Normal Human Placenta Research Articles

Single-cell and spatial transcriptomics: Discovery of human placental development and disease.

The human placenta is a vital organ, encompassing many distinct cell types, that maintains the growth and development of the fetus and is essential for substance exchange, defense, synthesis, and immunity. Abnormalities in placental cells can lead to various pregnancy complications, but the mechanisms remain largely unclear. Single-cell and spatial transcriptomics technologies have been developed in recent years to demonstrate placental cell heterogeneity and spatial molecular localization. Here, we review and summarize the current literature, demonstrating these technologies and showing the heterogeneity of various placenta cells and cell-cell communication of normal human placenta, as well as placenta-related diseases, such as preeclampsia, gestational diabetes mellitus, advanced maternal age, recurrent pregnancy loss, and placenta accreta spectrum disorders. Meanwhile, current weaknesses and future direction were discussed.

Combining RNAscope, Immunohistochemistry (IHC) and Digital Image Analysis to Assess Podoplanin (PDPN) Protein and PDPN_mRNA Expression on Formalin-Fixed Paraffin-Embedded Normal Human Placenta Tissues.

The expression and function of podoplanin (PDPN) in the normal human placenta has been debated in placental evaluation. This study emphasizes the importance of a multimodal approach of PDPN expression in normal human placentas. A complete examination is performed using immunohistochemistry, RNAscope and automated Digital Image examination (DIA) interpretation. QuPath DIA-based analysis automatically generated the stromal and histological scores of PDPN expression for immunohistochemistry and RNAscope stains. The umbilical cord's isolated fibroblasts and luminal structures expressed PDPN protein and PDPN_mRNA. RNAscope detected PDPN_mRNA upregulation in syncytial placental knots trophoblastic cells, but immunohistochemistry did not certify this at the protein level. The study found a significant correlation between the IHC and RNAscope H-Score (p = 0.033) and Allred Score (p = 0.05). A successful multimodal strategy for PDPN assessment in human placentas confirmed PDPN expression heterogeneity in the full-term human normal placenta and umbilical cord at the protein and mRNA level. In placental syncytial knots trophoblastic cells, PDPN showed mRNA overexpression, suggesting a potential role in placenta maturation.

Downregulation of SPARC Expression Enhances the Fusion of BeWo Choriocarcinoma Cells.

Syncytiotrophoblasts, which are formed by the fusion of villous cytotrophoblasts, play an essential role in maintaining a successful pregnancy. Secreted protein acidic and rich in cysteine (SPARC) is a non-structural Ca2+-binding extracellular matrix glycoprotein involved in tissue remodeling and cell proliferation, differentiation, and migration. Previous studies have revealed that SPARC is expressed in villous and extravillous cytotrophoblasts in the first trimester and that RNA interference targeted at SPARC significantly inhibited invasion of human extravillous trophoblast HTR8/SVneo cells. However, the involvement of SPARC in cytotrophoblast fusion remains unknown. This study aimed to investigate the role of SPARC in cytotrophoblast fusion, using the BeWo choriocarcinoma cell line as a model of villous cytotrophoblasts. Immunohistochemical analysis was conducted to assess SPARC expression in normal human placentas using placental tissues obtained during the first and third trimesters of pregnancy. We investigated the effects of SPARC knockdown on trophoblast differentiation markers and cell fusion in BeWo cells using small interfering RNA. Immunohistochemical analysis revealed that SPARC expression was high in the early gestational chorionic villi and low in the late gestational chorionic villi. SPARC knockdown increased the expressions of human chorionic gonadotropin and Ovo-like transcriptional repressor 1; however, glial cells missing transcription factor 1, syncytin-1, and syncytin-2 showed no significant changes. The assessment revealed that SPARC knockdown significantly enhanced cell fusion compared to the non-silencing control. Our data suggest that SPARC plays a vital role in regulating trophoblast fusion and differentiation during placental development.

