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Objective: To investigate the mechanism and effect of rat perinatal exposure to bisphenol A (BPA) resulting in midbrain dopaminergic neuronal cell apoptosis and tyrosine hydroxylase expression of male offspring., Methods: Rat dams were randomLy divided into 4 groups on gestational day(GD) 10 and given orally the bisphenol A doses as 0, 0.5, 5, 50 mg/kg x d from GD10 to weaning. The brains of male offspring were obtained for detecting, with immunohistochemistry protocol, the Caspase-3, Bcl-2 and tyrosine hydroxylase expression in the midbrain on postnatal day 21 or 30 respectively, and the midbrain apoptotic neuronal cell were detected by TUNEL on PND21., Results: The expression of Caspase-3 in the midbrain of rat male offspring were increased but bcl-2 were decreased on PND21 and 30, respectively. On PND21, apoptotic neuronal cell were found in the midbrain of high and medium doses groups. TH protein expression was decreased., Conclusion: Perinatal exposure to bisphenol A can induce the apoptosis of midbrain dopaminergic neuron in the male rat offspring even after weaning, and concomitantly decrease the midbrain TH immunoreactivity, this may cause the abnormal function of dopaminergic pathway of rat male offspring.

Objective: To study the effects of Flutamide (Flu) on the full gene expression of early testis development of embryonic period in male SD rats., Methods: Flu was injected subcutaneously (sc) with NS at doses of 6.25 mg/kg x d from 12th to 17th gestation days (GD), and in control group with tales doses NS. Fifty pregnant SD rats were sacrificed on GD 20. The mRNA were extracted respectively from fetus testis with treated and untreated Flu, then transcribed reversely to cDNA and labeled with cy5 and cy3 fluorescence. Subsequently, cDNA probes were hybridized to the Mouse40S cDNA microarray and fluorescence signals were analyzed. Meanwhile, we took out of the right testes of male offspring to make the detection of immunohistochemistry (IHC). Furthermore, we determined the level of testosterone by enzyme-linked immunosorbentassay (ELISA)., Results: Compared with the control group, a total of 31 identified genes expressed differently, including 14 down-regulated and 17 upregulated genes. However, the function of 5 down-regulated and 10 upregulated difference expression genes are unknown in Genbank. This finding has provided important reference basis for the molecular mechanism of reproductive development of Flu in the future. The activity of two important enzymes was cut down by IHC. The level of testosterone was decreased by ELISA., Conclusion: The results indicated that Flu administrated to F0 pregnant rats can damage apparently to the reproductive development of embryonic period male SD rats in many ways, and can cause the abnormality of sexual differentiation and sexual development in rats.

Objective: To study the mechanism of nonylphenol on testis tissue development and apoptosis of F1 generation male SD rats in weaning period., Methods: Nonylphenol was administrated via gastropipe to pregnant rats at doses of 50, 100 and 200 mg/kg, respectively, from the 7th day of preganacy to weaning. F1 generation male SD rats were sacrificed in the weaning period; their serum levels of nonylpheol were determined. The histopathological examination and immunohistochemical analysis(Bax,Bcl-2,Caspase-3) on testis were also carried out., Results: Compared with control, a higher serum nonylpheol level was found in F1 generation rats treated with nonylpheol 100 and 200 mg/kg. Histopathological examination revealed that rats treated with 200 mg/kg nonylphenol had much smaller diameter of testis seminiferous tubules when compared with those in the control group. Immunohistological analysis showed that the expression of Caspase-3 and Bax in the testis seminiferous tubules increased while the expression of Bcl-2 decreased in three experiment groups., Conclusion: All the above data proved that nonylphenol, after being given to pregnant rats, caused apparent damage to the development of testis of F1 generation male SD rats in weaning period.

Objective: To investigate the adverse effect of flutamide on the sex differentiation of F1 generation male rats and to establish a rat model of hypospadis., Methods: Sixty pregnant SD rats were divided randomly into one control group and five treatment groups. Flutamide was injected subcutaneously (sc) with normal salin (NS) at a dose of 0, 4882.8, 5078.1, 5273.4, 5468.7, and 6250.0 microg/kg x d to the rats in the above groups respectively during the period of gestation day (GD) 12 to 17. The fetuses of two rats from each group were anatomized on GD18 to extract genital tubercle (GT) to make pathological sections. Meanwhile, the position of testes and the development of prostates were examined. The position of urethral opening and testes were examined on GD2, 13 and 28., Results: The incidence of hypospadias increased with the dose of flutamide (0, 29.00%, 40.00%, 63.68%, 78.72%, and 100% for the above groups respectively). The incidence of single cryptorchidism also increased with the dose of flutamide (0, 4.23%, 6.23%, 9.23%, 11.52%, and 16.28% for the above groups respectively). Flutamide led to 100% incidence of prostate agenesis,100% incidence of female-like areola of nipple, and 100% incidence of female-like anogenital distance (AGD)., Conclusion: Flutamide can cause abnormal sex differentiation and sex development and induce the experimental model of hypospaias in rats.

