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Estrogenic and DNA-Damaging Activity of Red No. 3 in Human Breast Cancer Cells
Abstract
Exposure to pesticides, dyes, and pollutants that mimic the growth promoting effects of estrogen may cause breast cancer. The pesticide DDT and the food colorant Red No. 3 were found to increase the growth of HTB 133 but not estrogen receptor (ER) negative human breast cells (HTB 125) or rat liver epithelial cells (RLE). Red No. 3, beta-estradiol, and DDT increase ER site-specific DNA binding to the estrogen response element in HTB 133 cells and increase cyclin-dependent kinase 2 activity in MCF-7 breast cancer cells. Site-specific DNA binding by p53 in RLE, HTB 125, HTB 133, and MCF-7 cells was increased when they were treated with Red No. 3, which suggests that cellular DNA was damaged by this colorant. Red No. 3 increased binding of the ER from MCF-7 cells to the estrogen-responsive element. Consumption of Red No. 3, which has estrogenlike growth stimulatory properties and may be genotoxic, could be a significant risk factor in human breast carcinogenesis.
... There has been an increasing concern among the general and medical community in relation to the potential environmental hazard that estrogens present to health in general, although the mechanisms by which they affect homeostasis are still barely known123. It has been known for several years that exposure to some xenohormones increases the production of reactive oxygen species, which in turn could inflict structural damage to cell DNA of target organs, as well as to DNA from other systemic cells [4,5]. The genotoxic action of contaminating agents affects human health directly, damaging the genetic material, which is considered to play an important role in oncogenesis [6,7]. ...
... Xenohormones can have a bi-functional behavior, depending on their mechanism of action, either through a genetic or hormonal route; this depends on their structure, concentration and exposure period [4]. On the other hand, it has been reported that exposure to some xenoestrogens and to estrogenic metabolites promotes the production of free radicals through the hydrogen peroxide (H 2 O 2 ) reduction cycle, by means of the P 450 oxidase and reductase enzymes, giving origin to the production of oxygen reactive species which might damage the genetic material [33]. ...
The genotoxicity of some environmental contaminants may affect human health directly by damaging genetic material and thus plays an important role in cancer development. Xenoestrogens are one kind of environmental pollutants that may alter hormonal routes or directly affect DNA. The number of available biomarkers used to assess genetic risk and cancer is very extensive. The present study evaluated genotoxicity produced by the pesticide DDT on systemic and mammary gland cells obtained from adult female Wistar rats. Oral mucosa cells micronuclei were assessed; the comet assay in peripheral blood-isolated lymphocytes and mammary epithelial cells was also carried out. Additionally, oxidative stress was studied in mammary tissue through a lipid peroxidation assay. Our data showed an increase in lipid peroxidation, product of an increase in free oxygen radical levels, which leads to an oxidative stress status. Our results suggest that DDT is genotoxic, not only for lymphocytes but also to mammary epithelial cells.
... In contrast, synthetic dyes tend to be chemically more stable and maintain their color across a broad range of food conservation conditions, tolerating greater variations in, for example, temperature, acidity, oxygen concentration, and light intensity. Despite their generally higher chemical stability, in the last decades, a number of potential health risks have been associated with the consumption of synthetic food dyes, including behavioral and neurological disorders (McCann et al. 2007), higher incidence of allergies (Koutsogeorgopoulou et al. 1998), and potential carcinogenesis (Dees et al. 1997). Such health-related concerns have raised the interest in finding alternative sources of natural colorants for the food industry. ...
Carrot is an economically important vegetable crop worldwide. Its storage root, the consumed organ, varies broadly within the carrot germplasm, exhibiting different colors due to the accumulation of anthocyanin and carotenoid pigments, as well as extensive variation for phytochemicals composition and consumer-quality traits. Anthocyanins and other phenolics, carotenoids, polyacetylenes, and terpenes represent the major carrot nutraceutical classes. In recent years, the use of next-generation sequencing technologies has facilitated the application of “multi-omics” approaches, in combination with transgenics and classical genetic tools, for studying the genetics underlying the accumulation of these phytochemicals in the carrot root. In purple carrot, such approaches allowed the identification and mapping of simply inherited and quantitative trait loci (QTLs) conditioning anthocyanin pigmentation in different tissues and genetic backgrounds, and the discovery of key genes conditioning anthocyanin biosynthesis, glycosylation, and acylation. Glycosylation and acylation influence the chemical stability and bioavailability of anthocyanins, and therefore their potential use as food colorants or nutraceutical agents, respectively. Similarly, important advances were made for two major loci conditioning carotenoids accumulation in white, yellow, and orange roots, namely Y and Y2. With the sequencing of the carrot genome, a candidate for the Y gene involved in photosystem development and carotenoid storage was described, whereas fine mapping of Y2 drastically reduced the genomic region of interest to 650-kb, but a clear candidate was not identified. Another gene, Or, which regulates chromoplasts development, was associated with carotenoids presence in the carrot root. Besides these nonstructural genes, progress towards understanding the role of several carotenoid biosynthetic genes has been made. The genetics of carrot polyacetylenes is also becoming increasingly understood. Candidate fatty acid desaturase 2 (FAD2) genes with specific desaturase and/or acetylenase activities have been identified by QTLs analysis and proposed as catalyzers of different steps in the polyacetylene pathway, and their genomic organization was described. Similarly, gene members of the large carrot terpene synthase family were catalogued, partially associated with QTLs for characteristic carrot root monoterpenes like sabinene, and functionally characterized in vitro. This chapter reviews and discusses recent advances in genetics and genomics of the main carrot nutraceuticals.
... Some herbicides and pesticides can even increase estrogen biosynthesis (Cooke et al., 2013). The effects of overpromotion of estrogen signaling are related to breast cancer (Dees et al., 1997), as well as various morphological changes in the mammary glands (Mandrup et al., 2016;Soto et al., 2013;Colerangle and Roy, 1997). Exposure to BPA also impairs milk yield, lipid fraction, and protein synthesis (Kass et al., 2012), (Altamirano et al., 2015). ...
During pregnancy, the maternal body undergoes a considerable transformation regarding the anatomy, metabolism, and immune profile that, after delivery, allows for protection and nourishment of the offspring via lactation. Pregnancy hormones are responsible for the development and functionality of the mammary gland for breast milk production, but little is known about how hormones control its immune properties. Breast milk composition is highly dynamic, adapting to the nutritional and immunological needs that the infant requires in the first months of life and is responsible for the main immune modeling of breastfed newborns. Therefore, alterations in the mechanisms that control the endocrinology of mammary gland adaptation for lactation could disturb the properties of breast milk that prepare the neonatal immune system to respond to the first immunologic challenges. In modern life, humans are chronically exposed to endocrine disruptors (EDs), which alter the endocrine physiology of mammals, affecting the composition of breast milk and hence the neonatal immune response. In this review, we provide a landscape of the possible role of hormones in the control of passive immunity transferred by breast milk and the possible effect of maternal exposure to EDs on lactation, as well as their impacts on the development of neonatal immunity.
... Natural and 2. Artificial. In the present world, natural pigments acceptability is rising immensely because of health and environmental awareness (Bridle and Timberlake', 1997;Dees et al., 1997;Downham and Collins, 2000). Purple corn husk is an excellent source of natural pigment which called anthocyanin pigment mostly used in food factory and human being health issues (Tsuda et al., 2003). ...
Corn husk is generated as a by-product during the processing and collection of corn, the second largest agricultural crop of the whole planet. The use of these by-products is currently being studied due to their rich availability, less expense, and biodegradability. In association with extraction of nanocellulose, reinforcement in composites and utilization as adsorbents, corn husk has been explored in technical applications. There are a variety of advantages of corn husk reinforced composites, including enhanced mechanical properties, heat resistance, sound absorption, and flame resistance. In this review, we present an overview of the by-products of corn plants and discuss how chemical composition of corn husk is affected by the source and chemical treatment. The utilization of corn husk in a wide variety of fields, like nanocomposites, bioplastics, wood plastic composites, flame retardant and polymer composites, paper production, as potential source of anthocyanins, adsorbent, acoustic panel, electrochemical applications, infrastructure, and medicine are discussed comprehensively. In addition to evaluating the limitations and prospects of corn husk, the directions for future research have also been explored in detail.
... Red 3 causes DNA damage (cancer) in animals, and there is other evidence showing that several other dyes are also carcinogenic. The consumption of Red No. 3 was reported with estrogen-like growth stimulatory properties and resulted in being genotoxic, and it had significant risk factors in human breast carcinogenesis [20]. Three dyes, Red 40, Yellow 5, and Yellow 6, have also been reported to be adulterate with other carcinogens or benzidine [21]. ...
Synthetic food colors are important ingredients in the food industry. These synthetic food colorants are azo dyes, majorly acidic in nature such as Allura red and Tartrazine. They are present in sweets, carbonated drinks, meat products, and candies to attract the consumers. This review article is an attempt to explain the adverse effects of azo dyes and their association with oral cavities and cardiovascular disorders. These synthetic dyes (azo dyes) have staining effects on dentin. Poor dental care accelerates the bacterial accumulation on the dental crown (Gram-negative bacteria P. gingivalis, T. denticola, and T. forsythia and Gram-positive bacteria Strep. Gordonii), causing the washing of enamel, forming dental plaque. Bacterial pathogens (P. ginigivalis and F. nacleatum) release different chemicals (FadA and Fap2) that bind to protein on the cell by producing an inflammatory response through different line-host defenses, such as Gingival epithelial cells (ECs), Hemi-desmosomes, and desmosomes, which helps the bacterium migration from the cell–cell junction. This makes the junctions slightly open up and makes the whole vessel permeable, through which the bacterium enters into the blood stream line. This leads to different major arteries, such as the carotid artery, and causes the accumulation of plaque in major cardiac arteries, which causes different cardiovascular disorders. These bacterial species present in gums cause cardiovascular diseases, such as ischemic heart disease, coronary artery disease, heart attacks and strokes, and arrhythmias, which can lead to death.
... To elucidate how estrogenic compounds can compete with DDT by their estrogenic potential (108,109) some studies have yielded several molecular mechanisms explaining this interaction. For instance, it has been shown that DTT increased the growth of HTB 133 cells by enhancing the ER-DNA specific binding to the specific ERE (estrogen response elements) and also potentiates the activity of cyclin-dependent kinase 2 in MCF-7 breast cancer cells but not ER-negative HTB 125 breast cells or rat liver epithelial cells (110). Besides, there is evidence in vivo and in vitro that shows the influence of DTT in the interaction of ER and specific genes promoting cell proliferation (11,109), in fact, DTT can also bind to ERs in different places of the body other than female reproductive sites. ...
