目的:妊娠期高血压疾病(HDP)是一种常见的妊娠期疾病。NANOG和细胞周期蛋白依赖性激酶1(CDK1)对于调节细胞增殖和凋亡的功能至关重要。然而,HDP的作用机制尚不清楚.
方法:从GEO数据库下载微阵列数据集GSE6573。Emt相关基因集从上皮-间充质转化基因数据库下载2.0,通过生物信息学分析筛选差异表达基因。PathwayCommons和Scansite4.0数据库用于预测蛋白质之间的相互作用。从HDP患者和怀孕顺利的患者收集胎盘组织样品。RT-qPCR,Westernblot和免疫组化检测NANOG的表达,CDK1、MMP-2、MMP-9、EMT标志物和JAK/STAT3通路蛋白。转染NANOG过表达/敲低,和CDK1敲低进入人绒毛膜滋养层细胞(HTR-8/Svneo)。CCK-8,Transwell和伤口愈合试验用于评估细胞增殖,入侵和迁移。CO-IP和GST下拉测定用于确认蛋白质相互作用。
结果:共获得7个EMT相关差异表达基因,其中NANOG,NODAL和LIN28A存在蛋白质相互作用。在HDP患者组织中发现NANOG和CDK1表达较低。NANOG过表达促进HTR-8/Svneo增殖,迁移和EMT,而NANOG击倒则有相反的效果。此外,STAT3和CDK1与NANOG之间的蛋白质相互作用。NANOG过表达下调JAK/STAT3通路促进HTR-8/Svneo增殖,迁移和EMT,被CDK1敲低逆转。
结论:NANOG下调JAK/STAT3通路促进滋养细胞增殖,通过蛋白质与CDK1相互作用进行迁移和EMT。
OBJECTIVE: Hypertensive disorders of pregnancy (HDP) are a common pregnancy disease.
NANOG and Cyclin-dependent kinase 1 (CDK1) are essential for regulating the function of cell proliferation and apoptosis. However, the mechanism of action in HDP is yet unclear.
METHODS: The microarray dataset GSE6573 was downloaded from the GEO database. Emt-related gene set was downloaded from Epithelial-Mesenchymal Transition gene database 2.0 were screened differentially expressed genes by bioinformatics analysis. Pathway Commons and Scansite 4.0 databases were used to predict the interaction between proteins. Placental tissue samples were collected from HDP patients and patients with uneventful pregnancies. RT-qPCR, Western blot and immunohistochemistry were used to detect the expression of
NANOG, CDK1, MMP-2, MMP-9, EMT markers and the JAK/STAT3 pathway proteins. Transfection NANOG overexpression/knockdown, and CDK1 knockdown into the human chorionic trophoblast cells (HTR-8/Svneo). CCK-8, Transwell and Wound-healing assay were used to evaluate cell proliferation, invasion and migration. CO-IP and GST pull-down assays were used to confirm the protein interaction.
RESULTS: A total obtained seven EMT-related differentially expressed genes, wherein NANOG, NODAL and LIN28A had protein interaction. In the HDP patients\' tissue found that
NANOG and CDK1 had lower expression.
NANOG overexpression promoted HTR-8/Svneo proliferation, migration and EMT, while
NANOG knockdown had the opposite effect. Further a protein interaction between STAT3 and CDK1 with
NANOG. NANOG overexpression downregulated the JAK/STAT3 pathway to promote HTR-8/Svneo proliferation, migration and EMT, which was reversed by CDK1 knockdown.
CONCLUSIONS: NANOG downregulated the JAK/STAT3 pathway to promote trophoblast cell proliferation, migration and EMT through protein interaction with CDK1.