mRNA 발현량 분석 결과에서도 LPS 처리군에 비해 그 발현양이 감소하였다. 또한 세포질에 존재하는 $NF-{\kappa}B$와 $I-{\kappa}B$ 복합체는 LPS신호에 의한 $I-{\kappa}B$의 인산화 및 ubiquitination로 인해 $NF-{\kappa}B$가 이탈되기 때문에, 리바우디오사이드 A에 의한 $pNF-{\kappa}B$, $pI-{\kappa}B$의 단백질 발현을 비교 분석한 결과 $NF-{\kappa}B$ 단백질의 인산화가 농도 의존적으로 감소하였고, $I-{\kappa}B$의 인산화 또한 저해되는 것을 확인 하였다. 최종적으로 리바우디오사이드 A는 LPS처리 조건에서 MAPK중 특이적으로 extracellular signal-regulated kinase (ERK1/2)의 인산화를 농도 의존 방식으로 감소시킴으로써 $NF-{\kappa}B$ 조절 기작에 관여함을 알 수 있었다. 따라서 본 연구 결과들을 통해 우리는 리바우디오사이드 A가 RAW264.7 세포에서 LPS에 의해 활성화 되는 MAPK 및 $NF{\kappa}B$의 발현 억제를 통해 염증이 억제될 수 있음을 확인하였다.

