Investigational New Drugs

, Volume 37, Issue 5, pp 865–875 | Cite as

A mediator of phosphorylated Smad2/3, evodiamine, in the reversion of TAF-induced EMT in normal colonic epithelial cells

  • Wanbin Yang
  • Xiuli Gong
  • Xiulian Wang
  • Chao HuangEmail author


Purpose Transdifferentiation exists within stromal cells in the tumour microenvironment. Transforming growth factor-β (TGF-β) secreted by tumour-associated fibroblasts (TAFs) affects the differentiation states of epithelial cells, including epithelial-mesenchymal transition (EMT). Evodiamine, a natural drug, can regulate differentiation. However, the specific effects and relative mechanisms of evodiamine remain unknown. Design We used four models to observe the influence of TAF-like CCD-18Co cells on the colon epithelial cell line HCoEpiC: the 3D- and 2D-mono-culture system, Transwell and direct co-culture model. Additionally, we established conditioned medium from CCD-18Co cells. The TGF-β pathway inhibitor LY364947 and evodiamine were added. Morphological changes and classical EMT markers were observed and detected using phase contrast microscopy and immunofluorescence. Cell migration was measured by the wound-healing assay. Western blotting was performed to detect the TGF-β/Smad signalling pathway. Results CCD-18Co cells induced EMT-like changes in the 2D- and 3D-cultured epithelial cell line HCoEpiC, accompanied by high expression of ZEB1 and Snail and the enhancement of migration. Moreover, CCD-18Co-derived conditioned medium caused dysfunction of TGF-β/Smad signalling in EMT. Evodiamine inhibited these EMT-like HCoEpiC and their migration. Additionally, evodiamine down-regulated the expression of ZEB1/Snail and up-regulated the expression of phosphorylated Smad2/3 (pSmad2/3). Evodiamine also increased the ratios of pSmad2/Smad2 and pSmad3/Smad3. Conclusion Based on our observations, evodiamine can reverse the TAF-induced EMT-like phenotype in colon epithelial cells, which may be associated with its mediation of phosphorylated Smad2 and Smad3 expression.


Epithelial-mesenchymal transition Tumour-associated fibroblasts Tumour microenvironment Transdifferentiation Transforming growth factor-β Evodiamine 



Bo Liu, PhD, and Cun-jia Zhang, PhD, provided medical writing support.


The work was supported by the Natural Science Foundation of Guangdong Province (2018A030310060) and PhD Initial Foundation of Hospital (20170916).

Compliance with ethical standards

Conflict of interest

Wan bin Yang declares that he has no conflict of interest. Xiu li Gong declares that she has no conflict of interest. Xiu lian Wang declares that she has no conflict of interest. Chao Huang declares that he has no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Informed consent

For this type of study, formal consent is not required.


