Search

Background/Aims: Circular RNAs (circRNAs) are transcribed prevalently in the genome; however, their potential roles in multiple cardiovascular diseases, particularly preeclampsia (PE), are not yet well understood. This study investigated the expression profiles of circRNAs and explored circRNA-mediated pregnancy-associated plasma protein A (PAPP-A) expression as a potential biomarker for PE before 20 weeks of pregnancy. Methods: A nested case-control two-phase screening/validation study was performed in pregnant women before 20 weeks of gestation (before clinical diagnosis) at Guangzhou Women and Children's Medical Center from 2012 to 2015. In the screening phase, circRNA expression profiles of blood cells were assessed using a human circRNA microarray, which was designed to detect simultaneously 5396 circRNAs, in 5 patients with PE and 5 age- and gestational week-matched controls. In the validation phase, 18 circRNAs in blood cells predicted by bioinformatics tools were validated by quantitative reverse transcription PCR in a cohort of 60 patients (PE and age-, gestational week-, and sample storage time-matched controls). Then, we examined the involvement of circRNAs in PE-related pathways via interactions with miRNAs by multiple bioinformatics approaches. Bioinformatics analysis predicted that hsa_circ_0004904 and hsa_circ_0001855 miRNA sponges directly target PAPP-A. PAPP-A was verified in the serum of the same cohort of patients using an enzyme-linked immunosorbent assay. Finally, we combined PAPP-A with circRNAs to create a novel preclinical diagnostic model for PE with logistic regression and evaluated the efficiency of this model with receiver operating curve analysis. Results: Volcano plot analysis using various parameters showed that circRNAs were differentially expressed among both groups (P < 0.01, fold change > 2). In the screening phase, we found that 2178 circRNAs were differentially expressed between the control and PE groups, in which 884 circRNAs were downregulated and 1294 circRNAs were upregulated in the PE group compared with thecontrol group. In the validation phase, two circRNAs, hsa_circ_0004904 and hsa_circ_0001855, were significantly upregulated in PE patients compared with healthy pregnant women (P < 0.05). PAPP-A expression levels, related to the two circRNAs based on bioinformatics prediction, were increased in the PE group compared with the control group. The area under the curve of the combined model was 0.94 in the predicted PE subjects. Conclusions: This is the first study to report circRNA profiling in patients with PE prior to the onset of symptoms. Our data suggested that hsa_circ_0004904 and hsa_circ_0001855 combined with PAPP-A might be promising biomarkers for the detection of PE. Moreover, circRNAs may provide new insights into the potential mechanisms underlying the pathophysiology of PE. [ABSTRACT FROM AUTHOR]

