Convergencia de vías de señalización en un modelo de apoptosis

La sepsis es un evento inflamatorio generalizado del organismo inducido por un daño causado generalmente por un agente infeccioso. El patógeno más frecuentemente asociado con esta entidad es el Staphylococcus aureus, responsable de la inducción de apoptosis en células endoteliales debida a la producción de ceramida. Se ha descrito el efecto protector de la proteína C activada (PCA) en sepsis y su relación con la disminución de la apoptosis de las células endoteliales. En este trabajo se analizó la activación de las quinasas AKT, ASK1, SAPK/JNK y p38 en un modelo de apoptosis endotelial usando las técnicas de Western Blotting y ELISA. Las células endoteliales (EA.hy926), se trataron con C2-ceramida (130μM) en presencia de inhibidores químicos de cada una de estas quinasas y PCA. La supervivencia de las células en presencia de inhibidores químicos y PCA fue evaluada por medio de ensayos de activación de las caspasas 3, 7 y 9, que verificaban la muerte celular por apoptosis. Los resultados evidencian que la ceramida reduce la activación de AKT y aumenta la activación de las quinasas ASK, SAPK/JNK y p38, en tanto que PCA ejerce el efecto contrario. Adicionalmente se encontró que la tiorredoxina incrementa la activación/fosforilación de AKT, mientras que la quinasa p38 induce la defosforilación de AKT.

Long-chain n−3 PUFA: plant v. marine sources

Increasing recognition of the importance of the long-chain n-3 PUFA, EPA and DHA, to cardiovascular health, and in the case of DHA to normal neurological development in the fetus and the newborn, has focused greater attention on the dietary supply of these fatty acids. The reason for low intakes of EPA and DHA in most developed countries (0 center dot 1-0 center dot 5hairspg/d) is the low consumption of oily fish, the richest dietary source of these fatty acids. An important question is whether dietary intake of the precursor n-3 fatty acid, alpha-linolenic acid (alpha LNA), can provide sufficient amounts of tissue EPA and DHA by conversion through the n-3 PUFA elongation-desaturation pathway. alpha LNA is present in marked amounts in plant sources, including green leafy vegetables and commonly-consumed oils such as rape-seed and soyabean oils, so that increased intake of this fatty acid would be easier to achieve than via increased fish consumption. However, alpha LNA-feeding studies and stable-isotope studies using alpha LNA, which have addressed the question of bioconversion of alpha LNA to EPA and DHA, have concluded that in adult men conversion to EPA is limited (approximately 8%) and conversion to DHA is extremely low (< 0 center dot 1%). In women fractional conversion to DHA appears to be greater (9%), which may partly be a result of a lower rate of utilisation of alpha LNA for beta-oxidation in women. However, up-regulation of the conversion of EPA to DHA has also been suggested, as a result of the actions of oestrogen on Delta 6-desaturase, and may be of particular importance in maintaining adequate provision of DHA in pregnancy. The effect of oestrogen on DHA concentration in pregnant and lactating women awaits confirmation.

Endocrine regulation of ovarian antral follicle development in cattle

Antral follicle growth in cattle occurs in two distinct phases; the first 'slow' growth phase spans the time from antrum acquisition to a size of approximately 3 mm detectable by transrectal ultrasound, and the second 'fast' phase is gondadotrophin-dependent and includes cohort growth, dominant follicle (DF) selection, and DF growth. This review summarises current concepts of the relative roles FSH and LH, ovarian and metabolic hormones play mainly in the second phase of antral follicle growth in animals of different reproductive and nutritional states. It is proposed that differential FSH response may enable one cohort follicle to become selected, and that follicular secretions, particularly inhibin, suppress FSH and thus are responsible for DF selection and dominance. Acute dependence of the DF on LH pulses will determine DF lifespan, and the LH pulse profile can be influenced by metabolic hormones such as leptin, providing one possible link for nutritional state and reproduction. Direct ovarian effects of acute and chronic changes in growth hormone, insulin and insulin-like growth factor (IGF)-I have been described on cohort follicles, DF oestrogen activity and on DF growth. Influences of metabolic hormones on early antral follicles undergoing their first 'slow' growth phase are less well described, yet metabolic hormones appear to enhance growth into the cohort available for FSH-induced emergence, and may influence subsequent developmental competence of oocytes. (C) 2003 Elsevier Science B.V. All rights reserved.

