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Environmental factors that have been linked to an increased endometriosis risk include exposure to di-(2-ethylhexyl)-phthalate (DEHP), an endocrine disruptor. This study aims to investigate whether DEHP in vitro exposure in primary endometrial stromal cells (EnSC), primary endometrial epithelial cells (EnEC), and the human endometrial adenocarcinoma cell line Ishikawa properly mimics alterations described in the eutopic endometrium of women with endometriosis. Primary EnSC and EnEC, isolated from six fertile egg donors, and Ishikawa cells were exposed to DEHP (0.1, 1, and 10 µM) and were assessed for viability, endometriosis markers (IL-6, VEGF-A, HOXA10, EZH2, and LSD1), steroid receptor gene expressions (ER-1, ER-2, PR-T, PR-B, and PGRMC1), and invasive capacity. Viability after 72 h of DEHP exposure was not significantly affected. None of the endometriosis markers studied were altered after acute DEHP exposure, nor was the expression of steroid receptors. The invasive capacity of EnSC was significantly increased after 10 µM of DEHP exposure. In conclusion, acute DEHP exposure in primary endometrial cells does not fully phenocopy the changes in the viability, expression of markers, or steroidal receptors described in endometriosis. However, the significant increase in EnSC invasiveness observed after DEHP exposure could be a link between DEHP exposure and increased endometriosis likelihood. [ABSTRACT FROM AUTHOR]

Objective:We aimed to investigate a possible role of the endocrine disruptors phthalates, di-2-ethylhexyl phthalate (DEHP) and mono (2-ethylhexyl) phthalate (MEHP), in polycystic ovary syndrome (PCOS) aetiopathogenesis. We also wished to evaluate the relationship between phthalates and metabolic disturbances in adolescents with PCOS.Methods:A total of 124 adolescents were included. Serum MEHP and DEHP levels were determined by high-performance liquid chromatography. Insulin resistance was evaluated using homeostasis model assessment-insulin resistance, quantitative Insulin Sensitivity Check Index, fasting glucose/insulin ratio, Matsuda index, and total insulin levels during oral glucose tolerance test. Participants were further subdivided into lean and obese subgroups according to body mass index (BMI).Results:Sixty-three PCOS and 61 controls, (mean age 15.2±1.5; range: 13-19 years) were enrolled. Serum DEHP and MEHP concentrations were not significantly different between PCOS and control groups. The mean (95% confidence interval) values of DEHP and MEHP were 2.62 (2.50-2.75) μg/mL vs 2.71 (2.52-2.90) μg/mL and 0.23 (0.19-0.29) μg/mL vs 0.36 (0.18-0.54) μg/mL in PCOS and the control groups respectively, p>0.05. Correlation analysis, adjusted for BMI, showed that both phthalates significantly correlated with insulin resistance indices and serum triglycerides in adolescents with PCOS.Conclusion:Serum DEHP and MEHP concentrations were not different between adolescents with or without PCOS. However, these phthalates are associated with metabolic disturbances such as dyslipidemia and insulin resistance, independently of obesity, in girls with PCOS.

Introduction: Esters of phthalates, particularly di-(2-ethylhexyl)-phthalate (DEHP) generally utilized in plastic industry are environmental hazardous pollutants, has toxic effects on body systems, including reproductive system. The aim of this study was to evaluate the effects of Achillea millefolium on rats’ sperm exposed to DEHP. Materials & Methods: In this study, 45 male rats were randomly divided into 9 groups. 1- Control, 2- sham, received corn oil, 3- received 75mg/kg/day hydro alcoholic extract of Achillea millefolium inflorescence, 4- Low-dose diethyl-hexyl–phthalate (DEHP1), 5- Low-dose diethyl-hexyl–phthalate+Achillea millefolium, (DEHP1+A) 6- Medium-dose diethyl-hexyl–phthalate (DEHP2), 7- Medium-dose diethyl-hexyl–phthalate + Achillea millefolium, (DEHP2+A) 8- High-dose diethyl-hexyl–phthalate (DEHP3), 9- High-dose diethyl-hexyl–phthalate + Achillea millefolium (DEHP3+A). All of groups except the control group were treated by gavage for 40 days. After euthanized, the epididymal sperms are collected to investigate the sperm parameters. Data were analyzed using ANOVA and Tukey post hoc tests. Findings: di-(2-ethylhexyl)-phthalate significantly (P

Exposure to the widely-used phthalate plasticizer di-(2-ethylhexyl)-phthalate (DEHP) has been shown to be closely related to an increased prevalence of allergic diseases in infants and juveniles. Earlier work in our laboratory found that DEHP-related anaphylactic responses could be ascribed to T-follicular helper (Tfh) cell hyperfunction directly. The Tfh cell, a newly identified CD4+ TH cell subset, until recently has been considered as a key player in humoral immunity. Tfh cells can respond to stimulation through various receptors. Signaling lymphocytic activation molecule family member-1 (SLAMF1, CD150) is a surface co-stimulatory receptor that can bind to an intracytoplasmic adaptor signaling lymphocytic activation molecule-associated protein (SAP) to initiate downstream signaling cascades, regulating some events of immune response. The present study explored the role of SLAMF1 in Tfh cell differentiation and cytokine secretion under the condition of DEHP exposure. Using a weanling mice model of DEHP gavage with ovalbumin (OVA) sensitization, it was found that DEHP acted as an immunoadjuvant to elevate SLAMF1 and SAP expression in host Tfh cells. Ex vivo studies of effects from DEHP exposure on Tfh cells from OVA-sensitized hosts showed that DEHP acted in an adjuvant-like manner to promote the expression of adaptor protein SAP, transcription factors Bcl-6 and c-MAF, and cytokines interleukin (IL)-21 and IL-4 in Tfh cells. Transfection of these Tfh cells with Slamf1 small interfering RNA prior to exposure to the DEHP attenuated the over-expression of these molecules that was caused by the DEHP. In conclusion, this study demonstrated that DEHP, via a SLAMF1-mediated pathway, can impact on Tfh cell differentiation and their ability to form select cytokines.