Study Question: Are there any phenotypic and structural/architectural changes in the vessels of endometrium and superficial myometrium during the normal menstrual cycle in healthy women and those with heavy menstrual bleeding (HMB)?Summary Answer: Spatial and temporal differences in protein levels of endothelial cell (EC) markers and components of the extracellular matrix (ECM) were detected across the menstrual cycle in healthy women and these are altered in HMB.What Is Known Already: HMB affects 30% of women of reproductive age with ~50% of cases being idiopathic. We have previously shown that the differentiation status of endometrial vascular smooth muscle cells (VSMCs) is altered in women with HMB, suggesting altered vessel maturation compared to controls. Endometrial arteriogenesis requires the co-ordinated maturation not only of the VSMCs but also the underlying ECs and surrounding ECM. We hypothesized that there are spatial and temporal patterns of protein expression of EC markers and vascular ECM components in the endometrium across the menstrual cycle, which are altered in women with HMB.Study Design, Size, Duration: Biopsies containing endometrium and superficial myometrium were taken from hysterectomy specimens from both healthy control women without endometrial pathology and women with subjective HMB in the proliferative (PP), early secretory (ESP), mid secretory (MSP) and late secretory (LSP) phases (N = 5 for each cycle phase and subject group). Samples were fixed in formalin and embedded in paraffin wax.Participants/materials, Setting, Methods: Serial sections (3μm thick) were immunostained for EC markers (factor VIII related antigen (F8RA), CD34, CD31 and ulex europaeus-agglutinin I (UEA-1) lectin), structural ECM markers (osteopontin, laminin, fibronectin and collagen IV) and for Ki67 to assess proliferation. Immunoreactivity of vessels in superficial myometrium, endometrial stratum basalis, stratum functionalis and luminal region was scored using either a modified Quickscore or by counting the number of positive vessels.Main Results and the Role Of Chance: In control samples, all four EC markers showed a dynamic expression pattern according to the menstrual cycle phase, in both endometrial and myometrial vessels. EC protein marker expression was altered in women with HMB compared with controls, especially in the secretory phase in the endometrial luminal region and stratum functionalis. For example, in the LSP expression of UEA-1 and CD31 in the luminal region decreased in HMB (mean quickscore: 1 and 5, respectively) compared with controls (3.2 and 7.4, respectively) (both P = 0.008), while expression of F8RA and CD34 increased in HMB (1.4 and 8, respectively) compared with controls (0 and 5.8, respectively) (both P = 0.008). There was also a distinct pattern of expression of the vascular structural ECM protein components osteopontin, laminin, fibronectin and collagen IV in the superficial myometrium, stratum functionalis and stratum basalis during the menstrual cycle, which was altered in HMB. In particular, compared with controls, osteopontin expression in HMB was higher in stratum functionalis in the LSP (7.2 and 11.2, respectively P = 0.008), while collagen IV expression was reduced in stratum basalis in the MSP (4.6 and 2.8, respectively P = 0.002) and in stratum functionalis in the ESP (7 and 3.2, respectively P = 0.008).Limitations, Reasons For Caution: The protein expression of vascular EC markers and ECM components was assessed using a semi-quantitative approach in both straight and spiral arterioles. In our hospital, HMB is determined by subjective criteria and levels of blood loss were not assessed.Wider Implications Of the Findings: Variation in the protein expression pattern between the four EC markers highlights the importance of choice of EC marker for investigation of endometrial vessels. Differences in expression of the different EC markers may reflect developmental stage dependent expression of EC markers in endometrial vessels, and their altered expression in HMB may reflect dysregulated vascular development. This hypothesis is supported by altered expression of ECM proteins within endometrial vessel walls, as well as our previous data showing a dysregulation in VSMC contractile protein expression in the endometrium of women with HMB. Taken together, these data support the suggestion that HMB symptoms are associated with weaker vascular structures, particularly in the LSP of the menstrual cycle, which may lead to increased and extended blood flow during menstruation.Study Funding/competing Interest(s): This study was funded by Wellbeing of Women (RG1342) and Newcastle University. There are no competing interests to declare.Trial Registration Number: Not applicable. [ABSTRACT FROM AUTHOR]

Macrophages perform many specific functions including host defense, homeostasis, angiogenesis, and tissue development. Macrophages are the second most abundant leukocyte population in the non-pregnant endometrium and pregnant decidua and likely play a central role in the establishment and maintenance of normal pregnancy. Importantly, aberrantly activated uterine macrophages can affect trophoblast function and placental development, which may result in various adverse pregnancy outcomes ranging from pre-eclampsia to fetal growth restriction or demise. Only by fully understanding the roles of macrophage in pregnancy will we be able to develop interventions for the treatment of these various pregnancy complications. This review discusses the general origin and classification of monocytes and macrophages and focuses on the phenotype and functional roles of decidual macrophage at the maternal-fetal interface in normal pregnancy, as well as discussing the potential contribution of the abnormal state of these cells to various aspects of pregnancy pathologies. [ABSTRACT FROM AUTHOR]