Oestrogenic activity of benzyl salicylate, benzyl benzoate and butylphenylmethylpropional (Lilial) in MCF7 human breast cancer cells in vitro

Benzyl salicylate, benzyl benzoate and butylphenylmethylpropional (Lilial) are added to bodycare cosmetics used around the human breast. We report here that all three compounds possess oestrogenic activity in assays using the oestrogen-responsive MCF7 human breast cancer cell line. At 3 000 000-fold molar excess, they were able to partially displace [H-3]oestradiol from recombinant human oestrogen receptors ER alpha and ER beta, and from cytosolic ER of MCF7 cells. At concentrations in the range of 5 x 10(-5) to 5 x 10(-4) M, they were able to increase the expression of a stably integrated oestrogen-responsive reporter gene (ERE-CAT) and of the endogenous oestrogen-responsive pS2 gene in MCF7 cells, albeit to a lesser extent than with 10(-8) M 17 beta-oestradiol. They increased the proliferation of oestrogen-dependent MCF7 cells over 7 days, which could be inhibited by the antioestrogen fulvestrant, suggesting an ER-mediated mechanism. Although the extent of stimulation of proliferation over 7 days was lower with these compounds than with 10(-8) M 17 beta-oestradiol, given a longer time period of 35 days the extent of proliferation with 10(-4) M benzyl salicylate, benzyl benzoate or butylphenylmethylpropional increased to the same magnitude as observed with 10(-8) M 17 beta-oestradiol over 14 days. This demonstrates that benzyl salicylate, benzyl benzoate and butylphenylmethylpropional are further chemical components of cosmetic products which give oestrogenic responses in a human breast cancer cell line in culture. Further research is now needed to investigate whether oestrogenic responses are detectable using in vivo models and the extent to which these compounds might be absorbed through human skin and might enter human breast tissues. Copyright (C) 2009 John Wiley & Sons, Ltd.

Evidence for an inhibitory role of bone morphogenetic protein(s) in the follicular-luteal transition in cattle

Bone morphogenetic proteins (BMPs) and their receptors are expressed in ovarian theca cells (TC) and granulosa cells (GC) and BMPs have been implicated in the regulation of several aspects of follicle development including thecal androgen production and granulosal oestrogen production. Their potential involvement in luteal function has received less attention. in this study, we first compared relative abundance of mRNA transcripts for BMPs, activin-beta A and BMP/activin receptors in bovine corpus luteum (CL) and follicular theca and granulosa layers before undertaking functional in vitro experiments to test the effect of selected ligands (BMP6 and activin A) on luteinizing bovine TC and GC. Relative to P-actin transcript abundance, CL tissue contained more BMP4 and -6 mRNA than granulosa, more BMP2 mRNA than theca but much less activin-beta A mRNA than both granulosa and theca. Transcripts for all seven BMP/activin receptors were readily detected in each tissue and two transcripts (BMPRII, ActRIIA) were more abundant in CL than either theca or granulosa, consistent with tissue responsiveness. In vitro luteinization of TC and GC from antral follicles (4-6 mm) was achieved by culturing with 5% serum for 6 days. Treatment with BMP6 (0, 2, 10, and 50 ng/ml) and activin A (0, 2, 10 and 50 ng/ml) under these conditions dose-dependently suppressed forskolin-induced progesterone (P-4) secretion from both cell types without affecting cell number. BMP6 reduced forskolin-stimulated upregulation of STAR mRNA and raised 'basal' CYP17A1 mRNA level in theca-lutein cells without affecting expression of CYP11A1 or hydroxy-Delta-5-steroid dehydrogenase, 3 beta- and steroid Delta-isomerase 1 (HSD3B1). In granulosa-lutein cells, STAR transcript abundance was not affected by BMP6, whereas forskolin-induced expression of CYP11A1, HSD3B1, CYP19A1 and oxytocin transcripts was reduced. In both cell types, follistatin attenuated the suppressive effect of activin A and BMP6 on forskolin-induced P4 secretion but had no effect alone. Granulosa-lutein cells secreted low levels of endogenous activin A (similar to 1 ng/ml); BMP6 reduced this, while raising follistatin secretion thus decreasing activin A:follistatin ratio. Collectively, these findings support inhibitory roles for BMP/activin signalling in luteinization and steroidogenesis in both TC and GC.

