Division of Reproductive, Stem Cell and Perinatal Biology

Overall Question of Interest
The Winn Laboratory research program revolves around understanding the process of normal and abnormal human placentation. Acquiring knowledge about human placentation will provide important insights about pregnancy complications such as preeclampsia, intrauterine growth restriction (IUGR), placenta accreta and gestational trophoblastic disease. As a physician scientist, my ultimate goal is to see this knowledge translate into improved clinical care resulting in healthier mothers and infants. Current projects include:

Dissecting the role of pre-clampsia related molecules on cytotrophoblast differentiation and invasion.

The ultimate goal of this project is to shed light on the pathophysiology of preeclampsia; a pregnancy complication characterized by maternal hypertension, proteinuria and edema with the hope of providing better diagnostic markers and improved therapeutic approaches. The underlying pathophysiology of preeclampsia still remains elusive although recent advances have been made with the identification of the role of sFlt-1 and sEndoglin. One clearly established principle is that the placenta plays a pivotal role. Preeclampsia only occurs in the presence of trophoblastic tissue and the clinical condition resolves with the delivery of the placenta. The placental basal plate is the site of known anatomical abnormalities in preeclampsia. Shallow CTB invasion and limited remodeling of the uterine spiral arteries is noted in the basal plate region of preeclamptic placentas. Hence, this region has been the focus of our studies. We have recently determined the gene expression profile of this region from normal second trimester and term samples utilizing microarray technology (Affymetrix GeneChips). This analysis has identified those genes that are expressed in this region and those which have altered expression over gestation. Additionally, we have determined the gene expression profile of the basal plate region from preterm preeclamptic placentas and preterm labor placentas serving as controls. Analysis of the resulting gene profiles has identified potential pathways and molecules altered within the basal plate of preeclampsia. Our hypothesis is that those differentially expressed genes in preeclampsia that are not also regulated over gestation may contribute to the abnormal CTB invasion seen in preeclampsia. Ongoing projects in the lab are confirming the expression pattern of the identified molecules and to determine which cell-type(s) is (are) expressing the molecules of interest (i.e. CTBs, decidua or immune cells) in the basal plate. This is being done by using immunohistochemistry where antibodies are available or alternatively by in situ hybridization on banked basal plate biopsies from normal, preterm labor and preeclamptic placentas. Then, the functional significance of molecules in regards to CTB invasion will be determined. Functional perturbation studies using our in vitro model systems (as discussed above). Perturbations will be performed by using either blocking/stimulating antibodies where appropriate or by transfection using adeno/lentiviral systems to either over-express or knockdown expression of the target gene. Ultimately the results of these studies should identify key molecules and molecular pathways that could potentially serve as therapeutic targets for the prevention and/or treatment of preeclampsia.


Impact of Preeclampsia on Endothelial Progenitor Cell Biology
Preeclampsia (PE), a pregnancy-specific disease, affects 8 million women-infant pairs annually world-wide resulting in significant morbidity and mortality for both mothers and infants. The only curative treatment for PE is delivery of the inciting organ--the placenta, often at the expense of a premature infant. The immediate and chronic morbidities of premature birth are well known. However, increasing evidence suggests that PE may have a long-term impact on the infant (i.e. fetal origins of adult disease) and is associated with increased risk of later cardiovascular events for the mother and offspring.
A central feature of PE pathogenesis is maternal endothelial cell dysfunction. While much has been accomplished in characterizing the systemic endothelial dysfunction of preeclampsia, the role of endothelial progenitor cells (EPCs) in the mechanism is not fully understood and may provide a potential cellular target for novel therapies. Endothelium adaptations during pregnancy include the release of vasoactive agents such as nitric oxide (NO) to increase blood flow to the uteroplacental unit, and endothelium-mediated relaxation resulting in a fall in peripheral vascular resistance. These changes in endothelial function can be measured by brachial artery flow mediated dilation (FMD), which is a non-invasive ultrasound based technique. Studies have shown that FMD is reduced in woman with PE compared to controls (1). More recently endothelial function can be measured by EndoPAT, which has been used to assess function in pregnancy and PE (Carty 2014).

It was recently shown that maternal circulating EPCs increase during normal pregnancy but are reduced in women with preeclampsia suggesting a maladaptation in EPC biology related to PE pathogenesis (2). The concept that alterations in EPC biology impacts endothelial cell function is supported by recent studies showing that decreased numbers of circulating EPCs correlate with impaired endothelial function as measured by FMD (3, 4). If this connection holds true for PE then learning how to modulate EPCs may open up novel treatment modalities.

Therefore, we hypothesize that impairment in EPC biology contributes to the maternal endothelial cell dysfunction in preeclampsia and that similar impairments occur in fetal EPCs, which may impact neonatal outcomes.

We are testing this hypothesis through the following specific aims:

Aim 1) Determine if alterations in EPC biology correlate with the endothelial cell dysfunction in preeclampsia. This will be assessed by characterizing maternal EPC subtype numbers and function using flow cytometry, culture and functional assays. In parallel with EPC characterization, EndoPAT measurements will be analyzed in both preeclamptic and normotensive pregnancies in the third trimester and postpartum after resolution of the disease.

Aim 2) Investigate the impact of preeclampsia on fetal EPC biology. This will be assessed by characterizing venous cord blood EPC subtype numbers and function by flow cytometry and culture assays from pregnancies complicated by PE compared to controls. We will then correlate these results to the maternal EPC phenotypes.
Gaining insight into the role of EPCs in the mechanism of PE endothelial cell dysfunction may reveal not only potential preventative or therapeutic strategies geared at modulating EPCs but may provide a measure for the long-term impact of PE on maternal and neonatal health related to EPC biology.

Pregnancy Induced Rheumatoid Arthritis Amelioration.

Another more recent research direction in my laboratory is exploring the mechanism of pregnancy-induced amelioration of rheumatoid arthritis, the hypothesis being that the mechanism resulting in amelioration is likely the same mechanism employed for the establishment of maternal-fetal tolerance. In collaboration with Dr. Dragone we have established a murine model to study this biological process. Our first manuscript reporting on the model and the identification of a serum biomarker, SSA3, to use to measure inflammatory state is listed.

a. Shaw, LA, AL Stefanski, KK Rumer, A Vondracek, TL Phang, S Sakaguchi, VD Winn* and LL Dragone. (2012) Pregnancy Amelioration of Arthritis in SKG Mice Corresponds with Alterations in Serum Amyloid A3 levels. Am J Clin Exp Immunol. 1(1):12-19 (*co-senior author). PMID 23097751.

Biology of endothelial progenitor cells along pregnancy and postpartum

In collaboration with Dr. Frauke von Versen-Höynck, visiting assisted professor from Germany and Dr. Valerie Baker, Associate Professor of Obstetrics and Gynecology (REI) we study endothelial health and the biology of endothelial progenitor cells in our cohort along pregnancy and postpartum. While comparing the effects of different treatments and the number of corpora lutea during these treatments with regards to endothelial function we will better understand the effect of assisted reproduction on maternal and fetal physiology with the long-term goal to use the gained information to improve current treatments and pregnancy outcomes.