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Objective: To explore the effects of bisphenol A (BPA) exposure on toxicity characteristic and OCT4 and SOX2 gene expression of mouse embryonic stem cells (mESC)., Methods: mESC were cultured, and treated with the doses of 10(-8), 10(-7), 10(-6), 10(-5), 10(-4) mol/L respectively of BPA and DMSO (the solvent control group)for 24 hours, and three groups of cells were treated with the same method. The morphological changes of mESC in the control and exposure groups were observed through an inverted microscope. Cell counting kit 8 (CCK8) was used to detect the effects of BPA on proliferation of mESC, and based on the results, the half inhibitory concentration (IC50) was calculated. Real-time fluorescent quantitative polymerase chain reaction (RT-QPCR) and western blotting were used to detect the expression of OCT4 and SOX2., Results: BPA had certain toxicity on mESC, the treatment of BPA significantly increased cell toxicity in a concentration-dependent manner, and the IC50 was 4.3×10(-4) mol/L, combined with the BPA exposure concentration of the environment and the related literature, eventually taking the five concentrations of 10(-8), 10(-7), 10(-6), 10(-5), 10(-4) mol/L as the experimental groups. The mESC morphology were effected after the treatment of BPA for 24 h, compared with the control group, the number of cells decreased, appearing some floating cells, and the cell cloning became irregular and differentiation in the higher concentration groups. The OCT4 mRNA expression level in the 10(-7) mol/L (1.146 ± 0.087), 10(-6) mol/L (1.156 ± 0.030), 10(-5) mol/L (1.158 ± 0.103) and the 10(-4) mol/L (1.374 ± 0.053) dose group were all significantly higher than the control group (1.000 ± 0.000) (t values were -2.384, -2.953, -3.203, -4.021 respectively, P value all < 0.05). Meanwhile, the SOX2 mRNA expression level in the 10(-4) mol/L (1.113 ± 0.052) were higher than the control group (1.000 ± 0.000) (t value was -2.765, P value < 0.05). Moreover, the OCT4 protein expression level in the 10(-5) mol/L (1.360 ± 0.168) and 10(-4) mol/L (1.602 ± 0.151) were all significantly higher than the control group (1.000 ± 0.000) (t values were -3.538, -4.002 respectively, P value all < 0.05), while no obvious change of the SOX2 protein expression level was detected in all treated groups., Conclusion: BPA in a certain dose range could upregulate the expression of OCT4 gene in mouse embryonic stem cells while had no significant effect on the expression of SOX2 gene.

Objective: To investigate DNA methylation variation in human cells induces by B(a)P, and to explore the role of PARP1 during this process., Methods: The changes of DNA methylation of 16HBE and its PARP1-deficient cells exposed to B(a)P (1.0, 2.0, 5.0, 10.0, 15.0, 30.0 µmol/L) were investigated by immunofluorescence and high performance capillary electrophoresis, and simultaneously, the expression level of PARP 1 and DNMT 1 were monitored dynamically., Results: The percentage of methylated DNA of overall genome (mCpG%) in 16HBE and 16HBE-shPARP1 cells were separately (4.04 ± 0.08)% and (9.69 ± 0.50)%. After being treated by 5-DAC for 72 hours, mCpG% decreased to (3.15 ± 0.14)% and (6.07 ± 0.54)%. After both being exposed to B(a)P for 72 hours, the mCpG% in 16HBE group (ascending rank) were separately (5.10 ± 0.13), (4.25 ± 0.10), (3.91 ± 0.10), (4.23 ± 0.27), (3.70 ± 0.15), (3.08 ± 0.07); while the figures in 16HBE-shPARP1 group (ascending rank) were respectively (10.63 ± 0.60), (13.08 ± 0.68), (9.75 ± 0.55), (7.32 ± 0.67), (6.90 ± 0.49) and (6.27 ± 0.21). The difference of the results was statistically significant (F values were 61.67 and 60.91, P < 0.01). For 16HBE group, expression of PARP 1 and DNMT 1 were 141.0%, 158.0%, 167.0%, 239.0%, 149.0%, 82.9% and 108.0%, 117.0%, 125.0%, 162.0%, 275.0%, 233.0% comparing with the control group, whose difference also has statistical significance (t values were 11.45, 17.32, 32.24, 33.44, 20.21 and 9.87, P < 0.01). For 16HBE-shPARP1 group, expression of PARP 1 and DNMT 1 were 169.0%, 217.0%, 259.0%, 323.0%, 321.0%, 256.0% and 86.0%, 135.0%, 151.0%, 180.0%, 229.0%, 186.0% comparing with the control group, with statistical significance (t values were 9.06, 15.92, 22.68, 26.23, 37.19 and 21.15, P < 0.01). When the dose of B(a)P reached 5.0 µmol/L, the mRNA expression of DNMT 1 in 16HBE group (ascending rank) were 125.0%, 162.0%, 275.0%, 233.0% times of it in control group, with statistical significance (t values were 12.74, 24.92, 55.11, 59.07, P < 0.01); while the dose of B(a)P reached 2.0 µmol/L, the mRNA expression of DNMT 1 in 16HBE-shPARP1 group were 135.0%, 151.0%, 180.0%, 229.0%, 186.0% of the results in control group, and the differences were statistically significant (t values were 23.82, 40.17, 32.69, 74.85, 46.76, P < 0.01)., Conclusion: The hypomethylation of 16HBE cells induced by B(a)P might be one important molecular phenomenon in its malignant transformation process. It suggests that PARP1 could regulate DNA methylation by inhibiting the enzyme activity of DNMT1, and this effect could be alleviated by PARP1-deficiency.

