Euonymus alatus (Thunb.) Siebold leaf extract enhanced immunostimulatory effects in a cyclophosphamide-induced immunosuppressed rat model

  • Dong Yeop Shin Department of Companion and Laboratory Animal Science, Kongju National University
  • Byeong Soo Kim Department of Companion and Laboratory Animal Science, Kongju National University
  • Hak Yong Lee INVIVO Co. Ltd.
  • Young Mi Park INVIVO Co. Ltd.
  • Yong Wan Kim Daegu Cancer Center, Research and Development Unit, DongSung Pharmaceuticals Co. Ltd.
  • Min Jung Kim Korea Food Research Institute
  • Hye Jeong Yang Korea Food Research Institute
  • Mi Seong Kim Department of Oral Biochemistry, and Institute of Biomaterial-Implant, College of Dentistry, Wonkwang University
  • Jun Sang Bae Department of Pathology, College of Korean Medicine, Wonkwang University
Keywords: Euonymus alatus (Thunb.) Siebold, Immune enhancement, Cyclophosphamide, Macrophage, Immunosuppressed rat


Background: Euonymus alatus (Thunb.) Siebold (EA) is a medicinal plant used in some Asian countries to treat various diseases, including cancer, hyperglycemia, diabetes, urticaria, dysmenorrhea, and arthritis. Owing to the wide range of pharmacological applications of EA, various roles of EA are being studied.

Objective: We evaluated the immune-enhancing effect of EA treatment in a cyclophosphamide (Cy)-induced immunosuppressed rat model.

Design: We analyzed the immune enhancement effect of EA on macrophages by western blotting. In addition, cell viability and natural killer (NK) cell activity were analyzed in splenocytes following EA treatment. For in vivo studies, analysis of weekly body weight, spleen weight, immune cell count, cytokine levels, and spleen histological findings was performed following EA administration in Cy-induced immunocompromised rats.

Results: EA significantly increased cell viability and phospho-nuclear factor-kappa B and phospho-extracellular signal-regulated kinase protein levels in the macrophages. EA significantly increased NK cell activity in splenocytes compared with the control group. In Cy-induced immunosuppressed rats, EA administration increased spleen tissue weight and the contents of leukocytes, lymphocytes, granulocytes, intermediate cells, and plasma cytokines (tumor necrosis factor-α and interferon-γ). In addition, improvement in the damaged spleen tissue was observed.

Conclusions: These findings confirm that EA exerts an immune-enhancing effect, thereby suggesting its potential as an immunostimulatory agent or functional food.


Download data is not yet available.


Dunkelberger JR, Song WC. Complement and its role in innate and adaptive immune responses. Cell Res 2010; 20: 34–50. doi: 10.1038/cr.2009.139

Haddad PS, Azar GA, Groom S, Boivin M. Natural health products, modulation of immune function and prevention of chronic diseases. Evid Based Complement Alternat Med 2005; 2: 513–20. doi: 10.1093/ecam/neh125

Maggini S, Pierre A, Calder PC. Immune function and micronutrient requirements change over the life course. Nutrients 2018; 10: 1531. doi: 10.3390/nu10101531

Venter C, Eyerich S, Sarin T, Klatt KC. Nutrition and the immune system: a complicated tango. Nutrients 2020; 12: 818. doi: 10.3390/nu12030818

Wainwright CL, Teixeira MM, Adelson DL, Buenz EJ, David B, Glaser KB, et al. Future directions for the discovery of natural product-derived immunomodulating drugs: an IUPHAR positional review. Pharmacol Res 2022; 177: 106076. doi: 10.1016/j.phrs.2022.106076

Du B, Xu B. Editorial: immune-boosting effects of dietary bioactive polysaccharides. Front Nutr 2022; 9: 1102641. doi: 10.3389/fnut.2022.1102641

