Comparison of Anti-SARS-CoV-2-Specific Antibody Signatures in Maternal and Infant Blood after COVID-19 Infection versus COVID-19 Vaccination during Pregnancy.

OBJECTIVE: The Advisory Committee on Immunization Practices and The American College of Obstetricians and Gynecologists recommend coronavirus disease 2019 (COVID-19) vaccine for pregnant persons to prevent severe illness and death. The objective was to examine levels of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) IgG, IgM, and IgA against spike protein receptor binding domain (RBD) and nucleocapsid protein (NCP) in maternal and infant/cord blood at delivery after COVID 19 vaccination compared with SARS-CoV-2 infection at in mother-infant dyads at specified time points. STUDY DESIGN: Mothers with SARS-CoV-2 infection (n = 31) or COVID-19 vaccination (n = 25) during pregnancy were enrolled between July 2020 and November 2021. Samples were collected at delivery and IgG, IgM, and IgA to RBD of spike and NCPs compared in the infected and vaccinated groups. Timing of infection/vaccination prior to delivery and correlation with antibody levels was performed. RESULTS: The majority of participants received vaccination within 90 days of delivery and over half received the Pfizer BioNTech vaccine. There were no significant correlations between antibody levels and timing of infection or vaccination. Infant IgG levels to the RBD domain of spike protein were higher in the vaccinated group (n = 25) as compared with the infants born to mothers with infection (n = 31). Vaccination against COVID-19 during pregnancy was associated with detectable maternal and infant anti-RBD IgG levels at delivery irrespective of the timing of vaccination. CONCLUSION: Timing of vaccination had no correlation to the antibody levels suggesting that the timing of maternal vaccination in the cohort did not matter. There was no IgM detected in infants from vaccinated mothers. Infants from vaccinated mothers had robust IgG titers to RBD, which have a lasting protective effect in infants. KEY POINTS: · COVID-19 vaccination during pregnancy had detectable antibody.. · No correlation between antibody levels and timing of vaccination.. · Infants from vaccinated mothers had robust IgG titers to RBD..

Maternal, Infant, and Breast Milk Antibody Response Following COVID-19 Infection in Early Versus Late Gestation.

BACKGROUND: Coronavirus disease 2019 [severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)] infection at varying time points during the pregnancy can influence antibody levels after delivery. We aimed to examine SARS-CoV-2 IgG, IgM and IgA receptor binding domain of the spike protein and nucleocapsid protein (N-protein) reactive antibody concentrations in maternal blood, infant blood and breastmilk at birth and 6 weeks after SARS-CoV-2 infection in early versus late gestation. METHODS: Mothers with SARS-CoV-2 infection during pregnancy were enrolled between July 2020 and May 2021. Maternal blood, infant blood and breast milk samples were collected at delivery and 6 weeks postpartum. Samples were analyzed for SARS-CoV-2 spike and N-protein reactive IgG, IgM and IgA antibodies. Antibody concentrations were compared at the 2 time points and based on trimester of infection ("early" 1st/2nd vs. "late" 3rd). RESULTS: Dyads from 20 early and 11 late trimester infections were analyzed. For the entire cohort, there were no significant differences in antibody levels at delivery versus 6 weeks with the exception of breast milk levels which declined over time. Early gestation infections were associated with higher levels of breastmilk IgA to spike protein ( P = 0.04). Infant IgG levels to spike protein were higher at 6 weeks after late infections ( P = 0.04). There were strong correlations between maternal and infant IgG levels at delivery ( P < 0.01), and between breastmilk and infant IgG levels. CONCLUSIONS: SARS-CoV-2 infection in early versus late gestation leads to a persistent antibody response in maternal blood, infant blood and breast milk over the first 6 weeks after delivery.

Cytokine levels in maternal and infant blood after COVID-19 vaccination during pregnancy in comparison with unvaccinated controls.

