Investigating ionizing radiation-induced changes in breast cancer cells using stimulated Raman scattering microscopy.

Significance: Altered lipid metabolism of cancer cells has been implicated in increased radiation resistance. A better understanding of this phenomenon may lead to improved radiation treatment planning. Stimulated Raman scattering (SRS) microscopy enables label-free and quantitative imaging of cellular lipids but has never been applied in this domain. Aim: We sought to investigate the radiobiological response in human breast cancer MCF7 cells using SRS microscopy, focusing on how radiation affects lipid droplet (LD) distribution and cellular morphology. Approach: MCF7 breast cancer cells were exposed to either 0 or 30 Gy (X-ray) ionizing radiation and imaged using a spectrally focused SRS microscope every 24 hrs over a 72-hr time period. Images were analyzed to quantify changes in LD area per cell, lipid and protein content per cell, and cellular morphology. Cell viability and confluency were measured using a live cell imaging system while radiation-induced lipid peroxidation was assessed using BODIPY C11 staining and flow cytometry. Results: The LD area per cell and total lipid and protein intensities per cell were found to increase significantly for irradiated cells compared to control cells from 48 to 72 hrs post irradiation. Increased cell size, vacuole formation, and multinucleation were observed as well. No significant cell death was observed due to irradiation, but lipid peroxidation was found to be greater in the irradiated cells than control cells at 72 hrs. Conclusions: This pilot study demonstrates the potential of SRS imaging for investigating ionizing radiation-induced changes in cancer cells without the use of fluorescent labels.

Insights into a Cancer-Target Demethylase: Substrate Prediction through Systematic Specificity Analysis for KDM3A.

Jumonji C (JmjC) lysine demethylases (KDMs) catalyze the removal of methyl (-CH3) groups from modified lysyl residues. Several JmjC KDMs promote cancerous properties and these findings have primarily been in relation to histone demethylation. However, the biological roles of these enzymes are increasingly being shown to also be attributed to non-histone demethylation. Notably, KDM3A has become relevant to tumour progression due to recent findings of this enzyme's role in promoting cancerous phenotypes, such as enhanced glucose consumption and upregulated mechanisms of chemoresistance. To aid in uncovering the mechanism(s) by which KDM3A imparts its oncogenic function(s), this study aimed to unravel KDM3A substrate specificity to predict high-confidence substrates. Firstly, substrate specificity was assessed by monitoring activity towards a peptide permutation library of histone H3 di-methylated at lysine-9 (i.e., H3K9me2). From this, the KDM3A recognition motif was established and used to define a set of high-confidence predictions of demethylation sites from within the KDM3A interactome. Notably, this led to the identification of three in vitro substrates (MLL1, p300, and KDM6B), which are relevant to the field of cancer progression. This preliminary data may be exploited in further tissue culture experiments to decipher the avenues by which KDM3A imparts cancerous phenotypes.

Development and characterization of a DNA aptamer for MLL-AF9 expressing acute myeloid leukemia cells using whole cell-SELEX.

Current classes of cancer therapeutics have negative side effects stemming from off-target cytotoxicity. One way to avoid this would be to use a drug delivery system decorated with targeting moieties, such as an aptamer, if a targeted aptamer is available. In this study, aptamers were selected against acute myeloid leukemia (AML) cells expressing the MLL-AF9 oncogene through systematic evolution of ligands by exponential enrichment (SELEX). Twelve rounds of SELEX, including two counter selections against fibroblast cells, were completed. Aptamer pools were sequenced, and three candidate sequences were identified. These sequences consisted of two 23-base primer regions flanking a 30-base central domain. Binding studies were performed using flow cytometry, and the lead sequence had a binding constant of 37.5 + / - 2.5 nM to AML cells, while displaying no binding to fibroblast or umbilical cord blood cells at 200 nM. A truncation study of the lead sequence was done using nine shortened sequences, and showed the 5' primer was not important for binding. The lead sequence was tested against seven AML patient cultures, and five cultures showed binding at 200 nM. In summary, a DNA aptamer specific to AML cells was developed and characterized for future drug-aptamer conjugates.

