Journal of the Society of Cosmetic Scientists of Korea (대한화장품학회지)

Society of Cosmetic Scientists of Korea (대한화장품학회)
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Anti-oxidative Effect of Chungsimyeonja-um (CSYJE) via Nrf2/HO-1 Pathway Activity in Lipopolysaccharide (LPS) Induced RAW 264.7 Macrophages

대식세포에서 Nrf2/HO-1경로를 통한 청심연자음의 항산화효과

  • Jeon, Seon Hong (Department of Cosmetic Science, Semyung University) ;
  • Oh, Sol La (Department of Cosmetic Science, Semyung University) ;
  • Kim, So Jeong (Department of Cosmetic Science, Semyung University) ;
  • Jeon, Bo Hee (Department of Cosmetic Science, Semyung University) ;
  • Sung, Jin Young (Department of Cosmetic Science, Semyung University) ;
  • Kim, Yong Min (Department of Cosmetic Science, Semyung University)
  • 전선홍 (세명대학교 화장품과학과) ;
  • 오솔라 (세명대학교 화장품과학과) ;
  • 김소정 (세명대학교 화장품과학과) ;
  • 전보희 (세명대학교 화장품뷰티생명공학부) ;
  • 성진영 (세명대학교 화장품뷰티생명공학부) ;
  • 김용민 (세명대학교 화장품과학과)
  • Received : 2020.06.12
  • Accepted : 2020.07.21
  • Published : 2020.09.30
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Abstract

Reactive oxygen species (ROS) plays an important role in maintaining homeostasis. However, excessive ROS production damages cellular components such as proteins, lipids, and nucleic acids and promotes skin aging. In this study, we confirmed the antioxidant effect of CSYJE to prevent excessive oxidative stress. First, DPPH and ABTS assays were performed to confirm the antioxidant effect of CSYJE and the radical scavenging activity was confirmed depending on the concentration. As a result of performing the MTT assay to confirm the cell viability, it was confirmed that there was no cytotoxicity at a concentration of 1,000 ㎍/mL. As a result of western blotting to confirm the expression levels of the antioxidant-related proteins nuclear-E2-related factor 2 (Nrf2) and Heme oxygenase-1 (HO-1), it was confirmed that the expression was increased in a concentration-dependent manner. After inducing ROS with lipopolysaccharide (LPS), an intracellular ROS-causing substance, DCF-DA was performed to confirm the inhibitory effect of ROS production, and the inhibition of ROS production was confirmed to concentration-dependent. Real-time RT-PCR was performed to confirm the mRNA expression level of inflammatory cytokines and inflammatory mediator caused by ROS generation, mRNA expression was reduced in a dose dependent manner. Therefore, this study confirmed the antioxidant effect of CSYJE through the Nrf2/HO-1 signaling pathway, which suggests that CSYJE can be used as an antioxidant cosmetic material by inhibiting free radicals.

활성산소종(reactive oxygen species, ROS)은 우리 몸의 항상성 유지에 있어 중요한 역할을 한다. 그러나 과도한 ROS의 생성은 단백질, 지질, 핵산과 같은 세포 구성성분을 손상시키고 피부노화를 촉진시킨다. 이에 본 연구에서는 과도한 산화 스트레스를 예방하기 위해 Chungsimyeonja-um (CSYJE)의 항산화 효과를 확인하였다. 먼저 DPPH 및 ABTS assay를 실시하여 CSYJE의 항산화 효과를 확인한 결과 농도 의존적으로 radical 소거 활성을 확인하였다. 세포생존율 확인을 위해 MTT assay를 실시한 결과 1,000 ㎍/mL 농도에서 세포 독성이 없음을 확인하였다. 항산화 관련 단백질인 nuclear-E2-related factor 2 (Nrf2), Heme oxygenase-1 (HO-1)의 발현 수준을 확인하기 위해 western blotting을 실시한 결과 농도 의존적으로 발현이 증가하는 것을 확인하였다. 세포 내 ROS유발 물질인 lipopolysaccharide (LPS)로 ROS를 유도한 후, ROS생성 억제효과를 확인하기 위해 DCF-DA 염색법을 실시한 결과 농도 의존적으로 ROS 생성 억제효과를 확인하였으며 ROS의 생성으로 인한 염증성 사이토카인과 염증인자의 mRNA발현 수준을 확인하기 위해 real-time RT-PCR을 실시한 결과 농도 의존적으로 염증성 사이토카인과 염증인자의 mRNA 발현을 억제시켰다. 따라서, 본 연구는 Nrf2/HO-1 신호 전달 경로 활성을 통해 CSYJE의 항산화효과를 확인했으며 이는 CSYJE가 활성산소를 억제하여 항산화 화장품의 재료로서 사용될 수 있음을 시사한다.