Paraben esters: review of recent studies of endocrine toxicity, absorption, esterase and human exposure, and discussion of potential human health risks

This toxicology update reviews research over the past four years since publication in 2004 of the first measurement of intact esters of p-hydroxybenzoic acid (parabens) in human breast cancer tissues, and the suggestion that their presence in the human body might originate from topical application of bodycare cosmetics. The presence of intact paraben esters in human body tissues has now been confirmed by independent measurements in human urine, and the ability of parabens to penetrate human skin intact without breakdown by esterases and to be absorbed systemically has been demonstrated through studies not only in vitro but also in vivo using healthy human subjects. Using a wide variety of assay systems in vitro and in vivo, the oestrogen agonist properties of parabens together with their common metabolite (p-hydroxybenzoic acid) have been extensively documented, and, in addition, the parabens have now also been shown to possess androgen antagonist activity, to act as inhibitors of sulfotransferase enzymes and to possess genotoxic activity. With the continued use of parabens in the majority of bodycare cosmetics, there is a need to carry out detailed evaluation of the potential for parabens, together with other oestrogenic and genotoxic co-formulants of bodycare cosmetics, to increase female breast cancer incidence, to interfere with male reproductive functions and to influence development of malignant melanoma which has also recently been shown to be influenced by oestrogenic stimulation. Copyright (C) 2008 John Wiley & Sons, Ltd.

Oestrogenic and androgenic activity of triclosan in breast cancer cells

As a consequence of its widespread use as an antimicrobial agent in consumer goods, triclosan has become distributed ubiquitously across the ecosystem, and recent reports that it can cause endocrine disruption in aquatic species has increased concern. It is reported here that triclosan possesses intrinsic oestrogenic and androgenic activity in a range of assays in vitro which could provide some explanation for the endocrine disrupting properties described in aquatic populations. In terms of oestrogenic activity, triclosan displaced [H-3]oestradiol from oestrogen receptors (ER) of MCF7 human breast cancer cells and from recombinant human ER alpha/ER beta. Triclosan at 10(-5) M completely inhibited the induction of the oestrogen-responsive ERE-CAT reporter gene in MCF7 cells by 10(-10) M 17 beta-oestradiol and the stimulation of growth of MCF7 human breast cancer cells by 10(-10) M 17 beta-oestradiol. On its own, 1 mu M triclosan increased the growth of MCF7 cells over 21 days. Triclosan also had androgenic activity. It displaced [H-3]testosterone from binding to the ligand binding domain of the rat androgen receptor (AR). Triclosan was able to inhibit the induction of the androgen-responsive LTR-CAT reporter gene in S115 mouse mammary tumour cells by 10(-9) M testosterone and in T47D human breast cancer cells by 10(-8) M testosterone at concentrations of 10(-7) M and 10(-6) M, respectively. Triclosan at 2 x 10(-5) M antagonized the stimulation of the growth of S115+A mouse mammary tumour cells by 10(-9) M testosterone. The finding that triclosan has oestrogenic and androgenic activity warrants further investigation in relation to both endocrine disruption of aquatic wildlife and any possible impact on human health. Copyright (C) 2007 John Wiley & Sons, Ltd.

Comparison of the global gene expression profiles produced by methylparaben, n-butylparaben and 17 beta-oestradiol in MCF7 human breast cancer cells