Responses of Porcupine and Wntless proteins to oxidative, hypoxic and endoplasmic reticulum stresses.

The WNT (Wingless and Int-1) proteins play a role in stem cell development and cell differentiation. Mutations in the WNT proteins lead to the development of various tumours, including gastric tumours. Porcupine (PORCN) is a palmitoyltransferase and Wntless (WLS) is a dedicated WNT transport protein that modify and fold the WNT proteins respectively and are involved in their proper secretion and binding to the frizzled (FZD) receptor and the lipoprotein receptor-related protein 5 or 6 (LRP5/6). We investigated how modifications of PORCN and WLS result in changes in WNT expression and secretion from cells under stress conditions that occur in the tumour microenvironment (hypoxia, oxidative stress, endoplasmic reticulum (ER) stress). In the present study, we found the mRNA expression of both PORCN and WLS were significantly increased with treatments inducing oxidative stress (antimycin A) and proteasome inhibition (MG-132), in human colon cancer (HCT116) and human intestinal epithelial cell-6 (HIEC-6) cells. Treatment with ER stressors thapsigargin, tunicamycin, and dithiolthreitol significantly increased PORCN gene expression, while treatment with thapsigargin and dithiolthreitol increased WLS gene expression. The expression of PORCN and WLS proteins increased with hypoxia and ER stressor treatments in both HCT116 and HIEC-6 cells. All stressors used in this study increased beta-catenin (ß-catenin) expression in HCT116 cells. Our results suggest that these stressors alter PORCN, WLS and ß-catenin expression and function which may, in turn, alter WNT secretion. Silencing the expression of PORCN and WLS with siRNA expression reduced the expression of WLS and WNT3A in HCT116 cells. The possibility exists that PORCN specifically may be involved in a novel signaling pathway, independent of its palmitoleation of the WNT proteins and its role in their secretion, that is rate-limiting for cancer cell growth and tumorigenesis, within the tumour microenvironment.

Self-monitored and optically powered fiber-optic device for localized hyperthermia and controlled cell death in vitro.

Localized hyperthermia therapy involves heating a small volume of tissue in order to kill cancerous cells selectively and with limited damage to healthy cells and surrounding tissue. However, these features are only achievable through real-time control of the tissue temperature and heated volume, both of which are difficult to obtain with current heating systems and techniques. This work introduces an optical fiber-based active heater that acts both as a miniature heat source and as a thermometer. The heat-induced damage in the tissue is caused by the conductive heat transfer from the surface of the device, while the heat is generated in an absorptive coating on the fiber by near-infrared light redirected from the fiber core to the surface by a tilted fiber Bragg grating inscribed in the fiber core. Simultaneous monitoring of the reflection spectrum of the grating provides a measure of the local temperature. Localized temperature increases between 0°C and 100°C in 10 mm-long/5 mm-diameter cylindrical volumes are obtained with continuous-wave pump power levels up to 1.8 W. Computational and experimental results further indicate that the temperature rise and dimensions of the heated volume can be maintained at a nearly stable level determined by the input optical power.

Hypoxia-Inducible Lysine Methyltransferases: G9a and GLP Hypoxic Regulation, Non-histone Substrate Modification, and Pathological Relevance.

