Mother Infant Research Institute (MIRI)

The Wallingford Lab MAP project investigates developmental mechanisms that control subsequent placental blood vessel formation, vascular remodeling, and vascular mimicry during placentation. Dynamic vascular remodeling of the mouse placenta is well exemplified by examination of trophoblast, vascular and structural markers (Figure 1). Building on Dr. Wallingford’s previous peri-implantation development work (Wallingford et al 2013), the MAP group applies vascular morphometric and cellular phenotyping to developmental models and tractable 3D culture systems in order to determine how blood vessel formation, vascular remodeling, and vascular mimicry are controlled at the molecular level.

Figure 1. Vascular Remodeling in Mouse Placentation. A. Implanting mouse embryo at E4.0. Laminin (LMN) highlights vascular structures in the decidua and E-Cadherin (CDH1) highlights the breakdown of uterine luminal epithelium. B-C. E9.5 Mouse placenta labelled with antibodies against Slc20a2, trophoblast cytokeratin 7 (CK7), LMN and endothelial von Willebrand’s Factor (vWF) localization.  E-H. E15.5 placenta with PiT-2, CK7, LMN, vWF, and pericyte (CD13) localization. 

The placenta is the least understood human organ partly because access to the developing tissue is limited and there are marked differences in comparative anatomy between eutherian species. A recent publication from the NIH reported that pregnant women and their babies are frequently viewed as a vulnerable patient population, and as a result they have been relatively underserved and functionally neglected in biomedical research – constituting a deprived population in need of more effective therapeutic interventions. New approaches are greatly needed to advance clinical care options. 

One way in which our lab is directly addressing this need is by pioneering the development of 3D culture systems that model placental vascular development. We aim to combine embryology and bioengineering techniques to generate tractable tools that better recapitulate the physiology of in vivo systems. We are developing a 3D cell culture approach to investigate how vascular tissue and specialized niches in the placenta develop and function in order to more accurately and reliably model physiological placental vascular development and generate novel data they may be used to improve biomedical understanding of human placental development and disease, including preeclampsia (Figure 2). As we expand our systems, our lab would gladly serve as a unique resource to local investigators who wish to expand into collaborative placenta research.

Figure 2. Application of 3D Culture Systems to Advance Reproductive Health. Complimentary 3D cell culture vascular models will be used to examine the interaction of four developmental pathways on trophoblast vascular mimicry and patterning. Impact on vascular morphometrics and cellular phenotyping will be determined by multivariate analysis. Future applications are numerous, including analysis of genetic variant function, developmental therapeutics, and other investigations.

Phosphate is an essential molecule that plays numerous roles in human health.  It is a primary component of the mineral that our bones are made of, but it also performs many other important functions.  While a large body of scientific research describes phosphate biology in the adult, very little is known about how phosphate gets to the developing baby.  The Wallingford Lab M-FPT project is closing this knowledge gap by defining maternal-fetal phosphate transport routes, testing hypotheses on the basic science of phosphate transport across the placenta, and developing new biological assays for maternal-fetal phosphate homeostasis that may one day be useful for assessment and interpretation of maternal and fetal health in the clinic.  Their work has identified a critical role for the placental sodium-dependent phosphate transporter Slc20a2 in fetal growth and development and protection from placental vascular calcification (Wallingford et al. 2016 and Yamada and Wallingford et al. 2018a). LacZ expression driven by the Slc20a2 promoter highlights physiologically relevant expression domains of Slc20a2 in the mouse placenta (Figure 3).  Coinciding with initiation of impaired fetal bone ossification, Slc20a2 mice develop increased levels of ectopic mineralization in Slc20a2 deleted expression domains, providing the first genetic link to placental calcification.  Clinically, vascular calcification in the placenta has been observed, but the biomedical significance of placental calcification is poorly understood, and may underlie hemodynamic alterations in conditions such as preeclampsia, placental thrombosis, and intrauterine growth restriction. The biomedical significance and prognostic value of placenta calcification are discussed in a recent publication from our lab which also reports on novel microcalcification patterns in human placenta (Wallingford et al. 2018b). 



 

Figure 3. Slc20a2 is expressed at the mouse maternal-fetal interface. Regional Slc20a2 expression during mouse placentation is indicated in blue. Slc20a2 expression detected by beta-galactosidase staining of Slc20a2-driven LacZ was observed in the ectoplacental cone, chorion, allantois, parietal endoderm, fetal endothelial cells, trophoblasts, and pericytes.

