Gamma-tocotrienol induced apoptosis is associated with unfolded protein response in human breast cancer cells☆
Introduction
Vitamin E includes a family of lipophilic micronutrients consisting of four tocopherols (Ts) and four tocotrienols (T3s) (α, β, γ and δ) that consist of a chromanol ring and a side chain. Both Ts and T3s are found in various components of the human diet. Tocopherols are present primarily in nuts and vegetable oils, while T3s are minor plant constituents especially abundant in rice bran, cereal grain and palm oil. Tocopherols and T3s are well recognized for their antioxidative effects. In general, antioxidants are suggested to reduce cancer by arresting free-radical-induced DNA damage. Tocopherols have been studied in great detail for their antioxidative property and physiological relevance. However, limited studies have been performed on T3s. It is well documented that T3s possess more powerful anticancer, neuroprotective and cholesterol-lowering properties that are often not exhibited by Ts [1]. The accumulation of T3s in the cells is much greater than Ts; this might be one of the reasons that T3s have more significant physiological effects than Ts [2]. Tocotrienols, particularly γ-T3, suppress the activity of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, an enzyme involved in cholesterol biosynthesis in the liver, resulting in less cholesterol being manufactured by the liver cells and an overall reduction of plasma cholesterol levels [1], [3]. γ-T3 possesses a hormone-like natriuretic function and can potentially prevent hypertension and cardiovascular disease caused by high salt intake [4].
Tocotrienols have been shown to inhibit the growth of various cancer cells without affecting the growth of normal cells [5], [6]. In breast cancer cells, γ-T3s induce apoptosis irrespective of their estrogen response status [7], [8], [9]. Although T3s are likely to be one of the powerful cancer chemotherapeutic/preventive agents in the human diet, their exact mechanisms of action on cell death and other inhibitory pathways are unknown. Gene expression profiling in estrogen receptor positive, p53 wild-type MCF-7 and estrogen receptor-negative, p53 mutant MDA-MB 231 cells treated with tocotrienol-rich fraction (TRF) of palm oil suggested different mechanisms in the two cell lines [10]. Lipid peroxidation is one of the mechanisms suggested for its antiproliferative action [11]. Other mechanisms involve modulation of various signaling pathways including apoptosis by caspase-8 activation and mitochondrial dependency [12], [13], inhibition of cell proliferation [6], down-regulation of cyclins [14], reduction in the Pl3K/PDK-1/Akt signaling and NFκb activity [15], [16] and modulation of p53, Bax/Bcl2 [17]. Estrogens are known to be involved in breast carcinogenesis. Tocotrienols have been demonstrated as antioxidants against the E2-epoxide-induced breast cancer carcinogenesis, whereas α-tocopherol was found to be less effective [18]. The effects of γ-T3 have also been studied in other cancers such as colon and prostate cancer where γ-T3 has been found to modulate multiple signaling pathways and induce apoptotic cell death. Activation of p53 has been reported in RKO human colon carcinoma cells in response to TRF [17]. Recently, Yap and colleagues reported the modulation of ID family proteins and mesenchymal markers in prostate and breast cancer cells in response to γ-T3 [19], [20].
In the present study, we examined the whole genome transcription in MCF-7 breast cancer cells when exposed to γ-T3. We demonstrate that γ-T3-induced apoptosis is associated with induction of early response genes and ER stress transcriptional response in MCF-7 cells. We have characterized the ER stress response induced by γ-T3 in MCF-7 and MDA-MB 231 breast cancer cells.
Section snippets
Cell culture and media
Human breast cancer cells (MCF-7 and MDA-MB 231) and immortalized normal human breast mammary MCF-10A cells were obtained from Lombardi Comprehensive Cancer Center cell repository and grown in Dulbecco's modified Eagle medium supplemented with 5% heat-inactivated fetal bovine serum and 25 μg/ml gentamicin (Invitrogen, Carlsbad, CA, USA).
