Maternal Smoking in the First Trimester and its Consequence on the Early Placenta.

Smoking during pregnancy increases the risk of adverse pregnancy outcomes, such as stillbirth and fetal growth restriction. This suggests impaired placental function and restricted nutrient and oxygen supply. Studies investigating placental tissue at the end of pregnancy have revealed increased DNA damage as a potential underlying cause, which is driven by various toxic smoke ingredients and oxidative stress induced by reactive oxygen species (ROS). However, in the first trimester, the placenta develops and differentiates, and many pregnancy pathologies associated with reduced placental function originate here. Therefore, we determined DNA damage in a cohort of first-trimester placental samples of verified smokers and nonsmokers. In fact, we observed an 80% increase in DNA breaks (P < .001) and shortened telomeres by 5.8% (P = .04) in placentas exposed to maternal smoking. Surprisingly, there was a decrease in ROS-mediated DNA damage, ie, 8-oxo-guanidine modifications, in placentas of the smoking group (-41%; P = .021), which paralleled the reduced expression of base excision DNA repair machinery, which restores oxidative DNA damage. Moreover, we observed that the increase in placental oxidant defense machinery expression, which usually occurs at the end of the first trimester in a healthy pregnancy as a result of the full onset of uteroplacental blood flow, was absent in the smoking group. Therefore, in early pregnancy, maternal smoking causes placental DNA damage, contributing to placental malfunction and increased risk of stillbirth and fetal growth restriction in pregnant women. Additionally, reduced ROS-mediated DNA damage along with no increase in antioxidant enzymes suggests a delay in the establishment of physiological uteroplacental blood flow at the end of the first trimester, which may further add to a disturbed placental development and function as a result of smoking in pregnancy.

Platelet-derived factors dysregulate placental sphingosine-1-phosphate receptor 2 in human trophoblasts.

RESEARCH QUESTION: Sphingosine-1-phosphate (S1P) is an essential and bioactive sphingolipid with various functions, which acts through five different G-protein-coupled receptors (S1PR1-5). What is the localization of S1PR1-S1PR3 in the human placenta and what is the effect of different flow rates, various oxygen concentrations and platelet-derived factors on the expression profile of S1PR in trophoblasts? DESIGN: Expression dynamics of placental S1PR1-S1PR3 were determined in human first trimester (n = 10), pre-term (n = 9) and term (n = 10) cases. Furthermore, the study investigated the expression of these receptors in different primary cell types isolated from human placenta, verified the findings with publicly available single-cell RNA-Seq data from first trimester and immunostaining of human first trimester and term placentas. The study also tested whether the placental S1PR subtypes are dysregulated in differentiated BeWo cells under different flow rates, different oxygen concentrations or in the presence of platelet-derived factors. RESULTS: Quantitative polymerase chain reaction revealed that S1PR2 is the predominant placental S1PR in the first trimester and reduces towards term (P < 0.0001). S1PR1 and S1PR3 increased from first trimester towards term (P < 0.0001). S1PR1 was localized in endothelial cells, whereas S1PR2 and S1PR3 were predominantly found in villous trophoblasts. Furthermore, S1PR2 was found to be significantly down-regulated in BeWo cells when co-incubated with platelet-derived factors (P = 0.0055). CONCLUSION: This study suggests that the placental S1PR repertoire is differentially expressed across gestation. S1PR2 expression in villous trophoblasts is negatively influenced by platelet-derived factors, which could contribute to down-regulation of placental S1PR2 over time of gestation as platelet presence and activation in the intervillous space increases from the middle of the first trimester onwards.

Maternal COVID-19 causing intrauterine foetal demise with microthrombotic placental insufficiency: a case report.

