The Susceptibility of Chickens to Zika Virus: A Comprehensive Study on Age-Dependent Infection Dynamics and Host Responses.

Zika virus (ZIKV) remains a public health concern, with epidemics in endemic regions and sporadic outbreaks in new areas posing significant threats. Several mosquito-borne flaviviruses that can cause human illness, including West Nile, Usutu, and St. Louis encephalitis, have associations with birds. However, the susceptibility of chickens to ZIKV and their role in viral epidemiology is not currently known. We investigated the susceptibility of chickens to experimental ZIKV infection using chickens ranging from 1-day-old chicks to 6-week-old birds. ZIKV caused no clinical signs in chickens of all age groups tested. Viral RNA was detected in the blood and tissues during the first 5 days post-inoculation in 1-day and 4-day-old chicks inoculated with a high viral dose, but ZIKV was undetectable in 6-week-old birds at all timepoints. Minimal antibody responses were observed in 6-week-old birds, and while present in younger chicks, they waned by 28 days post-infection. Innate immune responses varied significantly between age groups. Robust type I interferon and inflammasome responses were measured in older chickens, while limited innate immune activation was observed in younger chicks. Signal transducer and activator of transcription 2 (STAT2) is a major driver of host restriction to ZIKV, and chicken STAT2 is distinct from human STAT2, potentially contributing to the observed resistance to ZIKV infection. The rapid clearance of the virus in older chickens coincided with an effective innate immune response, highlighting age-dependent susceptibility. Our study indicates that chickens are not susceptible to productive ZIKV infection and are unlikely to play a role in the ZIKV epidemiology.

Inactivation of HCoV-NL63 and SARS-CoV-2 in aqueous solution by 254 nm UV-C.

Ultraviolet germicidal irradiation (UVGI) is a highly effective means of inactivating many bacteria, viruses, and fungi. UVGI is an attractive viral mitigation strategy against coronaviruses, including the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of the coronavirus disease-2019 (COVID-19) pandemic. This investigation measures the susceptibility of two human coronaviruses to inactivation by 254 nm UV-C radiation. Human coronavirus NL63 and SARS-CoV-2 were irradiated in a collimated, dual-beam, aqueous UV reactor. By measuring fluence and integrating it in real-time, this reactor accounts for the lamp output transients during UVGI exposures. The inactivation rate constants of a one-stage exponential decay model were determined to be 2.050 cm2/mJ and 2.098 cm2/mJ for the NL63 and SARS-CoV-2 viruses, respectively. The inactivation rate constant for SARS-CoV-2 is within 2% of that of NL63, indicating that in identical inactivation environments, very similar UV 254 nm deactivation susceptibilities for these two coronaviruses would be achieved. Given the inactivation rate constant obtained in this study, doses of 1.1 mJ/cm2, 2.2 mJ/cm2, and 3.3 mJ/cm2 would result in a 90%, 99%, and 99.9% inactivation of the SARS-CoV-2 virus, respectively. The inactivation rate constant obtained in this study is significantly higher than values reported from many 254 nm studies, which suggests greater UV susceptibility to the UV-C than what was believed. Overall, results from this study indicate that 254 nm UV-C is effective for inactivation of human coronaviruses, including SARS-CoV-2.

The epidermal growth factor receptor in healthy pregnancy and preeclampsia.

The epidermal growth factor receptor (EGFR) is expressed robustly in the placenta, and critical processes of pregnancy such as placental growth and trophoblast fusion are dependent on EGFR function. However, the role that aberrant EGFR signaling might play in the etiology and/or maintenance of preeclampsia (PE) remains largely unexplored. Recently, we have shown that overexpression of EGFR in cultured uterine artery endothelial cells (UAEC), which express little endogenous EGFR, remaps responsiveness away from vascular endothelial growth factor receptor (VEGFR) signaling and toward EGFR, suggesting that endothelial EGFR expression may be kept low to preserve VEGFR control of angiogenesis. Here we will consider the evidence for the possibility that the endothelial dysfunction observed in PE might in some cases result from elevation of endothelial EGFR. During pregnancy, trophoblasts are known to synthesize large amounts of EGFR protein, and the placenta regularly releases syncytiotrophoblast-derived exosomes and microparticles into the maternal circulation. Although there are no reports of elevated EGFR gene expression in preeclamptic endothelial cells, the ongoing shedding of placental vesicles into the vascular system raises the possibility that EGFR-rich vesicles might fuse with endothelium, thereby contributing to the symptoms of PE by interrupting angiogenesis and blocking pregnancy-adapted vasodilatory function.