Since the alkyl esters of p-hydroxybenzoic acid (parabens) can be measured intact in the human breast and possess oestrogenic properties, it has been suggested that they could contribute to an aberrant burden of oestrogen signalling in the human breast and so play a role in the rising incidence of breast cancer. However, although parabens have been shown to regulate a few single genes (reporter genes, pS2, progesterone receptor) in a manner similar to that of 17 beta-oestradiol, the question remains as to the full extent of the similarity in the overall gene profile induced in response to parabens compared with 17 beta-oestradiol. The GE-Amersham CodeLink 20 K human expression microarray system was used to profile the expression of 19881 genes in MCF7 human breast cancer cells following a 7-day exposure to 5 x 10(-4) m methylparaben, 10(-5) m n-butylparaben and 10(-8) m 17 beta-oestradiol. At these concentrations, the parabens gave growth responses in MCF7 cells of similar magnitude to 17 beta-oestradiol. The study identified genes which are upregulated or downregulated to a similar extent by methylparaben, n-butylparaben and 17 beta-oestradiol. However, the majority of genes were not regulated in the same way by all three treatments. Some genes responded differently to parabens from 17 beta-oestradiol, and furthermore, differences in expression of some genes could be detected even between the two individual parabens. Therefore, although parabens possess oestrogenic properties, their mimicry in terms of global gene expression patterns is not perfect and differences in gene expression profiles could result in consequences to the cells that are not identical to those following exposure to 17 beta-oestradiol. Copyright (c) 2006 John Wiley & Sons, Ltd.

Metalloestrogens: an emerging class of inorganic xenoestrogens with potential to add to the oestrogenic burden of the human breast

Many compounds in the environment have been shown capable of binding to cellular oestrogen receptors and then mimicking the actions of physiological oestrogens. The widespread origin and diversity in chemical structure of these environmental oestrogens is extensive but to date such compounds have been organic and in particular phenolic or carbon ring structures of varying structural complexity. Recent reports of the ability of certain metal ions to also bind to oestrogen receptors and to give rise to oestrogen agonist responses in vitro and in vivo has resulted in the realisation that environmental oestrogens can also be inorganic and such xenoestrogens have been termed metalloestrogens. This report highlights studies which show metalloestrogens to include aluminium, antimony, arsenite, barium, cadmium, chromium (Cr(II)), cobalt, copper, lead, mercury, nickel, selenite, tin and vanadate. The potential for these metal ions to add to the burden of aberrant oestrogen signalling within the human breast is discussed. Copyright (c) 2006 John Wiley & Sons, Ltd.

Environmental oestrogens, cosmetics and breast cancer

The established role of oestrogen in the development and progression of breast cancer raises questions concerning a potential contribution from the many chemicals in the environment which can enter the human breast and which have oestrogenic activity. A range of organochlorine pesticides and polychlorinated bipheryls possess oestrogen-mimicking properties and have been measured in human breast adipose tissue and in human milk. These enter the breast from varied environmental contamination of food, water and air, and due to their lipophilic properties can accumulate in breast fat. However, it is emerging that the breast is also exposed to a range of oestrogenic chemicals applied as cosmetics to the underarm and breast area. These cosmetics are left on the skin in the appropriate area, allowing a more direct dermal absorption route for breast exposure to oestrogenic chemicals and allowing absorbed chemicals to escape systemic metabolism. This review considers evidence in support of a functional role for the combined interactions of cosmetic chemicals with environmental oestrogens, pharmacological oestrogens, phyto-oestrogens and physiological oestrogens in the rising incidence of breast cancer.

Aluminium, antiperspirants and breast cancer

Aluminium salts are used as the active antiperspirant agent in underarm cosmetics, but the effects of widespread, long term and increasing use remain unknown, especially in relation to the breast, which is a local area of application. Clinical studies showing a disproportionately high incidence of breast cancer in the upper outer quadrant of the breast together with reports of genomic instability in outer quadrants of the breast provide supporting evidence for a role for locally applied cosmetic chemicals in the development of breast cancer. Aluminium is known to have a genotoxic profile, capable of causing both DNA alterations and epigenetic effects, and this would be consistent with a potential role in breast cancer if such effects occurred in breast cells. Oestrogen is a well established influence in breast cancer and its action, dependent on intracellular receptors which function as ligand-activated zinc finger transcription factors, suggests one possible point of interference from aluminium. Results reported here demonstrate that aluminium in the form of aluminium chloride or aluminium chlorhydrate can interfere with the function of oestrogen receptors of MCF7 human breast cancer cells both in terms of ligand binding and in terms of oestrogen-regulated reporter gene expression. This adds aluminium to the increasing list of metals capable of interfering with oestrogen action and termed metal I oestrogens. Further studies are now needed to identify the molecular basis of this action, the longer term effects of aluminium exposure and whether aluminium can cause aberrations to other signalling pathways in breast cells. Given the wide exposure of the human population to antiperspirants, it will be important to establish dermal absorption in the local area of the breast and whether long term low level absorption could play a role in the increasing incidence of breast cancer. (c) 2005 Elsevier Inc. All rights reserved.

