Increased urinary excretion of nephrin, podocalyxin, and βig-h3 in women with preeclampsia

Emerging evidence has shown that podocyte injury and reduced specific podocyte protein expressions contribute to proteinuria in preeclampsia. We collected urine specimens from women with preeclampsia to study whether podocyte-specific protein shedding is associated with renal barrier dysfunction. Urine specimens from women with normal pregnancies and from pregnant women complicated by chronic hypertension were used for comparison. We determined soluble podocyte slit protein nephrin levels in the urine specimens. Podocalyxin, βig-h3, and VEGF concentrations were also measured. We found that nephrin and podocalyxin were barely detectable in the urine specimens from normal pregnant women and from women with chronic hypertension. In preeclampsia, urinary nephrin and podocalyxin concentrations were significantly increased and highly correlated to each other, r(2) = 0.595. Nephrin and podocalyxin were also correlated with urine protein concentrations. βig-h3 was detected in the urine specimens from women with preeclampsia, and it is highly correlated with nephrin and podocalyxin concentrations in preeclampsia. βig-h3 was undetectable in normal pregnancy and pregnancy complicated by chronic hypertension. Elevated VEGF levels were also found in women with preeclampsia compared with those of normal pregnancy and pregnancy complicated by chronic hypertension. These results provide strong evidence that podocyte protein shedding occurs in preeclampsia, and their levels are associated with proteinuria. The finding of urinary βig-h3 excretion in preeclampsia suggests that increased transforming growth factor activity might also be involved in the kidney lesion in this pregnancy disorder.     

Serum concentrations of immunoglobulins and acute phase proteins in Nigerian women with preeclampsia

Preeclampsia is a pregnancy-specific condition that increases maternal and infant mortality and morbidity. It is diagnosed based on a triad of hypertension, significant proteinuria and rapidly increasing edema during gestation. The factors that initiate preeclampsia are unknown and still a subject of intense clinical research. The objective of this study is to provide additional immunological information about preeclampsia. To achieve this, humoral immunochemical parameters such as three classes of immunoglobulin (IgA, IgG and IgM) and three acute phase proteins (alpha 2-macroglobulin, haptoglobin and transferrin) were measured by single radial immunodifussion method in 32 pregnant women with preeclampsia, 36 pregnant women without preeclampsia and 24 non-pregnant women (controls). Total protein in the urine was also determined by spectrophotometric method. In women with preeclampsia, the levels of IgG, IgA, transferrin and alpha 2-macroglobulin were significantly reduced compared with subjects with normal pregnancy, but the level of haptoglobin was significantly raised in preeclampsia compared with women having normal pregnancy. Urinary total protein and IgG were significantly raised in Nigerian women with preeclampsia compared with non-pregnant controls. There were significant negative correlations between serum IgG, IgA and urinary protein. The possible involvement of immunoglobulins and acute phase proteins in preeclampsia is discussed.     

Preeclampsia: a bioinformatics approach through protein-protein interaction networks analysis

ABSTRACT: BACKGROUND: In this study we explored preeclampsia through a bioinformatics approach. We create a comprehensive genes/proteins dataset by the analysis of both public proteomic data and text mining of public scientific literature. From this dataset the associated protein-protein interaction network has been obtained. Several indexes of centrality have been explored for hubs detection as well as the enrichment statistical analysis of metabolic pathway and disease. RESULTS: We confirmed the well known relationship between preeclampsia and cardiovascular diseases but also identified statistically significant relationships with respect to cancer and aging. Moreover, significant metabolic pathways such as apoptosis, cancer and cytokine-cytokine receptor interaction have also been identified by enrichment analysis. We obtained FLT1, VEGFA, FN1, F2 and PGF genes with the highest scores by hubs analysis; however, we also found other genes as PDIA3, LYN, SH2B2 and NDRG1 with high scores. CONCLUSIONS: The applied methodology not only led to the identification of well known genes related to preeclampsia but also to propose new candidates poorly explored or completely unknown in the pathogenesis of preeclampsia, which eventually need to be validated experimentally. Moreover, new possible connections were detected between preeclampsia and other diseases that could open new areas of research. More must be done in this area to resolve the identification of unknown interactions of proteins/genes and also for a better integration of metabolic pathways and diseases.     