BACKGROUND: Pregnant women have an increased risk of getting infected with SARS-CoV-2 and are more prone to severe illness. Data on foetal demise in affected pregnancies and its underlying aetiology is scarce and pathomechanisms remain largely unclear. CASE: Herein we present the case of a pregnant woman with COVID-19 and intrauterine foetal demise. She had no previous obstetric or gynaecological history, and presented with mild symptoms at 34 + 3 weeks and no signs of foetal distress. At 35 + 6 weeks intrauterine foetal death was diagnosed. In the placental histopathology evaluation, we found inter- and perivillous fibrin depositions including viral particles in areas of degraded placental anatomy without presence of viral entry receptors and SARS-CoV-2 infection of the placenta. CONCLUSION: This case demonstrates that maternal SARS-CoV-2 infection in the third trimester may lead to an unfavourable outcome for the foetus due to placental fibrin deposition in maternal COVID-19 disease possibly via a thrombogenic microenvironment, even when the foetus itself is not infected.

Early human trophoblast development: from morphology to function.

Human pregnancy depends on the proper development of the embryo prior to implantation and the implantation of the embryo into the uterine wall. During the pre-implantation phase, formation of the morula is followed by internalization of blastomeres that differentiate into the pluripotent inner cell mass lineage, while the cells on the surface undergo polarization and differentiate into the trophectoderm of the blastocyst. The trophectoderm mediates apposition and adhesion of the blastocyst to the uterine epithelium. These processes lead to a stable contact between embryonic and maternal tissues, resulting in the formation of a new organ, the placenta. During implantation, the trophectoderm cells start to differentiate and form the basis for multiple specialized trophoblast subpopulations, all of which fulfilling specific key functions in placentation. They either differentiate into polar cells serving typical epithelial functions, or into apolar invasive cells that adapt the uterine wall to progressing pregnancy. The composition of these trophoblast subpopulations is crucial for human placenta development and alterations are suggested to result in placenta-associated pregnancy pathologies. This review article focuses on what is known about very early processes in human reproduction and emphasizes on morphological and functional aspects of early trophoblast differentiation and subpopulations.

Candesartan Does Not Activate PPARγ and Its Target Genes in Early Gestation Trophoblasts.

Angiotensin II receptor 1 blockers are commonly used to treat hypertension in women of childbearing age. While the fetotoxic effects of these drugs in the second and third trimesters of pregnancy are well documented, their possible impacts on placenta development in early gestation are unknown. Candesartan, a member of this group, also acts as a peroxisome proliferator-activated receptor gamma (PPARγ) agonist, a key regulator shown to be important for placental development. We have previously shown that trophoblasts do not express the candesartan target-receptor angiotensin II type 1 receptor AGTR1. This study investigated the possible role of candesartan on trophoblastic PPARγ and its hallmark target genes in early gestation. Candesartan did not affect the PPARγ protein expression or nuclear translocation of PPARγ. To mimic extravillous trophoblasts (EVTs) and cytotrophoblast/syncytiotrophoblast (CTB/SCT) responses to candesartan, we used trophoblast cell models BeWo (for CTB/SCT) and SGHPL-4 (EVT) cells as well as placental explants. In vitro, the RT-qPCR analysis showed no effect of candesartan treatment on PPARγ target genes in BeWo or SGHPL-4 cells. Treatment with positive control rosiglitazone, another PPARγ agonist, led to decreased expressions of LEP and PPARG1 in BeWo cells and an increased expression of PPARG1 in SGHPL-4 cells. Our previous data showed early gestation-placental AGTR1 expression in fetal myofibroblasts only. In a CAM assay, AGTR1 was stimulated with angiotensin II and showed increased on-plant vessel outgrowth. These results suggest candesartan does not negatively affect PPARγ or its target genes in human trophoblasts. More likely, candesartan from maternal serum may first act on fetal-placental AGTR1 and influence angiogenesis in the placenta, warranting further research.

Flow-through isolation of human first trimester umbilical cord endothelial cells.

Human umbilical vein and artery endothelial cells (HUVEC; HUAEC), placental endothelial cells (fpAEC), and endothelial colony-forming cells (ECFC) from cord blood are a widely used model for researching placental vascular development, fetal and placental endothelial function, and the effect of adverse conditions in pregnancy thereon. However, placental vascular development and angiogenesis start in the first weeks of gestation, and adverse conditions in pregnancy may also affect endothelial function before term, suggesting that endothelial cells from early pregnancy may respond differently. Thus, we established a novel, gentle flow-through method to isolate pure human umbilical endothelial cells from first trimester (FTUEC). FTUEC were characterized and their phenotype was compared to the umbilical endothelium in situ as well as to other fetal endothelial cell models from term of gestation, i.e. HUVEC, fpAEC, ECFC. FTUEC possess a CD34-positive, juvenile endothelial phenotype, and can be expanded and passaged. We regard FTUEC as a valuable tool to study developmental processes as well as the effect of adverse insults in pregnancy in vitro.

