| Objective1-nitropyrene(1-NP)is a representative nitro-polycyclic aromatic hydrocarbon pollutant ubiquitously distrubuted in exhaust particles of internal combustion engines.The mechanism of intrauterine growth restriction induced by 1-NP exposure during pregnancy remains unclear.In this study,the dose-response relationship of 1-NP exposure induced intrauterine growth restriction was analyzed through animal experiments,and the effects on placental structure,transport and endocrine functions as well as spiral artery remodeling were further explored.The findings of this study could provide evidence and new ideas for the exploration of specific mechanisms of 1-NP exposure induced intrauterine growth restriction during pregnancy.MethodsThis study has been approved by the Experimental Animal Ethics Committee of Anhui Medical University(No.: LLSC20221239),and the animals used were purchased from Zhejiang Vital River Laboratory Animal Technology Co.,Ltd.Wild-type SPF C57BL/6N mice(about 19g)at 8 weeks of age were acquired.They were reared in SPF Laboratory Animal Center of Anhui Medical University and reached 10 weeks of age(20-21g)after 1 week of quarantine and 1 week of adaptive feeding.According to the ratio of 2:4 male to female,conception was performed at 9:10 PM,and vaginal embolism was examined at 7:10 am the next day.The 0 day of pregnancy(GD0)was defined for the female mice whose vaginal embolism was detected.Pregnant mice were fed 4 in each cage,and their feed,water intake and weight were measured daily.Sixty pregnant mice were randomly divided into 4 groups,with 15 mice in each group.From day 0 of pregnancy(GD0),the mice were exposed to 1-NP(100μg/kg,50μg/kg,25μg/kg)and 1-NP free solvents(DMSO and Tween80)every 6 days,respectively.Five pregnant mice in each group were sacrificed on the 12 th day of pregnancy(GD12).Placental tissues were collected and the relationship between 1-NP exposure in the first and second trimesters and trophoblast invasion of placenta and spiral artery remodeling was analyzed.The remaining pregnant mice were sacrificed on the 18 th day of gestation(GD18)to analyze the relationship between 1-NP exposure during gestation and general and pregnancy status of pregnant mice,placental structure and function(transport and endocrine),as well as intrauterine growth restriction of fetal mice.ResultsThis study demonstrated that 1-NP exposure during pregnancy leads to intrauterine growth restriction in fetal mice.There was no significant difference between the groups in general and conception conditions.Compared with the solvent control group,high dose(100μg/kg)1-NP exposure significantly reduced fetal body weight,crown-rump length and placental diameter.Meanwhile,the placental structure and spiral artery remodeling of pregnant rats exposed to 1-NP concentration gradient air pollutants during the first and second trimester were evaluated.The overall structure of the placenta was changed by medium and high doses(50μg/kg and 100μg/kg)of 1-NP exposure,but the structural composition ratio did not change significantly.Immunohistochemistry results of the trophoblast spiral artery showed that 1-NP exposure in each dose group inhibited the remodeling of spiral artery in the first and second trimesters of pregnancy,which showed that the number of residual trophoblast cells in the wall of spiral artery significantly decreased and the number of smooth muscle cells significantly increased,indicating that 1-NP exposure in the first and second trimesters induced the defect of spiral artery remodeling.1-NP exposure during pregnancy significantly changed placental structure and function.Hematoxylin-eosin staining(HE)showed that exposure to 1-NP at a high dose(100μg/kg)changed the structure of placenta and vascular lumen,and caused changes in the area of blood sinuses in the labyrinthic layer.q RT-PCR results showed that compared with solvent control,low-medium doses(25μg/kg and 50μg/kg)of 1-NP exposure down-regulated the expression of vascular endothelial growth factor receptor 1(Vegfr1),vascular endothelial growth factor receptor 2(Vegfr2),insulin-like growth factor 2(Igf2),and insulin-like growth factor 2 receptors(Igf2r).However,exposure to 1-NP at high doses(100μg/kg)up-regulated placental growth factor(Plgf),vascular endothelial growth factor α(Vegf-α),and Vegfr1.These results suggest that although 1-NP exposure during pregnancy can inhibit spiral artery recast,while high doses(100μg/kg)of 1-NP exposure may activate the compensatory mechanism and promote the release of placental growth factor,so as to reduce the damage to placental growth and development through compensatory mechanism,but break the balance of its gene expression.The expression of glucose transporter 1(Glut1)and fatty acid transporter 1(Fatp1)was down-regulated,and the expression of sodium-coupled neutral amino acid transporter 2(Snat2)was up-regulated,indicating that glucose and fatty acid transport were inhibited,while amino acid transport was increased.ConclusionHigh dose of 100μg/kg 1-NP during pregnancy inhibited trophoblast invasion and smooth muscle cells regression,and induced spiral artery remodeling defects.The gene expression of vascular growth factor Plgf,Vegf-α and its receptor Vegfr1 was up-regulated.The expression of Glut1 and Fatp1 genes was down-regulated,and the placental structure was changed.Thus,these results suggest that 1-NP exposure during pregnancy inhibits trophoblastic invasion,induces spiral artery remodeling defects,inhibits glucose and protein transporter expression,disrupts the balance of gene expression between growth factors and their receptors,and leads to changes in placental structure and function,ultimately leading to intrauterine growth restriction. |
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