Placenta

Abstract1IUGR is associated with increased risk of fetal compromise, yet can be difficult to detect and phenotype with routine clinical surveillance. Given placental insufficiency and hypoxia can remodel placental energy metabolism, methods that directly assess placental metabolic function may improve identification and phenotyping of growth-restricted pregnancies. We combined structural, body composition, and hyperpolarized metabolic MRI to characterize a near-term spontaneous IUGR (spIUGR) phenotype in the guinea pig. Twenty-two pregnant guinea pig sows (71 fetuses) underwent 1H and hyperpolarized 13C MRI at 60 {+/-} 1 days gestation to quantify fetal and placental volumes, maternal/fetal body composition, and quantify placental pyruvate metabolism. Fetuses were classified as spIUGR when [≥] 3 of 5 established markers were present (body weight, brain-body, brain-liver, brain-placenta ratios, and body weight relative to pregnancy mean); corresponding volume cut-offs were derived from weight cut-offs. MRI-derived fetal and placental volumes correlated strongly with collection weights and classified spIUGR consistently with weight-based criteria. Maternal adiposity (subcutaneous and visceral) was inversely associated with fetal adipose tissue volume, and maternal visceral fat PDFF was negatively associated with fetal adipose PDFF. Hyperpolarized MRI demonstrated IUGR phenotype-dependent placental pyruvate routing: LPR increased with asymmetric (brain-sparing) growth, showing a positive association with brain-body volume ratio (p = 0.02) and brain-body weight ratio (p = 0.04). In contrast, BPR was not significantly related to brain-body ratios (p = 0.09-0.11) but was inversely associated with absolute fetal size (body volume and weight, p = 0.02). These findings validate MRI volumetry for non-invasive identification of placental insufficiency spIUGR and link growth restriction severity and asymmetry to distinct placental metabolic signatures measurable in vivo.

BackgroundThe relationship between maternal risk factors (MRFs) (particularly pre-gravid obesity, diabetes, and hypertension) and congenital heart disease (CHD) to placental and fetal brain outcomes is poorly understood. Here, we tested the hypothesis that MRF and CHD would be associated with reduced intrinsic placental and fetal brain function using a novel non-invasive technique. MethodsPregnant participants with and without MRF and fetal CHD were prospectively recruited and underwent feto-placental MRI. Using intrinsic properties of blood oxygen level dependent imaging (BOLD) we quantified spatiotemporal variance of placenta and fetal brain. MRFs and CHD were correlated with functional characteristics of the placenta and fetal brain. ResultsCo- morbid MRF (hypertension, diabetes, and obesity) reduced spatiotemporal functional variance of placenta and fetal brain (p < 0.05). CHD predicted reduced fetal brain temporal variance compared to controls (p<0.05). Interaction of MRF and CHD status was associated with reduced intrinsic pBOLD temporal variance (p=0.047). There were no significant interactions of MRFs and CHD status on either temporal or spatial variance of intrinsic brain BOLD. ConclusionMRF and CHD reduced functional characteristic of placenta and brain in fetuses. MRF modification and management during pregnancy may have the potential to not only provide additional risk stratification but may also improve neurodevelopmental outcomes.

The placenta undergoes morphological and functional adaptions to adverse exposures during pregnancy. The effects of suboptimal maternal body mass index (BMI), preterm birth, and infection on placental histopathological phenotypes remain unclear, despite the association between these conditions and poor offspring outcomes. We hypothesized that suboptimal maternal prepregnancy BMI and preterm birth (with and without infection) would associate with altered placental maturity and morphometry, and that altered placental maturity would associate with poor birth outcomes. Clinical data and human placentae were collected from 96 pregnancies where mothers were underweight, normal weight, overweight, or obese, without other major complications. Placental histopathological characteristics were scored with an anatomical pathologist. Associations between maternal BMI, placental pathology (immaturity and hypermaturity), placental morphometry, and infant outcomes were investigated at term and preterm, with and without infection. Fetal vascular endothelium volumetric proportion was decreased, whereas syncytial knot volumetric proportion was increased, in placentae from preterm pregnancies with chorioamnionitis compared to term placentae. At term and preterm, pregnancies with overweight and obesity had a high percentage increase in proportion of immature placentae compared to normal weight. Placental maturity did not associate with infant birth outcomes. We observed placental hypermaturity and altered placental morphometry among preterm pregnancies with chorioamnionitis, suggestive of altered placental development, which may inform about pregnancies susceptible to preterm birth and infection. Our data increase our understanding of how common metabolic exposures and preterm birth, in the absence of other comorbidities or perinatal events, potentially contribute to poor pregnancy outcomes and the programming of offspring development.

