Shannon A. Bainbridge

Inflammation-mediated preeclampsia (I-PE) is associated with increased placental NAD ⁺ -dependent ADP-ribosylation and NAD ⁺ depletion. We investigated whether PARP1/2- or tankyrase-sensitive ADP-ribosylation contributes to inflammation-associated trophoblast dysfunction and whether pharmacological inhibition improves trophoblast function under inflammatory stress. We profiled mRNA expression of genes encoding enzymes that catalyze poly-ADP-ribosylation (PARP1, PARP2, TNKS1, TNKS2) and selected enzymes that catalyze mono-ADP-ribosylation ( PARP7 , PARP10 , PARP12 , and PARP14 ), in human I-PE placentas (control n=23; I-PE n=10) and an LPS-induced rat model of I-PE (control n=8, LPS n=8). HTR-8/SVneo cells were treated with TNF-α alone or with olaparib, (PARP1/2 inhibitor) or XAV939 (TNKS1/2 inhibitor). Global mono- and poly-ADP-ribosylation (MAR/PAR), including a PARP1-associated ∼120-kDa ADP-ribosylation signal and pathway enrichment of proteins identified from this region, were assessed alongside mitochondrial respiration, OXPHOS protein abundance, 4-hydroxynonenal, migration, and viability. In human I-PE placentas, mRNA abundance of poly-ADP-ribosylating enzymes did not differ significantly from controls, whereas mono-ADP-ribosylating enzymes were increased in LPS-exposed rat placentas. In HTR-8/SVneo cells, TNF-α induced a prominent PARP1 associated ∼120-kDa MAR/PAR signal that was suppressed by olaparib, but not XAV939. Proteins identified from this ∼120-kDa region were enriched for known PARP1 targets and immune/stress-response pathways. Olaparib, but not XAV939, improved basal and maximal respiration, ATP production, and spare respiratory capacity. TNF-α did not significantly alter oxidative phosphorylation (OXPHOS) subunits, 4-HNE abundance, viability, or migration at 24 h. These data support PARP1/2-sensitive ADP-ribosylation, most consistent with PARP1-linked activity, as a contributor to inflammation-mediated trophoblast bioenergetic dysfunction. The PARP1/2-NAD ⁺ axis may therefore represent a mechanism-directed target for future studies of I-PE. Graphical Highlights: TNF-α-induced PARP1-associated trophoblast ADP-ribosylation (MAR/PAR) Olaparib suppresses the TNF-α-induced, PARP1-associated ∼120-kDa MAR/PAR signal PARP1/2 inhibition improves TNF-α-impaired mitochondrial respiration Tankyrase inhibition does not rescue inflammation-driven trophoblast dysfunction

Opioid and cannabis co-use during pregnancy is increasingly common, yet the placental mechanisms linking combined exposure to adverse fetal outcomes remain poorly defined. Using a controlled mouse model of gestational drug exposure, we tested how fentanyl, Δ ⁹ -tetrahydrocannabinol (THC), or their combination altered placental structure, immune signaling, and gene expression and how these changes related to fetal growth. Drug exposure produced fetal growth restriction and reduced placental efficiency, with the greatest impairment in the combined fentanyl+THC group. Placental alterations were detectable by mid-gestation, when fentanyl exposure increased nucleated red blood cells within the labyrinth, consistent with hypoxic stress. By term, placentas showed compartment-specific remodeling, with THC selectively expanding the labyrinth and combined fentanyl+THC exposure increasing decidual area. Labyrinth composition and proliferative activity were altered, ultrastructural analysis revealed disruption of the maternal-fetal exchange interface, and placental interleukin-10 and interferon-β levels were reduced across exposure groups. Transcriptomic analyses identified suppression of innate immune and antiviral defense pathways together with treatment-specific stress responses, and integration of placental gene expression with fetal weight showed coordinated repression of vascular and developmental regulators and activation of hypoxia- and metabolic stress-associated genes. These findings identify the placenta as a key mediator of adverse fetal outcomes associated with prenatal polysubstance exposure. Significance Statement: Opioids and cannabis are increasingly used together during pregnancy, but how this combination harms the developing fetus is unclear. Using a mouse model of chronic fentanyl and THC exposure, we show that combined exposure disrupts placental structure, suppresses immune signaling, and reprograms gene networks linked to fetal growth. These placental abnormalities appear before or alongside fetal growth restriction and are most severe with combined exposure. Our findings identify the placenta as a key biological target of prenatal polysubstance exposure and provide a mechanistic framework for understanding why opioid-cannabis co-use may worsen pregnancy outcomes.

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.

The COVID-19 pandemic intensified food insecurity (FI) and stress for many pregnant individuals, which may have contributed to adverse fetal developmental programming. This study aimed to identify key social determinants of health associated with pandemic-related FI and stress, and their association with gestational weight gain (GWG) and newborn birth weight in a Canadian pregnant cohort. Data were collected retrospectively from 273 pregnant individuals who delivered infants in Canada during the pandemic (March 2020–March 2023). Validated questionnaires were used to assess FI and pandemic-related stress, and GWG and infant birth weight were self-reported. FI was experienced by 55.7% of the participants, while 33.7% and 19.7% reported heightened stress related to COVID-19 infection and pregnancy preparedness, respectively. Participants from food-secure and food-insecure households differed significantly in parental structure, age, sexual orientation, housing status, household income, number of children in the household and pregnancy planning (all p values < 0.01). Heightened stress for both pregnancy preparedness and COVID-19 infection was also significantly associated with these same factors (all p values < 0.05) but not for age and housing status. FI and heightened stress were not associated with GWG outside the recommended range. However, significantly higher likelihood of birth weight extremes was observed with heightened COVID-19 infection-related stress (OR, 95% CI 1.50, 1.05–2.12, p = 0.02) and pregnancy preparedness-related stress (1.60, 1.10–2.31, p = 0.01), but not with FI. These findings underscore the influence of psychosocial factors on FI and stress during pregnancy, which may negatively impact infant health outcomes during the pandemic.

