The effect of maternal exposure to endocrine disrupting chemicals on fetal and neonatal development: A review on the major concerns

There is a widespread exposure of general population, including pregnant women and developing fetuses, to the endocrine disrupting chemicals (EDCs). These chemicals have been reported to be present in urine, blood serum, breast milk, and amniotic fluid. Endocrine disruptions induced by environmental toxicants have placed a heavy burden on society, since environmental exposures during critical periods of development can permanently reprogram normal physiological responses, thereby increasing susceptibility to disease later in life—a process known as developmental reprogramming. During development, organogenesis and tissue differentiation occur through a continuous series of tightly‐regulated and precisely‐timed molecular, biochemical, and cellular events. Humans may encounter EDCs daily and during all stages of life, from conception and fetal development through adulthood and senescence. Nevertheless, prenatal and early postnatal windows are the most critical for proper development, due to rapid changes in system growth. Although there are still gaps in our knowledge, currently available data support the urgent need for health and environmental policies aimed at protecting the public and, in particular, the developing fetus and women of reproductive age. Birth Defects Research (Part C) 108:224–242, 2016. © 2016 Wiley Periodicals, Inc.     

Clinical and public health research using methylated DNA Immunoprecipitation (MeDIP): A comparison of commercially available kits to examine differential DNA methylation across the genome

The methylated DNA immunoprecipitation method (MeDIP) is a genome-wide, high-resolution approach that detects DNA methylation with oligonucleotide tiling arrays or high throughput sequencing platforms. A simplified high-throughput MeDIP assay will enable translational research studies in clinics and populations, which will greatly enhance our understanding of the human methylome. We compared three commercial kits, MagMeDIP Kit TM (Diagenode), Methylated-DNA IP Kit (Zymo Research) and Methylamp? Methylated DNA Capture Kit (Epigentek), in order to identify which one has better reliability and sensitivity for genomic DNA enrichment. Each kit was used to enrich two samples, one from fresh tissue and one from a cell line, with two different DNA amounts. The enrichment efficiency of each kit was evaluated by agarose gel band intensity after Nco I digestion and by reaction yield of methylated DNA. A successful enrichment is expected to have a 1:4 to 10:1 conversion ratio and a yield of 80% or higher. We also evaluated the hybridization efficiency to genome-wide methylation arrays in a separate cohort of tissue samples. We observed that the MagMeDIP kit had the highest yield for the two DNA amounts and for both the tissue and cell line samples, as well as for the positive control. In addition, the DNA was successfully enriched from a 1:4 to 10:1 ratio. Therefore, the MagMeDIP kit is a useful research tool that will enable clinical and public health genome-wide DNA methylation studies.     

Effects of venlafaxine on the expression level and methylation status of genes involved in oxidative stress in rats exposed to a chronic mild stress

Recent human and animal studies indicate that oxidative and nitrosative stress may play a role in the aetiology and pathogenesis of depression. This study investigates the effect of chronic administration of the serotonin-norepinephrine reuptake inhibitor, venlafaxine, on the expression and methylation status of SOD1, SOD2, GPx1, GPx4, CAT, NOS1 and NOS2 in the brain and blood of rats exposed to a chronic mild stress (CMS) model of depression. Separate groups of animals were exposed to CMS for 2 or 7 weeks; the second group received saline or venlafaxine (10 mg/kg/d, IP) for 5 weeks. After completion of both stress conditions and drug administration, the mRNA and protein expression of selected genes and the methylation status of their promoters were measured in peripheral mononuclear blood cells (PBMCs) and in brain structures (hippocampus, amygdala, hypothalamus, midbrain, cortex, basal ganglia) with the use of TaqMan Gene Expression Assay, Western blot and methylation-sensitive high-resolution melting techniques. CMS caused a decrease in sucrose consumption, and this effect was normalized by fluoxetine. In PBMCs, SOD1, SOD2 and NOS2 mRNA expression changed only after venlafaxine administration. In brain, CAT, Gpx1, Gpx4 and NOS1 gene expression changed following CMS or venlafaxine exposure, most prominently in the hippocampus, midbrain and basal ganglia. CMS increased the methylation of the Gpx1 promoter in PBMCs, the second Gpx4 promoter in midbrain and basal ganglia, and SOD1 and SOD2 in hippocampus. The CMS animals treated with venlafaxine displayed a significantly higher CAT level in midbrain and cerebral cortex. CMS caused an elevation of Gpx4 in the hippocampus, which was lowered in cerebral cortex by venlafaxine. The results indicate that CMS and venlafaxine administration affect the methylation of promoters of genes involved in oxidative and nitrosative stress. They also indicate that peripheral and central tissue differ in their response to stress or antidepressant treatments. It is possible that that apart from DNA methylation, a crucial role of expression level of genes may be played by other forms of epigenetic regulation, such as histone modification or microRNA interference. These findings provide strong evidence for thesis that analysis of the level of mRNA and protein expression as well as the status of promoter methylation can help in understanding the pathomechanisms of mental diseases, including depression, and the mechanisms of action of drugs effective in their therapy.     