The objective of this study was to compare maternal and infant cytokine profiles at delivery among those vaccinated against SARS-CoV-2 during pregnancy to unvaccinated controls. Mother-infant dyads were enrolled in this prospective cohort study, and maternal blood and infant and/or cord blood collected. Samples were analyzed utilizing a LEGENDplex 13-plex human anti-viral response cytokine panel. Maternal IP-10 and IFN-λ2/3 were lower in the vaccinated cohort. In the infants, levels were lower for IL-1ß, IFN-λ2/3, and GM-CSF, and higher for IFN-λ1 in the vaccinated cohort. Vaccination against SARS-CoV-2 during pregnancy did not lead to elevations in cytokines in mothers or infants.

Understanding the role of placental pathophysiology in the development of bronchopulmonary dysplasia.

The associations between bronchopulmonary dysplasia (BPD) and the gestational pathologies of chorioamnionitis (CA) and hypertensive disorders of pregnancy (HDP) have become increasingly well recognized. However, the mechanisms through which these antenatal conditions cause increased risk of BPD remain less well characterized. The objective of this review is to discuss the role of the placenta in BPD predisposition as a primary driver of intrauterine alterations adversely impacting fetal lung development. We hypothesize that due to similarities in structure and function, placental disorders during pregnancy can uniquely impact the developing fetal lung, creating a unique placental-pulmonary connection. In the current review, we explore this hypothesis through analysis of clinical literature and preclinical model systems evaluating BPD predisposition, discussion of BPD phenotypes, and an overview on strategies to incorporate placental investigation into research on fetal lung development. We also discuss important concepts learned from research on antenatal steroids as a modulator fetal lung development. Finally, we propose that the appropriate selection of animal models and establishment of in vitro lung developmental model systems incorporating primary human placental components are key in continuing to understand and address antenatal predisposition to BPD.

The impact of maternal SARS-CoV-2 vaccination and first trimester infection on feto-maternal immune responses.

PROBLEM: COVID-19 infection during pregnancy increases maternal and fetal morbidity and mortality. Infection in the second or third trimester leads to changes in the decidual leukocyte populations. However, it is not known whether COVID-19 infection in the first trimester or COVID-19 vaccination during pregnancy alters the decidual immune environment. METHOD OF STUDY: We examined decidual biopsies obtained at delivery from women who had COVID-19 in the first trimester (n = 8), were fully vaccinated against COVID-19 during pregnancy (n = 17), or were neither infected nor vaccinated during pregnancy (n = 9). Decidual macrophages, NK cells, and T cells were quantified by immunofluorescence. Decidual IL-6, IL-10, and IP-10 were quantified by ELISA. RESULTS: There were no differences in decidual macrophages, NK cells, T cells, or cytokines between the first trimester COVID-19 group and the control group. The vaccinated cohort had lower levels of macrophages and NK cells compared to the control group. There were no differences in cytokines between the vaccinated and control groups. CONCLUSIONS: COVID-19 infection in the first trimester did not cause significant decidual leukocyte or cytokine changes at the maternal-fetal interface. Additionally, vaccination was not associated with decidual inflammation, supporting the safety of SARS-CoV-2 vaccination during pregnancy.

Effect of Prenatal Opioid Exposure on the Human Placental Methylome.

Prenatal exposure to addictive drugs can lead to placental epigenetic modifications, but a methylome-wide evaluation of placental DNA methylation changes after prenatal opioid exposure has not yet been performed. Placental tissue samples were collected at delivery from 19 opioid-exposed and 20 unexposed control full-term pregnancies. Placental DNA methylomes were profiled using the Illumina Infinium HumanMethylationEPIC BeadChip. Differentially methylated CpG sites associated with opioid exposure were identified with a linear model using the 'limma' R package. To identify differentially methylated regions (DMRs) spanning multiple CpG sites, the 'DMRcate' R package was used. The functions of genes mapped by differentially methylated CpG sites and DMRs were further annotated using Enrichr. Differentially methylated CpGs (n = 684, unadjusted p < 0.005 and |∆ß| ≥ 0.05) were mapped to 258 genes (including PLD1, MGAM, and ALCS2). Differentially methylated regions (n = 199) were located in 174 genes (including KCNMA1). Enrichment analysis of the top differentially methylated CpG sites and regions indicated disrupted epigenetic regulation of genes involved in synaptic structure, chemical synaptic transmission, and nervous system development. Our findings imply that placental epigenetic changes due to prenatal opioid exposure could result in placental dysfunction, leading to abnormal fetal brain development and the symptoms of opioid withdrawal in neonates.