Keywords

References

  1. Kundu JK, Surh YJ (2008) Inflammation: gearing the journey to cancer. Mutat Res Rev Mutat Res 659: 15-30 https://doi.org/10.1016/j.mrrev.2008.03.002
  2. Kim CH, Lee MA, Kim TW, Jang JY, Kim HJ (2012) Anti-inflammatory effect of Allium hookeri root methanol extract in LPS-induced RAW264.7 cells. J Korean Soc Food Sci Nutr 41: 1645-1648 https://doi.org/10.3746/jkfn.2012.41.11.1645
  3. Middleton E (1998) Effect of plant flavonoids on immune and inflammatory cell function. In: Flavonoids in the living system. Springer, Boston
  4. Nijveldt RJ, Van Nood ELS, Van Hoorn DE, Boelens, PG Van Norren K, Van Leeuwen PA (2001) Flavonoids: a review of probable mechanisms of action and potential applications. Am. J Clin Nutr 74: 418-425 https://doi.org/10.1093/ajcn/74.4.418
  5. Shutenko Z, Henry Y, Pinard E, Seylaz J, Potier P, Berthet F, Girard P, Sercombe R (1999) Influence of the antioxidant quercetin in vivo on the level of nitric oxide determined by electron paramagnetic resonance in rat brain during global ischemia and reperfusion. Biochem Pharmacol 57: 199-208 https://doi.org/10.1016/S0006-2952(98)00296-2
  6. Friesenecker B, Tsai AG, Allegra C, Intaglietta M (1994) Oral administration of purified micronized flavonoid fraction suppresses leukocyte adhesion in ischemia-reperfusion injury: in vivo observations in the hamster skin fold. J Vasc Res 14: 50-55
  7. Hamid Q, Boguniewicz M, Leung DY (1994) Differential in situ cytokine gene expression in acute versus chronic atopic dermatitis. J Clin Investig 94: 870-876 https://doi.org/10.1172/JCI117408
  8. Howell MD, Gallo RL, Boguniewicz M, Jones JF, Wong C, Streib JE, Leung DY (2006) Cytokine milieu of atopic dermatitis skin subverts the innate immune response to vaccinia virus. Immunity 24: 341-348 https://doi.org/10.1016/j.immuni.2006.02.006
  9. Wong CK, Leung KML, Qiu HN, Chow JYS, Choi AOK, Lam CWK (2012) Activation of eosinophils interacting with dermal fibroblasts by pruritogenic cytokine IL-31 and alarmin IL-33: implications in atopic dermatitis. PloS one 7: e29815 https://doi.org/10.1371/journal.pone.0029815
  10. Ruimi N, Rwashdeh H, Wasser S, Konkimalla B, Efferth T, Borgatti M, Roberto G, Mahajna J (2010) Daedalea gibbosa substances inhibit LPSinduced expression of iNOS by suppression of $NF-{\kappa}B$ and MAPK activities in RAW264.7 macrophage cells. Int J Mol Med 25: 421-432
  11. Bode JG, Ehlting C, Haussinger D (2012) The macrophage response towards LPS and its control through the p38MAPK-STAT3 axis. Cellular signalling 24: 1185-1194 https://doi.org/10.1016/j.cellsig.2012.01.018
  12. Karin M, Ben-Neriah Y (2000) Phosphorylation meets ubiquitination: the control of $NF-{\kappa}B$ activity. Annu Rev Immunol 18: 621-663 https://doi.org/10.1146/annurev.immunol.18.1.621
  13. Surh YJ, Chun KS, Cha HH, Han SS, Keum YS, Park KK, Lee SS (2001) Molecular mechanisms underlying chemopreventive activities of anti-inflammatory phytochemicals: down-regulation of COX-2 and iNOS through suppression of $NF-{\kappa}B$ activation. Mutat Res 480: 243-268
  14. Roberts PJ, Der CJ (2007) Targeting the Raf-MEK-ERK mitogenactivated protein kinase cascade for the treatment of cancer. Oncogene 26: 3291-3310 https://doi.org/10.1038/sj.onc.1210422
  15. Reber L, Vermeulen L, Haegeman G, Frossard N (2009) Ser276 phosphorylation of $NF-{\kappa}B$ p65 by MSK1 controls SCF expression in inflammation. PloS one 4: e4393 https://doi.org/10.1371/journal.pone.0004393
  16. Stoyanova S, Geuns J, Hideg E, Van Den Ende W (2011) The food additives inulin and stevioside counteract oxidative stress. Int J Food Sci Nutr 62: 207-214 https://doi.org/10.3109/09637486.2010.523416
  17. Fengyang L, Yunhe F, Bo L, Zhicheng L, Depeng L, Dejie L, Wen Z, Yongguo C, Naisheng Z, Xichen Z, Zhengtao Y (2012) Stevioside suppressed inflammatory cytokine secretion by downregulation of $NF-{\kappa}B$ and MAPK signaling pathways in LPS-stimulated RAW264.7 cells. Inflammation 35: 1669-1675 https://doi.org/10.1007/s10753-012-9483-0
  18. Cho UM, Hwang HS (2017) Anti-inflammatory Effects of rebaudioside A in LPS-stimulated RAW264.7 Macrophage Cells. J Soc Cosmet Scientists Korea 43: 157-164
  19. Chi G, Wei M, Xie X, Soromou LW, Liu F, Zhao S (2013) Suppression of MAPK and $NF-{\kappa}B$ pathways by limonene contributes to attenuation of lipopolysaccharide-induced inflammatory responses in acute lung injury. Inflammation 36: 501-511 https://doi.org/10.1007/s10753-012-9571-1
  20. Shi Y, Tu Z, Tang D, Zhang H, Liu M, Wang K, Calderwood SK, Xiao X (2006) The inhibition of LPS-induced production of inflammatory cytokines by HSP70 involves inactivation of the $NF-{\kappa}B$ pathway but not the MAPK pathways. Shock 26: 277-284 https://doi.org/10.1097/01.shk.0000223134.17877.ad
  21. Rachmilewitz D, Stamler J S, Karmeli F, Mullins M E, Singel D J, Loscalzo J, Podolsky D K (1993) Peroxynitrite-induced rat colitis-a new model of colonic inflammation. Gastroenterology 105: 1681-1688 https://doi.org/10.1016/0016-5085(93)91063-N
  22. Pande V, Ramos M J (2005) $NF-{\kappa}B$ in human disease: current inhibitors and prospects for de novo structure based design of inhibitors. Current medicinal chemistry 12: 357-374 https://doi.org/10.2174/0929867053363180
  23. Johnson G L, Lapadat R (2002) Mitogen-activated protein kinase pathways mediated by ERK, JNK, and p38 protein kinases. Science 298: 1911-1912 https://doi.org/10.1126/science.1072682

Cited by

  1. Anti-inflammation activity of brazilin in TNF-α induced human psoriasis dermatitis skin model vol.62, pp.1, 2018, https://doi.org/10.1186/s13765-019-0455-z
  2. 만성 역류성 식도염에서 황련과 오수유 혼합물이 식도 점막에 미치는 효과 vol.51, pp.4, 2020, https://doi.org/10.22889/kjp.2020.51.4.349
  3. Rebaudioside A Enhances Resistance to Oxidative Stress and Extends Lifespan and Healthspan in Caenorhabditis elegans vol.10, pp.2, 2018, https://doi.org/10.3390/antiox10020262