Evaluation of carcinogenic substances from the environment is a challenge for scientists. Recently, a novel approach based on 10 key characteristics of human carcinogens classified by the International Agency for Research on Cancer (IARC) has emerged. Carcinogenesis depends on different mechanisms and factors, including genetic, infectious (bacteria, viruses) and environmental (chemicals) factors. Endocrine disruptors are exogenous chemicals that can interfere and impair the function of the endocrine system due to their interaction with estrogen receptors or their estrogen signaling pathways inducing adverse effects in the normal mammary development, originating cancer. They are heterogeneous chemicals and include numerous synthetic substances used worldwide in agriculture, industry and consumer products. The most common are plasticizers, such as bisphenol A (BPA), pesticides, such as dichlorodiphenyltrichloroethane, and polychlorinated biphenyls (PCBs). Xenoestrogens appear to serve an important role in the increased incidence of breast cancer in the United States and numerous other countries. Several studies have demonstrated the role of organochlorine xenoestrogens in breast cancer. Therefore, the overall cumulative exposure of women to estrogens results in an increased risk for this type of cancer. Factors like lifestyle and diet also serve a role in the increased incidence of this disease. The aim of the present study was to analyze these chemical compounds based on the key characteristics given by the IARC, with a special focus on breast cancer, to establish whether these compounds are carcinogens, and to create a model for future analysis of other endocrine disruptors.
... Furthermore, it is associated with irritability, restlessness, and sleep disturbance in hyperactive children aged between two and fourteen years [18]. Moreover, Sunset yellow and Ponceau 4R have also been implicated to have adverse reactions in patients with chronic urticaria [34], and erythrosine has estrogen-like growth stimulatory properties which may be genotoxic and could be a significant risk factor in human breast cancer [35]. erefore, it is essential to make internationally accepted strategies to implement food safety and quality to control the usage of Tartrazine, sunset yellow, erythrosine, and Ponceau 4R. ...
Colour is a key component to enhance the ultimate appetizing value and consumer acceptance towards foods and beverages. Synthetic food colours have been increasingly used than natural food colours by food manufacturers to attain certain properties such as low cost, improved appearance, high colour intensity, more colour stability, and uniformity. Varied foods and beverages available in the market may contain some nonpermitted synthetic colours and overuse of permitted synthetic colours. This may lead to severe health problems such as mutations, cancers, reduced haemoglobin concentrations, and allergic reactions. According to the Food Act, 2011 (No. 26 of 1980), Sri Lanka, only nine synthetic food colours are permitted and the colour concentration cannot exceed 100 ppm as a single component or in combination. This study aims to identify the synthetic food colours in confectioneries and beverages available in Jaffna district, Sri Lanka. Randomly collected 110 samples from eleven Medical Officers Of Health areas in Jaffna district were analyzed by using thin layer chromatography and UV-visible spectrophotometry. According to the results, 100% beverages and 85% confectioneries contained permitted synthetic food colours. Out of all, 7% of the confectioneries did not contain any synthetic food colour and 8% of the confectioneries contained nonpermitted colours which do not comply with any of the permitted synthetic food colours. Tartrazine (E102) (41%) was the most used synthetic food colour in both confectioneries and beverages. Moreover, 60% of the beverages violated the label requirement without including proper colour ingredients. The study concluded that there is a high tendency to use synthetic food colours in confectioneries and beverages and some confectioneries contain unidentified colours including a textile dye. Therefore, the implementation of regulations and awareness programs of food colours for consumers and food manufacturers are highly recommended.
... Cytotoxicity encompassing alterations in the dividing cell frequency and chromosome architecture is induced by the heavy metals (Kiran and Şahin 2006, Ünyayar et al. 2006, Pandey and Upadhyay 2010, Bhat et al. 2011, Kwankua et al. 2012, Tommonaro et al. 2015, Kumbhakar et al. 2018, Pramanik et al. 2019) and azo-dyes (Dwivedi andKumar 2015, Kumbhakar et al. 2018) in plant species providing insight to subcellular mode of interaction (cellular damages caused). Accumulated toxicants in plant species can be transmitted to consumers, especially human beings, through the edible product(s) causing serious health hazards (Chandra and Nagaraja 1987, Dees et al. 1997, Miller et al. 2002, Soucy et al. 2003, Banerjee and Giri 2014. ...
Allium cepa (2n=16) assay is used to determine cytotoxicity of environmental pollutants like heavy metal arsenic (in the form of arsenic trioxide—concentration used: 0.010, 0.050, 0.075 and 0.100 mg L⁻¹ for 24 h duration) and azo-dye metanil yellow (concentration used: 100, 150, 200 and 400 mg L⁻¹ for 24 h duration) with an objective to understand the toxic effects of the test materials on cells and chromosomes of a plant-based system. Assessment of cytotoxicity reveals that arsenic trioxide can induce chromosomal breakages, affects spindle organization and causes cellular metabolic defects; whereas, metanil yellow predominantly affects cellular metabolism. Cytological disturbances are mostly dose-dependent, and arsenic trioxide depicts pronounced effectivity in inducing mitotic aberrations in root tip cells of A. cepa than metanil yellow (in relation to employed doses). Furthermore, aqueous plant extracts (used due to its operational simplicity and cost-effectivity) of the leaf (Coriandrum sativum L., Ocimum tenuiflorum L., and Pteris vittata L.) and seed (Nigella sativa L.) samples are used to ascertain their amelioration potentiality against the environmental toxicants. The ameliorative study (decrement in their observed values) involves attributes like mitotic index, total abnormal dividing cell frequency and frequency of giant and anucleate cells in resting stages. Results suggest that all the employed extracts are ameliorative, and can be explored further for their role in bioremediation.
... The latter (synthetic food dyes) are chemically stable under a broad range of temperature, pH, light intensity, and oxygen concentration, making them suitable for a wide range of food conservation conditions. However, there is increasing public concern regarding potential health problems associated with the consumption of synthetic dyes, including allergic reactions (Koutsogeorgopoulou et al. 1998), behavioral and neurological adverse effects (McCann et al. 2007), and potential carcinogenesis (Dees et al. 1997). Thus, because of these health issues, alternative colorants from natural sources are desired. ...
Purple carrots (Daucus carota ssp. sativus var. atrorubens Alef.) accumulate anthocyanins in their roots, petioles, and other plant parts. These flavonoid pigments represent an excellent dietary source of antioxidant and anti-inflammatory agents. In addition, carrot anthocyanins are also used as food dyes. Compositional variation in carrot root, mainly with regard to the content of acylated (AA) and non-acylated anthocyanins (NAA), strongly influences the bioavailability and chemical stability of these pigments, therefore conditioning their potential use as nutraceutical agents or as food colorants. In this context, genetic diversity analysis for root anthocyanin composition is relevant for selecting materials for either purpose. Also, knowledge on the genetic basis underlying anthocyanin biosynthesis and modification is expected to aid in the development of new varieties with high nutraceutical or for extracting food dyes. In the last decades, germplasm collections have been characterized for anthocyanin content and composition. Various simply inherited traits for root and petiole anthocyanin pigmentation and acylation, including P1, P3 and Raa1, and QTL for root anthocyanins, have been described and mapped to two regions of chromosome 3, in different genetic backgrounds. Recent advances in high-throughput sequencing and bioinformatic analyses have facilitated the discovery of candidate regulatory genes for root and petiole pigmentation associated with the P3 region in chromosome 3, as well as structural genes involved in anthocyanin glycosylation and acylation. In this chapter, we reviewed recent advances in diversity, genetic, and genomic studies related to carrot anthocyanin pigmentation.
... [6] Studies have reported the mixing of Curcuma zedoaria, a wild relative of turmeric, into turmeric powder due to its close resemblance with turmeric. [6,7] Similarly, metanil yellow (C18H14N3NaO3S) is a toxic azo dye that has been added to turmeric powder to mimic the appearance of curcumin [8] when the actual curcumin content is low. [9] Toxicologically, metanil yellow is classified as a CII category substance by the Joint FAO/WHO Expert committee on Food Additives, and it implies that it is a compound for on rats show that long term consumption of metanil yellow causes neurotoxicity [11] , hepatocellular carcinoma [12] , tumor development [8] , deleterious effect on gastric mucin [13] , and lymphocytic leukemia. ...
... Tsuda et al. (2001) observed colon DNA damage associated with intake of three azo dyes, amaranth (Red 2), allura red (Red 40) and acid red. Dees et al. (1997) reported that Red No. 3, which has estrogen like growth stimulatory properties, could be a significant risk factor in human breast carcinogenesis. ...
... But few studies have focused on the estrogenic activity of food dyes. Studies concern mainly food colors such as tartazine (E 102), sudan I and erythrosine B (E 127) which have effects on the chromosomal structure and can increase Estrogen Receptor (ER) site-specific DNA binding effect to Estrogen Response Element (ERE) in HTB 133 cells [12,13] and in the E-screen test [14]. Axon et al. [15] helped to identify several modulator of the human ER among food additives such as tartazine and sunset yellow. ...
The toxicity (mutagenic and carcinogenic effects) of food dyes (used like food additive) has been well documented. However the endocrine disrupting effects of these dyes have been poorly investigated. We studied five commercial
food dyes for their agonistic estrogen activity and do a comparison with estrogenic activity of some paraben (other food additives compounds). Total estrogenic activities were measured using the Yeast Estrogen Screen bioassay (YES). The estrogenic potency of the food dyes was measured by dose-response curves and compared to a doseresponse curve of 17-β-estradiol (E2), (reference compound). Estrogen EC50 values have been calculated using standard linear regression methods. Among these compounds, three food dyes, indigotine (E 132), the monoazo dye Ponceau 4R (E 124) and the inorganic dye titanium dioxide (E 171), revealed weak estrogenic activity. Negative results were obtained for Quinoline Yellow (E 104).
... • Kamel 2011 • Gao 2011 • Shimada 2010 • Amin 2010 • Hashem 2010 • Moutinho 2007 • Lau 2006 • Murphy 2006 • Ashida 2000 • Aboel • Aoshima 1997 • Dees 1997 • Reyes 1996 • Voorhees 1983 • Tanaka 1993Tanaka , (1980 and Lafferman and Silbergeld (1979) are all but ignored by the reviewers. ...
This book explains what the Feingold Diet does, who it can help, and the research supporting it.
... Instead of natural pigment, the artificial synthetic pigments are still of great importance to the food industry. But the toxicological status of all these colours has been already studied456. Therefore, there has been a continuing trend over the last few years to replace artificially [2]. ...
The semicontinuous production of red pigment by immobilized cells ofBacillus sp. BH-99 was investigated in comparison with free cells. The red pigment produced highest productivity under the conditions
of aeration of 0.2 mL/min and 2 mm diameter of gel beads by using 3.0% sodium alginate. Semicontinuous production by immobilized
cells showed the highest productivity with replacement of fresh production medium in every 72 h for fourth fermentation cycle
following the conditions of red pigment productivity.