Evaluation of maternal-infant dyad inflammatory cytokines in pregnancies affected by maternal SARS-CoV-2 infection in early and late gestation.

OBJECTIVE: SARS-CoV-2 infection induces significant inflammatory cytokine production in adults, but infant cytokine signatures in pregnancies affected by maternal SARS-CoV-2 are less well characterized. We aimed to evaluate cytokine profiles of mothers and their infants following COVID-19 in pregnancy. STUDY DESIGN: Serum samples at delivery from 31 mother-infant dyads with maternal SARS-CoV-2 infection in pregnancy (COVID) were examined in comparison to 29 control dyads (Control). Samples were evaluated using a 13-plex cytokine assay. RESULTS: In comparison with controls, interleukin (IL)-6 and interferon gamma-induced protein 10 (IP-10) were higher in COVID maternal and infant samples (p < 0.05) and IL-8 uniquely elevated in COVID infant samples (p < 0.05). Significant elevations in IL-6, IP-10, and IL-8 were found among both early (1st/2nd Trimester) and late (3rd Trimester) maternal SARS-CoV-2 infections. CONCLUSIONS: Maternal SARS-CoV-2 infections throughout gestation are associated with increased maternal and infant inflammatory cytokines at birth with potential to impact long-term infant health.

Decidual immune response following COVID-19 during pregnancy varies by timing of maternal SARS-CoV-2 infection.

While COVID-19 infection during pregnancy is common, fetal transmission is rare, suggesting that intrauterine mechanisms form an effective blockade against SARS-CoV-2. Key among these is the decidual immune environment of the placenta. We hypothesize that decidual leukocytes are altered by maternal SARS-CoV-2 infection in pregnancy and that this decidual immune response is shaped by the timing of infection during gestation. To address this hypothesis, we collected decidua basalis tissues at delivery from women with symptomatic COVID-19 during second (2nd Tri COVID, n = 8) or third trimester (3rd Tri COVID, n = 8) and SARS-CoV-2-negative controls (Control, n = 8). Decidual natural killer (NK) cells, macrophages and T cells were evaluated using quantitative microscopy, and pro- and anti-inflammatory cytokine mRNA expression was evaluated using quantitative reverse transcriptase PCR (qRT-PCR). When compared with the Control group, decidual tissues from 3rd Tri COVID exhibited significantly increased macrophages, NK cells and T cells, whereas 2nd Tri COVID only had significantly increased T cells. In evaluating decidual cytokine expression, we noted that IL-6, IL-8, IL-10 and TNF-α were significantly correlated with macrophage cell abundance. However, in 2nd Tri COVID tissues, there was significant downregulation of IL-6, IL-8, IL-10, and TNF-α. Taken together, these results suggest innate and adaptive immune responses are present at the maternal-fetal interface in maternal SARS-CoV-2 infections late in pregnancy, and that infections earlier in pregnancy show evidence of a resolving immune response. Further studies are warranted to characterize the full scope of intrauterine immune responses in pregnancies affected by maternal COVID-19.

Acute Severe Acute Respiratory Syndrome Coronavirus 2 Infection in Pregnancy Is Associated with Placental Angiotensin-Converting Enzyme 2 Shedding.