Objective: We aimed to investigate a possible role of the endocrine disruptors phthalates, di-2-ethylhexyl phthalate (DEHP) and mono (2-ethylhexyl) phthalate (MEHP), in polycystic ovary syndrome (PCOS) aetiopathogenesis. We also wished to evaluate the relationship between phthalates and metabolic disturbances in adolescents with PCOS. Methods: A total of 124 adolescents were included. Serum MEHP and DEHP levels were determined by high-performance liquid chromatography. Insulin resistance was evaluated using homeostasis model assessment-insulin resistance, quantitative Insulin Sensitivity Check Index, fasting glucose/insulin ratio, Matsuda index, and total insulin levels during oral glucose tolerance test. Participants were further subdivided into lean and obese subgroups according to body mass index (BMI). Results: Sixty-three PCOS and 61 controls, (mean age 15.2±1.5; range: 13-19 years) were enrolled. Serum DEHP and MEHP concentrations were not significantly different between PCOS and control groups. The mean (95% confidence interval) values of DEHP and MEHP were 2.62 (2.50-2.75) μg/mL vs 2.71 (2.52-2.90) μg/mL and 0.23 (0.19-0.29) μg/mL vs 0.36 (0.18-0.54) μg/mL in PCOS and the control groups respectively, p>0.05. Correlation analysis, adjusted for BMI, showed that both phthalates significantly correlated with insulin resistance indices and serum triglycerides in adolescents with PCOS. Conclusion: Serum DEHP and MEHP concentrations were not different between adolescents with or without PCOS. However, these phthalates are associated with metabolic disturbances such as dyslipidemia and insulin resistance, independently of obesity, in girls with PCOS. [ABSTRACT FROM AUTHOR]

The plasticizer di-(2-ethylhexyl)-phthalate (DEHP) is the most significant phthalate in production, usage, and environmental occurrence. DEHP is found in products such as personal care products, furniture materials, cosmetics, and medical devices. DEHP is noncovalently bind with plastic therefore, repeated uses lead to leaching out of it. Exposure to DEHP plasticizers leads to toxicity in essential organs of the body through various mechanisms. The main objective of this review article is to focus on the DEHP-induced endoplasmic reticulum (ER) stress pathway implicated in the testis, brain, lungs, kidney, heart, liver, and other organs. Not only ER stress, PPAR-related pathways, oxidative stress and inflammation, Ca2+ homeostasis disturbances in mitochondria are also identified as the relative mechanisms. ER is involved in various critical functions of the cell such as Protein synthesis, protein folding, calcium homeostasis, and lipid peroxidation but, DEHP exposure leads to augmentation of misfolded/unfolded protein. This review complies with various recently reported DEHP-induced toxicity studies and some pharmacological interventions that have been shown to be effective through ER stress pathway. DEHP exposure does assess health risks and vulnerability to populations across the globe. This study offers possible targets and approaches for addressing various DEHP-induced toxicity. [Display omitted] • Over 1000 articles emphasizing DEHP toxicity published during last 5 years. • DEHP is the most widely used plasticizer in the medical devices and PVC pipes. • ER stress pathway is the underlying mechanism that elicits DEHP toxicity. • Along with ER stress, other signaling pathways have also been reported. [ABSTRACT FROM AUTHOR]

Exposure to the widely-used phthalate plasticizer di-(2-ethylhexyl)-phthalate (DEHP) has been shown to be closely related to an increased prevalence of allergic diseases in infants and juveniles. Earlier work in our laboratory found that DEHP-related anaphylactic responses could be ascribed to T-follicular helper (Tfh) cell hyperfunction directly. The Tfh cell, a newly identified CD4+ TH cell subset, until recently has been considered as a key player in humoral immunity. Tfh cells can respond to stimulation through various receptors. Signaling lymphocytic activation molecule family member-1 (SLAMF1, CD150) is a surface co-stimulatory receptor that can bind to an intracytoplasmic adaptor signaling lymphocytic activation molecule-associated protein (SAP) to initiate downstream signaling cascades, regulating some events of immune response. The present study explored the role of SLAMF1 in Tfh cell differentiation and cytokine secretion under the condition of DEHP exposure. Using a weanling mice model of DEHP gavage with ovalbumin (OVA) sensitization, it was found that DEHP acted as an immunoadjuvant to elevate SLAMF1 and SAP expression in host Tfh cells. Ex vivo studies of effects from DEHP exposure on Tfh cells from OVA-sensitized hosts showed that DEHP acted in an adjuvant-like manner to promote the expression of adaptor protein SAP, transcription factors Bcl-6 and c-MAF, and cytokines interleukin (IL)-21 and IL-4 in Tfh cells. Transfection of these Tfh cells with Slamf1 small interfering RNA prior to exposure to the DEHP attenuated the over-expression of these molecules that was caused by the DEHP. In conclusion, this study demonstrated that DEHP, via a SLAMF1-mediated pathway, can impact on Tfh cell differentiation and their ability to form select cytokines. [ABSTRACT FROM AUTHOR]

