Subcellular localization of PP5/TFPI-2 in human placenta: a possible role of PP5/TFPI-2 as an anti-coagulant on the surface of syncytiotrophoblasts

Placental protein 5 (PP5)/tissue factor pathway inhibitor-2 (TFPI-2), a serine proteinase inhibitor, is homologous to tissue factor pathway inhibitor (TFPI) and commonly found in peripheral blood of pregnant woman. Although TFPI is well known to be synthesized primarily in endothelium and to play an important role in regulation of the extrinsic pathway of blood coagulation, the function of PP5/TFPI-2 remains unclear. Our previous report demonstrated that PP5/TFPI-2 expression is ubiquitous and extremely high in growing placental tissue. Using newly generated polyclonal anti-PP5/TFPI-2 antibody, and by immunohistochemistry and immunoelectromicroscopy, we examined precise localization of PP5/TFPI-2 in placenta especially in syncytiotrophoblasts, which had been shown to produce PP5/TFPI-2 mRNA by our previous study using in situ hybridization.Immunoelectromicroscopy revealed PP5/TFPI-2 localizing on the surface of the microvilli and the membrane of endoplasmic reticulum of syncytiotrophoblasts. To confirm the cell surface association of PP5/TFPI-2, placental villi were incubated with heparin and resultant soluble fraction was analysed by Western blotting. Heparin liberating PP5/TFPI-2 from villi suggested that PP5/TFPI-2 might be retained on the microvilli surface through the binding to membrane-anchored proteoglycans such as glypican and/or syndecan family members. We also examined the relationship between the presence of cell layer of syncytiotrophoblasts and the coagulation using clinical specimens, and revealed that the fibrin depositions were specifically observed on the regions lacking syncytiotrophoblasts cell layer in placental villi. Therefore, it is likely that during pregnancy PP5/TFPI-2 may be retained on the surface of placental villi via proteoglycans, and may play an important role to maintain intervillous blood flow and the patency of microvasculature in feto-maternal blood system mediated by the inhibition of serine proteinases involved in the blood coagulation.     

Vascular endothelial growth factor, its receptors, and the tie receptors in the placental bed of women with preeclampsia, diabetes, and intrauterine growth retardation

Placental bed vascular function is compromised in pregnancies complicated by preeclampsia (PE), intrauterine growth retardation (IUGR), and maternal diabetes mellitus (DM). We performed an immunohistochemical analysis of vascular endothelial growth factor (VEGF), its receptors (VEGFR) -1 and -2, and the Tie-1 and -2 receptors in cryostat tissue sections of the placental bed from healthy women (n = 5) and women with PE (n = 5), IUGR (n = 5), and DM (n = 5). VEGF immunoreactivity was stable between the study groups. VEGFR-1 immunoreactivity in the myometrial vascular smooth muscle cells was strongest in the controls. In the myometrial cells, the strongest VEGFR-2 immunoreactivity was seen in women with IUGR. In the decidual vascular endothelial cells, the strongest Tie-1 immunoreactivity was observed in healthy women and in those with DM. Alterations in the placental bed expression of VEGFR-1, VEGFR -2, and Tie-1, but not of VEGF and Tie-2, may be associated with PE, IUGR, or DM.     

Decreased Pathological Placental Findings in Aspirin-Treated Pregnant Women at Risk of Hypertensive Complications

Objective: The small controlled trials reporting large reductions in the incidence of preeclampsia and intrauterine growth restriction (IUGR) in highrisk pregnant women treated with low-dose aspirin have recently been followed by large clinical trials suggesting less beneficial results. The effect of low-dose aspirin on placental lesions associated with preeclampsia and IUGR has not yet been studied.

Methods: We participated in the large multicenter randomized collaborative low-dose aspirin study in pregnancy (CLASP) trial of low-dose aspirin for the prevention and treatment of preeclampsia and intrauterine growth restriction. As part of this study, we evaluated placentae submitted from 25 women treated with aspirin and 28 with placebo.

Results: More of the pathological findings classically described in preeclampsia and IUGR were demonstrated in the placentae from the placebo group than from the aspirin group (54% vs. 16%, P = 0.02). The placental findings did not correlate with clinical pregnancy outcome or Doppler flow parameters of the fetal umbilical artery in either group.

Conclusions: Our results support the assumption that aspirin may have some inhibitory effect on the uteroplacental circulatory ischemic changes typically occurring in preeclampsia and IUGR.     

