Morphological alterations in human skeletal muscle cells exposed to di-(2-Ethylhexyl) Phthalate (dehp) in primary cell culture conditions / Alteraciones morfológicas en células musculares esqueléticas humanas expuestas a Ftalato de di-(2-etilhexilo) (dehp) en condiciones de cultivo celular primario

SUMMARY: Di-(2-ethylhexyl) phthalate (DEHP) is among the most common plasticizer additives that humans are in contact with daily. DEHP can be released from plastic and enter the human body, whereby it is metabolized and transformed into oxidative hydrophilic molecules. Clinical follow-ups in patients exposed to this phthalate and investigations in cultures of several cell types have provided information on its effects. For example, it is associated with inhibition of diploid human cell development and morphological changes in cultured germ cells. Although skeletal muscle represents around 50 % of the human body mass, knowledge about the effects of DEHP on this tissue is poor. Cultured skeletal muscle cells were exposed to DEHP (1 mM) for 13 days with the aim of exploring and evaluating some of the potential morphological effects. Three culture development parameters and nine cell characteristics were monitored during the bioassay. At 13 days, growth area, cell viability, and concentration of total proteins were lower in DEHP exposed than in control cells. Cell width and area, as well as the diameter of the nucleus and nucleolus, were greater in exposed cells than in control cells. These are interpreted as signs of cytotoxicity and suggest potential adverse effects on the development of skeletal muscle cells from DEHP exposure, as reported for other cell types.

RESUMEN: Diariamente los seres humanos tenemos contacto con aditivos plastificantes, el di-(2-etilhexil) ftalato (DEHP) se encuentra entre los más comunes. El DEHP puede liberarse del plástico e ingresar al cuerpo humano, donde es metabolizado y transformando en moléculas hidrofílicas oxidativas. Seguimientos en pacientes expuestos a este ftalato e investigaciones en cultivos de varios tipos celulares han aportado información sobre sus efectos. El DEHP es asociado con la inhibición del desarrollo de células humanas diploides y cambios morfológicos en células germinales en cultivo. Sin embargo, aún es poco lo que se sabe sobre los efectos en el músculo esquelético, a pesar de que este tejido representa alrededor del 50 % de la masa corporal del humano. Para explorar y evaluar algunos efectos morfológicos en células de músculo esquelético, cultivos primarios fueron expuestos a DEHP (1 mM) durante 13 días. Se dio seguimiento a tres parámetros de desarrollo del cultivo y nueve características celulares. Al término de 13 días de exposición, los valores del área de crecimiento, viabilidad celular y concentración de proteínas totales fueron inferiores con respecto a los cultivos control. Se observaron cambios morfométricos en las células expuestas. Particularmente, el ancho y área celular, así como los diámetros del núcleo y nucleolos, fueron mayores a los registros en las células control. Estos resultados se interpretan como signos de citotoxicidad y sugieren efectos potencialmente adversos en el desarrollo de las células del músculo esquelético ante una exposición al DEHP, como se ha registrado para otros tipos celulares.

Effects of Shoutai pills on immune function and oxidative stress in pregnant rats with di(2-ethylhexyl) phthalate exposure / 南方医科大学学报