Expression of the Na(+)-H+ exchanger isoform-1 and cyclooxygenases in human placentas: their implications in preeclampsia.

The sodium hydrogen exchanger isoform, NHE-1 plays an important role in electrolyte and water homeostasis. These functions are compromised in pregnancies complicated with preeclampsia. At present it is not known whether NHE-1 expression is altered during preeclampsia. In the present study the placental level of NHE-1 protein was measured using immunoblotting. Since prostaglandins regulate the secretory and absorptive functions, the levels of prostaglandin E-2 as well as the expression of cyclooxygenase-1 and -2 were also estimated. The amount of NHE-1 protein and cyclooxygenase-2 was reduced in preeclamptic placentas, whereas the level of cyclooxygenase-1 remained unaltered. In contrast, prostaglandin E-2 concentration was higher in preeclampsia. Suppression of NHE-1 might render the placenta with impaired uptake of water and electrolytes and therefore may be involved in the pathogenesis of preeclampsia. While prostaglandin E-2 may play a role in preeclampsia, these findings discount the induction of cyclooxygenase-genes for this increase.     

Association of angiotensin-converting enzyme insertion-deletion polymorphism with preeclampsia

The aim of this study was to determine if insertion-deletion polymorphism of angiotensin-converting enzyme is a risk factor for the development of preeclampsia. Sixty women with preeclampsia and 50 normotensive pregnant women were included in this study. Preeclampsia was defined as blood pressure >140/90 mmHg in a previously normotensive women with proteinuria >300 mg/L in a 24-hours. Twelve women also had preeclampsia in previous pregnancy. The genotyping of polymorphism in the intron 16 of the angiotensin-converting enzyme was performed by the polymerase chain reaction followed by the agarose electrophoresis. The patients were divided into three groups according to the presence (I) or absence (D) of insertional polymorphism (II, ID, and DD). Genotype distribution and allele frequencies were compared by Mantel-Haenszel chi2 testing. The frequency of DD genotype was not significantly higher in women with preeclampsia (26/60) than in the control group (14/50, p=0.096). The D allele frequency was significantly higher in 17 women with preeclampsias who required delivery before 34 weeks of pregnancy (0.735), than in 43 women in whom obstetric complications took place after 34 weeks of pregnancy (0.56, p=0.036). The D allele frequency was 0.83 in women having recurrent preeclampsia, i.e. significantly higher compared with women, who were for the first time, experienced preeclampsia (0.57, p=0.013). This study showed a significantly positive association between D allele frequency and risk of recurrent preeclampsia and preterm delivery before 34 weeks of pregnancy. The deletion genotype could be an important contributing factor for an early onset and recurrent preeclampsia.     

Pro-angiogenic therapeutics for preeclampsia

Preeclampsia is a pregnancy-induced hypertensive disorder resulting from abnormal placentation, which causes factors such as sFlt-1 to be released into the maternal circulation. Though anti-hypertensive drugs and magnesium sulfate can be given in an effort to moderate symptoms, the syndrome is not well controlled. A hallmark characteristic of preeclampsia, especially early-onset preeclampsia, is angiogenic imbalance resulting from an inappropriately upregulated sFlt-1 acting as a decoy receptor binding vascular endothelial growth factor (VEGF) and placental growth factor (PlGF), reducing their bioavailability. Administration of sFlt-1 leads to a preeclamptic phenotype, and several models of preeclampsia also have elevated levels of plasma sFlt-1, demonstrating its role in driving the progression of this disease. Treatment with either VEGF or PlGF has been effective in attenuating hypertension and proteinuria in multiple models of preeclampsia. VEGF, however, may have overdose toxicity risks that have not been observed in PlGF treatment, suggesting that PlGF is a potentially safer therapeutic option. This review discusses angiogenic balance as it relates to preeclampsia and the studies which have been performed in order to alleviate the imbalance driving the maternal syndrome.     

