Animals ; Benzo(a)pyrene/toxicity* ; Cytokines ; Dibutyl Phthalate/toxicity* ; Female ; Liver ; Male ; NF-kappa B ; Rats ; Rats, Sprague-Dawley

Objective: To induce hypospadias in male rat offspring by maternal exposure to di-n-butyl phthalate (DBP) during late pregnancy and further investigate its mechanisms. METHODS: We randomly divided 20 pregnant rats into a DBP exposure and a control group, the former treated intragastrically with DBP while the latter with soybean oil at 750 mg per kilogram of the body weight per day from gestation days (GD) 14 to 18. On postnatal day (PND) 1, we recorded the incidence rate of hypospadias and observed the histopathological changes in the genital tubercle of the hypospadiac rats. We also measured the level of serum testosterone (T) by radioimmunoassay and determined the mRNA and protein expressions of the androgen receptor (AR), sonic hedgehog (Shh), bone morphogenetic protein 4 (Bmp4) and fibroblast growth factor 8 (Fgf8) in the genital tubercle by real-time PCR and Western blot. RESULTS: No hypospadiac male rats were found in the control group. The incidence rate of hypospadias in male offspring was 43.6% in the DBP-treatment group. Histological analysis confirmed hypospadiac malformation. The serum testosterone concentration was decreased in the hypospadiac male rats as compared with the controls (［0.49 ± 0.05］ vs ［1.12 ± 0.05］ ng/ml, P <0.05). The mRNA expressions of AR, Shh, Bmp4 and Fgf8 in the genital tubercle were significantly lower in the hypospadiac male rats than in the controls (AR: 0.50 ± 0.05 vs 1.00 ± 0.12, P <0.05; Shh: 0.65 ± 0.07 vs 1.00 ± 0.15, P <0.05; Bmp4: 0.42 ± 0.05 vs 1.00 ± 0.13, P <0.05; Fgf8: 0.46 ± 0.04 vs 1.00 ± 0.12, P <0.05), and so were their protein expressions (AR: 0.34 ± 0.05 vs 1.00 ± 0.09, P <0.05; Shh: 0.51 ± 0.07 vs 1.00 ± 0.12, P <0.05; Bmp4: 0.43 ± 0.05 vs 1.00 ± 0.11, P <0.05; Fgf8: 0.57 ± 0.04 vs 1.00 ± 0.13, P <0.05). CONCLUSIONS Maternal exposure to DBP during late pregnancy can induce hypospadias in the male rat offspring. DBP affects the development of the genital tubercle by reducing the serum T concentration and expressions of AR, Shh, Bmp4 and Fgf8 in the genital tubercle, which might underlie the mechanism of DBP inducing hypospadias.
Animals ; Body Weight ; Bone Morphogenetic Protein 4 ; blood ; Dibutyl Phthalate ; toxicity ; Female ; Fibroblast Growth Factor 8 ; blood ; Hedgehog Proteins ; blood ; Hypospadias ; blood ; chemically induced ; pathology ; Male ; Maternal Exposure ; Plasticizers ; toxicity ; Pregnancy ; RNA, Messenger ; blood ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, Androgen ; blood ; Soybean Oil ; Testosterone ; blood

OBJECTIVETo investigate the relationship between atopic allergy and depression and the role of DBP in the development of depression.

METHODSBALB/c mice were randomly divided into eight groups: saline; ovalbumin (OVA)-immunized; saline+DBP (0.45 mg/kg•d); saline+DBP (45 mg/kg•d); DBP (0.45 mg/kg•d) OVA-immunized; DBP (45 mg/kg•d) OVA-immunized; saline+hydrocortisone (30 mg/kg•d); and hydrocortisone (30 mg/kg•d)-exposed OVA-immunized. Behavior (e.g. open-field, tail suspension, and forced swimming tests), viscera coefficients (brain and spleen), oxidative damage [e.g. reactive oxygen species (ROS), malondialdehyde (MDA), and glutathione (GSH)], as well as levels of IgE and IL-4, were then analyzed.

