Preliminary Study Of The Placental Mechanism Of Insulinlike Growth Factor-1 Mediated Intrauterine Growth Restriction Induced By Nutrition Restriction

[Research Background]Fetal growth restriction refers to impaired and insufficient intrauterine growth potential caused by a variety of adverse factors,which is a serious perinatal complication leads to fetal or neonatal mortality and morbidity.FGR results from numerous causes and its pathogenesis has not been fully understood while recently more and more researchers has begun to focus on the placenta,the only link between fetus and mother.Placenta is a vital organ that play key roles in fetal development.Moreover,clinically there is lack of effective diagnosis,predictive or monitoring technique for FGR and there is no effective prevention or treatment for FGR which is diagnosed during pregnancy too.So there are calls for more studies for FGR.It has been confirmed that the IGF system plays an important role in the pathogenesis of FGR,which can also affect the placental structures and functions.Any adverse factor damages normal placental growth lead to placental insufficiency could directly damage fetal growth and development.During early period of placenta implantation,it is a vital for trophoblast invading into uterine decidua and muscle layer,finishing uterine spiral artery remodeling.It is a process that including a number of important proteins and cytokines,among which matrix metalloproteinase-2 is one of the most important enzymes that involved in trophoblast invasion.During trophoblast invasion,MMP2 could dissolve extracellular matrix and basilar membrane,promote vascular surface growth factors etc so as to stimulate trophoblast invasion and placental vascular remodeling.Besides,it has been proved that maternal undernutrition is a cause of fetal growth restriction,which may affect normal placental structure and function development.Therefore,we intend to firstly build FGR rat models by prenatal nutrition restriction and to examine the relationship of fetal or placental growth and placental IGF1 or IGFBP3 expression.So as to prove the possible link between link between placental or fetal growth and the IGFs.Furtherly we carried out cell experiments to verifiy our finding and hypothesis.By what is mentioned above,we would like to find the possible mechanism of placenta IGF system mediated fetal growth restriction induced by nutrition restriction.It may be help for understanding the pathogenesis of FGR and also to offer theoretical references for future early detection and treatment for FGR.Part I Changes of placental IGF1 and IGFBP3 expression in FGR rats model induced by prenatal nutrition restriction[Objectives]To build the FGR rat model by prenatal nutritional restriction and to investigate the effects of nutritional restriction on placental IGF1 and IGFBP3 expression.And also to explore the possible relationships between IGF1/IGFBP3 and FGR.[Methods]1.Firstly to build the FGR rat model by feeding with a protein restricted diet of 8%protein so as to observe the effects of nutrient restriction on the development of rat offsprings and placentas;2.To value the effect of nutrition restriction on placentas structure by HE staining and pathological examination;3.Fluorescence quantitative PCR was carried out for detection of placental IGF1 and IGFBP3 mRNA expression,immunohistochemistry staining for placental IGF 1 and IGFBP3 protein expression.[Results]1.Nutrition restriction lead to fetal rats growth restriction with smaller placentas and increased FGR rate(P<0.05);2.HE staining pathological examination showed that the placentas of research group were underdeveloped with disordered placental structure and narrowed labyrinth areas;3.Fluorescence quantitative PCR showed that comparing with the control group,placental IGF1 and IGFBP3 mRNA expression in the research group were significantly decreased(P<0.05);4.Immunohistochemical staining showed that IGF1 and IGFBP3 mainly express in trophoblast of placental basal layer and the labyrinth area.Compared with the control group,placental IGF 1 and IGFBP3 expression were down regulated in the research group.[Conclusions]Prenatal nutrition restriction impried normal fetus and placentas growth with downregulated placental IGF1 and IGFBP3 expressions,which may a cause leads to placental dysfunction and fetal growth restriction.Part ? Effect of FBS restriction on IGF-1 expression and cell invasiveness in human trophoblast cell line HTR-8/SVneo cells[Objectives]To explore the effect of FBS restriction on the expression of IGF1 and MMP2 and cell invasion ability of human trophoblast cell line HTR-8/SVneo cells.[Methods]1.HTR-8/SVneo cells were cultured with low concentration of FBS(1%),medium concentration of FBS(5%)and high concentration of FBS(10%),fluorescence quantitative PCR were carried out for detection of IGF1 and IGFBP3 mRNA expression,immunofluorescence to detect IGF1 and IGFBP3 protein expression in HTR-8/SVneo cells.By mentioned methods we tend to value the effects of different concentrations of FBS on HTR-8/SVneo cells IGF 1 and IGFBP3 expressions;2.Same treatment as mentioned above,fluorescence quantitative PCR were carried out to detect MMP2 mRNA expression,immunofluorescence for MMP2 protein and Transwell assays for cell invasiveness.3.Furtherly HTR-8/SVneo cells were with different concentration of human recombinant IGF 1,CCK-8 assay were carried out for cell growth,fluorescence quantitative PCR were carried out to detect MMP2 mRNA expression,immunofluorescence for MMP2 protein and Transwell assays for cell invasiveness.[Results]1.Along with the decreasing FBS concentrations,cell proliferation and invasion ability decreased significantly(P<0.05);2.FBS restriction inhibited mRNA and protein expression of IGF1 and MMP2 of HTR-8/SVneo cells(P<0.05);3.rhIGF1 could significantly up-regulate MMP2 expression of in HTR-8/SVneo cells(P<0.05)and promote HTR-8/SVneo cells invasion(P<0.05).[Conclusion]FBS restriction downregulated IGF1 and IGFBP3 expression in human trophoblast cells line HTR-8/SVneo cells,which might lead to decreased expression of MMP2 in HTR-8/SVneo cells and thereby inhibit cell invasiveness.Suplement with rhIGFl contributes to the upregulated expression of MMP2 in HTR-8/SVneo cells and promote cell invasion.