Simple method of thawing cryo-stored samples preserves ultrastructural features in electron microscopy.

Preservation of ultrastructural features in biological samples for electron microscopy (EM) is a challenging task that is routinely accomplished through chemical fixation or high-pressure freezing coupled to automated freeze substitution (AFS) using specialized devices. However, samples from clinical (e.g. "biobanking" of bulk biopsies) and preclinical (e.g. whole mouse tissues) specimens are often not specifically prepared for ultrastructural analyses but simply immersed in liquid nitrogen before long-term cryo-storage. We demonstrate that ultrastructural features of such samples are insufficiently conserved using AFS and developed a simple, rapid, and effective method for thawing that does not require specific instrumentation. This procedure consists of dry ice-cooled pre-trimming of frozen tissue and aldehyde fixation for 3 h at 37 °C followed by standard embedding steps. Herein investigated tissues comprised human term placentae, clinical lung samples, as well as mouse tissues of different composition (brown adipose tissue, white adipose tissue, cardiac muscle, skeletal muscle, liver). For all these tissues, we compared electron micrographs prepared from cryo-stored material with our method to images derived from directly prepared fresh tissues with standard chemical fixation. Our protocol yielded highly conserved ultrastructural features and tissue-specific details, largely matching the quality of fresh tissue samples. Furthermore, morphometric analysis of lipid droplets and mitochondria in livers of fasted mice demonstrated that statistically valid quantifications can be derived from samples prepared with our method. Overall, we provide a simple and effective protocol for accurate ultrastructural and morphometric analyses of cryo-stored bulk tissue samples.

Automated Quantitative Image Evaluation of Antigen Retrieval Methods for 17 Antibodies in Placentation and Implantation Diagnostic and Research.

Immunostaining in clinical routine and research highly depends on standardized staining methods and quantitative image analyses. We qualitatively and quantitatively compared antigen retrieval methods (no pretreatment, pretreatment with pepsin, and heat-induced pretreatment with pH 6 or pH 9) for 17 antibodies relevant for placenta and implantation diagnostics and research. Using our newly established, comprehensive automated quantitative image analysis approach, fluorescent signal intensities were evaluated. Automated quantitative image analysis found that 9 out of 17 antibodies needed antigen retrieval to show positive staining. Heat induction proved to be the most efficient form of antigen retrieval. Eight markers stained positive after pepsin digestion, with ß-hCG and vWF showing enhanced staining intensities. To avoid the misinterpretation of quantitative image data, the qualitative aspect should always be considered. Results from native placental tissue were compared with sections of a placental invasion model based on thermo-sensitive scaffolds. Immunostaining on placentas in vitro leads to new insights into fetal development and maternal pathophysiological pathways, as pregnant women are justifiably excluded from clinical studies. Thus, there is a clear need for the assessment of reliable immunofluorescent staining and pretreatment methods. Our evaluation offers a powerful tool for antibody and pretreatment selection in placental research providing objective and precise results.

Changes in Maternal Platelet Physiology during Gestation and Their Interaction with Trophoblasts.

Upon activation, maternal platelets provide a source of proinflammatory mediators in the intervillous space of the placenta. Therefore, platelet-derived factors may interfere with different trophoblast subtypes of the developing human placenta and might cause altered hormone secretion and placental dysfunction later on in pregnancy. Increased platelet activation, and the subsequent occurrence of placental fibrinoid deposition, are linked to placenta pathologies such as preeclampsia. The composition and release of platelet-derived factors change over gestation and provide a potential source of predicting biomarkers for the developing fetus and the mother. This review indicates possible mechanisms of platelet-trophoblast interactions and discusses the effect of increased platelet activation on placenta development.