While the human placenta may be infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the rate of fetal transmission is low, suggesting a barrier at the maternal-fetal interface. Angiotensin-converting enzyme (ACE)2, the main receptor for SARS-CoV-2, is regulated by a metalloprotease cleavage enzyme, a disintegrin and metalloprotease domain 17 (ADAM17). ACE2 is expressed in the human placenta, but its regulation in relation to maternal SARS-CoV-2 infection in pregnancy is not well understood. This study evaluated ACE2 expression, ADAM17 activity, and serum ACE2 abundance in a cohort of matched villous placental and maternal serum samples from control pregnancies (SARS-CoV-2 negative, n = 8) and pregnancies affected by symptomatic maternal SARS-CoV-2 infections in the second trimester [2nd Tri coronavirus disease (COVID), n = 8] and third trimester (3rd Tri COVID, n = 8). In 3rd Tri COVID compared with control and 2nd Tri COVID villous placental tissues, ACE2 mRNA expression was remarkably elevated; however, ACE2 protein expression was significantly decreased with a parallel increase in ADAM17 activity. Soluble ACE2 was also significantly increased in the maternal serum from 3rd Tri COVID infections compared with control and 2nd Tri COVID pregnancies. These data suggest that in acute maternal SARS-CoV-2 infections, decreased placental ACE2 protein may be the result of ACE2 shedding and highlights the importance of ACE2 for studies on SARS-CoV-2 responses at the maternal-fetal interface.

Antenatal Mesenchymal Stromal Cell Extracellular Vesicle Therapy Prevents Preeclamptic Lung Injury in Mice.

In preeclamptic pregnancies, a variety of intrauterine alterations lead to abnormal placentation, release of inflammatory and/or antiangiogenic factors, and subsequent fetal growth restriction with significant potential to cause a primary insult to the developing fetal lung. Thus, modulation of the maternal intrauterine environment may be a key therapeutic avenue to prevent preeclampsia-associated developmental lung injury. A biologic therapy of interest is mesenchymal stromal cell-derived extracellular vesicles (MEx), which we have previously shown to ameliorate preeclamptic physiology through intrauterine immunomodulation. To evaluate the therapeutic potential of MEx to improve developmental lung injury in experimental preeclampsia, using the heme oxygenase-1-null mouse (Hmox1-/-) model, preeclamptic pregnant dams were administered intravenous antenatal MEx treatment during each week of pregnancy followed by analysis of fetal and postnatal lung tissues, amniotic fluid protein profiles, and lung explant and amniotic fluid cocultures in comparison with control and untreated preeclamptic pregnancies. We first identified that a preeclamptic intrauterine environment had a significant adverse impact on fetal lung development, including alterations in fetal lung developmental gene profiles in addition to postnatal alveolar and bronchial changes. Amniotic fluid proteomic analysis and fetal lung explant and amniotic fluid cocultures further demonstrated that maternally administered MEx altered the expression of multiple inflammatory mediators in the preeclamptic intrauterine compartment, resulting in the normalization of fetal lung branching morphogenesis and developmental gene expression. Our evaluation of fetal and postnatal parameters overall suggests that antenatal MEx treatment may provide a highly valuable preventative therapeutic modality for amelioration of lung development in preeclamptic disease.