... El " geinstein " , de igual forma, tiene influencia en la proliferación de células aberrantes. A niveles farmacológicos, estos compuestos que operan mediante mecanismos independie ntes al receptor a estrógenos, pueden inhibir cinasas y ADN topoisomerasas y así, afectar otros blancos bioquímicos intracelulares [28, 29]. Cerca de 15% de mujeres portadoras del gene BRCA1 mutado aparentemente no desarrollan cáncer de mama; en esos casos, las xenohormonas benéficas y otros factores exógenos pueden tener un papel positivo, mediante la promoción de detoxificación enzimática de carcinógenos potenciales, procesos de reparación de ADN, formación de antioxidantes o bien, aumentando la capacidad de las células, de rechazar señales que puedan promover un crecimiento descontrolado [30, 31]. ...
There are xenobiotic compounds, which are able to increase estrogen synthesis by matching active sites with estogen receptors. These kinds of compounds perform a similar action to endogenic estrogens, also known as xenoestrogens, which can be classified in general as xenohormones. They are constituted by chemical, synthetical or natural compounds from plants, which can interfere with the endocrinal system´s functions. These kinds of compounds vary in their ways of behavior. One is the simply union between receptor and estrogen. Two by biochemical signal paths, and three is through complex independent mechanism that joins receptor and estrogen. There are studies showing that the endocrinal system in certain fishes and wild species have been altered by chemical substances and pollutants of their microenvironment. These substances are related to the reproduction and endocrinal system´s disturbances as well as incidences and developments of different malignant tumors. Another kind of xenohormones can be beneficial, such as those that we find in the natural forms of plants and fishes. The global consensus among experts in this topic is that new researches must be launched. This should be done with rigorous precision about toxicity and carcinogenicity related to hormonal activity for all current and new chemical products before marketing them
Science remains an eminently social institution, though the interactions between science and society are often poorly understood. This article addresses the context of recent efforts to identify environmental contributions to cancer. The authors first examine cancer incidence and discuss how incidence patterns may be related to air pollution and occupational and general toxic chemical and xenestrogen exposures. They then discuss the social context in which cancer research and treatment occurs, including the dominating role of the biomedical model and socioeconomic factors, including regulatory strategies that address single chemicals, corporate conflicts of interest, and the manipulation of public opinion. Last, they consider the broad context out of which cancer arises and discuss the merits of applying the precautionary principle to sustainable social policies. Progress in reducing cancer may be fruitfully made by returning our attention to broad-scale factors such as those affecting the quality of air, water, workplace, household environments, and the global climate.
Endocrine-disrupting chemicals (EDCs) are exogenous chemicals that interfere with hormone action, thereby increasing the risk of adverse health outcomes, including cancer, reproductive impairment, cognitive deficits and obesity. A complex literature of mechanistic studies provides evidence on the hazards of EDC exposure, yet there is no widely accepted systematic method to integrate these data to help identify EDC hazards. Inspired by work to improve hazard identification of carcinogens using key characteristics (KCs), we have developed ten KCs of EDCs based on our knowledge of hormone actions and EDC effects. In this Expert Consensus Statement, we describe the logic by which these KCs are identified and the assays that could be used to assess several of these KCs. We reflect on how these ten KCs can be used to identify, organize and utilize mechanistic data when evaluating chemicals as EDCs, and we use diethylstilbestrol, bisphenol A and perchlorate as examples to illustrate this approach. In this Expert Consensus Statement, the authors define 10 key characteristics (KCs) for endocrine-disrupting chemicals. They further describe the logic by which these KCs are identified and the assays that could be used to assess several of these KCs.
Erythrosine B (ErB) is a cherry pink food colorant and is widely used in foods, drugs, and cosmetics. Quinoline yellow (QY) is a chinophthalon derivative used in cosmetic compositions for application to the skin, lips, and/or body surface. Previously, ErB and QY synthetic dyes were found to induce DNA damage in HepG2 cells. The aim of this study was to investigate the molecular basis underlying the genotoxicity attributed to ErB and QY using the RT² Profiler polymerase chain reaction array and by analyzing the expression profile of 84 genes involved in cell cycle arrest, apoptosis, and DNA repair in HepG2 cells. ErB (70 mg/L) significantly decreased the expression of two genes (FEN1 and REV1) related to DNA base repair. One gene (LIG1) was downregulated and 20 genes related to ATR/ATM signaling (ATR, RBBP8, RAD1, CHEK1, CHEK2, TOPB1), nucleotide excision repair (ERCC1, XPA), base excision repair (FEN1, MBD4), mismatch repair (MLH1, MSH3, TP73), double strand break repair (BLM), other DNA repair genes (BRIP1, FANCA, GADD45A, REV1), and apoptosis (BAX, PPP1R15A) were significantly increased after treatment with QY (20 mg/L). In conclusion, our data suggest that the genotoxic mechanism of ErB and QY dyes involves the modulation of genes related to the DNA repair system and cell cycle.
When I first read these words, they prompted me to think about writing a book that would address the gap between knowledge about cancer trends and knowledge about environmental carcinogens. Five years later, I published Living Downstream: A Scientist’s Personal Investigation of Cancer and the Environment, which represents my best attempt as both a biologist and a former cancer patient to examine the evidence—however preliminary—that we do have for an environmental link to human cancers. How much evidence is there? How should we take action in light of it?
This article reviews the effects of Endocrine Disrupters (EDs) on cancer incidence in humans, the analysis being practically restricted to the influence of xenoestrogens on the occurrence of breast cancer. Although receptor-mediated mechanisms play an important role in eliciting the effects of (xeno)oestrogens (XEs), it should not be overlooked that other pathways exist, and that any evaluation must therefore consist of the net result of all influences. It is not acceptable to base the evaluation of the effects of xenoestrogens, or their inclusion in the category of EDs, solely on their property to bind the oestrogen receptor (ER). Other mechanisms contribute in eliciting the oestrogenic response and are briefly reviewed. They include serum factors; the binding to carrier proteins; alterations in metabolic pathways; interactions between oestrogens; growth factors and their receptors and oncogenes; disturbances in signal transduction pathways and interference with several accessory mechanisms of carcinogenesis. The particular carcinogenesis mechanisms underlying the action of (xeno)oestrogens are discussed. This mechanism implicates the absence of any threshold and-consequently-inducing effects at the lowest possible concentrations: one molecule. Other factors influencing the effects of XEs are represented by the occurrence of a membrane oestrogen receptor, different from the traditional nuclear ER, participating in the regulation of Prolactin production by the hypophysis after stimulation of the hypothalamic-hypophyseal pathways. Strong co-operation also exists with thyroid hormones. Many epidemiological studies, of which some are critically reviewed, support the basic role of XEs as one of the causative factors of breast cancer. This only serves to underscore the compulsory application of the precautionary principle, once a compound has been proven to exert endocrine disrupting properties, and the necessity to evaluate all modes of action of EDs.
We report here about the practice of using metanil yellow, a non permitted synthetic dye, in the adulteration of some food items produced by the organized and unorganized sectors located in different districts in West Bengal, India. We considered three food items- turmeric powder, ladoo and besan for the detection of the presence of metanil yellow. We observed that 58 of the 253 samples i.e. 20.94% of total samples contain metanil yellow in which 36.21% of the positive samples contained the metanil yellow below the maximum permissible limit i.e., below the 100 mg kg-1 food samples and 63.79% of the positive samples contained above the maximum permissible limit i.e., above the 100 mg kg-1 food samples as specified in the Prevention of Food Adulteration Act of India (PFA, 2008). We observed insignificant presence of metanil yellow in besan samples. We did not observe significant presence of metanil yellow in the same food samples collected from the organized sectors. We also found that all the positive samples i.e., the samples containing significant amount of metanil yellow were prepared from the food items collected from the unorganized sectors. From the study it is concluded that the unorganized sectors practice to use metanil yellow indiscriminately to adulterate the food items. We suggest strict governmental vigilance to prevent food adulteration with metanil yellow to avoid human health hazards.
This new clinical resource clearly explains how to approach integrated care in a way that combines Chinese herbal medicine with Western medicine to enhance and improve medical care for patients with cancer without undermining or negatively impacting patients' medical treatment. Each chapter covers a different type of cancer, first introducing the conventional medical understanding of that cancer including its etiology, diagnosis, and treatment according to staging and type. The chapter then covers that cancer from the perspective of Oriental medicine. Case studies illustrate the integration of treatment for each cancer type, raising important issues and considerations associated with specific cancers and treatments.
Currently, natural food colorants and preservatives are being used for their general health benefits. Monascus koji, the product of certain fungi that grow on rice grains, has been added to many foods for coloring and preservation. In this study, the antimicrobial activities of Monascus koji ethanol extracts were investigated. Six Monascus strains (M. araneosus KFRI 00371, M. kaoliang ATCC 46597, M. pilosus IFO 4520, M. purpureus IFO 4482, M. ruber IFO 32318 and M. sp. ATCC 16437) were selected based on their relative intensity of red pigment. Two Monascus extracts, M. kaoliang ATCC 46597 and M. purpureus IFO 4482, displayed antimicrobial activities against Bacillus subtilis, B. cereus, Micrococcus luteus, Staphylococcus aureus and Salmonella typhimurium in concentration-dependent manners. The two extracts showed their strongest antimicrobial activity against S. typhimurium, a cause of food poisoning. Therefore, these results suggest that Monascus koji could be used as a natural food colorant and preservative.
In this paper, the capacity of a soil to retain the dye Erythrosine B from aqueous solution by sorption has been studied. Batch adsorption experiments were conducted to investigate the sorption of the dye from aqueous solutions onto soil with particles of different size. Different models were used to describe the kinetic data, to calculate the corresponding rate constants and to predict the theoretical capacities of soil for dye sorption.
Novel coated graphite electrode (CGE) for the determination of quinoline yellow (QY), an artificial food colorant, was evaluated based on the use of tetraphenylphosponium-QY ion-pair complex as the electroactive substance. The electrode revealed linear emf-pQY response over wide concentration ranges (1.0 × 10−1 ∼ 5.0 × 10−5 M) with a slope −30.1 ± 0.2 mV decade−1 and a limit of detection of 4.0 × 10−5 M. The electrode showed good stability, reproducibility and fast response (<5 s). It can be used in wide pH range (4.7–10.7) and was successfully applied to determination of quinoline yellow in artificial mixtures and commercial soft drinks. The results obtained with the electrode were in good agreement with the value obtained by using HPLC measurements, as an official method.