Gene expression and DNA methylation changes in the hypothalamus and hippocampus of adult rats developmentally exposed to bisphenol A or ethinyl estradiol: a CLARITY-BPA consortium study

Bisphenol A (BPA), an endocrine disrupting chemical (EDC), is a ubiquitous pollutant. As part of the Consortium Linking Academic and Regulatory Insights on BPA Toxicity (CLARITY-BPA), we sought to determine whether exposure of Sprague-Dawley rats to 2,500 μg/kg/day BPA (BPA) or 0.5 μg/kg/day ethinyl estradiol (EE) from gestational day 6 through postnatal day 21 induces behavior-relevant gene expression and DNA methylation changes in hippocampus and hypothalamus at adulthood. RNA and DNA were isolated from both regions. Expression of ten genes (Dnmt1, Dnmt3a, Dnmt3b, Esr1, Esr2, Avp, Ar, Oxt, Otr, and Bdnf) presumably altered by early-life BPA/EE exposure was examined. Three genes (Bdnf, Dnmt3b, and Esr1) were studied for DNA methylation changes in their putative 5? promoter regions. Molecular changes in hippocampus were correlated to prior Barnes maze performance, including sniffing correct holes, distance traveled, and velocity. Exposure to BPA and/or EE disrupted patterns of sexually dimorphic gene expression/promoter DNA methylation observed in hippocampus and hypothalamus of controls. In the hippocampus of female offspring, BPA exposure resulted in hypermethylation of the putative 5? promoter region of Bdnf, while EE exposure induced hypomethylation. Bdnf methylation was weakly associated with Bdnf expression in hippocampi of female rats. Hippocampal Bdnf expression in females showed a weak negative association with sniffing correct hole in Barnes maze. Hippocampal expression of Avp, Esr2, Oxt, and Otr was strongly associated with velocity of control rats in Barnes maze. Findings suggest BPA exposure induced non-EE-like gene expression and epigenetic changes in adult rat hippocampi, a region involved in spatial navigation.     

Promoter methylation status of VEGF receptor genes: A possible epigenetic biomarker to anticipate the efficacy of intracellular-acting VEGF-targeted drugs in cancer cells

We evaluated whether the inhibitory effects of vascular endothelial growth factor (VEGF)-targeted drugs on the proliferation of cancer cells differed according to VEGF receptor (VEGFR) genes, Flt1 and KDR, promoter methylation status. Five hyper-VEGFR-methylation and six no-VEGFR-methylation cancer cells were used for the present study, together with human umbilical endothelial cells (HUVECs) as a control. No-VEGFR-methylation cancer cells showed higher expression of Flt1 and KDR than hyper-VEGFR-methylation cancer cells. Hyper-VEGFR-methylation cancer cells only showed increased expression and protein levels of Flt1 and KDR after treatment with the demethylase 5-aza-2’-deoxycytidine. Two drugs (a VEGF-specific-antibody, bevacizumab, and a KDR-specific-antibody) targeting extracellular VEGF-VEGFR signaling and two VEGF-specific-tyrosine kinase inhibitors (PTK/ZK and sunitinib) targeting intracellular VEGFR signaling were used in the cell proliferation assay. HUVECs showed dose- and time-dependent proliferation decrease with all tested drugs over a 72 h incubation period. No- or hyper-VEGFR-methylation cancer cells showed no significant proliferation differences after treatment with VEGF-specific-antibody or VEGFR2-specific-antibody. After PTK/ZK or sunitinib treatment, no-VEGFR-methylation cancer cells showed dose- or time-dependent decreases in proliferation. Hyper-VEGFR-methylation cancer cells also showed proliferation inhibition by VEGF-specific-tyrosine kinase inhibitors after demethylation of Flt1 and KDR. Proliferation inhibition synergistically increased after combination of demethylation with PTK/ZK in hyper-VEGF-methylation cancer cells. We observed that intracellular targeting of VEGF-VEGFR signaling could be more effective than extracellular targeting of the pathway in the suppression of proliferation of some cancer cells. In particular, the efficacy of intracellular targeting of VEGF-specific-tyrosine kinase inhibitors might be influenced by the epigenetic alteration of VEGFRs.     