Combining 2nd-trimester maternal serum homocysteine levels and uterine artery Doppler for prediction of preeclampsia and isolated intrauterine growth restriction

AIM: To evaluate the efficacy of a combined 2nd-trimester maternal serum homocysteine and uterine artery Doppler screening at 20 weeks of gestation for complications of pregnancy: preeclampsia, isolated intrauterine growth restriction (IUGR), placental abruption, and stillbirths. METHODS: Consecutive singleton pregnancies without previous risk factors who had homocysteine measured as part of a serum-screening program for trisomy 21 had uterine artery Doppler performed. Sensitivity, specificity, positive and negative predictive values, odds ratio, and positive and negative likelihood ratios for the subsequent development of preeclampsia, isolated IUGR, placental abruption, stillbirth, and preterm delivery were calculated for the following methods (1) homocysteine cut-off level 6.3 micromol/l (95th centile); (2) on Doppler ultrasound bilateral notches with a mean resistance index (RI) >0.55 (50th centile), all unilateral notches with a mean RI >0.65 (80th centile), and absence of notches with a mean RI >0.7 (95th centile), and (3) Doppler ultrasound notch evaluation (bilateral, unilateral, absence as in method 2) combined with the homocysteine cut-off level of 6.3 micromol/l. RESULTS: By using a logistic regression model, methods 1 and 2 predicted preeclampsia (p < 0.001), isolated IUGR (p < 0.01), and "any complication" (p < 0.01). The sensitivity for prediction of preeclampsia using the combined method (3) was 61.3% for a false-positive rate of 2%, better than that for isolated IUGR (54%) below the 5th centile and "any complication" (56%). CONCLUSION: This prospective study confirms the potential of a combined method of elevated homocysteine and uterine artery Doppler screening for preeclampsia, isolated IUGR, and any obstetric complication.     

Perinatal outcome in women with severe pregnancy complications and multiple thrombophilias

Hypercoagulability leading to placental thrombosis has been implicated in severe pregnancy complications. We compared the perinatal outcome in women with severe preeclampsia, intrauterine growth retardation (IUGR) and severe abruptio placentae and multiple acquired and inherited thrombophilias (study group, n=22) to matched women with similar complications and single thrombophilia (control group, n=22). Gestational age at delivery and birth weight were significantly lower in the study group compared to the control group (p<0.01) and among the study women with severe preeclampsia and IUGR. Severe pregnancy complications may occur earlier during pregnancy and more seriously affect perinatal outcome in women with multiple thrombophilias.     

Vascular Endothelial Growth Factor Receptor 1 (Flt1) and Apoptosis in the Preeclamptic Placenta and Effects of in vivo Anti-hypertensive Exposure

Objective: To test the hypothesis that vascular endothelial growth factor receptor 1 (Flt1) is negatively correlated with apoptosis in preeclampsia placentae, and to examine the effects of antihypertensive medication on apoptosis. Methods: Flt1 and TUNEL immunoreactivity were quantitatively compared in the stromal decidual cells, villous trophoblasts, and endothelial cells of placentae from uncomplicated pregnancies (NP, n = 34) to those in patients with preeclampsia (PE, n = 30), and those in patients with preeclampsia with superimposed intrauterine growth restriction (PE + IUGR, n = 7). Further analyses determined any correlations with the antepartum use of the antihypertensives clonidine and hydralazine. Results: There was no difference in either Flt1 or TUNEL when comparing PE placentae (with or without IUGR) with NP. There were no correlations with the use of the antihypertensives. Conclusion. Apoptotic levels do not correlate with Flt1 in preeclampsia placentae and are not regulated by invivo exposure to the antihypertensives clonidine and hydralazine.     

Plasma fibronectin receptor levels during pregnancy complicated by preeclampsia and abruptio placentae.