Experimentally Induced Hyperinsulinemia Fails to Induce Polycystic Ovary Syndrome-like Traits in Female Rhesus Macaques.

As in women with polycystic ovary syndrome (PCOS), hyperinsulinemia is associated with anovulation in PCOS-like female rhesus monkeys. Insulin sensitizers ameliorate hyperinsulinemia and stimulate ovulatory menstrual cycles in PCOS-like monkeys. To determine whether hyperinsulinemia (>694 pmol/L), alone, induces PCOS-like traits, five PCOS-like female rhesus monkeys with minimal PCOS-like traits, and four control females of similar mid-to-late reproductive years and body mass index, received daily subcutaneous injections of recombinant human insulin or diluent for 6−7 months. A cross-over experimental design enabled use of the same monkeys in each treatment phase. Insulin treatment unexpectedly normalized follicular phase duration in PCOS-like, but not control, females. In response to an intramuscular injection of 200 IU hCG, neither prenatally androgenized nor control females demonstrated ovarian hyperandrogenic responses while receiving insulin. An intravenous GnRH (100 ng/kg) injection also did not reveal evidence of hypergonadotropism. Taken together, these results suggest that experimentally induced adult hyperinsulinemia, alone, is insufficient to induce PCOS-like traits in female rhesus monkeys and to amplify intrinsic PCOS-like pathophysiology.

Perinatal Research Society's Young Investigator Workshop Prepares the Next Generation of Investigators.

Sustaining impactful research within the field of perinatal biology requires training and retention of the next generations of physician-scientists and basic-scientists. Professional societies such as the Perinatal Research Society (PRS) have a unique role to play in training and retention of perinatal biologists. Here we report outcomes for an innovative Young Investigator Training Workshop created for the PRS. The PRS Workshop uses immersive, active-writing, and active-oral presentation design, with one-on-one feedback from NIH-funded faculty-mentors drawn from the PRS membership. Young investigator data were collected by anonymous surveys of young investigators, NIH RePORTER, and individual young investigator follow-up. Ninety-seven young investigators attended the Workshops over the period 2013-2018. Young investigators were physician- (73%) and PhD- (27%) scientists at the rank of clinical fellow/postdoctoral fellow (27%) or instructor/assistant professor (73%). Participation by underrepresented minority (URM) young investigators was 14%. Young investigators received NIH and non-NIH funding, with 80% of young investigators receiving new funding since the Workshop that they attended. NIH funding was received by 31% of young investigators in the form of K-series awards, R01 equivalents, and other NIH awards. In conclusion, our PRS young investigator Workshop serves as a model to facilitate training of emerging physician- and basic-scientists by scientific societies.

Differential control of uterine artery endothelial monolayer integrity by TNF and VEGF is achieved through multiple mechanisms operating inside and outside the cell - Relevance to preeclampsia.

Uterine artery endothelium undergoes a form of functional adaptation during pregnancy because of an increase in Cx43 communication, resulting in increased Ca2+/IP3 exchange and more synchronous and sustained vasodilator production. We have shown previously that acute exposure to growth factors and TNF can block this adaptation through ERK and/or Src-mediated Cx43 phosphorylation. In preeclampsia such adapted function is already missing, but while elevated TNF is associated with this condition, particularly after 28 weeks (late PE), elevated circulating VEGF165 is not. Given PE is a long term condition emerging in the second half of pregnancy, and is often associated with added edema, we now compare the chronic effects of these two factors on the cell monolayer in order to establish if the breakdown of junctional adherens and tight junctional assemblies in which Cx43 resides could also explain loss of vasodilatory function. We report that while TNF can degrade monolayer integrity even in the 0.1-1 ng/ml physiologic range, VEGF up to 10 ng/ml does not. In addition, the progressive action of TNF is mediated through Src and ERK signaling to promote internalization and destruction of VE-Cadherin (VE-Cad) and ZO-1, as well as the expression and secretion of a variety of proteases. At least one protein degraded from the extracellular space is VE-Cad, resulting in release of a shed VE-Cad protein product, and consistent with monolayer breakdown being sensitive to both Src and MEK/ERK kinase inhibitors and the general protease inhibitor GM6001. We conclude that the greater association of TNF with 'late' PE is as much due to its longer term destabilizing effects on junctional assemblies as it is to acute closure of Cx43 channels themselves. New therapies aimed at stabilizing these junctional assemblies may help treat this hypertensive condition.

