Study On Disturbance Of Glucose Transport In Placenta Of Conditionally Inducible Knockout Of Syncytin-a Gene Mouse

Objective:To explore the mechanism of Syncytin-a gene in placental and fetal development through conditionally inducible Syncytin-a gene knockout mice,and to reveal the possible relationship between fetal growth restriction and the decrease of placental glucose transport function caused by Syncytin-a deletion.Material and methods:1.Animal model and grouping:the male and female Cre ER^T2/Syn A^fl/fl mice were cohabited during 16:00 to 20:00,and then checked the virginal plugs at 8:00 the next day.The female mice with virginal plugs were fed in a single cage.Syncytin-a gene knockout of pregnant mice was induced by intraperitoneal injection of tamoxifen in E11.5（embryonic day11.5）in gene knockout group,and the mice continued to be raised until E17.5 and the placentas and embryos were collected at E17.5.The pregnant mice in normal group were raised as usual and then the placentas and embryos were collected at E17.5.2.Methods and contents:（1）The placenta and fetal phenotype（placental size,placental color,fetal morphology and fetal weight,fetal crown-rump length,number of dead embryos,etc.）were observed and recorded in E17.5 in normal group and knockout group.（2）The function of placental transport was detected by rhodamine123 accumulation assay;（3）The morphology and structure of placenta was observed by HE staining（hematoxylin-eosin staining）;（4）The ultrastructure of placenta was observed by electron microscope;（5）The micro-vessel density in labyrinthine layer of placenta was observed by CD34 immunohistochemical staining;（6）The localization and expression of GLUT1（glucose transporter 1）,GLUT3（glucose transporter 3）and Cx26（connexin 26）in placenta of the two groups were detected by immunofluorescence staining（semi-quantitative）;（7）The protein expressions of GLUT1,GLUT3 and Cx26 in the total placental proteins of the two groups were detected by western blotting.3.Data collection and software analysis:SPSS22.0 was used to analyze the experimental data,including analyzing the normality and homogeneity of variance,and t-test and nonparametric test were used to analyze and compare the differences between the two groups and multiple groups.P<0.05 indicates that the results is significant.Results:1.Phenotypic observation showed that in E17.5,the placentas of Syncytin-a knockout mice were paler than normal group,the placental weight was lighter than normal group,and the placental diameter was not significant difference between normal group and Syncytin-a knockout group.The embryos of Syncytin-a knockout mice showed partial or even total absorption,lighter body weight,shorter crown-rump length,higher mortality and the incidence of FGR（fetal growth restriction）,and decreased fetal/placental weight ratio.2.Rhodamine123 accumulation assay showed that the total placental transport capacity decreased in the knockout group compared to the normal group.3.The morphological study showed that in the knockout group,the area of the labyrinthine layer of placenta was decreased,the number of fetal blood vessels in the labyrinthine layer were decreased,and the structure of placental maternal-fetal exchange interface was destroyed,that unfused trophoblasts increased,the shape of cells in syncytiotrophoblast layer were irregular and couldn’t fit closely.4.The study of glucose transporters showed that compared with the normal group,the expression of glucose transporter in the labyrinthine layer of placenta was decreased in the knockout group,and the results of western blot showed that there was no significant difference of GLUT1,GLUT3 and Cx26 between normal and Syncytin-a knockout group.Conclusion:The deletion of Syncytin-a gene leads to the decrease of glucose transport between mother and fetus by reducing the placental glucose transport capacity,which might involve in embryonic malnutrition,even fetal growth restriction and intrauterine embryo death,but the molecular mechanism of gene deletion leading to the decrease of transport capacity needs to be further studied.