In future work the M-FPT project will evaluate actions of Slc20a2 at the molecular level during placentation and embryonic development.  This project is supported by NIH/NICHD R00HD090198 (PI: Wallingford, MC) Determination of maternal-fetal phosphate transport mechanisms and the role of sodium-dependent phosphate transporters in extraembryonic tissues and has previously received support from NIH/NHLBI T32HL007828 (PI: Dichek, D) Phosphate Transporters and Cardiovascular Development and Disease in the Mouse.

The PVS-FR project is a suite of beta testing initiatives coordinated by Dr. Wallingford.  PVS-FR initiatives and collaborators include:

• New Approaches to Placental Ultrasound Interpretation 
  -Manjiri Dighe, Department of Radiology, University of Washington 
  -Ezgi Mercan, Seattle Children’s Research Institute 
• Evaluation of Triad Ovine Physiology (Maternal, Placentome, Fetal). Martin Frasch, Department of Obstetrics, University of Washington
• Quantitative Placenta Phenotyping Assessment. Murat Maga, Center for Developmental Biology and Regenerative Medicine, Seattle Children’s Research Institute
• Development of Noninvasive Imaging Techniques. Howard Chen, Molecular Cardiology Research Institute (MCRI), Tufts Medical Center.

 
Neonatal outcomes are of utmost importance for overall health and quality of life in a given community. Developmental outcomes set the stage for long-term health of offspring and influence increased risk for adverse health conditions and significant global health challenges, including metabolic disorders, cardiovascular disease, and chronic kidney disease.

1. Physicians and scientists have begun to meet the challenge of advancing mechanistic understanding and treatment options for neonatal patients by developing rapid genomic sequencing and analysis methods that identify human genomic variants linked with specific neonatal outcomes. These groundbreaking multi-center initiatives, such as the Genomic Medicine for Ill Neonates and Infants (GEMINI) led by former MIRI Executive Director Dr. Jill Maron have provided an immense genetic foundation for the development of precision medicine approaches. However, before this genetic information can be truly harnessed, the newly identified variants must be functionally validated and characterized.
   
   
2. In order to meet this need, MIRI investigator Dr. Mary Wallingford has spearheaded the establishment of Development in situ (DEV-IS), which is an emerging in vitro-in vivo research platform for the study of pregnancy and mammalian development at Tufts Medicine. The DEV-IS platform is supported by a collaborative network of Tufts investigators who have established cutting edge methods, resources, and expertise that, in combination, are ideally suited for in depth functional characterization and pharmaceutical assessment of human genomic variants that regulate human health and development. Developmental progression and tissue-specific effects in vivo are established through the use of specialized mouse embryology research approaches in the Wallingford Lab (Tufts Medical Center MIRI). Research is directed towards translational relevance to human pregnancy health through analysis of comparative multi-omics datasets and by critical guidance and resources provided by MIRI Interim Executive Director Dr. Perrie O'Tierney-Ginn . These methods and resources are coupled with real time in vitro assays and therapeutic drug treatment protocols are used to evaluate cell-type specific efficacy of candidate treatment strategies in the Pulakat Lab (Tufts Medical Center MCRI). Mechanistic questions regarding parental genetic and epigenetic contributions are pursued in collaboration with Dr. Larry Feig (Tufts University School of Medicine GSBS). Fetal and placental health are studied through noninvasive body imaging approaches including ultrasound (US) in the Blanton Lab   (Tufts Medical Center MCRI), laser speckle contrast imaging (LSCI) in the Good Lab (Tufts Medical Center MCRI), and photoacoustic ultrasound (PAI) in the Mallidi Lab (Tufts University Bioengineering).

Overall, the DEV-IS platform is poised to have an immense impact on uncovering essential gene and gene variant functions that are crucial for a healthy pregnancy and healthy perinatal development. Ongoing DEV-IS pilot programs pursue developmental mechanisms regulated by: 1) maternal dietary protein intake; 2) phosphate homeostasis and placental phosphate transport; 3) genes causative for the vascular development disorder HHT (hereditary hemorrhagic telangiectasia); 4) critical cardiovascular genes identified through the GEMINI study; and 5) mRNA splicing and congenital birth defects.