Vascular endothelial growth factor (VEGF) is a potent endothelial cell mitogen and angiogenic growth factor that enhances endothelial cell invasion through the extracellular matrix (ECM). While various cell types express VEGF receptors, little is known about the biological actions of VEGF on nonendothelial cells. Therefore, the main objective of the present study was to determine the effect of VEGF on the in vitro invasiveness and proliferation of human MDA-MB-231 breast carcinoma cells and human HTR-8/SVneo trophoblast cells. Reverse-transcriptase polymerase chain reaction analysis demonstrated the presence of transcripts encoding VEGF receptors (VEGFR) -1, -2, and -3 as well as neuropilins-1 and -2 in the trophoblast cells, and the presence of transcripts encoding VEGFR-2 and neuropilins-1 and -2 in the breast carcinoma cells. Both cell lines also expressed transcripts for VEGF-A, -B, -C and -D, as well as for placenta growth factor (PlGF). Although incubation with exogenous VEGF-A165 or VEGF-A121 did not affect the rate of proliferation of either the trophoblast or the breast carcinoma cells, incubation with these molecules reduced their ability to invade through reconstituted ECM (Matrigel). The effect of VEGF-A165 on the invasiveness of both cell lines was inhibited by the inclusion of a neutralizing antibody to VEGF. Exogenous VEGF-A165 also decreased the cell surface expression of the urokinase-type plasminogen activator (a molecule required for invasion) by the breast carcinoma and trophoblast cells. These results indicate that the biological actions of VEGF on certain cell types may differ from the effects of this molecule on vascular endothelial cells, and therefore are relevant to angiogenesis-based therapies. [Copyright &y& Elsevier]

During early pregnancy, fetal-derived extravillous trophoblast cells (EVT) from the placenta invade the maternal decidua and inner third of the uterus where they establish fetal tolerance and remodel the uterine spiral arteries, which ensures establishment of a successful pregnancy. Aberrant EVT invasion and spiral artery remodeling is associated with a number of pregnancy complications including miscarriage, preeclampsia, fetal growth restriction, and placenta accrete. During invasion of the maternal tissues, the EVT interact with a number of different cell types including the decidual leukocytes. EVT express HLA-C, HLA-G, HLA-E, and HLA-F and interact with uterine natural killer (uNK) cells through a series of different receptors. Epidemiological evidence suggests that different combinations of HLA-C and killer cell Ig-like receptor (KIR) haplotypes impact pregnancy success. Therefore, there is much interest in the functional consequence of interactions between EVT and uNK cells, and several different methodologies have been used to isolate these different cell types and their co-culture., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Introduction: Premature placental aging is implicated in a number of complications of pregnancy including preeclampsia. A placenta knockout mouse model has shown a relationship between SIRT1, aging and placental dysfunction. The role of SIRT1 in cellular senescence has been extensively studied in various cell types, but its role in trophoblast senescence is almost unknown., Methods: Human placental samples were obtained from preeclampsia-affected women and healthy controls. The placental aging profiles were assessed by Doppler ultrasound, placental histopathology, and evaluation of senescence- and ECM-related markers. The SIRT1 expression pattern relevant to placental aging profiles was studied in premature aging placenta with preeclampsia (32-37 weeks gestation, n = 10) and healthy controls (37-40 weeks gestation, n = 10). Using cell culture, the effects of activation and knockdown of SIRT1 or its downstream target molecules in syncytialized BeWo cells were evaluated., Results: SIRT1 was expressed by syncytiotrophoblast across normal gestation. In preeclamptic premature aging placentas, SIRT1 was significantly downregulated, while senescence- and extracellular matrix (ECM) -related protein levels were upregulated compared to controls. Immunohistochemistry showed these changes to be confined to syncytiotrophoblast. In vitro, SIRT1 activation in response to resveratrol (RSV) abrogated senescence in forskolin-induced syncytialization of BeWo cells via regulation of senescence- and ECM-related proteins, and filamentous actin (F-actin). These effects were restored by SIRT1 siRNA., Discussion: The downregulation of SIRT1 may accelerate senescence of syncytiotrophoblast via targets contributing to regulation of the cell cycle, ECM production and cytoskeleton reorganization leading to premature placental aging observed in preeclampsia., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Spontaneous preterm birth (sPTB, delivery <37 weeks gestation), accounts for approximately 10% of births worldwide; the aetiology is multifactorial with intra-amniotic infection being one contributing factor. This study aimed to determine whether asymptomatic women with a history of sPTB or cervical surgery have altered levels of inflammatory/antimicrobial mediators and/or microflora within cervical fluid at 22-24 weeks gestation. External cervical fluid was collected from women with history of previous sPTB and/or cervical surgery at 22-24 weeks gestation (n = 135). Cytokine and antimicrobial peptides were measured on a multiplex platform or by ELISA. qPCR was performed for detection of 7 potentially pathogenic bacterial species. IL-8 and IL-1β levels were lower in women who delivered preterm compared to those who delivered at term (IL-8 P = 0.02; IL-1β P = 0.04). There were no differences in elafin or human beta defensin-1 protein levels between the two groups. Multiple bacterial species were detected in a higher proportion of women who delivered preterm than in those who delivered at term (P = 0.005). Cervical fluid IL-8 and IL-1β and microflora have the potential to be used as biomarkers to predict sPTB in high risk women.