Abstract Cross-sectional studies have described an association between exposure to phthalate esters and cardiovascular risk factors. However, the association with coronary heart disease (CHD) is still unclear. A total of 180 subjects randomly selected from 336 CHD patients, and 360 age- and sex-matched non-CHD controls were included from 2008 to 2011. Urinary metabolites of phthalate esters were measured by liquid chromatography–tandem mass spectrometry. The geometric means of urinary phthalates metabolites were significantly higher for the three Di-(2-ethylhexyl)-phthalate (DEHP) metabolites, mono‐2‐ethylhexyl phthalate (MEHP), mono‐(2‐ethyl‐5‐hydroxyhexyl) phthalate, and mono‐(2‐ethyl‐5‐oxohexyl) phthalate among CHD patients in-hospital than those of being discharged. Excluding 89 CHD patients of in-hospital and hospital discharge within 2 days, we found the urinary concentrations of MEHP, mono‐n‐butyl phthalate (MnBP), and mono-isobutyl phthalate (MiBP) of 91 CHD patients discharged ≥ 3 days were higher than those of controls. Among 451 participants, those with higher tertile levels of urinary MEHP, MnBP, and MiBP showed an increased risk for CHD compared to those with lowest tertile levels; the corresponding odds ratios (95% CI) were 2.77 (1.22–6.28), 2.90 (1.32–6.4), and 3.19 (1.41–7.21), respectively, after adjustment for confounders. Higher levels of hs-CRP, fibrinogen, and D-dimer were linked with increased levels of all DEHP metabolites in CHD patients. In conclusion, exposure to DEHP and dibutyl phthalates was positively associated with CHD and this relationship may be probably mediated via atherothrombosis. Graphical abstract Data was adjusted for age, gender, body mass index, diabetes mellitus, hypertension, and hypercholesterolemia, use of statins, smoking, and alcohol consumption according to tertiles of urinary phthalate levels. fx1 Highlights • Urinary MEHP, MiBP, and MnBP are associated with coronary heart disease (CHD). • DEHP metabolites are associated with hs-CRP, fibrinogen, and D-dimer in CHD patients. • Regulation to reduce phthalates exposure, particularly DEHP and DBP, is important. • The mechanism of phthalates exposure and CHD awaits further investigations. [ABSTRACT FROM AUTHOR]

Abstract: Accumulating evidence has suggested a link between maternal di‐(2‐ethylhexyl)‐phthalate (DEHP) exposure and various developmental abnormalities. However, the evidence regarding the effect of maternal DEHP exposure on fetal cardiac development is scarce. The present study aimed to determine the effect of maternal DEHP exposure on fetal cardiac development in mice and explore the possible involved mechanism preliminarily. The C57BL mice were randomly divided into four groups: the vehicle group (corn oil, n = 10), 250 mg kg–1 DEHP group (n = 15), 500 mg kg–1 DEHP group (n = 20) and 1 g kg–1 DEHP group (n = 20). Pregnant dams in different group received respective intervention by gavage once daily from embryonic day (E)6.5 to E14.5. Maternal weights were monitored every day and samples were collected at E15.5. Hematoxylin and eosin staining was used to examine fetal cardiac malformations. Real‐time quantitative polymerase chain reaction and western blot were applied to detect peroxisome proliferator‐activated receptor (PPAR)α/PPARγ/Nkx2.5/Gata4/Tbx5/Mef2c/Chf1 mRNA and protein expression, respectively. Maternal DEHP exposure significantly decreased maternal body weight, fetal weight and placental weight, and remarkably elevated fetal cardiac malformations rate. The phenotypes of cardiac anomalies mainly include septal defects, ventricular myocardium noncompaction and cardiac hypoplasia. Higher doses DEHP (500 mg kg–1 and 1 g kg–1) could significantly decreased fetal cardiac Gata4/Mef2c/Chf1 expression, while PPARγ expression was upregulated. Maternal exposure to higher doses of DEHP could result in fetal cardiac development malformations in mice and it might have resulted from the inhibition of cardiac GATA4/Mef2c/Chf1 expression via PPARγ activation. [ABSTRACT FROM AUTHOR]

As a plasticizer, di-(2-ethylhexyl)-phthalate (DEHP) is widely added in various commercial products. Some researchers had suggested that DEHP has adverse effects on the glucose metabolism, but the mechanisms remain unclear. Adolescent Wistar rats were divided into four groups and administered DEHP by gavage at 0, 5, 50, and 500 mg kg−1 d−1 for 28 days. ELISA was used to quantify the serum insulin and leptin levels; RT-PCR, immunohistochemistry, and Western blot were used to detect the mRNA and protein expressions of Janus-activated kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), suppressor of cytokine signaling 3 (SOCS3), leptin receptor (Ob-R), and insulin receptor (IR) in liver and pancreas In comparison to the control group, the DEHP-treated rats showed the following: (1) higher organ coefficient of liver; (2) higher fasting blood glucose levels, higher fasting serum insulin and leptin levels, higher insulin resistance index homeostasis model assessment; (3) lower protein levels of Ob-R and IR in the liver and pancreas; (4) higher protein levels of JAK2 and STAT3 in the liver; and (5) higher protein and mRNA levels of SOCS3 in the liver and pancreas. Exposure to DEHP induced glucose metabolic disorder in the adolescent rats, and the mechanism is that DEHP may interfere with the JAK2/STAT3/SOCS3 pathway, regulated the sensitivity of the insulin receptor and leptin receptor. [ABSTRACT FROM AUTHOR]

Final oocyte maturation is the key step to successful spawning and fertilization. Quantitative real-time PCR (qPCR) is the technique of election to quantify the abundance of functional genes in such study. Reference gene is essential for correct interpretation of qPCR data. However, an ideal universal reference gene that is stable under all experimental circumstances has not been described. Researchers should validate their reference genes while performing qPCR analysis. The expression of 6 candidate reference genes: 18s rRNA , 28s rRNA , Cathepsin Z , Elongation factor 1-α , Glyceraldehyde-3-phosphate dehydrogenase and β-actin were investigated during final oocyte maturation induced by different compounds (DES and DEHP) in common carp ( Cyprinus carpio ). Four softwares (Bestkeeper, geNorm, NormFinder and RefFinder) were used to screen the most stable gene in order to evaluate their expression stability. The results revealed that EF1α was highly stable expressed when final oocyte maturation was induced by DES, while gapdh was the most stable gene when final oocyte maturation was induced by DEHP. Stable expressed reference gene selection is critical for all qPCR analysis to get accurate target gene mRNA expression information. [ABSTRACT FROM AUTHOR]

