Trophoblast deportation part I: review of the evidence demonstrating trophoblast shedding and deportation during human pregnancy

Trophoblast deportation was first described before the turn of the 20th Century by the German Scientist Schmorl and is now considered a normal physiological process during human pregnancy. Increased shedding and deportation of placental trophoblast is well documented in preeclampsia, one of the most common diseases of pregnancy. This review summarises the seminal historical and contemporary publications that have contributed to our knowledge of trophoblast deportation to the maternal lungs, their presence and quantity in the maternal circulation during normal pregnancy and during preeclampsia, and the range of morphologies deported trophoblasts display. Finally, the contentious nature of the deported multinucleated trophoblasts’ current nomenclature (syncytial knots vs. sprouts) is considered.     

Expression of serum amyloid A4 in human trophoblast-like choriocarcinoma cell lines and human first trimester/term trophoblast cells

Trophoblast invasion into uterine tissues represents a hallmark of first trimester placental development. As expression of serum amyloid A4 (SAA4) occurs in tumorigenic and invasive tissues we here investigated whether SAA4 is present in trophoblast-like human AC1-M59/Jeg-3 cells and trophoblast preparations of human first trimester and term placenta. SAA4 mRNA was expressed in non-stimulated and cytokine-treated AC1-M59/Jeg-3 cells. In purified trophoblast cells SAA4 mRNA expression was upregulated at weeks 10 and 12 of pregnancy. Western-blot and immunohistochemical staining of first trimester placental tissue revealed pronounced SAA4 expression in invasive trophoblast cells indicating a potential role of SAA4 during invasion.     

Expression of DAB2IP in human trophoblast and its role in trophoblast invasion

Objective: DAB2IP is a growth inhibitor present in many types of cancer cells and is associated with epigenetic regulations controlling tumor development. The primary objective of this study is to determine whether DAB2IP participates in the invasion and migration of trophoblasts during placental development.

Methods: The expressions of DAB2IP in human placentas (10 villi, 18 term placentas and 20 pre-eclampsia placentas) were determined by immunohistochemistry, Western blotting and quantitative RT-PCR. HTR8/SVneo cells were treated with hypoxia–reoxygenation (H/R) to test how DAB2IP expression would affect the invasion and migration of trophoblasts. JEG-3 andHTR8/SVneo cells were treated with 5-aza-2-deoxycytidine (5-aza-dC) to study the role of DAB2IP promoter methylation in trophoblasts.

Results: DAB2IP was strongly expressed in human villi and extravillous trophoblasts as well as in HTR8/SVneo cells, but not in pre-eclampsia placentas. DAB2IP expression increased after H/R treatment, but the invasive and migratory abilities of trophoblasts were reduced. DAB2IP expression in JEG-3 cells also increased after treatment with 5-aza-dC.

Conclusions: These findings strongly suggest that DAB2IP is an important negative regulator at the maternal–fetal interface during early pregnancy. Excessive oxidative stress can increase DAB2IP expression in trophoblasts. The mechanism of DNA methylation may involve in its function during the development of pathologic pregnancy.     

Expression of EGF,EGFR, and proliferation in placentas from pregnancies complicated with preeclampsia

Objective: To investigate proliferation, EGF and EGFR expression of villous trophoblast (VTB), decidual cells (DC), and extravillous trophoblast (EVTB) in the placentas from pregnancies complicated with preeclampsia (PE) and to compare them with placentas from normal pregnancies. Methods: Twenty-nine PE placentas and 19 control placentas were studied for EGF and EGFR immunohistochemical expression (noted as week, moderate or strong). Proliferation was expressed as the proliferation index. The CK7 antibody was used to distinguish DC from EVTB. Results: DC and EVTB proliferation was significantly higher in PE placentas. EGFR and EGF expression showed no significant difference. Conclusion: Higher DC and EVTB proliferation in PE could contribute to PE development.     