Limited window for donation of convalescent plasma with high live-virus neutralizing antibody titers for COVID-19 immunotherapy.

Millions of individuals who have recovered from SARS-CoV-2 infection may be eligible to participate in convalescent plasma donor programs, yet the optimal window for donating high neutralizing titer convalescent plasma for COVID-19 immunotherapy remains unknown. Here we studied the response trajectories of antibodies directed to the SARS-CoV-2 surface spike glycoprotein and in vitro SARS-CoV-2 live virus neutralizing titers (VN) in 175 convalescent donors longitudinally sampled for up to 142 days post onset of symptoms (DPO). We observed robust IgM, IgG, and viral neutralization responses to SARS-CoV-2 that persist, in the aggregate, for at least 100 DPO. However, there is a notable decline in VN titers ≥160 for convalescent plasma therapy, starting 60 DPO. The results also show that individuals 30 years of age or younger have significantly lower VN, IgG and IgM antibody titers than those in the older age groups; and individuals with greater disease severity also have significantly higher IgM and IgG antibody titers. Taken together, these findings define the optimal window for donating convalescent plasma useful for immunotherapy of COVID-19 patients and reveal important predictors of an ideal plasma donor.

Newborn Screening for Congenital Adrenal Hyperplasia: Review of Factors Affecting Screening Accuracy.

Newborn screening for 21-hydroxylase deficiency (21OHD), the most common form of congenital adrenal hyperplasia, has been performed routinely in the United States and other countries for over 20 years. Screening provides the opportunity for early detection and treatment of patients with 21OHD, preventing salt-wasting crisis during the first weeks of life. However, current first-tier screening methodologies lack specificity, leading to a large number of false positive cases, and adequate sensitivity to detect all cases of classic 21OHD that would benefit from treatment. This review summarizes the pathology of 21OHD and also the key stages of fetal hypothalamic-pituitary-adrenal axis development and adrenal steroidogenesis that contribute to limitations in screening accuracy. Factors leading to both false positive and false negative results are highlighted, along with specimen collection best practices used by laboratories in the United States and worldwide. This comprehensive review provides context and insight into the limitations of newborn screening for 21OHD for laboratorians, primary care physicians, and endocrinologists.

SARS-CoV-2 RBD Neutralizing Antibody Induction is Enhanced by Particulate Vaccination.

The receptor-binding domain (RBD) of the SARS-CoV-2 spike protein is a candidate vaccine antigen that binds angiotensin-converting enzyme 2 (ACE2), leading to virus entry. Here, it is shown that rapid conversion of recombinant RBD into particulate form via admixing with liposomes containing cobalt-porphyrin-phospholipid (CoPoP) potently enhances the functional antibody response. Antigen binding via His-tag insertion into the CoPoP bilayer results in a serum-stable and conformationally intact display of the RBD on the liposome surface. Compared to other vaccine formulations, immunization using CoPoP liposomes admixed with recombinant RBD induces multiple orders of magnitude higher levels of antibody titers in mice that neutralize pseudovirus cell entry, block RBD interaction with ACE2, and inhibit live virus replication. Enhanced immunogenicity can be accounted for by greater RBD uptake into antigen-presenting cells in particulate form and improved immune cell infiltration in draining lymph nodes. QS-21 inclusion in the liposomes results in an enhanced antigen-specific polyfunctional T cell response. In mice, high dose immunization results in minimal local reactogenicity, is well-tolerated, and does not elevate serum cobalt levels. Taken together, these results confirm that particulate presentation strategies for the RBD immunogen should be considered for inducing strongly neutralizing antibody responses against SARS-CoV-2.

Influenza C and D viral load in cattle correlates with bovine respiratory disease (BRD): Emerging role of orthomyxoviruses in the pathogenesis of BRD.