The level of human fibronectin receptor (FNR) in plasma was measured by enzyme-linked immunosorbent assay in samples from normal pregnant women in the 1st trimester (n = 5), 2nd trimester (n = 7), 3rd trimester (n = 23), normal postpartum women day 1 (n = 4), day 2 (n = 5), day 3 (n = 8), nonpregnant women (n = 18), 20 preeclamptic patients in the 3rd trimester, and 8 patients with abruptio placentae in the 3rd trimester. In normal pregnancy, the mean value of FNR was 1.4 +/- 0.4 micrograms/ml in the 1st, 1.4 +/- 0.2 micrograms/ml in the 2nd, and 1.9 +/- 0.3 micrograms/ml (p less than 0.05) in the 3rd trimester. FNR values increased with pregnancy. During the puerperium, its level decreased with time, being 1.4 +/- 0.5 micrograms/ml (p less than 0.01) on day 1, 1.0 +/- 0.3 micrograms/ml on day 2, and 0.8 +/- 0.2 micrograms/ml on day 3. The level in preeclamptic patients was 2.0 +/- 0.4 micrograms/ml, and that in abruptio placentae was 2.7 +/- 0.4 micrograms/ml. There were significant differences between the levels in abruptio placentae versus preeclampsia (p less than 0.05) and 3rd-trimester normal pregnant women (p less than 0.01). In the immunohistochemical study, the surface of normal decidual cells stained weakly for FNR, and the decidual cell membranes of the cases of preeclampsia stained moderately or strongly. Decidual cells and their extracellular matrix close to hematomas of abruptio placentae stained very strongly for FNR.(ABSTRACT TRUNCATED AT 250 WORDS)     

Expression of serine proteinase inhibitor PP5/TFPI-2/MSPI decreases the invasive potential of human choriocarcinoma cells in vitro and in vivo

OBJECTIVE: PP5/TFPI-2/MSPI is a Kunitz-type serine proteinase inhibitor with broad inhibitory spectra, abundantly produced by placenta and detected in the blood of pregnant women. Expression of PP5/TFPI-2/MSPI is exclusively detected in syncytiotrophoblasts of placenta, but is barely detectable in choriocarcinoma cells, a trophoblast-derived malignant tumor. Chromosome 7, in which the PP5/TFPI-2/MSPI gene is localized, is frequently lost in various types of tumors. We attempted to elucidate the relation between PP5/TFPI-2/MSPI expression and the malignant properties of choriocarcinoma cells. METHODS: Human choriocarcinoma cells, JAR, were transfected with either a human PP5/TFPI-2/MSPI expression vector or an empty vector, and stable clones were obtained. Messenger RNA expression, protein secretion/localization, growth rate, and plating efficiency were evaluated. In vitro migration and invasive activity were determined by transwell chamber experiments. In vivo tumor growth was evaluated by the subcutaneous injection of cells to nude mice and followed by histological examination. RESULTS: Expression of mRNA and protein of PP5/TFPI-2/MSPI were confirmed, and a high producing clone and a low producing clone were chosen for further analysis. The majority of secreted PP5/TFPI-2/MSPI protein was revealed to associate with the extracellular matrix. Expression of PP5/TFPI-2/MSPI did not affect the growth and migration of the tumor cells, but enhanced their plating efficiency. Its expression significantly inhibited invasion through the Matrigel. Invasive growth into the subcutaneous muscle layer was not evident in the nude mouse tumors of the PP5/TFPI-2/MSPI-expressing cells. CONCLUSION: PP5/TFPI-2/MSPI-expressing choriocarcinoma cells showed suppressed potential of invasion in vitro and in vivo. It is suggested that loss or suppression of PP5/TFPI-2/MSPI expression may result in the acquisition of invasiveness in choriocarcinoma cells.     

Metabolism of vitamin D3 in the placental tissue of normal and preeclampsia complicated pregnancies and premature births

The aim of this study was to analyze the hormonal basis for low 1,25(OH)2D3 circulating levels in patients with preeclampsia and/or preterm deliveries. The activity and expression of the 1 alphaOHase, 25-OHase, 24-OHase and VDR in the placental tissue of normal pregnancies, preeclampsia-complicated pregnancies and premature births were investigated. The mRNA of the enzymes was detected in the placental tissue from preeclamptic pregnancies and compared to those of normal placental tissue. Real time PCR analysis showed a significant increased 1 alpha-OHase gene expression in preeclamptic patients, and the gene expression of 24-OHase was significantly decreased. With regard to the 25-OHase the median value of the normal placental tissue was significantly higher than in the placental tissue of preeclamptic patients. The real time analysis of all target genes also showed significant differences in normal placental tissue compared to placental tissue from premature births (VDR: p = 0.041; 1 alpha-OHase: p = 0.013; 24-OHase: p = 0.007; 25-OHase p = 0.027). Our observation of reduced VDR expression on mRNA level in placental tissue indicates a possible dependence of the modulation of VDR expression from proliferation and differentiation processes. It can be speculated whether the down-regulation of VDR in the examined placenta cells was the result of an altered production of calcitriol by these cells. We found a significantly higher 1 alpha-OHase-expression in the placental tissue of pregnant women with preeclampsia or preterm birth compared to healthy pregnant women, whereas the expression of 25-OHase was significantly reduced. These results correlate with other studies and support the significance of the placenta regarding metabolism malfunctions as they were observed in the calcium metabolism for preeclampsia. That a placenta with preeclampsia expresses less 1 alpha-OHase-mRNA and shows less 1 alpha-OHase-activity than in placental samples of inconspicuous placentae, can be granted as a specific alteration in the placental ability to synthesize adequate amounts of 1,25(OH)2D3.     