The expression of major histocompatibility complex antigens by trophoblast in ectopic tubal pregnancy

The reactivity of the various trophoblast populations found in ectopic fallopian tube pregnancy with established trophoblast-reactive markers and monoclonal antibodies to MHC antigens has been studied. In ectopic tubal pregnancy fetal trophoblast shows an identical reaction pattern with these antibodies to that seen in intrauterine pregnancy, suggesting that ectopic implantation is not related to an inherent immunological abnormality of fetal trophoblast. However, from this and other studies, it appears that extravillous trophoblast displays an unusual class I MHC antigenic structure. This observation may explain the ability of class I MHC--bearing fetal trophoblast--to survive both in the uterus and at an abnormal implantation site.     

The role of serum markers and uterine artery Doppler in identifying at-risk pregnancies

Measures of placental dysfunction, including maternal serum analytes and Doppler studies, have been linked to adverse pregnancy outcomes, although the predictive ability of any single one is poor. Improved knowledge of the multifactorial nature of many of the adverse outcomes of pregnancy has sparked interest in the use of multi-parameter models that combine maternal serum analytes with measures of placental structure and blood flow. The combination of various first-trimester and second-trimester analytes and uterine artery Doppler screening show promise as potential screening tools, but large prospective studies are needed to further define their role in clinical practice.     

Preeclampsia associates with RECK-dependent decrease in human trophoblasts migration and invasion

IntroductionPreeclampsia is characterized by reduced invasion capacity of trophoblasts involving lower matrix metalloproteinase (MMP) activity. Cell invasion is reduced by reversion-inducing-cysteine-rich protein with Kazal motifs (RECK), a plasma membrane protein that inhibits MMP in several cell types. However, it is unknown whether this mechanism happens in the human placenta from preeclampsia. The hypothesis of this study sustains that RECK expression is increased leading to reduced trophoblasts invasion in preeclampsia.MethodsRECK expression in the human first trimester trophoblast cell line HTR8/SvNeo and in placentas from normal (n = 4) and preeclampsia (n = 4) pregnancies was evaluated by Western blot and immunofluorescence. MMP-dependent gelatin hydrolyzation was measured by in situ zymography and gelatinase assay in placental and cell extracts. RECK was overexpressed (plasmidial vector transfection) or partially reduced (shRNA) to evaluate its role in HTR8/SVneo cell migration and invasion.ResultsRECK was expressed in trophoblasts layer in human placentas. Preeclampsia resulted in higher placental RECK protein abundance, reduced MMP function, and higher level of fibronectin (a MMP substrate) compared with placentas from normal pregnancies. RECK is also expressed in HTR-8/SVneo cells. Reduced RECK expression resulted in higher MMP-dependent gelatin hydrolyzation, associated to higher migration and invasion of HTR8/SVneo cells. However, RECK overexpression associated with reduced hydrolyzation, cell migration and invasion.DiscussionRECK is overexpressed in human trophoblasts from preeclampsia and may be responsible of this disease-associated lower migration and invasion of this cell type.     

Wnt5a inhibited human trophoblast cell line HTR8/SVneo invasion: implications for early placentation and preeclampsia

Objective: Wnt5a and Wnt signaling play potential roles in human placental and fetal development. The objective of this study is to explore the role of Wnt5a in the invasion of the human trophoblast cell line HTR8/SVneo and the probable mechanism of early placentation and preeclampsia in which Wnt5a is involved.

Methods: Human first trimester villous tissues from normal pregnancies and third trimester placentas from pregnancies with or without preeclampsia (PE) were used in the detection of the expression and subcellular location of Wnt5a. The human trophoblast cell line HTR8/SVneo was treated with 0–400?ng/ml recombinant Wnt5a to investigate the role of Wnt5a in human trophoblast invasion.

Results: Human first trimester villous is accompanied by the decreased expression of Wnt5a compared with term placenta. Upregulated Wnt5a was detected in PE placenta compared with the normal control. Wnt5a inhibited the migration and invasion of HTR8/SVneo cells with decreased integrin β1, α5 and N-cadherin. Moreover, Wnt5a downregulated β-catenin in HTR8/SVneo cells.

Conclusions: These findings strongly suggest that Wnt5a inhibits the invasion of HTR8/SVneo cells. Decreased Wnt5a facilitates early placentation, whereas increased Wnt5a contributes to the pathogenesis of PE with insufficient trophoblast invasion. Aberrant Wnt5a may function by impairing Wnt non-canonical/β-catenin signaling pathway in trophoblasts.     

