Effects Of Long-term Low-dose Bisphenol A Exposure On Reproductive Toxicity In Mice

Bisphenol A(BPA)is a raw material for the manufacture of plastic products and coatings which is considered to have reproductive toxicity.Exposure to BPA can lead to sugar metabolism disorders,hormonal disorders and death of offspring.The purpose of this research is to investigate the reproductive damage and abnormal gene expression related to reproduction caused by BPA in mice.The ovaries and testes of the 21 d and 45 d pups were subjected to transcriptomics sequencing to analyze the mechanism of BPA damage to the reproductive organs of offspring mice.The ovarian granulosa cells and testicular mesenchymal cells were cultured in vitro to confirm the differences of gene expression caused by BPA in animal experiments.1.Study on the reproductive toxicity of BPA on pregnant mice:Reproductive toxicity of BPA on pregnant mice:Based on the results of previous studies,the dose of BPA administered to pregnant mice was 50 mg·kg-1·d-1 and the dams were sacrificed at 3,6,9,12,15,18 days of pregnancy.This study counted the number of embryos at different stages of pregnancy in female mice and the apoptosis of ovary and uterus at different stages of pregnancy and estrogen receptor ?(ER?),estrogen receptor ?(ER?).The results showed that the serum BPA content of pregnant mice increased significantly after exposure to BPA(P<0.05).However,there was no severe pathological damage to the ovaries and uterus.On the other hand,due to the estrogenic properties of BPA,the expression of estrogen receptors in the ovaries and uterus of pregnant mice was changed after exposure to BPA.The expression of ER?(after pregnant day(PND)15)and ER?(whole pregnant)in the ovary was significantly increased while the expression of ER?(after PND 6)in the uterus was significantly increased(P<0.05).BPA caused a significant increase in ovarian cell apoptosis in pregnant mice(PND 3 to 9)and a significant decrease in uterine cell apoptosis(PND 3)(P<0.05).This study confirmed that BPA can affect uterine and ovarian ER expression,and further revealed that BPA can cause abortion in pregnant mice at PND 18(P<0.05).2.BPA on the reproductive toxicity of male offspring:Reproductive toxicity of BPA on male offspring:This study counted the pathological damage,apoptosis and expression of androgen receptor(AR)and synaptic complex protein 3(Scp3)in testes of 21-day-old and 45-day-old pups testis,while the sperm quality of 45-day-old pups was tested.At the same time,transcriptome sequencing was performed on the testes of 21-day and 45-day-old mice to observe the development of testis of offspring.The result showed that BPA exposure can cause testis abnormalities in pups.After exposure to BPA,BPA was significantly increased in the serum and testes of offspring male mice at 21 and 45 d(P<0.05),cause pathological changes,increased testicular germ cell apoptosis(P<0.05),and reduced AR expression(P<0.05).In addition,low-dose BPA can significantly reduce the testicular sperm quality(P<0.05).The decrease of SCP3 protein which related to spermatogenic cell differentiation further confirmed the effect of BPA on spermatogenesis(P<0.05).Transcriptome sequencing showed that BPA affected testicular spliceosome function,in which the testicular Snrnp40 was up-regulated and Hnrnpu down-regulated at 21 d mice,and the expressions of Snrpc and Hnrnpu were down-regulated at 45 d mice.Hnrnpu plays a connecting role when the mRNA precursor is combined with the spliceosome,and the down-regulation may cause the first step of post-transcriptional modification of mRNA to be blocked.The vitro culture of testicular mesenchymal cells confirms the effect of BPA on Hnrnpu.3.BPA on the reproductive toxicity of female offspring:The study counted the pathological damage to the reproductive organs,apoptosis and the expression of ERa and ER? in the ovaries and uterus of 21-day-old and 45-day-old pups.Meanwhile,transcriptomics sequencing was performed on the ovary of 21-day and 45-day-old pups to explore the changes of reproductive organ development and maturation and reproductive function of BPA on offspring.It confirmed that less than 50 mg·kg-1·d-1 BPA exposure can also cause ovarian abnormalities in pups.After exposure to BPA,BPA was significantly increased in the serum and ovarian of offspring female mice at 21 and 45 d(P<0.05).On the 21st day,the ovarian organ index of the pups increased(P<0.05),and the follicle granulosa cell layer was damaged.However,on the 45th day,the ovarian organ index of the pups did not show significant changes and pathological damage.BPA can increase the apoptosis of granulosa cells in ovaries of 21 d and 45 d mice(P<0.05),and the expression of ERa and ER? is disordered(P<0.05).Transcriptome sequencing showed that BPA affected ovarian ribosomal gene expression,in which the Rpl3,Rpl21 and Rpsa were significantly down-regulated at 21 d,and Rps2,Rps28,Rpl26,Rp132 and Rpl10a were significantly up-regulated.At born 45 d,the ovary Rpl21 and Rpsa were significantly down-regulated,while Rpl3,Rps2,Rpl7a,Rps26,Rpl26 were significantly increased.The abnormal ovarian development of the offspring may be related to the impaired ribosome function.The vitro culture results of ovarian granulosa cells confirmed the effect of BPA on Rps2.In summary,BPA can cause abortion in pregnant mice and abnormalities in testes and ovaries of their offspring.For offspring male testis development,BPA blocked the function of spliceosome and Hnrnpu might be the key gene;for offspring female ovarian,BPA affects the ribosome function,and Rps2 possibly was the key gene.