High-throughput mRNA sequencing of human placenta shows sex differences across gestation

IntroductionFetal sex affects fetal and maternal health outcomes in pregnancy, but this connection remains poorly understood. As the placenta is the route of fetomaternal communication and derives from the fetal genome, placental gene expression sex differences may explain these outcomes. ObjectivesWe utilized next generation sequencing to study the normal human placenta in both sexes in first and third trimester to generate a normative transcriptome based on sex and gestation. Study designWe analyzed 124 first trimester (T1, 59 female and 65 male) and 43 third trimester (T3, 18 female and 25 male) samples for sex differences within each trimester and sex-specific gestational differences. ResultsPlacenta shows more significant sexual dimorphism in T1, with 94 T1 and 26 T3 differentially expressed genes (DEGs). The sex chromosomes contributed 60.6% of DEGs in T1 and 80.8% of DEGs in T3, excluding X/Y pseudoautosomal regions. There were 6 DEGs from the pseudoautosomal regions, only significant in T1 and all upregulated in males. The distribution of DEGs on the X chromosome suggests genes on Xp (the short arm) may be particularly important in placental sex differences. Dosage compensation analysis of X/Y homolog genes shows expression is primarily contributed by the X chromosome. In sex-specific analyses of first versus third trimester, there were 2815 DEGs common to both sexes upregulated in T1, and 3263 common DEGs upregulated in T3. There were 7 female-exclusive DEGs upregulated in T1, 15 female-exclusive DEGs upregulated in T3, 10 male-exclusive DEGs upregulated in T1, and 20 male-exclusive DEGs upregulated in T3. DiscussionThis is the largest cohort of placentas across gestation from healthy pregnancies defining the normative sex dimorphic gene expression and sex common, sex specific and sex exclusive gene expression across gestation. The first trimester has the most sexually dimorphic transcripts, and the majority were upregulated in females compared to males in both trimesters. The short arm of the X chromosome and the pseudoautosomal region is particularly critical in defining sex differences in the first trimester placenta. As pregnancy is a dynamic state, sex specific DEGs across gestation may contribute to sex dimorphic changes in overall outcomes.

The number of trophoblast-derived particles in the intervillous space is higher in female than in male normal human placentas

The number of trophoblast-derived particles in the intervillous space is higher in female than in male normal human placentas

ODP434 Maternally-Derived Pancreatic Extracellular Vesicle Encompassed miRNAs Influence Placental Development in Pregnancies Complicated by Gestational Diabetes

Abstract Pregnancies affected by gestational diabetes (GDM) commonly result in large-for-gestational-age (LGA) infants; these infants have an increased risk of developing cardiometabolic complications compared to appropriate-for-gestational-age (AGA) infants. The mechanisms responsible are unclear but GDM/LGA is associated with altered placental development/function. We have recently reported altered levels of extracellular vesicle (EV) encapsulated miRNAs, including miR-375 (a pancreatic-specific miRNA) in maternal serum prior to the onset of LGA. We propose that the EVs containing miR-375 are produced by maternal pancreas, and that increased miR-375 in maternal circulation contributes to LGA by influencing placental development/function. Human placental tissue was collected from GDM pregnancies that delivered AGA (n=14) or LGA infants (n=10). Levels of mature miR-375 and primary transcript (pri-miR-375) were measured by QPCR. Mature miR-375 was increased in GDM-LGA (4.74-fold; p<0. 01) compared to GDM AGA placentae. Mature miR-375, but not pri-miR-375, was detected in human placenta suggesting that miR-375 is not produced in the placenta but instead is transported, potentially via EVs in maternal circulation. Intact human pancreatic islets (20,000 IEQ/patient) were obtained through the Integrated Islet Distribution Program (IIDP) from female donors of reproductive age. Islets were cultured in EV-depleted media containing 5.5mM (normoglycemia) or 7mM (mild hyperglycaemia) glucose for 3 days. Islet purity and viability were confirmed using dithizone incorporation and trypan blue exclusion, respectively. Media was collected at 24-hour intervals, pooled from all time points, concentrated and EVs were isolated by size exclusion chromatography. Confirmation of the presence of EVs was confirmed by electron microscopy (typical cup-shape morphology of EVs), nanoparticle tracking analysis (NTA; mean size of 83.5-163.2nm; concentration of 1.28×109-1.82×1010 particles/ml), Western blotting (EV-enriched proteins). QPCR confirmed that mature miR-375 was present in islet-derived EVs and revealed that levels were altered following culture in 7mM glucose compared to 5.5mM. Islet-derived EVs were labelled with maleimide-488 and uptake into explants of normal human term placenta was confirmed using fluorescent microscopy=3). miR-375 was overexpressed in normal human placental explants (by 30-fold, p<0. 05; n=6) using specific mimics (100nM for 72 hours) and TMT mass spectrometry was performed to assess the impact of miR-375 overexpression on the placental proteome. Functional enrichment analysis (FEA) was performed on differentially expressed proteins (DEP; P<0. 05; -0.41≥log2foldchange≥0.58) using over representation analysis and Ingenuity Pathway Analysis. This revealed that DEGs were significantly enriched in pathways associated with placental growth, mitochondrial function and glucose metabolism. Our data demonstrates that miR-375 is increased in the placenta in GDM/LGA and that this influences placental development/function. miR-375 does not appear to be produced in the placenta but instead, is likely released in EVs from maternal pancreas under conditions of hyperglycaemia and transported to the placenta, via maternal circulation. This provides insight into the potential mechanisms contributing to LGA in pregnancies complicated by GDM. Presentation: No date and time listed