The increasing incidence of male reproductive impairments has been associated with di-(2-ethylhexyl)-phthalate (DEHP) exposure. However, mechanisms involved are lacking. We exposed 4-week-old male C57BL/6j mice to DEHP by gavage at 0, 125, 250 or 500 mg/kg body weight/day for 28 consecutive days. Our data showed that pubertal exposure to DEHP induces sperm count reduction as well as histological abnormalities in seminiferous epithelium and apoptosis of post-meiotic germ cells, and these effects are concomitant with reduction of testosterone levels and its steroidogenic gene expression. Moreover, the expressions of estrogen receptor ERβ and nuclear receptors Nr0b1, Nr0b2 are increased. The expression of Nr5a2 which is the inducer of steroidogenesis is significantly reduced. Furthermore, spermatogonial stem cell (SSC) self-renewal, differentiation and meiosis were significantly impaired, and the epigenetic regulator G9a-mediated histone methylation was decreased following DEHP exposure. Our results suggest that the DEHP-induced male reproductive impairments may depend on its estrogenic action on estrogen receptor and nuclear receptor, and involve inhibition of steroidogenesis, SSC self-renewal and meiosis, which may be attributed to the down-regulation of G9a-mediated histone methylation. [ABSTRACT FROM AUTHOR]

The environmental risk of phthalic acid esters (PAEs) is of great concern. We investigated the reproductive impairment of di-(2-ethylhexyl)-phthalate (DEHP) on Chinese rare minnow, an endemic fish inhabiting the upper streams of the Yangtze River. Chinese rare minnow larvae were exposed to environmentally relevant concentrations of DEHP (0, 4.2, 13.3, and 40.8 μg/L) for 6 months. Plasma testosterone and 17β-estradiol levels decreased in females, accompanied by downregulation of cyp19a and cyp17 gene transcription in ovary. Increases in plasma testosterone concentration were observed in males, accompanied by downregulation of cyp19a gene transcription in testes. Hepatic VTG gene transcription was upregulated in males and females. Exposure to DEHP reduced egg production and inhibited oocyte maturation in females and retarded spermiation in males. Decreased egg protein content was measured in F1 embryos. These results indicate that long-term exposure to low concentrations of DEHP (13.3 μg/L) causes endocrine disruption and impairs fish reproduction. [ABSTRACT FROM AUTHOR]

Diethylhexyl phthalate (DEHP) and dibutyl phthalate (DBP) are the two most widely used plasticizers (PAEs) that may act as endocrine disruptors and cause developmental toxicity. We measured intrauterine exposure to DEHP and DBP which are the two most widely used phthalates (PAEs) in the cord blood of 50 postpartum women using ultra-HPLC-tandem mass spectrometry and solid-phase extraction. The embryotoxicity of DEHP and DBP was evaluated using the human embryonic stem cell test (hEST). Based on the intrauterine exposure concentration of DEHP and DBP in pregnant women and the reference point of toxic effects in hEST, we used the reference point index (RPI) method to assess the cumulative risk of DBP and DEHP. The mean concentrations of DEHP and DBP were 99.9 μg/L and 24.7 μg/L, respectively. DEHP and DBP were weakly embryotoxic, and the benchmark dose lower confidence intervals were 29.99 and 0.99 μg/mL, respectively, as determined using hEST. Both DEHP and DBP inhibited embryonic development via PPAR/PTEN/Akt signal pathway. The findings suggest that the cumulative risk in pregnant women with a high level of exposure should receive more attention in the future. • DEHP and DBP intrauterine co-exposure was very common in Chinese pregnant women. • DEHP and DPB were evaluated as weak embryotoxic chemicals by hEST. • DBP triggered PPARγ and induced apoptosis via PTEN/Akt pathway. • The cumulative risk assessment suggested more attention should be paid in pregnant women. [ABSTRACT FROM AUTHOR]

Di-(2-ethylhexyl)-phthalate (DEHP) is a potential hazard to human health. The effects of occupational high level DEHP exposure on human health were evaluated by measuring the plasma cholinesterase, residues, renal and hepatic biochemical markers. The study was conducted in three representative polyvinyl chloride manufacturing facilities from large size (S1), medium side (S2) to small size (S3). Total 456 adult males including 352 exposed workers (occupational) and 104 control workers (background) were selected. The average DEHP concentrations in respirable particulate matter were 233, 291, and 707 μg m −3 for S1–S3, respectively, compared with 0.26 μg m −3 in the background atmosphere (labeled by S4). The results showed significant decreases in post exposure plasma cholinesterase (PChE) levels (<30%) from the exposed workers as compared to baseline. These exposed workers had been evaluated for plasma DEHP residues. Regression analyses explored that PChE decreased significantly with increasing plasma DEHP residues. Serum aspartate aminotransferase, alanine aminotransferase, creatinine, urea, gamma glutamyltransferase, malondialdehyde, total antioxidant and C-reactive protein were significantly raised as compared to the controls. Of the 352 exposed workers, 116 (33.0%) had a daily DEHP intake 22.7 μg kg bw −1 d −1 , which is more than 20 μg kg bw −1 d −1 specified by the US Environmental Protection Agency. The study demonstrated that occupational phthalate exposure produces health hazards. [ABSTRACT FROM AUTHOR]

Both the detrimental effects of early life adversity and the beneficial effects of exercise on the hypothalamic-pituitary-adrenal (HPA) axis have been reported. Early life exposure to di-(2-ethylhexyl)-phthalate (DEHP) may impair the development of endocrine system. In this study, we investigated the effects of lactational DEHP exposure on stress responses in late adolescent female rats and examined the protective role of treadmill running. Sprague-Dawley dams were fed with DEHP (10 mg/kg per day) or vehicle during lactation. After weaning, the female offspring rats were trained to exercise on a treadmill for 5 weeks and then stressed by exploring on an elevated plus maze. The activities of HPA axis were evaluated by measuring the plasma levels of ACTH and corticosterone, the expressions of adrenal enzymes cholesterol side-chain cleavage enzyme (CYP11A1) and cytochrome P-450 11β-hydroxylase (CYP11B1), and the expression of hypothalamic glucocorticoid receptors (GR). The results demonstrate that DEHP-exposed rats exhibited enhanced anxiety-like behaviors. Increased hypothalamic GR and plasma ACTH levels, but decreased adrenal CYP11A1 and corticosterone levels, were observed in DEHP-exposed animals under stressed condition. Importantly, in DEHP-exposed animals, exercise during childhood-adolescence reduced anxiety-like behaviors by normalizing stress-induced alterations in ACTH level and adrenal CYP11A1 expression. The findings of this study suggest that treadmill running may provide beneficial effects on ameliorating the dysregulation of HPA axis in lactational DEHP-exposed adolescent female rats. [ABSTRACT FROM AUTHOR]

