| Objective:Acrylamide,a widely used chemical and industrial material in our life,recently has been discovered in various high temperatures cooked carbohydrate-rich foods.The compound can be absorbed by inhalation,ingestion and dermal contact in daily life,and do harm to animal and human many systems,especially reproductive system.Previous studies have reported that maternal exposure to acrylamide significantly increased resorptions,decreased fetal weights of offspring and increased the number of abnormal fetus.However,no relevant information is available about the effects of acrylamide on the placental growth and development during pregnancy.Therefore,the study was designed to explore the effects of acrylamide on the placenta of pregnant mice and its mechanism by constructing animal models and giving different doses of acrylamide by gavage.Methods:1.Establishment of animal models and treatment: The virgin females(adult Kunming strain mice,approximately 25 to 30 g)were mated with the fertile males to induce the pregnancy in the same cage,and the day of finding the vaginal plug was regarded as gestational day(GD 1).The pregnant mice were given different doses of acrylamide(0,10,50 mg/kg/day)by forced gavage from the third day of gestational days(GD 3)according to the body weight,and we collected the uterus,placenta,ovary,liver and kidney specimens of mice at the 8th day of gestation(GD 8)and the13 th day of gestation(GD 13).2.Placental structure of pregnant mice exposed to acrylamide were observed by H&E staining on GD 8 and GD 13.3.The expression levels of placental growth and development marker genes of pregnant mice exposed to acrylamide were analyzed by real-time fluorescence quantitative PCR on GD 8 and GD 13.4.Placental labyrinth vascular of pregnant mice exposed to acrylamide wasobserved by immunohistochemical method on GD 13.5.The proliferation of placental cells of pregnant mice exposed to acrylamide was observed by immunohistochemical method on GD 8 and GD 13.6.Placental apoptosis and possible signaling pathways of pregnant mice exposed to acrylamide were explored by Western blot on GD 8 and GD 13.Results:1.There were no significant differences in the liver and kidney weight and organ coefficient between the control and acrylamide-treated groups on GD 8 and GD 13(P>0.05),but the ovary weight and organ coefficient were obviously decreased(P<0.05).Compared with the control group,exposure of acrylamide did not significantly affect the numbers of viable and resorbed embryos,the uterine weight and the ratio of uterus versus body weight on GD 8(P>0.05).However,on GD 13,the numbers of viable embryos markedly decreased(P<0.05)and the numbers of resorbed embryos significantly increased in the high-dose group compared with the control group(P<0.01).As the concentration of acrylamide increased,the weight of uterus,placenta and embryo and the ratio of uterus versus body weight significantly decreased compared with the control group(P<0.01,or P<0.001).2.Compared with the control group,the results of H&E staining showed that acrylamide treatment significantly inhibited the growth of the ectoplacental cone(EPC)on GD 8.On GD 13,the total area of placenta,the area of spongiotrophoblast,and the area of labyrinth in acrylamide-treated groups were dramatically reduced compared with the control group(P<0.01,or P<0.001).3.Real-time quantitative PCR data indicated that expression levels of Esx1,Hand2,and Hand1 mRNA were significantly lower in mice treated with acrylamide than that of control group on GD 8 and GD 13(P<0.01,or P<0.001).On GD 8,compared with the control group,acrylamide treatment significantly increased the levels of Ascl2 and Fra1 mRNA(P<0.001);but the expression levels of Eomes mRNA markedly decreased in the low-dose group and increased in the high-dose group(P<0.001).On GD 13,mice administered acrylamide had significantly inhibited Eomes and Ascl2 mRNA levels(P<0.001)。4.Compared with the control group,immunohistochemical results showed that acrylamide exposure significantly inhibited the formation of placental labyrinth blood vessels,and resulted in the collapse of fetal vessels and no intricate formation of branched vessels on GD 13.5.Compared with the control group,immunohistochemical results showed that low-dose acrylamide did not significantly affect the number of Ki67-positive cells(P>0.05),whereas the number of Ki67-positive cells dramatically decreased in the high-dose group on GD 8(P<0.05).On GD 13,acrylamide treatment group markedly decreased numbers of Ki67-positive cells compared to control group(P<0.001).6.Compared with the control group,western blot analysis results demonstrated that the apoptotic proteins Bax,Cleaved-caspase-3 and Cleaved-caspase-8significantly increased(P<0.01,or P<0.001);but anti-apoptotic protein Bcl-2markedly decreased in the high-dose acrylamide-treated groups(P<0.05)and there was no significantly difference between control group and low-dose group on GD 8and GD 13(P>0.05).The Caspase-3 and Caspase-8 were active in acrylamidetreatment group compared to control group.7.Western blot showed that there were no obviously difference of the phosphoErk1/2 levels between control group and low-dose group(P>0.05),but increased markedly in the high-dose group(P<0.01)on GD 8 and GD 13.Conclusions:Maternal exposure to acrylamide significantly inhibits the placental development and growth by the destruction of placental labyrinth vessels,down-regulation of placental development-related genes,inhibition of placental cell proliferation and induction of apoptosis。... |
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