Calnexin Is Involved in Forskolin-Induced Syncytialization in Cytotrophoblast Model BeWo Cells.

Calnexin (CNX), a membrane-bound molecular chaperone, is involved in protein folding and quality control of nascent glycoproteins in the endoplasmic reticulum. We previously suggested critical roles of calreticulin, a functional paralogue of CNX, in placentation, including invasion of extravillous trophoblasts and syncytialization of cytotrophoblasts. However, the roles of CNX in placentation are unclear. In human choriocarcinoma BeWo cells, which serve as an experimental model of syncytialization, CNX knockdown suppressed forskolin-induced cell fusion and β-human chorionic gonadotropin (β-hCG) induction. Cell-surface luteinizing hormone/chorionic gonadotropin receptor, a β-hCG receptor, was significantly down-regulated in CNX-knockdown cells, which suggested the presence of a dysfunctional autocrine loop of β-hCG up-regulation. In this study, we also found abundant CNX expression in normal human placentas. Collectively, our results revealed the critical role of CNX in the syncytialization-related signaling in a villous trophoblast model and suggest a link between CNX expression and placenta development.

Renalase and its receptor, PMCA4b, are expressed in the placenta throughout the human gestation

Placental function requires organized growth, transmission of nutrients, and an anti-inflammatory milieu between the maternal and fetal interface, but placental factors important for its function remain unclear. Renalase is a pro-survival, anti-inflammatory flavoprotein found to be critical in other tissues. We examined the potential role of renalase in placental development. PCR, bulk RNA sequencing, immunohistochemistry, and immunofluorescence for renalase and its binding partners, PMCA4b and PZP, were performed on human placental tissue from second-trimester and full-term placentas separated into decidua, placental villi and chorionic plates. Quantification of immunohistochemistry was used to localize renalase across time course from 17 weeks to term. Endogenous production of renalase was examined in placental tissue and organoids. Renalase and its receptor PMCA4b transcripts and proteins were present in all layers of the placenta. Estimated RNLS protein levels did not change with gestation in the decidual samples. However, placental villi contained more renalase immunoreactive cells in fetal than full-term placental samples. RNLS co-labeled with markers for Hofbauer cells and trophoblasts within the placental villi. Endogenous production of RNLS, PMCA4b, and PZP by trophoblasts was validated in placental organoids. Renalase is endogenously expressed throughout placental tissue and specifically within Hofbauer cells and trophoblasts, suggesting a potential role for renalase in placental development and function. Future studies should assess renalase’s role in normal and diseased human placenta.

Ex-Vivo MRI of the Normal Human Placenta: Structural-Functional Interplay and the Association With Birth Weight.

