Effect of dietary GLA+/-tamoxifen on the growth, ER expression and fatty acid profile of ER positive human breast cancer xenografts

Gamma linolenic acid (GLA) possesses a number of selective anti-tumour properties including modulation of steroid receptor structure and function. We have investigated the effect of dietary GLA on the growth, oestrogen receptor (ER) expression and fatty acid profile of ER+ve human breast cancer xenografts. Experimental diets A, B, C, D were commenced after subcutaneous implantation of 40 female nude mice with the MCF-7 B1M cell line (Group A = control diet: B = control diet + GLA supplement: C = control diet + tamoxifen: D = control diet + GLA + tamoxifen; 10 mice/group). The mice were terminated when tumour cross-sectional area reached 250 mm(2). ER H-scores were assessed by immunohistochemical assay and fatty acid profiles by gas-liquid chromatography of termination tumour samples. Groups C and D displayed significantly slower tumour growth (p =.0002, p =.0006) with trend for slower growth in B (p =.065) compared to control Group A. ER was significantly reduced in all groups compared to A (p <.0001) with Group D (combined therapy) displaying markedly lower ER expression than with either therapy alone (p =.0002). There were significantly raised levels of tumour GLA and metabolites in the two groups (B and D) receiving GLA (p <.0001). This xenograft model of ER+ve breast cancer has demonstrated significantly lower tumour ER expression in those groups receiving GLA, an effect which appears to be additive to the reduced ER expression resulting from tamoxifen alone. The effects of GLA on ER function and the possibility of synergistic inhibitory action of GLA with tamoxifen via enhanced down-regulation of the ER pathway require further investigation.     

Menopause and the skin

Women live one-third of their lives in the post-menopausal state. Significant hormonal alterations occur at the time of menopause, leading to a range of physiological disorders affecting multiple organ systems in the body. The effects of menopause on the skin have been underresearched. Many skin changes occur at the time of menopause and the cutaneous effects of hormone replacement therapy are significant. Menopausal changes in hormones may alter the biomechanical properties of the skin and certain disorders are more common in menopausal women, such as lichen sclerosus, atrophic vulvovaginitis, flushing and dysaesthetic vulvodynia. Hair and oral changes may also be associated. As the average life expectancy increases, dermatologists need to be familiar with skin diseases affecting women in this age group.     

Oestrogen receptor beta-immunoreactivity in gonadotropin releasing hormone-expressing neurones: regulation by oestrogen

Double-label immunohistochemistry was employed to establish whether immunoreactivity for the beta subtype of the oestrogen receptor (ER beta-IR) is present in gonadotropin releasing hormone (GnRH)-containing cells. In the immortalized GnRH cell line, GT1-7, almost all nuclei were immunoreactive for ER beta. In the preoptic area of ovariectomized rats, more than one-half of the GnRH neurones (52.0-63.5%) contained ER beta-IR within the nucleus; a smaller proportion of these neurones (5-10%) displayed a particularly intense nuclear signal for ER beta. The presence of ER beta-IR in the nuclei of GT1-7 cells and GnRH neurones is consistent with recent reports of ER beta mRNA in these cells. Oestrogen treatment reduced the percentage of GnRH neurones with detectable ER beta-IR. The range of signal intensity for ER beta and the incidence of the ER beta signal in GnRH neurones were comparable following double-label immunohistochemistry involving either bright field or fluorescent techniques. These findings raise the possibility that ER beta receptors mediate direct effects of oestrogen on GnRH neurones.     

Rapid Action of Oestrogen in Luteinising Hormone-Releasing Hormone Neurones: The Role of GPR30

Previously, we have shown that 17β-oestradiol (E₂) induces an increase in firing activity and modifies the pattern of intracellular calcium ([Ca²⁺]_i) oscillations with a latency < 1 min in primate luteinising hormone-releasing hormone (LHRH) neurones. A recent study also indicates that E₂, the nuclear membrane impermeable oestrogen, oestrogen-dendrimer conjugate, and the plasma membrane impermeable oestrogen, E₂-BSA conjugate, all similarly stimulated LHRH release within 10 min of exposure in primate LHRH neurones, indicating that the rapid action of E₂ is caused by membrane signalling. The results from a series of studies further suggest that the rapid action of E₂ in primate LHRH neurones appears to be mediated by GPR30. Although the oestrogen receptor antagonist, ICI 182, 780, neither blocked the E₂-induced LHRH release nor the E₂-induced changes in [Ca²⁺]_i oscillations, E₂ application to cells treated with pertussis toxin failed to result in these changes in primate LHRH neurones. Moreover, knockdown of GPR30 in primate LHRH neurones by transfection with human small interference RNA for GPR30 completely abrogated the E₂-induced changes in [Ca²⁺]_i oscillations, whereas transfection with control siRNA did not. Finally, the GPR30 agonist, G1, resulted in changes in [Ca²⁺]_i oscillations similar to those observed with E₂. In this review, we discuss the possible role of G-protein coupled receptors in the rapid action of oestrogen in neuronal cells.     

