Breast cancer is the most common cancer in women worldwide. The growth of breast cancer cells is either hormone-dependent or hormone-independent. Both types are represented in vitro by the estrogen-receptor positive (ER+) MCF-7 and the estrogen-receptor negative (ER-) MDA-MB-231 cell lines, respectively. The pS2 gene is an estrogen-regulated gene and serves as a marker for the ER+ tumours. Carotenoids are pigments with anti-cancer properties besides having pro-vitamin A, antioxidant and free-radical quenching effects. This study was designed firstly, to compare the effect of palm oil carotene concentrate with retinoic acid on the growth of the ER+ MCF-7 and the ER- MDA-MB-231 cells; and secondly to evaluate the effect of the palm oil carotene concentrate on the regulation of pS2 mRNA. The growth experiments were performed with monolayer cells seeded in phenol red free RPMI 1640 culture media and subsequently treated with varying concentrations of either retinoic acid or palm oil carotenoids. The cell numbers were determined at the start of each experiment and then at successive time intervals. The results showed that the palm oil carotene concentrate caused dose-dependent inhibition of estradiol-stimulated growth of MCF-7 cells but did not affect the proliferation of MDA-MB-231 cells. Retinoic acid caused similar, albeit more potent effects, as significant inhibition was observed at lower concentrations than the palm oil carotenoids. In the pS2 gene expression experiment, cell monolayers were treated with the carotene concentrate (10(-6) M), either with or without supplemented estradiol (10(-8) M), and subsequently the RNA was extracted. Northern blotting was performed and the regulation of pS2 mRNA determined using a 32P-labelled pS2 cDNA probe. The results showed that the palm oil carotene concentrate did not affect the expression of pS2 mRNA and are therefore independent of the estrogen-regulated pathway.
Oestrogen is important in the development of breast cancer. Oestrogen receptor positive breast cancers are associated with a better prognosis than oestrogen-receptor negative breast cancers since they are more responsive to hormonal treatment. Oestrone sulphate acts as a huge reservoir for oestrogens in the breast. It is converted to the potent oestrogen, oestradiol (E(2)) by the enzymes oestrone sulphatase and oestradiol-17beta hydroxysteroid dehydrogenase (E(2)DH). Retinoic acid and carotenoids have been shown to have chemopreventive activity against some cancers. The aim of our study was to determine and compare the effects of retinoic acid and palm oil carotenoids on growth of and oestrone sulphatase and E(2)DH activities in the oestrogen receptor positive, MCF-7 and oestrogen receptor negative, MDA-MB-231 breast cancer cell lines. Retinoic acid and carotenoids inhibited MCF-7 cell growth but had no effect on MDA-MB-231 cell growth. Both retinoic acid and carotenoids stimulated oestrone sulphatase activity in the MCF-7 cell line. E(1) to E(2) conversion was inhibited by 10(-7) M carotenoids but was stimulated at 10(-6) M in the MCF-7 cell line. Retinoic acid had no effect on E(1) to E(2) conversion at 10(-7) M but stimulated E(1) to E(2) conversion at 10(-6) M. Retinoic acid and carotenoids had no effect on E(2) to E(1) conversion in the MCF-7 cell line. Retinoic acid stimulated E(1) to E(2) conversion in the MDA-MB-231 cell line but had no effect on oestrone sulphatase activity or E(2) to E(1) conversion in this cell line. Both oestrone sulphatase and E(2)DH activity were not affected by carotenoids in the MDA-MB-231 cell line. In conclusion, retinoic acid and carotenoids may prevent the development of hormone-dependent breast cancers since they inhibit the growth of the MCF-7 cell line.