IntroductionPlacenta accreta spectrum (PAS) occurs when the placenta is pathologically adherent to the myometrium. An intact retroplacental clear space (RPCS) is a marker of normal placentation, but visualization with conventional imaging techniques is a challenge. In this study, we investigate use of an FDA-approved iron oxide nanoparticle, ferumoxytol, for contrast-enhanced magnetic resonance imaging of the RPCS in mouse models of normal pregnancy and PAS. We then demonstrate the translational potential of this technique in human patients presenting with severe PAS (FIGO Grade 3C), moderate PAS (FIGO Grade 1), and no PAS. MethodsA T1-weighted gradient recalled echo (GRE) sequence was used to determine the optimal dose of ferumoxytol in pregnant mice. Pregnant Gab3-/- mice, which demonstrate placental invasion, were then imaged at day 16 of gestation alongside wild-type (WT) pregnant mice which do not demonstrate invasion. Signal-to-noise ratio (SNR) was computed for placenta and RPCS for all fetoplacental units (FPUs) with ferumoxytol-enhanced magnetic resonance imaging (Fe-MRI) and used for the determination of contrast-to-noise ratio (CNR). Fe-MRI was also performed in 3 pregnant subjects using standard T1 and T2 weighted sequences and a 3D magnetic resonance angiography (MRA) sequence. RPCS volume and relative signal were calculated in all three subjects. ResultsFerumoxytol administered at 5 mg/kg produced strong T1 shortening in blood and led to strong placental enhancement in Fe-MRI images. Gab3-/- mice demonstrated loss of hypointense region characteristic of the RPCS relative to WT mice in T1w Fe-MRI. CNR between RPCS and placenta was lower in FPUs of Gab3-/- mice compared to WT mice, indicating higher degrees of vascularization and interruptions throughout the space. In human patients, Fe-MRI at a dose of 5 mg/kg enabled high uteroplacental vasculature signal and quantification of the volume and signal profile in severe and moderate invasion of the placenta relative to a non-PAS case. DiscussionFerumoxytol, an FDA-approved iron oxide nanoparticle formulation, enabled visualization of abnormal vascularization and loss of uteroplacental interface in a murine model of PAS. The potential of this non-invasive visualization technique was then further demonstrated in human subjects. Diagnosis of placental invasion using Fe-MRI may provide a sensitive method for clinical detection of PAS.

Studying the human placenta through in vitro cell culture methods is necessary due to limited access and amenability of human placental tissue to certain experimental methods as well as distinct anatomical and physiological differences between animal and human placentas. Selecting an in vitro culture model of the human placenta is challenging due to representation of different trophoblast cell types with distinct biological roles and limited comparative studies that define key characteristics of these models. Therefore, the aim of this research was to create a comprehensive transcriptomic comparison of common in vitro models of the human placenta compared to bulk placental tissue from the CANDLE and GAPPS cohorts (N=1083). We performed differential gene expression analysis on publicly available RNA sequencing data from 6 common in vitro models of the human placenta (HTR-8/SVneo, BeWo, JEG-3, JAR, Primary Trophoblasts, and Villous Explants) and compared to CANDLE and GAPPS bulk placental tissue or cytotrophoblast, syncytiotrophoblast, and extravillous trophoblast cell types derived from bulk placental tissue. All in vitro placental models had a substantial number of differentially expressed genes (DEGs, FDR<0.01) compared to the CANDLE and GAPPS placentas (Average DEGs=10,873), and the individual trophoblast cell types (Average DEGs=5,346), indicating that there are vast differences in gene expression compared to bulk and cell-type specific human placental tissue. Hierarchical clustering identified 53 gene clusters with distinct expression profiles across placental models, with 22 clusters enriched for specific KEGG pathways, 7 clusters enriched for high-expression placental genes, and 7 clusters enriched for absorption, distribution, metabolism, and excretion genes. In vitro placental models were classified by fetal sex based on expression of Y-chromosome genes that identified HTR-8/SVneo cells as being of female origin, while JEG-3, JAR, and BeWo cells are of male origin. Overall, none of the models were a close approximation of the transcriptome of bulk human placental tissue, highlighting the challenges with model selection. To enable researchers to select appropriate models, we have compiled data on differential gene expression, clustering, and fetal sex into an accessible web application: "Comparative Transcriptomic Placental Model Atlas (CTPMA)" which can be utilized by researchers to make informed decisions about their selection of in vitro placental models.