Fan L, Zhang C, Ai L, Wang L, Li L, Fan W, et al. Traditional uses, botany, phytochemistry, pharmacology, separation and analysis technologies of Euonymus alatus (Thunb.) siebold: a comprehensive review. J Ethnopharmacol 2020; 259: 112942. doi: 10.1016/j.jep.2020.112942

Zhai X, Lenon GB, Xue CC, Li CG. Euonymus alatus: a review on its phytochemistry and antidiabetic activity. Evid Based Complement Alternat Med 2016; 2016: 9425714. doi: 10.1155/2016/9425714

Lin XL, Li K, Yang Z, Chen B, Zhang T. Dulcitol suppresses proliferation and migration of hepatocellular carcinoma via regulating SIRT1/p53 pathway. Phytomedicine 2020; 66: 153112. doi: 10.1016/j.phymed.2019.153112

Kim KW, Suh SJ, Kim JD, Kim SS, Lee IS, Kim JK, et al. Effects on lipid peroxidation and antioxidative enzymes of Euonymus A. latus in cultured rat hepatocytes. Basic Clin Pharmacol Toxicol 2009; 104: 60–70. doi: 10.1111/j.1742-7843.2007.00152.x

Ahmed AR, Hombal SM. Cyclophosphamide (Cytoxan). A review on relevant pharmacology and clinical uses. J Am Acad Dermatol 1984; 11: 1115–26. doi: 10.1016/s0190-9622(84)80193-0

Noh EM, Kim JM, Lee HY, Song HK, Joung SO, Yang HJ, et al. Immuno-enhancement effects of Platycodon grandiflorum extracts in splenocytes and a cyclophosphamide-induced immunosuppressed rat model. BMC Complement Altern Med 2019; 19: 322. doi: 10.1186/s12906-019-2724-0

Yan H, Lu J, Wang J, Chen L, Wang Y, Li L, et al. Prevention of cyclophosphamide-induced immunosuppression in mice with traditional Chinese Medicine Xuanfei Baidu Decoction. Front Pharmacol 2021; 12: 730567. doi: 10.3389/fphar.2021.730567

Ogino MH, Tadi P. Cyclophosphamide. Treasure Island, FL: StatPearls Publishing; 2022. Available from: [cited 4 July 2022].

Zhong W, Huang H, Yang Z, Chang P. rhCNB improves cyclophosphamide-induced immunodeficiency in BALB/c mice. Evid Based Complement Alternat Med 2022; 2022: 4891399. doi: 10.1155/2022/4891399

Fan KJ, Li YW, Wu J, Li J, Zhang J, Wang QS, et al. The traditional Chinese Medicine Fufang Shatai Heji (STHJ) enhances immune function in cyclophosphamide-treated mice. Evid Based Complement Alternat Med 2020; 2020: 3849847. doi: 10.1155/2020/3849847

Park YM, Lee HY, Shin DY, Kim DS, Yoo JJ, Yang HJ, et al. Immune-enhancing effects of co-treatment with Kalopanax pictus Nakai Bark and Nelumbo nucifera Gaertner leaf extract in a cyclophosphamide-induced immunosuppressed rat model. Front Nutr 2022; 9: 898417. doi: 10.3389/fnut.2022.898417

Atanasov AG, Zotchev SB, Dirsch VM, International Natural Product Sciences Taskforce, Supuran CT. Natural products in drug discovery: advances and opportunities. Nat Rev Drug Discov 2021; 20: 200–16. doi: 10.1038/s41573-020-00114-z

Lee TK, Lee JY, Kim DI, Lee YC, Kim CH. Differential regulation of protein kinase C activity by modulating factors and Euonymus alatus (Thunb.) Sieb in human myometrial and uterine leiomyomal smooth muscle cells. Int J Gynecol Cancer 2005; 15: 349–58. doi: 10.1111/j.1525-1438.2005.15228.x

Cha BY, Park CJ, Lee DG, Lee YC, Kim DW, Kim JD, et al. Inhibitory effect of methanol extract of Euonymus alatus on matrix metalloproteinase-9. J Ethnopharmacol 2003; 85: 163–7. doi: 10.1016/s0378-8741(02)00373-2

