Brien, Marie‐Eve

Background: Preeclampsia (PE) is a hypertensive disorder of pregnancy defined by new-onset hypertension occurring after 20 weeks of pregnancy, with end organ damage. Pro-inflammatory changes are also observed in these patients. Although the clinical features of PE are resolved soon after delivery, affected patients face a higher risk and earlier onset of cardiovascular disease (CVD). It is unknown what leads to this persistent elevated risk, and long-term alterations of the maternal immune system could be involved. Study objective: Our goal was to address the immune alterations and functional changes in the first year postpartum in PE as compared to normotensive (NT) pregnancies. Hypothesis: We hypothesize that the maternal immune profile is altered in the postpartum period in PE vs NT patients, with a proinflammatory bias and altered function in PE patients. Brief Statement of Methods: We prospectively followed 31 women (PE=15, NT=16) and collected blood 24 hours, and 3, 6, and 12 months postpartum. Plasma and peripheral blood mononuclear cells (PBMC) were isolated and stored for analysis. Inflammatory mediators were quantified through multiplex assays, and PBMCs profile were defined through flow cytometry as well as following inflammatory stimuli to assess their functional changes through intracellular cytokine staining and expression of activation markers. Summary of Results: PE patients had elevated CD3+ T cell frequencies at 3 and 12 months postpartum compared to NT patients. Both groups showed reduced CD14+ monocytes across postpartum timepoints compared with 24 hours. PE was also associated with increased amounts of pro-inflammatory cytokines (such as IL1α, IL9, IFNγ), chemokines and growth factors at multiple timepoints throughout the first year postpartum; NT patients had decreased amounts of pro-inflammatory cytokines. Intracellular staining showed altered immune cell functions in PE, with decreased IL-6 production following polyI:C stimulation and decreased IL-10+ monocytes after LPS and polyI:C at 3 months postpartum. At 3 months, PE patients had increased markers of activation (i.e. CD69+) in both NK cells and T cells, the latter following PMA/Ionomycin stimulation. Statement of Conclusions: Pregnancy itself induces lasting immune changes persisting to 12 months postpartum, even in NT patients, whilst PE complicated pregnancies have sustained inflammation and altered immune function in the postpartum period. Overall, this work will provide essential insight into an understudied period that many women experience and help to identify long lasting immunological changes that could impact long term maternal health. Future work will explore the impact of this sustained inflammation on epigenetic modifications in immune cells. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Abstract SARS-CoV-2 infection disrupts iron homeostasis in organs such as the lung. Emerging evidence suggests similar dysregulation in the placenta may impair function and fetal growth. Ferroptosis, an iron-dependent form of cell death, is driven by the accumulation of labile ferrous iron, catalyzing the peroxidation of polyunsaturated fatty acids in the absence of sufficient iron storage or antioxidant defenses. This pathway has been implicated in SARS-CoV-2-related tissue damage in other organs and may contribute to placental dysfunction during pregnancy. However, the influence of infection timing, symptom severity, and fetal sex on placental iron regulation remains unexamined. We analyzed placental samples from a cohort of SARS-CoV-2-exposed pregnancies in the first (n = 6), second (n = 14), or third trimester (n = 30), classified by symptom severity (asymptomatic n = 23, symptomatic n = 27) and fetal sex, alongside unexposed controls (n = 47). We assessed placental iron deposition, lipid peroxidation, and mRNA and protein expression of ferroptosis markers. Publicly available RNA-sequencing datasets from human placenta tissue, exposed or not to SARS-CoV-2, were analyzed to assess ferroptosis-related gene expression. SARS-CoV-2 infection, particularly in symptomatic cases, was associated with reduced placental weight and birth weight. Asymptomatic cases showed increased placental iron deposition, which correlated with lower birthweight and was accompanied by elevated expression of nuclear receptor coactivator 4, a cytosolic adaptor protein that mediates the selective autophagic degradation of ferritin (ferritinophagy) and iron release. Placentas from symptomatic patients exhibited evidence of altered iron transport and sex-specific down-regulation of antioxidant defenses. Transcriptomic analyses further suggested widespread disruption of ferroptosis pathways in placentas from infected patients. Our findings reveal that SARS-CoV-2 infection alters placental iron homeostasis and is associated with ferroptosis-related changes, with distinct molecular responses based on timing of infection, symptom severity, and fetal sex.