SARS-CoV-2 in infant urine and fecal samples after in utero COVID-19 exposure.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is a pandemic that has and will continue to affect many pregnant women. Knowledge regarding the risk of vertical transmission is limited. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) real-time reverse transcriptase-polymerase chain reaction (RT-PCR) of nasopharyngeal swabs typically have been used to confirm the diagnosis among infants, but whether the virus can be detected in other biological specimens, and therefore potentially transmitted in other ways, is unknown. Positive SARS-CoV-2 RT-PCR has been reported from feces and urine from adult patients. We hypothesize that the presence of SARS-CoV-2 in infant urine and fecal samples after prenatal COVID-19 exposure is low. METHODS: We examined the presence of SARS-CoV-2 RNA using RT-PCR in urine and fecal samples among 42 infants born to SARS-CoV-2-infected mothers during different stages of pregnancy. RESULTS: A urine sample was collected from 39 of 42 infants and fecal samples from all 42 infants shortly after birth. Although the majority of the women had the symptomatic disease (85.6%), we were unable to detect the presence of SARS-CoV-2 virus from any infant urine or fecal samples. CONCLUSIONS: SARS-CoV-2 was not detected in infant urine or feces after maternal infection during pregnancy, providing further evidence for low rates of perinatal transmission. IMPACT: SARS-CoV-2 was not detected in the urine or feces of infants of mothers with COVID-19 during various time points in pregnancy. This study provides further evidence for low rates of perinatal transmission of SARS-CoV-2. Results help to provide guidance on perinatal care practices for infants exposed to COVID-19 in utero.

Antenatal mesenchymal stromal cell extracellular vesicle treatment preserves lung development in a model of bronchopulmonary dysplasia due to chorioamnionitis.

Antenatal stressors such as chorioamnionitis (CA) increase the risk for bronchopulmonary dysplasia (BPD). Studies have shown that experimental BPD can be ameliorated by postnatal treatment with mesenchymal stromal cell-derived extracellular vesicles (MEx). However, the antenatal efficacy of MEx to prevent BPD is unknown. To determine whether antenatal MEx therapy attenuates intrauterine inflammation and preserves lung growth in a rat model of CA-induced BPD. At embryonic day (E)20, rat litters were treated with intra-amniotic injections of saline, endotoxin (ETX) to model chorioamnionitis, MEx, or ETX plus MEx followed by cesarean section delivery with placental harvest at E22. Placental and lung evaluations were conducted at day 0 and day 14, respectively. To assess the effects of ETX and MEx on lung growth in vitro, E15 lung explants were imaged for distal branching. Placental tissues from ETX-exposed pregnancies showed increased expression of inflammatory markers NLRP-3 and IL-1ß and altered spiral artery morphology. In addition, infant rats exposed to intrauterine ETX had reduced alveolarization and pulmonary vessel density (PVD), increased right ventricular hypertrophy (RVH), and decreased lung mechanics. Intrauterine MEx therapy of ETX-exposed pups reduced inflammatory cytokines, normalized spiral artery architecture, and preserved distal lung growth and mechanics. In vitro studies showed that MEx treatment enhanced distal lung branching and increased VEGF and SPC gene expression. Antenatal MEx treatment preserved distal lung growth and reduced intrauterine inflammation in a model of CA-induced BPD. We speculate that MEx may provide a novel therapeutic strategy to prevent BPD due to antenatal inflammation.

Acute SARS-CoV-2 infection in pregnancy is associated with placental ACE-2 shedding.

Human placental tissues have variable rates of SARS-CoV-2 invasion resulting in consistently low rates of fetal transmission suggesting a unique physiologic blockade against SARS-CoV-2. Angiotensin-converting enzyme (ACE)-2, the main receptor for SARS-CoV-2, is expressed as cell surface and soluble forms regulated by a metalloprotease cleavage enzyme, ADAM17. ACE-2 is expressed in the human placenta, but the regulation of placental ACE-2 expression in relation to timing of maternal SARS-CoV-2 infection in pregnancy is not well understood. In this study, we evaluated ACE-2 expression, ADAM17 activity and serum ACE-2 abundance in a cohort of matched villous placental and maternal serum samples from Control pregnancies (SARS-CoV-2 negative, n=8) and pregnancies affected by symptomatic maternal SARS-CoV-2 infections in the 2 nd trimester ("2 nd Tri COVID", n=8) and 3rd trimester ("3 rd Tri COVID", n=8). In 3 rd Tri COVID as compared to control and 2 nd Tri-COVID villous placental tissues ACE-2 mRNA expression was remarkably elevated, however, ACE-2 protein expression was significantly decreased with a parallel increase in ADAM17 activity. Soluble ACE-2 was also significantly increased in the maternal serum from 3 rd Tri COVID infections as compared to control and 2 nd Tri-COVID pregnancies. These data suggest that in acute maternal SARS-CoV-2 infections, decreased placental ACE-2 protein may be the result of ACE-2 shedding. Overall, this work highlights the importance of ACE-2 for ongoing studies on SARS-CoV-2 responses at the maternal-fetal interface.