Placental Endocrine Activity: Adaptation and Disruption of Maternal Glucose Metabolism in Pregnancy and the Influence of Fetal Sex.

The placenta is an endocrine fetal organ, which secretes a plethora of steroid- and proteo-hormones, metabolic proteins, growth factors, and cytokines in order to adapt maternal physiology to pregnancy. Central to the growth of the fetus is the supply with nutrients, foremost with glucose. Therefore, during pregnancy, maternal insulin resistance arises, which elevates maternal blood glucose levels, and consequently ensures an adequate glucose supply for the developing fetus. At the same time, maternal ß-cell mass and function increase to compensate for the higher insulin demand. These adaptations are also regulated by the endocrine function of the placenta. Excessive insulin resistance or the inability to increase insulin production accordingly disrupts physiological modulation of pregnancy mediated glucose metabolism and may cause maternal gestational diabetes (GDM). A growing body of evidence suggests that this adaptation of maternal glucose metabolism differs between pregnancies carrying a girl vs. pregnancies carrying a boy. Moreover, the risk of developing GDM differs depending on the sex of the fetus. Sex differences in placenta derived hormones and bioactive proteins, which adapt and modulate maternal glucose metabolism, are likely to contribute to this sexual dimorphism. This review provides an overview on the adaptation and maladaptation of maternal glucose metabolism by placenta-derived factors, and highlights sex differences in this regulatory network.

Defective Lysosomal Lipolysis Causes Prenatal Lipid Accumulation and Exacerbates Immediately after Birth.

Cholesterol and fatty acids are essential lipids that are critical for membrane biosynthesis and fetal organ development. Cholesteryl esters (CE) are degraded by hormone-sensitive lipase (HSL) in the cytosol and by lysosomal acid lipase (LAL) in the lysosome. Impaired LAL or HSL activity causes rare pathologies in humans, with HSL deficiency presenting less severe clinical manifestations. The infantile form of LAL deficiency, a lysosomal lipid storage disorder, leads to premature death. However, the importance of defective lysosomal CE degradation and its consequences during early life are incompletely understood. We therefore investigated how defective CE catabolism affects fetus and infant maturation using Lal and Hsl knockout (-/-) mouse models. This study demonstrates that defective lysosomal but not neutral lipolysis alters placental and fetal cholesterol homeostasis and exhibits an initial disease pathology already in utero as Lal-/- fetuses accumulate hepatic lysosomal lipids. Immediately after birth, LAL deficiency exacerbates with massive hepatic lysosomal lipid accumulation, which continues to worsen into young adulthood. Our data highlight the crucial role of LAL during early development, with the first weeks after birth being critical for aggravating LAL deficiency.

Kidney Injury Caused by Preeclamptic Pregnancy Recovers Postpartum in a Transgenic Rat Model.

Preeclampsia (PE) is characterized by the onset of hypertension (≥140/90 mmHg) and presence of proteinuria (>300 mg/L/24 h urine) or other maternal organ dysfunctions. During human PE, renal injuries have been observed. Some studies suggest that women with PE diagnosis have an increased risk to develop renal diseases later in life. However, in human studies PE as a single cause of this development cannot be investigated. Here, we aimed to investigate the effect of PE on postpartum renal damage in an established transgenic PE rat model. Female rats harboring the human-angiotensinogen gene develop a preeclamptic phenotype after mating with male rats harboring the human-renin gene, but are normotensive before and after pregnancy. During pregnancy PE rats developed mild tubular and glomerular changes assessed by histologic analysis, increased gene expression of renal damage markers such as kidney injury marker 1 and connective-tissue growth factor, and albuminuria compared to female wild-type rats (WT). However, four weeks postpartum, most PE-related renal pathologies were absent, including albuminuria and elevated biomarker expression. Only mild enlargement of the glomerular tuft could be detected. Overall, the glomerular and tubular function were affected during pregnancy in the transgenic PE rat. However, almost all these pathologies observed during PE recovered postpartum.

