Regulation of Angiogenic Gene Expression in Human Trophoblast by Cellular Stressors: Implications in Preeclampsia

Author

Mark Wesley Schroeder, Southern Illinois University CarbondaleFollow

Date of Award

8-1-2017

Degree Name

Master of Science

Department

Molecular Biology Microbiology and Biochemistry

First Advisor

Torry, Donald

Abstract

Preeclampsia (PE) affects 5-8% of pregnancies worldwide and is one of the leading contributors of maternal and fetal morbidity and mortality during pregnancy. Its high prevalence coupled with its significant contribution to health crises during pregnancy emphasizes the importance of research and development of a cure to its clinical pathology. It is believed that insufficient remodeling of the maternal spiral arteries of the uterus lead to decreased perfusion of the placenta and a relatively hypoxic, inflammatory, and endoplasmic reticulum stressed placenta. This hypoxic stress has been shown to decrease the secretion of the proangiogenic molecule placental growth factor (PlGF), as well as increase the production of its antiangiogenic decoy receptor, soluble fms-like tyrosine kinase (sFlt1) by trophoblast. This results in what is commonly referred to as the “angiogenic imbalance of PE.” Hypertension and proteinuria are the clinical manifestations of PE and represent the effect of decreased systemic vasodilation and compromised glomerular endothelial integrity in the mother caused by the angiogenic imbalance. Recently, is has been shown that endoplasmic reticulum stress in the syncytiotrophoblast of the placenta correlates with a down regulation of PlGF protein expression by these cells. The molecular mechanisms which cause the decrease in PlGF production by trophoblast during PE are not known. The aims of this thesis were to elucidate whether hypoxia, inflammation, and ER stress lead to decreases in the expression and function of one of the main transcription factors of PlGF in trophoblast, glial cell missing-1(GCM1). Furthermore, we wanted to know if these insults worked in conjunction or independently of one another to decrease GCM1 and PlGF expression. Treatment of JEG3 choriocarcinoma cells with inflammatory cytokine TNFα and overexpression of pro-inflammatory transcription factor, NFκBp65, did not significantly alter the expression of GCM1 or PlGF mRNA. Treatment of JEG3 cells with ER stress inducer tunicamycin significantly decreased GCM1 and PlGF mRNA expression. ER stress induction in JEG3 cells transduced with a 1.5kb 5`UTR of PlGF linked to a GFP reporter significantly decreased GFP expression. These results suggest that ER stress decreases transcription in the 1.5kb 5`UTR of PlGF. This region of the 5`UTR of PlGF was recently shown to be under transcriptional regulation by GCM1. Finally, 1%O2 treatment of JEG3 choriocarcinoma cells significantly decreased GCM1 and PlGF gene expression while significantly increasing the expression of an ER stress marker, CHOP. These data indicate that hypoxia can lead to ER stress in trophoblast which can decrease GCM1 expression. As GCM1 is a principle transcription factor of PlGF in trophoblast, this results in decreased transcription and expression of PlGF. Thus, inhibiting the occurrence of ER stress in trophoblast of preeclamptic pregnancies, possibly though drug delivery to the preeclamptic mother, may provide a novel therapeutic approach. By reducing ER stress of trophoblast we hypothesize that expression of PlGF will be increased in maternal serum. This may function to correct the “angiogenic imbalance of PE” and may alleviate preeclamptic symptoms, allowing for prolongation of pregnancy and therefore decreased risk of maternal and fetal morbidity and mortality.