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Research Programs: Uterine, Placental and Fetal Physiology

Transcriptional Regulation of Placental Hormones

The placenta is directly responsible for mediating and/or modulating the maternal environment necessary for normal fetal development. As an active endocrine organ, the placenta is capable of secreting a plethora of hormones, growth factors and cytokines. Some of these are true placental hormones since they are not produced by other organs and have known or inferred functions during pregnancy. Included in this category are members of the growth hormone (GH) - prolactin (PRL) gene family. Placental members of this family have been implicated in modulating maternal and fetal metabolism, luteal function and mammogenesis.

We have characterized the biosynthesis and structure of one member of this family, ovine placental lactogen (oPL), and have structurally characterized the gene encoding oPL. The 5'-flanking region of the oPL gene is being examined to determine what cis-acting elements and trans-acting factors are responsible for its expression only by chorionic binucleate cells. We now know that this regulation involves a splice variant of activator protein-2a, GATA-2, and at least two other transcription factors not previously identified in the placenta. The first of these is PURa, a single-strand DNA-binding protein that is involved in the transcriptional regulation of neural genes. We are in the process of identifying the other trans-acting factor that interacts with the oPL gene providing binucleate cell-specific expression. Additionally, we are generating transgenic sheep pregnancies, by nuclear transfer of stably transfected fetal fibroblast cells into enucleated oocytes, to determine what domains of the oPL gene 5'-flanking sequence are required for chorionic cell-specific expression in vivo.

Binucleate cell image

The image to the right shows dual immunostaining of an ovine placentome for ovine placental lactogen in purple and the transcription factor AP-2 (activator protein-2) in brown. These results indicate that binculeate cells (red arrow) contain AP-2, but are not the only cells in the fetal placenta containing AP-2.

It is now known that the utero-placenta of sheep express GH gene for a limited period of time during early pregnancy (days 35 to 55 post-coitus). This time period of oGH gene expression coincides with the period of maximal placental development during sheep pregnancy. In our model of placental insufficiency-intrauterine growth restriction (PI-IUGR, see below), oGH expression is virtually abolished, implicating its role in normal placental growth and development. The gene expressed in the utero-placenta is the same gene expressed in the anterior pituitary, raising the question as to what allows this gene to be expressed for only a defined period of time, by non-pituitary tissue. We are beginning to address this question by in vitro transfection and DNase I footprinting experiments. Combined, our studies will not only allow us to understand the transcriptional regulation of the oPL and oGH genes, but will provide insight into transcriptional regulation of other genes expressed solely by the placenta and their respective functions.

Participants: Russell V. Anthony, Sean W. Limesand, Kimberly M. Jeckel, Amy S. Erickson and Meredith Holtzen

Recent publication: Liang R, Limesand SW, Anthony RV. 1999. Structure and transcriptional regulation of the ovine placental lactogen gene. Eur J Biochem 265:883-895.

Placental Function and Fetal Development

The placenta is a multifaceted organ that plays critical roles in maintaining and protecting the developing fetus. These roles include nutrient transfer from the mother to the fetus and waste secretion from the fetus to the mother, acting as a barrier for the fetus against pathogens and the maternal immune system, and as an active endocrine organ. Placental insufficiency-intrauterine growth restriction (PI-IUGR) is a major cause of increased perinatal morbidity and mortality in humans, and has recently been tied to the predisposition of adult onset of diabetes, hypertension, stroke and coronary heart disease. Using a pregnant sheep model of PI-IUGR, we are studying the vascular and endocrine development of the placenta in PI-IUGR pregnancies. The overall aim of these studies is to determine what aspects of placental development are insufficient or incorrect, thereby setting the stage for an IUGR outcome. It is our hope that, through these basic studies, key developmental mechanisms can be evaluated, such that potential interventive methods can be derived.

We have determined that placental vascular structure is altered, and that expression of vascular endothelial growth factor is enhanced in early PI-IUGR pregnancies. Utero-placental expression of GH is virtually abolished, at the time of maximal placental growth. Maternal concentrations of progesterone and oPL are depressed in PI-IUGR pregnancies, although the ability of chorionic binucleate cells to synthesize these hormones is not altered, indicating that the ability of these cells to migrate and fuse with the maternal syncytium is impaired. Regardless of the cause, the endocrine environment of PI-IUGR pregnancies is altered, thereby interfering with normal placental and fetal development. Coinciding with these efforts is our work on developing "knockout" mice in which either the mouse placental lactogen I or II genes have been functionally disrupted. We are also developing similar approaches in sheep that will be used to functionally disrupt the oPL gene.

Scanning electron micrographs of fetal-placental vascular casts

The image above depicts scanning electron micrographs of fetal-placental vascular casts. (A) Drawing of a sheep placentome, showing the approximate location of fetal placental villi from which the photomicrographs were obtained. (B) Low power magnification (x100) of fetal vasculature extending from the intermediate zone (top) to the basal plate zone (bottom). (C) Normal fetal-placental vasculature in the intermediate zone at higher magnification (x1,000) showing organized outgrowth of vessels which remain relatively straight. (D) Fetal-placental vasculature (x1,000 magnification) in the intermediate zone derived from an intra-uterine growth restricted pregnancy. Note the increase in vessel coiling and overall disorganized arrangement of vessels. An increase in the number of sinusoidal dilations is also observed.

Participants: Russell V. Anthony, Sean W. Limesand, Timothy R.H. Regnault and Randall B. Wilkening.

Recent publication: Regnault TRH, Battaglia FC, Wilkening RB, Anthony RV. 1999. Altered arterial concentrations of placental hormones during maximal placental growth in a model of placental insufficiency. J Endocrinol 163:433-442.