Advanced magnetic resonance imaging (MRI) methods are increasingly being used to assess the human placenta. Yet, the structure-function interplay in normal placentas and their associations with pregnancy risks are not fully understood. To characterize the normal human placental structure (volume and umbilical cord centricity index (CI)) and function (perfusion) ex-vivo using MRI, to assess their association with birth weight (BW), and identify imaging-markers for placentas at risk for dysfunction. Prospective. Twenty normal term ex-vivo placentas. 3 T/ T1 and T2 weighted (T1 W, T2 W) turbo spin-echo, three-dimensional susceptibility-weighted image, and time-resolved angiography with interleaved stochastic trajectories (TWIST), during passage of a contrast agent using MRI compatible perfusion system that mimics placental flow. Placental volume and CI were manually extracted from the T1 W images by a fetal-placental MRI scientist (D.L., 7 years of experience). Perfusion maps including bolus arrival-time and full-width at half maximum were calculated from the TWIST data. Mean values, entropy, and asymmetries were calculated from each perfusion map, relating to both the whole placenta and volumes of interest (VOIs) within the umbilical cord and its daughter blood vessels. Pearson correlations with correction for multiple comparisons using false discovery rate were performed between structural and functional parameters, and with BW, with P < 0.05 considered significant. All placentas were successfully perfused and scanned. Significant correlations were found between whole placenta and VOIs perfusion parameters (mean R=0.76 ± 0.06, range=0.67-0.89), which were also significantly correlated with CI (mean R=0.72 ± 0.05, range=0.65-0.79). BW was correlated with placental volume (R=0.62), but not with CI (P=0.40). BW was also correlated with local perfusion asymmetry (R=-0.71). Results demonstrate a gradient of placental function, associated with CI and suggest several ex-vivo imaging-markers that might indicate an increased risk for placental dysfunction. 1 TECHNICAL EFFICACY: Stage 1.

Proteome profiling of human placenta reveals developmental stage-dependent alterations in protein signature

IntroductionPlacenta is a complex organ that plays a significant role in the maintenance of pregnancy health. It is a dynamic organ that undergoes dramatic changes in growth and development at different stages of gestation. In the first-trimester, the conceptus develops in a low oxygen environment that favors organogenesis in the embryo and cell proliferation and angiogenesis in the placenta; later in pregnancy, higher oxygen concentration is required to support the rapid growth of the fetus. This oxygen transition, which appears unique to the human placenta, must be finely tuned through successive rounds of protein signature alterations. This study compares placental proteome in normal first-trimester (FT) and term human placentas (TP).MethodsNormal human first-trimester and term placental samples were collected and differentially expressed proteins were identified using two-dimensional liquid chromatography-tandem mass spectrometry.ResultsDespite the overall similarities, 120 proteins were differently expressed in first and term placentas. Out of these, 72 were up-regulated and 48 were down-regulated in the first when compared with the full term placentas. Twenty out of 120 differently expressed proteins were sequenced, among them seven showed increased (GRP78, PDIA3, ENOA, ECH1, PRDX4, ERP29, ECHM), eleven decreased (TRFE, ALBU, K2C1, ACTG, CSH2, PRDX2, FABP5, HBG1, FABP4, K2C8, K1C9) expression in first-trimester compared to the full-term placentas and two proteins exclusively expressed in first-trimester placentas (MESD, MYDGF).ConclusionAccording to Reactome and PANTHER softwares, these proteins were mostly involved in response to chemical stimulus and stress, regulation of biological quality, programmed cell death, hemostatic and catabolic processes, protein folding, cellular oxidant detoxification, coagulation and retina homeostasis. Elucidation of alteration in protein signature during placental development would provide researchers with a better understanding of the critical biological processes of placentogenesis and delineate proteins involved in regulation of placental function during development.

The jury is still out on the existence of a placental microbiome.

The human microbiome is crucial for regulating normal development, but the exact point when it is established remains unknown. A sterile placenta was traditionally considered a prerequisite for a healthy pregnancy, but studies have revealed that the placenta harbours microbial communities, even under normal conditions. However, reports have failed to provide evidence for the consistent presence of bacteria in the normal human placenta, challenging the in utero colonisation hypothesis. This mini review examines our understanding of the potential placental microbial colonisation in normal healthy pregnancies. This may impact the metabolic and immune functions of the growing foetus and have long-term consequences.

