Inotodiol inhibits cells migration and invasion and induces apoptosis via p53-dependent pathway in HeLa cells

Phytomedicine. 2019 Jul;60:152957. doi: 10.1016/j.phymed.2019.152957. Epub 2019 May 13.


Background: Inonotus obliquus, namely as Chaga mushroom, is a medicinal and edible fungus, which is widely used in food and medical fields. Inotodiol, a natural lanostane-type triterpenoid with remarkable pharmacological activities, was isolated from Inonotus obliquus, which its potential anti-tumor molecular mechanism was elaborated poorly.

Purpose: The aim of the present study was to investigate the effect of Inotodiol on HeLa cell migration, invasion and apoptosis through p53-dependent pathway.

Study design and methods: The potential mechanisms of Inotodiol on HeLa cell anti-metastatic and pro-apoptosis via wound healing assay, trans-well invasion assay, flow cytometry, caspase-3 activity assay and western blot analysis were studied, as well as the involvement of p53 signaling pathway in anti-metastatic and pro-apoptosis of Inotodiol. Besides, the function of tumor suppressor p53 was further verified by small interfering RNA.

Results: Firstly, the cell viability assay showed that low-concentration of Inotodiol had no cytotoxicity to HeLa cells and whereas the concentration above 25 μM significantly inhibited HeLa cell growth and even induced apoptosis. This result was further demonstrated by cell proliferation and morphology assay. Secondly, in vitro wound healing and trans-well invasion assays reported that low-concentration treatment of Inotodiol significantly inhibited cells migration and invasion in a dose-dependent manner, the western blot analysis of matrix mettalloprotinase-2 (MMP2) and matrix mettalloprotinase-9 (MMP9) levels were also decreased. Moreover, Inotodiol notably induced tumor cell apoptosis by Annexin-V-FITC apoptosis assay, which is associated with activation pro-apoptotic proteins of PARP, cleaved caspase-3 and Bax expression, inhibition anti-apoptotic protein Bcl-2 expression. Finally, the anti-tumor activity of Inotodiol was attenuated by silencing p53 tumor suppressor, the result revealed that pre-treatment with p53-specific small interfering RNA (si-p53) markedly inhibited Intodiol-indeuced HeLa cell apoptosis and decreased the caspase-3 activity. What is more, the inhibitory effect of Inotodiol on tumor migration and invasion was blocked under p53 knockdown.

Conclusion: To sum up, the present study indicated that Inotodiol possessed the potential to prevent malignant tumor migration and invasion, and it might be a natural active compound candidate for clinical treatment of human cervical cancer.

Keywords: Apoptosis; Inotodiol; Invasion; Migration; p53-dependent pathway.

MeSH terms

  • Agaricales / chemistry*
  • Antineoplastic Agents / pharmacology*
  • Apoptosis / drug effects*
  • Caspase 3 / genetics
  • Caspase 3 / metabolism
  • Cell Movement / drug effects
  • Cell Proliferation / drug effects
  • Cell Survival / drug effects
  • Female
  • HeLa Cells
  • Humans
  • Lanosterol / analogs & derivatives*
  • Lanosterol / pharmacology
  • Neoplasm Invasiveness
  • Proto-Oncogene Proteins c-bcl-2 / genetics
  • Proto-Oncogene Proteins c-bcl-2 / metabolism
  • Signal Transduction / drug effects
  • Tumor Suppressor Protein p53 / genetics
  • Tumor Suppressor Protein p53 / metabolism*
  • Uterine Cervical Neoplasms / drug therapy*


  • Antineoplastic Agents
  • BCL2 protein, human
  • Proto-Oncogene Proteins c-bcl-2
  • TP53 protein, human
  • Tumor Suppressor Protein p53
  • inotodiol
  • Lanosterol
  • CASP3 protein, human
  • Caspase 3