Potential interaction of brain natriuretic peptide with hyperadiponectinemia in preeclampsia

Adiponectin was reported recently to have roles in the pathophysiology of preeclampsia. Moreover, elevation of adiponectin and brain natriuretic peptide (BNP) has been observed in preeclampsia. We examined the possible links between adiponectin and BNP in the pathophysiology of preeclampsia. We performed a cross-sectional study in 56 preeclampsia patients and 56 controls matched for gestational age and body mass index. The BNP, leptin, and adiponectin levels were measured by ELISA, and their mRNA expressions were evaluated in omental adipose tissue by real-time PCR. The effects of BNP on adiponectin and leptin mRNA expression and secretion were investigated in primary cultures of adipocytes from obese and normal-weight women. The BNP, adiponectin, and leptin levels were significantly higher in preeclampsia patients compared with controls. The adiponectin level was increased significantly in normal-weight preeclampsia patients compared with overweight preeclampsia patients. Adiponectin mRNA expression was increased significantly in adipose tissues of preeclampsia patients compared with controls and was also increased significantly in normal-weight preeclampsia patients compared with overweight preeclampsia patients, whereas leptin was not. BNP and adiponectin showed significant positive correlations in both normal-weight and overweight preeclampsia patients. BNP had a significantly weaker effect on adiponectin in overweight compared with normal-weight preeclampsia patients. Moreover, BNP had a weaker effect on adiponectin production in adipocytes from overweight women compared with adipocytes from normal-weight women using primary culture of human adipocytes. These data suggested that BNP may play a role in hyperadiponectinemia of preeclampsia patients. The weaker effect of BNP on adiponectin production may participate in the pathophysiology of overweight preeclampsia patients.     

LncRNAs Expression in Preeclampsia Placenta Reveals the Potential Role of LncRNAs Contributing to Preeclampsia Pathogenesis

Background

Long non-coding RNAs (lncRNAs) are an important class of pervasive genes involved in a variety of biological functions. They are aberrantly expressed in many types of diseases. In this study, we aimed to investigate the lncRNA profiles in preeclampsia. Preeclampsia has been observed in patients with molar pregnancy where a fetus is absent, which demonstrate that the placenta is sufficient to cause this condition. Thus, we analyzed the lncRNA profiles in preeclampsia placentas.

Methodology/Principal Findings

In this study, we described the lncRNA profiles in six preeclampsia placentas (T) and five normal pregnancy placentas (N) using microarray. With abundant and varied probes accounting for 33,045 LncRNAs in our microarray, 28,443 lncRNAs that were expressed at a specific level were detected. From the data, we found 738 lncRNAs that were differentially expressed (≥1.5-fold-change) among preeclampsia placentas compared with controls. Coding-non-coding gene co-expression networks (CNC network) were constructed based on the correlation analysis between the differentially expressed lncRNAs and mRNAs. According to the CNC network and GO analysis of differentially expressed lncRNAs/mRNAs, we selected three lncRNAs to analyze the relationship between lncRNAs and preeclampsia. LOC391533, LOC284100, and CEACAMP8 were evaluated using qPCR in 40 preeclampsia placentas and 40 controls. These results revealed that three lncRNAs were aberrantly expressed in preeclampsia placentas compared with controls.

Conclusions/Significance

Our study is the first study to determine the genome-wide lncRNAs expression patterns in preeclampsia placenta using microarray. These results revealed that clusters of lncRNAs were aberrantly expressed in preeclampsia placenta compared with controls, which indicated that lncRNAs differentially expressed in preeclampsia placenta might play a partial or key role in preeclampsia development. Misregulation of LOC391533, LOC284100, and CEACAMP8 might contribute to the mechanism underlying preeclampsia. Taken together, this study may provide potential targets for the future treatment of preeclampsia and novel insights into preeclampsia biology.     

The role of the immune system in preeclampsia

Recent data demonstrate that an altered immune response may play a key role in the development of preeclampsia. Some epidemiological findings and animal models support this idea. In this article, we review the innate immune system and adaptive immune system in preeclampsia and discuss the pathophysiology of preeclampsia from an immunological viewpoint. The most characteristic immunological finding in preeclampsia is the activation of both the innate and adaptive immune system. Activated neutrophils, monocytes, and NK cells initiate inflammation which induce endothelial dysfunction, and activated T cells may support inadequate tolerance during pregnancy. The cytokine profile in preeclampsia shows that the production of type 1 cytokines, which induce inflammation, is dominant while the production of type 2 cytokines, which regulates inflammation, is suppressed. Furthermore, the immunoregulatory system is down-regulated in preeclampsia and persistent inflammation reduces regulatory T cell function. Therefore, systematical immunoactivation may be one cause of preeclampsia.     

