Study On The Effects And Related Mechanisms Of Bisphenol A On Reproductive And Endocrine System In Male Adult Rats

Bisphenol A (BPA) is the one of the most important organic chemical materials. As the derivant of phenol and acetonic, it is mainly used to produce polymers, such as polycarbonate, epoxide resin, polysulfone resin, polyphenylene oxide resin, and is also used in the production of plasticizer, fire retardant, antioxidant, thermal stabilizer, rubber antioxidant, pesticide, paints and so on. Now, BPA has become one of the highest production-volume chemicals in the world wide, and it is used extensively in lining food, beverage containers, cans inner packaging, tableware, milk bottles, and even dental sealant and fillers.In the past several years, BPA was generally considered as an environmental endocrine disruptor. Today many studies reported that BPA existed not only in many environmental mediums and lower organisms, but also in many human samples from different countries, like blood, urine, semen, amniotic fluid, milk and so on. Acute toxicity experiment shows that BPA is a low toxic substance, but many data of chronic and subchronic experiments from different laboratories indicate that BPA could cause lots of system damages from embryo to adult. And as a male reproductive endocrine disruptor, the studies of BPA have become one of the hotspot, but the mechanisms have not been confirmed.In the present study, we established BPA exposure model of male adult rats, in order to observe subchronic toxicity and the effects of reproductive endocrine system. Then we detected the changes in gene expression using cDNA microarray analysis, and combined with cluster analysis and bioinformatics database, we examined the expression of effect-related genes in target organ to analysis the mechanism preliminary. It would be helpful to provide the evidence and data for the effects and mechanisms of BPA's toxicity, and also provided the clue for early biomarker at gene level.PartⅠStudy on the effects of Bisphenol A on the reproductive endocrine system in male adult rats Objectives To estimate subchronic toxicity and the effects of reproductive endocrinesystem by establishing male adult SD rats model with different doses of BPA exposure. Methods Based on some toxicity parameters, including level of environmental exposure, LOAEL and NOAEL, 70 healthy male adult rats were divided into 5 groups, including one corn oil control group and four BPA treatment groups (0.0005, 0.5, 5, 50mg/kg). During oral administrating for 8 continuous weeks, we measured the body weights. After the treatment, we weighted the organ, detected the serum biochemical indicator (ALT, TP, BUN, CHOL), the levels of SOD and GSH-Px in plasma, serum T, E2, FSH, LH and E2/T ratio, so well as sperm count and its motility rate. Results During the treatment, the body weights of 50mg/kg group increased extremely slowly, and after 8 weeks, the final weights were significant lower than control group's. The plasma SOD and GSH-Px level showed downward trend, and GSH-Px level had decreased significantly at 5 and 50mg/kg. The influence on male reproductive endocrine had showed that serum T had increased with the increasing dose of BPA, and the level was obvious higher than control at 50mg/kg group, at the same time, E2/T ratio showed downward trend. The sperm number declined obviously in dose-dependent manner, but the motility rate had no significant difference.PartⅡStudy on the mechanisms of Bisphenol A induced reproductive endocrine toxicity in male adult ratsObjectives Using cDNA microarray analysis technology, we observed the changes of gene expression profile in liver tissue of male rats with low-dose BPA treatment. Combined with the effects of reproductive endocrine system, we also examined the expression of effect-related genes in target organ, to investigate the mechanisms of reproductive endocrine toxicity preliminary. Methods On the basis of previous experiment, we compared the gene expression between 0.5mg/kg BPA group and control group using gene chips, obtained 3 rats livers in each group by random, extracted RNA in low temperature and RNase free condition, reverse transcripted, then performed the chip hybridization, standardized the original data using gene spring software, meanwhile monitored and estimated the quality of the data. We screened the genes in certain conditions, classified the genes that we had chosen and validated them by real-time PCR. In addition, we evaluated the mRNA levels of the steroid hormones-related genes and spermatogenesis and development-related genes by real-time PCR. Results The quality of RNA met the experimental request, that detected by ultraviolet spectrophotometer and gel electrophoresis, and gene chips data had better repetitiveness and reliability. We obtained 334 genes up-regulated and 224 genes down-regulated, and found that steroid hormones-related genes and steroid hormones-regulated genes were up-regulated together, spermatogenesis and development-related genes were down-regulated together after cluster analysis. These genes were related to the effects of reproductive endocrine system. In steroid hormones synthesis and metabolism enzymes of testis, the mRNA levels of stAR, p450scc, p450c17 were obvious higher than that of control at high doses of BPA, meanwhile 3β-HSD and 17β-HSD had decreased significantly when BPA treated. In testis and liver, the mRNA levels of p450arom had showed a dose-dependent decline. Stard5 and shbg had no obvious change. In testis, we also found that the mRNA levels of AR and spermatogenesis and development-related genes, including Odf1 and Tnp1, showed significant down regulation, Mmp14 had minor changes.Conclusions In the rats model of subchronic toxicity, we found 50mg/kg BPA could lead to body weight increasing slowly, the levels of plasma antioxidase decreased, including SOD and GSH-Px. The effects of reproductive endocrine system, on one hand, BPA could disrupt the sex hormone of male adult rats, especially increased serum T, and the mechanism was related to the change of key enzymes of steroid hormones synthesis and metabolism, such as stAR, p450scc, p450c17, p450arom. And the changes appeared in lower dose than the effects. On other hand, BPA also could reduce the sperm number of male adult rats, it was possible concerned with the descent of transcriptional level of AR, and it also may be related to the spermatogenesis and development-related genes, such as Odf1 and Tnp1.