| After 40 years of development,assisted reproductive technology(ART)has become the most effective way to treat infertility.However,with the hope brought to infertile families,ART also leads to higher risk of adverse pregnancy outcomes and cardiovascular and metabolic diseases in adulthood.As the crucial organ that links the mother and fetus,the placenta plays a key role in maintaining the material exchange between the mother and the fetus and ensuring the adequate development of fetus.The placental dysfunction will not only affect the intrauterine development of the fetus,but even affect the long-term health.These non-physiological manipulations on gametes and embryos may,as an iatrogenic stress factor,affect the development of the embryo,thus affecting the development of the placenta and fetal development.A multitude of studies have shown that there are sexual differences in the formation,function and adaption of placenta,which have different effects on the health of the offspring.Therefore,in this study,we use a mouse model to explore whether ART has a sexual dimorphism effect on the growth,development,morphology,transcriptomics and X linked gene expression of the placenta,in order to provide a new theoretical basis for pregnancy complications and offspring health risks caused by ART.Part 1 Sexual dimorphism effect of ART on placental growth,development and morphologyObjective: To explore the sexually different effects of ART on fetal and placental growth and development and placental tissue morphology of mice,and to analyze whether it leads to selective loss of embryos of different sexes,thus leading to skewed sex ratio.Methods: ICR mice were used as experimental model animals.We used IVF group as the experimental group,and embryo transferred after naturally fertilization(NET)as control group to balance the litter size.At E12.5 and E18.5 the placenta and fetuses were collected,respectively.Firstly,we analyzed the implantation rate,the living fetus rate and the embryo abortion rate in the two groups.Secondly,PCR was used to identify the sex of aborted embryos and fetuses at E12.5.Then the abortion rate of male and female embryos in the two groups was compared.Finally,we compared the fetus and placenta weight and placenta efficiency,and the morphology of mice placenta at E18.5.Results:(1)Compared with NET group,at E12.5,the implantation rate and embryo survival rate in the IVF group were significantly lower(P<0.05),and the embryo abortion rate was significantly higher(P<0.05).(2)There was no significant difference in the sex ratio in E12.5 and E18.5,but the abortion rate of female embryos in IVF group was higher than that of male embryos(P<0.05).(3)Compared with NET group,at E12.5,there was no significant difference in male placenta weight in IVF group(P>0.05),but the fetal weight decreased by 10.1%(P<0.05),and the placental efficiency decreased by 14.9%(P<0.05);female placental weight increased by 13.9%(P<0.05),fetal weight decreased by 9.1%(P<0.05),and placental efficiency decreased by 20.8%(P<0.05).(4)Compared with NET group,at E18.5,IVF group male placental weight increased by 34.4%(P<0.05),placental efficiency decreased by 23.4%(P<0.05),but there was no significant difference in fetal weight(P>0.05);Among females,in IVF group,placental weight increased by 23.2%(P<0.05),placental efficiency decreased by 19.7%(P<0.05),and there was no significant difference in fetal weight either(P>0.05).(5)Compared with the control group,at E18.5,it was found that the SPA staining area of IVF male placenta increased by 24.6%(P<0.05),while that of IVF female placenta increased by 19.7%(P<0.05).Conclusion: IVF leads to the increase of abortion rate of female embryos after implantation,and induced significant increase of female placental weight,but no significant effect on male placental weight at E12.5.Male placental hyperplasia and morphological changes was more obvious than female at E18.5.These findings suggest that ART has differential effects on male and female placenta in mice.Part 2 Sexual dimorphism effects of ART on placental transcriptomics and X chromosome inactivationObjective: To detect the transcriptomics and X-linked genes of placenta at E12.5 to explore the sexual different effects of IVF on mice placenta.Methods: We used transcriptome to detect the expression level of genome in placenta,and q RT-PCR to detect X-linked Chm,Cited,Esx1,Nrk,Plac1,Ftx,Jpx,Tsix and Xist.Results:(1)Compared with NET group,at E12.5,there were 338 differentially expressed genes in IVF male placenta,among which 222 were up-regulated and 116 were down-regulated;In contrast,212 genes were differentially expressed in IVF female placenta,among which 45 genes were up-regulated and 167 genes were down-regulated.More than 80% of the differentially expressed genes were different between males and females.(2)There are great differences in the biological processes involved in the differentially expressed genes between males and females.(3)Through the study of X-linked genes that are crucial to placental function we found that relative transcription level of Cited,Esx1,Nrk,Plac1 were significantly lower in IVF male placenta(P < 0.05),while only the relative transcription level of Esx1 was significantly lower in IVF female placenta(P < 0.05).(4)Through the transcriptional level detection of Lnc RNA related to X chromosome activation,it was found that the expression of Xist of both female and male placenta significantly increased(P<0.05),while the Tsix,which negatively regulates the expression of Xist,was significantly decreased in both male and female(P<0.05).Conclusion: There are differenct effects of IVF on transcriptomics of male and female placenta.Moreover,there are huge differences in gene function between male and female differentially expressed genes.Although IVF affected the X chromosome activation of both male and female placenta,IVF had a more significant effect on the X-linked gene expression of male placenta than that of female placenta. |
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