Abstract Since the beginning of the COVID-19 pandemic, vulnerable populations such as pregnant persons have been at higher risk of severe symptoms and poor outcomes. Although reports of SARS-CoV-2 vertical transmission remain rare, several studies showed that maternal infection during pregnancy can induce histomorphological and inflammatory alterations in the placenta. However, the permissiveness of human trophoblasts to various variants of the virus remains poorly characterized. In this study, human primary villous cytotrophoblasts isolated from term placentas, along with trophoblastic cell lines BeWo, JEG-3, and HIPEC-65 were infected with the ancestral SARS-CoV-2 strain, which disseminated worldwide in early 2020. Permissiveness was assessed with quantitative RT-PCR, immunostaining of viral protein Nucleocapsid, and plaque assays. To investigate viral entry routes, cells were treated with Camostat mesylate (an inhibitor of the co-entry factor TMPRSS2) or chloroquine phosphate (an endosomal entry inhibitor) and viral fitness was assessed by plaque assays. Primary villous cytotrophoblasts and JEG-3 cells were also tested for infection with three pre-omicron SARS-CoV-2 variants of concern. Our results show that primary villous cytotrophoblasts are permissive to all tested SARS-CoV-2 strains in vitro. Infection with the ancestral SARS-CoV-2 strain relies mainly on a non-canonical endosomal entry pathway. Notably, JEG-3 cells represent an appropriate and convenient model for studying trophoblast infection by SARS-CoV-2, as they exhibit high permissiveness to the ancestral strain, and the SARS-CoV-2 entry pathway is similar to that in villous cytotrophoblasts. Overall, this study reveals that the cytotrophoblastic permissiveness to SARS-CoV-2 depends on the viral genetic nature and provides new insights into its entry route in human trophoblasts.

Abstract Description Hypertensive disorders of pregnancy (HDP) include preeclampsia (PE) and gestational hypertension (gHTN), both suspected to be immune-driven pathologies. Our data shows that both PE and gHTN involve systemic immune cell activation and increased proinflammatory cytokines however it is unclear how these immune profiles play a role in the pathogenesis of disease. We hypothesize that pregnancies complicated with HDP have dysregulated immune profiles, including hyperactive T cells which are activating the vascular endothelium. We have investigated the circulating immune profiles of patients with normotensive (NT), PE or gHTN pregnancies at the RNA, protein and cellular levels, which show significant alterations in T cells, among others. We have seen increased levels of IL-6, CXCL9 and CXCL10 in the plasma of HDP patients vs NT, suggesting the polarization and recruitment of T cells. Using cytometry by time of flight (CyTOF), we have seen changes in expression of CD11a and CXCR3 on CD4+, CD8+ and yd T cells in HDP compared to NT. Single cell RNA sequencing of CD4+ and CD8+ T cells from PE vs NT shows increased expression of genes related to pathways such as allograft rejection, angiogenesis and cytokine signaling. Our in vitro co-culture experiments examine the ability of immune cells, specifically T cells, to activate endothelial cells. These studies will address the functional contribution of T cells to endothelial activation, which is a hallmark of the hypertension seen in HDP. Topic Categories Cellular Adhesion, Migration, and Inflammation (CAM)

Abstract Description Preeclampsia (PE) is a complication of pregnancy characterized by de novo hypertension after 20 weeks of gestation with end-organ damage. Even though symptoms of PE rapidly resolve after delivery, patients have higher incidence of cardiovascular diseases in the years postpartum. Immunological changes have been observed in PE but it is unknown if these are sustained in the postpartum period. Our goal was to address the immune changes in the first year postpartum in PE as compared to normotensive (NT) as well as nulliparous women. We saw that both PE and NT pregnancies had altered immune system, especially decreased CD14+ monocytes and increased circulating pro-inflammatory cytokines when compared to nulliparous patients even one year after delivery. Furthermore, our intracellular cytokine staining analysis following stimulation suggest altered response in the postpartum period in PE with a predominant anti-inflammatory response vs NT. Overall this work suggest that the immune system remains different from nulliparous women at 1 year postpartum, even in cases of normotensive pregnancies, and is further altered following PE-complicated pregnancies. Although further analysis is ongoing, this shows that the immune system does not go back to pre-pregnancy levels in the postpartum period and that changes induced by pregnancy are at least in part sustained. We will further expand this work to address the functional changes in immune cells and the mechanisms underlying these changes. Topic Categories Mucosal and Regional Immunology (MUC)