Oxygen sensing is inherent among most animal lifeforms and is critical for organism survival. Oxygen sensing mechanisms collectively trigger cellular and physiological responses that enable adaption to a reduction in ideal oxygen levels. The major mechanism by which oxygen-responsive changes in the transcriptome occur are mediated through the hypoxia-inducible factor (HIF) pathway. Upon reduced oxygen conditions, HIF activates hypoxia-responsive gene expression programs. However, under normal oxygen conditions, the activity of HIF is regularly suppressed by cellular oxygen sensors; prolyl-4 and asparaginyl hydroxylases. Recently, these oxygen sensors have also been found to suppress the function of two lysine methyltransferases, G9a and G9a-like protein (GLP). In this manner, the methyltransferase activity of G9a and GLP are hypoxia-inducible and thus present a new avenue of low-oxygen signaling. Furthermore, G9a and GLP elicit lysine methylation on a wide variety of non-histone proteins, many of which are known to be regulated by hypoxia. In this article we aim to review the effects of oxygen on G9a and GLP function, non-histone methylation events inflicted by these methyltransferases, and the clinical relevance of these enzymes in cancer.

Hepatotoxicity of Cadmium Telluride Quantum Dots Induced by Mitochondrial Dysfunction.

The aim of this study was to investigate the detailed mechanisms of hepatotoxicity induced by cadmium telluride quantum dots (CdTe-QDs) in BALB/c mice after intravenous injection. The study investigated oxidative stress, apoptosis, and effects on mitochondria as potential mechanistic events to elucidate the observed hepatotoxicity. Oxidative stress in the liver, induced by CdTe-QD exposure, was demonstrated by depletion of total glutathione, an increase in superoxide dismutase activity, and changes in the gene expression of several oxidative stress-related biomarkers. Furthermore, CdTe-QD treatment led to apoptosis in the liver via both intrinsic and extrinsic apoptotic pathways. Effects on mitochondria were evidenced by the enlargement and increase in the number of mitochondria in hepatocytes of treated mice. CdTe-QDs also caused changes in the levels and gene expression of electron transport chain enzymes, depletion of ATP, and an increase in the level of the peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), a regulator of mitochondrial biogenesis. The findings from this study suggest that CdTe-QDs-induced hepatotoxicity might have originated from mitochondrial effects which resulted in oxidative stress and apoptosis in the liver cells. This study provides insight into the biological effects of CdT-QDs at the tissue level and the detailed mechanisms of their toxicity in animals. The study also provides important data for bridging the gap between in vitro and in vivo testing and risk assessment of these NPs.

Antioxidant, Physicochemical, and Cellular Secretion of Glucagon-Like Peptide-1 Properties of Oat Bran Protein Hydrolysates.

The aim of this work was to determine the physicochemical and biological activities of hydrolyzed proteins from sonicated oat brans. In addition to the control bran sample, two types of pre-treatment procedures-namely, ultrasonic bath and probe-type sonication-were performed to extract proteins, followed by hydrolysis with various proteases. Physicochemical analyses showed that Flavourzyme-hydrolysates had greater amounts of aromatic amino acids, Papain-hydrolysates low surface charges (-0.78 to -1.32 mV) compared to the others (-3.67 to -9.17 mV), and Alcalase-hydrolysates a higher surface hydrophobicity. The hydrolysates had good radical scavenging activities but, as the ultrasonic pre-treatment of the brans showed, in certain cases there was a reduction in activities of up to 22% for ROO• and HO• and 15% for O2•- radicals. In anti-diabetic tests, the maximum inhibition of α-amylase was 31.8%, while that of dipeptidyl peptidase-4 was 53.6%. In addition, the secretion of glucagon-like peptide-1 in NCI-H716 cells was enhanced by 11.5% in the presence of hydrolysates.

Insights into The Function and Regulation of Jumonji C Lysine Demethylases as Hypoxic Responsive Enzymes.

Cellular responses to hypoxia (low oxygen) are governed by oxygen sensitive signaling pathways. Such pathways, in part, are controlled by enzymes with oxygen-dependent catalytic activity, of which the role of prolyl 4-hydroxylases has been widely reviewed. These enzymes inhibit hypoxic response by inducing the oxygen-dependent degradation of hypoxia-inducible factor 1α, the master regulator of the transcriptional hypoxic response. Jumonji C domain-containing lysine demethylases are similar enzymes which share the same oxygen-dependent catalytic mechanism as prolyl 4- hydroxylases. Traditionally, the role of lysine demethylases has been studied in relation to demethylation activity against histone substrates, however, within the past decade an increasing number of nonhistone protein targets have been revealed, some of which have a key role in survival in the hypoxic tumor microenvironment. Within this review, we highlight the involvement of methyllysine in the hypoxic response with a focus on the HIF signaling pathway, the regulation of demethylase activity by oxygen, and provide insights into notable areas of future hypoxic demethylase research.