OBJECTIVE@#To investigate the effects of Shoutai pills (a traditional Chinese medicinal preparation) on immune functions and oxidative stress in pregnant rats exposed to di(2-ethylhexyl) phthalate (DEHP).@*METHODS@#Thirty-six mature female SD rats were randomly divided into 3 groups (=12). After pregnancy was confirmed, the rats were given 10 mL/kg corn oil +10 mL/kg saline (control group), 500 mg/kg DEHP+10 mL/kg saline (model group), and 500 mg/kg DEHP+10 mL/kg Shoutai pills (treatment group). At 19 days of gestation, the rats were sacrificed and the fetal rats were weighed and the numbers of live and stillborn fetal rats were recorded. Serum levels of interleukin-6 (IL-6), interleukin-2 (IL-2), tumor necrosis factor-ɑ (TNF-ɑ), estradiol (E2) and progesterone (P) levels were detected. The appearance, color and quality of the placenta in each group were recorded, and the placental tissues were examined pathologically. The total antioxidant capacity (T-AOC), superoxide dismutase (SOD), glutathione peroxidase (GSH- Px), catalase (CAT), reactive oxygen species (ROS) and malondialdehyde (MDA) in the placental tissues were measured.@*RESULTS@#Compared with the control group, the rats with DEHP exposure showed slow weight gain in the middle and late gestation period and significantly lower fetal weight ( < 0.05) with lowered serum levels of IL-2, IL-6 and TNF-ɑ, increased estradiol level ( < 0.05), decreased placental T-AOC, GSH-Px, SOD and CAT levels, and increased ROS and MDA levels ( < 0.01). Compared with the model group, the rats treated with Shoutai pills had significantly increased weight gain in mid and late pregnancy and greater fetal weight ( < 0.05) with significantly increased serum IL-2 and IL-6 levels, decreased estradiol level ( < 0.05), slightly increased TNF-ɑ expression (> 0.05), increased placenta T-AOC, GSH- Px and CAT levels, decreased MDA level ( < 0.05), and slightly increased SOD and decreased ROS levels (>0.05). No significant difference was found in progesterone levels among the groups (>0.05). HE staining showed that the trophoblast in the placental tissue sponge in the model group was loose and irregular with numerous vacuoles. In the treatment group, the structure of the placenta remained intact with clearly visible labyrinth zone, sponge trophoblast and giant cell trophoblast, and the cell distribution in each layer was better than that in the model group.@*CONCLUSIONS@#Shoutai pills can regulate the immune function of DEHP-exposed pregnant rats possibly by antagonizing the estrogenlike effect of DEHP and regulating serum immune factors; Shoutai pills can also reduce placental tissue damage and improve pregnancy outcome by correcting DEHP-induced imbalance of oxidative stress in the placental tissues.

Di(2-ethylhexyl) phthalate-induced toxicity and peroxisome proliferator-activated receptor alpha: a review

The plasticizer di(2-ethylhexyl) phthalate (DEHP) has been widely used in the manufacture of polyvinyl chloride-containing products such as medical and consumer goods. Humans can easily be exposed to it because DEHP is ubiquitous in the environment. Recent research on the adverse effects of DEHP has focused on reproductive and developmental toxicity in rodents and/or humans. DEHP is a representative of the peroxisome proliferators. Therefore, peroxisome proliferator-activated receptor alpha (PPARα)-dependent pathways are the expected mode of action of several kinds of DEHP-induced toxicities. In this review, we summarize DEHP kinetics and its mechanisms of carcinogenicity and reproductive and developmental toxicity in relation to PPARα. Additionally, we give an overview of the impacts of science policy on exposure sources.

Effects of Di(2-ethylhexyl)phthalate on Bone Metabolism in Ovariectomized Mice

BACKGROUND: The molecular pathways of how endocrine disruptors affect bone mineral density (BMD) and bone remodeling are still unclear. The purpose of this experimental study is to determine the effects of di(2-ethylhexyl)phthalate (DEHP) on bone metabolism in ovariectomized mice. METHODS: Twenty-six-month-old female CD-1 mice were divided into 4 groups: control, low-dose DEHP, high-dose DEHP, and estrogen groups (n=5, each group). All mice were subjected to ovariectomy for the induction of artificial menopause and then exposed to corn oil, DEHP, and estrogen for 2 months. Micro-computed tomography (Micro-CT) of the bone and analysis of blood samples for bone markers were performed to observe the changes in bone metabolism. RESULTS: Osteocalcin level was decreased in the control, low-dose and high-dose DEHP group, the reduction width was greater in the high-dose DEHP group (−0.219 ng/mL) than control group (−0.077 ng/mL, P<0.05). C-terminal telopeptide of type I collagen level was increased in the control, low-dose and high-dose DEHP group, the increase range of low-dose DEHP group (0.329 ng/mL) showed greater than control group (0.093 ng/mL, P<0.05). Micro-CT analysis revealed that the BMD was significantly lower in the high-dose DEHP group (19.8×10⁻² g/cm³) than control group (27.2×10⁻² g/cm³, P<0.05). The structure model index was significantly higher in the high-dose DEHP group (2.737) than low-dose DEHP group (2.648) and estrogen group (2.63, P<0.05). It means the progression of osteoporosis in the high-dose DEHP group. CONCLUSIONS: These results confirm the negative effects of DEHP on bone health in ovariectomized mice. Further continuous studies on genetic pathways and other endocrine disruptors will be necessary to validate these findings.