Abstract Background and objective The placenta is crucial for fetal development; altered function is associated with complications and adverse neonatal outcomes. Our goal was to ascertain changes in the placental transcriptome in relation to neonatal outcome and fetal/placental sex. Study design Seventy-two mother-baby dyads were included. Demographic, obstetrical, neonatal, and infant health data were obtained through medical charts. Adverse neonatal outcome was defined as the presence of a pulmonary, cardiac, neurological or other health complication. Bulk RNA-sequencing of placental biopsies was obtained. P-value <0.05 was considered statistically significant. Results Neonates experiencing adverse outcomes were more likely to be premature or have lower birth weights. Analysis of the placental transcriptome revealed a predominant inflammatory profile in pregnancies associated with adverse neonatal outcomes with the top pathways being related to immune and inflammatory responses. Among differently expressed genes (DEGs), 1237 were upregulated and 239 were downregulated in adverse vs no adverse outcomes. Furthermore, sex-specific differences in gene expression were observed and indicated that male and female placentas displayed unique DEGs in association with adverse outcomes. Indeed, no DEG was observed in female placentas when comparing those without vs with adverse neonatal outcomes, as opposed to 1279 DEGs in male placentas, of which 91% were up-regulated in adverse subgroups. Conclusions These findings highlight that inflammatory pathways are upregulated in placentas in association with adverse neonatal outcomes, and showcase the importance of the fetal sex in understanding neonatal health. The placenta provides a unique tool for early identification of high-risk infants rapidly after delivery.

Abstract BACKGROUND Postpartum preeclampsia (PPPE) is a maternal condition characterized by de novo hypertension in the postpartum period with end-organ damage. We investigated the use of perinatal immune changes, through routine complete blood count (CBC), to identify high-risk individuals before PPPE development. METHODS We performed a retrospective matched case-control study of 100 individuals with PPPE, 200 term pregnancies (Ctrl) and 200 antenatal preeclampsia (PE). Detailed demographic, obstetrical, and laboratory data were retrieved from medical records. Statistical analysis was performed using one-way ANOVA, multivariate regression, and paired or unpaired t-tests, as appropriate. RESULTS Individuals who developed PPPE were significantly older and predominantly Black vs Ctrl and PE. Both PE and PPPE had higher pre-pregnancy BMI and increased personal and family history of hypertension/PE vs Ctrl (P < 0.001). Before delivery, individuals who later developed PPPE, had lower total leukocyte counts vs Ctrl (9.61 vs 10.75 × 109/L, P < 0.05) whereas monocytes percentage was elevated (8.07 vs 7.27%, P < 0.01). Comparing the postpartum/antenatal ratio in-between each condition revealed elevated leukocyte ratio in PPPE vs Ctrl (P < 0.001). The Neutrophils ratio was also increased, whereas lymphocytes and monocytes were decreased in PPPE vs both Ctrl and PE. After adjusting for race, maternal age, pre-pregnancy BMI, personal history of PE/HT, and diabetes mellitus, perinatal immune changes were still significantly associated with PPPE. CONCLUSIONS Globally, perinatal immune changes were observed in individuals with a seemingly uncomplicated pregnancy prior to the development of PPPE. This strongly supports that such changes could be used to identify high-risk individuals prior to disease onset.