Chemicals, reagents and antibodies
Tocotrienols (>95% pure) were from EISAI Corporation (Woodcliff Lake, NJ, USA) and Carotech (Edison, NJ, USA) and dissolved in dimethyl sulfoxide (DMSO). Other
Effects of α- and γ-T3s on the proliferation of human breast cancer cells
We compared the effects of α- and γ-T3 on the viability of MCF-7 and MDA-MB 231 cells using WST-1 assay. Both α- and γ-T3 inhibited cell proliferation in a dose-dependent manner when treated with 10–40 μM of each T3 for 24 h (Fig. 1A, B). γ-T3 exhibited a slightly better cell growth inhibitory effect at 20–40 μM as compared with α-T3 in both the cell lines (P<.05). The inhibitory effect of γ-T3 on MCF-7 and MDA-MB 231 cells was also confirmed by trypan blue dye exclusion assay (data not shown).
Discussion
Tocotrienols have gained attention due to their powerful anticancer, neuroprotective and cholesterol-lowering properties not exhibited by Ts [1]. The accumulation of T3s in the cells is much greater than Ts; this might be one of the reasons that T3s have more significant effects than Ts [2]. Tocotrienols inhibit the growth of various cancer cell lines without affecting the growth of normal cells [5], [6]; our results confirmed that, at higher concentrations of γ-T3, only the growth of breast
Acknowledgments
The studies were conducted using the Tissue Culture, Flow Cytometry and Cell Sorting and Macromolecular Analysis Shared Resources of the Lombardi Comprehensive Cancer Center. We thank Karen Creswell, Xiaojun Zou and James Li for providing excellent technical assistance.
References (39)
- et al.
Tocotrienols: vitamin E beyond tocopherols
Life Sci
(2006) - et al.
Inhibition of THP-1 cell adhesion to endothelial cells by alpha-tocopherol and alpha-tocotrienol is dependent on intracellular concentration of the antioxidants
Free Radic Biol Med
(2003) - et al.
Gamma-tocotrienol, a vitamin E homolog, is a natriuretic hormone precursor
J Lipid Res
(2003) - et al.
Tocotrienol-rich fraction of palm oil induces cell cycle arrest and apoptosis selectively in human prostate cancer cells
Biochem Biophys Res Commun
(2006) - et al.
Identities and differences in the metabolism of tocotrienols and tocopherols in HepG2 cells
J Nutr
(2002) - et al.
Disruption of mitochondria during tocotrienol-induced apoptosis in MDA-MB-231 human breast cancer cells
Biochem Pharmacol
(2004) - et al.
The effect of alpha- and gamma-tocopherol and their carboxyethyl hydroxychroman metabolites on prostate cancer cell proliferation
Arch Biochem Biophys
(2004) - et al.
Evidence for the preventive effect of the polyunsaturated phytol side chain in tocotrienols on 17beta-estradiol epoxidation
Cancer Detect Prev
(2005) - et al.
Identification of a novel tumor necrosis factor-alpha-inducible gene, SCC-S2, containing the consensus sequence of a death effector domain of fas-associated death domain-like interleukin-1beta-converting enzyme-inhibitory protein
J Biol Chem
(2000) - et al.
SCC-112, a novel cell cycle-regulated molecule, exhibits reduced expression in human renal carcinomas
Gene
(2004)
Tocotrienols: the emerging face of natural vitamin E
Vitam Horm
gamma-Tocotrienol induces mitochondria-mediated apoptosis in human gastric adenocarcinoma SGC-7901 cells
JNutrBiochem
Proteasome-dependent degradation of cyclin D1 in 1-methyl-4-phenylpyridinium ion (MPP+)-induced cell cycle arrest
J Biol Chem
Effects of administration of alpha-tocopherol and tocotrienols on serum lipids and liver HMG CoA reductase activity
Int J Food Sci Nutr
Induction of apoptosis by tocotrienol in rat hepatoma dRLh-84 cells
Anticancer Res
Tocotrienols inhibit the growth of human breast cancer cells irrespective of estrogen receptor status
Lipids
Tocotrienols inhibit growth of ZR-75-1 breast cancer cells
Int J Food Sci Nutr
Tocotrienol-rich fraction from palm oil affects gene expression in tumors resulting from MCF-7 cell inoculation in athymic mice
Lipids
Studies on the biological activity of tocotrienols
Chem Pharm Bull (Tokyo)
Cited by (0)
- ☆
Financial support (DK): NCI UDC-LCCC U56, UDC Agricultural Experiment Station, MBRS SCORE, NSF HBCU-UP program.
- 1
These authors contributed equally.