Association between urinary phthalate metabolites and obesity in adult Korean population: Korean National Environmental Health Survey (KoNEHS), 2012–2014

BACKGROUND: Phthalate is a chemical that is commonly used as a plasticizer in processing plastic products and as a solvent in personal care products. Although previous experimental studies have reported that phthalate metabolites are associated with obesity, epidemiological study results have been inconsistent and insufficient. The objective of the present study was to investigate the association between urinary phthalate metabolites and obesity in adult Korean population. METHODS: The present study selected 4,752 Korean adults aged 19 years or older from the 2012–2014 Korean National Environmental Health Survey data. The concentrations of urinary di-(2-ethyl-5-carboxypentyl) phthalate (DEHP) metabolites—i.e., mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP) and mono-(2-ethyl-5-carboxypentyl) phthalate—mono-benzyl phthalate (MBzP) and mono-n-butyl phthalate (MnBP) were adjusted using the urinary creatinine. We used logistic regression analysis to investigate the association between urinary phthalate metabolite concentration and body mass index (BMI) with respect to sex and age. RESULTS: Among women, urinary MEHHP and DEHP concentrations were found to have statistically significantly positive associations with obesity (Q4 versus Q1; odds ratio (OR): 1.72, 95% confidence interval (CI): 1.19–2.49 for MEHHP and OR: 1.52, 95% CI: 1.04–2.21 for DEHP). Among men, urinary MnBP concentration was found to have statistically significantly negative association with obesity (Q4 versus Q1; OR: 0.71, 95% CI: 0.50–0.99). In the analysis stratified by sex and age, women aged ≥ 50 years showed statistically significantly positive associations between the concentrations of urinary DEHP metabolites, DEHP, MBzP, and obesity (Q4 versus Q1; OR: 1.94, 95% CI: 1.28–2.94 for MEHHP, OR: 1.88, 95% CI: 1.21–2.94 for MEOHP, OR: 2.04, 95% CI: 1.31–3.18 for DEHP, and Q3 versus Q1; OR: 1.45, 95% CI: 1.02–2.05 for MBzP). Meanwhile, men aged ≥ 50 years showed no significant associations between urinary phthalate concentrations and obesity. CONCLUSIONS: In the present study, we found differences in the associations between urinary phthalate metabolites and BMI according to sex and age. However, because the present study was cross-sectional in nature, additional support through prospective studies is needed to estimate the causal associations.

Experimental Study on Migration Parameters of DEHP in PVC Infusion / 中国医疗器械杂志

The work explored the DEHP migration parameters in PVC infusion in clinic,based on the previous research on the test model of DEHP migrated from PVC infusion,to assess the safety of PVC infusion.The leaching solution samples in different conditions were evaluated by analysis of the DEHP in leaching solution using GC-MS under simulated clinical transfusion way.The release behavior of DEHP was significantly affected by the storage time,storage temperature,surrounding temperature,dripping speed,sterilization process,volume of the leaching solution,and the property of the leaching solution.

Di (2-ethylhexyl) phthalate-induced hypospadias in SD rats is related with Mafb expression: a transcriptome profiling-based study / 南方医科大学学报

OBJECTIVE@#To investigate the transcriptome profile of genital tubercles (GTs) in male SD rats and explore the mechanism of hypospadias induced by Di (2-ethylhexyl) phthalate (DEHP).@*METHODS@#Forty time-pregnant SD rats were randomly divided into 4 equal groups, namely GD16 group and GD19 group (in which the male GTs were collected on gestation day[GD]16 and GD19 for RNA-seq, respectively), control group and DEHP exposure group (with administration of oil and 750 mg/kg DEHP by gavage from GD12 to GD19, respectively).In the control and DEHP exposure groups, the GTs were collected from the male fetuses on GD19.5, and scanning electron microscopy and HE staining were used to observe the morphological changes.The differentially expressed genes (DEGs) in the GTs were screened using lllumina HiSeq 2000 followed by GO and KEGG enrichment analyses to characterize the transcriptome profile.Immunofluorescence assay was performed to verify the DEGs (Mafb) identified by RNA-seq results.Immunofluorescence assay and Western blotting were used to examine the expression levels of Mafb in the penile tissue.@*RESULTS@#A total of 1360 DEGs were detected in the GTs between GD16 group and GD19 group by RNA-seq.Among these genes, 797 were up-regulated and 563 were down-regulated.These DEGs were mainly enriched in the cell adhesion plaque signaling pathway, axon guidance signaling pathway, and extracellular matrix receptor signaling pathway.Compared with that in GD16 group, Mafb was significantly up-regulated in GD19 group, which was consistent with the sequencing results.Mafb and β-catenin were significantly down-regulated in DEHP-exposed group compared with the control group ( < 0.01).@*CONCLUSIONS@#Mafb expression increases progressively with the development of GTs in male SD rats.DEHP exposure causes significant down-regulation of Mafb and β-catenin, suggesting that β-catenin signaling pathway that affects Mafb is related to DEHP-induced hypospadias in SD rats.