The Effect of Cellular Stress on T and B Cell Memory Pathways in Immunized and Unimmunized BALB/c Mice

Immunological memory is a fundamental function of vaccination. The antigenic breakdown products of the vaccine may not persist, and undefined tonic stimulation has been proposed to maintain the specific memory. We have suggested that cellular stress agents to which the immune cells are constantly exposed may be responsible for tonic stimulation. Here we have studied four stress agents: sodium arsenite, an oxidative agent; Gramicidin, eliciting K⁺ efflux and calcium influx; dithiocarbamate, a metal ionophore; and aluminum hydroxide (alum), an immunological adjuvant. The aims of this study are to extend these investigations to T and B cell responses of unimmunized and ovalbumin (OVA)-immunized BALB/c mice, and furthermore, to ascertain whether stress is involved in optimal expression of memory B cells, as demonstrated in CD4⁺ T cells. Examination of the homeostatic pathway defined by IL-15/IL-15R (IL-15 receptor) interaction and the inflammasome pathway defined by the IL-1-IL-1R interaction between dendritic cells (DC) and CD4⁺ T cells suggests that both pathways are involved in the development of optimal expression of CD4⁺CD45RO⁺ memory T cells in unimmunized and OVA-immunized BALB/c mice. Furthermore, significant direct correlation was found between CD4⁺CD44⁺ memory T cells and both IL-15 of the homeostatic and IL-1β of the inflammasome pathways. However, CD19⁺CD27⁺ memory B cells in vivo seem to utilize only the IL-15/IL-15R homeostatic pathway, although the proliferative responses are enhanced by the stress agents. Altogether, stress agents may up-regulate unimmunized and OVA-immunized CD4⁺CD44⁺ memory T cells by the homeostatic and inflammasome pathways. However, the CD19⁺CD27⁺ memory B cells utilize only the homeostatic pathway.     

ROS and pentose phosphate pathway: mathematical modelling of the metabolic regulation in response to xenobiotic-induced oxidative stress and the proposed Impact of the gluconate shunt

Abstract

Elevated intracellular levels of reactive oxygen species (ROS), e.g. resulting from exposure to xenobiotics, can cause severe damages. Antioxidant defence mechanisms, which involve regulation of enzyme activities, protect cells to a certain extent. Nevertheless, continuous or increased exposure can overwhelm this system resulting in an adverse cellular state. To simulate exposure scenarios and to investigate the transition to an adverse cellular state, a mathematical model for the dynamics of ROS in response to xenobiotic-induced oxidative stress has been developed. It is based on exposure experiments of human urothelial cells (RT4) to the nitrated polycyclic aromatic hydrocarbon 3-nitrobenzanthrone (3-NBA), a component of diesel engine exhaust, and takes into account the following metabolic pathways of the antioxidant defence system: glutathione redox cycle scavenging directly ROS, the pentose phosphate pathway and the gluconate shunt as NADPH supplier and the beginning of glycolysis. In addition, ROS generation due to the bioactivation of 3-NBA has been implemented. The regulation of enzyme activities plays an important role in the presented mathematical model. The in silico model consists of ordinary differential equations on the basis of enzyme kinetics and mass action for the metabolism of 3-NBA. Parameters are either estimated from performed in vitro experiments via least-squares fitting or obtained from the literature. The results underline the importance of the pentose phosphate pathway to cope with oxidative stress and suggest an important role of the gluconate shunt during low-dose exposure.     

The impact of climate change on the future distribution of priority crop wild relatives in Indonesia and implications for conservation planning