Estrogen receptor (ER) transactivation assays were initially designed to study endogenous mechanisms of steroid hormone action, but more recently have been used to assess industrial chemicals for potential estrogenic activity. Given the diverse spectrum of physicochemical properties of these chemicals, we examined the effects of pH (a weak organic and strong inorganic acid and base), hyperosmolality (NaCl, mannitol) and two different vehicles (DMSO, Triton X-100) on responses to estradiol-17β (E2) in an ER transactivation assay. MCF-7 human breast cancer cells were transiently transfected with a chimeric estrogen receptor (Gal4-HEG0) and a Gal4-regulated luciferase reporter gene (17m5-G-Luc), treated with E2 under various test conditions, and then assessed for ER-mediated luciferase activity. Maximal E2-induced reporter activity was observed at pH 7.8 (pre-incubation), but was markedly reduced at pH ⩽7.5 or ⩾8.0 (P < 0.001), even though there was no evidence of cytotoxicity. Hyperosmolality induced by addition of mannitol (⩾25 mM) resulted in significant decreases in overall assay responsiveness, whereas NaCl (⩾80 mM) decreased the sensitivity of the assay by increasing the no-observed-effect level for E2 compared to control cultures (330 mOsm). The maximal DMSO concentration that resulted in consistently high E2-induced reporter activity was 0.1%, whereas concentrations of Triton X-100 above 1 ppm inhibited E2-induced reporter responses and were cytotoxic above 10 ppm. These results indicate that various physicochemical factors have the potential to confound assay data if not kept within predefined operational limits. Copyright © 2000 John Wiley & Sons, Ltd.
In this research, the construction and general performance characteristics of a sunset yellow sensor based on sunset yellow-cetyl pyridinum (SY-CPY) ion pair as an ion exchanger were described. A coated platinum wire electrode (CPE) was prepared and compared with coated graphite (CGE) and membrane electrode (PME). The CPE exhibited a rapid and Nernstian response (-29.77 ± 0.2 mV decade-1) to SY concentration range from 3.16 × 10-7 to 3.16 × 10-3 mol dm-3 within pH 4.5-9.5. Interfering effects of some foreign substances were reported. The optimized matrix was successfully applied to the determination of SY in artificial mixtures and commercial soft drinks. The results showed good agreements with the determination made by use of high-performance liquid chromatography.
Science remains an eminently social institution, though the interactions between science and society are often poorly understood. This article addresses the context of recent efforts to identify environmental contributions to cancer. The authors first examine cancer incidence and discuss how incidence patterns may be related to air pollution and occupational and general toxic chemical and xenestrogen exposures. They then discuss the social context in which cancer research and treatment occurs, including the dominating role of the biomedical model and socioeconomic factors, including regulatory strategies that address single chemicals, corporate conflicts of interest, and the manipulation of public opinion. Last, they consider the broad context out of which cancer arises and discuss the merits of applying the precautionary principle to sustainable social policies. Progress in reducing cancer may be fruitfully made by returning our attention to broad-scale factors such as those affecting the quality of air, water, workplace, household environments, and the global climate.
In order to set up an accurate quality criteria for the Boraginaceae that have been traditionally used for medical purposes and food colorant, and to assess its viability as functional food ingredient, antioxidant tests were conducted on the wild and cultivated plants. Variety of indicators including total contents of phenol, DPPH, SOD-liked effect, hydroxy radical-scavenging effect, lecithin oxidation inhibitory effect, etc were analyzed. Wild and cultivated gromwell's total contents of phenol in their methanol extracts were 0.14% and 0.13%, while they were most active in ethyl acetate extracts and n-hexane extracts, respectively. values of methanol extract of the wild and cultivated plants were 794.41 /mL and 971.86 /mL, indicating that the wild plant is more responsive (p
Acid Red 51, commonly named as Erythrosine B (Ery B), a water-soluble xanthene class of dye, is
widely used in cosmetics, foodstuffs, medicines and textiles. Some procedures for textile
wastewater remediation, such as photochemical and chemical degradation are not recommended
mainly due to the formation of toxic by-products. Therefore, development of low-cost and
efficient methods, such as sorption on agricultural waste is a good alternative. The present paper
aims to investigate the removal of Ery B by sorption on agro-waste. Pumpkin seeds hull (PSH) has
been chosen as a sorbent. Batch experiments were conducted for the study of the influence of
some parameters on dye adsorption efficiency, such as: dye concentration (5 to 50 mg L-1),
temperature (20, 30, 40, 50ºC) and the dosage of adsorbents (5 to 30 g L-1). Kinetic studies of Ery
B adsorption on PSH were carried out at 25ºC, using aqueous solutions with 10, 20 and 30 mg L-1
of dye. All the experiments were conducted at the natural pH of the solution, 5.6. The results
showed that the hazardous water-soluble dye Ery B can be efficiently removed from the aqueous
solutions by adsorption over PSH. Using 20 g L-1 of PSH the amount of dye uptake was around 3.5
mg g-1. Adsorption has been correlated with the different adsorption isotherms and based on the
data free energy of adsorption (ΔGº), enthalpy (ΔHº), and entropy (ΔSº) were calculated.
We baked low-calorie bread by mixing charred cellulose granules with wheat flour, using the charred cellulose granules to eliminate toxic xanthene food dyes contained in processed foods from the alimentary canal. The size of the charred cellulose granules played an important role in determining good breadmaking properties in respect of the bread height (mm) and specific volume (SV, cm(3)/g). Charred cellulose granules with a diameter above 270 μm were blended with wheat flour at 10% to obtain bread with a lower caloric content (1020 kcal/gram of bread) than the control bread (1126 kcal) made solely from wheat flour. The charred cellulose granules taken out from the bread adsorbed toxic xanthene food dyes at around pH 6.5, such that toxic food dyes taken into the alimentary canal were excreted in the feces with the non-digestible cellulose granules.
This chapter highlights representative research accomplishments in the area of fundamental and applied studies of the tunable
LSPR. Specific applications in the Van Duyne laboratory include exploitation of the LSPR as a signal transduction mechanism
for sensing applications and optimization of SERS signals. The optical properties of metallic nanostructures will find future
application in the areas of dichroic filters, plasmonic waveguides, data storage, biological labels and sensors. Commercialization
of nanoparticle devices relies on better understanding of the electromagnetic interaction between nanoparticles as well as
the development of techniques that will preserve nanoparticle optical activity in adverse environments.
Anthocyanin pigments are extracted from various plants and used for diverse purposes. The overall goal of this study was to develop high-anthocyanin corn to enhance the economic efficiency of anthocyanin production. We determined and compared the anthocyanin contents from the different parts of purple corn in various breeding lines. Our results revealed that purple corn produced the anthocyanin pigment throughout the plant, especially high in the husk and cob regions, although anthocyanin levels varied significantly among different plant parts. We analyzed the 295 selected lines from the 2006 breeding population, and it showed that anthocyanin levels of husks ranged from 17.3% to 18.9% of dry weight, roughly 10 times more than the standard current purple corn kernel content, 1.78%. LC-MS/MS analysis demonstrated that the main components of purple corn husk anthocyanin were cyanidin derivatives, and the most prevalent constituents were cyanidin-3-glucoside, cyanidin-3-succinylglucoside and pelargonidin-3-(6''-malonylglucoside). The results suggested that high-anthocyanin corn will boost the purple corn pigment production far more than its current level.
It has been suggested that dietary estrogens neutralize the effect of synthetic chemicals that mimic the effects of estrogen (i.e., xenoestrogens, environmental estrogens). Genistein, a dietary estrogen, inhibits the growth of breast cancer cells at high doses but additional studies have suggested that at low doses, genistein stimulates proliferation of breast cancer cells. Therefore, if dietary estrogens are estrogenic at low doses, one would predict that they stimulate estrogen-receptor positive breast cancer cells to enter the cell cycle. Genistein and the fungal toxin zearalenone were found to increase the activity of cyclin dependent kinase 2 (Cdk2) and cyclin D1 synthesis and stimulate the hyperphosphorylation of the retinoblastoma susceptibility gene product pRb105 in MCF-7 cells. The steroidal antiestrogen ICI 182,780 suppressed dietary estrogen-mediated activation of Cdk2. Dietary estrogens not only failed to suppress DDT-induced Cdk2 activity, but were found to slightly increase enzyme activity. Both zearalenone and genistein were found to stimulate the expression of a luciferase reporter gene under the control of an estrogen response element in MVLN cells. Our findings are consistent with a conclusion that dietary estrogens at low concentrations do not act as antiestrogens, but act like DDT and estradiol to stimulate human breast cancer cells to enter the cell cycle.
As inherited germ line mutations, such as loss of BRCA1 or AT, account for less than 5% of all breast cancer, most cases involve acquired somatic perturbations. Cumulative lifetime exposure to bioavailable estradiol links most known risk factors (except radiation) for breast cancer. Based on a series of recent experimental and epidemiologic findings, we hypothesize that the multistep process of breast carcinogenesis results from exposure to endogenous or exogenous hormones, including phytoestrogens that directly or indirectly alter estrogen metabolism. Xenohormones are defined as xenobiotic materials that modify hormonal production; they can work bifunctionally, through genetic or hormonal paths, depending on the periods and extent of exposure. As for genetic paths, xenohormones can modify DNA structure or function. As for hormonal paths, two distinct mechanisms can influence the potential for aberrant cell growth: compounds can directly bind with endogenous hormone or growth factor receptors affecting cell proliferation or compounds can modify breast cell proliferation altering the formation of hormone metabolites that influence epithelial-stromal interaction and growth regulation. Beneficial xenohormones, such as indole-3-carbinol, genistein, and other bioflavonoids, may reduce aberrant breast cell proliferation, and influence the rate of DNA repair or apoptosis and thereby influence the genetic or hormonal microenvironments. Upon validation with appropriate in vitro and in vivo studies, biologic markers of the risk for breast cancer, such as hormone metabolites, total bioavailable estradiol, and free radical generators can enhance cancer detection and prevention.
Controversy exists over the role that environmental estrogens, such as pesticides or polychlorinated biphenyls (PCBs), might play as risk factors for breast cancer. Several laboratory studies have suggested that these chemicals function as weak estrogens, binding to the estrogen receptor and inducing various measures of estrogen response. However, epidemiologic studies assessing the link between exposure to pesticides or PCBs and breast cancer have generally not shown enhanced breast cancer risk with higher levels of xenoestrogen exposure. These findings heighten our uncertainty about the relevance of the preclinical findings to human breast cancer risk.
Nowadays, the need to have a realistic characterization of industrial effluents in the environment has become more and more recognized. A palette of different analytical methods both for sample extraction and instrumental analysis are available today, some older, others introduced more recently. The aim of this research is to compare a number of these techniques. To do this we studied a real leachate from an industrial landfill and carried out chemical analyses for organic pollutants, using different extraction methods based on solid-phase extraction and solid-phase microextraction and different instrumental techniques such as GC-MS, LC-MS, NMR and LC-NMR. Results show the performances of the different techniques, which are complementary.