Di (2-ethylhexyl) phthalate Disorders Lipid Metabolism via TYK2/STAT1 and Autophagy in Rats / 生物医学与环境科学（英文）

OBJECTIVE@#Previous studies have indicated that the plasticizer di (2-ethylhexyl) phthalate (DEHP) affects lipid accumulation; however, its underlying mechanism remains unclear. We aim to clarify the effect of DEHP on lipid metabolism and the role of TYK2/STAT1 and autophagy.@*METHODS@#In total, 160 Wistar rats were exposed to DEHP [0, 5, 50, 500 mg/(kg•d)] for 8 weeks. Lipid levels, as well as mRNA and protein levels of TYK2, STAT1, PPARγ, AOX, FAS, LPL, and LC3 were detected.@*RESULTS@#The results indicate that DEHP exposure may lead to increased weight gain and altered serum lipids. We observed that DEHP exposure affected liver parenchyma and increased the volume or number of fat cells. In adipose tissue, decreased TYK2 and STAT1 promoted the expression of PPARγ and FAS. The mRNA and protein expression of LC3 in 50 and 500 mg/(kg•d) groups was increased significantly. In the liver, TYK2 and STAT1 increased compensatorily; however, the expression of FAS and AOX increased, while LPL expression decreased. Joint exposure to both a high-fat diet and DEHP led to complete disorder of lipid metabolism.@*CONCLUSION@#It is suggested that DEHP induces lipid metabolism disorder by regulating TYK2/STAT1. Autophagy may play a potential role in this process as well. High-fat diet, in combination with DEHP exposure, may jointly have an effect on lipid metabolism disorder.

Low Dose Exposure to Di-2-Ethylhexylphthalate in Juvenile Rats Alters the Expression of Genes Related with Thyroid Hormone Regulation

Phthalates widely used in the manufacture of plastics have deeply penetrated into our everyday lives. Recently, a concern over the toxicity of phthalates on thyroid, has been raised but in most of cases, the doses employed were unrealistically high. To investigate the effects of phthalates on thyroid, we investigated the effects of the repeated oral exposure to low to high doses (0.3, 3, 30 and 150 mg/kg) di-2-ethylhexylphthalate (DEHP) from weaning to maturity for 90 days in juvenile rats on the thyroid. The histological examination revealed that DEHP significantly induced hyperplasia in the thyroid from the doses of 30 mg/kg, which was confirmed with Ki67 staining. In line with this finding, increased mRNA expression of thyrotropin releasing hormone (Trh) was observed in the thyroid of female at 0.3 mg/kg and 150 mg/kg as determined by RNAseq analysis. Moreover, significantly increased expression of parathyroid hormone (Pth) in the female at 0.3 mg/kg, and thyroglobulin (Tg) and thyroid hormone responsive (Thrsp) in the male at 0.3 mg/kg were noted in the blood, of which changes were substantially attenuated at 150 m/kg, alluding the meaningful effects of low dose DEHP on the thyroid hormone regulation. Urinary excretion of mono-2-ethylhexyl-phthalate (MEHP), a major metabolite of DEHP was determined to be 4.10 and 12.26 ppb in male, 6.65 and 324 ppb in female at 0.3 and 30 mg/kg DEHP, respectively, which fell within reported human urine levels. Collectively, these results suggest a potential adverse effects of low dose phthalates on the thyroid.

Research on Experimental Model of Security Assessment of DEHP Migration from PVC-Tubes Medical Devices / 中国医疗器械杂志

As we all know, DEHP is seriously harmful to human health and consequently has been acquired critical attention. DEHP is able to migrate from PVC medical devices for the non-chemically bound to PVC, thus contact with user and patient. The DEHP migration is influenced by various parameters. In order to assess the security of PVC-tubes medical devices scientifically of DEHP migration, we develop an experimental model by analyzing the parameters comprehensively and systematically, taking into account the clinical practices. For example, assessing the security of DEHP migration from infusion sets by utilizing this model.

