Actions of plant estrogens on breast cancer cells as demonstrated by flow cytometry

Date

2009-11-19T21:57:37Z

Authors

Cosby, Cheri Charlene

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Abstract

The activity of plant estrogens, genistein (GEN) and enterolactone (ENL), on mammalian cells is not well understood. Previously, they have been shown to exhibit a biphasic effect, being stimulatory at low doses and inhibitory at high doses. Therefore, consumption of plant estrogens by women with breast cancer has become controversial. Simplified flow cytometry applications to quantify the degree of proliferation and apoptosis were used to measure differentiation and growth inhibitory properties of GEN and/or ENL on two estrogen receptor-positive, MCF-7 and T47D, and two estrogen receptor-negative, MDA-MB-231 and MDA-MB-468, breast cancer cell lines in the presence and absence of I7β-estradiol (E2). Activity in the presence of 4-hydroxytamoxifen (OHT) was also evaluated. A live cell assay using propidium iodide (PI) and fluorescein diacetate was used for evaluation of live, apoptotic and necrotic populations. The biological activity of ENL and GEN in tissue culture was confirmed by MTT assay. An Annexin V/PI live cell assay confirmed the presence and degree of apoptosis. No effect was seen at lower doses, ie. less than 10µM and 5011M in estrogen receptor-positive, ER (+), cells and decreased viability was seen at ENL 10 µM in one estrogen receptor-negative, ER (-), cell line. A significant reduction in viability was seen with treatments of 100 µM of ENL and GEN in both ER (+) cell lines. The effect of combining E2 with treatments of GEN 50 and 100 µM was a reduction in viability in one ER (+) cell line. The effect of combining treatments of low doses of ENL and GEN (10 MM) resulted in a reduction in viability in one ER (+) and one ER (-) cell line. Treatment of ER (+) breast cancer cell lines with low dose GEN (10 µM) and high dose ENL and GEN (50 MM) combined with low dose OHT (10 µM ) reduced viability in excess OHT 10 µM alone. High dose OHT (50 µM ) significantly reduced viability in all cell lines. However, when high dose OHT was combined with low dose ENL and GEN, there was an effect of rescuing ER (-) cells from OHT-induced apoptosis and necrosis. Although the use of high dose OHT is not typically used to treat ER (-) breast cancer, these results suggest the action of plant estrogens is more varied than previously thought. This work supports the notion that there may be dissimilarity in the actions between ENL and GEN which may warrant separate recommendations regarding dietary consumption, especially in women with breast cancer.

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Keywords

phytoestrogens, breast cancer

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