Increased placental apoptosis in intrauterine growth restriction

OBJECTIVES: Our purpose was to investigate a possible role for apoptosis in the pathophysiologic mechanisms of intrauterine growth restriction. STUDY DESIGN: Placental samples were obtained from 43 uncomplicated third-trimester pregnancies and from 26 pregnancies complicated by intrauterine growth restriction. The definition used to identify cases of intrauterine growth restriction depended on three criteria: clinical evidence of suboptimal growth, ultrasonographic evidence of deviation from an appropriate growth percentile, and individualized birth weight ratios <10th percentile. Light microscopy was used to quantify the incidence of apoptosis. Electron microscopy and TUNEL (terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick end labeling) staining were used to confirm the occurrence of apoptosis. RESULTS: Quantification of apoptosis (medians and interquartile ranges) resulted in the following values: normal third trimester (n = 43) 0.14% of cells (0.08% to 0.20%) and intrauterine growth restriction third trimester (n  = 26) 0.24% of cells (0.16% to 0.29%). The incidence of apoptosis was significantly higher in placentas from pregnancies with intrauterine growth restriction compared with normal third-trimester placentas (p < 0.01, Mann Whitney U test). CONCLUSIONS: These results suggest that apoptosis may play a role in the pathophysiologic mechanisms of intrauterine growth restriction.(Am J Obstet Gynecol 1997;177:401)     

Prevention of cardiovascular risk in women who had hypertension during pregnancy after 36 weeks gestation

Objective. The aim of our study was to investigate a possible correlation between the expression of the placenta-secreted hormones, β-subunit of human chorionic gonadotrophin (βhCG) and pregnancy-associated plasma protein A (PAPP-A), during the first trimester screening and the development of preeclampsia. Methods. A total of 155 patients between 11 + 0 and 13 + 6 weeks of gestation were enrolled in this study. PAPP-A and βhCG levels were measured using the KRYPTOR® system. Results. The serum levels of βhCG were significantly higher in pregnancies which subsequently developed preeclampsia. The PAPP-A concentration did not differ significantly in pregnancies complicated by preeclampsia than in uncomplicated pregnancies. Conclusion. These results might contribute to developing new tests in the prediction of preeclampsia.     

The common prothrombotic factors in nulliparous women do not compromise blood flow in the feto-maternal circulation and are not associated with preeclampsia or intrauterine growth restriction

OBJECTIVE: In this study we evaluated the associations between common prothrombotic factors and increased blood flow resistance in the feto-maternal circulation, intrauterine growth restriction, small for gestational age, or preeclampsia. STUDY DESIGN: A prospective study was conducted in healthy nulliparous women with spontaneous singleton pregnancy. Blood was tested for the common prothrombotic factors, i.e., factor V Leiden, factor II G20210A, methylenetetrahydrofolate reductase C677T, anticardiolipin, and lupus anticoagulant. Blood flow resistance in the uterine, placental, and umbilical arteries were assessed by multigate Doppler and compared between women with and without prothrombotic factors. The maternal, fetal, and neonatal clinical courses were also compared among these subgroups. RESULTS: Prothrombotic factors were detected in 191 of 637 (30%) subjects. No significant difference in resistance to blood flow in the feto-maternal unit was discernible between women with and without prothrombotic factors. Pregnancy-induced hypertension or preeclampsia occurred in 10 of 191 (5.2%) and in 19 of 446 (4.3%) of women with and without a prothrombotic factor respectively ( P = .59). Intrauterine growth restriction was detected at 31 weeks in 13 of 164 (7.9%) and in 42 of 377 (11.1%) fetuses of women with and without a prothrombotic factor ( P = .26), and small for gestational age at delivery was observed in 19 of 187 (10.2%) and in 41 of 413 (9.9%) of mothers with and without prothrombotic markers, respectively. CONCLUSION: The presence of prothrombotic factors in healthy nulliparous women does not compromise blood flow in the feto-maternal unit, nor is it associated with preeclampsia, intrauterine growth restriction, or small for gestational age .     