Objective: To elucidate the mechanism of the toxicity of nonylphenol (NP) to reproductive development by analyzing the PCNA and Aromatase expression of the F1 generation male reproductive system in perinatal period., Methods: Nonylphenol was administrated to pregnant rats at doses of 50, 100 and 200 mg/kg, respectively, for 7 to 20 days of pregnancy. The rats were sacrificed at 20 days of pregnancy and 2 days after parturition. Their serum levels of estradiol and nonylphenol were determined. The histopathological examination and immune histochemical analysis on testis and epididymis were also performed., Results: Significantly higher serum estradiol and nonylphenol levels were found in the pregnant rats treated with nonylphenol. Immune histochemical analysis showed that the PCNA expression levels of testis tissues of F1 generation rats, both 20-day-old fetus and 2-day-old neonatal rats, in 100 mg/kg and 200 mg/kg experiment groups were much lower than those in the control group. Both PCNA expression of epididymis and Aromatase expression of testis of the F1 generation rats after 2 days of birth in 200 mg/kg group were also lower than those in the control group., Conclusion: Nonylphenol can impair the development of testis and epididymis of the F1 generation in perinatal period.

Objective: Human lymphoblastoid cell line TK6 was used to investigate TK gene mutation frequency and the loss of heterozygosity (LOH) induced by cyclophosphamide (CP)., Methods: Relative survival, mutation frequency at tk locus induced by CP (+S9) after 4 h treatment were detected. DNA isolated from mutants in the control culture and CP treatment group was analyzed for LOH., Results: Exposure to CP for 4 h decreased relative survival, induced TK gene mutation in a dose-dependent manner. Doubling time of normally and slowly growing mutants was (14.6 +/- 1.74) h and (35.8 +/- 3.78) h respectively. The percentage of hemizygous LOH in CP-induced mutants (50%) was 2.4 times than that of the control (20.7%)., Conclusion: Great damage to TK gene is the main cause of mutation induced by CPin TK6 cells.

Objective: To investigate the adverse effects of perinatal exposure to nonylphenol (NP) on the reproductive development of F1 male SD rats in sexual maturation period., Methods: The pregnant rats were randomly divided into control, 50 mg/kg, 100 mg/kg, and 200 mg/kg NP groups respectively. NP was administered to dams by gavage from gestation day 7 to weaning period. Rat pups were sacrificed at postnatal day 55. The concentration of serum testosterone was measured by radioimmunoassay. The epididymis was subjected to the determination of sperm count and motility while the testis was submitted to histopathological and immunohistochemical analyses., Results: Compared with control, the concentration of serum testosterone, sperm count and motility in the 200 mg/kg NP dose groups significantly decreased (P < 0.05), and the histopathological examination revealed that NP-treated groups had higher rates of maldeveloped siminiferous tubules, smaller amount of Sertoli cells and weaker spermatogenesis. The expression of proliferative cell nuclear antigen (PCNA) significantly decreased in the 200 mg/kg NP dose groups (P < 0.05). The expression of Arom in 100 mg/kg and 200 mg/kg NP dose groups was lower than that in control. There was no significant difference in ER expression between the control and NP-treated groups., Conclusion: These findings indicated that 200 mg/kg Nonylphenol apparently damaged the reproductive development of F1 male SD rats in the sexual maturation period.

Objective: Comparing the genotoxicity of gasoline-fueled vehicle exhaust (gasoline exhaust for short) with that of methanol-fueled vehicle exhaust (methanol exhaust for short) so as to provide a scientific basis for replacement of gasoline by methanol as fuel in vehicle., Methods: The MTT method was used first to evaluate the cytotoxicity of the two kinds of vehicle exhausts, and the concentration that had no obvious cytotoxicity would be used as the highest dose in the experiments. The A549 cells micronucleus test and single cell gel electrophoresis (SCGE) assay (comet assay) in vitro were applied to compare the genotoxicity of gasoline exhaust and methanol exhaust., Results: The MTT results showed that the gasoline-fueled vehicle exhaust exerted stronger cytotoxicity to A549 cells in both 2 h and 24 h exposure times, compared with the methanol exhaust. In A549 cells micronucleus test in vitro, at doses 0.025, 0.05, 0.1 and 0.2 L/ml, the micronucleated cell (MNC) rates were 2.65%, 4.35%, 4.95% and 5.85% respectively, which were higher than those (1.30% and 1.35%) of controls (P<0.01). For the methanol-fueled vehicle exhaust, there was no significant difference in the rate of A549 cells micronucleus between the test groups and control group. In the comet assay, gasoline-fueled vehicle exhaust could induce A549 cells DNA damage. The rate of caudate cell and the length of DNA migration increased with the escalation of dosing level. However, the methanol-fueled vehicle exhaust did not show any DNA damage to A549 cells., Conclusion: The results demanstrate that gasoline-fueled vehicle exhaust can induce DNA and chromosome damage, it has a distinct genotoxicity, whereas the methanol-fueled vehicle exhaust does not show any potential genotoxicity in both tests.

