Functional analysis of the mannose-binding lectin complement pathway in normal pregnancy and preeclampsia.

Preeclampsia is a severe complication of pregnancy characterized by hypertension and proteinuria developing after midgestation. Previous studies have shown increased complement activation in normal and preeclamptic pregnancies. We aimed to investigate the role of the mannose-binding lectin pathway in the initiation of pathological complement activation observed in patients with preeclampsia. The study included 60 preeclamptic patients, 60 healthy pregnant women and 56 healthy non-pregnant women. Functional activity of the complex of mannose-binding lectin and mannose-binding lectin-associated serine protease 2 (MBL-MASP2 complex) was determined by ELISA. Circulating levels of complement components and C-reactive protein (CRP) were also measured. MBL-MASP2 activity was significantly higher in healthy pregnant than non-pregnant women. However, increased activity of the MBL-MASP2 complex in preeclamptic patients was not observed, compared to healthy pregnant women. MBL-MASP2 activity showed no relationship with either the levels of complement parameters, or with the clinical data and level of CRP in patients with preeclampsia. In conclusion, the complement system is activated with increased terminal complex formation in the third trimester of normal human pregnancy, and is further activated in preeclampsia as shown by the elevated amounts of activation markers. The activity of MBL-MASP2 is also increased in normal pregnancy, to the same level seen in preeclampsia. In our study, no relationship between MBL-MASP2 activity and extent of complement activation was observed in preeclampsia. We tentatively conclude, albeit without an evaluation of local placental concentrations, that the mannose-binding lectin pathway may play only a minor role in pathological complement activation during preeclampsia.

Activation of the complement system in normal pregnancy and preeclampsia.

The purpose of this study was to explore the role of the complement system in normal human pregnancy and preeclampsia in a comprehensive manner, measuring circulating levels of complement proteins, their activation fragments and regulatory factors, as well as those of C-reactive protein (CRP). Sixty preeclamptic patients, 60 healthy pregnant women and 59 healthy non-pregnant women were involved in this case-control study. Circulating levels of complement components and CRP were determined with ELISA, radial immunodiffusion and particle enhanced immunoturbidimetric assay. Levels of CRP, C4d, C3a, SC5b9, C3, C9 and factor H antigen were significantly higher, while those of C1-inhibitor were significantly lower in healthy pregnant than non-pregnant women. In addition, preeclamptic patients had significantly higher CRP, C4d, C3a, SC5b9 levels and significantly lower C3 concentrations as compared to healthy pregnant women. Their CRP, C4d, C3a, SC5b9, C4, C3, C9 and factor H antigen levels were significantly higher, while C1-inhibitor concentrations were significantly lower compared with healthy non-pregnant women. However, no significant difference was found in Bb and C4b-binding protein levels among the three study groups. Preeclamptic patients with fetal growth restriction had significantly higher plasma SC5b9 levels than those without IUGR. There was a relative deficiency of C1-inhibitor and C4b-binding protein, and a relative abundance of factor H both in normal pregnancy and preeclampsia. Activation of the classical or lectin pathway (C4d) showed significant positive correlation to C3 activation (C3a) both in healthy pregnant women and preeclamptic patients. However, the correlation between C3 and terminal pathway activation was dominating only in patients with preeclampsia, but not in healthy pregnant women. In conclusion, the complement system is activated through the classical and/or lectin pathways with increased terminal complex formation in the third trimester of normal human pregnancy, and further in preeclampsia, as shown by the elevated amounts of activation markers in the systemic circulation. Excessive activation of the terminal pathway is associated with fetal growth restriction in preeclamptic women. However, additional studies are required to determine the cause and consequence of systemic complement activation in this pregnancy-specific disorder.

Plasma osteopontin concentrations in preeclampsia - is there an association with endothelial injury?