Trophoblast invasion is essential for normal placentation, with failure resulting in a fetal growth restriction (FGR) phenotype. Utilizing a calorie-restricted mouse model, we report progressive epigenetic, molecular, and phenotypic placental changes throughout gestation. Following maternal caloric restriction initiated at E9, we observed a significant reduction in fetal and placental weights beginning at E12.5, with persistent growth restriction at E14.5, E16.5, and E17.5. Immunohistochemistry of the decidual invasion site at E17.5 demonstrated reduced 1) decidual depth, 2) trophoblast invasion distance, and 3) trophoblast quantity within the decidua. Preceding these phenotypic changes, RT-qPCR revealed downregulation of trophoblast invasion and angiogenesis genes, including MMP2, MMP9, EFNA1, Rac1, Rras, ASCL2, TRAP2C, Prl7b1, VEGFa, VEGFb, PDGF, and AKT3, beginning as early as E14.5. Notably, microRNA sequencing at E12.5, prior to these transcriptional changes, identified significant upregulation of miR-503-5p, a predicted inhibitor of several of these pathways. The summation of these observations suggests miR-503-5p may be an early driver of placental dysfunction in FGR, linking maternal malnutrition to impaired trophoblast invasion and angiogenesis. These findings provide insight into the molecular mechanisms underlying placental insufficiency and highlight miR-503-5p as a potential therapeutic target for improving pregnancy outcomes in FGR.

Intrauterine growth restriction (IUGR) is an obstetrical outcome where a fetus has not achieved its genetic potential. A consequence of IUGR is a decrease in brain myelin content. Myelin water imaging (MWI) has previously assessed fetal myelin water fraction (MWF) and can potentially assess myelination changes associated with IUGR. Thus, this study aims to quantify and compare the MWF of non-IUGR and IUGR fetal guinea pigs (GPs) in late gestation. Our sample consisted of 22 pregnant Dunkin-Hartley GPs with 71 fetuses (34 male) [mean {+/-} standard deviation: 60 {+/-} 1.2 days gestation]. Eight SPGR volumes [flip angles (): 2{degrees} - 16{degrees}], and two sets of 8 bSSFP volumes (: 8{degrees} - 64{degrees}), at 0{degrees} and 180{degrees} phase increments were acquired at 3.0 T. MWF maps were generated for each fetal GP brain using multicomponent driven equilibrium single pulse observation of T1/T2 (mcDESPOT). Regions of interest (ROIs) were placed in the fetal corpus callosum (CC), fornix (FOR), and parasagittal white matter (PSW). Linear regression was performed between five fetal IUGR markers [body volume (BV), body-to-pregnancy volume ratio (BPrVR), brain-to-liver VR (BLVR), brain-to-placenta VR (BPlVR), and brain-to-BVR (BBVR)] and MWF for all regions (coefficient of determination, R2). A t-test with a linear mixed model compared the MWF of non-IUGR and IUGR fetal GPs for all three regions ( = 0.05). The MWF values are as follows: (mean {+/-} standard deviation): 0.23 {+/-} 0.02 (fetal CC), 0.19 {+/-} 0.02 (fetal CC - IUGR), 0.31 {+/-} 0.02 (fetal FOR), 0.27 {+/-} 0.01 (fetal FOR - IUGR), 0.28 {+/-} 0.02 (fetal PSW), and 0.24 {+/-} 0.03 (fetal PSW - IUGR). Significant differences in MWF were found between the non-IUGR and IUGR fetuses in every region. In conclusion, the mean MWF of IUGR fetal GPs is significantly lower than non-IUGR fetal GPs.