Expression profiling of RTL1 in human breast cancer tissues and cell lines

Breast cancer (BC) is the most common cancer in females. In this regard, the identification of molecular alterations driving BC is an immediate need for developing effective immunotherapeutic tools. Here we investigated the expression of a placenta-specific protein, Retrotransposon-like 1 (RTL1) in a series of BC tissues and cell lines. RTL1-specific polyclonal antibody was generated and characterized. Using tissue microarray immunohistochemistry, expression of RTL1 in a total of 147 BC and 36 non-malignant breast tissues was investigated and the association of patient's clinicopathological parameters with RTL1 expression was then examined. Expression of RTL1 in four BC cells was assessed by flow cytometry, immunofluorescent staining and Western blotting. We observed a mixture pattern of nuclear and cytoplasmic RTL1 expression in most tissues examined, however nuclear expression was found to be dominant pattern of expression. The level of nuclear RTL1 expression was significantly higher in BC tissues (P < 0.001). A statistically significant association between nuclear RTL1 expression and histological grade and vascular invasion was found (P < 0.001 and P < 0.05). All cell lines expressed RTL1 with varying degrees at their surface. The most invasive BC cell line MDA-MB-231, compared to T-47D, SKBR3 and MCF7 expressed higher levels of RTL1 at their surface. Cells with a low level of surface expression, expressed high levels of intracellular RTL1 expression. Our antibody reacted with a specific band of about 125 KD in normal human placenta and all cell lines examined. In contrast to placenta, two additional bands were also observed in cancer cell lines. Our results showed for the first time that RTL1 is differentially expressed in BC compared to non-malignant breast tissues and is associated with a higher grade and vascular invasion. In BC cells with high metastatic and invasive potential, this antigen is mostly confined to cell surface compartment indicating the possibility of using antibody-based immunotherapy for advanced metastatic BC patients.

Comparative Histological study of Normal human placentae with Hypertensive placentae in Western part of Tamilnadu state (Kongunadu region)

Background: Placenta forms an intimate bonding between the mother and the foetus. Apart from gaseous exchange, transport of nutrients from mother to the foetus, any disease or disorder affecting the mother also reflects the foetus. Hypertension during antenatal period produces maternal vasospasm and vascular damage due to luminal constriction of uterine spiral arterioles causing histological changes in the placenta and ultimately leads to foetal hypoxia and death. Aim of the study: To compare the histological features between the normal and hypertensive placenta. Materials and Methods: 30 normal and 30 hypertensive placentae were collected, processed in the Department of Anatomy, Karpagam Faculty of Medical Sciences and Research, Coimbatore, Tamilnadu, and visualised under light microscope and their histological features are noted. Results: Various histological features obtained are Syncytial knots, Fibrinoid necrosis, Calcification of Placenta, Placental infarction, Stromal fibrosis, Hypovascularity of Villous, Intervillous fibrin deposition and Basement membrane thickening. Conclusion: In our study, the histological features are more prominent and their areas in microscopic field is noted more in hypertensive placentae than normal placentae.Histtological features are mainly due to maternal vasospasm and vascular endothelial injury.The statistical analysis calculated for the histological features was found to be significant. KEY WORDS: Placenta, Hypertension, Histology, Syncytial knots, Calcification.

Andrographolide promotes proliferative and osteogenic potentials of human placenta-derived mesenchymal stem cells through the activation of Wnt/\u03b2-catenin signaling

IntroductionThe in vitro expansion and differentiation of mesenchymal stem cells derived from bone marrow (BM-hMSCs) are considered as potential therapeutic tools for clinical applications in bone tissue engineering and regenerative medicine. However, invasive sampling and reduction in number and proliferative capacity with age are the major limitations of BM-hMSCs. Recently, human placenta-derived MSCs (PL-hMSCs) obtained by a non-invasive procedure have attracted much interest. Attempts to increase the potential of PL-hMSCs would be an important paradigm in regenerative medicine. Herein, we examined the proliferative and osteogenic effect of andrographolide (AP) on PL-hMSCs.MethodsMesenchymal stem cells were isolated from full-term normal human placentas and were characterized before using. Cell cytotoxicity and proliferative effect of AP were examined by MTT and BrdU assay, respectively. The non-toxicity concentrations of AP were further assessed for osteogenic effect determined by alkaline phosphatase (ALP) expression and activity, alizarin red staining, and osteoblast-specific gene expressions. Screening of genes involved in osteogenic differentiation-related pathways modulated by AP was explored by a NanoString nCounter analysis.ResultsPL-hMSCs generated in this study met the MSC criteria set by the International Society of Cellular Therapy. The non-cytotoxic concentrations of AP on PL-hMSCs are up to 10 μM. The compound increased PL-hMSC proliferation concomitant with increases in Wnt/β-catenin level and activity. It also enhanced osteogenic differentiation in association with osteoblast-specific mRNA expression. Further, AP promoted bone formation and increased bone structural protein level, osteocalcin, in osteoblastic cells. Gene screening analysis showed the upregulation of genes related to Wnt/β-catenin, TGFβ/BMP, SMAD, and FGF signaling pathways.ConclusionWe demonstrated, for the first time, the potential role of AP in promoting proliferation, osteogenic differentiation, and osteoblast bone formation of PL-hMSCs. This study suggests that AP may be an effective novel agent for the improvement of PL-hMSCs and stem cell-based therapy for bone regeneration.