Evaluation of maternal and umbilical serum TNFalpha levels in preeclamptic pregnancies in the intrauterine normal and growth-restricted fetus.

OBJECTIVE: The aim of this study was to carry out a comparative analysis of the maternal and umbilical cord TNFalpha serum levels in pregnancies complicated by severe preeclampsia with normal intrauterine fetal growth, in preeclamptic pregnancies with intrauterine growth restriction (IUGR), and in normotensive pregnant patients. PATIENTS AND METHODS: The study was carried out on eight patients with severe preeclampsia complicated by IUGR and 18 preeclamptic patients with normal intrauterine fetal growth. The control group consisted of 18 healthy normotensive patients with singleton uncomplicated pregnancies. Maternal and umbilical serum TNFalpha concentrations were estimated using a sandwich ELISA assay. RESULTS AND CONCLUSIONS: Pregnant women with severe preeclampsia had significantly higher maternal and umbilical serum TNFalpha levels than those in the normotensive controls. Our findings and other reports indicate that TNFalpha may participate in the pathogenesis and sequelae of preeclampsia with and without IUGR. The results of excessive umbilical serum activity of tumor necrosis factor alpha (TNFalpha) in preeclamptic pregnancy complicated by intrauterine growth restriction (IUGR) may suggest additional changes and dysfunction of the placental-fetal unit and deterioration of placental function, leading to fetal hypotrophia in the course of preeclampsia.     

Preeclampsia with and without intrauterine growth restriction–Two pathogenetically different entities?

Objective: The objective of this study is to determine the differences in histopathological features of basal decidua and placenta in cases of preeclampsia with or without fetal intrauterine growth restriction (IUGR). Methods: A prospective case–control study included a study group consisting of 30 pregnant women with preeclampsia completed by cesarean section (CS), in 19 of whom preeclampsia was associated with IUGR, and in 11 it was not. The control group consisted of 20 healthy pregnant women delivered by elective CS. Placentas and samples of placental bed obtained during CS were histopathologically (HP) analyzed after hematoxylin–eosin staining and immunohistochemical labeling of Cytokeratin 7 (CK7) trophoblastic cells in decidua. Results: Regarding the HP changes in the spiral arteries in preeclampsia, the most frequent features were inadequate transformation of spiral arteries with poor trophoblastic invasion (70.0%) and fibrinoid necrosis of the media (66.7%), and rarely acute atherosis (33.3%) and thrombosis (30.0%). Villous hypermaturity was more frequently found in placentas of patients with preeclampsia with IUGR (p < 0.05), while there were no differences between subgroups of preeclampsia with and without IUGR regarding some of HP alterations of placental bed. Conclusion: Alterations of the placental bed in terms of decidual vasculopathy are more the characteristics of the preeclampsia itself than IUGR, while changes in placental villi primarily follow the presence of IUGR, which could indicate that preeclampsia with and without IUGR are two pathogenetically different entities.     

Increased endothelial thrombomodulin (TM) expression in pregnancies complicated by IUGR

Thrombomodulin (TM) is a cell surface receptor playing an important role in endothelial cell anticoagulant activity. TM is also known as a factor of angiogenesis; low TM activity correlates with impaired angiogenesis. Since vascular lesions with disorders of the placental coagulation and inadequate angiogenesis can be associated with IUGR, we hypothesized that thrombomodulin expression in the villous vasculature and syncytiotrophoblast of placentae complicated by IUGR might differ from those of normal pregnancies. Representative tissue samples of normal, IUGR as well as 1st and 2nd trimester (n = 12) placentae were collected. Immunohistochemistry (APAAP) of paraffin tissue sections was performed using monoclonal antibodies specific for TM and PECAM. The percentage of immunopositive vessels and the intensity of immunoreactivity was assessed. Vascular endothelium and syncytiotrophoblast stained positive for TM. Immunoreactivity for TM in villous vasculature rose significantly with gestational age. Villous vessels of IUGR placentae, showed a higher expression of TM, compared to placentae of appropriately grown fetus (p < 0.05). The number of terminal villi and the number of blood vessels per intermediate villi was significantly reduced in IUGR placentae (p < 0.05). These differences reflect inadequate vascularisation and impaired angiogenesis in IUGR.     