BackgroundSex differences in vulnerability to and severity of SARS-CoV-2 infection have been described in non-pregnant populations. ACE2 and TMPRSS2, host molecules required for viral entry, are regulated by sex steroids and expressed in the placenta. We sought to investigate whether placental ACE2 and TMPRSS2 expression vary by fetal sex and in the presence of maternal SARS-CoV-2 infection. MethodsPlacental ACE2 and TMPRSS2 were quantified in 68 pregnant individuals (38 SARS-CoV-2 positive, 30 SARS-CoV-2 negative) delivering at Mass General Brigham from April to June 2020. Maternal SARS-CoV-2 status was determined by nasopharyngeal RT-PCR. Placental SARS-CoV-2 viral load was quantified. RTqPCR was performed to quantify expression of ACE2 and TMPRSS2 relative to the reference gene YWHAZ. Western blots were performed on placental homogenates to quantify protein levels. The impact of fetal sex and SARS-CoV-2 exposure on ACE2 and TMPRSS2 expression was analyzed by 2-way ANOVA. ResultsSARS-CoV-2 virus was undetectable in all placentas. Maternal SARS-CoV-2 infection impacted TMPRSS2 placental gene and protein expression in a sexually dimorphic fashion (2-way ANOVA interaction p-value: 0.002). We observed no impact of fetal sex or maternal SARS-CoV-2 status on placental ACE2 gene or protein expression. Placental TMPRSS2 expression was significantly correlated with ACE2 expression in males (Spearmans {rho}=0.54, p=0.02) but not females ({rho}=0.23, p=0.34) exposed to maternal SARS-CoV-2. ConclusionsSex differences in placental TMPRSS2 but not ACE2 were observed in the setting of maternal SARS-CoV-2 infection. These findings may have implications for offspring vulnerability to placental infection and vertical transmission.These findings may have implications for offspring vulnerability to placental infection and vertical transmission.

SRC knockdown inhibits trophoblast cell proliferation, migration, and invasion while inducing apoptosis via activation of the PI3K/Akt/Bcl-2 signaling pathway. Trophoblast dysfunction is central to pregnancy disorders such as preeclampsia and miscarriage, yet the role of SRC, a non-receptor tyrosine kinase, in these cells remains poorly understood. This study aimed to elucidate the functional impact of SRC on trophoblast behavior and its underlying mechanism. Using siRNA-mediated knockdown in HTR8/SVneo cells, we confirmed efficient reduction of SRC mRNA and protein expression via RT-qPCR and Western blot. Functional assays demonstrated that SRC silencing significantly suppressed cell proliferation (CCK-8), migration (wound healing), and invasion (Transwell), while promoting apoptosis, evidenced by increased Annexin V-FITC/PI staining and upregulated Caspase-3 and Caspase-9 protein levels. Mechanistically, Western blot analysis revealed that SRC knockdown upregulated PI3K, Akt1, and Bcl-2 protein expression without altering IRS1 levels, indicating activation of the PI3K/Akt/Bcl-2 pro-survival pathway. This paradoxical activation appears to be a compensatory feedback insufficient to overcome SRC loss-induced dysfunction. Our findings identify SRC as a critical positive regulator of trophoblast proliferation, motility, and survival, acting through a non-canonical, IRS1-independent negative regulation of PI3K/Akt signaling. This study provides novel insights into trophoblast biology and suggests SRC as a potential therapeutic target for pregnancy complications; future in vivo studies are warranted to validate these mechanisms.

ObjectiveTo assess the vertebral pattern in a cohort of deceased fetuses and neonates, and to study the possible impact of DNA Copy Number Variations (CNVs) in coding regions and/or disturbing enhancers on the development of the vertebral pattern.\n\nMethodRadiographs of 445 fetuses and infants, deceased between 2009 and 2015, were assessed. Terminations of pregnancies, stillbirths and neonatal deaths were included. Patients were excluded if the vertebral pattern could not be determined. Copy number profiles of 265 patients were determined using single nucleotide polymorphism array.\n\nResults274/374 patients (73.3%) had an abnormal vertebral pattern. Cervical ribs were present in 188/374 (50.3%) and were significantly more common in stillbirths (69/128 (53.9%)) and terminations of pregnancies (101/188 (53.7%)), compared to live births (18/58, 31.0%, p = 0.006). None of the rare CNVs were recurrent or overlapped candidate genes for vertebral patterning.\n\nConclusionThe presence of an abnormal vertebral pattern, particularly in the cervical region, could be a sign of disruption at critical, highly interactive and conserved stages of embryogenesis. The vertebral pattern might provide valuable information regarding fetal and neonatal outcome. CNV analyses did not identify a mutual genetic cause for the occurrence of vertebral patterning abnormalities, indicating genetic heterogeneity.