ULTRASOUND CORRELATION OF PLACENTA PREVIA IN ANC FEMALES WITH PREVIOUS LSCS PREGNANCIES

The placenta is a highly specialized tissue that serves a number of crucial functions for survival of the foetus. It allows the foetus to receive oxygen and nutrition before it can breathe or nourish itself. The placenta releases multiple hormones that initiate and maintain the essential changes of pregnancy in both the mother and the foetus. (1, 2)The normal human placenta averages 22 cm (9 inch) in length and 2–2.5 cm (0.8–1 inch) in thickness, with the center being the thickest, and the edges being the thinnest.

Ethical and Regulatory Considerations of Placental Therapeutics

PurposePlacental therapeutics aim to treat placental disease; however, ethical and regulatory issues should be considered if the drug also potentially affects the fetus. Drugs that might transfer or edit genes carry a specific challenge because currently fetal gene editing and fetal gene therapy are considered unethical. MethodsThis article reviews the literature on ethical and regulatory considerations for placental therapeutics. FindingsProposals for maternal gene therapy, directed to the maternal side of the placenta, have been discussed with patients and stakeholders. No absolute ethical, legal, or regulatory barriers to this potential treatment were identified. Patients who have experienced placental disease, such as fetal growth restriction, are interested in these therapies; some would participate in first-in-human trials. Such trials need careful regulatory considerations, such as the steps required to indicate tolerability and efficacy in preclinical models and the optimal animals for reproductive toxicology studies. Ex vivo dual human placenta perfusion experiments and villous explant in vitro studies allow drugs to be tested in normal and diseased human placenta, providing short-term tolerability and toxicologic assessment. Testing drugs in nonhuman primates is an option but carries ethical and feasibility considerations. Selection of inclusion and exclusion criteria for clinical trial participants is important to ensure that the most suitable patients are exposed to a first-in-human drug. These patients will almost certainly be pregnant women with a high risk of perinatal loss and/or perinatal and maternal morbidity. Criteria should identify sufficient numbers of patients to make a trial feasible as well as a phenotype that will respond to the mechanism of action. How to dose escalate and to capture information on adverse events are also key to optimal clinical trial design. ImplicationsDeveloping placental therapeutics requires input from scientists, practitioners, and regulators and close liaison with patients to ensure that new drugs are tested as carefully as possible.

Alternative splicing in normal and pathological human placentas is correlated to genetic variants

Two major obstetric diseases, preeclampsia (PE), a pregnancy-induced endothelial dysfunction leading to hypertension and proteinuria, and intra-uterine growth-restriction (IUGR), a failure of the fetus to acquire its normal growth, are generally triggered by placental dysfunction. Many studies have evaluated gene expression deregulations in these diseases, but none has tackled systematically the role of alternative splicing. In the present study, we show that alternative splicing is an essential feature of placental diseases, affecting 1060 and 1409 genes in PE vs controls and IUGR vs controls, respectively, many of those involved in placental function. While in IUGR placentas, alternative splicing affects genes specifically related to pregnancy, in preeclamptic placentas, it impacts a mix of genes related to pregnancy and brain diseases. Also, alternative splicing variations can be detected at the individual level as sharp splicing differences between different placentas. We correlate these variations with genetic variants to define splicing Quantitative Trait Loci (sQTL) in the subset of the 48 genes the most strongly alternatively spliced in placental diseases. We show that alternative splicing is at least partly piloted by genetic variants located either in cis (52 QTL identified) or in trans (52 QTL identified). In particular, we found four chromosomal regions that impact the splicing of genes in the placenta. The present work provides a new vision of placental gene expression regulation that warrants further studies.