Keywords

References

  1. K. J. Trouba, H. K. Hamadeh, R. P. Amin, and D. R. Germolec, Oxidative stress and its role in skin disease, Antioxid. Redox Signal., 4(4), 665 (2002). https://doi.org/10.1089/15230860260220175
  2. J. W. Kaspar, S . K. Niture, and A. K. Jaiswal, Nrf2:inrf2 (keap1) signaling in oxidative stress, Free Radical Biology and Medicine, 47(9), 1304 (2009). https://doi.org/10.1016/j.freeradbiomed.2009.07.035
  3. J. A. Johnson, D. A. Johnson, A. D. Kraft, M. J. Calkins, R. J. Jakel, M. R. Vargas, and P. C. Chen, The nrf2-are pathway: an indicator and modulator of oxidative stress in neurodegeneration, Annals of The New York Academy of Sciences, 1147(1), 61 (2008). https://doi.org/10.1196/annals.1427.036
  4. L. Li, P. Grenard, J. T. Van Nhieu, B. Julien, A. Mallat, A. Habib, and S. Lotersztajn, Heme oxygenase-1 is an antifibrogenic protein in human hepatic myofibrobalsts, Gastroenterology, 125(2), 460 (2003). https://doi.org/10.1016/S0016-5085(03)00906-5
  5. A. Nakao, L. E. Otterbein, M. Overhaus, J. K. Sarady, A. Tsung, K. Kimizuka, and F. H. Bach, Biliverdin protects the functional integrity of a transplanted syngeneic small bowel, Gastroenterology, 127(2), 595 (2004). https://doi.org/10.1053/j.gastro.2004.05.059
  6. S. R. Yoo, H. K. Ha, N. R. Lee, H. K. Shin, C. S. Seo, A study on change of marker compounds and biological activity in chungsimyeonja-eum decoction depending on a storage temperature and periods, The Korea Journal of Herbology, 32(4), 25 (2017). https://doi.org/10.6116/kjh.2017.32.1.25.
  7. I. J. Rhee and D. M. Lee, The effect of some antipolydipsia oriental prescriptions on experimental diabetic rats, The Pharmaceutical Society of Korea, 38(4), 555 (1994).
  8. Y. Z. Yang, Y. Z. Tang, and Y. H. Liu, Wogonoside displays anti-inflammatory effects through modulating inflammatroy mediator expression using raw 264.7 cells, Journal of Ethnopharmacology, 148(1), 271 (2013). https://doi.org/10.1016/j.jep.2013.04.025
  9. X. L. Li, A. G. Zhou, L. Zhang, and W.J. Chen, Antioxidant status and immune activity of glycyrrhizin in allergic rhinitis mice, International Journal of Molecular Sciences, 12(2), 905 (2011). https://doi.org/10.3390/ijms12020905
  10. H. S. Lim, Y. J. Kim, C. S. Seo, S. R. Yoo, H. K. Shin, and S. J. Jeong, Chungsimyeonja-eum inhibits inflammatory responses in raw 264.7 macrophages and hacat keratinocytes, BMC Complementary and Alternative Medicine, 15(1), 371 (2015). https://doi.org/10.1186/s12906-015-0902-2
  11. P. Perer, Text of the immune system, third edition, 1, Lifescience, Seoul (2011).
  12. I. S. Kim, S. H. Lee, K. J. Lee, D. S. Lee, J. S. Park, and N. K. Lee, Text of essential immunology, 28, Daihak, Seoul (2017).
  13. J. A. Smith, A. Das, S. K. Ray, and N. L. Banik, Role of pro-inflammatory cytokines released from microglia in neurodegenerative diseases, Brain Research Bulletin, 87(1), 10 (2012). https://doi.org/10.1016/j.brainresbull.2011.10.004