Protein Profiling of Preeclampsia Placental Tissues

Preeclampsia is a multi-system disorder involved in pregnancy without an effective treatment except delivery. The precise pathogenesis of this complicated disorder is still not completely understood. The objective of this study is to evaluate the alterations of protein expression and phosphorylations that are important in regulating placental cell function in preterm and term preeclampsia. Using the Protein Pathway Array, 38 proteins in placental tissues were found to be differentially expressed between preterm preeclampsia and gestational age matched control, while 25 proteins were found to be expressed differentially between term preeclampsia and matched controls. Among these proteins, 16 proteins and their associated signaling pathways overlapped between preterm and term preeclampsia, suggesting the common pathogenesis of two subsets of disease. On the other hand, many proteins are uniquely altered in either preterm or term preeclampsia and correlated with severity of clinical symptoms and outcomes, therefore, providing molecular basis for these two subsets of preeclampsia. Furthermore, the expression levels of some of these proteins correlated with neonatal small for gestational age (PAI-1 and PAPP-A) and adverse outcomes (Flt-1) in women with preterm preeclampsia. These proteins could potentially be used as candidate biomarkers for predicting outcomes of preeclampsia.     

Plasma from pre‐eclamptic patients induces the expression of the anti‐angiogenic miR‐195‐5p in endothelial cells

We examined the effect of plasma incubation from preeclampsia pregnant on the antiangiogenic miR‐195‐5p expression. Higher miR‐195‐5p expression was found in cultures incubated with preeclampsia plasma compared to those incubated with healthy pregnant plasma. Next, as VEGF is a target of miR‐195‐5p we have quantified its expression by real‐time qPCR and ELISA. We found reduced VEGF levels in culture incubated with preeclampsia plasma. Therefore, we have concluded that the higher expression of miR‐195‐5p in endothelial cell cultures incubated with preeclampsia plasma may contribute to decreased expression of VEGFA (gene and protein) and increased antiangiogenic status in preeclampsia. Therefore, this miR may be an important target in preeclampsia.     

Preeclampsia‐associated stresses activate Gadd45a signaling and sFlt‐1 in placental explants

Accumulating evidence suggests that placental stresses during pregnancy can play an important role in the pathogenesis of preeclampsia. A common signal pathway that senses and converts placental stresses into intracellular stress response may be contributing to this pathology. Based on our previous findings, we extended our investigation to establish that Gadd45a stress signaling regulates sFlt‐1 levels, particularly in placenta, when exposed to various preeclampsia‐associated stresses including AT‐1 receptor agonist (Angiotensin II), hypoxia, and inflammatory cytokines. Using a placental explant model, we found that Gadd45a was induced in response to all the preeclampsia stresses stated above. Although stress induced Gadd45a was associated with the activation of its downstream effectors phospho‐p38 and phospho‐JNK, the subsequent regulation of sFlt‐1 levels occurred through either one of these effectors, but not both. These observations indicate that Gadd45a signaling may work as a hub connecting placental stresses and the pathogenesis of preeclampsia. It also provides evidence to justify testing the role of Gadd45 in the etiology of preeclampsia using in vivo mouse (i.e., Gadd45a null mice) models. J. Cell. Physiol. 228: 362–370, 2013. © 2012 Wiley Periodicals, Inc.     