The proprotein convertase furin in human trophoblast: Possible role in promoting trophoblast cell migration and invasion

Furin, a proprotein convertase (PC), is ubiquitously expressed and implicated in many physiological and pathological processes. This study is aimed to identify the role of furin in human trophoblast invasion and migration. Furin was found to be highly expressed in placental villi of both rhesus monkeys and human beings during early pregnancy. Specifically, furin was found in trophoblast column and trophoblast shell, regions where highly invasive cytotrophoblast cells invade the maternal decidua during human placentation. To determine whether furin plays any role in trophoblast invasion and migration, we employed human extravillous HTR8/SVneo cells in Matrigel invasion and transwell migration assays. Knocking-down furin expression by siRNA significantly inhibited invasion and migration of HTR8/SVneo cells (P < 0.01), with corresponding decrease of matrix metalloproteinase-9 (MMP-9) activities. In contrast, over-expression of furin markedly increased cell invasion and migration (P < 0.01), accompanied by significant increase of MMP-9 activities. Furthermore, furin siRNA significantly increased the levels of both tissue inhibitors of MMPs (TIMP)-1 and -2. Our results suggest that furin may play an important role in the invasion and migration of human trophoblast cells during early pregnancy.     

STOX1在子痫前期滋养细胞炎症反应中的作用研究#br#

Objective?To investigate the STOX1 (Storkhead box1) in the inflammatory response of trophoblast cells simulated by human choriocarcinoma cell line JEG3 cells. Methods?The human choriocarcinoma cell line JEG3 cell line was used to mimic trophoblast cells, and a stable transitional cell line model with overexpression/knockdown of STOX1 gene was constructed. 0.05 mmol/L aspirin was added to the experimental and control groups, respectively. Cell migration ability was measured by Transwell, and apoptosis was detected in each group by flow cytometry, and real-time quantitative PCR and protein immunoblotting were used to detect the expression of cell-associated inflammatory, hypoxic and apoptotic factors, biological behavior, and the relative expression of mRNA and protein of molecular biology. Results?Hypoxia-inducible factor α(HIF-1α), endothelial-type nitric oxide synthase (eNOS), and inducible nitric oxide synthase (iNOS), and apoptosis B lymphocytoma-2 (Bcl-2) were significantly reduced in the overexpression STOX1 trophoblast group (P<0.01); meanwhile, inflammatory response-related factor nuclear factor κB (NF-κB), tumor necrosis factor α (TNF-α), interleukin 8 (IL-8) and cysteine protease-3 (Caspase-3) were significantly increased at both gene and protein levels (P<0.05). The expression of hypoxia-inducible genes HIF-1α (P<0.01), eNOS (P<0.05), and iNOS (P<0.001) and anti-apoptotic genes Bcl-2 (P<0.01) was elevated after knockdown of STOX1, while inflammatory responses NF-κB (P<0.05), TNF-α (P<0.01), IL-8 (P<0.01), and Caspase-3 (P>0.05) levels were reduced. Conclusion?STOX1 induces inflammatory response, promotes hypoxia and apoptosis-related gene expression in JEG3 cell line, aspirin may inhibit the effect of STOX1, reduce apoptosis in JEG3 cells, decrease inflammatory response and improve hypoxia in trophoblast cells, and knockdown of STOX1 has synergistic effect with the use of aspirin.     

Apoptosis of extravillous trophoblast cells limits the trophoblast invasion in uterine but not in tubal pregnancy during first trimester