Woo Y, Lim JS, Oh J, Lee JS, Kim JS. Neuroprotective effects of Euonymus alatus extract on scopolamine-induced memory deficits in mice. Antioxidants (Basel) 2020; 9: 449. doi: 10.3390/antiox9050449

Bronte V, Pittet MJ. The spleen in local and systemic regulation of immunity. Immunity 2013; 39: 806–18. doi: 10.1016/j.immuni.2013.10.010

Klimp AH, de Vries EG, Scherphof GL, Daemen T. A potential role of macrophage activation in the treatment of cancer. Crit Rev Oncol Hematol 2002; 44: 143–61. doi: 10.1016/s1040-8428(01)00203-7

Kadl A, Meher AK, Sharma PR, Lee MY, Doran AC, Johnstone SR, et al. Identification of a novel macrophage phenotype that develops in response to atherogenic phospholipids via Nrf2. Circ Res 2010; 107: 737–46. doi: 10.1161/CIRCRESAHA.109.215715

Martinez FO, Helming L, Gordon S. Alternative activation of macrophages: an immunologic functional perspective. Annu Rev Immunol 2009; 27: 451–83. doi: 10.1146/annurev.immunol.021908.132532

Lumeng CN, Bodzin JL, Saltiel AR. Obesity induces a phenotypic switch in adipose tissue macrophage polarization. J Clin Invest 2007; 117: 175–84. doi: 10.1172/JCI29881

Wang LX, Zhang SX, Wu HJ, Rong XL, Guo J. M2b macrophage polarization and its roles in diseases. J Leukoc Biol 2019; 106: 345–58. doi: 10.1002/JLB.3RU1018-378RR

McAleer JP, Vella AT. Understanding how lipopolysaccharide impacts CD4 T-cell immunity. Crit Rev Immunol 2008; 28: 281–99. doi: 10.1615/critrevimmunol.v28.i4.20

Mburu S, Marnewick JL, Abayomi A, Ipp H. Modulation of LPS-induced CD4+ T-cell activation and apoptosis by antioxidants in untreated asymptomatic HIV infected participants: an in vitro study. Clin Dev Immunol 2013; 2013: 631063. doi: 10.1155/2013/631063

Ji C, Zhang Z, Chen J, Song D, Liu B, Li J, et al. Immune-enhancing effects of a novel glucan from Purple Sweet Potato Ipomoea batatas (L.) Lam on RAW264.7 macrophage cells via TLR2- and TLR4-mediated pathways. J Agric Food Chem 2021; 69: 9313–25. doi: 10.1021/acs.jafc.1c03850

Baeuerle PA, Henkel T. Function and activation of NF-kappa B in the immune system. Annu Rev Immunol 1994; 12: 141–79. doi: 10.1146/annurev.iy.12.040194.001041

Hayden MS, West AP, Ghosh S. NF-kappaB and the immune response. Oncogene 2006; 25: 6758–80. doi: 10.1038/sj.onc.1209943

Shin HY, Hwang KC, Mi XJ, Moon SK, Kim YJ, Kim H. Rhamnogalacturonan I-rich polysaccharide isolated from fermented persimmon fruit increases macrophage-stimulatory activity by activating MAPK and NF-κB signaling. J Sci Food Agric 2022; 102: 2846–54. doi: 10.1002/jsfa.11625

Park E-J, Lee Y-S, Kim SM, Jung AJ, Yoo J-H, Lee S-H, et al. Jeong and H.Lee, immune-enhancing effects of red Platycodon grandiflorus root extract via p38 MAPK-mediated NF-κB activation. Appl Sci 2020; 10: 5457. doi: 10.3390/app10165457

Lee J, Choi JW, Sohng JK, Pandey RP, Park YI. The immunostimulating activity of quercetin 3-O-xyloside in murine macrophages via activation of the ASK1/MAPK/NF-κB signaling pathway. Int Immunopharmacol 2016; 31: 88–97. doi: 10.1016/j.intimp.2015.12.008

