Date of Graduation

5-2010

Document Type

Dissertation (PhD)

Program Affiliation

Biomedical Sciences

Degree Name

Doctor of Philosophy (PhD)

Advisor/Committee Chair

Dr. Yang Xia, MD, PhD

Committee Member

Dr. Jeffrey K. Actor, PhD

Committee Member

Dr. Michael R. Blackburn, PhD

Committee Member

Dr. Russell R. Broaddus, MD, PhD

Committee Member

Dr. Susan M. Ramin, MD

Abstract

Preeclampsia (PE) is a disease of late pregnancy characterized by maternal hypertension and proteinuria. It is associated with preterm delivery and significant perinatal morbidity and mortality. Despite affecting ~7% of first pregnancies, there is no effective screening method to identify women at risk, nor is there a definitive treatment other than delivery of the baby and placenta. Though the pathogenesis of PE remains unclear, an imbalance in the renin-angiotensin and immune systems are thought to be major contributors. Bridging these two concepts, it has recently been shown that women with PE harbor specific autoantibodies: the angiotensin II type 1 receptor activating autoantibody (AT1-AA). These autoantibodies act as angiotensin II and stimulate the ubiquitous AT1 receptor. To elucidate the role of AT1-AA in the pathophysiology of PE, a model of adoptive transfer was generated wherein AT1-AA isolated from human sera are injected into pregnant mice. This autoantibody incites the key features of the disease in pregnant mice: increased blood pressure, proteinuria, renal and placental abnormalities and increases in the anti-angiogenic factors soluble fms-like tyrosine kinase (sFlt-1) and soluble endoglin (sEng). These experiments were also carried out in non-pregnant animals who did not share the same symptoms. This suggests that pregnancy is a requirement for the full spectrum of preeclamptic features. In addition, the placentas and fetuses of AT1-AA-injected mice are reduced in size. The pups demonstrated intrauterine growth restriction (IUGR) and organ immaturity, especially in their kidneys and livers. To elucidate the mechanism by which these preeclamptic symptoms arise in the mouse model, the effects of increased inflammation were investigated. A multi-analyte screen indicated that the autoantibody induced inflammatory cytokines. TNF-alpha, a potent pro-apoptotic cytokine, known to be increased in both the sera and placentas of preeclamptic women, was most elevated, and was therefore the focus of further research. When incubated with AT1-AA, human placental explants greatly increased their production of TNF-alpha. In addition, TNF-alpha-mediated apoptosis was increased due to AT1-receptor activation in both the mouse placenta and human villous explants. Autoantibody-induced apoptosis and TNF-alpha production could be specifically reduced by co-treatment of AT1-AA with an anti-TNF-alpha antibody, losartan (an AT1 receptor blocker) or an antibody-neutralizing peptide, 7-aa. In order to test the pathophysiologic relationship between AT1-AA and TNF-alpha in vivo, the established adoptive transfer mouse model was employed. Co-injection of AT1-AA and an anti-TNF-alpha antibody reduced the features of PE in pregnant mice, implying an important pathogenic role for this cytokine. Overall, when injected into pregnant mice, AT1-AA induces the clinical features of PE and results in increased TNF-alpha production, placental apoptosis and fetal anomalies. Blockade of these features can be partially diminished by anti-TNF-alpha treatment, an AT1 receptor blocking drug, or a peptide which specifically neutralizes autoantibody action. Significantly, these findings could lead to a screening tool for preeclampsia as well as a potential therapeutic strategy for this life-threatening disease of mother and child.

Keywords

hypertension, autoimmunity, pregnancy, angiotensin, mouse models, inflammation, fetal

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