The analysis of climate change impact is essential to include in conservation planning of crop wild relatives (CWR) to provide the guideline for adequate long-term protection under unpredictable future environmental conditions. These resources play an important role in sustaining the future of food security, but the evidence shows that they are threatened by climate change. The current analyses show that five taxa were predicted to have contraction of more than 30 % of their current ranges: Artocarpus sepicanus (based on RCP 4.5 in both no dispersal and unlimited dispersal scenario and RCP 8.5 in no dispersal scenario by 2050), Ficus oleifolia (RCP 4.5 5 in both no dispersal and unlimited dispersal scenario by 2080), Cocos nucifera and Dioscorea alata (RCP 8.5 in both no dispersal and unlimited dispersal scenario by 2050), and Ficus chartacea (RCP 8.5 in both no dispersal and unlimited dispersal scenario by 2050 and 2080). It shows that the climate change impact is species-specific. Representative Concentration Pathways (RCP) of greenhouse gas (GHG) emission and dispersal scenarios influence the prediction models, and the actual future distribution range of species falls in between those scenarios. Climate refugia, holdout populations, and non-analogue community assemblages were identified based on the Protected Areas (PAs) network. PAs capacity is considered an important element in implementing a conservation strategy for the priority CWR. In areas where PAs are isolated and have less possibility to build corridors to connect each other, such as in Java, unlimited dispersal scenarios are unlikely to be achieved and assisted dispersal is suggested. The holdout populations should be the priority target for the ex situ collection. Therefore, by considering the climate refugia, PAs capacity and holdout populations, the goal of keeping high genetic variations for the long-term conservation of CWR in Indonesia can be achieved.     

Ovarian Stem Cells (OSCs) from the Cryopreserved Ovarian Cortex: A Potential for Neo-Oogenesis in Women with Cancer-Treatment Related Infertility: A Case Report and a Review of Literature

Cancer treatment related infertility (CTRI) affects more than one third of young women undergoing anti-cancer protocols, inducing a premature exhaustion of the ovarian reserve. In addition to ovarian suppression by GnRHa, oocyte and cortex cryopreservation has gained interest in patients with estrogen-sensitive tumors for whom the hormonal burst to prompt the multiple follicular growth could provide a further pro-life tumor pulsing. On the other hand, cortex reimplantation implies a few drawbacks due to the unknown consistency of the follicles to be reimplanted or the risk of reintroducing malignant cells. The capability of ovarian stem cells (OCSs) from fresh ovarian cortex fragments to differentiate in vitro to mature oocytes provides a tool to overcome these drawbacks. In fact, since ovarian cortex sampling and cryopreservation is practicable before gonadotoxic treatments, the recruitment of OSCs from defrosted fragments could provide a novel opportunity to verify their suitability to be expanded in vitro as oocyte like cells (OLCs). Here, we describe in very preliminary experiments the consistency of an OSC population from a single cryopreserved ovarian cortex after thawing as well as both their viability and their suitability to be further explored in their property to differentiate in OLCs, thus reinforcing interest in stemness studies in the treatment of female CTRI.     

A note on pre-spawning maternal cortisol exposure in farmed Atlantic salmon and its impact on the behaviour of offspring in response to a novel environment

This study was designed to assess whether pre-spawning maternal cortisol administration and an additional stressor, here modelled by mild hyperthermia experienced during incubation, affected the behavioural responses of farmed Atlantic salmon (Salmo salar) offspring when exposed to a novel environment. Mature females were injected prior to stripping with 0, 50 or 100 mg/kg cortisol. After fertilization the eggs were kept at incubation temperatures of either 8 or 10 °C. Four months after hatching, the fish were introduced to a novel environment, i.e. an aquarium (38 l). In each trial two individuals originating from different exposure groups were video recorded and their activity (number of turns, time spent not swimming, time spent at the bottom) and feeding behaviour were measured. Maternal cortisol increased time spent non-swimming and also time spent at the bottom, while cortisol and temperature together additionally affected number of turns. To the best of our knowledge this study is the first in fish to demonstrate a possible link between the maternal endocrinological state at spawning and the offspring's behavioural reactivity to novel situations.     

Modeling and experimental study on the anaerobic/aerobic/anoxic process for simultaneous nitrogen and phosphorus removal: The effect of acetate addition

A mathematical model based on the simulation software AQUASIM was developed to validate an anaerobic/aerobic/anoxic (AOA) process that enables simultaneous nitrogen and phosphorus removal in a single reactor by adding external organic carbon to preclude excess aerobic phosphate uptake by polyphosphate-accumulating organisms (PAOs) and provide phosphate for denitrifying PAOs (DNPAOs). Aerobic batch tests after anaerobic phosphate release with different chemical oxygen demand (COD) concentrations indicated that the effect of COD concentration on the phosphate uptake preclusion could be expressed by a simple formula. The reduction factor reflecting the formula, which retards the aerobic phosphate uptake in the presence of COD, was added to the process rates of aerobic polyphosphate storage and PAOs growth in the model. The improved model, which included the reduction factor, reasonably matched the experimental result regarding aerobic phosphate uptake behavior whereas the model without it did not; thus, the former precisely predicts the AOA process behavior.     