Application and Safety Evaluation of DEHP in PVC Medical Devices / 中国医疗器械杂志

DEHP is largely used in soft PVC products as the plasticizer, which is also widely applied in medical devices. Due to its potential and widespread toxicity and medical devices' specific use, the safety of DEHP's application in medical devices has received extensive attention. In this paper, a comprehensive review of the application and potential toxicity of DEHP in PVC medical devices is made on the basis of the research results all over the world. Besides, the safety evaluation in medical devices is discussed and some possible coping strategies are explored.

Antagonistic effect of vitamin E on di-2-ethylhexyl phthalate-induced reproductive toxicity in male rats / 中华男科学杂志

<p><b>Objective</b>To explore the antagonistic effect of vitamin E (VE) on male reproductive toxicity induced by di-2-ethylhexyl phthalate (DEHP) in pubertal SD rats and its underlying mechanisms.</p><p><b>METHODS</b>Thirty 5-week-old male SD rats were randomly divided into five groups of equal number, corn oil control, low-dose (10 mg/kg/d), medium-dose (100 mg/kg/d) and high-dose DEHP exposure (500 mg/kg/d), and VE intervention (high-dose DEHP + VE ［100 mg/kg/d］), and treated respectively for 30 successive days. At 3 days after treatment, the testes of the animals were harvested for determination of the oxidative stress index, serum reproductive hormone levels, cauda epididymal sperm parameters, and expressions of cell apoptosis-related genes and proteins.</p><p><b>RESULTS</b>Compared with the control group, the rats of the medium- and high-dose DEHP groups showed significant decreases in the levels of such serum reproductive hormones as follicle-stimulating hormone (FSH), luteinizing hormone (LH) and testosterone (T), sperm parameters as average path velocity (VAP), straight line velocity (VSL), curvilinear velocity (VCL), straightness (STR), linearity (LIN) and wobble (WOB), and the activities of superoxide dismutase (SOD) and glutathione peroxide (GSH-Px), but significant increases were observed in the latter two groups in the content of malondialdehyde (MDA)(［3.32±0.87］ nmol/mg pro vs ［2.13±0.49］ nmol/ mg pro), mRNA expressions of Bad, Bax, Cytochrome C, Caspase-3 and the Bax/Bcl-2 ratio, and protein expressions of Cytochrome C and Caspase-3. In comparison with the high-dose DEHP group, the VE intervention group exhibited remarkably increased serum LH and T levels, sperm VAP, VSL, VCL, STR and WOB, and activities of SOD and GSH-Px, but markedly decreased mRNA expressions of Bad, Bax, Cytochrome C, Caspase-3 and the Bax/Bcl-2 ratio as well as the protein expressions of Cytochrome C and Caspase-3 in the testis tissue (P<0.05).</p><p><b>CONCLUSIONS</b>Exposure to DEHP induces androgen secretion disorders, causes oxidative damage to the testicular tissue, activates the mitochondrial apoptosis pathway in the testis, and ultimately reduces the quality of epididymal sperm, while VE can protect the rat testis from DEHP-induced reproductive toxicity.</p>

Developmental Exposure to Di-(2-ethylhexyl) Phthalate Induces Cerebellar Granule Cell Apoptosis via the PI3K/AKT Signaling Pathway

Di-(2-ethylhexyl) phthalate (DEHP) is an ubiquitous environmental contaminant because of its extensive use in plastics and its persistence. As an environmental endocrine disruptor, it is suspected to interfere with neurodevelopment in people. However, evidence of the effects of maternal DEHP exposure on cerebellar development in offspring is scarce. The objective of this study was to investigate maternal exposure to DEHP and its effect on apoptosis of cerebellar granule cells (CGCs) and related mechanisms. Pregnant Wistar rats were administrated DEHP (0, 30, 300 and 750 mg/kg/d) by gavage from gestational day (GD) 0 to postnatal day (PN) 21. Primary CGCs were also exposed to mono-(2-ethylhexyl) phthalate (MEHP), the main metabolite of DEHP, for 24 h with concentrations of 0, 25, 100 and 250 µM. The CGCs of male offspring from 300 and 750 mg/kg/d DEHP exposure groups showed significantly increased apoptosis. In addition, the PI3K/AKT signaling pathway was inhibited in the male offspring of the 300 and 750 mg/kg/d DEHP exposure groups. However, effects on female pups were not obvious. Apoptosis was also elevated and the PI3K/AKT signaling pathway was inhibited after primary CGCs were exposed to MEHP. Furthermore, apoptosis was reduced after treatment with the PI3K/AKT signaling pathway activator, insulin-like growth factor (IGF) 1, and increased after treatment with LY294002, an inhibitor of the PI3K/AKT signaling pathway. These results suggested that maternal DEHP exposure induced apoptosis in the CGCs of male pups via the PI3K/AKT signaling pathway, and the apoptosis could be rescued by IGF1 and aggravated by LY294002.