Rapid Detection of Circulating Breast Cancer Cells Using a Multiresonant Optical Fiber Aptasensor with Plasmonic Amplification.

The detection of circulating tumor cells (CTCs), which are responsible for metastasis in several forms of cancer, represents an important goal in oncological diagnosis and treatment. These cells remain extremely challenging to detect, despite numerous previous studies, due to their low concentration (1-10 cells/mL of blood). In this work, an all-fiber plasmonic aptasensor featuring multiple narrowband resonances in the near-infrared wavelength range was developed to detect metastatic breast cancer cells. To this aim, specific aptamers against mammaglobin-A were selected and immobilized as receptors on the sensor surface. In vitro assays confirm that the label-free and real-time detection of cancer cells [limit of detection (LOD) of 49 cells/mL] occurs within 5 min, while the additional use of functionalized gold nanoparticles allows a 2-fold amplification of the biosensor response. Differential measurements on selected optical resonances were used to process the sensor response, and results were confirmed by microscopy. The detection of only 10 cancer cells/mL was achieved with relevant specificity against control cells and with quick response time.

Biodistribution and Systemic Effects in Mice Following Intravenous Administration of Cadmium Telluride Quantum Dot Nanoparticles.

Quantum dots (QDs) are engineered nanoparticles (NPs) of semiconductor structure that possess unique optical and electronic properties and are widely used in biomedical applications; however, their risks are not entirely understood. This study investigated the tissue distribution and toxic effects of cadmium telluride quantum dots (CdTe-QDs) in male BALB/c mice for up to 1 week after single-dose intravenous injections. CdTe-QDs were detected in the blood, lung, heart, liver, spleen, kidney, testis and brain. Most CdTe-QDs accumulated in the liver, followed by the spleen and kidney. At high doses, exposure to CdTe-QDs resulted in mild dehydration, lethargy, ruffled fur, hunched posture, and body weight loss. Histological analysis of the tissues, upon highest dose exposures, revealed hepatic hemorrhage and necrotic areas in the spleen. The sera of mice treated with high doses of CdTe-QDs showed significant increases in alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin levels, as well as a reduction in albumin. CdTe-QD exposure also led to a reduced number of platelets and elevated total white blood cell counts, including monocytes and neutrophils, serum amyloid A, and several pro-inflammatory cytokines. These results demonstrated that the liver is the main target of CdTe-QDs and that exposure to CdTe-QDs leads to hepatic and splenic injury, as well as systemic effects, in mice. By contrast, cadmium chloride (CdCl2), at an equivalent concentration of cadmium, appeared to have a different pharmacokinetic pattern from that of CdTe-QDs, having minimal effects on the aforementioned parameters, suggesting that cadmium alone cannot fully explain the toxicity of CdTe-QDs.

Antioxidant and Anti-Apoptotic Properties of Oat Bran Protein Hydrolysates in Stressed Hepatic Cells.