Roughly 10% of all pregnancies are affected by fetal growth restriction (FGR). The primary etiology of FGR is placental insufficiency: the placenta not providing the appropriate amount of nutrients and oxygen to the fetus. There is currently no treatment for FGR or placental insufficiency. Because of the placentas pivotal role in FGR and supplying nutrients to the fetus, it offers an excellent target for therapeutic intervention. Using a guinea pig maternal nutrient restriction model and a repeated placental nanoparticle-mediated IGF1 treatment, placental IGF1 signaling and nutrient transport pathways were characterized to understand changes with FGR and treatment. This study elucidates the signaling mechanisms in which repeated placental nanoparticle-mediated IGF1 treatment leads to correct fetal growth. Overall, this study resulted in sex-specific kinase signaling and nutrient transporter changes within the placenta in both FGR and treatment groups. Combined with our previous studies using this treatment, we demonstrate the basic molecular signaling of this treatment and recapitulate the plausibility of this therapy for future human translation.

Early onset preeclampsia is a placental disorder characterized by shallow implantation, whereas placenta accreta spectrum is a placental disorder of deep placental attachment. This study compares the transcriptome of these two obstetric syndromes. By integrating available microarray and single-cell placenta/decidua transcriptomic datasets, we demonstrated that early onset preeclampsia genes are inversely expressed in placenta accreta, with the most marked differences noted in cell types of decidua, endothelial, and extravillous trophoblasts. Our findings highlight the key functions of trophoblast cell migration and invasion, decidua cell signaling, hypoxia pathways, and global growth factor and collagen contributions to these pregnancy disorders. This research provides new insights into the mechanisms of placentation and unifies these clinical siloes of disease by focusing on the fundamental biology of placental development at the maternal-fetal interface.

BackgroundPreeclampsia (PE) complicates 2-8% of pregnancies and is marked by placental hypoxia and HIF-pathway activation, especially in early-onset PE (eoPE). Integrating patient tissue analyses with experimental models may reveal common molecular markers of trophoblast hypoxic response. MethodsWe analyzed scRNA-seq data from 10 eoPE, 7 late-onset PE (loPE), and corresponding control placentas, identifying villous cytotrophoblast (VCT), syncytiotrophoblast (SCT), and extravillous trophoblast (EVT) subpopulations. BeWo b30 cells were treated for 24 h with CoCl2 (300 {micro}M) or an oxyquinoline derivative (OD, 5 {micro}M) to induce hypoxia. RNA and small RNA sequencing quantified mRNA and microRNA changes. PROGENy inferred pathway activities. ResultsSingle-cell analysis revealed highest hypoxia pathway activation in eoPE, with EVT showing maximum activity among trophoblast populations. Nine genes were upregulated across all trophoblast types in eoPE: EBI3, CST6, FN1, RFK, COL17A1, LDHA, PKP2, RPS4Y1, and RPS26. In vitro, OD induced more specific hypoxia responses than CoCl2, with 1,284 versus 3,032 differentially expressed genes respectively. Critically, EBI3, FN1, and COL17A1 showed concordant upregulation in both placental tissue and OD-treated cells, while CoCl2 treatment produced opposite expression patterns. MicroRNA analysis identified hsa-miR-27a-5p and hsa-miR-193b-5p as consistently elevated in both experimental conditions and previously reported in PE placental vesicles. We also identified isoforms of hsa-miR-9-5p and hsa-miR-92b-3p as hypoxia-associated in trophoblast. ConclusionsEBI3, COL17A1, hsa-miR-27a-5p, and hsa-miR-193b-5p emerge as trophoblast hypoxia markers in PE. Oxyquinoline derivatives offer a more physiologically relevant in vitro hypoxia model than CoCl2. This integrated approach advances understanding of PE pathophysiology and suggests candidate therapeutic targets.