Objective: To investigate the secular trends in the prevalence rate of hypospadias in Chinese perinatals and explore its possible influencing factors., Methods: Data were taken from Chinese Birth Defects Monitoring Network, a hospital-based congenital anomalies registry system. All live or still births at or beyond 28 weeks of gestation were assessed within 7 days after delivery in the period from 1987 to 2001., Results: 1999 cases of hypospadias were identified among 3,769,464 male births, the crude prevalence rate being 5.30 per 10,000 births. And an increasing secular trend was observed both in urban and rural areas. In recent years, an increase in rate of hypospadias was found in coastal provinces and inland provinces, but not in remote border provinces. Faster increase was seen in urban areas and coastal areas, compared with that in rural areas and inland areas respectively., Conclusion: The prevalence rates of hypospadias in Chinese perinatals from 1987 to 2001 showed an upward tendency.

Objective: To study the effect of pesticide N'N-methylene-bis on thyroid function in rats, and the time-effect relationship., Methods: N'N-methylene-bis dimethylsulfoxide solution was administered to rats by gavage at a dose of 50 mg/kg body weight for 1, 3, and 5 days respectively. The rats in the control group were given solvent dimethylsulfoxide by oral gavage at the same volume. All animals were sacrificed 1 day after the last dosage, and the levels of FT4 and TSH in serum were measured by radioimmunoassay. The histological changes of thyroid gland were observed, and the expression of PCNA in thyroid was detected by immunohistochemistry., Results: Compared with the solvent control group, the serum FT4 level in rats given N'N-methylene-bis for 3 days significantly decreased, while the TSH level significantly increased in the 3 d and 5 d groups (P<0.05). Hyperplasia of thyroid follicular epithelium was observed in the 3 d and 5 d groups. PCNA positive cells in thyroid of the 3 d and 5 d groups were significantly higher than that of control (P<0.05)., Conclusion: The effect of N'N-methylene-bis on the level of serum FT4 and TSH in rats appeared on the 2nd or 3rd day of the experiment. Histopathologic examination showed thyroid gland proliferation of follicular epithelium within 3 days. The combined use of serologic test and histopathologic examination for screening thyroid hormone disruptors may be an effective method.

Objective: To investigate the environmental degradation process of nonylphenol(NP):, Methods: The NP was detected by HPLC, and the system of ultraviolet light, ferrous ion and hydrogen peroxide was used to promote photo-degradation of NP in water., Results: The maximal photo-degradation of NP occurred at the 5.0 mg/L of Fe2+, pH 2.5 and 1 ml/L of H2O2, the environmental factors in order of importance to promoting the photo-degradation of NP were: illumination time, initial concentration of NP, pH value, concentration of H2O2, concentration of Fe2+., Conclusion: The system of ultraviolet light, ferrous ion and hydrogen peroxide was effective in promoting the photo-degradation of NP.

Objective: This study was directed to the development of a simple and highly sensitive method for determination of 4-nonylphenol (4-NP) and bisphenol A (BPA) in rat serum using HPLC with fluorescence detector., Methods: 4-NP and BPA in serum samples were extracted by a mixed solvent of n-hexane and diethyl ether (70:30), the organic layer was dried with nitrogen flow and dissolved with mobile phase. The operating conditions such as C18 column (250 mm x 4.6 mm, 5 microns), acetonitrile-ammonium acetate buffer (pH 4.5) as mobile phase at a flow rate of 1 ml/min and fluorescence detector with the excitation and emission wavelength at 227 nm and 313 nm respectively were used for the determination., Results: There was good linear relationship between the concentrations of the analytes in rat serum and their peak areas in the ranges of 0.013-5.0 micrograms/ml for 4-NP and 0.004-3.0 micrograms/ml for BPA. The detection limit of the method was 13 ng/ml for 4-NP and 4 ng/ml for BPA. Recoveries of rat serum samples were 83.8%-100.0% for 4-NP and 83.3%-100.0% for BPA. The intra-day precision was 1.70%-2.70%, the inter-day precision was 2.06%-9.78%., Conclusion: The results showed that the method is suitable for the determination of BPA and 4-NP in serum.