Bovine respiratory disease (BRD) is the costliest disease affecting the cattle industry globally. Orthomyxoviruses, influenza C virus (ICV) and influenza D virus (IDV) have recently been implicated to play a role in BRD. However, there are contradicting reports about the association of IDV and ICV to BRD. Using the largest cohort study (cattle, n = 599) to date we investigated the association of influenza viruses in cattle with BRD. Cattle were scored for respiratory symptoms and pooled nasal and pharyngeal swabs were tested for bovine viral diarrhea virus, bovine herpesvirus 1, bovine respiratory syncytial virus, bovine coronavirus, ICV and IDV by real-time PCR. Cattle that have higher viral loads of IDV and ICV also have greater numbers of co-infecting viruses than controls. More strikingly, 2 logs higher IDV viral RNA in BRD-symptomatic cattle that are co-infected animals than those infected with IDV alone. Our results strongly suggest that ICV and IDV may be significant contributors to BRD.

Convalescent plasma anti-SARS-CoV-2 spike protein ectodomain and receptor-binding domain IgG correlate with virus neutralization.

The newly emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) highlights the urgent need for assays that detect protective levels of neutralizing antibodies. We studied the relationship among anti-spike ectodomain (anti-ECD), anti-receptor-binding domain (anti-RBD) IgG titers, and SARS-CoV-2 virus neutralization (VN) titers generated by 2 in vitro assays using convalescent plasma samples from 68 patients with COVID-19. We report a strong positive correlation between both plasma anti-RBD and anti-ECD IgG titers and in vitro VN titers. The probability of a VN titer of ≥160, the FDA-recommended level for convalescent plasma used for COVID-19 treatment, was ≥80% when anti-RBD or anti-ECD titers were ≥1:1350. Of all donors, 37% lacked VN titers of ≥160. Dyspnea, hospitalization, and disease severity were significantly associated with higher VN titer. Frequent donation of convalescent plasma did not significantly decrease VN or IgG titers. Analysis of 2814 asymptomatic adults found 73 individuals with anti-ECD IgG titers of ≥1:50 and strong positive correlation with anti-RBD and VN titers. Fourteen of these individuals had VN titers of ≥1:160, and all of them had anti-RBD titers of ≥1:1350. We conclude that anti-RBD or anti-ECD IgG titers can serve as a surrogate for VN titers to identify suitable plasma donors. Plasma anti-RBD or anti-ECD titers of ≥1:1350 may provide critical information about protection against COVID-19 disease.

NLRC5 Serves as a Pro-viral Factor During Influenza Virus Infection in Chicken Macrophages.

Avian influenza viruses (AIVs) cause major economic losses to the global poultry industry. Many host factors have been identified that act as regulators of the inflammatory response and virus replication in influenza A virus (IAV) infected cells including nucleotide-binding oligomerization domain (NOD) like receptor (NLR) family proteins. Evidence is emerging that NLRC5, the largest NLR member, is a regulator of host immune responses against invading pathogens including viruses; however, its role in the avian immune system and AIV pathogenesis has not been fully explored. In this study, we found that NLRC5 is activated by a range of low and highly pathogenic AIVs in primary chicken lung cells and a chicken macrophage cell line. Further, siRNA mediated NLRC5 knockdown in chicken macrophages resulted in a significant reduction in AIV replication which was associated with the upregulation of genes associated with activated NFκB signaling pathway. The knockdown of NLRC5 enhanced the expression of genes known to be associated with viral defense and decreased innate cytokine gene expression following AIV infection. Overall, our investigation strongly suggests that NLRC5 is a pro-viral factor during IAV infection in chicken and may contribute to pathogenesis through innate cytokine regulation. Further studies are warranted to investigate the IAV protein(s) that may regulate activation of NLRC5.

Relationship between Anti-Spike Protein Antibody Titers and SARS-CoV-2 In Vitro Virus Neutralization in Convalescent Plasma.