Estrogen receptor (ER) transactivation assays were initially designed to study endogenous mechanisms of steroid hormone action, but more recently have been used to assess industrial chemicals for potential estrogenic activity. Given the diverse spectrum of physicochemical properties of these chemicals, we examined the effects of pH (a weak organic and strong inorganic acid and base), hyperosmolality (NaCl, mannitol) and two different vehicles (DMSO, Triton X-100) on responses to estradiol-17beta (E2) in an ER transactivation assay. MCF-7 human breast cancer cells were transiently transfected with a chimeric estrogen receptor (Gal4-HEG0) and a Gal4-regulated luciferase reporter gene (17m5-G-Luc), treated with E2 under various test conditions, and then assessed for ER-mediated luciferase activity. Maximal E2-induced reporter activity was observed at pH 7.8 (pre-incubation), but was markedly reduced at pH < or =7.5, or > or =8.0 (P < 0.001), even though there was no evidence of cytotoxicity. Hyperosmolality induced by addition of mannitol (> or =25 mM) resulted in significant decreases in overall assay responsiveness, whereas NaCl (> or =80 mM) decreased the sensitivity of the assay by increasing the no-observed-effect level for E2 compared to control cultures (330 mOsm). The maximal DMSO concentration that resulted in consistently high E(2)-induced reporter activity was 0.1%, whereas concentrations of Triton X-100 above 1 ppm inhibited E2-induced reporter responses and were cytotoxic above 10 ppm. These results indicate that various physicochemical factors have the potential to confound assay data if not kept within predefined operational limits.
Evidence from recent publications indicates that repeated exercise may enhance the quality of life of cancer patients. The lack of reported negative effects and the consistency of the observed benefits lead one to conclude that physical exercise may provide a low-risk therapy that can improve patients' capacity to perform activities of daily living and improve their quality of life. Repeated physical activity may attenuate the adverse effects of cancer therapy, prevent or reverse cachexia, and reduce risk for a second cancer through suppression of inflammatory responses or enhancement of insulin sensitivity, rates of protein synthesis, and anti-oxidant and phase II enzyme activities. These results most likely come about through the ability of physical exercise to attenuate a chronic inflammatory signaling process and to transiently activate the mitogen-activated protein kinase, c-Jun NH2-terminal kinase, c-Jun NH2-terminal kinase-mitogen-activated protein kinase, and nuclear factor-kappa B pathways and through its ability to enhance insulin sensitivity. Expanded molecular-based research into these areas may provide new insights into the biological mechanisms associated with cancer rehabilitation and endogenous risk.
A cross sectional study was designed to measure DDT residues and its metabolites in breast milk samples collected randomly from Saudi lactating mothers living in Al-Ehssa region; which was under leishmania control until 1995, and compare them to samples from mothers living in Riyadh region where no spraying activities was involved. p,p'-DDE, p,p'-DDD and p,p'-DDT residues were measured in 878 breast milk samples by Gas Chromatography/Electron Capture Detector (GC/ECD) and confirmed by Gas Chromatography/Mass Spectrometer Detector (GC/MSD). Variation in the DDT and its metabolites levels were investigated with respect to regional distribution. Wilcoxon rank sum tests showed that the average ranks of p,p'-DDE, p,p'-DDD, p,p'-DDT and sigma p,p'-DDT in lactating mothers from Al-Ehssa region were significantly higher than those living in Riyadh region. These differences supported our hypothesis that the implications of the spraying activities to control vector borne diseases in Al-Ehssa region are obvious. We estimated that 99.2% of infants of lactating mothers living in Al-Ehssa region had sigma p,p'-DDT daily intakes that exceeded 20 micrograms/Kg-day of body weight, the WHO/UNEP Acceptable Daily Intakes for a 5-Kg infant. Exposure of infants to these chemicals through breast-feeding is clearly a public health concern. Because the bulk of literature highlights the adverse health effects of DDT and its metabolites on children and infants, public health polices should enforce the ban of DDT use and advise pregnant and lactating women to avoid DDT containing food or any other type of exposure.
Prostate cancer has the highest prevalence of any nonskin cancer in the human body, with similar likelihood of neoplastic foci found within the prostates of men around the world regardless of diet, occupation, lifestyle, or other factors. Essentially all men with circulating androgens will develop microscopic prostate cancer if they live long enough. This review is a contemporary and comprehensive, literature-based analysis of the putative risk factors for human prostate cancer, and the results were presented at a multidisciplinary consensus conference held in Crystal City, Virginia, in the fall of 2002. The objectives were to evaluate known environmental factors and mechanisms of prostatic carcinogenesis and to identify existing data gaps and future research needs. The review is divided into four sections, including 1) epidemiology (endogenous factors [family history, hormones, race, aging and oxidative stress] and exogenous factors [diet, environmental agents, occupation and other factors, including lifestyle factors]); 2) animal and cell culture models for prediction of human risk (rodent models, transgenic models, mouse reconstitution models, severe combined immunodeficiency syndrome mouse models, canine models, xenograft models, and cell culture models); 3) biomarkers in prostate cancer, most of which have been tested only as predictive factors for patient outcome after treatment rather than as risk factors; and 4) genotoxic and nongenotoxic mechanisms of carcinogenesis. The authors conclude that most of the data regarding risk relies, of necessity, on epidemiologic studies, but animal and cell culture models offer promise in confirming some important findings. The current understanding of biomarkers of disease and risk factors is limited. An understanding of the risk factors for prostate cancer has practical importance for public health research and policy, genetic and nutritional education and chemoprevention, and prevention strategies.
Although intracranial dural metastasis of Ewing's sarcoma is a very rare finding, its imaging characteristics are similar to those of its primary form in the central nervous system. Thus, this tumor must be considered in the differential diagnosis of extra-axial dural masses.
Aromatase, a key steroidogenic enzyme that catalyses the conversion of androgens to estrogens, present a target for endocrine disrupting chemicals. However, little is known about the effect of pollutants on aromatase enzymes. In this study, we first optimized a non-radioisotope aromatase assay using rat ovarian microsomes in vitro and measuring the estrone level with an enzyme-linked immunosorbent assay (EIA method). The sensitivity of the EIA method was about ten times as high as that of the radioisotope (RI) method. A significant positive correlation was indicated between EIA and RI method. We used this EIA assay system to investigate the effects of aromatase activity on 45 chemicals that had previously been reported to act as endocrine disruptors or to have the possibility of having such an effect. Six of the chemicals, rose bengal, erythrosine, phloxine, allura red, gallic acid, and tributyltin, inhibited aromatase activity. The inhibitory effect of rose bengal was the strongest (IC50=2.4 x 10(-8) M), and its strength of inhibition was about 100 times that of a known aromatase inhibitor, 4-hydroxy-androstenedione (IC50=2.6 x 10(-6) M) but was about 1/25 that of fadrazole (IC50=1.0 x 10(-9) M). It is thought that this EIA method would be useful for measuring the aromatase activity of microstructures.
We investigated the accumulation of p53 proteins after heat stress and their association with HSP72/HSC73 using four human glioblastoma cell lines. Human glioblastoma cell lines U-87MG and A-172 exhibited no mutation in the region between the 2nd and 11th exons of the p53 gene, whereas A-7 and T98G had mutations in exon 5 and exon 7 of the p53 gene, respectively. In U-87MG and A-172, the levels of wild-type p53 protein were slightly increased by heat stress. Levels of mutant p53 protein were apparently increased by heat stress in A-7, but not in T98G. Furthermore, wild-type p53 proteins in both U-87MG and A-172 co-immunoprecipitated with antibody and HSP72 and HSC73 in them co-immunoprecipitated with anti-p53 antibody as did the mutant p53 proteins. These findings suggest that p53 proteins accumulated by heat stress are associated with HSP72 and HSC73.
Extracts containing wild-type or mutant human estrogen receptor (ER) have been used to study the binding of ER to its responsive element (ERE). Estradiol (E2) or the antiestrogen hydroxytamoxifen is required for ER binding as assayed by gel retardation. The DNA binding domain (DBD) encompasses the highly conserved region C. Both intact ER-E2 complexes and ER mutants truncated for the hormone binding domain (HBD) bind as dimers to an ERE. However, an HBD-truncated ER binds less tightly to an ERE than an intact ER-E2 complex. The DBD and HBD contain a constitutive and a stronger ER-induced dimerization function, respectively. Thus, in addition to inducing the activation function associated with the HBD, estrogen plays a crucial role in the formation of stable ER dimers that bind tightly to ERE.
Sewage, a complex mixture of organic and inorganic chemicals, is considered to be a major source of environmental pollution. A random screen of 20 organic man-made chemicals present in liquid effluents revealed that half appeared able to interact with the estradiol receptor. This was demonstrated by their ability to inhibit binding of 17 beta-estradiol to the fish estrogen receptor. Further studies, using mammalian estrogen screens in vitro, revealed that the two phthalate esters butylbenzyl phthalate (BBP) and di-n-butylphthalate (DBP) and a food antioxidant, butylated hydroxyanisole (BHA) were estrogenic; however, they were all less estrogenic than the environmental estrogen octylphenol. Phthalate esters, used in the production of various plastics (including PVC), are among the most common industrial chemicals. Their ubiquity in the environment and tendency to bioconcentrate in animal fat are well known. Neither BBP nor DBP were able to act as antagonists, indicating that, in the presence of endogenous estrogens, their overall effect would be cumulative. Recently, it has been suggested that environmental estrogens may be etiological agents in several human diseases, including disorders of the male reproductive tract and breast and testicular cancers. The current finding that some phthalate compounds and some food additives are weakly estrogenic in vitro, needs to be supported by further studies on their effects in vivo before any conclusions can be made regarding their possible role in the development of these conditions.
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It has been convincingly demonstrated that genotoxic stresses cause the accumulation of the tumor suppressor gene p53. One important consequence of increased p53 protein levels in response to DNA damage is the activation of a G1-phase cell cycle checkpoint. It has also been shown that G1-phase cell cycle checkpoints are activated in response to other stresses, such as lack of oxygen. Here we show that hypoxia and heat, agents that induce cellular stress primarily by inhibiting oxygen-dependent metabolism and denaturing proteins, respectively, also cause an increase in p53 protein levels. The p53 protein induced by heat is localized in the cytoplasm and forms a complex with the heat shock protein hsc70. The increase in nuclear p53 protein levels and DNA-binding activity and the induction of reporter gene constructs containing p53 binding sites following hypoxia occur in cells that are wild type for p53 but not in cells that possess mutant p53. However, unlike ionizing radiation, the accumulation of cells in G1 phase by hypoxia is not strictly dependent on wild-type p53 function. In addition, cells expressing the human papillomavirus E6 gene, which show increased degradation of p53 by ubiquitination and fail to accumulate p53 in response to DNA-damaging agents, do increase their p53 levels following heat and hypoxia. These results suggest that hypoxia is an example of a "nongenotoxic" stress which induces p53 activity by a different pathway than DNA-damaging agents.