Glucose metabolism in the human preterm and term placenta of IUGR fetuses

Many fetuses suffering from intrauterine growth restriction (IUGR) are hypoglycaemic. However, the underlying mechanisms are not well established. An increased placental glucose consumption in IUGR could impair glucose transfer across the placenta. In this study we used two different approaches to investigate glucose metabolism in preterm and term placentae of IUGR fetuses. We determined activity and protein expression of the three rate-limiting glycolytic enzymes phosphofructo kinase (PFK), pyruvate kinase (PK) and hexokinase (HXK) in a cytoplasmic fraction of homogenates of placentae obtained from IUGR and appropriate for gestational age (AGA) pregnancies. Protein expression was assessed using Western blot and enzyme activities were determined in a spectrophotometer by measuring the rate of NADH oxidation (PFK and PK) or NADP reduction (HXK) in enzyme reactions coupled to the respective enzyme. To determine the distribution of the glycolytic enzymes immunocytochemistry was performed. We also measured glucose consumption and lactate production in fresh placental villous tissue using a perifusion system. The expression of PFK, PK and HXK as well as the activity of PK and HXK was unaltered in IUGR placentae. The activity of PFK on the other hand was 32 per cent lower in IUGR placentae (n=24, P<0.05). Immunocytochemistry confirmed the distribution of the enzymes to the cytoplasm of the syncytiotrophoblast. Placental glucose consumption in IUGR [0.06+/-0.01 micromol/(min*g), n=5] was not different from AGA [0.06+/-0.005 micromol/(min*g), n=12], whereas lactate production was decreased by 28 per cent in IUGR. These results do not support the hypothesis of increased placental glucose consumption but suggest an altered glycolytic pathway in the IUGR placenta.     

Unbalanced placental expression of imprinted genes in human intrauterine growth restriction

Imprinted genes control fetal and placental growth in mice and in rare human syndromes, but the role of these genes in sporadic intrauterine growth restriction (IUGR) is less well-studied. We measured the ratio of mRNA from a maternally expressed imprinted gene, PHLDA2, to that from a paternally expressed imprinted gene, MEST, by Northern blotting in 38 IUGR-associated placentae and 75 non-IUGR placentae and found an increase in the PHLDA2/MEST mRNA ratio in IUGR (p=0.0001). Altered expression of PHLDA2 and MEST was not accompanied by changes in DNA methylation within their imprinting centers, and immunohistochemistry showed PHLDA2 protein appropriately restricted to villous and intermediate cytotrophoblast in the IUGR placentae. We next did a genome-wide survey of mRNA expression in 14 IUGR placentae with maternal vascular under-perfusion compared to 15 non-IUGR placentae using Affymetrix U133A microarrays. In this series six imprinted genes were differentially expressed by ANOVA with a Benjamini-Hochberg false discovery rate of 0.05, with increased expression of PHLDA2 and decreased expression of MEST, MEG3, GATM, GNAS and PLAGL1 in IUGR placentae. At lower significance, we found IGF2 mRNA decreased and CDKN1C mRNA increased in the IUGR cases. We confirmed the significant reduction in MEG3 non-translated RNA in IUGR placentae by Northern blotting. In addition to imprinted genes, the microarray data highlighted non-imprinted genes acting in endocrine signaling (LEP, CRH, HPGD, INHBA), tissue growth (IGF1), immune modulation (INDO, PSG-family genes), oxidative metabolism (GLRX), vascular function (AGTR1, DSCR1) and metabolite transport (SLC-family solute carriers) as differentially expressed in IUGR vs. non-IUGR placentae.     