Newly emerged pathogens such as SARS-CoV-2 highlight the urgent need for assays that detect levels of neutralizing antibodies that may be protective. We studied the relationship between anti-spike ectodomain (ECD) and anti-receptor binding domain (RBD) IgG titers, and SARS-CoV-2 virus neutralization (VN) titers generated by two different in vitro assays using convalescent plasma samples obtained from 68 COVID-19 patients, including 13 who donated plasma multiple times. Only 23% (16/68) of donors had been hospitalized. We also studied 16 samples from subjects found to have anti-spike protein IgG during surveillance screening of asymptomatic individuals. We report a strong positive correlation between both plasma anti-RBD and anti-ECD IgG titers, and in vitro VN titer. Anti-RBD plasma IgG correlated slightly better than anti-ECD IgG titer with VN titer. The probability of a VN titer ≥160 was 80% or greater with anti-RBD or anti-ECD titers of ≥1:1350. Thirty-seven percent (25/68) of convalescent plasma donors lacked VN titers ≥160, the FDA-recommended level for convalescent plasma used for COVID-19 treatment. Dyspnea, hospitalization, and disease severity were significantly associated with higher VN titer. Frequent donation of convalescent plasma did not significantly decrease either VN or IgG titers. Analysis of 2,814 asymptomatic adults found 27 individuals with anti-RBD or anti-ECD IgG titers of ≥1:1350, and evidence of VN ≥1:160. Taken together, we conclude that anti-RBD or anti-ECD IgG titers can serve as a surrogate for VN titers to identify suitable plasma donors. Plasma anti-RBD or anti-ECD titer of ≥1:1350 may provide critical information about protection against COVID-19 disease.

Iminosugars With Endoplasmic Reticulum α-Glucosidase Inhibitor Activity Inhibit ZIKV Replication and Reverse Cytopathogenicity in vitro.

Zika virus (ZIKV), a vector-borne virus of the family Flaviviridae, continues to spread and remains a significant global public health threat. Currently, there are no approved vaccines or antivirals against ZIKV. We investigated the anti-ZIKV ability of three iminosugars with endoplasmic reticulum α-glucosidase inhibitor (ER-AGI) activity, namely deoxynojirimycin (DNJ), castanospermine, and celgosivir. None of the three iminosugars showed any significant cytotoxicity in Vero or human microglia CHME3 cells when applied for 72 h at concentrations up to 100 µM. Iminosugar treatment of Vero or CHME3 cells prior to ZIKV infection resulted in significant inhibition of ZIKV replication over 48 h. Reduction in ZIKV replication in iminosugar-treated cells was not associated with any significant change in the expression levels of key antiviral genes. Following infection with three different strains of ZIKV, iminosugar-treated Vero or CHME3 cells showed no cell death, whereas vehicle-treated control cells exhibited 50-60% cell death at 72 h post-infection (hpi). While there was no significant difference in apoptosis between iminosugar-treated and control cells, iminosugar-treated cells exhibited a substantial reduction of necrosis at 72 hpi following ZIKV infection. In summary, iminosugars with ER-AGI activity inhibit ZIKV replication and significantly reduce necrosis without altering the antiviral gene expression and apoptosis of infected human cells. The results of this study strongly suggest that iminosugars are promising anti-ZIKV antiviral agents and such warrant further in vivo studies.

Conjugated linoleic acid improves endothelial Ca2+ signaling by blocking growth factor and cytokine-mediated Cx43 phosphorylation.

Sustained Ca2+ burst signaling is crucial for endothelial vasodilator production and is disrupted by growth factors and cytokines. Conjugated linoleic acid (CLA), a Src inhibitor in certain preparations, is generally regarded as safe during pregnancy by the FDA. Multiple CLA preparations; t10, c12 or c9, t11 CLA, or a 1:1 mixture of the two were administered before growth factor or cytokine treatment. Growth factors and cytokines caused a significant decrease in Ca2+ burst numbers in response to ATP stimulation. Both t10, c12 CLA and the 1:1 mixture rescued VEGF165 or TNFα inhibited Ca2+ bursts and correlated with Src-specific phosphorylation of connexin 43. VEGF165, TNFα, and IL-6 in combination at physiologic concentrations revealed IL-6 amplified the inhibitory effects of lower dose of VEGF165 and TNFα. Again, the 1:1 CLA mixture was most effective at rescue of function. Therefore, CLA formulations may be a promising treatment for endothelial dysfunction in diseases such as preeclampsia.

Adenoviral transduction of EGFR into pregnancy-adapted uterine artery endothelial cells remaps growth factor induction of endothelial dysfunction.

