綠茶多酚 EGCg 影響人類乳癌細胞株 MCF-7 細胞增生

Abstract

It has been reported that green tea polyphenol (e.g. (-)-epigallocatechin-3-gallate, EGCg）can inhibit human MCF-7 breast cancer cell growth via the signal transduction pathways to modify the activity of fatty acid synthesis. In addition, EGCg has been shown to cause inhibition of tumor cell growth by altering the lipid compositions on cell membranes such as to induce the lipid raft associated with membrane receptors including human estrogen receptor （HER）and epidermal growth factor receptor（EGFR）in many cancer cells. The main goal of this study was to investigate whether EGCg can inhibit human MCF-7 breast cancer cell proliferation via the membrane-evoked signaling pathway to modulate cytoplasmic Ca2+ homeostasis. Using the MTT assay, EGCg was found to attenuate mitochondria-dependent cell viability of MCF-7 cells. Analyses by using western blot, quantitative real time PCR、and immune-cellular-fluorescence microscopy revealed the chemo-protection of EGCg on inhibition of MCF-7 breast cancer cell proliferation. The anti-tumor effects of EGCg included (1) re-activation of tumor-suppressive gene, tropomyosin-1, (2) increases of pro-apoptotic activity of 20 kD caspase-3, (3) decreases of anti-apoptotic protein-gelsolin gene expression, (4) modulation of gene expression and cellular distribution of adherens junction proteins of E-cadherin,β-catenin, and (5) attenuating extracellular secretion of breast cancer cell marker protein, osteopontin. To examine further whether EGCg acts by the membrane-associated signaling pathway to modulate cytoplasmic Ca2+ homeostasis, we first forced EGFP expression in MCF-7 cell and measured the EGCg-dependent decrease in the EGFP fluorescence. The observation that the EGCg-induced decrease of fluorescence intensity was recovered after cell membrane removal indicates that membrane portion is required for the action of EGCg. Since a multitude of intracellular Ca2+ signaling is sensitive to membrane activation, the dosage effect of EGCg was also determined on changes in intracellular Ca2+. The specific receptor on the membrane was identified by applying a specific antibody to laminin receptor (LR) and genistein, an inhibitor of TPK and AKT receptors. Results obtained demonstrated that EGCg can act through the LR and Akt signaling pathway to influence the intracellular Ca2+. Taken together, EGCg acts by modifying the membrane signal transduction to alter intracellular Ca2+ levels, which might promote apoptosis and attenuate MCF-7 cell viability.