The effects of fatty acids consumption on OPG/RANKL/RANK system in cardiovascular diseases: Current status and future perspectives for the impact of diet-gene interaction

Cardiovascular disease (CVD) is an overall term that comprises a number of related pathologies, these include peripheral arterial disease, cerebrovascular disease, coronary heart disease (CHD), venous thromboembolism, and rheumatic and congenital heart diseases. Fatty acids in the diet have been reported to affect CVD. The OPG/RANKL/RANK system appears to have a role in CVD outcomes. However, there have been few studies on the impact of diet-gene interaction for effects of fatty acids consumption on the OPG/RANKL/RANK system in CVD. This review focuses on the effects of fatty acids on OPG/RANKL/RANK in CVD.     

Acidic and basic fibroblast growth factor bind with differing affinity to the same heparan sulfate proteoglycan on BALB/c 3T3 cells: Implications for potentiation of growth factor action by heparin

Heparan sulfate proteoglycans on the cell surface act as low affinity binding sites for acidic and basic fibroblast growth factor (FGF) [Moscatelli (1887): J Cell Physiol 131:123–130] and play an important role in the interaction of FGF with the FGF receptor (FGFR). In this study, several aspects of the interaction of FGFs with cell surface heparan sulfate proteoglycans were examined. Reciprocal cross blocking studies demonstrated that acidic FGF (aFGF) and basic FGF (bFGF) bind to identical or closely associated heparan sulfate motifs on BALB/c 3T3 cell surface heparan sulfate proteoglycans. However, the binding affinity of the two growth factros for these heparan sulfate proteoglycans differs considerably, competition binding data indicating that aFGF has a 4.7-fold lower affinity than bFGF for 3T3 heparan sulfate proteoglycan. Subsequent studies of dissociation kinetics demonstrated that bFGF dissociates form the FGFR at least 10-fold slower than aFGF, whereas, following removal of cell surface heparan sulfate proteoplycan. Subsequent studies of dissociation kinetic demonstrated that bFGF dissociates from the FGFR at least 10-fold slwer than aFGF, whereas, following removal of cell surface heparan sulfate proteoglycans by heparinase treatment, the dissociation rate of both FGFs is similar and rapid. These results support the concept that cell surface heparan sulfate proteoglycans stabilize the interactio fo FGF with FGFR, possibly by the formatin of a ternary complex. © Wiley-Liss, Inc.     

Mutations in amyloid precursor protein and presenilin-1 genes increase the basal oxidative stress in murine neuronal cells and lead to increased sensitivity to oxidative stress mediated by amyloid beta-peptide (1-42), HO and kainic acid: implications for Alzheimer's disease

Oxidative stress is observed in Alzheimer's disease (AD) brain, including protein oxidation and lipid peroxidation. One of the major pathological hallmarks of AD is the brain deposition of amyloid beta-peptide (Abeta). This 42-mer peptide is derived from the beta-amyloid precursor protein (APP) and is associated with oxidative stress in vitro and in vivo. Mutations in the PS-1 and APP genes, which increase production of the highly amyloidogenic amyloid beta-peptide (Abeta42), are the major causes of early onset familial AD. Several lines of evidence suggest that enhanced oxidative stress, inflammation, and apoptosis play important roles in the pathogenesis of AD. In the present study, primary neuronal cultures from knock-in mice expressing mutant human PS-1 and APP were compared with those from wild-type mice, in the presence or absence of various oxidizing agents, viz, Abeta(1-42), H2O2 and kainic acid (KA). APP/PS-1 double mutant neurons displayed a significant basal increase in oxidative stress as measured by protein oxidation, lipid peroxidation, and 3-nitrotyrosine when compared with the wild-type neurons (p < 0.0005). Elevated levels of human APP, PS-1 and Abeta(1-42) were found in APP/PS-1 cultures compared with wild-type neurons. APP/PS-1 double mutant neuron cultures exhibited increased vulnerability to oxidative stress, mitochondrial dysfunction and apoptosis induced by Abeta(1-42), H2O2 and KA compared with wild-type neuronal cultures. The results are consonant with the hypothesis that Abeta(1-42)-associated oxidative stress and increased vulnerability to oxidative stress may contribute significantly to neuronal apoptosis and death in familial early onset AD.     

The endothelium in health and disease: A discussion of the contribution of non-nitric oxide endothelium-derived vasoactive mediators to vascular homeostasis in normal vessels and in type II diabetes

Molecular and Cellular Biochemistry - Endothelial dysfunction is considered as a major risk factor of cardiovascular complications of type I and types II diabetes. Impaired endothelium-dependent...