  1. 1.
    Servais C, Erez N (2013) From sentinel cells to inflammatory culprits: cancer-associated fibroblasts in tumour-related inflammation. J Pathol 229:198–207CrossRefGoogle Scholar
  2. 2.
    Glentis A, Oertle P, Mariani P, Chikina A, El Marjou F, Attieh Y et al (2017) Cancer-associated fibroblasts induce metalloprotease-independent cancer cell invasion of the basement membrane. Nat Commun 8(1):924CrossRefGoogle Scholar
  3. 3.
    Kojima Y, Acar A, Eaton EN, Mellody KT, Scheel C, Ben-Porath I, onder TT, Wang ZC, Richardson AL, Weinberg RA, Orimo A (2010) Autocrine TGF-beta and stromal cell-derived factor-1 (SDF-1) signaling drives the evolution of tumor-promoting mammary stromal myofibroblasts. Proc Natl Acad Sci U S A 107:20009–20014CrossRefGoogle Scholar
  4. 4.
    Achyut BR, Yang L (2011) Transforming growth factor-β in the gastrointestinal and hepatic tumor microenvironment. Gastroenterology 141(4):1167–1178CrossRefGoogle Scholar
  5. 5.
    Neuzillet C, Tijeras-Raballand A, Cohen R, Cros J, Faivre S, Raymond E, de Gramont A (2015) Targeting the TGFβ pathway for cancer therapy. Pharmacol Ther 147:22–31CrossRefGoogle Scholar
  6. 6.
    Mukaida N, Sasaki S (2016) Fibroblasts, an inconspicuous but essential player in colon cancer development and progression. World J Gastroenterol 22(23):5301–5316CrossRefGoogle Scholar
  7. 7.
    Guarino M, Tosoni A, Nebuloni M (2009) Direct contribution of epithelium to organ fibrosis: epithelial-mesenchymal transition. Hum Pathol 40:1365–1376CrossRefGoogle Scholar
  8. 8.
    Yu HI, Chou HC, Su YC, Lin LH, Lu CH, Chuang HH, Tsai YT, Liao EC, Wei YS, Yang YT, Lee YR, Chan HL (2018) Proteomic analysis of evodiamine-induced cytotoxicity in thyroid cancer cells. J Pharm Biomed Anal 160:344–350CrossRefGoogle Scholar
  9. 9.
    Zhou Y, Hu J (2018) Evodiamine induces apoptosis, G2/M cell cycle arrest, and inhibition of cell migration and invasion in human osteosarcoma cells via Raf/MEK/ERK Signalling pathway. Med Sci Monit 24:5874–5880CrossRefGoogle Scholar
  10. 10.
    Huang C, Wen B (2016) Phenotype transformation of immortalized NCM460 colon epithelial cell line by TGF-β1 is associated with chromosome instability. Mol Biol Rep 43:1069–1078CrossRefGoogle Scholar
  11. 11.
    Bauer M, Su G, Casper C, He R, Rehrauer W, Friedl A (2010) Heterogeneity of gene expression in stromal fibroblasts of human breast carcinomas and normal breast. Oncogene 29(12):1732–1740CrossRefGoogle Scholar
  12. 12.
    Navab R, Strumpf D, Bandarchi B, Zhu CQ, Pintilie M, Ramnarine VR, Ibrahimov E, Radulovich N, Leung L, Barczyk M, Panchal D, To C, Yun JJ, der S, Shepherd FA, Jurisica I, Tsao MS (2011) Prognostic gene-expression signature of carcinoma-associated fibroblasts in non-small cell lung cancer. Proc Natl Acad Sci U S A 108(17):7160–7165CrossRefGoogle Scholar
  13. 13.
    De Wever O, Demetter P, Mareel M, Bracke M (2008) Stromal myofibroblasts are drivers of invasive cancer growth. Int J Cancer 123(10):2229–2238CrossRefGoogle Scholar
  14. 14.
    Calon A, Tauriello DV, Batlle E (2014) TGF-beta in CAF-mediated tumor growth and metastasis. Semin Cancer Biol 25:15–22CrossRefGoogle Scholar
  15. 15.
    Lakins MA, Ghorani E, Munir H, Martins CP, Shields JD (2018) Cancer-associated fibroblasts induce antigen-specific deletion of CD8 + T cells to protect tumour cells. Nat Commun 9(1):948CrossRefGoogle Scholar
  16. 16.
    Giménez-Bastida JA, Surma M, Zieliński H (2015) In vitro evaluation of the cytotoxicity and modulation of mechanisms associated with inflammation induced by perfluorooctanesulfonate and perfluorooctanoic acid in human colon myofibroblasts CCD-18Co. Toxicol in Vitro 29(7):1683–1691CrossRefGoogle Scholar
  17. 17.
    Pereira C, Araújo F, Barrias CC, Granja PL, Sarmento B (2015) Dissecting stromal-epithelial interactions in a 3D in vitro cellularized intestinal model for permeability studies. Biomaterials 56:36–45CrossRefGoogle Scholar
  18. 18.
    Chen Q, Liu G, Liu S, Su H, Wang Y, Li J, Luo C (2018) Remodeling the tumor microenvironment with emerging Nanotherapeutics. Trends Pharmacol Sci 39(1):59–74CrossRefGoogle Scholar
  19. 19.
    Papageorgis P (2015) TGFβ signaling in tumor initiation, epithelial-to-mesenchymal transition, and metastasis. J Oncol 2015:587193CrossRefGoogle Scholar
  20. 20.
    Fearon ER (2011) Molecular genetics of colorectal cancer. Annu Rev Pathol 6:479–507CrossRefGoogle Scholar
  21. 21.