Abstract Pre-eclampsia (PE) affects 5–8% of pregnancies and has detrimental effects on maternal–fetal health. PE is characterized by de novo hypertension after 20 wk of gestation and end-organ damage. Systemic inflammatory imbalance has been associated with PE, but its contribution to the pathology is poorly understood. Our objective was to investigate maternal systemic immune changes in early-onset PE (EOPE) and late-onset PE (LOPE) versus uncomplicated pregnancies (control [CTRL]), and their contribution to endothelial activation, hallmark of hypertension. Blood samples were analyzed by flow cytometry, multiplex assay, intracellular cytokine staining, and single-cell RNA sequencing. We performed cocultures between circulating immune cells and HUVECs to assess endothelial activation. We found that EOPE had decreased regulatory T cells (4.64±0.33, p < 0.05) and monocytes (33.92±3.08, p < 0.01), whereas LOPE had decreased regulatory T cells (4.60±0.30, p < 0.05) and Th2 cells (7.50±0.62, p < 0.01) versus CTRL. Compared to CTRL, elevated cytokines/chemokines, and growth factors were observed in LOPE, whereas EOPE primarily showed decreased levels. Using intracellular cytokine staining, we observed more monocytes producing IL-12, TNF-α, and IL-1β (all p < 0.05) in LOPE versus CTRL. At the transcriptomic level, we found differentially expressed genes between EOPE and CTRL, predominantly related to upregulation of immune activation pathways. Lastly, EOPE PBMCs induced heightened endothelial activation in vitro observed by increased ICAM-1 and ET-1 (p < 0.05), whereas LOPE PBMCs required LPS stimulation. Although significant proteomic changes are observed in the LOPE group, the EOPE displayed changes mostly at the transcriptomic levels and could induce endothelial activation in vitro.

AbstractProblemThe COVID‐19 pandemic has been shown to have a detrimental impact on the mental health of pregnant individuals, and chronic stress can alter the immune profile. However, the effects of the COVID‐19 pandemic on the immune system in pregnancy are still poorly understood. We aimed to evaluate the impact of pandemic exposure on the maternal immune profile in uncomplicated pregnancies.Method of studyWe collected blood and placenta samples from pregnant individuals exposed and unexposed to the pandemic to compare their immune and inflammatory profiles. We performed co‐culture with circulating maternal immune cells and endothelial cells to assess endothelial activation. Statistical analysis was performed using unpaired t‐test, Mann‐Whitney, or Fisher's exact test as appropriate.ResultsIn exposed individuals, we observed a decreased proportion of Th2 cells (p < .0001) and Treg/Th17 ratio (p < .05), as well as an increased Th1/Th2 ratio (p < .0001). Levels of IL‐1β (p < .01) and IL‐18 (p < .01) were increased in the circulation of exposed participants, whilst other mediators were significantly decreased (IFNγ, IL‐8, MCP‐1, amongst others). Furthermore, we observed increased production of ICAM, hallmark of endothelial activation, when we co‐cultured endothelial cells with immune cells from exposed individuals. Vaccination status impacted the cellular profile with increased proportions of Th1 and B cells in vaccinated participants.ConclusionOverall, we observed a pro‐inflammatory bias in the circulation of pregnant individuals exposed to the COVID‐19 pandemic, with otherwise uncomplicated pregnancies. Our work also supports an association between the increased risk of endothelial activation/hypertension and SARS‐CoV2 infection, which might be driven in part by exposure to the pandemic and associated stressors.

AbstractProblemIt is now recognized that SARS‐CoV‐2 infection and pandemic‐related stress impacts maternal health. However, their effects at the maternal‐fetal interface are still debated.Method of studyWe recruited 199 women between March 2020 and July 2021, 79 SARS‐CoV‐2+ and 120 negative (the latter exposed to pandemic stress only). We also included 40 historic controls (i.e. pre‐pandemic uncomplicated pregnancies recruited before March 2020). Placental samples were collected for protein and histological analysis.ResultsThe majority of SARS‐CoV‐2+ women were multiethnic, had higher pre‐pregnancy BMI and elevated preterm birth rate (17%) vs SARS‐CoV‐2– or historic control. Placental inflammatory profile revealed increased IL‐1Ra and CRP, independently of SARS‐CoV‐2 status, whilst MCP‐1, IL‐6 and IFNγ were elevated in the negative, but pandemic stress‐exposed, group. These changes were predominant in placentas with inflammatory lesions on histopathological analysis. Furthermore, we observed elevated immune cells (CD45+) in placentas from SARS‐CoV‐2+ and negative pregnancies vs historic controls, even when individuals with pregnancy complications were excluded.ConclusionsPlacental inflammatory profiles differed between SARS‐CoV‐2 statuses, namely exposed to pandemic stress +/‐ SARS‐CoV‐2 infection. This highlights the need to understand the differences between the effects of pandemic‐related stress and the added burden of SARS‐CoV‐2 infection on placental health.