Exposure to estrogens during critical periods of development induces teratogenic and carcinogenic lesions in the reproductive tracts of humans and experimental animals. It is important to determine the molecular and cellular targets of estrogenic chemicals and to establish the mechanisms by which interactions of estrogens with the developing genital tract results in permanent lesions of growth and differentiation. The experiments presented here were designed to examined the effects of neonatal estrogen exposure on the expression of two genes, lactoferrin and epidermal growth factor, that are subject to steroid hormone regulation. Using in situ and Northern RNA hybridization, immunoblotting, and immunohistochemistry, our data demonstrate that exposure to the synthetic estrogen, diethylstilbestrol, during a critical neonatal period results in the persistent ovary-independent induction of mRNA and protein encoded by these two genes in the mouse uterus and vagina. The constitutive expression of lactoferrin and EGF, and probably other estrogen-regulated genes, may contribute to the establishment of a permanently "estrogenized" phenotype which is then instrumental in the development of abnormal tissue morphogenesis, function, and neoplasia in the rodent reproductive tract.
Changes in documented risk factors for breast cancer and rates of screening cannot completely explain recent increases in incidence or mortality. Established risk factors for breast cancer, including genetics, account for at best 30% of cases. Most of these risk factors can be linked to total lifetime exposure to bioavailable estrogens. Experimental evidence reveals that compounds such as some chlorinated organics, polycyclic aromatic hydrocarbons (PAHs), triazine herbicides, and pharmaceuticals affect estrogen production and metabolism and thus function as xenoestrogens. Many of these xenoestrogenic compounds also experimentally induce mammary carcinogenesis. Recent epidemiologic studies have found that breast fat and serum lipids of women with breast cancer contain significantly elevated levels of some chlorinated organics compared with noncancer controls. As the proportion of inherited breast cancer in the population is small, most breast cancers are due to acquired mutations. Thus, the induction of breast cancer in the majority of cases stems from interactions between host factors, including genetics and environmental carcinogens. We hypothesize that substances such as xenoestrogens increase the risk of breast cancer by mechanisms which include interaction with breast-cancer susceptibility genes. A series of major epidemiologic studies need to be developed to evaluate this hypothesis, including studies of estrogen metabolism, the role of specific xenoestrogenic substances in breast cancer, and relevant genetic-environmental interactions. In addition, experimental studies are needed to evaluate biologic markers of suspect xenoestrogens and biologic markers of host susceptibility and identify pathways of estrogenicity that affect the development of breast cancer. If xenoestrogens do play a role in breast cancer, reductions in exposure will provide an opportunity for primary prevention of this growing disease.(ABSTRACT TRUNCATED AT 250 WORDS)
In mammalian cells inhibition of the cdc2 function results in arrest in the G2-phase of the cell cycle. Several cdc2-related gene products have been identified recently and it has been hypothesized that they control earlier cell cycle events. Here we have studied the relationship between activation of one of these cdc2 homologs, the cdk2 protein kinase, and the progression through the cell cycle in cultured human fibroblasts. We found that cdk2 was activated and specifically localized to the nucleus during S phase and G2. Microinjection of affinity-purified anti-cdk2 antibodies but not of affinity-purified anti-cdc2 antibodies, during G1, inhibited entry into S phase. The specificity of these effects was demonstrated by the fact that a plasmid-driven cdk2 overexpression counteracted the inhibition. These results demonstrate that the cdk2 protein kinase is involved in the activation of DNA synthesis.
We have investigated the effect of chemotherapeutic and DNA damaging agents on binding of the tumor suppressor phosphoprotein p53 to its consensus DNA sequence. Activation of p53-DNA binding was seen for treatment with radiation, hydrogen peroxide, actinomycin D, Adriamycin, etoposide, camptothecin, 5-fluorouracil, mitomycin C, and cisplatin. These results showed that DNA strand breaks were sufficient to lead to increased levels of p53. The protein synthesis inhibitor cycloheximide blocks the increase in p53 following DNA damage. The increase in p53 activation in camptothecin treated cells may result, at least in part, from an increased half-life of the protein and consequent increases in intracellular protein concentration.
The estrogen receptor (ER) can be activated as a transcription factor either by binding of cognate estrogenic ligand or, indirectly, by a variety of other extracellular signals. As a first step towards elucidating the mechanism of 'steroid-independent activation' of the ER by the epidermal growth factor (EGF), we have mapped the ER target domain and determined the signaling pathway. We show that the N-terminal transcriptional activation function AF-1, but not the C-terminal AF-2, is necessary for the EGF response. Both the EGF-induced hyperphosphorylation and the transcriptional activation of the unliganded ER depend on a phosphorylatable serine residue at position 118. However, its phosphorylation is not sufficient and, hence, there must be other target domains or proteins which fulfill an additional requirement for EGF signaling through the ER. Using dominant-negative Ras and MAP kinase kinase (MAPK kinase) and constitutively active MAPK kinase mutants, we show that EGF activates the ER by signaling through the MAPK pathway suggesting that MAPK directly phosphorylates the critical serine 118. Our results also imply that the steroid-independent activation of a variety of ER mutants, which arise during the malignant progression of breast tumors, may contribute to tamoxifen resistance.
Cyclin-dependent kinases (Cdk) act to regulate G1- to S-phase transition in mammalian cells. We have studied the effects of estradiol and the steroidal antiestrogen ICI 182, 780 on induction of Cdk activity and the consequent phosphorylation of retinoblastoma protein (Rb) in estrogen-responsive MCF-7 breast cancer cells. Treatment of growth-arrested MCF-7 cells with physiological concentrations of estradiol led to a time-dependent increase in Cdk2-associated and cyclin E-dependent kinase activity, which was accompanied by hyperphosphorylation of Rb and S-phase entry. Induction of both Cdk2 activity and DNA synthesis by estradiol was dose dependent and was inhibited by coadministration of ICI 182,780. Elicitation of Cdk2 activity was found to require prolonged (> 8h) estradiol exposure. Levels of cyclins E and A were unchanged in MCF-7 cells undergoing G1- to S-transit; however, synthesis and steady state levels of cyclin D1 protein were increased by estradiol. Cdk4-associated Rb kinase activity was evident in MCF-7 cells by 6 h after estradiol exposure and was inhibited by antiestrogen. Cdk2 and Cdk4 protein levels were not altered by estrogen treatment; however, faster migrating, phosphorylated Cdk2 forms increased in estradiol-treated MCF-7 cells by 12 after release from growth arrest. Cdtk-inhibitory activities, associated with p27kip-1, were eliminated from growth-arrested MCF-7 cells after treatment with estradiol but were not eliminated from cells cotreated with estradiol and ICI 182,780. These findings suggest that estradiol regulates G1 progression in MCF-7 cells through direct effects upon Cdk activation, Rb phosphorylation, and by inducing elimination of Cdk inhibitors.
It has been convincingly demonstrated that genotoxic stresses cause the accumulation of the tumor suppressor gene p53. One important consequence of increased p53 protein levels in response to DNA damage is the activation of a G1-phase cell cycle checkpoint. It has also been shown that G1-phase cell cycle checkpoints are activated in response to other stresses, such as lack of oxygen. Here we show that hypoxia and heat, agents that induce cellular stress primarily by inhibiting oxygen-dependent metabolism and denaturing proteins, respectively, also cause an increase in p53 protein levels. The p53 protein induced by heat is localized in the cytoplasm and forms a complex with the heat shock protein hsc70. The increase in nuclear p53 protein levels and DNA-binding activity and the induction of reporter gene constructs containing p53 binding sites following hypoxia occur in cells that are wild type for p53 but not in cells that possess mutant p53. However, unlike ionizing radiation, the accumulation of cells in G1 phase by hypoxia is not strictly dependent on wild-type p53 function. In addition, cells expressing the human papillomavirus E6 gene, which show increased degradation of p53 by ubiquitination and fail to accumulate p53 in response to DNA-damaging agents, do increase their p53 levels following heat and hypoxia. These results suggest that hypoxia is an example of a "nongenotoxic" stress which induces p53 activity by a different pathway than DNA-damaging agents.
Few processed foods can be found that do not contain one or more of the thousands of different chemicals and materials that are food additives. According to recent estimates, about $5.5 billion worth of additives are used widely in the $385 billion U.S. food and beverage market. Throughout the 1990s, the U.S. market for food additives is expected to grow 4 to 6% annuallyto as much as $7 billion by 1995. The development and use of food additives is frequently driven by consumer demand and often becomes a balancing act between consumers' seemingly contradictory desires—the desire for more "natural" foods versus that for prepared, often microwaveable, convenience foods. The situation is complicated further by the desire for low-calorie, lowfat, and low-cholesterol foods. Health concerns and the desire for convenience foods will drive market growth for food additives, particularly for new products such as high-intensity sweeteners and fat replacers. The impact of these trends on ...
The retinoblastoma gene product (the RB protein) is phosphorylated in a cell cycle-dependent manner and this modification is believed to be important for cells to progress through the cell cycle. We found that purified cdk2 (cyclin-dependent kinase/cell division kinase 2) can phosphorylate the RB protein in vitro at the sites phosphorylated in the cell. The timing of activation of cdk2 in the cell cycle was similar to that of the onset of phosphorylation of the RB protein. The kinase coprecipitated with the RB protein also exhibited a similar substrate specificity to cdk2 and a similar time course of activation during the cell cycle. We further showed that cdk2 formed a complex with the RB protein in vitro and that its formation was not competitively inhibited by the simian virus 40 large T antigen. These observations suggest that cdk2 or a cdk2-related protein is involved in the cell cycle-dependent phosphorylation of the RB protein.
This article has no abstract; the first 100 words appear below.
BREAST cancer is a major public health problem of great interest and importance to physicians in a variety of specialties. Since this topic was last reviewed in the Journal,¹ the incidence of the disease has increased dramatically, heightening concern among physicians and women in general. In addition, long-term results are now available from clinical trials initiated in the 1970s and 1980s to evaluate the usefulness of early detection with mammography and physical examination, breast-conserving treatment with limited breast surgery and irradiation, and adjuvant systemic therapy with hormonal therapy and chemotherapy. Furthermore, in the light of newly gained knowledge, new . . .
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From the Departments of Radiation Oncology, Beth Israel Hospital and the Dana–Farber Cancer Institute, and the Joint Center for Radiation Therapy, Harvard Medical School, Boston (J.R.H.); the Vincent T. Lombardi Cancer Research Center and the Departments of Medicine and Pharmacology, Georgetown University Medical Center, Washington, D.C. (M.E.L.); the Istituto Nazionale per lo Studio e la Cura dei Tumori, Milan, Italy (U.V.); and the Departments of Epidemiology and Nutrition, Harvard School of Public Health and the Channing Laboratory, Departments of Medicine, Harvard Medical School and Brigham and Women's Hospital, Boston (W.W.). Address reprint requests to Dr. Harris at the Harvard Joint Center for Radiation Therapy, 50 Binney St., Boston, MA 02115.