Extracellular Vesicles Protect the Neonatal Lung from Hyperoxic Injury through the Epigenetic and Transcriptomic Reprogramming of Myeloid Cells.

Rationale: Mesenchymal stem/stromal cell (MSC)-small extracellular vesicle (MEx) treatment has shown promise in experimental models of neonatal lung injury. The molecular mechanisms by which MEx afford beneficial effects remain incompletely understood. Objectives: To investigate the therapeutic mechanism of action through assessment of MEx biodistribution and impact on immune cell phenotypic heterogeneity. Methods: MEx were isolated from the conditioned medium of human umbilical cord Wharton's jelly-derived MSCs. Newborn mice were exposed to hyperoxia (HYRX, 75% O2) from birth and returned to room air at Postnatal Day 14 (PN14). Mice received either a bolus intravenous MEx dose at PN4 or bone marrow-derived myeloid cells (BMDMy) pretreated with MEx. Animals were killed at PN4, PN7, PN14, or PN28 to characterize MEx biodistribution or for assessment of pulmonary parameters. The therapeutic role of MEx-educated BMDMy was determined in vitro and in vivo. Measurements and Main Results: MEx therapy ameliorated core histological features of HYRX-induced neonatal lung injury. Biodistribution and mass cytometry studies demonstrated that MEx localize in the lung and interact with myeloid cells. MEx restored the apportion of alveolar macrophages in the HYRX-injured lung and concomitantly suppressed inflammatory cytokine production. In vitro and ex vivo studies revealed that MEx promoted an immunosuppressive BMDMy phenotype. Functional assays demonstrated that the immunosuppressive actions of BMDMy are driven by phenotypically and epigenetically reprogrammed monocytes. Adoptive transfer of MEx-educated BMDMy, but not naive BMDMy, restored alveolar architecture, blunted fibrosis and pulmonary vascular remodeling, and improved exercise capacity. Conclusions: MEx ameliorate hyperoxia-induced neonatal lung injury though epigenetic and phenotypic reprogramming of myeloid cells.

Intratracheal transplantation of trophoblast stem cells attenuates acute lung injury in mice.

BACKGROUND: Acute lung injury (ALI) is a common lung disorder that affects millions of people every year. The infiltration of inflammatory cells into the lungs and death of the alveolar epithelial cells are key factors to trigger a pathological cascade. Trophoblast stem cells (TSCs) are immune privileged, and demonstrate the capability of self-renewal and multipotency with differentiation into three germ layers. We hypothesized that intratracheal transplantation of TSCs may alleviate ALI. METHODS: ALI was induced by intratracheal delivery of bleomycin (BLM) in mice. After exposure to BLM, pre-labeled TSCs or fibroblasts (FBs) were intratracheally administered into the lungs. Analyses of the lungs were performed for inflammatory infiltrates, cell apoptosis, and engraftment of TSCs. Pro-inflammatory cytokines/chemokines of lung tissue and in bronchoalveolar lavage fluid (BALF) were also assessed. RESULTS: The lungs displayed a reduction in cellularity, with decreased CD45+ cells, and less thickening of the alveolar walls in ALI mice that received TSCs compared with ALI mice receiving PBS or FBs. TSCs decreased infiltration of neutrophils and macrophages, and the expression of interleukin (IL) 6, monocyte chemoattractant protein-1 (MCP-1) and keratinocyte-derived chemokine (KC) in the injured lungs. The levels of inflammatory cytokines in BALF, particularly IL-6, were decreased in ALI mice receiving TSCs, compared to ALI mice that received PBS or FBs. TSCs also significantly reduced BLM-induced apoptosis of alveolar epithelial cells in vitro and in vivo. Transplanted TSCs integrated into the alveolar walls and expressed aquaporin 5 and prosurfactant protein C, markers for alveolar epithelial type I and II cells, respectively. CONCLUSION: Intratracheal transplantation of TSCs into the lungs of mice after acute exposure to BLM reduced pulmonary inflammation and cell death. Furthermore, TSCs engrafted into the alveolar walls to form alveolar epithelial type I and II cells. These data support the use of TSCs for the treatment of ALI.