Background and Objectives: Vitamin D deficiency during pregnancy has been associated with many adverse pregnancy and birth outcomes. Low serum 25-hydroxyvitamin D (25OHD) levels (<30 nmol/L) increases the risk of nutritional rickets. This study aimed to investigate the concentration of cord serum 25OHD in a birth cohort in Guangzhou, China and determine whether maternal lifestyle factors had any effect on these levels., Methods and Study Design: A total of 854 pregnant women giving birth between Dec 2016 and Dec 2017 were recruited to this study. Basic information was obtained from the clinical database. A voluntary retrospective pregnancy lifestyle questionnaire was completed by 388 participants. The concentration of serum 25OHD, calcium, phosphorus, and alkaline phosphatase (ALP) were measured in umbilical cord blood., Results: The mean (SD) of cord serum 25OHD was 44.7 (16.7) nmol/L. The prevalence of cord 25OHD <30 nmol/L was 22.2% and 70.4% had levels <50 nmol/L. The prevalence of vitamin D deficiency is higher in infants born in winter months (31% <30 nmol/L and 76% <50 nmol/L), compared to those born in the summer (12% <30 nmol/L and 64% <50 nmol/L). Infants born to women taking a vitamin D containing supplement had approximately 10 nmol/L higher levels of 25OHD than those who did not supplement their diets., Conclusions: Summer born infants have higher serum 25OHD levels at birth, but there are still infants being born with vitamin D deficiency. Vitamin D containing supplement use during pregnancy was effective in raising cord serum vitamin D levels.

