NFκB REGULATION OF PLACENTA GROWTH FACTOR AND ITS PRIMARY TRANSCRIPTION FACTOR, GLIAL CELL MISSING 1

Author

Timothy Murphy, Southern Illinois University CarbondaleFollow

Date of Award

8-1-2014

Degree Name

Master of Science

Department

Molecular Biology Microbiology and Biochemistry

First Advisor

Torry, Donald

Abstract

Preeclampsia is one of the most common complications of human pregnancy. Though its pathophysiology is not fully understood, it is thought that shallow migration of cytotrophoblast and the subsequent insufficient conversion of the maternal spiral arteries into utero-placental arteries play important roles. As fetal demands for oxygen and nutrients increase during gestation, this aberrant remodeling of the vasculature at the maternal-fetal interface results in a relatively hypoxic and nutrient-deficient placenta bed. Hypoxia is known to decrease placenta growth factor (PGF) expression in trophoblast and may play a key role in the suppression of PGF levels associated with preeclampsia. Systemic vasodilation, also critical to facilitate a healthy pregnancy, is significantly correlated with higher serum levels of PGF. Dysregulation of certain pro-inflammatory pathways, including NFκB, has also been implicated in preeclampsia development. The goal of our studies was to determine the mechanism governing NFκB regulation of PGF in trophoblast. Over-expression of NFκBp65, an active subunit of the NFκB complex, significantly decreases PGF mRNA expression in trophoblast - an effect which is ablated by co-expression of a dominant negative (dn) IκB which inhibits NFκB complex activity. Similarly, over-expression of NFκBp65 functionally inhibits glial cell missing 1 (GCM1), the primary transcription factor of PGF in trophoblast. Previous reports show that GCM1 protein is degraded during hypoxia. Because hypoxia and activation of NFκB have similar effects on both PGF and its transcription factor, GCM1, we have investigated the possibility of cross-talk between these two pathways. Over-expression of NFκBp65 was able to inhibit GCM1 protein stability and expression, but transient transfection with dnIκB was unable to prevent hypoxia-induced inhibition of PGF reporter activity. Collectively, these results indicate that hypoxia and NFκB activation utilize independent pathways to decrease PGF expression in trophoblast. To facilitate future investigation of these pathways, we have also established a novel stable cell line which can be used to induce high levels of NFκBp65 expression in a controllable manner, and observe its effects without the need for co-transfection.