Effects of mono-[2-ethylhexyl] phthalate and DI-[2-ethylhexyl] phthalate administrations on oocyte meiotic maturation, apoptosis and gene quantification in mouse model

Objective: Phthalates, which are commonly used to render plastics into soft and flexible materials, have also been determined as developmental and reproductive toxicants in human and animals. The purpose of this study was to evaluate the effect of mono-[2-ethylhexyl] phthalate [MEHP] and di-[2-ethylhexyl] phthalate [DEHP] oral administrations on maturation of mouse oocytes, apoptosis and gene transcription levels

Materials and Methods: In this experimental study, immature oocytes recovered from Naval Medical Research Institute [NMRI] mouse strain [6-8 weeks], were divided into seven different experimental and control groups. Control group oocytes were retrieved from mice that received only normal saline. The experimental groups I, II or III oocytes were retrieved from mice treated with 50, 100 or 200 micro l DEHP [2.56 micro M] solution, respectively. The experimental groups IV, V or VI oocytes were retrieved from mouse exposed to 50, 100 or 200 micro l MEHP [2.56 micro M] solution, respectively. Fertilization and embryonic development were carried out in OMM and T6 medium. Apoptosis was assessed by annexin V-FITC/Dead Cell Apoptosis Kit, with PI staining. In addition, the mRNA levels of Pou5f1, Ccna1 and Asah1 were examined in oocytes. Finally, mouse embryo at early blastocyst stage was stained with acridine-orange [AO] and ethidium-bromide [EB], in order to access their viability

Results: The proportion of oocytes that progressed up to metaphase II [MII] and 2-cells embryo formation stage was significantly decreased by exposure to MEHP or DEHP, in a dose-dependent manner. Annexin V and PI positive oocytes showed greater quantity in the treated mice than control. Quantitative reverse transcriptase-polymerase chain reaction [qRT-PCR] revealed that expression levels of Pou5f1, Asah1 and Ccna1 were significantly lower in the treated mouse oocytes than control. The total cell count for blastocyst developed from the treated mouse oocytes was lower than the controls

Conclusion: These results indicate that oral administration of MEHP and DEHP could negatively affect mouse oocyte meiotic maturation and development in vivo, suggesting that phthalates could be risk factors for mammalians' reproductive health. Additionally, phthalate-induced changes in Pou5f1, Asah1 and Ccna1 transcription level could explain in part, the reduced developmental ability of mouse-treated oocytes

Risk assessment of di(2-ethylhexyl) phthalate in the workplace

OBJECTIVES: A hazard assessment of di(2-ethylhexyl) phthalate (DEHP), a commonly used workplace chemical, was conducted in order to protect the occupational health of workers. A literature review, consisting of both domestic and international references, examined the chemical management system, working environment, level of exposure, and possible associated risks. This information may be utilized in the future to determine appropriate exposure levels in working environments. METHODS: Hazard assessment was performed using chemical hazard information obtained from international agencies, such as Organization for Economic Cooperation and Development-generated Screening Information Data Set and International Program on Chemical Safety. Information was obtained from surveys conducted by the Minister of Employment and Labor (“Survey on the work environment”) and by the Ministry of Environment (“Survey on the circulation amount of chemicals”). Risk was determined according to exposure in workplaces and chemical hazard. RESULTS: In 229 workplaces over the country, 831 tons of DEHP have been used as plasticizers, insecticides, and ink solvent. Calculated 50% lethal dose values ranged from 14.2 to 50 g/kg, as determined via acute toxicity testing in rodents. Chronic carcinogenicity tests revealed cases of lung and liver degeneration, shrinkage of the testes, and liver cancer. The no-observed-adverse-effect level and the lowest-observed-adverse-effect level were determined to be 28.9 g/kg and 146.6 g/kg, respectively. The working environment assessment revealed the maximum exposure level to be 0.990 mg/m³, as compared to the threshold exposure level of 5 mg/m³. The relative risk of chronic toxicity and reproductive toxicity were 0.264 and 0.330, respectively, while the risk of carcinogenicity was 1.3, which is higher than the accepted safety value of one. CONCLUSIONS: DEHP was identified as a carcinogen, and may be dangerous even at concentrations lower than the occupational exposure limit. Therefore, we suggest management of working environments, with exposure levels below 5 mg/m³ and all workers utilizing local exhaust ventilation and respiratory protection when handling DEHP.