Comparative study of oestrogenic properties of eight phytoestrogens in MCF7 human breast cancer cells

Previous studies have compared the oestrogenic properties of phytoestrogens in a wide variety of disparate assays. Since not all phytoestrogens have been tested in each assay, this makes inter-study comparisons and ranking oestrogenic potency difficult. In this report, we have compared the oestrogen agonist and antagonist activity of eight phytoestrogens (genistein, daidzein, equol, miroestrol, deoxymiroestrol, 8-prenylnaringenin, coumestrol and resveratrol) in a range of assays all based within the same receptor and cellular context of the MCF7 human breast cancer cell line. The relative binding of each phytoestrogen to oestrogen receptor (ER) of MCF7 cytosol was calculated from the molar excess needed for 50 % inhibition of [H-3]oestradiol binding (IC50), and was in the order coumestrol (35x)/8-prenylnaringenin (45 x)/deoxymiroestrol (50 x) > miroestrol (260x) > genistein (1000x) > equol (4000x) > daidzein (not achieved: 40 % inhibition at 10(4)-fold molar excess) > resveratrol (not achieved: 10 % inhibition at 10(5)-fold molar excess). For cell-based assays, the rank order of potency (estimated in terms of the concentration needed to achieve a response equivalent to 50 % of that found with 17 beta-oestradiol (IC50)) remained very similar for all the assays whether measuring ligand ability to induce a stably transfected oestrogen-responsive ERE-CAT reporter gene, cell growth in terms of proliferation rate after 7 days or cell growth in terms of saturation density after 14 days. The IC50 values for these three assays in order were for 17 beta-oestradiol (1 x 10-(11) M, 1 x 10-(11) M, 2 x 10(-11) M), and in rank order of potency for the phytoestrogens, deoxymiroestrol (1 x 10(-10) M, 3 x 10(-11) M, 2 x 10(-11) M) > miroestrol (3 x 10(-10) M, 2 x 10(-11) M, 8 x 10(-11) M) > 8-prenylnaringenin (1 x 10(-9) M, 3 x 10(-10) M, 3 x 10(-10) M) > cournestrol (3 x 10(-8) M, 2 x 10(-8) M, 3 x 10(-8) M) > genistein (4 x 10(-8) M, 2 x 10(-8) M, 1 x 10(-8) M)/equol (1 x 10(-7) M, 3 x 10(-8) M, 2 x 10(-8) M) > daidzein (3 x 10(-7) M, 2 x 10(-7) M, 4 x 10(-8) M) > resveratrol (4 x 10(-6) M, not achieved, not achieved). Despite using the same receptor context of the MCF7 cells, this rank order differed from that determined from receptor binding. The most marked difference was for cournestrol and 8-prenylnaringenin which both displayed a relatively potent ability to displace [3H]oestradiol from cytosolic ER compared with their much lower activity in the cell-based assays. Albeit at varying concentrations, seven of the eight phytoestrogens (all except resveratrol) gave similar maximal responses to that given by 17 beta-oestradiol in cell-based assays which makes them full oestrogen agonists. We found no evidence for any oestrogen antagonist action of any of these phytoestrogens at concentrations of up to 10(-6) M on either reporter gene induction or on stimulation of cell growth. (c) 2005 Elsevier Ltd. All rights reserved.

Oestrogenic activity of p-hydroxybenzoic acid (common metabolite of paraben esters) and methylparaben in human breast cancer cell lines