Considerable work is being carried out on endometrial NK cells to determine whether they play a role in successful pregnancy outcome. In addition there is debate about whether measurements of uNK should be included in the clinical assessment for women with recurrent implantation failure or recurrent miscarriage. A hindrance to taking this forward is the fact that the density of uNK cells reported by different centres is very different. The aim of this study was to determine the reason for these differences and to develop a standardised method. Three centres participated in the study. Each centre exchanged five formalin fixed, wax embedded sections of endometrium from five women. Sections were immunostained for CD56. Images were taken of 10 random fields at ×400 magnification; total stromal and uNK cells were counted using Image J. Results were expressed as % positive uNK cells and the variation in counts obtained in each centre was compared. After initial analysis a standardised protocol was agreed and the process repeated. Significant variation was seen in the counts obtained after initial analysis (Centre A vs.B, mean difference=-0.72 P<0.001; A vs.C mean difference=-0.47 P<0.001; B vs.C, mean difference=0.25 P=0.085). Analysis suggested that differences may be due to duration of tissue fixation, the embedding and sectioning processes, selection of areas for assessment, definition of immunopositive cells and inclusion or exclusion of blood vessels. Adoption of a standardised protocol reduced the variation (Centre A vs.B mean difference=-0.105 P=0.744; A vs.C mean difference=0.219 P=0.150; B vs.C mean difference=0.32 P=0.031). Use of a standardised method is needed to establish a normal range for uNK cells and to develop a meaningful clinical test for uNK cell measurements., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Heavy menstrual bleeding (HMB) affects 30% of women of reproductive age and significantly interferes with quality of life. Altered endometrial vascular maturation has been reported in HMB and recurrent miscarriage, the latter associated with increased uterine natural killer (uNK) cell numbers. This study compared endometrial leukocyte populations in controls and women with HMB. Formalin-fixed paraffin-embedded endometrial biopsies from controls (without endometrial pathology) and HMB were immunostained for CD14 (macrophages), CD56 (uNK cells), CD83 (dendritic cells), FOXP3 (regulatory T cells/Tregs), CD3 and CD8 (T cells). Leukocyte numbers were analysed as a percentage of total stromal cells in five randomly selected fields of view in the stratum functionalis of each sample. In control women across the menstrual cycle, 2-8% of total stromal cells were CD3(+) cells, 2-4% were CD8(+) T cells and 6-8% were CD14(+) macrophages. Compared with controls, CD3(+) cells were reduced during the mid-secretory phase (4%, P<0.01) and increased in the late secretory phase (12%, P=0.01) in HMB. CD83(+) dendritic cells and FOXP3(+) Tregs were scarce throughout the menstrual cycle in both groups. In controls, 2% of stromal cells in proliferative endometrium were CD56(+) uNK cells, increasing to 17% during the late secretory phase. In HMB, CD56(+) uNK cells were increased in the proliferative (5%, P<0.01) and early secretory (4%, P<0.02) phases, but reduced (10%, P<0.01) in the late secretory phase. This study demonstrates dysregulation of uNK cells in HMB, the functional consequence of which may have an impact on endometrial vascular development and/or endometrial preparation for menstruation., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

The establishment of pregnancy requires the co-ordinated implantation of the embryo into the receptive decidua, placentation, trophoblast invasion of the maternal decidua and myometrium in addition to remodelling of the uterine spiral arteries. Failure of any of these steps can lead to a range of pregnancy complications, including miscarriage, pre-eclampsia, fetal growth restriction, placenta accreta and pre-term birth. Cytokines are small multifunctional proteins often derived from leucocytes and have primarily been described through their immunomodulatory actions. The maternal-fetal interface is considered to be immunosuppressed to allow development of the semi-allogeneic placental fetal unit. However, cytokine profiles of the decidua and different decidual cell types suggest that the in vivo situation might be more complex. Data suggest that decidual-derived cytokines not only play roles in immunosuppression, but also in other aspects of the establishment of pregnancy, including the regulation of trophoblast invasion and spiral artery remodelling. This review focuses on the potential role of decidua-derived cytokines in the aetiology of unexplained spontaneous miscarriage., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

The human endometrium contains a substantial population of leucocytes which vary in distribution during the menstrual cycle and pregnancy. An unusual population of natural killer (NK) cells, termed uterine NK (uNK) cells, are the most abundant of these cells in early pregnancy. The increase in number of uNK cells in the mid-secretory phase of the cycle with further increases in early pregnancy has focused attention on the role of uNK cells in early pregnancy. Despite many studies, the in vivo role of these cells is uncertain. This chapter reviews current information regarding the role of uNK cells in healthy human pregnancy and evidence indicating their importance in various reproductive and pregnancy problems. Studies in humans are limited by the availability of suitable tissues and the limitations of extrapolation from animal models.