Oestrogens play an important role in the development of breast cancer. Oestrone sulphate (E1S) acts as a huge reservoir of oestrogens in the breast and is converted to oestrone (E1) by oestrone sulphatase (E1STS). E1 is then reversibly converted to the potent oestrogen, oestradiol (E2) by oestradiol-17beta hydroxysteroid dehydrogenase (E2DH). The aim of this study was to assess the effects of transforming growth factor-beta1 (TGFbeta1), insulin-like growth factor-I (IGF-I) and insulin-like growth factor-II (IGF-II) on cell growth, E1STS and E2DH activities in the MCF-7 and MDA-MB-231 human breast cancer cell lines. TGFbeta1, IGF-I and IGF-II alone or in combination inhibited cell growth of both cell lines but no additive or synergistic effects were observed. The treatments significantly stimulated E1STS activity in the MCF-7 cell line, except for TGFbeta1 alone and TGFbeta1 and IGF-I in combination, where no effects were seen. Only TGFbeta1 and IGF-II acted synergistically to stimulate E1STS activity in the MCF-7 cells. There was no significant effect on E1STS activity in the MDA-MB-231 cells with any of the treatments. In the MCF-7 cells, TGFbeta1 and IGF-I, IGF-I and IGF-II, and TGFbeta1, IGF-I and IGF-II acted synergistically to stimulate the reductive E2DH activity, while only TGFbeta1, IGF-I and IGF-II synergistically stimulated the oxidative E2DH activity. There were no additive or synergistic effects on both oxidative and reductive E2DH activities in the MDA-MB-231 cells. In conclusion, TGFbeta1, IGF-I and IGF-II may have effects on oestrogen metabolism, especially in the MCF-7 cell line where they stimulated the conversion of E1S to E1 and E1 to E2 and, thus, may have roles to play in the development of breast cancer.
Oestrone sulphate is a major source of active oestrogens in the breast. It is converted to oestrone by oestrone sulphatase. Breast cyst fluid (BCF) is a rich source of sex hormones and growth factors. BCF obtained from British women has been shown to inhibit oestrone sulphatase activity in the MCF-7 oestrogen-receptor-positive breast cancer cell line. The aim of the present study was to assess whether BCF obtained from Malaysian women inhibited oestrone sulphatase activity in the MCF-7 and MDA-MB-231 breast cancer cell lines. The cell lines were grown in supplemented Dulbecco's Modified Eagle Medium for 3 days, following which a 3-day incubation with sterilised BCF was carried out. At the end of the treatment period the cell monolayers were assayed for oestrone sulphatase activity and the number of cell nuclei counted on a Coulter Counter. BCF was also fractionated on a Bio-Sil SEC 125-5 column by HPLC and the effects of the fractions collected on oestrone sulphatase activity in the MDA-MB-231 cell line were assessed. All 18 samples of BCF tested inhibited cell growth in the MDA-MB-231 cell line while 8 out of 10 samples inhibited MCF-7 cell growth; 15 out of 18 BCF samples inhibited oestrone sulphatase activity in the MDA-MB-231 cell line whereas 5 out of 10 samples stimulated oestrone sulphatase activity in the MCF-7 cell line. HPLC fractions corresponding to molecular weights of > 158 kDa and 28 kDa were found to inhibit oestrone sulphatase activity in the MDA-MB-231 cell line. Further work is required to fully characterise these substances as they may have roles to play in the prevention of breast cancer.
Transforming growth factor-beta (TGF-beta) has been shown to inhibit the growth of mammary epithelial cells and may play a protective role in mammary carcinogenesis. In contrast, oestrogens promote the development of breast cancer. Oestrone sulphate (E1S) is a huge reservoir of active oestrogens in the breast being converted to the weak oestrogen, oestrone (E1), by oestrone sulphatase. E1 is reversibly converted by oestradiol-17beta hydroxysteroid dehydrogenase to the potent oestrogen, oestradiol (E2). The aim of this study was to assess the effect of the TGF-beta1 isoform on growth and oestrogen metabolism in the hormone-dependent MCF-7 and hormone-independent MDA-MB-231 human breast cancer cell lines. The results showed that TGF-beta1 significantly inhibited cell growth and stimulated the conversion of E1S to E1 and E1 to E2 in the MCF-7 cell line. In the MDA-MB-231 cell line TGF-beta1 significantly stimulated cell growth and inhibited the interconversions between E1 and E2. In conclusion, the growth inhibitory effect of TGF-beta1 on the MCF-7 cell line would appear to confer a protective effect in breast cancer. However, its ability to increase the amount of E2 would increase the risk of breast cancer. Which of these effects predominates in vivo remains to be explored. The growth stimulatory effect of TGF-beta1 on the MDA-MB-231 cell line probably acts through a mechanism independent of the effect of TGF-beta1 on oestrogen concentrations since this cell line is hormone unresponsive.
