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Wallingford Lab Presentations at 2021 MCRI Retreat



Dr. Mary Wallingford is a Principal Investigator in the Mother Infant Research Institute (MIRI) who holds a dual appointment in the Molecular Cardiology Research Institute (MCRI). Her laboratory team includes Dr. Ana Correia-Branco, Nirmala Jayaraman, Vineetha Mathew, Radhika Sharma and Hamida Giwa, as well as Aaron Zou who is co-mentored by Dr. Robert Blanton, a cardiologist and physician-scientist in the MCRI.


The Wallingford Lab group presented three abstracts at the 2021 MCRI Retreat which was held in Woods Hole, MA from Sept 29-Oct 1. Their work, linked below, sheds new insight into mechanisms of placental vascular development and disease. Their comprehensive developmental mouse placenta proteome breaks new ground (Kashpur et al.)*, and is the first of its kind to identify temporal abundance patterns, global trends, and confirmed molecular targets. Some of the identified proteins have also been revealed by the lab to regulate vascular morphogenesis in the placenta, such as Endoglin which is associated with the vascular development disorder hereditary hemorrhagic telangiectasia (HHT) (Mei et al.),** as well as Slc20a2 which is required for homeostatic balance of phosphate homeostasis and molecular ER stress pathways in placenta (Correia-Branco et al.).***

The Wallingford group pursues biomedical and basic science research questions in the area of vascular biology of pregnancy (obstetric cardiovascular biology). From the vascular perspective, extensive remodeling and de novo vascular development is integral to a healthy pregnancy. Multiple systemic vascular processes occur in concert during pregnancy, ranging from maternal cardiovascular remodeling, to placental organogenesis, to development of the fetal cardiovascular system.  Placental vascular development and vascular disease are of particular interest to the lab, as the placenta is a highly vascularized organ that forms de novo, mediates maternal-fetal transport of nutrients, oxygen, and waste, and produces cells and molecules that impact maternal cardiovascular health.


Mary Wallingford

The developmental mouse placenta proteome identifies dynamic temporal changes.

Olga Kashpur1, Shiori Kuraoka2, Hideyuki Higashi2, Ariel Mei1, Andrew Joe1, Elena Aikawa2, Sasha Singh2, Mary Wallingford1,3

1. Mother Infant Research Institute, Tufts Medical Center, Boston, MA

2. Center for Interdisciplinary Cardiovascular Sciences, Brigham and Women's Hospital, Harvard Medical School, Boston, MA

2. Molecular Cardiology Research Institute, Tufts Medical Center, Boston, MA


Placental dysfunction is associated with adverse maternal and fetal clinical outcomes, such as preeclampsia, fetal growth restriction, and preterm birth. In the case of preeclampsia, a life-threatening pregnancy-specific hypertensive disorder, abnormal vascular remodeling of placental blood vessels is frequently observed. However, the developmental origin(s) that this cause abnormal remodeling are incompletely understood at the molecular level. Furthermore, no curative treatments nor early diagnostic biomarkers are available for preeclampsia. We propose that advanced understanding of how healthy placental blood vessels normally develop, grow, and age will better enable the preeclampsia research field to design future studies that determine when and how preeclamptic cases diverge from health cases.  In this study we employed proteomic analyses using tandem mass spectrometry to study temporal changes in mouse placenta protein abundance levels across the entirety of gestation (embryonic day (E) 8.5-E17.5). Mouse placenta samples and their progenitors (allantois and chorion) were collected at 12h intervals between E8.5-E12.5 and 24h intervals between E12.5-E17.5 (n=3/independent liters per stage, N=45). Decidua was removed to enrich for placental progenitors and the developing maternal-fetal interface (MFI). In total, 3792 distinct proteins were detected. Proteomes differed principally in accordance with gestational age. Protein abundance kinetics using the mClust function of Xina identified groups of co-occurring proteins with similar temporal abundance patterns. Functional enrichment analysis (GO and GSEA) identified biological processes, developmental stage-specific protein networks that characterize mouse placentation. Overall, temporal proteomic analyses identified a dynamic continuum between placental development and placental aging. In conclusion, we propose that gestational age is a critical factor across the entirety of pregnancy that must be carefully considered in study design, data interpretation, and the potential influence of environmental effects on placental and fetal health.