The objective of this work was to find out how the method to extract proteins and subsequent enzymatic hydrolysis affect the ability of hepatic cells to resist oxidative stress. Proteins were isolated from oat brans in the presence of Cellulase (CPI) or Viscozyme (VPI). Four protein hydrolysates were produced from CPI and four others from VPI when they treated with Alcalase, Flavourzyme, Papain, or Protamex. Apart from CPI-Papain that reduced the viability of cell by 20%, no other hydrolysate was cytotoxic in the hepatic HepG2 cells. In the cytoprotection test, VPI-Papain and VPI-Flavourzyme fully prevented the damage due to peroxyl radical while CPI-Papain and CPI-Alcalase enhanced the cellular damage. Cells treated with VPI-hydrolysates reduced intracellular reactive oxygen species (ROS) by 20-40% and, also increased the intracellular concentration of glutathione, compared to CPI-hydrolysates. In antioxidant enzyme assays, although all hydrolysates enhanced the activity of both superoxide dismutase and catalase by up to 2- and 3.4-fold, respectively relative the control cells, the largest increase was due to VPI-Papain and VPI-Flavourzyme hydrolysates. In caspase-3 assays, hydrolysates with reduced ROS or enhanced antioxidant enzyme activities were able to reduce the activity of the pro-apoptotic enzyme, caspase-3 indicating that they prevented oxidative stress-induced cell death.

Peptidomic analysis of hydrolyzed oat bran proteins, and their in vitro antioxidant and metal chelating properties.

Peptide profiles of hydrolyzed oat proteins and the susceptibility of their polypeptides to proteolytic cleavages were determined using peptidomic analysis. In addition, antioxidant activities were also measured. Proteins isolates were first extracted with carbohydrases, Viscozyme or Cellulase and then hydrolyzed with proteases (Alcalase, Papain, Protamex, Flavourzyme). Amongst the eight hydrolysates, Viscozyme-proteins hydrolyzed with Papain showed the highest ability to quench ABTS+ radicals (866.9 ±â€¯10.6 µM TE/g) and to chelate ferrous ions (75 ±â€¯0.4%) while displaying the second strongest activity for ROO radicals (396.7 ±â€¯14.0 µM TE/g). Peptidomics analysis showed that the higher activity of papain hydrolysate in most assays was related to its greater proteolytic action on main proteins (avenin, 11S- and 12S-globulins) compared to other proteases. In addition, the number of peptides identified in the Papain digest of proteins extracted with Viscozyme was about half relative to the number in proteins from bran treated with Cellulase and digested with the same protease. This was likely because the carbohydrases differently affected polypeptide secondary structures.

A Current Overview of the Biological and Cellular Effects of Nanosilver.

Nanosilver plays an important role in nanoscience and nanotechnology, and is becoming increasingly used for applications in nanomedicine. Nanosilver ranges from 1 to 100 nanometers in diameter. Smaller particles more readily enter cells and interact with the cellular components. The exposure dose, particle size, coating, and aggregation state of the nanosilver, as well as the cell type or organism on which it is tested, are all large determining factors on the effect and potential toxicity of nanosilver. A high exposure dose to nanosilver alters the cellular stress responses and initiates cascades of signalling that can eventually trigger organelle autophagy and apoptosis. This review summarizes the current knowledge of the effects of nanosilver on cellular metabolic function and response to stress. Both the causative effects of nanosilver on oxidative stress, endoplasmic reticulum stress, and hypoxic stress-as well as the effects of nanosilver on the responses to such stresses-are outlined. The interactions and effects of nanosilver on cellular uptake, oxidative stress (reactive oxygen species), inflammation, hypoxic response, mitochondrial function, endoplasmic reticulum (ER) function and the unfolded protein response, autophagy and apoptosis, angiogenesis, epigenetics, genotoxicity, and cancer development and tumorigenesis-as well as other pathway alterations-are examined in this review.

N-acetylcysteine manipulation fails to elicit an increase in glutathione in a teleost model.

Levels of oxidative stress can be affected by a range of compounds including toxins and pharmaceuticals. Antioxidants are important protective compounds which counteract the damaging effects of oxidative stress. Glutathione (GSH) is one of the main antioxidants for many organisms and can be synthesized from administered N-acetylcysteine (NAC). NAC has therefore often been used in a wide range of taxa to manipulate levels of GSH. Our objective was to validate this approach in a wild temperate teleost fish model, the brown trout (Salmo trutta). We used intracoelomic injections of NAC in saline and vegetable shortening, at two different concentrations (100 and 400 mg/kg), with the appropriate controls and shams, under controlled laboratory settings. We found that NAC failed to elicit an increase in GSH over three time periods and concluded that NAC is not an effective method to enhance GSH levels in teleost fish using the concentrations and vehicles tested here. We emphasize the importance of validation studies across all new species/taxa when possible and suggest that more investigation is required with regard to NAC manipulation in fish if this approach is to be used.