ObjectiveTo investigate novel biomarkers and the mechanism of obstetric antiphospholipid syndrome (OAPS). MethodsHTR8/SVneo cells line were treated with plasma from OAPS (OAPS group) and healthy (NC group) pregnant women, respectively. The changes induced by plasma treatment at the transcriptome level were examined by RNA sequencing of the cells. Results were analyzed with bioinformatics tools to elucidate the potential biomarkers. Reverse-transcription quantitative polymerase chain reaction (RT-qPCR), western blotting, hematoxylin and eosin (HE), immunohistochemistry (IHC), and immunofluorescence (IF) were used for subsequent validation. ResultsBioinformatic analysis revealed the expression of Fibronectin 1 (FN1) was significantly increased in OAPS group. On analyzing molecular function, OAPS plasma exposure mainly affected the expression of the genes related to extracellular matrix (ECM) structural constituent. Compared to the NC group, differently expressed genes were mainly annotated to the collagen-containing ECM matrix and the ECM organization. In OAPS group, the protein expression of FN1 was also increased in blood (p <0 .05). The mRNA and protein expression of FN1 in placenta tissue were increased (p <0 .05) in OAPS group. Massive degeneration and atrophy can be seen in placental villi, with a significant reduction or disappearance of syncytiotrophoblasts and excessive fibrinoid deposition in the villous stroma of OAPS placenta. Both IHC and IF results showed the staining area and intensity of FN1 in the placental villi and stroma were significantly higher in OAPS group. ConclusionsFN1 may play a potential role in the pathogenic mechanisms of OAPS. Highlights

IntroductionMatrix metalloproteinases (MMPs) specifically MMP-9 is a key regulator of vascular and uterine spiral artery remodelling and its activity is controlled at multiple levels, including gene transcription, activation of its latent forms and endogenous inhibition by tissue inhibitors of metalloproteinases [TIMPs (specifically TIMP-1)]. Alteration in MMP-9 and TIMP-1 expression may contribute to uterine and vascular dysfunction leading to adverse pregnancy outcomes such as Preeclampsia (PE). Depending on time, PE is classified as early-onset preeclampsia, which is diagnosed before 34 weeks of gestation, or late-onset preeclampsia, which is diagnosed after 34 weeks. Early-onset preeclampsia (EOPE) is a severe obstetrics disease which threatens mother and foetus. Therefore, in the present study, we aimed to investigate the levels of MMP-9 and its inhibitor TIMP-1 in the placentae of EOPE patients and their maternal age matched normotensive, non-proteinuric controls at both mRNA and protein levels. MethodsA total of 30 caesarean delivered placentae (15 EOPE patients and 15 controls) were collected from Department of Obstetrics and Gynaecology, AIIMS, New Delhi after taking permission from Institute Ethics Committee. MMP-9 and TIMP-1 protein expression was observed by immunohistochemistry and immunofluorescence stainings. Gelatin Gel Zymography was done to evaluate MMP-9 activity and Western Blot was done for the quantification of MMP-9 and TIMP-1 proteins. mRNA levels of MMP-9 and TIMP-1 were determined by qRT-PCR ResultsImmunohistochemistry and Immunofluorescence staining discerned stronger expression of MMP-9 in normotensive placentae as compared to EOPE placentae whereas stronger expression of TIMP-1 was seen in EOPE placentae in comparison to those of normotensive placentae. Gelatin Gel Zymography reflected that MMP-9 activity was found elevated in maternal placental side in normotensive placentae as compared to EOPE placentae. Western Blot analysis revealed that protein expression of MMP-9 was elevated whereas for that of TIMP-1 was reduced in normotensive placentae in comparison to EOPE placentae. mRNA expression of MMP-9 was found up-regulated whereas for that of TIMP-1 was down-regulated in normotensive placentae as compared to EOPE placentae. ConclusionThe present study implies that aberrant functioning of MMP-9 and TIMP-1 in EOPE patients contribute to impaired placentation which might be relevant for possible future screening programs in order to predict and to design therapies for early onset preeclamptic patients.