Di-(2-ethylhexyl)-phthalate (DEHP) is present in a wide variety of medical devices. Infants are more susceptible than the general population.To investigate histological and immnohistochemical changes in the lung of newborn rats in response to mothers’ exposure.Twenty nonpregnant adult female albino rats were used. After pregnancy, on the 15th day of pregnancy, dams were divided into two equal groups: the control group received corn oil and the treated group received DEHP (500 mg/kg/day) from the 15th day of pregnancy and during the 1st three weeks of lactation. Fifteen offspring from each group were subdivided into three subgroups, 1, 2, and 3, which were sacrificed at the 2nd, 7th, and 21st postnatal day, respectively. Lung specimens were processed for light microscope (H&E and immunohistochemical staining of proliferating cell nuclear antigen) and electron microscope examination.H&E-stained sections of treated lung on the 2nd day showed variable-sized saccules. Primary septa were significantly thicker than those of the control group. However, on the 7th day, focally dilated saccules with thin primary septa were observed. Saccules and secondary septa were fewer than those in the control group. On the 21st day, variable-sized alveoli and a significant increase in septal thickness were observed. Immunohistochemically, there was a significant increase in proliferating cell nuclear antigen-positive cells on both the 2nd day and the 21st day, whereas they decreased on the 7th day in comparison with the control group. Ultrastructurally, treated lung on the 2nd day showed a saccular wall with many layers of interstitial cells, some with shrunken nuclei, whereas on the 7th day, thin septa were observed. Type II pneumocytes and interstitial cells had heterochromatic shrunken nuclei, whereas on the 21st day, the presence of macrophages was observed in the lumen. Interstitials cells and type II pneumocytes showed heterochromatic nuclei and marked distortion of organelles.DEHP induced marked histological and immunohistochemical changes in the lung. Alternative plasticizers with less adverse impact could be used. Further studies on the possible protective effect of antioxidants are recommended. [ABSTRACT FROM AUTHOR]

Abstract: In the last years the concern about the negative effects of phthalates on reproduction significantly increased. Considering that, at date data available dealing with the adverse outcome of Di-(2-ethylhexyl)-phthalate (DEHP) on the reproduction of several species are still contrasting, in this study, the effects induced by DEHP (0.05, 0.1, 1, 10 and 100nM) and its active metabolite, phthalic acid (PA) (0.01, 0.1, 1 and 10μM), were analyzed in zebrafish, Danio rerio, primary hepatocyte cultures, using target molecules involved in fish reproduction (vitellogenin – vtg and estrogen receptors – ERα, β1 and β2) and metabolism (peroxisome proliferators activated receptors – PPAR α, β, γ). The use of in vitro culture, in fact, has the potential to significantly reduce the number of animals sacrificed for research allowing a precise control of the physical and chemical parameters that is often not possible in vivo. Moreover, since many toxicological studies revealed a sex specific response to toxicants, male and female primary hepatocyte cultures were set up to elucidate the possible gender specific effects of two common environmental phthalates. The increase of vtg levels observed in the culture media of male or female hepatocytes strongly evidenced the phthalates E2-like action. Moreover, the data obtained suggested that the observed different ERs isoforms modulation is otherwise associated with the vtg increase, depending on fish gender. Regarding PPARs, a similar trend of expression was found in both males and females. In conclusion, this study enforces the role of vtg as biomarker for evaluate the presence of environmental doses of DEHP and PA. Considering the similar gender modulation observed for vtg and PPARs, these molecules could be used for the rapid screening of the presence of DEHP and PA. Noteworthy the gender specific modulation observed for ERs opens a debate on the estrogenic mechanism of action of DEHP and PA and their role on vtg induction. [Copyright &y& Elsevier]

Abstract: A major question is whether exposure to mixtures of low-dose endocrine disruptors (EDs) having different action mechanisms affects neurodevelopment differently than exposure to EDs individually. We therefore investigated the effects of fetal and neonatal exposure to three typical EDs – bisphenol A (BPA), di-(2-ethylhexyl)-phthalate (DEHP), and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) – on the midbrain dopaminergic system associated with functions – including motor activity, emotion, and cognition – affected by neuropsychiatric diseases such as attention-deficit/hyperactivity disorder. ICR mouse dams and their pups were orally treated with BPA (5mg/(kgday)), DEHP (1mg/(kgday)), or TCDD (8ng/kg) individually, or with mixtures thereof, to compare the effects between sole and mixed administration. We analyzed tyrosine hydroxylase (TH)- and Fos-immunoreactive (ir) neurons as markers of dopamine and neuronal activation, respectively. The numbers of TH- and/or Fos-ir neurons and the intensity of TH-immunoreactivity within midbrain dopaminergic nuclei (A9, A10, and A8) of each sole administration group significantly differed from controls at 2, 4, and 6 weeks of age. In contrast, no significant differences were detected in the mixture groups, suggesting counteractions among those chemicals. These results indicate that ED mixtures as pollution have unique and elusive effects. Thyroid hormones and/or aryl hydrocarbon receptor-related mechanisms may be responsible for this counteraction. [Copyright &y& Elsevier]