Oestrogens play an important role in the development of breast cancer. A very important source of active oestrogens in the breast is oestrone sulphate which is converted to oestrone by oestrone sulphatase. The aim of this study was to assess the effects of IGF-I and IGF-II on oestrone sulphatase activity in, as well as cell growth of, MCF-7 and MDA-MB-231 human breast cancer cell lines. Cells were grown in supplemented DMEM and treated with varying concentrations of IGFs. At the end of the treatment period, intact cell monolayers were washed and assayed for oestrone sulphatase activity and the number of cell nuclei determined on a Coulter Counter. Oestrone sulphatase activity was significantly stimulated by IGF-I and II at concentrations of 100 ng/ml and 200 ng/ml in MCF-7 cells. IGF-I had no effect on oestrone sulphatase activity in MDA-MB-231 cells over the range of concentrations tested. Significant inhibition of oestrone sulphatase was observed in MDA-MB-231 cells at higher concentrations of IGF-II (50 ng/ml, 100 ng/ml and 200 ng/ml). Both IGF-I and IGF-II at higher concentrations (100 ng/ml and 200 ng/ml) significantly inhibited MCF-7 and stimulated MDA-MB-231 cell growth. Since IGF-I and II have effects on cell growth and oestrone sulphatase activity in breast cancer cell lines they may play a role in the development and progression of human breast cancer.
It has recently been shown that tocotrienols are the components of vitamin E responsible for inhibiting the growth of human breast cancer cells in vitro, through an estrogen-independent mechanism. Although tocotrienols act on cell proliferation in a dose-dependent manner and can induce programmed cell death, no specific gene regulation has yet been identified. To investigate the molecular basis of the effect of tocotrienols, we injected MCF-7 breast cancer cells into athymic nude mice. Mice were fed orally with 1 mg/d of tocotrienol-rich fraction (TRF) for 20 wk. At end of the 20 wk, there was a significant delay in the onset, incidence, and size of the tumors in nude mice supplemented with TRF compared with the controls. At autopsy, the tumor tissue was excised and analyzed for gene expression by means of a cDNA array technique. Thirty out of 1176 genes were significantly affected. Ten genes were downregulated and 20 genes up-regulated with respect to untreated animals, and some genes in particular were involved in regulating the immune system and its function. The expression of the interferon-inducible transmembrane protein-1 gene was significantly up-regulated in tumors excised from TRF-treated animals compared with control mice. Within the group of genes related to the immune system, we also found that the CD59 glycoprotein precursor gene was up-regulated. Among the functional class of intracellular transducers/effectors/modulators, the c-myc gene was significantly down-regulated in tumors by TRF treatment. Our observations indicate that TRF supplementation significantly and specifically affects MCF-7 cell response after tumor formation in vivo and therefore the host immune function. The observed effect on gene expression is possibly exerted independently from the antioxidant activity typical of this family of molecules.
Biological therapies are new additions to breast cancer treatment. Among biological compounds, beta-carotene has been reported to have immune modulatory effects, in particular, enhancement of natural killer cell activity and tumor necrosis factor-alpha production by macrophages. The objective of this study was to investigate the effect of palm carotene supplementation on the tumorigenicity of MCF-7 human breast cancer cells injected into athymic nude mice and to explore the mechanism by which palm carotenes suppress tumorigenesis. Forty-eight 4-wk-old mice were injected with 1 x 10(6) MCF-7 cells into their mammary fat pad. The experimental group was supplemented with palm carotene whereas the control group was not. Significant differences were observed in tumor incidence (P< 0.001) and tumor surface area and metastasis to lung (P< 0.005) between the two groups. Natural killer (NK) cells and B-lymphocytes in the peripheral blood of carotene-supplemented mice were significantly increased (P < 0.05 and P < 0.001, respectively) compared with controls. These results suggest that palm oil carotene is able to modulate the immune system by increasing peripheral blood NK cells and B-lymphocytes and suppress the growth of MCF-7 human breast cancer cells.