Objective: To explore the effects of zearalenone (ZEA) on proliferation and apoptosis in estrogen-dependent human breast cancer MCF-7 cells and the likely underlying molecular mechanisms., Methods: Cell viability was determined by MTT assay and cell cycle distribution by cytometry. Apoptosis was detected by Cell Death Detection ELISA and cytometry, respectively. The expressions of bax and bcl-2 were examined using multiple RT-PCR and Western-blot both at mRNA and protein level, respectively., Results: The current study confirmed the previous studies that ZEA could stimulate proliferation in MCF-7 cells with inducing a profound increase in S phase and a modest increase in G(2)/M phase that was accompanied by a decrease in G(0)/G(1) phase. ZEA could inhibit apoptosis in MCF-7 cells following estrogen ablation at a range of concentrations of 2 nmol/L -96 nmol/L. Western blot and RT-PCR analysis revealed that the anti-apoptotic bcl-2 was upregulated at both protein and mRNA level, together with the downregulation of pro-apoptotic bax., Conclusion: ZEA should have possessed comparative estrogenic activity and could promote the progression of MCF-7 cells through the cell cycle by a decreasing in the G(0)/G(1) phase and by a significant increasing in S-phase. The pro-proliferative activity of ZEA was due to inhibition of apoptosis through regulation of bax/bcl-2 expression.

Objective: To study the distribution of genotypes about alcohol dehydrogenase 2 (ADH2) and aldehyde dehydrogenase 2 (ALDH2) and its relationship with drinking-behaviors in Chinese Han healthy population as to providing a theoretic direction for filtering out high-risk and sensitive individuals and taking preventive measures to decrease the alcohol-related diseases., Methods: Using questionnaires to select subjects (201 persons, including men 104, women 97) for collecting blood samples and data about drinking-behaviors. Techniques of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) were used to detect the genotypes of ADH2 and ALDH2., Results: Heterozygous ADH2 and homozygous ALDH2 were the two dominant ones (respectively 53.23%, 68.16%). There were no statistical differences among the distributions of nine combinations of ADH2 genotypes and ALDH2 ones. The difference of distribution of homozygous ALDH2 between males having high and middle drinking-frequencies seemed to be statistically meaningful., Conclusion: The proportion of individuals carrying about "susceptible genotypes of alcohol-related diseases" in Chinese Han healthy population should be more than one half (68.16%), which calls on reinforcing the surveillance and preventing the alcohol-related diseases. Correlation between genotypes of ADH2 and ALDH2 and alcohol-related diseases should be more important.

Objective: To explore the effects of Bisphenol A in adult rats and its possible mechanisms., Methods: BPA (in corn oil) was administered orally to 9-week-old male Sprague-Dawley rats for 14 days (0, 1 and 5 g/kg bw), and incubated primary Sertoli cells from pubertal SD rats with 0, 10(-7), 10(-6), 10(-5), 10(-4) mol/L BPA., Results: After oral administration, a significant decrease in right testis weight was observed in 5 g/kg dose group, but not in the 1 g/kg bw dose group. Germ cells were detached from basement membrane of seminiferous tubules and Sertoli cells in BPA-treated groups. Administration of BPA at 1 g/kg bw and 5 g/kg bw produced both nucleus pycnosis and vacuolized nucleus in germ cells and Sertoli cells. A marked loss in vimentin staining in Sertoli cells from testis of BPA-treated rats was detected. No change in levels of serum estradiol and testosterone was observed after two-week exposure to BPA. In Sertoli cell primary culture, BPA destroyed the cytoskeleton and cell-cell junctions, and elongated Sertoli cells., Conclusion: These results suggest that BPA may injure reproductive function of male rats by destroying the cytoskeleton and changing the form of Sertoli cells.