Abstract: Nutrient availability is one of the major non-genetic factors determining embryonic growth and larval or fetal size. Due to the high human consumption of blood lipid regulators, fibrates have recently been reported as pollutants in rivers. Our study investigated the developmental toxicity of fibrates in zebrafish. Treatment with micromolar concentrations of clofibrate or gemfibrozil induced an embryonic malabsorption syndrome (EMS) with very little yolk consumption, resulting in small-sized larvae. This effect was reversible on removing the drug from the water. Clofibrate delayed hatching time and decreased the amount of oil red O lipid staining in the vasculature. It also induced higher density, round-shaped neuromuscular junctions associated with disorganization and less striation of muscular fibers, and pericardial edema, as well as impairing thyroid gland morphogenesis. acox1, apoa1 and mtp hybridization transcript signals were not affected in the yolk syncytial layer (YSL) after clofibrate exposure. Di-(2-ethylhexyl)-phthalate did not slow down yolk resorption, whereas brefeldin A induced EMS. These findings suggest that the inhibition of yolk sac resorption on exposure to fibrate is not at a pre-translational level or peroxisome proliferator-activated receptor alpha dependent and may be due to an inhibition of the YSL constitutive cell secretion. The effects of fibrates and the potential bioconcentration in eggs as well as the additive action of structurally related toxicants warrant an evaluation of the developmental impact of these compounds after long-term exposure at environmentally relevant concentrations. Fibrate-induced EMS in zebrafish seems useful for studying the morphogenetic consequences of impaired nutrient availability during the early stages of vertebrate development. [Copyright &y& Elsevier]

Abstract: The plasticizer di-(2-ethylhexyl)-phthalate (DEHP) is the most important phthalate with respect to its production, use and occurrence in the environment. In standard carcinogenicity experiments with F344 rats and B6C3F1 mice, DEHP has been shown to induce hepatocellular tumors. Moreover, DEHP is strongly suspected to be a developmental and reproductive toxicant. The present study aimed at determining the long-term toxic effects of lifetime exposure to low concentrations of DEHP in Sprague–Dawley rat strain. Seven hundred and thirty male rats, stratified into four groups, received DEHP with the diet, resulting in dosages of 300, 95, 30 and 0mg/kg per day for up to 159 weeks and were only sacrificed when moribund. All organs of the dead and sacrificed animals were histopathologically examined. Significantly increased tumor incidences after exposure to 300mg/kg per day DEHP (P = 0.04 for testes and 0.05 for liver) and a significant dose-related trend (PTrend = 0.02 for testes and 0.03 for liver) were detected in both organs liver and testes. Time to tumor analysis revealed that DEHP-induced testicular tumors developed earlier in lifetime than hepatocellular neoplasias, and their multiplicity increased with time. In addition, animals exposed to the highest DEHP dose showed a significantly increased rate of testicular tubular atrophy (P < 0.01). In conclusion, this study shows for the first time that the rat testes are a target organ of DEHP carcinogenicity in Sprague–Dawley rats upon lifetime exposure. This new finding indicates the importance of evaluating the effects of lifetime exposure in assessing the potential human health risks of DEHP. In addition, the carcinogenicity should be evaluated in rat strains with low spontaneous tumor incidence in the organs known as target of DEHP toxicity. [Copyright &y& Elsevier]

Since the 1930s, plasticizers have been used to impart flexibility to an otherwise rigid polyvinylchloride (PVC). Di-(2-ethylhexyl)-phthalate (DEHP) is the most widely used plasticizer in PVC formulations. However, DEHP leaks out from PVC items with time and use and consequently it is an ubiquitous environmental contaminant. To date, very little is known about the actual extent of DEHP exposure in infants, who are believed to be the most sensitive population, as they may be exposed to several different sources (breast milk, infant formula, baby food, indoor air, and by dermal and oral exposure via indoor dust containing DEHP) since early in life and since differences in DEHP pharmacokinetics and metabolism between children and adults have been well documented. Little information exists on DEHP dietary exposure, believed to represent a major source and orally administered DEHP is converted more rapidly to MEHP than other routes of exposure. Although DEHP has been shown to induce toxicity in experimental animals, a limited but suggestive human exposure data causes a serious concern that an early in life DEHP exposure may adversely affect male reproductive tract development. Here, we report a review on dietary phthalate exposure in babies. [Copyright &y& Elsevier]

Numerous epidemiological findings have shown that di-(2-ethylhexyl)-phthalate (DEHP), one of industrial plasticizers with endocrine-disrupting properties, positively contributes to high incidence of obesity. However, potential pathogenesis of dietary DEHP exposure-induced obesity remains largely unknown. Chronic DEHP exposure at different doses (0.05 and 5 mg/kg body weight) to mice had been continuously lasted for 14 weeks through the diet. A combination of targeted quantitative metabolomics (LC/GC-MS) with global 1H NMR-based metabolic profiling to explore the effects of dietary DEHP exposure with different doses on host lipid metabolism of mice. Metagenomics (16S rRNA gene sequencing) was also employed to examine the alterations of gut microbiota composition in the cecal contents of mice after dietary DEHP exposure. Dietary exposure to DEHP at both doses induced weight gain and hepatic lipogenesis of mice by promoting the uptake of fatty acids and disrupting phospholipids and choline metabolism. Dietary DEHP exposure altered the gut microbiota community with disruption of intestinal morphology and reduction of Firmicutes to Bacteroidetes ratio in the cecal contents of mice. Furthermore, DEHP exposure activated gut microbiota fermentation process producing excess short chain fatty acids of mice. These findings provide systematic evidence that long-term chronic DEHP exposure induces obesity through disruption of host lipid metabolism and gut microbiota in mice, which not only confirm the epidemiological results, but also expand our understanding of metabolic diseases caused by environmental pollutants exposure. [Display omitted] • Dietary DEHP induced body weight gain, lipid and triglyceride accumulation of mice. • Dietary DEHP induced abnormal metabolism of phospholipids and choline of mice. • DEHP disrupted microbial morphology and composition with promotion of fermentation. [ABSTRACT FROM AUTHOR]