Chung HS, Jeong HJ, Kim JS, Jeong SI, Kim KS, Kim KS, et al. Activation of inducible nitric oxide synthase by Euonymus alatus in mouse peritoneal macrophages. Clin Chim Acta 2002; 318: 113–20. doi: 10.1016/s0009-8981(01)00808-7

Caamaño J, Hunter CA. NF-kappaB family of transcription factors: central regulators of innate and adaptive immune functions. Clin Microbiol Rev 2002; 15: 414–29. doi: 10.1128/CMR.15.3.414-429.2002

Kos FJ, Engleman EG. Immune regulation: a critical link between NK cells and CTLs. Immunol Today 1996; 17: 174–6. doi: 10.1016/0167-5699(96)80616-5

Ghasemi M, Abbasi L, Ghanbari Naeini L, Kokabian P, Nameh Goshay Fard N, Givtaj N. Dendritic cells and natural killer cells: the road to a successful oncolytic virotherapy. Front Immunol 2023; 13: 950079. doi: 10.3389/fimmu.2022.950079

Huang GC, Wu LS, Chen LG, Yang LL, Wang CC. Immuno-enhancement effects of Huang Qi Liu Yi Tang in a murine model of cyclophosphamide-induced leucopenia. J Ethnopharmacol 2007; 109: 229–35. doi: 10.1016/j.jep.2006.07.023

Endo M, Beppu H, Akiyama H, Wakamatsu K, Ito S, Kawamoto Y, et al. Agaritine purified from Agaricus blazei Murrill exerts anti-tumor activity against leukemic cells. Biochim Biophys Acta 2010; 1800: 669–73. doi: 10.1016/j.bbagen.2010.03.016

Boyman O, Sprent J. The role of interleukin-2 during homeostasis and activation of the immune system. Nat Rev Immunol 2012; 12: 180–90. doi: 10.1038/nri3156

Wendelsdorf K, Bassaganya-Riera J, Hontecillas R, Eubank S. Model of colonic inflammation: immune modulatory mechanisms in inflammatory bowel disease. J Theor Biol 2010; 264: 1225–39. doi: 10.1016/j.jtbi.2010.03.027

Goldsby R, Kindt TJ, Osborne BA, Kuby J. Cells and organs of the immune system. In: Immunology. 5th ed. New York, NY: W. H. Freeman and Company; 2003, pp. 24–56.

Jeong DG, Seo JH, Heo SH, Choi YK, Jeong ES. Tumor necrosis factor-alpha deficiency impairs host defense against Streptococcus pneumoniae. Lab Anim Res 2015; 31: 78–85. doi: 10.5625/lar.2015.31.2.78

Woolard MD, Hudig D, Tabor L, Ivey JA, Simecka JW. NK cells in gamma-interferon-deficient mice suppress lung innate immunity against Mycoplasma spp. Infect Immun 2005; 73: 6742–51. doi: 10.1128/IAI.73.10.6742-6751.2005

Yang ML, Wang CT, Yang SJ, Leu CH, Chen SH, Wu CL, et al. IL-6 ameliorates acute lung injury in influenza virus infection. Sci Rep 2017; 7: 43829. doi: 10.1038/srep43829

Kawai T, Akira S. The role of pattern-recognition receptors in innate immunity: update on Toll-like receptors. Nat Immunol 2010; 11: 373–84. doi: 10.1038/ni.1863
How to Cite
Shin D. Y., Kim B. S., Lee H. Y., Park Y. M., Kim Y. W., Kim M. J., Yang H. J., Kim M. S., & Bae J. S. (2023). <em>Euonymus alatus</em&gt; (Thunb.) Siebold leaf extract enhanced immunostimulatory effects in a cyclophosphamide-induced immunosuppressed rat model. Food & Nutrition Research, 67.
Original Articles