RESULTSIn the saline and OVA groups, the degree of depression symptoms in mice increased with increasing DBP concentration. Additionally, the OVA-immunity groups were associated with more serious depressive behavior compared with the same exposure concentration in the saline group. Oxidative damage was associated with a dose-dependent increase in DBP in the different groups. IL-4 and IgE levels were associated with low-dose DBP stimulation, which changed to high-dose inhibition with increasing DBP exposure, possibly due to spleen injury seen at high DBP concentrations.

CONCLUSIONDevelopment of an atopic allergy has the potential to increase the risk of depression in mice, and it seems that DBP helps OVA to exert its effect in our present model. Moreover, the results of our study implicate a certain connection between brain oxidative stress and depression, which deserves a further exploration.

Animals ; Behavior, Animal ; drug effects ; Body Weight ; Depression ; blood ; chemically induced ; immunology ; Dibutyl Phthalate ; immunology ; toxicity ; Environmental Pollutants ; immunology ; toxicity ; Hydrocortisone ; Hypersensitivity, Immediate ; blood ; complications ; Immunization ; Immunoglobulin E ; blood ; Interleukin-4 ; blood ; Male ; Mice ; Mice, Inbred BALB C ; Ovalbumin ; Oxidative Stress

OBJECTIVETo observe the effects of dibutyl phthalate (DBP) and monobutyl phthalate (MBP) on the mRNA and protein expression of insulin-like factor 3 (INSL3) in the Leydig tumor cells (MA-10) of mice and the level of testosterone secreted from MA-10 cells.

METHODSThe MA-10 cells of mice, used as a cellular model, were exposed to DBP and MBP. The content of testosterone in the supernatant medium was measured by enzyme-linked immunosorbent assay; the mRNA and protein expression levels of INSL3 in MA-10 cells were measured by quantitative PCR and Western Blot.

RESULTSCompared with the control group, MA-10 cells showed increased synthesis of testosterone when exposed to low concentrations of DBP and MBP (10(-9) ∼ 10(-6) mol/L) and inhibited synthesis of testosterone when exposed to high concentrations of DBP and MBP (10(-3) mol/L), and the typical two-way effects became more significant as the time went one and the concentrations increased (P < 0.05). Compared with the control group, MA-10 cells showed significantly lower mRNA and protein expression levels of INSL3 when exposed to 10(-6) and 10(-4) mol/L DBP (P < 0.05); MA-10 cells showed increased protein expression of INSL3 when exposed to 10(-7) mol/L MBP, and the mRNA and protein expression levels of INSL3 decreased as the concentration of MBP increased.

CONCLUSIONDBP and MBP can inhibit the secretion of testosterone from MA-10 cells at high concentrations, but stimulate the secretion of testosterone at low concentrations. Both DBP and MBP have inhibitory effects on the mRNA and protein expression of INSL3 in MA-10 cells.

Animals ; Cell Line ; Dibutyl Phthalate ; toxicity ; Insulin ; metabolism ; Leydig Cells ; drug effects ; metabolism ; Male ; Mice ; Phthalic Acids ; toxicity ; Proteins ; metabolism ; Testosterone ; secretion

OBJECTIVETo evaluate the combined subchronic toxicity of bisphenol A (BPA) and dibutyl phthalate (DBP) in male Sprague Dawley (SD) rats.

METHODSForty 4-week-old male rats weighing 115-125 g were randomly divided into BPA-treated, DBP-treated group, BPA+DBP-treated and control groups and fed with a soy- and alfalfa-free diet containing 285.4 ppm BPA, 285.4 ppm DBP, 285.4 ppm BPA plus 285.4 ppm DBP, and a control diet, respectively, for 90 consecutive days. At the end of the study, the animals were sacrificed by exsanguination via the carotid artery under diethyl etherane aesthesia and weighed. Organs, including liver, kidneys, spleen, thymus, heart, brain, and testis underwent pathological examination. The androgen receptor (AR), gonadotropin-releasing hormone receptor (GNRHR), and progesterone hormone receptor (PR) genes from the hypothalamus were detected by real-time PCR. The biomedical parameters were analyzed.