In Vitro and in Silico Competitive Binding of Brominated Polyphenyl Ether Contaminants with Human and Gull Thyroid Hormone Transport Proteins.

Tetradecabromo-1,4-diphenoxybenzene (TeDB-DiPhOBz) is a highly brominated additive flame retardant (FR). Debrominated photodegradates of TeDB-DiPhOBz are hydroxylated in vitro in liver microsomal assays based on herring gulls (Larus argentatus), including one metabolite identified as 4″-OH-2,2',2″,4-tetrabromo-DiPhOBz. Chemically related methoxylated tetra- to hexabromo-DiPhOBzs are known contaminants in herring gulls. Collectively, nothing is currently known about biological effects of these polybrominated (PB) DiPhOBz-based compounds. The present study investigated the potential thyroidogenicity of 2,2',2″,4-tetrabromo-(TB)-DiPhOBz along with its para-methoxy (MeO)- and hydroxy-(OH)-analogues, using an in vitro competitive protein binding assay with the human thyroid hormone (TH) transport proteins transthyretin (hTTR) and albumin (hALB). This model para-OH-TB-DiPhOBz was found to be capable of competing with thyroxine (T4) for the binding site on hTTR and hALB. In silico analyses were also conducted using a 3D homology model for gull TTR, to predict whether these TB-DiPhOBz-based compounds may also act as ligands for an avian TH transport protein despite evolutionary differences with hTTR. This analysis found all three TB-DiPhOBz analogues to be potential ligands for gull TTR and have similar binding efficacies to THs. Results indicate structure-related differences in binding affinities of these ligands and suggest there is potential for these contaminants to interact with both mammalian and avian thyroid function.

Bisphenol A induces DSB-ATM-p53 signaling leading to cell cycle arrest, senescence, autophagy, stress response, and estrogen release in human fetal lung fibroblasts.

Experimental and/or epidemiological studies suggest that prenatal exposure to bisphenol A (BPA) may delay fetal lung development and maturation and increase the susceptibility to childhood respiratory disease. However, the underlying mechanisms remain to be elucidated. In our previous study with cultured human fetal lung fibroblasts (HFLF), we demonstrated that 24-h exposure to 1 and 100 µM BPA increased GPR30 protein in the nuclear fraction. Exposure to 100 µM BPA had no effects on cell viability, but increased cytoplasmic expression of ERß and release of GDF-15, as well as decreased release of IL-6, ET-1, and IP-10 through suppression of NFκB phosphorylation. By performing global gene expression and pathway analysis in this study, we identified molecular pathways, gene networks, and key molecules that were affected by 100, but not 0.01 and 1 µM BPA in HFLF. Using multiple genomic and proteomic tools, we confirmed these changes at both gene and protein levels. Our data suggest that 100 µM BPA increased CYP1B1 and HSD17B14 gene and protein expression and release of endogenous estradiol, which was associated with increased ROS production and DNA double-strand breaks, upregulation of genes and/or proteins in steroid synthesis and metabolism, and activation of Nrf2-regulated stress response pathways. In addition, BPA activated ATM-p53 signaling pathway, resulting in increased cell cycle arrest at G1 phase, senescence and autophagy, and decreased cell proliferation in HFLF. The results suggest that prenatal exposure to BPA at certain concentrations may affect fetal lung development and maturation, and thereby affecting susceptibility to childhood respiratory diseases.

In vitro selections of mammaglobin A and mammaglobin B aptamers for the recognition of circulating breast tumor cells.