Invasion of extravillous trophoblast cells (EVTs) into the maternal tissues is a key step in the development of a successful pregnancy, excessive and insufficient EVT invasion being associated with pregnancy complications. These pregnancy complications include preeclampsia and fetal growth restriction, both of which are associated with maternal and fetal morbidity and mortality at the time of birth and with increased risk of cardiovascular disease, diabetes and obesity in adult life for infants born from these conditions. In addition, women who develop preeclampsia are also at a greater risk of cardiovascular disease in later life. Many factors, protein and environmental, have been shown to both up- and down-regulate this process in vitro via different mechanisms. The redundancy observed in the regulation of this system suggests that dysregulation of one factor may not contribute to the pathological conditions of EVT invasion and that the relative local concentrations of many different factors may be more important. This review article explores the possibility that the modulation of EVT invasion as a therapeutic target for pregnancies affected by preeclampsia and fetal growth restriction may not be possible or needs to concentrate on the modulation of cell activity as a whole and not of one particular factor., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Background: Extravillous trophoblast cell (EVT) invasion of decidua and inner third of the myometrium is critical for a successful pregnancy. Many decidual factors are likely to play a role in regulating this process, including uterine natural killer (uNK) cell-derived cytokines., Hypotheses: 1) uNK cells are a major source of IFN gamma (IFN-gamma) and 2) IFN-gamma inhibits EVT invasion via an increase in EVT apoptosis and/or a decrease in active protease levels., Methods: Total decidual and uNK cells from 8-10 wk and 12-14 wk gestational age were cultured. IFN-gamma mRNA (real-time RT-polymerase chain reaction) and protein levels (FastQuant multicytokine analysis) were determined. EVT invasion in the presence of IFN-gamma or anti-IFN-gamma-neutralizing antibodies was assessed. Trophoblast apoptosis and proliferation was assessed in explants by immunohistochemistry for M30 and Ki67. Substrate zymography was performed to determine levels of secreted MMP2, MMP9, and uPA., Results: mRNA and protein for IFN-gamma was detected in both total decidual and uNK cell fractions. Trophoblast invasion was inhibited by IFN-gamma. The level of M30-positive EVT was increased in the presence of IFN-gamma whereas levels of secreted MMP2 were decreased., Conclusions: uNK cells are a source of IFN-gamma within early human pregnancy decidua. Mechanisms of IFN-gamma inhibition of EVT invasion include both increased EVT apoptosis and reduced levels of active proteases.

Tumour hypoxia is associated with resistance to therapy and with increased invasion and metastatic potential. Recent studies in our laboratory have shown that the hypoxic up-regulation of tumour cell invasiveness and chemoresistance is in part due to reduced nitric oxide (NO) signaling. Using B16F10 murine melanoma cells, we demonstrate here that the increased metastatic potential associated with exposure to hypoxia is mediated by a reduction in cGMP-dependent NO-signaling. Pre-incubation of B16F10 cells in hypoxia (1% vs. 20% O(2)) for 12 hr increased lung colonization ability by over 4-fold. This effect of hypoxia on metastasis was inhibited by co-incubation with low concentrations of the NO-mimetic drugs glyceryl trinitrate (GTN) and diethylenetriamine NO adduct (DETA/NO). In a manner similar to hypoxia, pharmacological inhibition of NO synthesis resulted in a significant increase in lung nodule formation, an effect that was prevented by co-incubation with GTN. An important NO-signaling pathway involves the activation of soluble guanylyl cyclase and the consequential generation of cGMP. Culture in the presence of a non-hydrolysable cGMP analogue (8-Br-cGMP) abrogated the hypoxia-induced lung nodule formation, suggesting that the effects of NO on metastasis are mediated via a cGMP-dependent pathway. These findings suggest that a novel mechanism whereby hypoxia regulates metastatic potential involves a downstream inhibition of cGMP-dependent NO signaling., (Copyright 2004 Wiley-Liss, Inc.)