Objective: To explore the effects of environmental estrogens (n-4-noniphenol, NP; bisphenol, BisA; and dibutylphthalate, DBP) on apoptosis induced by estrogen depletion in breast cancer T47D cells., Methods: Human T47D breast cancer cells were grown in DMEM medium containing 10% bovine serum. Four days before adding the test compounds, the cells were washed in phosphate-buffered saline, and the medium was substituted with a phenol red-free DMEM medium containing 5% dextral charcoal-stripped FBS. Respective test compound was added in fresh medium and the control cell received only the vehicle (ethanol). Apoptotic features in T47D cell were analyzed by light microscope that was commonly used to define apoptosis. DNA integrity of T47D cells was examined by agarose gel electrophoresis. Hypodiploid population was detected by flow cytometry., Results: The typical characters of apoptosis in T47D cells were observed after estrogen deletion and then disappeared following exposure to T47D cells at 32 x 10(-7) mol/L Np and 32 x 10(-7) mol/L BisA respectively. Inhibition of apoptosis at 32 x 10(-6) mol/L DBP was not shown in our study., Conclusion: N-4-noniphenol and Bisphenol A could inhibit apoptosis induced by estrogen deletion in breast cancer T47D cells. This result suggests that these environmental estrogens might involve in signal transduction connected with apoptosis.

Objective: The objective of this study was to investigate the estrogenic activity of genistein and zearalenone through their effects on the proliferative capacity of human ovarian PEO4., Methods: Estrogen receptor-positive PEO4 cell was grown in DMEM medium containing 10% bovine serum. Five days before the addition of the test compounds, the cells were washed in phosphate-buffered saline, and the medium was substituted with a phenol red-free DMEM medium containing 5% dextran charcoal-stripped FBS. The respective test compound was added in fresh medium and the control cell received only the vehicle (ethanol). Cell proliferation was detected respectively by MTT assay, (3)H-TdR incorporation and flow cytometry., Results: Compared with vehicle control, 96 x 10(-6) mol/L GS significantly inhibited PEO4 cell proliferation and DNA synthesis as measured by MTT and (3)H-TdR incorporation after treatment for 24 h. Alao, 32 x 10(-6) mol/L GS could exert inhibition on PEO4 cell growth as time extension to 48 h. 32 x 10(-6) mol/L approximately 96 x 10(-6) mol/L GS induced G(2)/M arrest. At low dose (< 8 x 10(-6) mol/L=, GS promoted proliferation in PEO4 cells. ZEA enhanced proliferation, promoted DNA synthesis and increased the S phase population in PEO4 cells., Conclusions: Genistein possess estrogenic activity and zearalenone have anti-estrogenic activity. They play different effects on the proliferation of human ovarian cancer cell. Genistein enhanced the proliferation of PEO4. Zearalenone inhibited its the proliferation. These results implied that genistein and zearalenone elicit different signal-transduction channel.

Objective: To explore the effect of environmental estrogens on the proliferation of breast cancer cell line MCF-7., Methods: The tested compounds were n-4-nonyphenol, Bisphenol A and dibutylphthalate. Human estradiol-dependent MCF-7 breast cancer cells were grown in DMEM medium containing 10% bovine serum. Five days before the addition of the test compounds, the cells were washed by phosphate-buffered saline, and the medium was substituted with a phenol red-free DMEM medium containing 5% dextral charcoal-stripped FBS. The respective test compound was added in fresh medium and the control cell received only the vehicle (ethanol). The proliferation of MCF-7 was analyzed by the MTT assay, (3)H-TdR incorporation assay and flow cytometry., Results: Compared with the ethanol control, the proliferation and DNA synthesis of the test cells treated with n-4-nonyphenol (8 x 10(-7) mol/L, 96 h), Bisphenol A (8 x 10(-7) mol/L, 96 h) or dibutylphthalate (32 x 10(-6) mol/L, 96 h) treatment was markedly enhanced in a time-dependent and dose-dependent manner., Conclusion: n-4-Nonyphenol, Bisphenol A and dibutylphthalate enhanced the proliferation of human breast cancer cell in vitro, which may demonstrate an estrogenic activity.