  14. M. Hori, M. Kita, S. Torihashi, S. Miyamoto, K. J. Won, K. Sato, and H. Karaki, Upregulation of inos by cox-2 in muscularis resident macrophage of rat intestine stimulated with lps, American Journal of Physiology-Gastrointestinal and Liver Physiology, 280(5), 930 (2001). https://doi.org/10.1152/ajpgi.2001.280.5.G930
  15. C. A. Dinarello, Biologic basis for interleukin-1 in disease, American Society of Hematology, 87(6), 2095 (1996).
  16. T. Kishimoto, S. Akira, M. Narazaki, and T. Taga, Interleukin-6 family of cytokines and gp130, American Society of Hematology, 86(4), 1243 (1995).
  17. D. Y. Kim, Anti-allergic effect of sapindus mukorossi fruits extract in human mast cells, Journal of Investigative Cosmetology, 15(4), 389 (2019).
  18. X. Xu, P. Yin, C. Wan, X. Chong, M. Liu, P. Cheng, J. Chen, F. Liu, and J. Xu, Punicalagin inhibits inflammation in lps-induced raw 264.7 macrophages via the suppression of tlr4-mediated mapks and nf-${\kappa}b$ activation, Inflammation, 37(3), 956 (2014). https://doi.org/10.1007/s10753-014-9816-2
  19. T. Lawrence, C. Fong, The resolution of inflammation: anti-inflammatory roles for nf-${\kappa}b$, The International Journal of Biochemistry & Cell Biology, 42(4), 519 (2010). https://doi.org/10.1016/j.biocel.2009.12.016
  20. H. K. Kim, Role of erk/mapk signalling pathway in anti-inflammatory effects of ecklonia cava in activated human mast cell line-1 cells, Asian Pacific Journal of Tropical Medicine, 7(9), 703 (2014). https://doi.org/10.1016/S1995-7645(14)60120-6
  21. M. Yoshizumi, T. Nakamura, M. Kato, T. Ishioka, K. Kozawa, K. Wakamatsu, and H. Kimura, Release of cytokines/chemokines and cell death in uvb-irradiated human keratinocytes, hacat, Cell Biology International, 32(11), 1405 (2008). https://doi.org/10.1016/j.cellbi.2008.08.011
  22. S. N. Lee, The antioxidant effect of rutin in human dermal fibroblasts damaged by reactive oxygen species, Asian Journal of Beauty & Cosmetology. 12(6), 831 (2014).
  23. X. Xu, H. Li, X. Hou, D. Li, S. He, C. Wan, P. Yin, M. Liu, F. Liu, and J. Xu, Punicalagin induces nrf2/ho-1 expression via upregulation of pi3k/akt pathway and inhibits lps-induced oxidative stress in raw 264.7 macrophages, Mediators of Inflammation. 1, 15 (2015). https://doi.org/10.1155/S0962935192000048
  24. J. P. Roach, E. E. Moore, D. A. Partrick, S. S. Damle, C. C. Silliman, R. C. Mclntyre, and A. Banerjee, Heme oxygenase-1 induction in macrophages by a hemoglobin-based oxygen carrier reduces endotoxin-stimulated cytokine secretion, Shock, 31(3), 251 (2009). https://doi.org/10.1097/SHK.0b013e3181834115
  25. N. Li, J. Alam, M. I. Venkatesan, A.E. Fernandez, D. Schmitz, E. D. Stefano, N. Slauther, E. K. Xiaorong wang, A. Huang, M. Wang, A. H. Miguel, and A.E. Nelj, Nrf2 is a key transcription factor that regulates antioxidant defense in macrophages and epithelial cells: protecting against the proinflammatory and oxidizing effects of diesel exhaust chemicals, Immunology, 173(5), 3467 (2004). https://doi.org/10.4049/jimmunol.173.5.3467