RESULTSNo significant difference was found in food intake, body weight, tissue weight, organ/brain weight ratio, and biomedical parameters among the four groups (P>0.05). However, BPA and DBP up-regulated AR, PR and GNRHR expression levels in rats and showed a synergistic or an additive effect in the BPA+DBP group.

CONCLUSIONThe combined subchronic toxicity of BPA and DBP is synergistic or additive in male SD rats.

Animals ; Benzhydryl Compounds ; administration & dosage ; toxicity ; Body Weight ; drug effects ; Dibutyl Phthalate ; administration & dosage ; toxicity ; Drug Interactions ; Eating ; drug effects ; Environmental Pollutants ; administration & dosage ; toxicity ; Male ; Phenols ; administration & dosage ; toxicity ; Random Allocation ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo detect the differential expression of Notch1 in the genital tubercle (GT) of fetal male rats with hypospadias induced by maternal exposure to Di-n-butyl phthalate (DBP) and that in normal control fetal rats in order to further explore the role of Notch1 in DBP-induced hypospadias.

METHODSTwenty pregnant SD rats were equally and randomly divided into an experimental and a control group, the former given DBP and the latter soybean oil intragastrically at 800 mg/(kg x d) and 2 ml/d respectively from gestation day (GD) 14 to GD 18. On GD 19, the birth weight (BW), anogenital distance (AGD) and hypospadias incidence were recorded, GTs of the fetal male rats collected, and the expression of Notch1 analyzed by Western blot and immunohistochemistry.

RESULTSThe BW of the fetal male rats was (4.40 +/- 0.30) g in the experimental group, significantly lower than (6.11 +/- 0.40) g in the control (P <0.05), and the AGD was (2.17 +/- 0.18) mm in the former, markedly shorter than (3.28 +/- 0.16) mm in the latter (P<0.05). The incidence of hypospadias was 42.9%. The relative expression of Notch1 was remarkably lower in the hypospadiac rats than in the normal controls (0.671 +/- 0.021 vs 1.327 +/- 0.031, P<0.05), and it was mainly located in the epithelial cells of the GT. The staining intensity was obviously weaker in the hypospadias than in the normal control group.

CONCLUSIONDBP has an obvious toxic effect on fetal male rats and can change the expression of Notch1 in the GT. It possibly affects cell proliferation and apoptosis and epithelial-to-mesenchymal transition (EMT), resulting in the occurrence of hypospadias.

Animals ; Dibutyl Phthalate ; toxicity ; Female ; Fetus ; Hypospadias ; chemically induced ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Receptor, Notch1 ; metabolism

OBJECTIVETo evaluate the effect of dibutyl phthalate (DBP) and di-(2-ethylhexyl) phthalate (DEHP) on urine superoxide dismutase (SOD) activity and malondialdehyde (MDA) content in rats.

METHODSAccording to 2×2 factorial analysis, 60 adult male SD rats were randomized into 10 groups (n=6), including a control group (fed with sesame oil), 3 DBP groups (fed with DBP at the doses of 30, 100 and 300 mg/kg), 3 DEHP groups (with DEHP at 50, 150, and 450 mg/kg), and 3 DBP+DEHP groups (with 30 mg/kg DBP+50 mg/kg DEHP, 100 mg/kg DBP+150 mg/kg DEHP, and 300 mg/kg DBP +450 mg/kg DEHP). The agents were administered in a single dose through gavage in a volume of 2 ml. After the treatments, the 24, 48, 72, and 96 h urine samples were collected to determine the SOD activity and MDA content.

RESULTSDBP and DEHP, either alone or in combination, significantly decreased SOD activity and increased MDA content in the urine collected at 24 h but not at the other time points. Such changes were gradually reversed with time.

CONCLUSIONDBP or DEHP treatment alone can result in significant oxidative damage in the kidney of rats, and the toxic effect of the combined exposure is even more obvious.