Circulating and placental endoglin concentrations in pregnancies complicated by intrauterine growth restriction and preeclampsia

Inadequate trophoblast invasion and spiral artery remodeling leading to poor placental perfusion and hypoxia are believed to underlie preeclampsia (PE) and intrauterine growth restriction (IUGR). Recent studies implicate increased circulating endoglin as a contributor to the pathogenesis of PE. The objective of this study was to determine whether placental and circulating endoglin concentrations are altered in pregnancies complicated by intrauterine growth restricted (IUGR) infants and to address the role of hypoxia on the regulation of placental endoglin. We analyzed 10 placentas each from normal pregnant (NP), PE, and IUGR subjects. Endoglin levels were 2.5-fold higher in preeclamptic placentas compared to NP (15.4+/-2.6 versus 5.7+/-1.0, p<0.01). In contrast, endoglin levels were similar in NP and IUGR placentas (5.7+/-1.0 vs 5.9+/-1.1, p=NS). Placentas from pregnancies with both PE and IUGR exhibited endoglin levels comparable to the PE group and significantly different from normotensive pregnancies with and without IUGR pregnancies (mean 14.9+/-4.0, n=9, p=0.013). Soluble endoglin concentrations in maternal plasma were comparable in NP and IUGR, but higher in women with PE (n=10 per group, p<0.05). Despite a 2-fold increase in hypoxia inducible factor, HIF-1alpha, we did not observe endoglin upregulation in NP, PE, or IUGR placental villous explants exposed to hypoxia (2% oxygen). In contrast to PE, placental or circulating endoglin is not increased in normotensive women delivering small, asymmetrically grown (IUGR) infants at term. The placentas of women with IUGR appear to be fundamentally different from PE women with respect to endoglin, despite the proposed common pathology of deficient trophoblast invasion/spiral artery remodeling and poor placental perfusion.     

Evaluation of maternal and umbilical serum TNFα levels in preeclamptic pregnancies in the intrauterine normal and growth-restricted fetus

Objective.?The aim of this study was to carry out a comparative analysis of the maternal and umbilical cord TNFα serum levels in pregnancies complicated by severe preeclampsia with normal intrauterine fetal growth, in preeclamptic pregnancies with intrauterine growth restriction (IUGR), and in normotensive pregnant patients.

Patients and methods.?The study was carried out on eight patients with severe preeclampsia complicated by IUGR and 18 preeclamptic patients with normal intrauterine fetal growth. The control group consisted of 18 healthy normotensive patients with singleton uncomplicated pregnancies. Maternal and umbilical serum TNFα concentrations were estimated using a sandwich ELISA assay.

Results and conclusions.?Pregnant women with severe preeclampsia had significantly higher maternal and umbilical serum TNFα levels than those in the normotensive controls. Our findings and other reports indicate that TNFα may participate in the pathogenesis and sequelae of preeclampsia with and without IUGR. The results of excessive umbilical serum activity of tumor necrosis factor α (TNFα) in preeclamptic pregnancy complicated by intrauterine growth restriction (IUGR) may suggest additional changes and dysfunction of the placental–fetal unit and deterioration of placental function, leading to fetal hypotrophia in the course of preeclampsia.     

Increased production of tumor necrosis factor-alpha TNF-alpha by IUGR human placentae

OBJECTIVE: To evaluate the effect of pathological placental conditions such as intrauterine growth restriction (IUGR) or exposure to angiotensin II (AII) on TNF-alpha secretion in the vasculature of isolated human placental cotyledons. STUDY DESIGN: Isolated placental cotyledons from 10 normal and four intrauterine growth restricted fetuses were dually perfused. Perfusate samples from the fetal circulation were collected every 30 min during 120 min. TNF-alpha levels in the fetal-placental perfusate were evaluated using specific commercial ELISA kits. In three additional normal placentae, bolus injections of angiotensin II (10(-9)-10(-4) mol/l) were given into the fetal-placental circulation and perfusate samples were collected. Statistical significance of difference TNF-alpha levels between different conditions was determined by analysis of variance (ANOVA) and paired t-test. RESULTS: TNF-alpha levels were significantly higher in the perfusate of IUGR placentae as compared with normal placentae after 120 min of perfusion (mean 410+/-121 vs. 39+/-14 pg/ml, P=0.005). There was a significant dose-dependent increase in TNF-alpha levels in the placental perfusate after a bolus injection of AII 66 pg/ml with AII 10(-9) mol/l vs. 97 pg/ml with AII 10(-5) mol/l (P=0.004), respectively. CONCLUSIONS: Placental pathology related to condition IUGR might induce the secretion of proinflammatory cytokines such as TNF-alpha, which may enhance the vasoconstriction of the fetal placental vascular bed.     