The expression patterns of IL-1β and IL-10 and their relation to CYP epoxygenases in normal human placenta

IntroductionThe success of pregnancy depends on the regulation of immunological processes in the placenta. Important mediators of an immune response include pro- and anti-inflammatory interleukins which may be regulated by CYP epoxygenases and their metabolites. The relation between interleukins and CYP epoxygenases expression in human placenta has not yet been studied vastly. Material and methodsWe investigated the expression patterns of IL-1β and IL-10 in embryonic (n=8), early foetal (n=16) and term (n=7) human placenta tissue by an immunohistochemical method and evaluated the results by Kruskal-Wallis test. The obtained data was correlated using Spearman’s correlation coefficient to our previously published data of CYP epoxygenases expression in the same samples. To confirm that Hofbauer cells express IL-10 and IL-1β as well as CYP2C8 and IL-10 together, and thus there is a relation between proteins of interests, we used multiplex immunofluorescent staining. ResultsThe expression of IL-1β decreased with gestational age in cytotrophoblast, syncytiotrophoblast, as well as in Hofbauer cells whilst IL-10 decreased in cytotrophoblast, remained at the same levels in syncytiotrophoblast and increased in Hofbauer cells. In trophoblast cells, we found a statistically significant positive correlation between the expression of CYP2J2 and CYP2C9 with IL-1β, whereas there was no relation between IL-10 and any of the tested CYP epoxygenases. In Hofbauer cells, we found a significant positive correlation between CYP2C8 and IL-10 and a significant negative correlation between CYP2C8 and IL-1β. ConclusionOur results showed that the exact role and relation of interleukins and CYP epoxygenases and their metabolites is dependent on their respective cellular context. Because of IL-10, IL-1β, as well as HBCs play a role in various pathological conditions, further investigation of the exact role of CYP epoxygenase, interleukins and their relations is needed.

Morphometric analysis of Hofbauer cells in normal placenta and chorioamnionitis in humans.

Hofbauer cells are macrophages residing in the stroma of placental villi and play a number of roles during normal pregnancy, as well as pathological conditions. A morphometric analysis of Hofbauer cells, in particular to investigate the number of cells, their size and shape in samples of normal human placenta from 1st trimester, term and with chorioamnionitis was performed. Tissue samples were immunostained for CD206 antigen and evaluated using ImageJ software. We detected significant changes in number and morphology of HBCs between normal placenta and placenta with chorioamnionitis samples. In chorioamnionitis, the cells were unevenly distributed within the villi, generally present in higher numbers, larger and more elongated than those in normal 1st trimester and term placenta.

MiR-18a Contributes to Preeclampsia by Downregulating Smad2 (Full Length) and Reducing TGF-β Signaling

The study investigated the regulation of Smad2 by miR-18a and its role in preeclampsia (PE). Bioinformatics analysis showed that both Smad2 and Smad3 were the predicted targets for miR-18a. Mass spectrum analysis showed that two mature Smad2 isoforms existed in human placenta: full length, Smad2(FL), and that lacking exon3, Smad2(Δexon3). The protein level of Smad2(FL), but not Smad2(Δexon3) or Smad3, was significantly increased in severe PE (sPE) placenta, which was inversely correlated with the level of miR-18a. Elevated Smad2(FL) phosphorylation level appeared in sPE placenta, and Smad2 was colocalized with miR-18a in various subtypes of trophoblasts in human placenta. Smad2(FL) was validated as the direct target of miR-18a in HTR8/SVneo cells. miR-18a enhanced trophoblast cell invasion, which was blocked by the overexpression of Smad2(FL). Furthermore, overexpression of miR-18a repressed Smad2 activation and the inhibition of trophoblast cell invasion by transforming growth factor-β (TGF-β). In conclusion, our results suggest that miR-18a inhibits the expression of Smad2(FL), but not Smad2(Δexon3) or Smad3, which can reduce TGF-β signaling, leading to the enhancement of trophoblast cell invasion. A lack of miR-18a, which results in the upregulation of Smad2(FL), contributes to the development of PE.