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Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins

Chalcones have been shown to exhibit anti-cancer properties by targeting multiple molecular pathways. It was, therefore, of interest to investigate flavokawain C (FKC), a naturally occurring chalcone, which can be isolated from Kava (Piper methysticum Forst) root extract. The aim of this study was t...

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Autores principales: Phang, Chung-Weng, Karsani, Saiful Anuar, Abd Malek, Sri Nurestri
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Medknow Publications & Media Pvt Ltd 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5538174/
https://www.ncbi.nlm.nih.gov/pubmed/28808400
http://dx.doi.org/10.4103/0973-1296.210180
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author Phang, Chung-Weng
Karsani, Saiful Anuar
Abd Malek, Sri Nurestri
author_facet Phang, Chung-Weng
Karsani, Saiful Anuar
Abd Malek, Sri Nurestri
author_sort Phang, Chung-Weng
collection PubMed
description Chalcones have been shown to exhibit anti-cancer properties by targeting multiple molecular pathways. It was, therefore, of interest to investigate flavokawain C (FKC), a naturally occurring chalcone, which can be isolated from Kava (Piper methysticum Forst) root extract. The aim of this study was to investigate the inhibitory effect of FKC on the growth of HT-29 cells and its underlying mechanism of action. Cell viability of HT-29 cells was assessed by Sulforhodamine B assay after FKC treatment. Induction of apoptosis was examined by established morphological and biochemical assays. ROS generation was determined by dichlorofluorescein fluorescence staining, and superoxide dismutase activity was measured using the spectrophotometric method. Western blotting was used to examine the changes in the protein levels. FKC markedly decreased the cell viability of HT-29 cells and the cells showed dramatic changes in cellular and nuclear morphologies with typical apoptotic features. The induction of apoptosis correlated well with the externalization of phosphatidylserine, DNA fragmentation, decreased mitochondrial membrane potential, activation of caspases, and PARP cleavage. This was associated with an increase in reactive oxygen species and a decrease in SOD activity. The protein levels of XIAP, c-IAP1, and c-IAP2 were downregulated, whereas the GADD153 was upregulated after FKC treatment. FKC induced cell cycle arrest at the G(1) and G(2)/M phases via upregulation of p21 and p27 in a p53-independent manner. Our results provide evidence that FKC has the potential to be developed into chemotherapeutic drug for the treatment of colon adenocarcinoma. SUMMARY: Flavokawain C inhibited the growth of HT-29 human colon adenocarcinoma cells. Flavokawain C induced apoptosis in HT-29 cells, associated with an increase in reactive oxygen species and a decrease in SOD activity. Flavokawain C induced cell cycle arrest at the G(1) and G(2)/M phases via upregulation of p21 and p27 in HT-29 cells. HT-29 cells treated with flavokawain C caused downregulation of XIAP, c-IAP1, and c-IAP2, and upregulation of GADD153. Abbreviations used: FKC: Flavokawain C; SRB: Sulforhodamine B; ROS: Reactive oxygen species; SOD: Superoxide dismutase; PARP: Poly(ADP-ribose) polymerase; ER: Endoplasmic reticulum; IAPs: Inhibitor of apoptosis proteins; TUNEL: Transferase dUTP nick end labeling; Annexin V-FITC: Annexin V conjugated with fluorescein isothicyanate
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spelling pubmed-55381742017-08-14 Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins Phang, Chung-Weng Karsani, Saiful Anuar Abd Malek, Sri Nurestri Pharmacogn Mag Original Article Chalcones have been shown to exhibit anti-cancer properties by targeting multiple molecular pathways. It was, therefore, of interest to investigate flavokawain C (FKC), a naturally occurring chalcone, which can be isolated from Kava (Piper methysticum Forst) root extract. The aim of this study was to investigate the inhibitory effect of FKC on the growth of HT-29 cells and its underlying mechanism of action. Cell viability of HT-29 cells was assessed by Sulforhodamine B assay after FKC treatment. Induction of apoptosis was examined by established morphological and biochemical assays. ROS generation was determined by dichlorofluorescein fluorescence staining, and superoxide dismutase activity was measured using the spectrophotometric method. Western blotting was used to examine the changes in the protein levels. FKC markedly decreased the cell viability of HT-29 cells and the cells showed dramatic changes in cellular and nuclear morphologies with typical apoptotic features. The induction of apoptosis correlated well with the externalization of phosphatidylserine, DNA fragmentation, decreased mitochondrial membrane potential, activation of caspases, and PARP cleavage. This was associated with an increase in reactive oxygen species and a decrease in SOD activity. The protein levels of XIAP, c-IAP1, and c-IAP2 were downregulated, whereas the GADD153 was upregulated after FKC treatment. FKC induced cell cycle arrest at the G(1) and G(2)/M phases via upregulation of p21 and p27 in a p53-independent manner. Our results provide evidence that FKC has the potential to be developed into chemotherapeutic drug for the treatment of colon adenocarcinoma. SUMMARY: Flavokawain C inhibited the growth of HT-29 human colon adenocarcinoma cells. Flavokawain C induced apoptosis in HT-29 cells, associated with an increase in reactive oxygen species and a decrease in SOD activity. Flavokawain C induced cell cycle arrest at the G(1) and G(2)/M phases via upregulation of p21 and p27 in HT-29 cells. HT-29 cells treated with flavokawain C caused downregulation of XIAP, c-IAP1, and c-IAP2, and upregulation of GADD153. Abbreviations used: FKC: Flavokawain C; SRB: Sulforhodamine B; ROS: Reactive oxygen species; SOD: Superoxide dismutase; PARP: Poly(ADP-ribose) polymerase; ER: Endoplasmic reticulum; IAPs: Inhibitor of apoptosis proteins; TUNEL: Transferase dUTP nick end labeling; Annexin V-FITC: Annexin V conjugated with fluorescein isothicyanate Medknow Publications & Media Pvt Ltd 2017-07 2017-07-11 /pmc/articles/PMC5538174/ /pubmed/28808400 http://dx.doi.org/10.4103/0973-1296.210180 Text en Copyright: © 2017 Pharmacognosy Magazine http://creativecommons.org/licenses/by-nc-sa/3.0 This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 3.0 License, which allows others to remix, tweak, and build upon the work non-commercially, as long as the author is credited and the new creations are licensed under the identical terms.
spellingShingle Original Article
Phang, Chung-Weng
Karsani, Saiful Anuar
Abd Malek, Sri Nurestri
Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins
title Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins
title_full Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins
title_fullStr Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins
title_full_unstemmed Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins
title_short Induction of Apoptosis and Cell Cycle Arrest by Flavokawain C on HT-29 Human Colon Adenocarcinoma via Enhancement of Reactive Oxygen Species Generation, Upregulation of p21, p27, and GADD153, and Inactivation of Inhibitor of Apoptosis Proteins
title_sort induction of apoptosis and cell cycle arrest by flavokawain c on ht-29 human colon adenocarcinoma via enhancement of reactive oxygen species generation, upregulation of p21, p27, and gadd153, and inactivation of inhibitor of apoptosis proteins
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5538174/
https://www.ncbi.nlm.nih.gov/pubmed/28808400
http://dx.doi.org/10.4103/0973-1296.210180
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