Biological characteristics of breast cancer at the primary tumour and the involved lymph nodes

Diminished oestrogen receptor (ER) expression in the involved axillary lymph nodes (ALN) in breast cancer compared with the primary tumour has been reported in previous studies. We have assessed a wider spectrum of tumour markers (ER, progesterone receptor (PgR), p53, Ki-67 and HER-2/neu) and compared extent and staining intensities at the primary tumour and the involved ALN on specimens of 22 cases with invasive ductal breast cancer. At the involved ALN, both the quantity of positive staining cells and the staining intensities for ER and PgR were decreased (p < 0.001 and p = 0.003, respectively). In contrast, the quantity of positive staining cells (p < 0.004) and the staining intensities for Ki-67 were increased. The differences for HER-2/neu and p53 staining at both sites were insignificant. The immunohistochemical staining properties of both the primary tumour and the ALN metastases showed no correlation with the number of involved ALN (p > 0.05). This study suggested that ALN metastasis might indicate a more unfavourable expression pattern of ER, PgR and Ki-67 in invasive ductal breast cancer.     

Previous Midlife Oestradiol Treatment Results in Long‐Term Maintenance of Hippocampal Oestrogen Receptor α Levels in Ovariectomised Rats: Mechanisms and Implications for Memory

Ovariectomised rats that have received previous administration of oestradiol in midlife display enhanced cognition and increased hippocampal levels of oestrogen receptor (ER)α months after oestradiol treatment ended compared to ovariectomised controls. The present study aimed to investigate the mechanisms by which ERα levels are maintained following midlife oestradiol exposure and the role of ERα in memory in ageing females in the absence of circulating oestrogens. Unliganded ERα has increased interaction with the ubiquitin ligase, C‐terminus of Hsc‐70 interacting protein (CHIP), leading to increased degradation of the receptor. In our first experiment, we tested the hypothesis that midlife oestradiol exposure in ovariectomised rats results in decreased interaction between CHIP and hippocampal ERα, leading to increased levels of ERα. Middle‐aged rats were ovariectomised and received oestradiol or vehicle implants. After 40 days, implants were removed. One month later, rats were killed and hippocampi were processed for whole protein western blotting and co‐immunoprecipitation, in which ERα was immunoprecipitated from lysate. As expected, ERα protein expression was increased in rats previously treated with oestradiol compared to vehicle‐treated rats. In rats treated with oestradiol, there was a decrease in CHIP–ERα interaction, suggesting that previous oestradiol treatment reduces interaction, slowing the degradation of ERα. In a second experiment, we determined the impact on memory of antagonism of ER in the absence of circulating oestrogens. Rats were ovariectomised and implanted with oestradiol capsules. Capsules were removed after 40 days. Rats received chronic i.c.v. infusion of ER antagonist, ICI 182 780, or artificial cerebrospinal fluid vehicle and were tested on a spatial memory radial‐maze task. Rats treated with ICI 182 780 had significantly worse performance (more errors). These experiments provide evidence that previous midlife oestradiol treatment maintains hippocampal ERα by decreasing its interaction with CHIP and that activation of these receptors provides cognitive benefits in the absence of circulating oestrogens.     

Normal bone physiology,remodelling and its hormonal regulation

The skeleton has structural and locomotor functions, and is a mineral reservoir. Bone turnover by osteoclasts and osteoblasts is a lifelong process, incorporating growth, modelling and remodelling to repair microdamage and access the mineral reservoir. Signalling between bone cells is essential for the co-ordination of these processes. Osteoblasts regulate osteoclast activity through the RANK/RANK ligand/OPG system, and osteocytes regulate osteoblast activity through sclerostin secretion. If resorption and formation are balanced there is no net change in bone mass after each remodelling cycle, but with ageing and some disease states resorption exceeds formation, leading to negative bone balance, decreased bone mass and loss of microstructural integrity. The rate of remodelling is determined by factors including mechanical loading and endocrine influences. The most important endocrine regulator of bone turnover is oestrogen, but other hormones regulating bone metabolism include IGF-1, PTH and gut and adipocyte hormones.