Identification of Placental Vascular Malformations in Alk1 and Eng Null Mice

Vineetha Mathew

Ariel Mei*, Vineetha Mathew*, Olga Kashpur, Molly Mann, Andrew Joe, Jacqueline Garcia, Shreyas A. Bhave, Miranda E. Good, Navin Kapur, Mary C. Wallingford

The placenta is a transient organ crucial for sustaining fetal development by proper nutrient, gas, and waste exchange between the maternal and fetal circulations. This study aimed to characterize preclinical mouse models of pregnancy and placentation in hereditary hemorrhagic telangiectasia (HHT), an autosomal dominant disease distinguished by malformed blood vessels. Previous studies have identified crucial roles for HHT-associated genes Eng, Smad4, Alk1, and Bmp9 in vascular health, but their function in the placenta remains poorly understood. The study presented herein evaluated expression of HHT genes in placenta through histologic, proteomic, and in silico approaches. Fetal viability and maternal reproductive capacity were assessed for Eng (HHT1), Alk1 (HHT2), and Bmp9 (HHT5) null mouse strains. Placental morphogenesis was also evaluated at E9.5 in Eng and Alk1 wildtype (WT) and knockout (KO) mice. Proteomics analyses, Reactome data, and immunofluorescence findings supported that: 1) ENG, ALK1, and BMP9 are present in the placenta throughout development, 2) ENG colocalizes with CD31 in both placental and decidual endothelial cells and 3) ALK1 is abundant in the placenta, far exceeding expression levels observed in other organs. In agreement with published studies, global Eng or Alk1 loss resulted in embryonic lethality, but Bmp9 KO mice were viable. Finally, we present identification of placental vascular malformations observed in Eng or Alk1 KO placenta. The results of this study are significant in determining potential interactions of HHT in pregnancy and placentation, forming the basis for the investigation of cell-type specific roles and translational relevance of HHT genes in placenta. On-going work aims to characterize the cellular composition and developmental origins of Eng and Alk1 null placenta phenotypes and to evaluate postpartum maternal cardiac physiology in these models. Translational relevance will be assessed through a comparative analysis of human HHT placenta (HHT1 and HHT2) obtained from the MGH HHT Center of Excellence. Vineetha Matthew pictured at right, also received second place in the 2021 Earle P. Charlton Poster Competition for a presentation on "Characterization of Placental Vascular Health in Hereditary Hemorrhagic Telangiectasia.





The Phosphate Transporter Slc20a2 Regulates the Unfolded Protein Response to Endoplasmic Reticulum Stress (UPRER) through Attenuation of eIF2alpha Signaling in Placenta

Ana Correia-BrancoAna Correia-Branco1,2, Eugene W. Hinderer III5, Olga Kashpur1,2, Ciara Benson3, Ariel Mei1,2, Nirmala Jayaraman1,2, Mark C. Blaser4, Hideyuki Higashi4, Shiori Kuraoka 4, Elena Aikawa 4, Sasha A. Singh4, Mary C. Wallingford1,2

1Mother Infant Research Institute, Tufts Medical Center, Boston, MA, USA, 2Molecular Cardiology Research Institute, Tufts Medical Center, Boston, MA, USA, 3Global Alliance to Prevent Prematurity and Stillbirth, Seattle, WA, USA, 4Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA, 5Tufts Analytic Platform (TAP), Clinical and Translational Science Institute-CTSI, Tufts Medical Center, MA, USA.

Placental dysfunction can result in preeclampsia, a pregnancy-specific hypertensive disorder which lacks a curative treatment due to insufficient understanding of the molecular etiology. Maternal-fetal inorganic phosphate (Pi) performs numerous critical functions during embryonic development, but chronic exposure to Pi is detrimental to vascular health. Expression of the sodium-dependent placental Pi transporter Slc20a2 is reduced in early preeclampsia, and the homeostatic endoplasmic stress unfolded protein response (UPRER) is activated. Our previous work supports  that loss of Slc20a2 hastens epigenetic aging in placenta and increases ectopic placental calcification through unknown mechanisms. In this study, we hypothesized that Slc20a2 loss contributes to cellular stress in placenta. We tested this hypothesis through proteomic analysis of Slc20a2 control (WT) and knockout (KO) mouse placenta collected before (E13.5) and after (E17.5) calcification onset (E13.5). After filtering and normalization, a total of 3,050 unique proteins were identified. Interactome analysis of proteins with altered abundance patterns at both time points (n=23) were analyzed with StringDB, and revealed a cluster of endoplasmic reticulum and Golgi-associated proteins. We then tested the hypothesis that Slc20a2 protects the placenta from ectopic calcification by maintaining homeostatic Pi levels in balance with UPRER through molecular approaches. Although robust alterations in Xbp1 mRNA splicing were not observed at E17.5, we found that Slc20a2 loss significantly increases eukaryotic translation initiation factor 2A (eIF2A), which  activates the molecular UPRER molecular chaperone protein disulfide isomerase (PDI). We now propose that Slc20a2 protects the placenta from cellular stress and ectopic calcification through attenuating eIF2A-mediated PDI activation in trophoblasts. Ongoing work investigates the timing and cell type-specificity of UPRER activation in mouse placenta with PDI immunofluorescence, as well as the translational relevance of this novel SLC20A2/eIF2A molecular mechanism in human placenta.