Type 1 Diabetes Mellitus and the First Trimester Placenta: Hyperglycemia-Induced Effects on Trophoblast Proliferation, Cell Cycle Regulators, and Invasion.

Type 1 diabetes mellitus (T1DM) is associated with reduced fetal growth in early pregnancy, but a contributing role of the placenta has remained elusive. Thus, we investigated whether T1DM alters placental development in the first trimester. Using a protein array, the level of 60 cell-cycle-related proteins was determined in human first trimester placental tissue (gestational week 5-11) from control (n = 11) and T1DM pregnancies (n = 12). Primary trophoblasts (gestational week 7-12, n = 32) were incubated in the absence (control) or presence of hyperglycemia (25 mM D-glucose) and hyperosmolarity (5.5 mM D-glucose + 19.5 mM D-mannitol). We quantified the number of viable and dead trophoblasts (CASY Counter) and assessed cell cycle distribution (FACS) and trophoblast invasion using a transwell assay. T1DM was associated with a significant (p < 0.05) downregulation of Ki67 (-26%), chk1 (-25%), and p73 (-26%). The number of viable trophoblasts was reduced under hyperglycemia (-23%) and hyperosmolarity (-18%), whereas trophoblast invasion was increased only under hyperglycemia (+6%). Trophoblast cell death and cell cycle distribution remained unaffected. Collectively, our data demonstrate that hyperglycemia decreases trophoblast proliferation as a potential contributing factor to the reduced placental growth in T1DM in vivo.

Go with the Flow-Trophoblasts in Flow Culture.

With establishment of uteroplacental blood flow, the perfused fetal chorionic tissue has to deal with fluid shear stress that is produced by hemodynamic forces across different trophoblast subtypes. Amongst many other cell types, trophoblasts are able to sense fluid shear stress through mechanotransduction. Failure in the adaption of trophoblasts to fluid shear stress is suggested to contribute to pregnancy disorders. Thus, in the past twenty years, a significant body of work has been devoted to human- and animal-derived trophoblast culture under microfluidic conditions, using a rather broad range of different fluid shear stress values as well as various different flow systems, ranging from commercially 2D to customized 3D flow culture systems. The great variations in the experimental setup reflect the general heterogeneity in blood flow through different segments of the uteroplacental circulation. While fluid shear stress is moderate in invaded uterine spiral arteries, it drastically declines after entrance of the maternal blood into the wide cavity of the intervillous space. Here, we provide an overview of the increasing body of evidence that substantiates an important influence of maternal blood flow on several aspects of trophoblast physiology, including cellular turnover and differentiation, trophoblast metabolism, as well as endocrine activity, and motility. Future trends in trophoblast flow culture will incorporate the physiological low oxygen conditions in human placental tissue and pulsatile blood flow in the experimental setup. Investigation of trophoblast mechanotransduction and development of mechanosome modulators will be another intriguing future direction.

Maternal Obesity Alters Placental Cell Cycle Regulators in the First Trimester of Human Pregnancy: New Insights for BRCA1.

In the first trimester of pregnancy, placental development involves a wide range of cellular processes. These include trophoblast proliferation, fusion, and differentiation, which are dependent on tight cell cycle control. The intrauterine environment affects placental development, which also includes the trophoblast cell cycle. In this work, we focus on maternal obesity to assess whether an altered intrauterine milieu modulates expression and protein levels of placental cell cycle regulators in early human pregnancy. For this purpose, we use first trimester placental tissue from lean and obese women (gestational week 5+0-11+6, n = 58). Using a PCR panel, a cell cycle protein array, and STRING database analysis, we identify a network of cell cycle regulators increased by maternal obesity in which breast cancer 1 (BRCA1) is a central player. Immunostaining localizes BRCA1 predominantly to the villous and the extravillous cytotrophoblast. Obesity-driven BRCA1 upregulation is not able to be explained by DNA methylation (EPIC array) or by short-term treatment of chorionic villous explants at 2.5% oxygen with tumor necrosis factor α (TNF-α) (50 mg/mL), leptin (100 mg/mL), interleukin 6 (IL-6) (100 mg/mL), or high glucose (25 nM). Oxygen tension rises during the first trimester, but this change in vitro has no effect on BRCA1 (2.5% and 6.5% O2). We conclude that maternal obesity affects placental cell cycle regulation and speculate this may alter placental development.