Mesenchymal Stromal Cell-Derived Extracellular Vesicles Restore Thymic Architecture and T Cell Function Disrupted by Neonatal Hyperoxia.

Treating premature infants with high oxygen is a routine intervention in the context of neonatal intensive care. Unfortunately, the increase in survival rates is associated with various detrimental sequalae of hyperoxia exposure, most notably bronchopulmonary dysplasia (BPD), a disease of disrupted lung development. The effects of high oxygen exposure on other developing organs of the infant, as well as the possible impact such disrupted development may have on later life remain poorly understood. Using a neonatal mouse model to investigate the effects of hyperoxia on the immature immune system we observed a dramatic involution of the thymic medulla, and this lesion was associated with disrupted FoxP3+ regulatory T cell generation and T cell autoreactivity. Significantly, administration of mesenchymal stromal cell-derived extracellular vesicles (MEx) restored thymic medullary architecture and physiological thymocyte profiles. Using single cell transcriptomics, we further demonstrated preferential impact of MEx treatment on the thymic medullary antigen presentation axis, as evidenced by enrichment of antigen presentation and antioxidative-stress related genes in dendritic cells (DCs) and medullary epithelial cells (mTECs). Our study demonstrates that MEx treatment represents a promising restorative therapeutic approach for oxygen-induced thymic injury, thus promoting normal development of both central tolerance and adaptive immunity.

Differential Expression of Rab5 and Rab7 Small GTPase Proteins in Placental Tissues From Pregnancies Affected by Maternal Coronavirus Disease 2019.

PURPOSE: The majority of pregnancies affected by maternal coronavirus disease 2019 (COVID-19) do not result in fetal transmission. However, several studies have identified parenchymal changes in their placental tissues, suggesting a placental response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) at the maternal-fetal interface. Although many COVID-19 placental studies have focused on the expression of the canonical SARS-CoV-2 entry proteins angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2, further characterization of subcellular molecules involved in viral trafficking have not yet been investigated in these tissues. Of interest are Rab proteins, a family of small GTPase proteins that direct intracellular transport between different endocytic organelles. Rab5 and Rab7 in particular have previously been implicated in HIV and cytomegalovirus invasion of placental trophoblast cells in vitro; the localization of these molecules has not been fully characterized within the human maternal-fetal interface, however, or within placental tissues from SARS-CoV-2-infected pregnancies. METHODS: Using fluorescent immunohistochemistry, Rab5 and Rab7 placental localization and comparative fluorescence intensity were explored in a cohort of placental tissues from pregnancies affected by maternal COVID-19 disease (COVID, n = 15) compared with contemporary control subjects (Control, n = 10). Fluorescence intensity was quantified by using corrected total cell fluorescence values. FINDINGS: Within placental villi, Rab5 was consistently localized in syncytiotrophoblast and cytotrophoblast cells. Rab5 had significantly higher mean (SEM) fluorescence intensity in the COVID cohort (Control, 1.96 [0.16]; COVID, 2.62 [0.09]; P = 0.0014). In contrast, although Rab7 was also localized within placental villous syncytiotrophoblast and cytotrophoblast cells, mean (SEM) Rab7 fluorescence intensity was significantly downregulated in COVID vs Control placentas (Control, 35.9 [4.1]; COVID, 20.1 [0.52]; P = 0.0001). IMPLICATIONS: This differential expression of Rab5 and Rab7 suggests that placental endocytic pathways may be altered at the maternal-fetal interface in pregnancies affected by maternal SARS-CoV-2 infection. As key molecules governing intracellular vesicle transport, including viral trafficking, Rab GTPase proteins may be of interest for ongoing studies examining placental responses to COVID-19 in pregnancy.

