Basic Study
Copyright ©The Author(s) 2016. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Apr 21, 2016; 22(15): 3952-3961
Published online Apr 21, 2016. doi: 10.3748/wjg.v22.i15.3952
Gallic acid induced apoptotic events in HCT-15 colon cancer cells
Aruna Priyadharshni Subramanian, Saravana Kumar Jaganathan, Mahitosh Mandal, Eko Supriyanto, Ida Idayu Muhamad
Aruna Priyadharshni Subramanian, Saravana Kumar Jaganathan, Eko Supriyanto, Ida Idayu Muhamad, IJN-UTM Cardiovascular Engineering Centre, Faculty of Biosciences and Medical Engineering, Universiti Teknologi Malaysia, Johor Bahru 81310, Malaysia
Mahitosh Mandal, School of Medical Science and Technology, Indian Institute of Technology, Kharagpur 721302, West Bengal, India
Author contributions: Subramanian AP and Jaganathan SK contributed equally to this work; Jaganathan SK and Mandal M designed the research; Subramanian AP and Jaganathan SK performed the research; Mandal M, Supriyanto E and Muhamad II contributed new reagents/analytic tools; Subramanian AP, Jaganathan SK and Mandal M analyzed the data; and Subramanian AP and Jaganathan SK wrote the paper.
Supported by the Research University Grant scheme with the Grant Vot No. Q.J130000.2509.10H13 (partly).
Conflict-of-interest statement: The authors of the manuscript do not have any conflicts of interest.
Data sharing statement: No additional data are available.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Dr. Saravana Kumar Jaganathan, IJN-UTM Cardiovascular Engineering Centre, Faculty of Biosciences and Medical Engineering, Universiti Teknologi Malaysia, Johor Bahru 81310, Malaysia. jaganathaniitkgp@gmail.com
Telephone: +60-7-5558563 Fax: +60-7-5558553
Received: September 3, 2015
Peer-review started: September 10, 2015
First decision: September 29, 2015
Revised: October 26, 2015
Accepted: December 8, 2015
Article in press: December 8, 2015
Published online: April 21, 2016
Processing time: 208 Days and 5 Hours
Abstract

AIM: To investigate the inhibitory action of diet-derived phenolic compound gallic acid (GA) against HCT-15 colon cancer cells.

METHODS: The antiproliferative effect of GA against colon cancer cells was determined by performing thiazolyl blue tetrazolium bromide (MTT) assay. The colony forming ability of GA treated colon cancer cells was evaluated using the colony forming assay. The cell cycle changes induced by GA in HCT-15 cells were analyzed by propidium iodide staining. Levels of reactive oxygen species (ROS) and mitochondrial membrane potential of HCT-15 exposed to GA was assessed using 2’,7’-dichlorfluorescein-diacetate and rhodamine-123 respectively, with the help of flow cytometry. Morphological changes caused by GA treatment in the colon cancer cells were identified by scanning electron microscope and photomicrograph examination. Apoptosis was confirmed using flow cytometric analysis of GA treated HCT-15 cells after staining with Yo-Pro-1.

RESULTS: MTT assay results illustrated that GA has an inhibitory effect on HCT-15 cells with IC50 value of 740 μmol/L. A time-dependent inhibition of colony formation was evident with GA treatment. Cell cycle arrest was evident from the accumulation of GA treated HCT-15 cells at sub-G1 phase (0.98 ± 1.03 vs 58.01 ± 2.05) with increasing exposure time. Flow cytometric analysis of GA treated HCT-15 cells depicted early events associated with apoptosis like lipid layer breakage and fall in mitochondrial membrane potential apart from an increase in the generation of ROS which were in a time dependent manner. SEM and photomicrograph images of the GA-treated cells displayed membrane blebbing and cell shrinking characteristics of apoptosis. Further apoptosis confirmation by Yo-Pro-1 staining also showed the time-dependent increase of apoptotic cells after treatment.

CONCLUSION: These results show that GA induced ROS dependent apoptosis and inhibited the growth of colon cancer cells.

Keywords: Gallic acid, Colon cancer, Apoptosis, Cell cycle, Reactive oxygen species, Lipid layer break

Core tip: This article describes the inhibitory effect of gallic acid (GA), against colon cancer cells. GA treatment suppressed the proliferation and colony formation of HCT-15 cells and the anti-cancerous effect of GA was found to follow reactive oxygen species dependent apoptosis. Early events associated with apoptosis like lipid layer breakage and fall in mitochondrial membrane potential were induced by GA treatment in HCT-15 cells along with cell cycle arrest. Further, morphological changes like membrane blebbing and cell shrinkage were seen in the colon cancer cells after GA. Therefore, our results propel the role of GA as a possible anticancer agent.