Plasma l-arginine is markedly reduced in pregnant women affected by preeclampsia

The objective of this study was to determine the concentration of free l-amino acids and in particular of l-arginine in the plasma of pregnant women affected by preeclampsia compared to healthy pregnant women in order to know if an alteration in the concentrations of these amino acids occurs in preeclamspia. Twelve pregnant women affected by preeclampsia and twelve pregnant control women, ages 28–35 years old and at the 35–36 weeks of pregnancy were studied. The blood analysis of free amino acids was carried out by using a high performance liquid chromatographic (HPLC) fluorometric method and OPA-NAC as derivatizing agent for the amino acid determination. In the blood of women affected by preeclampsia l-arginine is markedly reduced compared to controls (about five-fold lower, P<0.01). The other amino acids also are significantly reduced, but to lesser extents (about 1.5 times lower, P<0.05). Thus, the determination of l-arginine in the blood of pregnant women could potentially constitute an additional marker for the early diagnosis of preeclampsia.     

Increased expression of high mobility group box 1 (HMGB1) in the cytoplasm of placental syncytiotrophoblast from preeclamptic placentae

BackgroundPreeclampsia is a pregnancy-specific disorder characterised by an inappropriate maternal inflammatory response during pregnancy. High mobility group box 1 (HMGB1) was originally characterised as a nuclear protein but when released into the extracellular environment following necrotic cell death, it is proinflammatory. HMGB1 is expressed in the syncytiotrophoblast of human placenta. Higher levels of uric acid are reported in preeclampsia. The aim of this study was to investigate whether the expression of HMGB1differed between early onset and late onset preeclampsia or severe and mild preeclampsia and whether its expression correlated with the levels of uric acid.Methods74 preeclamptic placentae and 110 normotensive placentae were included in this study. The levels of uric acid in women with preeclampsia were measured. The expression of HMGB1 in preeclamptic placentae or in first trimester and term placentae that had been treated with uric acid was measured.ResultsHMGB1 was expressed predominantly in the syncytiotrophoblast of the placenta and the expression of HMGB1 in the cytoplasm of the syncytiotrophoblast was significantly increased in both severe preeclampsia and early onset preeclampsia compared to normotensive pregnancies. The circulating levels of uric acid were significantly increased in preeclampsia and correlated with the expression of HMGB1. Increased levels of HMGB1 were significantly correlated with the severity and the time of onset of preeclampsia, but pathologic levels of uric acid did not increase the expression of HMGB1.ConclusionOur data provides a better understanding of the function of HMGB1, a danger molecule in the pathogenesis of preeclampsia.     

Erythrocyte omega-3, omega-6 and trans fatty acids in relation to risk of preeclampsia among women delivering at Harare Maternity Hospital, Zimbabwe

We sought to examine the association between maternal erythrocyte omega-3, omega-6 and trans fatty acids and risk of preeclampsia. We conducted a case-control study of 170 women with proteinuric, pregnancy-induced hypertension and 185 normotensive pregnant women who delivered at Harare Maternity Hospital, Harare, Zimbabwe. We measured erythrocyte omega-3, omega-6 and trans fatty acid as the percentage of total fatty acids using gas chromatography. After multivariate adjustment for confounding factors, women in the highest quartile group for total omega-3 fatty acids compared with women in the lowest quartile experienced a 14% reduction in risk of preeclampsia (odds ratio 0.86, 95% confidence interval 0.45 to 1.63). For total omega-6 fatty acids the odds ratio was 0.46 (95% confidence interval 0.23 to 0.92), although there was suggestion of a slight increase in risk of preeclampsia associated with high levels of arachidonic acid. Among women in the highest quartile for arachidonic acid the odds ratio was 1.29 (95% confidence interval 0.66 to 2.54). A strong statistically significant positive association of diunsaturated fatty acids with a trans double bond with risk of preeclampsia was observed. Women in the upper quartile of 9-cis 12-trans octadecanoic acid (C(18:2n6ct)) compared with those in the lowest quartile experienced a 3-fold higher risk of preeclampsia (odds ratio = 3.02, 95% confidence interval 1.41 to 6.45). Among women in the highest quartile for 9-trans 12-cis octadecanoic acid (C(18:2n6tc)) the odds ratio was 3.32 (95% confidence interval 1.55 to 7.13). Monounsaturated trans fatty acids were also positively associated with the risk of preeclampsia, although of much reduced magnitude. We observed a strong positive association of trans fatty acids, particularly diunsaturated trans fatty acids, with the risk of preeclampsia. We found little support for the hypothesized inverse association between omega-3 fatty acids and preeclampsia risk in this population. Polyunsaturated fatty acids, particularly omega-3 fatty acids, were comparatively lower in Zimbabwean than among US pregnant women. Given the limited inter-person variation in omega-3 fatty acids among Zimbabwean women, our sample size may be too small to adequately assess the relation in this population.     