During the first trimester of pregnancy extravillous trophoblast cells (EVT) invade the maternal decidua. Invasion normally is reduced from the second trimester onwards and stops in the inner third of the myometrium. By contrast, in extrauterine tubal pregnancy, trophoblast invasion may even penetrate the tubal wall, which ultimately leads to the rupture of the fallopian tube. Induction of apoptosis of EVT cells, by maternal immune competent cells, may be an important mechanism to limit EVT invasion in uterine pregnancy. Tissue specimens from first and second trimester uterine pregnancy and first trimester tubal pregnancy were analyzed for apoptosis by TUNEL- and M30-staining. By immunohistochemical double labelling, maternal leukocyte subtypes were co-localized to apoptotic cells and in this context, the number of CD56(+)NK cells was analyzed. Our data show that apoptosis is confined to the decidua basalis. Most apoptotic cells are single cytokeratin-positive epithelial cells residing in the stromal compartment. Consequently these cells can only be EVT cells. Maternal leukocytes are not apoptotic. They are located in close contact to apoptotic cells. The number of apoptotic cells in the second trimester (1.8+/-0.7 per cent) is reduced compared to first trimester (5.6+/-0.7 per cent) of uterine pregnancy. In parallel, the number of NK cells declines from first (24.4+/-2.9) to second (12.4+/-1.8) trimester. Furthermore, apoptosis is significantly reduced in ectopic (0.9+/-0.3 per cent) compared to eutopic first trimester pregnancies. Consequently, we suggest that in first trimester uterine pregnancy, induction of EVT cell apoptosis by the maternal immune system is one mechanism to limit EVT invasion. During the second trimester, in parallel to declining numbers of NK cells, the mechanism changes. However, in tubal pregnancy due to differing immunological microenvironments at the ectopic implantation site, apoptosis induction fails, which deleteriously may result in uncontrolled invasion and penetration of the tubal wall.     

The Evaluation of the Oxidative State of Low-Density Lipoproteins in Intrauterine Growth Restriction and Preeclampsia

Objective. To evaluate the oxidative state of lipoproteins in pregnancies complicated by intrauterine growth restriction (IUGR) in comparison to preeclampsia (PE) and healthy pregnant control subjects (CN). Methods. Maternal serum of 20 PE, 29 IUGR, and 29 gestational age-matched CN were analyzed. Total cholesterol (TC), low-density lipoprotein (LDL)-bound cholesterol (LDL-C), and oxidized LDL (oxLDL) concentration were measured once between 25 and 34 weeks of gestation. Statistical estimates were performed by Student's t-test. Results. Serum concentrations of LDL-C and TC were significantly reduced in IUGR [LDL-C: CN – mean = 146 mg/dL, SD = ± 40.1; IUGR – mean = 102 mg/dL, SD = ± 27.3 (p < 0.0001); PE – mean = 130 mg/dL, SD = 38.8 mg/dL; TC: CN – mean = 259/dL, SD = ± 46.8; IUGR – mean = 218 mg/dL, SD = ± 35.0 (p < 0.001); PE – mean = 244 mg/dL, SD = 48.2]. There was no significant difference in oxLDL/LDL-C ratio within the three groups (CN: mean = 0.76, SD = 0.24; IUGR: mean = 0.74, SD = 0.12; PE: mean = 0.77, SD = 0.22). Conclusion. Our results show a lower maternal LDL-C and TC concentration in IUGR pregnancies. These data contribute to the hypothesis of a decreased cholesterol supply to the fetus in IUGR. However, we could not confirm the hypothesis of an altered oxidative state in neither IUGR nor PE.     

Remodeling of myometrial radial arteries in preeclampsia

OBJECTIVE: This study was undertaken to test for structural differences between myometrial radial arteries isolated from women having normal pregnancies and pregnancies complicated by preeclampsia and intrauterine growth restriction. STUDY DESIGN: Pressure myography was used to study myometrial radial arteries obtained at cesarean section. With the use of a transilluminating system, lumen diameter, wall thickness, wall/lumen ratio, distensibility and stress-strain relationship were studied through a range of pressures. Arteries were then fixed in glutaraldehyde, embedded in resin, cross-sectioned, and studied in greater detail by light and electron microscopy. RESULTS: Pressure myography showed that arteries from women with preeclampsia had a reduced lumen diameter, thicker wall, and greater wall/lumen ratio compared with vessels isolated from women with normal pregnancy. Light microscopy indicated an identical media content remodeled around a smaller lumen. Electron microscopy indicated enlarged extracellular spaces in the media but no change in myocyte profile size or number. There was no clear evidence of structural changes in myometrial radial arteries isolated from women with intrauterine growth restriction compared with normal pregnancy. No differences in vessel distensibility or stress-strain relationships were detected in complicated pregnancies. CONCLUSION: The changes observed in myometrial radial arteries isolated from women with preeclampsia are due to inward eutrophic remodeling. Alterations in these vessels may contribute to increased uterine vascular resistance in preeclampsia.