Breast cancer is the second leading cause of death by cancer among women in the United States. The total cost of illness for breast cancer has been estimated to be $3.8 billion, of which $1.8 billion represents medical care costs. It has been estimated that breast cancer detected early is considerably less expensive than when the tumor is discovered at a later stage. Mass screening using mammography can improve early detection by as much as 15-35%. Cost-effectiveness studies have estimated that the costs of breast cancer screening range between $13,200 and $28,000 per year of life saved. The cost-effectiveness of breast cancer screening in the 40-49-year-old age group is controversial. Mass screening for breast cancer will probably increase total health care costs, but when all economic costs are included, screening appears to be more cost-effective than not screening.
The growing proportion of processed foods in the average daily diet in industrialized countries is considered as a challenge to the food industry, not only to provide more variety to the food assortment, but also to cope with high nutritional quality standards. Industrial handling of food therefore includes the monitoring of numerous operations and conditions from the agricultural source, through processing, packaging and distribution. Various types of food raw materials are processed by different kinds of technological treatments, the main objectives being to guarantee wholesomeness, taste, nourishment and convenience. New processing methods combined with the cold chain distribution reduce micronutrient losses to a certain extent. We shall examine some of the critical technological operations with regard to losses of vitamins and minerals in representative examples of foods like milk, leafy vegetables, potatoes, etc. and compare the classical food preservation with more modern food handling. With the increasing demand of pre-prepared fresh, refrigerated or frozen foods, the contribution of the maintained micronutrients is of interest, especially with reference to modern eating habits, i.e. lowering the total food intake and improving the nutrient density in the normal diet plan.
We have learned through the study of human breast cancer that this disease is the result of a combination of factors, among them external factors such as ionizing radiation, diet, socioeconomic status and endocrinologic, familial or genetic factors. Among all of these factors, the development of breast cancer is influenced by the reproductive history of the individual; lower risk of breast cancer has been reported in women that have had an early full-term pregnancy (109, 110, 154, 162).
Synopsis
We review information highlighting the multiple roles of both steroidal (primarily oestrogen) and polypeptide regulators of mammary epithelial cell growth, emphasising the work of our laboratory. Effects of both classes of hormones are complex and involve multiple interactions with non-tumour, host tissue. Oestrogen may induce growth regulatory polypeptide growth factors and interact with them in hormone dependent breast cancer. Progression of hormone-dependent breast cancer to hormone independence may involve multiple genetic mechanisms of oncogene activation, loss of the oestrogen receptor, or loss of hormone responsivity of other gene products. Initial carcinogenesis and progression of mammary epithelium to cancer probably also requires both proliferative stimuli (oestrogen, polypeptide growth factors) and genetic damage, leading to qualitatively different hormonal responses (hormone responsive cancer). Future therapies should be designed to block hormonal stimulation better and to interfere with necessary activated or induced components of malignant progression such as oncogenes or polypeptide growth factors receptor systems.
This chapter discusses that the development of malignancy depends on interactions of inherited genetic factors, exposure to chemical carcinogens, damaging radiation, oncogenic viruses, mitogenic hormones, and other promotional agents. Studies of experimental animal model systems have allowed considerable insight into the mechanisms at work in the action of each of these components; however, the exact etiology of any human cancer is yet to be established. The chapter discusses the mechanisms of systemic estrogen actions in the breast cancer process. It explores mechanisms of loss of endocrine control in experimental and clinical breast cancer, commonly observed following systemic therapy. Loss of estrogenic control of breast cancer growth during malignant progression implies the existence of other growth control processes which take over in its place. Genetic events which evoke the malignant phenotype probably involve activation of dominant oncogenes and inactivation of dominant cancer suppressive genes. The mutation of cellular protooncogenes to yield highly active oncogenes is extremely important in chemical- and radiation-induced carcinogenesis.
Erythrosine (diNa, tetraiodofluorescein) was nonmutagenic to the Ames/Salmonella typhimurium strains TA97a, TA98, TA100, TA102, and TA104, to a concentration of 2 mg/plate. No mutative intermediates were detected on metabolism by rat caecal cell-free extracts or rat liver S9 mixture; or on incubation with the comutagens, harman and norharman (+/- S9). Instead, an unexpected dose-dependent suppression in spontaneous reversion frequencies was observed (maximum approximately equal to 35% decrease). Erythrosine was antimutagenic to benzo[a]pyrene, but it did not decrease the mutagenicity of the other adduct-forming mutagen, 4-nitroquinoline N-oxide. The food dye was strongly antimutagenic to the bifunctional alkylating agent, mitomycin C, though it did not exhibit a similar effect on the mutagenicity of the corresponding monofunctional agent, methyl methanesulphonate. It partially depressed the mutagenic potentials of sodium azide. The antimutagenic effect of erythrosine on an intercalating agent, ethidium bromide, was discernible only at the highest dose (2 mg/plate). These results have been interpreted in terms of a genointeractive role of erythrosine. Erythrosine produced differential toxic effects in repair-deficient (TA97a, TA98, TA100) and repair-proficient (TA102, TA104) Salmonella tester strains; survival of the repair-deficient strains was found to be decreased. Photoinduced potentiation of erythrosine toxicity was observed, although light irradiation in the presence of erythrosine did not modify the reversion frequencies of the tester strains. The evidence strongly suggests that erythrosine, which exhibits nonmutagenicity in the Ames/Salmonella test, can interact with DNA repair enzymes and/or with DNA.
The effects of erythrosine (Red 3), rose bengal B (Red 105) and thyroidectomy on the development of thyroid tumor were examined in male Wistar rats treated with N-bis(2-hydroxypropyl)nitrosamine (DHPN). Red 3 and Red 105 were used at 4% in the basal diet and were administered for 19 weeks from week 2 to 20. Thyroidectomy was performed by resection of the left lobe at week 4. Single injection of DHPN was performed intraperitoneally at 280 mg per 100 g body weight at the beginning of the experiment. Red 3 and Red 105 significantly promoted the development of thyroid tumors in thyroidectomized rats given DHPN, but had no significant effect in non-thyroidectomized rats. The incidence of thyroid tumors was 91% in rats with partial thyroidectomy, Red 3 and DHPN, 100% in rats with partial thyroidectomy, Red 105 and DHPN, and 64% in rats with partial thyroidectomy and DHPN. Serum TSH was 5.5 +/- 3.1 ng/ml in rats with partial thyroidectomy, Red 3 and DHPN, 2.1 +/- 2.2 ng/ml in rats with partial thyroidectomy, Red 105 and DHPN, and 1.5 +/- 0.5 ng/ml in rats with partial thyroidectomy and DHPN.
Erythrosine increased the yield of sporulation minus mutants of Bacillus subtilis excision repair-proficient strain 168 by approximately equal to 400% at a concentration of 1 mg/ml under ambient light conditions. This mutagenic response was dose-dependent (0-1 mg/ml). Significantly, the food dye did not mutate the corresponding repair-deficient B. subtilis, hcr-9 (exc-). A decrease in the mutagenicity of erythrosine to B. subtilis strain 168 was apparent on metabolism by rat liver S9 mixture or by rat caecal cell-free extracts. Erythrosine did not exhibit differential toxicity to B. subtilis excision repair-proficient (168) and -deficient (hcr-9) strains, although it was highly toxic to both strains. This indicated non-involvement of excision repair in the dye-mediated toxic reactions.
Although much attention has been paid to the removal of hormones from sera and to the development of serum-free media for studies on hormone-responsive cells in culture, little consideration has been given to the possibility that the media components themselves may have hormonal activity. We have found that phenol red, which bears a structural resemblance to some nonsteroidal estrogens and which is used ubiquitously as a pH indicator in tissue culture media, has significant estrogenic activity at the concentrations (15-45 microM) at which it is found in tissue culture media. Phenol red binds to the estrogen receptor of MCF-7 human breast cancer cells with an affinity 0.001% that of estradiol (Kd = 2 X 10(-5) M). It stimulates the proliferation of estrogen receptor-positive MCF-7 breast cancer cells in a dose-dependent manner but has no effect on the growth of estrogen receptor-negative MDA-MB-231 breast cancer cells. At the concentrations present in tissue culture media, phenol red causes partial estrogenic stimulation, increasing cell number to 200% and progesterone receptor content to 300% of that found for cells grown in phenol red-free media, thereby reducing the degree to which exogenous estrogen is able to stimulate responses. The antiestrogens tamoxifen and hydroxytamoxifen inhibit cell proliferation below the control level only when cells are grown in the presence of phenol red; in the absence of phenol red, the antiestrogens do not suppress growth. The estrogenic activity of phenol red should be considered in any studies that utilize estrogen-responsive cells in culture.
MT2 cells, a clonal cell line of MTW9/PL cells originally obtained from a mammary adenocarcinoma, from estrogen-responsive tumors in Wistar-Furth rats. o,p'-DDT supports MT2 tumor growth at a rate similar to 17 beta-estradiol. This effect of o,p'-DDT is dose-dependent and specific, since the DDT congener p,p'-DDD, which does not bind to tumor estrogen receptors, does not support tumor growth. This is the first demonstration that DDT can support the growth of an estrogen-responsive tumor.
Serine 118 is definitively identified as a major site of phosphorylation in the human estrogen receptor expressed in COS-1 cells treated with estradiol or phorbol ester. At least 30% of the estrogen receptor appears to be phosphorylated on serine 118 after treatment with estradiol or phorbol ester. Human estrogen receptor was expressed in COS-1 cells and labeled in vivo with [32P]orthophosphate in the presence of estradiol or phorbol ester. Immunopurified receptor was digested with cyanogen bromide. The most heavily labeled peptide (7 kilodaltons) was identified as amino acids 110-174 by microsequencing. Manual Edman degradation released a major portion of the 32P-label in the peptide at serine 118. A mutant with serine 118 replaced by alanine (S118A) had 80% less 32P-label in the 7 kilodalton peptide. Estrogen receptor labeled in vivo with [32P]-orthophosphate in the presence of estradiol or phorbol ester migrates electrophoretically as a doublet. The major difference between the bands is phosphorylation of serine 118 in the upshifted band. The mutant S118A does not show an upshifted band. Labeling of the estrogen receptor with [35S]methionine indicates that > or = 30% of the receptor is upshifted and suggests that > or = 30% of the receptor is phosphorylated on serine 118.
The phosphorylation of the human estrogen receptor (ER) serine residue at position 118 is required for full activity of the
ER activation function 1 (AF-1). This Ser118 is phosphorylated by mitogen-activated protein kinase (MAPK) in vitro and in cells treated with epidermal growth factor (EGF)
and insulin-like growth factor (IGF) in vivo. Overexpression of MAPK kinase (MAPKK) or of the guanine nucleotide binding protein
Ras, both of which activate MAPK, enhanced estrogen-induced and antiestrogen (tamoxifen)-induced transcriptional activity
of wild-type ER, but not that of a mutant ER with an alanine in place of Ser118. Thus, the activity of the amino-terminal AF-1 of the ER is modulated by the phosphorylation of Ser118 through the Ras-MAPK cascade of the growth factor signaling pathways.