Plasma -arginine is markedly reduced in pregnant women affected by preeclampsia

The objective of this study was to determine the concentration of free -amino acids and in particular of -arginine in the plasma of pregnant women affected by preeclampsia compared to healthy pregnant women in order to know if an alteration in the concentrations of these amino acids occurs in preeclamspia. Twelve pregnant women affected by preeclampsia and twelve pregnant control women, ages 28–35 years old and at the 35–36 weeks of pregnancy were studied. The blood analysis of free amino acids was carried out by using a high performance liquid chromatographic (HPLC) fluorometric method and OPA-NAC as derivatizing agent for the amino acid determination. In the blood of women affected by preeclampsia -arginine is markedly reduced compared to controls (about five-fold lower, P<0.01). The other amino acids also are significantly reduced, but to lesser extents (about 1.5 times lower, P<0.05). Thus, the determination of -arginine in the blood of pregnant women could potentially constitute an additional marker for the early diagnosis of preeclampsia.     

Preeclampsia inactivates glucose-6-phosphate dehydrogenase and impairs the redox status of erythrocytes and fetal endothelial cells

Inactivation of glucose-6-phosphate dehydrogenase (G6PD) may contribute to vascular dysfunction in preeclampsia, and oxidative stress has been implicated in the pathogenesis of this disease. We have compared the susceptibility of erythrocytes and human umbilical vein endothelial cells (HUVEC) to oxidative stress in women with normotensive or preeclamptic pregnancies. The redox status of erythrocytes was also correlated with neutrophil-mediated superoxide (O₂⁻) production in women recruited to the “Vitamins in Preeclampsia” (VIP) trial. Erythrocytes and HUVEC from women with preeclampsia demonstrated impaired redox regulation and diminished response to glucose, detectable at 14–20 weeks gestation prior to onset of the clinical disease. Hexokinase and G6PD activities were decreased in erythrocytes and G6PD activity was decreased in HUVEC from preeclamptic pregnancies. Phorbol-ester-stimulated O₂⁻ was enhanced in preeclamptic neutrophils. Impaired redox regulation in erythrocytes and HUVEC in preeclampsia may be due to diminished hexokinase and G6PD activities resulting from increased release of reactive oxygen species from activated neutrophils. Our findings provide the first evidence that decreased G6PD activity in preeclampsia is associated with impaired redox regulation in erythrocytes and fetal endothelial cells. The deficiency in G6PD in preeclampsia potentially accounts for the lack of protection against oxidative stress afforded by antioxidant vitamin C/E supplementation in the VIP trial.     

Pathophysiology of hypertension during preeclampsia: linking placental ischemia with endothelial dysfunction

Studies over the last decade have provided exciting new insights into potential mechanisms underlying the pathogenesis of preeclampsia. The initiating event in preeclampsia is generally regarded to be placental ischemia/hypoxia, which in turn results in the elaboration of a variety of factors from the placenta that generates profound effects on the cardiovascular system. This host of molecules includes factors such as soluble fms-like tyrosine kinase-1, the angiotensin II type 1 receptor autoantibody, and cytokines such as tumor necrosis factor-alpha, which generate widespread dysfunction of the maternal vascular endothelium. This dysfunction manifests as enhanced formation of factors such as endothelin, reactive oxygen species, and augmented vascular sensitivity to angiotensin II. Alternatively, the preeclampsia syndrome may also be evidenced as decreased formation of vasodilators such as nitric oxide and prostacyclin. Taken together, these alterations cause hypertension by impairing renal pressure natriuresis and increasing total peripheral resistance. Moreover, the quantitative importance of the various endothelial and humoral factors that mediate vasoconstriction and elevation of arterial pressure during preeclampsia remains to be elucidated. Thus identifying the connection between placental ischemia/hypoxia and maternal cardiovascular abnormalities in hopes of revealing potential therapeutic regimens remains an important area of investigation and will be the focus of this review.