Downregulation of p53 drives autophagy during human trophoblast differentiation.

The placental barrier is crucial for the supply of nutrients and oxygen to the developing fetus and is maintained by differentiation and fusion of mononucleated cytotrophoblasts into the syncytiotrophoblast, a process only partially understood. Here transcriptome and pathway analyses during differentiation and fusion of cultured trophoblasts yielded p53 signaling as negative upstream regulator and indicated an upregulation of autophagy-related genes. We further showed p53 mRNA and protein levels decreased during trophoblast differentiation. Reciprocally, autophagic flux increased and cytoplasmic LC3B-GFP puncta became more abundant, indicating enhanced autophagic activity. In line, in human first trimester placenta p53 protein mainly localized to the cytotrophoblast, while autophagy marker LC3B as well as late autophagic compartments were predominantly detectable in the syncytiotrophoblast. Importantly, ectopic overexpression of p53 reduced levels of LC3B-II, supporting a negative regulatory role on autophagy in differentiating trophoblasts. This was also shown in primary trophoblasts and human first trimester placental explants, where pharmacological stabilization of p53 decreased LC3B-II levels. In summary our data suggest that differentiation-dependent downregulation of p53 is a prerequisite for activating autophagy in the syncytiotrophoblast.

Maternal Platelets­Friend or Foe of the Human Placenta?

Human pregnancy relies on hemochorial placentation, including implantation of the blastocyst and deep invasion of fetal trophoblast cells into maternal uterine blood vessels, enabling direct contact of maternal blood with placental villi. Hemochorial placentation requires fast and reliable hemostasis to guarantee survival of the mother, but also for the neonates. During human pregnancy, maternal platelet count decreases gradually from first, to second, and third trimester. In addition to hemodilution, accelerated platelet sequestration and consumption in the placental circulation may contribute to a decline of platelet count throughout gestation. Local stasis, turbulences, or damage of the syncytiotrophoblast layer can activate maternal platelets within the placental intervillous space and result in formation of fibrin-type fibrinoid. Perivillous fibrinoid is a regular constituent of the normal placenta which is considered to be an important regulator of intervillous hemodynamics, as well as having a role in shaping the developing villous trees. However, exaggerated activation of platelets at the maternal-fetal interface can provoke inflammasome activation in the placental trophoblast, and enhance formation of circulating platelet-monocyte aggregates, resulting in sterile inflammation of the placenta and a systemic inflammatory response in the mother. Hence, the degree of activation determines whether maternal platelets are a friend or foe of the human placenta. Exaggerated activation of maternal platelets can either directly cause or propagate the disease process in placenta-associated pregnancy pathologies, such as preeclampsia.

Placental CX3CL1 is Deregulated by Angiotensin II and Contributes to a Pro-Inflammatory Trophoblast-Monocyte Interaction.

CX3CL1, which is a chemokine involved in many aspects of human pregnancy, is a membrane-bound chemokine shed into circulation as a soluble isoform. Placental CX3CL1 is induced by inflammatory cytokines and is upregulated in severe early-onset preeclampsia. In this study, the hypothesis was addressed whether angiotensin II can deregulate placental CX3CL1 expression, and whether CX3CL1 can promote a pro-inflammatory status of monocytes. qPCR analysis of human placenta samples (n = 45) showed stable expression of CX3CL1 and the angiotensin II receptor AGTR1 throughout the first trimester, but did not show a correlation between both or any influence of maternal age, BMI, and gestational age. Angiotensin II incubation of placental explants transiently deregulated CX3CL1 expression, while the angiotensin II receptor antagonist candesartan reversed this effect. Overexpression of recombinant human CX3CL1 in SGHPL-4 trophoblasts increased adhesion of THP-1 monocytes and significantly increased IL8, CCL19, and CCL13 in co-cultures with human primary monocytes. Incubation of primary monocytes with CX3CL1 and subsequent global transcriptome analysis of CD16⁺ subsets revealed 81 upregulated genes, including clusterin, lipocalin-2, and the leptin receptor. Aldosterone synthase, osteopontin, and cortisone reductase were some of the 66 downregulated genes present. These data suggest that maternal angiotensin II levels influence placental CX3CL1 expression, which, in turn, can affect monocyte to trophoblast adhesion. Release of placental CX3CL1 could promote the pro-inflammatory status of the CD16⁺ subset of maternal monocytes.