During pregnancy, uterine vascular vasodilation is enhanced through adapted Ca2+ signaling, facilitated through increased endothelial connexin 43 (Cx43) gap junctional communication (GJC). In preeclampsia (PE), this adaptive response is missing. Of note, the angiogenic factor VEGF can also act via Src and ERK to close Cx43 gap junctions. While VEGFR2 is necessary for such closure, a role VEGFR1 is less clear. We reasoned if VEGFR2 is acting alone, then substituting another growth factor receptor with VEGFR2-like signaling should have the same effect. In uterine artery endothelial cells derived from pregnant sheep (P-UAEC), endogenous EGFR expression is very low. When we used adenovirus to raise EGFR, we also dose-dependently induced EGF-sensitive Cx43 phosphorylation mainly via ERK, and corresponding loss of Ca2+ bursts, but eliminated VEGF effects on phosphorylation of Cx43 or loss of Ca2+ bursting. This surprising observation suggests that while activated EGFR may indeed substitute for VEGFR2, it also sequesters a limited pool of effector molecules needed for VEGFR2 to phosphorylate Cx43. Thus, low endogenous EGFR expression in P-UAEC may be a necessary strategy to allow VEGFR-2 control of GJC, a first step in initiating angiogenesis in healthy pregnancy. Of further note, trophoblasts are rich in EGFR, and we have demonstrated shed PLAP+/EGFR + extracellular vesicles in maternal circulation in first trimester plasma samples using nanoscale high resolution flow cytometry. Collectively our data suggest that placenta derived exosomes positive for EGFR should be further considered as a possible cause of endothelial dysfunction in women with PE.

Evaluation of a motion-robust 2D chemical shift-encoded technique for R2* and field map quantification in ferumoxytol-enhanced MRI of the placenta in pregnant rhesus macaques.

BACKGROUND: 3D chemical shift-encoded (CSE)-MRI techniques enable assessment of ferumoxytol concentration but are unreliable in the presence of motion. PURPOSE: To evaluate a motion-robust 2D-sequential CSE-MRI for R2* and B0 mapping in ferumoxytol-enhanced MRI of the placenta. STUDY TYPE: Prospective. ANIMAL MODEL: Pregnant rhesus macaques. FIELD STRENGTH/SEQUENCE: 3.0T/CSE-MRI. ASSESSMENT: 2D-sequential CSE-MRI was compared with 3D respiratory-gated CSE-MRI in placental imaging of 11 anesthetized animals at multiple timepoints before and after ferumoxytol administration, and in ferumoxytol phantoms (0 µg/mL-440 µg/mL). Motion artifacts of CSE-MRI in 10 pregnant women without ferumoxytol administration were assessed retrospectively by three blinded readers (4-point Likert scale). The repeatability of CSE-MRI in seven pregnant women was also prospectively studied. STATISTICAL TESTS: Placental R2* and boundary B0 field measurements (ΔB0) were compared between 2D-sequential and 3D respiratory-gated CSE-MRI using linear regression and Bland-Altman analysis. RESULTS: In phantoms, a slope of 0.94 (r2 = 0.99, concordance correlation coefficient ρ = 0.99), and bias of -4.8 s-1 (limit of agreement [LOA], -41.4 s-1 , +31.8 s-1 ) in R2*, and a slope of 1.07 (r2 = 1.00, ρ = 0.99) and bias of 11.4 Hz (LOA -12.0 Hz, +34.8 Hz) in ΔB0 were obtained in 2D CSE-MRI compared with 3D CSE-MRI for reference R2* ≤390 s-1 . In animals, a slope of 0.92 (r2 = 0.97, ρ = 0.98) and bias of -2.2 s-1 (LOA -55.6 s-1 , +51.3 s-1 ) in R2*, and a slope of 1.05 (r2 = 0.95, ρ = 0.97) and bias of 0.4 Hz (LOA -9.0 Hz, +9.7 Hz) in ΔB0 were obtained. In humans, motion-impaired R2* maps in 3D CSE-MRI (Reader 1: 1.8 ± 0.6, Reader 2: 1.3 ± 0.7, Reader 3: 1.9 ± 0.6), while 2D CSE-MRI was motion-free (Reader 1: 2.9 ± 0.3, Reader 2: 3.0 ± 0, Reader 3: 3.0 ± 0). A mean difference of 0.66 s-1 and coefficient of repeatability of 9.48 s-1 for placental R2* were observed in the repeated 2D CSE-MRI. DATA CONCLUSION: 2D-sequential CSE-MRI provides accurate R2* and B0 measurements in ferumoxytol-enhanced placental MRI of animals in the presence of respiratory motion, and motion-robustness in human placental imaging. LEVEL OF EVIDENCE: 1 Technical Efficacy: Stage 1 J. Magn. Reson. Imaging 2020;51:580-592.