Effect of di-(2-ethylhexyl) phthalate exposure on placental development in pregnant mice / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effect of di-(2-ethylhexyl) phthalate (DEHP) exposure on the growth and development of placenta, uterine natural killer (uNK) cell number and angiogenesis at the maternal-fetal interface in pregnant mice.</p><p><b>METHODS</b>From day 1 of pregnancy, pregnant mice were exposed daily to DEHP by oral gavage at 125, 250, or 500 mg/kg for 13 consecutive days. The uterine and placental tissues were then harvested for HE staining and immunohistochemistry to examine the effect of DEHP exposure on the growth and development of the placenta and angiogenesis and uNK cell number at the maternal-fetal interface.</p><p><b>RESULTS</b>Compared with the control group, the mice exposed to 500 mg/kg DEHP, but not those exposed to 125 and 250 mg/kg, showed significantly reduced number of embryo implantation (P<0.05). DEHP exposure significantly increased the rate of abortion. DEHP exposure at 125, 250, and 500 mg/kg significantly and dose-dependently lowered the placental weight compared with that in the control group (0.0637±0.0133, 0.0587±0.0176, 0.0524±0.0183 g vs 0.0786±0.0143 g, respectively; P<0.01), and significantly reduced the total area of the placenta and area of spongiotrophoblasts. DEHP exposure resulted in a significant reduction in the number of fetal vascular branches, and collapse and atresia of blood vessels. The mice exposed to DEHP at 125, 250, and 500 mg/kg had significantly lowered numbers of uNK cells (83.2±10.3, 60.7±12.4, and 50.4±14.5/HP, respectively) as compared with the control group (105.1±14.2/HP) at the maternal-fetal interface (P<0.01).</p><p><b>CONCLUSION</b>DEHP exposure significantly affects the growth and development of the placenta in mice possibly by suppressing angiogenesis and reducing uNK cell number at the maternal-fetal interface during pregnancy.</p>

Dietary intake and risk assessment of diethylhexyl phthalate in Chinese populations / 中华预防医学杂志

<p><b>OBJECTIVE</b>To assess dietary exposure of diethylhexyl phthalate(DEHP) among Chinese population, including general population, children aged 2-6 years, adolescent aged 7-12, young people aged 13-17, adults aged 18-59 years old as well as older people aged 60 and above and its health risk.</p><p><b>METHODS</b>A total of 6 650 food samples were collected during 2011 to 2013 from 140 local markets of 14 provinces in China, which covered major foods in China. Samples were detected by GC-MS and categorized into 22 food groups. Food consumption data were taken from China National Nutrition and Health Survey performed in 2002 including 68 959 subjects. Mean concentrations of DEHP in food were combined with individual food consumption data to estimate dietary exposure, and food contributors to dietary DEHP intake were also calculated. Then, the exposure was compared with the tolerable daily intake (TDI, 50 µg·kg(-1)-d(-1)) of DEHP.</p><p><b>RESULTS</b>DEHP level in foods (n = 6 650) was in the range of not detected to 43.80 mg/kg. Mean dietary intakes of DEHP in general population was 2.07 (95% CI: 0.06-4.09) µg·kg(-1)·d(-1), accounting for 4.14 percent of TDI (50 µg·kg(-1)·d(-1)). Mean dietary intake for population aged 2-6, 7-12, 13-17, 18-59 as well as elderly aged 60 and above were 3.92 (95% CI: 0.83-7.01), 3.02 (95% CI: 0.69-5.36), 2.17 (95% CI: 0.54-3.81), 1.83 (95% CI: 0.46-3.21) and 1.66 (95% CI: 0.38-2.94) µg·kg(-1)·d(-1) respectively. The 97.5 percentile intakes in the general populations was 4.73 µg·kg(-1)·d(-1), accounting for 9.46% of TDI. Main food sources of DEHP were rice (28.4% (0.59/2.07)), melon solanaceous vegetables (14.7% (0.30/2.07)) and flour (13.2% (0.27/2.07)) for the general population.</p><p><b>CONCLUSION</b>The results suggested that dietary exposure to DEHP among Chinese population was lower than tolerable daily intake of DEHP and there were no health concerns based on generally accepted exposure limits. Rice, melon solanaceous vegetables and flour were main food contributors of DEHP dietary intake for Chinese populations.</p>