Mammaglobin B (MGB2) and mammaglobin A (MGB1) are proteins expressed in metastatic breast cancers. The early detection of circulating tumor cells (CTCs) in breast cancer patients is crucial to decrease mortality rate. Herein, novel aptamers were successfully selected and characterized against MGB2 and MGB1 proteins using a hybrid SELEX approach. The potential use of the selected aptamers in breast CTC detection was studied using spiked breast cancer cells in whole blood lysate. The results obtained from this study showed that the selected aptamers (MAMB1 and MAMA2) bind to their target breast cancer cell lines with high affinity (low nanomolar Kd values) and specificity. They also bind to their free recombinant target proteins and show minimal non-specific binding to normal and other cancer cell lines. Additionally, they were able to distinguish a low number of breast cancer cells spiked in whole blood lysate containing normal blood cells. The results obtained in this study indicate the great potential for the use of aptamers to detect MGB1 and MGB2 protein biomarkers, expressed on the surface of breast CTCs.

Short-term and long-term effects of transient exogenous cortisol manipulation on oxidative stress in juvenile brown trout.

In the wild, animals are exposed to a growing number of stressors with increasing frequency and intensity, as a result of human activities and human-induced environmental change. To fully understand how wild organisms are affected by stressors, it is crucial to understand the physiology that underlies an organism's response to a stressor. Prolonged levels of elevated glucocorticoids are associated with a state of chronic stress and decreased fitness. Exogenous glucocorticoid manipulation reduces an individual's ability to forage, avoid predators and grow, thereby limiting the resources available for physiological functions like defence against oxidative stress. Using brown trout (Salmo trutta), we evaluated the short-term (2 weeks) and long-term (4 months over winter) effects of exogenous cortisol manipulations (versus relevant shams and controls) on the oxidative status of wild juveniles. Cortisol caused an increase in glutathione over a 2 week period and appeared to reduce glutathione over winter. Cortisol treatment did not affect oxidative stress levels or low molecular weight antioxidants. Cortisol caused a significant decrease in growth rates but did not affect predation risk. Over-winter survival in the stream was associated with low levels of oxidative stress and glutathione. Thus, oxidative stress may be a mechanism by which elevated cortisol causes negative physiological effects.

Bisphenol A exposure alters release of immune and developmental modulators and expression of estrogen receptors in human fetal lung fibroblasts.

Bisphenol A (BPA) has been shown to exert biological effects through estrogen receptor (ER)-dependent and ER-independent mechanisms. Recent studies suggest that prenatal exposure to BPA may increase the risk of childhood asthma. To investigate the underlying mechanisms in the actions of BPA, human fetal lung fibroblasts (hFLFs) were exposed to varying doses of BPA in culture for 24hr. Effects of BPA on localization and uptake of BPA, cell viability, release of immune and developmental modulators, cellular localization and expression of ERα, ERß and G-protein coupled estrogen receptor 30 (GPR30), and effects of ERs antagonists on BPA-induced changes in endothelin-1 (ET-1) release were examined. BPA at 0.01-100µmol/L caused no changes in cell viability after 24hr of exposure. hFLFs expresses all three ERs. BPA had no effects on either cellular distribution or protein expression of ERα, however, at 100µmol/L (or 23µmol/L intracellular BPA) increased ERß protein levels in the cytoplasmic fractions and GPR30 protein levels in the nuclear fractions. These paralleled with increased release of growth differentiation factor-15, decreased phosphorylation of nuclear factor kappa B p65 at serine 536, and decreased release of ET-1, interleukin-6, and interferon gamma-induced protein 10. ERs antagonists had no effects on BPA-induced decrease in ET-1 release. These data suggest that BPA at 100µmol/L altered the release of immune and developmental modulators in hFLFs, which may negatively influence fetal lung development, maturation, and susceptibility to environmental stressors, although the role of BPA in childhood asthma remains to be confirmed in in vivo studies.