Quantitative ferumoxytol-enhanced MRI in pregnancy: A feasibility study in the nonhuman primate.

OBJECTIVES: To assess the feasibility of ferumoxytol-enhanced MRI in pregnancy with a nonhuman primate model. MATERIALS AND METHODS: In this prospective study, eleven pregnant rhesus macaques at day 98 ±â€¯5 of gestation were divided into three groups, untreated control (UC) (n = 3), saline control (SC) (n = 4) and interleukin 1 beta (IL-1ß) treated (IT) (n = 4), which were administered with either saline or IL-1ß into the amniotic fluid. All animals were imaged at multiple time points before and after ferumoxytol administration (4 mg/kg). Longitudinal R2* and susceptibility of tissues were obtained using region-of-interest analysis and the longitudinal changes were assessed using linear mixed models and Student's t-test. RESULTS: In fetuses, a slope of 0.3 s-1/day (P = 0.008), 0.00 ppm/day (P = 0.699) and - 0.2 s-1/day (P = 0.023) was observed in liver R2*, liver susceptibility, and lung R2*, respectively. In placentas, R2* and susceptibility increased immediately after ferumoxytol administration (P < 0.001) and decreased to baseline within two days. The mean change from baseline showed no significant difference between the SC group and the IT group at all scan time points. In maternal livers, R2* increased immediately after ferumoxytol administration, further increased at one-day, and then decreased but remained elevated (P < 0.001). The mean change from baseline showed no significant difference between the SC group and the IT group at all scan time points. CONCLUSIONS: This work demonstrates the feasibility of quantitative ferumoxytol-enhanced MRI to measure dynamics of ferumoxytol delivery and washout in the placenta. Stable MRI measurements indicated no evidence of iron deposition in fetal tissues of nonhuman primates after maternal ferumoxytol exposure.

Sexual Dimorphisms of Preeclampsia-Dysregulated Transcriptomic Profiles and Cell Function in Fetal Endothelial Cells.

Preeclampsia impairs fetoplacental vascular function and increases risks of adult-onset cardiovascular disorders in children born to preeclamptic mothers, implicating that preeclampsia programs fetal vasculature in utero. However, the underlying mechanisms remain elusive. We hypothesize that preeclampsia alters fetal endothelial gene expression and disturbs cytokines- and growth factors-induced endothelial responses. RNA sequencing analysis was performed on unpassaged human umbilical vein endothelial cells (HUVECs) from normotensive and preeclamptic pregnancies. Functional assays for endothelial monolayer integrity, proliferation, and migration were conducted on passage 1 HUVECs from normotensive and preeclamptic pregnancies. Compared with normotensive cells, 926 and 172 genes were dysregulated in unpassaged female and male HUVECs from preeclamptic pregnancies, respectively. Many of these preeclampsia-dysregulated genes are associated with cardiovascular diseases (eg, heart failure) and endothelial function (eg, cell migration, calcium signaling, and endothelial nitric oxide synthase signaling). TNF (tumor necrosis factor)-α-, TGF (transforming growth factor)-ß1-, FGF (fibroblast growth factor)-2-, and VEGFA (vascular endothelial growth factor A)-regulated gene networks were differentially disrupted in unpassaged female and male HUVECs from preeclamptic pregnancies. Moreover, preeclampsia decreased endothelial monolayer integrity in responses to TNF-α in both female and male HUVECs. Preeclampsia decreased TGF-ß1-strengthened monolayer integrity in female HUVECs, whereas it enhanced FGF-2-strengthened monolayer integrity in male HUVECs. Preeclampsia promoted TNF-α-, TGF-ß1-, and VEGFA-induced cell proliferation in female, but not in male HUVECs. Preeclampsia inhibited TNF-α-induced cell migration in female HUVECs, but had an opposite effect on male HUVECs. In conclusion, preeclampsia differentially dysregulates cardiovascular diseases- and endothelial function-associated genes/pathways in female and male fetal endothelial cells in association with the sexual dimorphisms of preeclampsia-dysregulated fetal endothelial function.