Determination of Plasticizer DEHP Released from Medical Injection Equipment / 中国医疗器械杂志

To established an effective GC-MS /MS method for the contents determination of the residual DEHP in injection equipment, and investigate the effect of the pretreatment on the measurement. To simulate the clinical conditions of use, under the condition of 37 degrees C balance extraction, extract liquor by chloroform extraction, then the extract followed by analysis of GC-MS /MS. The method was simple, rapid, sensitive and accurate. The limits of quantitation (LOQ, S/N = 5) of cyclohexanone was 0.075 μg/mL, The spiked average recoveries ranged from 92% to 98%. The relative standard deviations (RSDs) of the method ranged from 1.01% to 1.61%, The method was simple, fast, sensitive and accurate, and may serve as a mass control method for residual DEHP in injection equipment.

The effects of DEHP on morphology and function of progenitor Leydig cell in rat / 中国应用生理学杂志

<p><b>OBJECTIVE</b>To investigate the effects and mechanisms of diethylhexylphthalate (DEHP) on morphology and function of progenitor Leydig cells (PLC) in rats.</p><p><b>METHODS</b>Twenty pregnant SD rats were randomly divided into 4 groups ( n = 5): normal control group, DEHP low dose group , middle dose group, and high dose group, which were treated from postnatal day (PND) 1 to PND 21 of the pubs with DEHP at the doses of 0, 10, 100, 750 mg/(kg · d) in 0.5 ml of corn oil by gavage respectively. At the end of the treatment, the male pups were killed and blood samples were collected for determination of serum testosterone concentration by chemiluminescence method. The body weight, testis weight and anogenital distance (AGD) were measured. The morphology of PLC was observed by light and transmission electron microscopy. The protein expression of steroidogenic acute regulatory protein(StAR) in PLC was determined by immunohistochemistry. The mRNA expression of insulin-like growth factor-I (IGF-I) in the testis was assayed by real-time PCR.</p><p><b>RESULTS</b>Compared with normal control group, the serum testosterone and AGD of male pubs from the middle and high dose groups were declined significantly (P < 0.01), the testis weight and body weight from high dose group were decreased significantly (P < 0.01), while the testis weight increased in the low dose group (P < 0.05). Under light microscope, PLC showed hyperplasia and cluster aggregation in the low dose group and focal hyperplasia in the middle and high dose group. The spermatogenic cells in seminiferous tubules showed decrease, apoptosis and unfix in the high dose group. Under transmission electron microscope, the PLC showed decreased lipid droplets, smooth endoplasmic reticulum and mitochondriae in the treated group. The mRNA expression of IGF-I increased in the low dose group, and the protein expression of StAR decreased in the middle and high dose group.</p><p><b>CONCLUSION</b>Lactating exposure to DEHP may interfere with the synthesis of testosterone of PLC in male pubs, the decrease of StAR and the damage of PLC may be involved in it.</p>

Differential toxicological endpoints of di(2-ethylhexyl) phthalate (DEHP) exposure in MCF-7 and MDA-MB-231 cell lines: Possible estrogen receptor α (ERα) independent modulations.

Wide spread use of Di-(2-ethylhexyl) phthalate (DEHP) has made it a ubiquitous contaminant in today’s environment, responsible for possible carcinogenic and endocrine disrupting effects. In the present investigation an integrative toxico-proteomic approach was made to study the estrogenic potential of DEHP. In vitro experiments carried out with DEHP (0.1-100 μM) induced proliferations (E-screen assay) in human estrogen receptors-α (ERα) positive MCF-7 and ERα negative MDA-MB-231 breast cancer cells irrespective of their ERα status. Further, DEHP suppressed tamoxifen (a potent anti-breast cancer drug) induced apoptosis in both cell types as shown by flowcytometric cell cycle analysis. Label-free quantitative proteomics analysis of the cell secretome of both the cell lines indicated a wide array of stress related, structural and receptor binding proteins that were affected due to DEHP exposure. The secretome of DEHP treated MCF-7 cells revealed the down regulation of lactotransferrin, an ERα responsive iron transport protein. The results indicated that toxicological effects of DEHP did not follow an ERα signaling pathway. However, the differential effects in MCF-7 and MDA-MB-231 cell lines indicate that ERα might have an indirect modulating effect on DEHP induced toxicity.