This paper addresses the question of whether p-hydroxybenzoic acid, the common metabolite of parabens, possesses oestrogenic activity in human breast cancer cell lines. The alkyl esters of p-hydroxybenzoic acid (parabens) are used widely as preservatives in consumer products to which the human population is exposed and have been shown previously to possess oestrogenic activity and to be present in human breast tumour tissue, which is an oestrogen-responsive tissue. Recent work has shown p-hydroxybenzoic acid to give an oestrogenic response in the rodent uterotrophic assay. We report here that p-hydroxybenzoic acid possesses oestrogenic activity in a panel of assays in human breast cancer cell lines. p-Hydroxybenzoic acid was able to displace [H-3]oestradiol from cytosolic oestrogen receptor of MCF7 human breast cancer cells by 54% at 5 x 10(6)-fold molar excess and by 99% at 10(7)-fold molar excess. It was able to increase the expression of a stably integrated oestrogen responsive reporter gene (ERE-CAT) at a concentration of 5 x 10(-4) M in MCF7 cells after 24 h and 7 days, which could be inhibited by the anti-oestrogen ICI 182 780 (Faslodex, fulvestrant). Proliferation of two human breast cancer cell lines (MCF7, ZR-75-1) could be increased by 10(-5) M p-hydroxybenzoic acid. Following on from previous studies showing a decrease in oestrogenic activity of parabens with shortening of the linear alkyl chain length, this study has compared the oestrogenic activity of p-hydroxybenzoic acid where the alkyl grouping is no longer present with methylparaben, which has the shortest alkyl group. Intrinsic oestrogenic activity of p-hydroxybenzoic acid was similar to that of methylparaben in terms of relative binding to the oestrogen receptor but its oestrogenic activity on gene expression and cell proliferation was lower than that of methylparaben. It can be concluded that removal of the ester group from parabens does not abrogate its oestrogenic activity and that p-hydroxybenzoic acid can give oestrogenic responses in human breast cancer cells. Copyright (C) 2005 John Wiley & Sons, Ltd.

Endocrine disrupters and human health: Could oestrogenic chemicals in body care cosmetics adversely affect breast cancer incidence in women? A review of evidence and call for further research

In the decade that has elapsed since the suggestion that exposure of the foetal/developing male to environmental oestrogens could be the cause of subsequent reproductive and developmental effects in men, there has been little definitive research to provide conclusions to the hypothesis. Issues of exposure and low potency of environmental oestrogens may have reduced concerns. However, the hypothesis that chemicals applied in body care cosmetics (including moisturizers, creams, sprays or lotions applied to axilla or chest or breast areas) may be affecting breast cancer incidence in women presents a different case scenario, not least in the consideration of the exposure issues. The specific cosmetic type is not relevant but the chemical ingredients in the formulations and the application to the skin is important. The most common group of body care cosmetic formulation excipients, namely p-hydroxybenzoic acid esters or parabens, have been shown recently to be oestrogenic in vitro and in vivo and now have been detected in human breast tumour tissue, indicating absorption (route and causal associations have yet to be confirmed). The hypothesis for a link between oestrogenic ingredients in underarm and body care cosmetics and breast cancer is forwarded and reviewed here in terms of. data on exposure to body care cosmetics and parabens, including dermal absorption; paraben oestrogenicity; the role of oestrogen in breast cancer; detection of parabens in breast tumours; recent epidemiology studies of underarm cosmetics use and breast cancer; the toxicology database; the current regulatory status of parabens and regulatory toxicology data uncertainties. Notwithstanding the major public health issue of the causes of the rising incidence of breast cancer in women, this call for further research may provide the first evidence that environmental factors may be adversely affecting human health by endocrine disruption, because exposure to oestrogenic chemicals through application of body care products (unlike diffuse environmental chemical exposures) should be amenable to evaluation, quantification and control. The exposure issues are clear and the exposed population is large, and these factors should provide the necessary impetus to investigate this potential issue of public health. Copyright (C) 2004 John Wiley Sons, Ltd.

Underarm cosmetics and breast cancer

Although risk factors are known to include the loss of function of the susceptibility genes BRCA1/BRCA2 and lifetime exposure to oestrogen, the main causative agents in breast cancer remain unaccounted for. It has been suggested recently that underarm cosmetics might be a cause of breast cancer, because these cosmetics contain a variety of chemicals that are applied frequently to an area directly adjacent to the breast. The strongest supporting evidence comes from unexplained clinical observations showing a disproportionately high incidence of breast cancer in the upper outer quadrant of the breast, just the local area to which these cosmetics are applied. A biological basis for breast carcinogenesis could result from the ability of the various constituent chemicals to bind to DNA and to promote growth of the damaged cells. Multidisciplinary research is now needed to study the effect of long-term use of the constituent chemicals of underarm cosmetics, because if there proves to be any link between these cosmetics and breast cancer then there might be options for the prevention of breast cancer. Copyright D 2003 John Wiley Sons, Ltd.