Animals ; Dibutyl Phthalate ; toxicity ; Diethylhexyl Phthalate ; toxicity ; Environmental Pollutants ; toxicity ; Kidney ; drug effects ; physiopathology ; Male ; Malondialdehyde ; urine ; Oxidative Stress ; drug effects ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase ; metabolism ; urine

OBJECTIVEThe aim of the present study was to investigate the effects of paternal Di-N-butyl-phthalate (DBP) exposure pre- and postnatally on F1 generation offspring, and prenatally on F2 generation offspring.

METHODSMale mice were exposed to either 500 mg/kg or 2 000 mg/kg of DBP for 8 weeks, and mated with non-exposed females. Three-quarters of the females were sacrificed a day prior to parturition, and examined for the number of living and dead implantations, and incidence of gross malformations. Pups from the remaining females were assessed for developmental markers, growth parameters, as well as sperm quantity and quality.

RESULTSThere were no changes in the fertility of parents and in intrauterine development of the offspring. Pups of DBP-exposed males demonstrated growth-retardation. Following paternal exposure to 500 mg/kg bw of DBP, there were almost twice the number of males than females born in the F1 generation. F1 generation females had a 2.5-day delay in vaginal opening. Paternal exposure to 2 000 mg/kg bw of DBP increased the incidence of sperm head malformations in F1 generation males; however, there were no changes in the fertility and viability of foetuses in the F2 generation.

CONCLUSIONPaternal DBP exposure may disturb the sex ratio of the offspring, delay female sexual maturation, and deteriorate the sperm quality of F1 generation males.

Abnormalities, Drug-Induced ; Animals ; Dibutyl Phthalate ; toxicity ; Female ; Male ; Mice ; Paternal Exposure ; adverse effects ; Plasticizers ; toxicity ; Pregnancy ; Sex Ratio ; Sexual Development ; drug effects ; Sperm Head ; drug effects ; pathology

OBJECTIVES: Maintaining the constant exposure to hydrophobic organic compouds in acute toxicity tests is one of the most difficult issues in the evaluation of their toxicity and corresponding risks. Passive dosing is an emerging tool to keep constant aqueous concentration because of the overwhelming mass loaded in the dosing phase. The primary objectives of this study were to develop the constant exposure condition for an acute mortality test and to compare the performance of the passive dosing method with the conventional spiking with co-solvent. METHODS: A custom cut polydimethylsiloxane (PDMS) tubing loaded with benzyl butyl phthalate (BBP) was placed in each well of a 24-well plate containing assay medium. The rate of the release of BBP from PDMS was evaluated by measuring the change in the concentration of BBP in the assay medium. The efficiency of maintaining constant exposure condition was also evaluated using a simple two-compartment mass transport model employing a film-diffusion theory. An acute mortality test using 10 C. elegans in each well was conducted for the evaluation of the validity of passive dosing and the comparative evaluation of the passive dosing method and the conventional spiking method. RESULTS: Free concentration in the assay medium reached 95% steady state value within 2.2 hours without test organisms, indicating that this passive dosing method is useful for an acute toxicity test in 24 hours. The measured concentration after the mortality test agreed well with the estimated values from partitioning between PDMS and the assay medium. However, the difference between the nominal and the free concentration became larger as the spiked concentration approached water solubility, indicating the instability of the conventional spiking with a co-solvent. CONCLUSIONS: The results in this study support that passive dosing provides a stable exposure condition for an acute toxicity test. Thus, it is likely that more reliable toxicity assessment can be made for hydrophobic chemicals using passive dosing.
Benzophenones ; Biological Availability ; Boronic Acids ; Caenorhabditis ; Caenorhabditis elegans ; Dibutyl Phthalate ; Dimethylpolysiloxanes ; Phthalic Acids ; Solubility ; Toxicity Tests, Acute

Animals ; Dibutyl Phthalate ; toxicity ; Female ; Hippocampus ; cytology ; growth & development ; Male ; Mitochondrial Proton-Translocating ATPases ; metabolism ; Neurons ; drug effects ; metabolism ; Pregnancy ; Rats ; Rats, Wistar