Fetal growth depends on placental function, which requires energy from mitochondria. Here we investigated whether mitochondrial function in the placenta relates to growth of the lightest and heaviest fetuses of each sex within the litter of mice. Placentas from the lightest and heaviest fetuses were taken to evaluate placenta morphology (stereology), mitochondrial energetics (high-resolution respirometry), and mitochondrial regulators, nutrient transporters, hormone handling and signalling pathways (qPCR and western blotting). We found that mitochondrial complex I and II oxygen consumption rate was greater for placentas supporting the lightest female fetuses, although placental complex I abundance of the lightest females and complexes III and V of the lightest males were decreased compared to their heaviest counterparts. Expression of mitochondrial biogenesis (Nrf1) and fission (Drp1 and Fis1) genes was lower in the placenta from the lightest females, whilst biogenesis-related gene Tfam was greater in the placenta of the lightest male fetuses. Additionally, placental morphology and steroidogenic gene (Cyp17a1 and Cyp11a1) expression was aberrant for the lightest females, but glucose transporter (Glut1) expression was lower in only the lightest males versus their heaviest counterparts. Differences in intra-litter placental phenotype were related to sex-dependent changes in the expression of hormone responsive (androgen receptor) and metabolic signalling (AMPK, AKT, PPAR{gamma}) pathways. Thus, in normal mouse pregnancy, placental structure, function and mitochondrial phenotype are differentially responsive to growth of the female and the male fetus. This study may inform the design of sex- specific therapies for placental insufficiency and fetal growth abnormalities with life-long benefits for the offspring.

BackgroundThere is a growing body of evidence indicating the importance of endogenous retrovirus (ERV) derived proteins during early development and reproduction in mammals. Recently, a protein derived from the youngest ERV in humans, ERVK (HML2), was shown to be expressed during human placentation. Since a number of highly similar ERVK proviral loci exist across the human genome, locus-specific analysis of ERVK transcription and identification of the coding sequence expressed in the human placenta is difficult. Thus, despite its activity in early human development, the native expression and function of ERVK in the human placenta remains largely uncharacterized. ResultsIn this study, we comprehensively examined locus-specific ERVK transcription across several human placental tissues and cell types. Through a combination of RNA-seq and siRNA knock-down analyses, we identified the expression of a single ERVK locus, ERVK11q23.3, as (1) being significantly upregulated in preterm compared to term placenta, (2) predominantly expressed by mononuclear trophoblasts, (3) capable of encoding a truncated viral-like envelope protein, and (4) contributing to the expression cytokines involved in both antiviral and anti-inflammatory innate immune responses in human placental trophoblasts and BeWo choriocarcinoma cells, respectively. ConclusionsCollectively, the results of this study highlight the utility of studying locus-specific ERVK expression, provide a thorough characterization of locus-specific ERVK transcription from human placental tissues, and indicate that altered expression of placental ERVK11q23.3 influences interferon antiviral response, which may contribute to preterm birth and other pregnancy complications.

This study focuses on developing new approaches to identify and quantify glycogen trophoblast cells in the placenta. Due to limitations with currently available methods, alternative markers for these cells are being explored. Here we report on one promising candidate, Aldh1a3, a gene that was recently shown to be expressed in the mouse placenta only in glycogen trophoblast cells and their progenitors. This study validates ALDH1A3 protein as glycogen trophoblast marker with high specificity when compared to CDKN1C. This marker will be useful for the isolation of the glycogen trophoblast subpopulation in order to examine its function in placental programming, and for understanding its role in pathological pregnancy models.

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 2nd trimester ("2ndTri COVID", n=8) and 3rd trimester ("3rdTri COVID", n=8). In 3rdTri COVID as compared to control and 2ndTri-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 3rdTri COVID infections as compared to control and 2ndTri-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.

IntroductionStereological estimates of villous membrane thickness and surface area are widely used to infer the diffusive exchange capacity of the human placenta. A key geometric determinant of exchange capacity can be expressed as an effective diffusive length scale. Here we combine virtual histological sections with computational modelling in realistic villous geometries to assess the accuracy of classical stereological estimates of this diffusive length scale. MethodsTwo terminal villi, reconstructed from three-dimensional imaging, were digitally sectioned to generate random two-dimensional geometries containing fetal capillaries and surrounding villous tissue. For each section, we simulated steady diffusive transport between the fetal capillary and intervillous space boundaries to obtain a physics-based diffusive length scale as a reference case. Using the same geometries, we applied standard line-intercept stereology to measure harmonic-mean barrier thickness and boundary-length densities, from which a stereological estimate of diffusive length scale was derived. ResultsAcross both villi, stereology systematically overestimated the diffusive length scale by approximately 15-25%, depending on villus and section. We identified sources of this discrepancy, including interface curvature and assumptions underpinning the stereological correction factors, using idealised models of villus structure. ConclusionThese findings highlight the need for stereological approaches that account for curvature when interpreting placental structure-function relationships.