1 Di-(2-ethylhexyl)-phthalate (DEHP) possesses a great industrial value as a plasticizing agent and has become an ubiquitous environmental contaminant. In most species it is rapidly metabolized to mono-(2-ethylhexyl)-phthalate (MEHP) and 2-ethylhexanoic acid (2-EHA). Evaluation of toxicity of DEHP and its primary metabolites has been focussed on reproductive toxicity and hepatocarcinogenic properties. The aim of this study was to determine the nephrotoxic potential of both DEHP metabolites by use of cultured kidney epithelial cells (Opossum kidney cells; OK cells). 2 For this purpose, OK cells were exposed for 3 days to MEHP and 2-EHA at concentrations ranging from 0.1 – 500 μmol/L and the toxicity as well as the effects on migratory activity and intracellular cytoskeleton were studied by cell biological, morphological and morphometric methods. 3 When compared with corresponding controls, treatment of OK cells with MEHP and 2-EHA, respectively, showed marked differences in cell viability between both DEHP metabolites. MEHP caused a dose-dependent decrease in cell viability (ED[sub 50] = 25 μmol/L) accompanied by a moderate swelling of the cells at concentrations up to 25 μmol/L. MEHP concentrations higher than 25 μmol/L caused a dose-dependent shrinkage of the cells and the occurrence of a high amount of cell debris as a result of cell lysis. 2-EHA did not cause a reduced viability or an altered cell volume. The migratory activity of OK cells was not significantly influenced by both metabolites. Moreover, MEHP toxicity resulted in a largely reduced and altered organization of F-actin (stress fibers), but not of myosin, microtubules and vimentin. 4 The study indicates that cultured epithelial cells can be used as a prescreening system to assess the nephrotoxicity of hazardous substances such as DEHP. As demonstrated in this study, only MEHP, but not 2-EHA, has a marked nephrotoxic effect in vitro. [ABSTRACT FROM AUTHOR]

Both the detrimental effect of prenatal exposure to di-(2-ethylhexyl)-phthalate (DEHP) and the beneficial effects of physical exercise on brain functions have been reported. The oxytocin pathway has been implicated in the onset of maternal behaviors. Epigenetic modification of the oxytocin receptor gene (OXTR) through DNA methylation has been associated with the pathogenesis of neuropsychiatric disorders. The purpose of this study was to investigate the effects of prenatal DEHP exposure on oxytocin-regulated maternal behaviors and to examine the protective effect of exercise. Pregnant rats (F0) were fed with vehicle or DEHP during gestation and the offspring females (F1) were assessed for their maternal behaviors by pup retrieval test at postpartum. The results showed that reduced pup retrieval activities without significant alteration of stress responses were observed in the prenatally DEHP-exposed females. Prenatal DEHP exposure decreased the expressions of oxytocin, Oxtr mRNA, and oxytocin receptor, and increased Oxtr methylation in the hypothalamus of postpartum female rats. There were no significant effects of exercise on behavioral, biochemical, and epigenetic measurements. These results suggest that prenatal DEHP exposure has a long-term adverse effect on maternal behaviors; Oxtr hyper-methylation may be a potential epigenetic mechanism for this alteration, which cannot be prevented by physical exercise during childhood. [ABSTRACT FROM AUTHOR]

• Prenatal DEHP exposure impairs spatial learning and memory in the late-adolescent male rats. • Down-regulations of BDNF, NMDA receptor, Arc, and synaptophysin are found in the DEHP-exposed male rats. • Exercise prevents the impairment of learning and memory in prenatally DEHP exposed male rats. • Exercise improves the hippocampal expressions of plasticity-related proteins in the DEHP-exposed male rats. Regular exercise has been identified to facilitate neuroplasticity that maximize functional outcome after brain injuries. Brain-derived neurotrophic factor (BDNF) has emerged as a key facilitator of neuroplasticity after exercise. The activity-regulated cytoskeleton associated protein (Arc) is induced by BDNF and N-methyl- d -aspartic acid receptor (NMDAR), contributing to functional modification of neuroplasticity in the hippocampus. Meanwhile, early-life exposure to neuroendocrine disruptor di-(2-ethylhexyl)-phthalate (DEHP) is a risk factor for behavioral deficits, but the mechanisms responsible for DEHP-induced neurotoxicity are not well understood. The purpose of this study is to investigate whether hippocampal Arc expression is impaired by DEHP exposure and to examine the protective role of exercise in the prenatally DEHP-exposed male rats. Sprague Dawley dams were fed with vehicle or DEHP during gestation. The male offspring were trained to treadmill running for 5 weeks followed by examination of behavioral and biochemical outcomes. The results showed that DEHP-exposed rats exhibited impairment of spatial learning and memory as well as down-regulations of BDNF, NMDAR, Arc, and synaptophysin. Importantly, aerobic exercise during childhood-adolescence prevented the impairment of learning and memory by recovering the expressions of BDNF, NMDAR, Arc, and synaptophysin. These findings suggest that exercise may provide beneficial effects on ameliorating the impairment of neuroplasticity in the prenatally DEHP-exposed male rats at late adolescence. [ABSTRACT FROM AUTHOR]

Abstract: Irradiation with 20–25kGy is a process commonly used for sterilizing poly(vinyl chloride) (PVC) medical devices. Moreover, whole blood and blood components undergo additional irradiation with 25–50Gy to inhibit the proliferative capacity of lymphocytes and reduce the risk of transfusion-associated graft-vs-host disease (GVHD). Di-(2-ethylhexyl)-phthalate (DEHP) plasticized PVC is extensively used for the production of flexible medical devices including blood bags, but since DEHP is not covalently bound to PVC, it tends to migrate and leach out of the medical device, with harmful consequences for the patients. In this study, the effects of different doses of gamma irradiation on DEHP migration from PVC blood bags was investigated using differential scanning calorimetry (DSC) analysis. Our findings indicate that irradiation with 25–100Gy reduces the ability of DEHP to migrate from the blood bags, and in the case of a primary container a correlation between the doses of gamma ray irradiation was also observed. In particular, a decrease in DEHP leachability was obtained by increasing the dose of gamma ray irradiation. [Copyright &y& Elsevier]