IGF2 stimulates fetal growth in a sex- and organ-dependent manner.

BackgroundInsulin-like growth factor 2 (IGF2) is a key determinant of fetal growth, and the altered expression of IGF2 is implicated in fetal growth disorders and maternal metabolic derangements including gestational diabetes. Here we studied how increased levels of IGF2 in late pregnancy affect fetal growth.MethodsWe employed a rat model of repeated intrafetal IGF2 administration in late pregnancy, i.e., during GD19-GD21, and measured the consequences on fetal organ weight and expression of insulin/IGF-axis components.ResultsIGF2 treatment tended to increase fetal weight, but only weight increase of the fetal stomach reached significance (+33±9%; P<0.01). Sex-dependent data analysis revealed a sexual dimorphism of IGF2 action. In male fetuses, IGF2 administration significantly increased fetal weight (+13±3%; P<0.05) and weight of fetal stomach (+42±10%; P<0.01), intestine (+26±5%; P<0.05), liver (+13±4%; P<0.05), and pancreas (+25±8%; P<0.05). Weights of heart, lungs, and kidneys were unchanged. In female fetuses, IGF2 increased only stomach weight (+26±9%; P<0.05). Furthermore, gene expression of insulin/IGF axis in the heart, lungs, liver, and stomach was more sensitive toward IGF2 treatment in male than in female fetuses.ConclusionData suggest that elevated circulating IGF2 in late pregnancy predominantly stimulates organ growth of the digestive system, and male fetuses are more susceptible toward the IGF2 effects than female fetuses.

CD74-Downregulation of Placental Macrophage-Trophoblastic Interactions in Preeclampsia.

RATIONALE: We hypothesized that cluster of differentiation 74 (CD74) downregulation on placental macrophages, leading to altered macrophage-trophoblast interaction, is involved in preeclampsia. OBJECTIVE: Preeclamptic pregnancies feature hypertension, proteinuria, and placental anomalies. Feto-placental macrophages regulate villous trophoblast differentiation during placental development. Disturbance of this well-balanced regulation can lead to pathological pregnancies. METHODS AND RESULTS: We performed whole-genome expression analysis of placental tissue. CD74 was one of the most downregulated genes in placentas from preeclamptic women. By reverse transcriptase-polymerase chain reaction, we confirmed this finding in early-onset (<34 gestational week, n=26) and late-onset (≥34 gestational week, n=24) samples from preeclamptic women, compared with healthy pregnant controls (n=28). CD74 protein levels were analyzed by Western blot and flow cytometry. We identified placental macrophages to express CD74 by immunofluorescence, flow cytometry, and RT-PCR. CD74-positive macrophages were significantly reduced in preeclamptic placentas compared with controls. CD74-silenced macrophages showed that the adhesion molecules ALCAM, ICAM4, and Syndecan-2, as well as macrophage adhesion to trophoblasts were diminished. Naive and activated macrophages lacking CD74 showed a shift toward a proinflammatory signature with an increased secretion of tumor necrosis factor-α, chemokine (C-C motif) ligand 5, and monocyte chemotactic protein-1, when cocultured with trophoblasts compared with control macrophages. Trophoblasts stimulated by these factors express more CYP2J2, sFlt1, TNFα, and IL-8. CD74-knockout mice showed disturbed placental morphology, reduced junctional zone, smaller placentas, and impaired spiral artery remodeling with fetal growth restriction. CONCLUSIONS: CD74 downregulation in placental macrophages is present in preeclampsia. CD74 downregulation leads to altered macrophage activation toward a proinflammatory signature and a disturbed crosstalk with trophoblasts.