Mesenchymal stromal cell-derived extracellular vesicle therapy prevents preeclamptic physiology through intrauterine immunomodulation†.

Human umbilical cord-derived mesenchymal stromal cells (MSCs) are a widely recognized treatment modality for a variety of preclinical disease models and have been transitioned to human clinical trials. We have previously shown in neonatal lung disease that the therapeutic capacity of MSCs is conferred by their secreted extracellular vesicles (MEx), which function primarily through immunomodulation. We hypothesize that MEx have significant therapeutic potential pertinent to immune-mediated gestational diseases. Of particular interest is early-onset preeclampsia, which can be caused by alterations of the maternal intrauterine immune environment. Using a heme-oxygenase-1 null mouse model of pregnancy loss with preeclampsia-like features, we examined the preventative effects of maternal MEx treatment early in pregnancy. Heme oxygenase-1 null females (Hmox1-/-) or wild-type control females were bred in homozygous matings followed by evaluation of maternal and fetal parameters. A single dose of MEx was administered intravenously on gestational day (GD)1 to Hmox1-/- females (Hmox1-/- MEx). Compared with untreated Hmox1-/- females, Hmox1-/- MEx-treated pregnancies showed significant improvement in fetal loss, intrauterine growth restriction, placental spiral artery modification, and maternal preeclamptic stigmata. Biodistribution studies demonstrated that MEx localize to a subset of cells in the preimplantation uterus. Further, mass cytometric (CyTOF) evaluation of utero-placental leukocytes in Hmox1-/- MEx versus untreated pregnancies showed alteration in the abundance, surface marker repertoire, and cytokine profiles of multiple immune populations. Our data demonstrate the therapeutic potential of MEx to optimize the intrauterine immune environment and prevent maternal and fetal sequelae of preeclamptic disease.

Consistent localization of SARS-CoV-2 spike glycoprotein and ACE2 over TMPRSS2 predominance in placental villi of 15 COVID-19 positive maternal-fetal dyads.

INTRODUCTION: While the COVID-19 pandemic continues to have a significant global health impact, rates of maternal to infant vertical transmission remain low (<5%). Parenchymal changes of placentas from COVID-19 infected mothers have been reported by several groups, but the localization and relative abundance of SARS-CoV-2 viral proteins and cellular entry machinery has not been fully characterized within larger placental tissue cohorts. METHODS: An extended placental tissue cohort including samples from 15 COVID-19 positive maternal-fetal dyads (with n = 5 cases with evidence of fetal transmission) in comparison with 10 contemporary COVID-19 negative controls. Using comparative immunofluorescence, we examined the localization and relative tissue abundance of SARS-CoV2 spike glycoprotein (CoV2 SP) along with the co-localization of two SARS-CoV2 viral entry proteins angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2). RESULTS/CONCLUSIONS: CoV2 SP was present within the villous placenta in COVID-19 positive pregnancies with and without evidence of fetal transmission. We further identified the predominance of ACE2 expression in comparison with TMPRSS2. Importantly, both CoV2 SP and ACE2 expression consistently localized primarily within the outer syncytiotrophoblast layer placental villi, a key physiologic interface between mother and fetus. Overall this study provides an important basis for the ongoing evaluation of SARS-CoV-2 physiology in pregnancy and highlights the importance of the placenta as a key source of primary human tissue for ongoing diagnostic and therapeutic research efforts to reduce the global burden of COVID-19.