β-sitosterol induces G1 arrest and causes depolarization of mitochondrial membrane potential in breast carcinoma MDA-MB-231 cells

Shanthi Sri Vundru,Raosaheb K Kale,Rana P Singh

doi:10.1186/1472-6882-13-280

Abstract

BackgroundsIt is suggested that dietary phytosterols, such as β-sitosterol (ST), have cancer chemopreventive effects; however, studies are limited to support such claims. Here, we evaluated the efficacy of ST on three different human cancer cell lines including skin epidermoid carcinoma A431 cells, lung epithelial carcinoma A549 cells and breast adenocarcinoma MDA-MB-231.MethodsCell growth assay, cell cycle analysis, FACS, JC-1 staining, annexin V staining and immunoblotting were used to study the efficacy of ST on cancer cells.ResultsST (30–90 μM) treatments for 48 h and 72 h did not show any significant effect on cell growth and death in A431 cells. Whereas similar ST treatments moderately inhibited the growth of A549 cells by up to 13% (p ≤ 0.05) in 48 h and 14% (p ≤ 0.05-0.0001) in 72 h. In MDA-MB-231 cells, ST caused a significant dose-dependent cell growth inhibition by 31- 63% (p ≤ 0.0001) in 48 h and 40-50% (p ≤ 0.0001) in 72 h. While exploring the molecular changes associated with strong ST efficacy in breast cancer cells, we observed that ST induced cell cycle arrest as well as cell death. ST caused G0/G1 cell cycle arrest which was accompanied by a decrease in CDK4 and cyclin D1, and an increase in p21/Cip1and p27/Kip1 protein levels. Further, cell death effect of ST was associated with induction of apoptosis. ST also caused the depolarization of mitochondrial membrane potential and increased Bax/Bcl-2 protein ratio.ConclusionsThese results suggest prominent in vitro anti-proliferative and pro-apoptotic effects of ST in MDA-MB-231 cells. This study provides valuable insight into the chemopreventive efficacy and associated molecular alterations of ST in breast cancer cells whereas it had only moderate efficacy on lung cancer cells and did not show any considerable effect on skin cancer cells. These findings would form the basis for further studies to understand the mechanisms and assess the potential utility of ST as a cancer chemopreventive agent against breast cancer.

Highlights

Among many cancers, breast cancer in females and lung cancer in males are the most frequently diagnosed cancers and the leading cause of cancer death for each sex in both economically developed and developing countries [1]
Cell lines and reagents A431 and A549 cells were from ATCC (Manassas, USA) and MDA-MB-231 cells were from NCCS Pune, India
We observed an increase in p21/Cip1 and p27/Kip1 protein levels which were relatively higher at 90 μM concentration of ST and may explain the effect for G0/G1 arrest compared to control (Figure 2A). These results suggest that ST induced G0/G1 arrest in MDA-MB-231 cells could be mediated via modulation of cyclin-dependent kinase (CDK)-cyclin-Cyclin-dependent kinase inhibitors (CDKI) protein levels

Summary

Introduction

Breast cancer in females and lung cancer in males are the most frequently diagnosed cancers and the leading cause of cancer death for each sex in both economically developed and developing countries [1]. The use of natural, synthetic or biological agents to prevent, reverse or suppress the growth and progression of cancer is referred as chemoprevention of cancer [3]. It is one of the most promising strategies for cancer control, and is accomplished by various means including chemoprevention by phytochemicals from vegetables, fruits, spices, teas, herbs and medicinal plants making it as one of the most feasible means of cancer control [4]. There has been a considerable interest in secondary plant metabolites because of their potential preventative effects on chronic diseases including cancer

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