Background: Despite the effectiveness of hepatitis B virus (HBV) antiviral treatment therapy in reducing the risk of liver-related complications, such as cirrhosis and hepatocellular carcinoma among chronically infected patients, medication non-adherence continues to hamper the successful management of the infection. The extent of HBV treatment adherence, associated facilitators, and barriers in South Sudan is not established. This study aimed to explore the predictors of medication non-adherence among HBV patients attending a public health facility in, South Sudan. Methods: We conducted a facility-based cross-sectional study of 392 convenience-selected patients using a pretested interviewer-administered questionnaire premised on the information-motivation-behavioral skills (IMB) adherence model between December 2023 and March 2024. The relationship between medication non-adherence and antecedent variables was ascertained by logistic regression analysis. Results: The sample was predominantly male (64.3%), and the mean age was 31.06 (30.19– 31.93) years, with 28.1% reporting no formal education. The patients demonstrated inadequate HBV information (4.33± 1.93), low motivation (8.20± 2.69), and inadequate behavioral skills toward medication adherence (8.45± 2.99), as measured on their respective rating scales. Further, more than two-thirds of the patients (70.2%) were HBV medication non-adherent. Younger age (AOR = 4.74, 95% CI = 2.13– 10.56), being currently unmarried (AOR = 3.25, 95% CI = 1.76– 6.01), unemployment (AOR = 4.19, 95% CI = 1.84– 9.56), and increased behavioral skills (AOR = 1.12, 95% CI = 1.84– 9.56) significantly influenced medication non-adherence. Lower education (AOR = 0.21, 95% CI = 0.10– 0.46) and information adequacy (AOR = 0.63, 95% CI = 0.53– 0.75) were associated with lower odds of non-adherence. Conclusion: The study highlights key factors influencing the concerning rate of medication non-adherence among HBV patients in South Sudan. While these identified factors may explain the lingering burden of HBV-related complications, targeted interventions addressing demographic, socioeconomic barriers, and HBV-specific education are essential to enhance adherence and improve health outcomes.

The placenta plays a critical role in nutrient and oxygen exchange during pregnancy, yet the effects of medicinal drugs on this selective barrier remain poorly understood. To overcome this, this study presents a cost-effective bioimpedance spectroscopy (BIS) system to assess tight junction integrity and monolayer formation in BeWo b30 cells, a widely used model of the multinucleated maternal–fetal exchange surface of the placental barrier. Cells were cultured on collagen-coated porous membranes and treated with forskolin to induce controlled syncytialization. Electrical impedance was measured using an entry level impedance analyzer, while immunofluorescence staining was used to confirm monolayer formation and syncytialization. The measurements and staining confirmed the formation of a confluent monolayer on day 4. In fact, the electrical resistance tripled for treated samples indicating a more electrically restrictive barrier. This resistance remained constant for treated samples reflecting the intact barrier’s integrity over the next 3 days. The measurements show that, on day 4, the electrical capacitance of the cells decreased for the treated samples as opposed to the untreated samples. This reflects that the surface area of the BeWo b30 cells decreased when the samples were treated with forskolin. Finally, a COMSOL model was developed to explore the effects of electrode positioning, depth, and distance on TEER measurements, explaining discrepancies in the literature. In fact, there was a substantial 97% and 39.4% difference in the obtained TEER values. This study demonstrates the AD2 device’s feasibility for monitoring placental barrier integrity and emphasizes the need for standardized setups for comparable results. The system can hence be used to analyze drug effects and nutrient transfer across the placental barrier.

Preeclampsia (PE) is a hypertensive disorder of pregnancy and a major cause of maternal/perinatal adverse health outcomes with no effective therapeutic strategies. Our group previously identified distinct subclasses of PE, one of which exhibits heightened placental inflammation (inflammation-driven PE). In non-pregnant populations, chronic inflammation is associated with decreased levels of cellular NAD⁺, a vitamin B3 derivative involved in energy metabolism and mitochondrial function. Interestingly, specifically in placentas from women with inflammation-driven PE, we observed the increased activity of NAD⁺-consuming enzymes, decreased NAD⁺content, decreased expression of mitochondrial proteins, and increased oxidative damage. HTR8 human trophoblasts likewise demonstrated increased NAD⁺-dependent ADP-ribosyltransferase (ART) activity, coupled with decreased mitochondrial respiration rates and invasive function under inflammatory conditions. Such adverse effects were attenuated by boosting cellular NAD⁺levels with nicotinamide riboside (NR). Finally, in an LPS-induced rat model of inflammation-driven PE, NR administration (200 mg/kg/day) from gestational days 1–19 prevented maternal hypertension and fetal/placental growth restriction, improved placental mitochondrial function, and reduced inflammation and oxidative stress. This study demonstrates the critical role of NAD⁺in maintaining placental function and identifies NAD⁺boosting as a promising preventative strategy for PE.