Induction of cell cycle arrest and apoptosis by betulinic acid-rich fraction from Dillenia suffruticosa root in MCF-7 cells involved p53/p21 and mitochondrial signalling pathway

Jhi Biau Foo, Latifah Saiful Yazan, Yin Sim Tor, Agustono Wibowo, Norsharina Ismail, Chee Wun How, Nurdin Armania, Su Peng Loh, Intan Safinar Ismail, Yoke Kqueen Cheah, Rasedee Abdullah

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Abstract

Ethnopharmacological relevance Dillenia suffruticosa (Family: Dilleniaceae) or commonly known as "Simpoh air" in Malaysia, is traditionally used for treatment of cancerous growth including breast cancer. Aim of the study D. suffruticosa root dichloromethane extract (DCM-DS) has been reported to induce G0/G1 phase cell cycle arrest and apoptosis in caspase-3 deficient MCF-7 breast cancer cells. The present study was designed to investigate the involvement of p53/p21 and mitochondrial pathway in DCM-DS-treated MCF-7 cells as well as to identify the bioactive compounds responsible for the cytotoxicity of DCM-DS. Materials and methods Extraction of D. suffruticosa root was performed by the use of sequential solvent procedure. GeXP-based multiplex system was employed to investigate the expression of p53, p21, Bax and Bcl-2 genes in MCF-7 cells treated with DCM-DS. The protein expression was then determined using Western blot analysis. The bioactive compounds present in DCM-DS were isolated by using column chromatography. The structure of the compounds was elucidated by using nuclear magnetic resonance spectroscopy. The cytotoxicity of the isolated compounds towards MCF-7 cells was evaluated by using MTT assay. The percentage of betulinic acid (BA) in DCM-DS was determined by HPLC analysis. Results The expression of p53 was significantly up-regulated at protein level. The expression of p21 at both gene and protein levels was significantly up-regulated upon treatment with DCM-DS, suggesting that the induction of G0/G1 phase cell cycle arrest in MCF-7 cells was via p53/p21 pathway. Bcl-2 protein was down-regulated with no change at the mRNA level, postulating that post-translational modification has occurred resulting in the degradation of Bcl-2 protein. Overall, treatment with DCM-DS increased the ratio of Bax/Bcl-2 that drove the cells to undergo apoptosis. A total of 3 triterpene compounds were isolated from DCM-DS. Betulinic acid appears to be the most major and most cytotoxic compound in DCM-DS. Conclusion DCM-DS induced cell cycle arrest and apoptosis in MCF-7 cells via p53/p21 pathway. In addition, DCM-DS induced apoptosis by increasing the ratio of Bax/Bcl-2. Betulinic acid, which is one of the major compounds, is responsible for the cytotoxicity of the DCM-DS. Therefore, BA can be used as a marker for standardisation of herbal product from D. suffruticosa. DCM-DS can also be employed as BA-rich extract from roots of D. suffruticosa for the management of breast cancer.

Original languageEnglish
Pages (from-to)270-278
Number of pages9
JournalJournal of Ethnopharmacology
Volume166
DOIs
Publication statusPublished - 26 May 2015
Externally publishedYes

Keywords

  • Apoptosis
  • Betulinic acid
  • Breast cancer
  • Dillenia suffruticosa
  • Triterpene

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