Chemotherapeutic and DNA-damaging agents were found to increase p53 site-specific DNA binding in human breast and rat liver epithelial WBrasII cells which produce mutant p53. Increased p53 site-specific DNA binding by chemotherapeutic or DNA-damaging agents was also induced in transfected Saos-2 cells producing wild type or transforming mutant p53. Therefore, exposure of cells containing a transforming p53 mutant to chemotherapeutic or DNA-damaging agents may potentially enhance their transformation state and tumorigenic potential.
Upon cellular DNA damage, the p53 tumor suppressor protein transmits a signal to genes that control the cell cycle and apoptosis. One function of p53 that is important for its role in this pathway is its ability to function as a sequence-specific transcriptional activator. We demonstrate here that short single DNA strands can markedly stimulate the ability of human and murine p53 proteins to bind specifically to a p53 response element in supercoiled DNA. We also show that single-stranded DNA does not stimulate binding by a truncated p53 that lacks the C-terminal domain. Finally, we establish that a peptide spanning the p53 C-terminus has the ability in trans to stimulate sequence-specific DNA binding by p53 dramatically. These data taken together suggest a model in which the p53 C-terminus can recognize DNA structures resulting from damage-induced lesions, and this interaction can be propagated to regulate positively p53 sequence-specific DNA binding.
Insertion/deletion (IDL) mismatches in DNA are lesions consisting of extra bases on one strand. Here, the binding of p53 and its 14 kDa C-terminal domain to DNAs containing one or three 3-cytosine IDL mismatches was examined. Electron microscopy showed that both p53 forms bound predominantly as tetramers at the lesions while single-stranded binding proteins did not bind. Gel retardation assays showed that p53 formed highly stable complexes when the DNA contained the IDL mismatches, but only unstable complexes when the DNA lacked lesions (but did contain free ends). The highly stable complexes had a half-life of > 2 hr, suggesting that upon encountering lesions, p53 may recruit other proteins to the site, providing a signal for DNA damage.
Charles J. Sherr Howard Hughes Medical Institute Department of Tumor Cell Biology St. Jude Children’s Research Hospital 332 North Lauderdale Memphis, Tennessee 38104 Recent advances in our understanding of the cell division cycle are now tying the functions of Gl phase regulators to diverse processes involving signal transduction, differ- entiation, senescence, apoptosis, and malignant transfor- mation. What determines the rate of Gl phase progression, and how do cells integrate mitogenic and antiproliferative signals with the cell cycle machinery? Lessons From Budding Yeast In Saccharomyces cerevisiae, a single 34 kDa cyclin- dependent kinase (cdk) (p34cDCZB/cdc2, also known as cdkl) serves as a master controller of the cell cycle, assembling sequentially into active holoenzyme complexes with Gl, S phase, or mitotic cyclins temporally to direct distinct transitions (reviewed by Nasmyth, 1993; Reed, 1992). In the presence of appropriate nutrients, Gl cells that reach a critical size initiate DNA replication, form buds, and dupli- cate their spindle bodies in preparation for subsequent division. Gl cyclins (Clnl, Cln2, and Cln3) are required for these processes (Richardson et al., 1989) (see Figure I), and their overexpression contracts Gl phase and de- creases cell size. Cln3-Cdc28 is present throughout Gl, and its kinase activity appears necessary for the subse- quent transcriptional activation of the CLN7 and CLN2 genes (Tyers et al., 1993). In turn, the induced Clnl and Cln2 proteins associate with Cdc28, whose kinase activity further stimulates CLN7 and CLN2 transcription. CLN7 and CLN2 gene expression is controlled by a heterodimeric transcription factor composed of Swi4 and Swi6, and Cln- CdcPm Schwab and Nasmyth, 1993). The kinase activities of Clb-Cdc28 complexes are held in check by an inhibitory protein (p40sfc’) (Mendenhall, 1993) that accumulates early in Gl and is degraded shortly before S phase (Schwab et al., 1994). Phosphorylation of ~40~‘~’ by Clnl,Cln2-Cdc28 might trigger its ubiquitin- mediated degradation, thereby enabling the Cln-regulated kinases to control S phase entry indirectly. Haploid Gl phase cells can also undergo cell cycle ar- rest and mate to form diploids. Conjugation is provoked by pheromones (a and a factors), secreted by cells of oppo- site mating types, that trigger a receptor-mediated sig- naling pathway (serpentine receptor-heterotrimeric G
Five small case-control studies have examined the relationship between exposure to organochlorines and the risk of breast cancer and have found inconsistent results. In these studies, organochlorine levels in breast cancer patients were measured after (or at most 6 months before) diagnosis.
We tested the hypothesis that organochlorines are a risk factor for breast cancer, using prospectively gathered data on serum levels of DDE [1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene] (the main metabolite of the pesticide DDT [2,2-bis(p-chlorophenyl)-1,1,1-trichloroethane]) and polychlorinated biphenyls (PCBs).
Study subjects belonged to a cohort of 57,040 women (46,629 white, 8123 black, and 2288 Asian) from the San Francisco Bay Area who took a multiphasic health examination, independent of concern about risk of breast cancer, in the late 1960s. At that time, a sample of blood was obtained, then frozen and stored. Follow-up was through December 31, 1990. We conducted a nested case-control study of 150 case patients and 150 matched control subjects. A random sample of 50 women per racial/ethnic group who had been diagnosed with breast cancer more than 6 months after the multiphasic examination (mean follow-up = 14.2 years) was selected, and each case patient was matched to a cancer-free control subject.
Matched analyses found no differences in the case patients' and control subjects' serum levels of DDE (mean difference = 0.2 parts per billion [ppb]; 95% confidence interval [CI] = -6.7, 7.2) or PCBs (mean difference = -0.4 ppb; 95% CI = -0.8, 0.1). DDE levels, however, tended to be higher among black case patients compared with black controls (mean difference = 5.7 ppb; 95% CI = -3.3, 14.8), and PCBs were lower among white case patients compared with white controls (mean difference = -0.6 ppb; 95% CI = -1.2, -0.1). Organochlorine levels were significantly higher among black and Asian women compared with white women. The mean difference for DDE was 11.0 ppb for black women (95% CI = 4.3, 17.6) and 12.6 ppb for Asian women (95% CI = 6.0, 19.2); for PCBs, the respective differences were 0.8 ppb for black women (95% CI = 0.2, 1.4) and 1.4 ppb for Asian women (95% CI = 0.8, 1.9). The results were not altered by adjusting for relevant confounders, and the lack of association between exposure to organochlorines and breast cancer was present regardless of length of follow-up, year of diagnosis, or the case patient's menopausal and estrogen-receptor status.
The data do not support the hypothesis that exposure to DDE and PCBs increases risk of breast cancer.
Future investigations must consider the biologic mechanisms involved and variations in exposure to chemical pollutants and of breast cancer incidence rates among diverse groups of women.
Many recent studies have implicated dietary factors in the cause and prevention of important diseases, including cancer, coronary heart disease, birth defects, and cataracts. There is strong evidence that vegetables and fruits protect against these diseases; however, the active constituents are incompletely identified. Whether fat per se is a major cause of disease is a question still under debate, although saturated and partially hydrogenated fats probably increase the risk of coronary heart disease. One clear conclusion from existing epidemiologic evidence is that many individuals in the United States have suboptimal diets and that the potential for disease prevention by improved nutrition is substantial.
Damage to cellular DNA greatly increases the levels of the tumor-suppressor gene p53 and induces cell cycle arrest in G1. A critical function of wild-type p53 is its ability to bind to specific DNA sequences. The effect of DNA damage on the sequence-specific DNA-binding properties of cellular p53 was investigated using DNA gel mobility-shift assays with nuclear extracts from NIH-3T3 cells. DNA damage (initiated by radiation) induced a rapid, cycloheximide-sensitive increase in the levels of nuclear p53-DNA binding activity and an increase in the half-life of the p53 protein. Increased p53-DNA binding activity could be detected at low (0.2 Gy), non-lethal doses of radiation. The tumor promoter 12-O-tetradecanoyl phorbol 13-acetate (TPA) attenuated the DNA damage-induced increase in p53-DNA binding activity by decreasing the half-life of the p53 protein. The tumor promoter properties of TPA may therefore be mediated by interfering with the cellular p53 response to DNA damage. The increased levels of p53 bound to specific DNA sequences following DNA damage may induce cell cycle arrest. p53-mediated growth arrest could occur by inhibition of DNA replication and/or alterations in transcription of cell cycle genes.
Certain chemicals in the environment are estrogenic. The low potencies of these compounds, when studied singly, suggest that
they may have little effect on biological systems. The estrogenic potencies of combinations of such chemicals were screened
in a simple yeast estrogen system (YES) containing human estrogen receptor (hER). Combinations of two weak environmental estrogens,
such as dieldrin, endosulfan, or toxaphene, were 1000 times as potent in hER-mediated transactivation as any chemical alone.
Hydroxylated polychlorinated biphenyls shown previously to synergistically alter sexual development in turtles also synergized
in the YES. The synergistic interaction of chemical mixtures with the estrogen receptor may have profound environmental implications.
These results may represent a previously uncharacterized level of regulation of estrogen-associated responses.
Estrogens play a critical role in the etiology of found breast cancer. Estradiol promotes the growth of breast cancer cells in vivo and in vitro. Exogenous estrogens in both the environment and in the human diet increase the growth of breast cancer cells in vitro. A role for xenoestrogens in breast cancer etiology has been proposed but remains controversial. We examined the effects of the xenoestrogenic pesticide 1,1,1-trichloro-2,2-bis(chlorophenyl)ethane (DDT) on estrogen-receptor (ER)-positive MCF-7 and T-47D human breast cancer cells as well as on ER-negative HS 578Bst breast cancer cells and rat liver cells. Estradiol and DDT were found to increase the growth of MCF-7 cells in the presence of insulin. The activity of cyclin-dependent kinase (Cdk)2 increased in growth-arrested T-47D and MCF-7 cells treated with beta-estradiol or DDT. The steroidal antiestrogen ICI 182,780 prevented both growth and Cdk2 activation induced by estradiol or DDT. Increased phosphorylation of Cdk2 and the retinoblastoma protein (pRb1O5) was observed in ER-positive cells treated with DDT or estradiol. Cdk2 activity was not affected by DDT or estradiol in ER-negative HS 578Bst breast cancer cells or in rat liver epithelial cells. Cyclin D1 protein synthesis was increased by DDT and estradiol in MCF-7 cells. DDT and estradiol-induced ER-dependent transcriptional activation of estrogen response elements (EREs) in stably transfected MVLN cells, and ERE activation by low doses of DDT was increased by insulin. These findings suggest that DDT can stimulate breast cancer cells to enter into the cell cycle by directly affecting key regulatory elements. The relative potency of DDT in inducing cell-cycle progression appears to be only 100-300 times less than that of estradiol when measured in the presence of insulin. Therefore, the cancer risks associated with DDT exposure may be greater than first thought, especially when additional mitogenic stimuli are present.
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