    Lyu G, Guan Y, Zhang C, Zong L, Sun L, Huang X, Huang L, Zhang L, Tian XL, Zhou Z, Tao W (2018) TGF-β signaling alters H4K20me3 status via miR-29 and contributes to cellular senescence and cardiac aging. Nat Commun 9(1):2560CrossRefGoogle Scholar
  22. 22.
    Wei J, Li Z, Yuan F (2014) Evodiamine might inhibit TGF-beta1-induced epithelial-mesenchymal transition in NRK52E cells via Smad and PPAR-gamma pathway. Cell Biol Int 38(7):875–880CrossRefGoogle Scholar
  23. 23.
    Massagué J (2008) TGFbeta in Cancer. Cell 134(2):215–230CrossRefGoogle Scholar
  24. 24.
    Peng X, Zhang Q, Zeng Y, Li J, Wang L, Ai P (2015) Evodiamine inhibits the migration and invasion of nasopharyngeal carcinoma cells in vitro via repressing MMP-2 expression. Cancer Chemother Pharmacol 76(6):1173–1184CrossRefGoogle Scholar
  25. 25.
    Wen Z, Feng S, Wei L, Wang Z, Hong D, Wang Q (2015) Evodiamine, a novel inhibitor of the Wnt pathway, inhibits the self-renewal of gastric cancer stem cells. Int J Mol Med 36(6):1657–1663CrossRefGoogle Scholar
  26. 26.
    David CJ, Huang YH, Chen M, Su J, Zou Y, Bardeesy N, Iacobuzio-Donahue CA, Massagué J (2016) TGF-β tumor suppression through a lethal EMT. Cell 164(5):1015–1030CrossRefGoogle Scholar
  27. 27.
    Vera-Ramirez L, Sanchez-Rovira P, Ramirez-Tortosa CL, Quiles JL, Ramirez-Tortosa MC, Alvarez JC, Fernandez-Navarro M, Lorente JA (2010) Gene-expression profiles, tumor microenvironment, and cancer stem cells in breast cancer: latest advances towards an integrated approach. Cancer Treat Rev 36:477–484CrossRefGoogle Scholar
  28. 28.
    Bussard KM, Mutkus L, Stumpf K, Gomez-Manzano C, Marini FC (2016) Tumor-associated stromal cells as key contributors to the tumor microenvironment. Breast Cancer Res 18(1):84CrossRefGoogle Scholar
  29. 29.
    Klingberg F, Hinz B, White ES (2013) The myofibroblast matrix: implications for tissue repair and fibrosis. J Pathol 229:298–309CrossRefGoogle Scholar
  30. 30.
    Werner S, Grose R (2003) Regulation of wound healing by growth factors and cytokines. Physiol Rev 83:835–870CrossRefGoogle Scholar
  31. 31.
    Giménez-Bastida JA, Laparra-Llopis JM, Baczek N, Zielinski H (2018) Buckwheat and buckwheat enriched products exert an anti-inflammatory effect on the myofibroblasts of colon CCD-18Co. Food Funct 9(6):3387–3397CrossRefGoogle Scholar
  32. 32.
    McDonald LT, LaRue AC (2012) Hematopoietic stem cell derived carcinoma-associated fibroblasts: a novel origin. Int J Clin Exp Pathol 5(9):863–873Google Scholar
  33. 33.
    Ji Q, Liu X, Han Z, Zhou L, Sui H, Yan L, Jiang H, Ren J, Cai J, Li Q (2015) Resveratrol suppresses epithelial-to-mesenchymal transition in colorectal cancer through TGF-β1/Smads signaling pathway mediated snail/E-cadherin expression. BMC Cancer 15:97CrossRefGoogle Scholar
  34. 34.
    Dong F, Liu T, Jin H, Wang W (2018) Chimaphilin inhibits human osteosarcoma cell invasion and metastasis through suppressing the TGF-β1-induced epithelial-to-mesenchymal transition markers via PI-3K/Akt, ERK1/2, and Smad signaling pathways. Can J Physiol Pharmacol 96(1):1–7CrossRefGoogle Scholar
  35. 35.
    Ioannou M, Kouvaras E, Papamichali R, Samara M, Chiotoglou I, Koukoulis G (2018) Smad4 and epithelial-mesenchymal transition proteins in colorectal carcinoma: an immunohistochemical study. J Mol Histol 49(3):235–244CrossRefGoogle Scholar
  36. 36.
    Dong Z, Tai W, Lei W, Wang Y, Li Z, Zhang T (2016) IL-27 inhibits the TGF-β1-induced epithelial-mesenchymal transition in alveolar epithelial cells. BMC Cell Biol 17:7CrossRefGoogle Scholar
  37. 37.
    Su T, Yang X, Deng JH, Huang QJ, Huang SC, Zhang YM, Zheng HM, Wang Y, Lu LL, Liu ZQ (2018) Evodiamine, a novel NOTCH3 methylation stimulator, significantly suppresses lung carcinogenesis in vitro and in vivo. Front Pharmacol 9:434. CrossRefGoogle Scholar
  38. 38.
    Yao X, Yu T, Zhao C, Li Y, Peng Y, Xi F, Yang G (2018) Evodiamine promotes differentiation and inhibits proliferation of C2C12 muscle cells. Int J Mol Med 41(3):1627–1634Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Institute of Tropical DiseasesGuangzhou University of Chinese MedicineGuangzhouPeople’s Republic of China
  2. 2.Department of Spleen and Stomach DiseasesSecond Hospital of Traditional Chinese Medicine of GuangdongGuangzhouPeople’s Republic of China
  3. 3.Department of Traditional Chinese Medicine, Affiliated Bao’an Hospital of Traditional Chinese Medicine of ShenzhenGuangzhou University of Chinese MedicineShenzhenPeople’s Republic of China
  4. 4.Central Laboratory, Affiliated Bao’an Hospital of ShenzhenSouthern Medical UniversityShenzhenPeople’s Republic of China

Personalised recommendations