Early-life exposure to di-(2-ethylhexyl)-phthalate (DEHP) has been suggested to relate to hyperactivity, lack of attention, and working memory deficits in school-age children. Brain-derived neurotrophic factor (BDNF) and endocannabinoids are induced by aerobic exercises to provide beneficial effects on brain functions. This study investigated the mechanisms underlying working memory impairment and the protective role of exercise in prenatal DEHP-exposed male rats. Sprague Dawley dams were fed with vehicle or DEHP during gestation. The male offspring were trained to exercise on a treadmill for 5 weeks, which was followed by an assessment of their working memory with a T-maze delayed non-match-to-sample task. The expressions of BDNF, dopamine D1 receptor (D1R), cannabinoid receptor 1 (CB1R), and fatty acid amide hydrolase (FAAH) in the prefrontal cortex were detected by Western blot. The results showed that DEHP-exposed rats exhibited working memory impairments without significant alterations in locomotor activities. The reduced expressions of prefrontal BDNF and CB1R were obtained in the DEHP-exposed rats, while D1R and FAAH were barely affected. Importantly, aerobic exercise during childhood-adolescence prevented the impairment of working memory in the DEHP-exposed rats by recovering the BDNF and CB1R expressions in the prefrontal cortex. These findings suggest that exercise may provide beneficial effects in ameliorating the impairment of working memory in the prenatal DEHP-exposed male rats at late adolescence. [ABSTRACT FROM AUTHOR]

Objective: To investigate the relationship between di-(2-ethyl hexyl) phthalate (DEHP) and male infertility by detecting the concentration of DEHP in the seminal plasma of the patient with idiopathic asthenozoospermia (IAS)., Methods: This study included 45 infertile males with diagnosed IAS in the observation group and another 45 men with normal sperm parameters as controls. We obtained the general baseline data on the subjects, determined the concentration of DEHP in the seminal plasma, the ROS level and the sperm DNA fragmentation index (DFI), and compared them between the two groups of males., Results: There were no statistically significant differences between the two groups of subjects in age, living habits and other general in baseline data (P > 0.05). The IAS patients, in comparison with the normal controls, showed significantly increased DEHP concentration in the seminal plasma (［0.45 ± 0.09］ vs ［0.23 ± 0.05］ μg/ml, P < 0.05), ROS level (［569.4 ± 45.3］ vs ［317.6 ± 27.8］ pmol/106 sperm, P < 0.05) and sperm DFI (［22.1 ± 8.3］% vs ［10.5 ± 6.7］%, P < 0.05). The concentration of DEHP in the seminal plasma was correlated positively with the ROS level (r = 0.77, P < 0.05) and sperm DFI (r = 0.75, P < 0.05) but negatively with the percentage of progressively motile sperm (r = －0.81, P < 0.05)., Conclusions: The DEHP level is escalated in the seminal plasma of the IAS patient, which may be responsible for the reduced sperm motility and increased DFI of the patient.

Di-(2-ethylhexyl)-phthalate (DEHP) accumulates in the environment, and its exposure is possibly associated with endocrine-related disease in women of reproductive age. The effects of DEHP on human endometrial cells are unknown. We treated human endometrial stromal cells with 10, 100, and 1000 pmol of DEHP and measured reactive oxygen species (ROS) generation, expression levels of antioxidant enzymes, alteration of MAPK/NF-κB signaling and hormonal receptors. DEHP increased reactive oxygen species (ROS) generation and decreased expression of superoxide dismutase (SOD), glutathione peroxidase (GPX), heme oxygenase (HO), and catalase (CAT). By DEHP exposure, p-ERK/p-p38 and NF-κB mediated transcription was increased. Additionally, DEHP induced estrogen receptor-α (ER-α) expression in a dose-dependent manner. This study shows the need for future mechanistic studies of oxidative stress, MAPK/NF-κB signaling, and ER-α as molecular mediators of DEHP-associated endometrial stromal cell alterations, which may be associated with the development of endocrine-related disease such as endometriosis., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Concern has increased regarding the adverse effects of di-(2-ethylhexyl)-phthalate (DEHP) on reproduction. However, limited information is available on the effects of DEHP in marine organisms. The aim of the present study was to examine whether long-term exposure to DEHP and its active metabolite mono-(2-ethylhexyl)-phthalate (MEHP) disrupts endocrine function in marine medaka (Oryzias melastigma). Marine medaka larvae were exposed to either DEHP (0.1 and 0.5mg/L) or MEHP (0.1 and 0.5mg/L) for 6 months, and the effects on reproduction, sex steroid hormones, liver vitellogenin (VTG), gonad histology and the expression of genes involved in the hypothalamic-pituitary-gonad (HPG) axis were investigated. Exposure to DEHP, but not MEHP, from hatching to adulthood accelerated the start of spawning and decreased the egg production of exposed females. Moreover, exposure to both DEHP and MEHP resulted in a reduction in the fertilization rate of oocytes spawned by untreated females paired with treated males. A significant increase in plasma 17β-estradiol (E2) along with a significant decrease in testosterone (T)/E2 ratios was observed in males, which was accompanied by the upregulation of ldlr, star, cyp17a1, 17βhsd, and cyp19a transcription in the testis. Increased concentrations of T and E2 were observed in females, which was consistent with the upregulation of ldlr. The expression of brain gnrhr2, fshβ, cyp19b and steroid hormone receptor genes also corresponded well with hormonal and reproductive changes. The liver VTG level was significantly increased after DEHP and MEHP exposure in males. DEHP induced histological changes in the testes and ovaries: the testes displayed a reduced number of spermatozoa, and the ovaries displayed an increased number of atretic follicles. In addition, the tissue concentrations of MEHP, MEHHP and MEOHP in DEHP-exposed groups were much higher than those in MEHP-exposed groups, and there were no dose- or sex-specific effects. Thus, DEHP exerts more obvious toxic effects compared with MEHP. There were some commonalities in the toxic effects and molecular mechanisms of DEHP and MEHP, suggesting that some of the toxic effects of DEHP may be induced by both DEHP itself and DEHP metabolites (including MEHP). Taken together, these results indicate that exposure to DEHP and MEHP from hatching to adulthood causes endocrine disruption with sex-specific effects in marine medaka, with males being more sensitive than females., (Crown Copyright © 2013. Published by Elsevier B.V. All rights reserved.)