Placental imbalance of Th1- and Th2-type cytokines in preeclampsia

OBJECTIVES: To characterize the changes in the level of T helper 1 (Th1)- [interleukin (IL)-2 and tumor necrosis factor (TNF)-alpha] and Th2-type cytokine (IL-10) and the ratios of Th1/Th2 (IL-2/IL-10 and TNF-alpha/IL-10) in placentae from women with preeclampsia and women with gestational hypertension. METHODS: Placental levels of IL-2, TNF-alpha, and IL-10 were determined with radioimmunoassay and Th1/Th2 ratios (IL-2/IL-10 and TNF-alpha/IL-10) calculated in the placentae from 22 women with preeclampsia, 15 women with gestational hypertension, and 32 normal term pregnant women. RESULTS: Although preeclampsia had the trend of the increase in the placental levels of IL-2 and TNF-alpha and the trend of the decrease in placental IL-10, there were not significant difference in placental levels of IL-2, IL-10, and TNF-alpha among preeclampsia, gestational hypertension, and normal pregnancy (P > 0.05 for all). Placental ratios of IL-2/IL-10 and TNF-alpha/IL-10 were significantly higher in preeclampsia than in normal pregnancy (P = 0.035 and P = 0.005, respectively). No differences of Th1/Th2 ratios were found between preeclampsia and gestational hypertension and between gestational hypertension and normal pregnancy (P > 0.05 for all). CONCLUSIONS: Alterations of placental balances of cytokines with Th1 predominance were demonstrated in preeclampsia. These associations may offer insights into the pathogenesis of preeclampsia.     

Higher Mitochondrial DNA Content in Human IUGR Placenta

IUGR has been associated to a specific placental phenotype with reduced uptake of specific nutrients. Recently, it has been hypothesized that IUGR may be determined during early gestation. This period is characterized by decidual trophoblast invasion and by intense cellular growth, replication and differentiation. Since a huge energetic availability is required during gestation, we hypothesize that mitochondria may play a crucial role in this process being the main energetic producer in the cell.The aim of this study was to investigate the role of mitochondria in IUGR pathogenesis, evaluating the number of mitochondrial DNA copies (mtDNA) in IUGR placentae compared to controls.Placental samples were collected from 50 singleton pregnancies at the time of elective caesarean section. Twenty-six pregnancies were controls with normal intrauterine growth (AGA) and 24 were studied after the in utero diagnosis of IUGR. All samples were analyzed by real-time quantitative PCR and statistical analysis was performed by non-parametric tests.The median value of mitochondrial DNA content (IQR) in AGA and IUGR placentae was significantly different (455 and 698, respectively, p = 0.004). The cell types responsible for the difference observed is unknown and it is possible that changes observed in the proportion of cell types may influence this measurement. Moreover, a significant negative relationship was observed between mtDNA and umbilical venous pO₂, with the highest levels detected in the most severe IUGR cases according to Doppler findings and to the presence of preeclampsia.These data suggest a relationship between the pathogenesis of IUGR and increased placental mtDNA copies. From our results we can speculate that increased mtDNA represents an adaptation of the metabolic placental mechanism to the calorie restriction of the fetus. Furthermore, we found that this rise was inversely related to oxygen tension in the umbilical vein. Although no specific pathogenetic role can be implied, mtDNA increases with hypoxia in placentas of IUGR.     

Chronic villitis of unknown aetiology and decidual maternal vasculopathies in sustained chronic hypertension

Ninety-six full-term placentae were examined in the study. Of the 96 pregnancies, 72 were normotensive and 24 were complicated by hypertension. Of the 24 hypertensive pregnancies, 19 patients had chronic hypertension only, and 5 had chronic hypertension with superimposed preeclampsia. The birth weights of babies were over the 25th centile of our ponderal curve in all cases. Chronic villitis of unknown aetiology (CVUE) was found in 25% of control placentae and in 26% of placentae from chronic hypertension without preeclampsia. Eighty percent of placentae of chronic hypertension with superimposed preeclampsia presented the lesion. The incidence of CVUE was significantly higher in the latter than in the other groups. The same was observed about the proportion of inflamed villi. A higher frequency of maternal vascular lesions was observed in placentae of chronic hypertension with superimposed preeclampsia. Similar incidence and severity of the above-mentioned placental lesions have been recently described in preeclamptic pregnancies. These results suggest that these lesions (CVUE and maternal vasculopathies) are related to preeclampsia and not only to maternal hypertension.