UNLABELLED: Abstract Background: It has been previously reported that plasma osteopontin (OPN) concentrations are increased in cardiovascular disorders. The goal of the present study was to determine plasma OPN concentrations in healthy pregnant women and preeclamptic patients, and to investigate their relationship to the clinical characteristics of the study subjects and to markers of inflammation [C-reactive protein (CRP)], endothelial activation [von Willebrand factor antigen (VWF:Ag)] or endothelial injury (fibronectin), oxidative stress [malondialdehyde (MDA)] and trophoblast debris (cell-free fetal DNA). METHODS: Forty-four patients with preeclampsia and 44 healthy pregnant women matched for age and gestational age were involved in this case-control study. Plasma OPN concentrations were measured with ELISA. Serum CRP concentrations were determined with an autoanalyzer using the manufacturer's reagents. Plasma VWF:Ag was quantified by ELISA, while plasma fibronectin concentrations were measured by nephelometry. Plasma MDA concentrations were estimated by the thiobarbituric acid-based colorimetric assay. The amount of cell-free fetal DNA in maternal plasma was determined by quantitative real-time PCR analysis of the sex-determining region Y (SRY) gene. For statistical analyses, non-parametric methods were applied. RESULTS: Serum levels of CRP, as well as plasma concentrations of VWF:Ag, fibronectin, MDA and cell-free fetal DNA were significantly higher in preeclamptic patients than in healthy pregnant women. There was no significant difference in plasma OPN concentrations between controls and the preeclamptic group. However, preeclamptic patients with plasma fibronectin concentrations in the upper quartile had significantly higher plasma OPN concentrations than those below the 75th percentile, as well as healthy pregnant women [median (interquartile range): 9.38 (8.10-11.99) vs. 7.54 (6.31-9.40) and 7.40 (6.51-8.80) ng/mL, respectively, p<0.05 for both]. Furthermore, in preeclamptic patients, plasma OPN concentrations showed a significant positive linear association with plasma fibronectin (Spearman R=0.38, standardized regression coefficient (beta)=0.41, p<0.05 for both). CONCLUSIONS: Plasma OPN concentrations are increased in preeclamptic patients with extensive endothelial injury. However, further studies are warranted to explore the relationship between OPN and endothelial damage. Clin Chem Lab Med 2010;48:181-7.

Preeclampsia is associated with decreased serum alpha(2)-HS glycoprotein (fetuin-A) concentration.

The purpose of this study was to determine serum alpha(2)-HS glycoprotein (AHSG) concentration and its diagnostic accuracy in preeclampsia. In this case-control study, the serum C-reactive protein (CRP) and AHSG levels were measured in 93 preeclamptic patients and in 127 healthy pregnant women by immunoturbidimetry and radial immunodiffusion. The serum CRP levels were significantly higher, whereas the serum AHSG concentrations were significantly lower in the preeclamptic group than in the control group (median (25th to 75th percentile), CRP: 6.71 mg l(-1) (2.76-12.69) vs. 3.38 mg l(-1) (1.69-7.27), respectively; AHSG: 660 microg ml(-1) (612-768) vs. 744 microg ml(-1) (660-816), respectively; P<0.001 for both). In preeclamptic patients, the serum AHSG concentrations showed significant inverse correlations with systolic blood pressure and serum CRP levels. A low serum AHSG level (< or = 720 microg ml(-1)) was significantly associated with preeclampsia (adjusted odds ratio (95% confidence interval): 3.69 (1.82-7.51); P<0.001). According to the receiver operating characteristic curves, the measurement of serum AHSG concentrations was as accurate as that of serum CRP levels to detect preeclampsia. In conclusion, serum AHSG concentration is decreased and reflects-at least partly-systemic inflammation in preeclampsia.

Overrepresentation of BclI polymorphism of the glucocorticoid receptor gene in pregnant women with HELLP syndrome.

BACKGROUND: Because the pathological background of preeclampsia and its severe variant, HELLP syndrome (hemolysis, elevated liver enzymes and low platelet counts) appears to involve a pathological maternal-fetal immune adaptation, we examined whether any association could exist between these disorders and polymorphisms of the glucocorticoid receptor (GR) gene. METHODS: The BclI, N363S, and ER22/23EK polymorphisms of the GR gene were determined in 300 healthy pregnant women, 150 pregnant women with severe preeclampsia including 17 pregnant women with HELLP syndrome. RESULTS: There were no significant differences in carrier and allelic frequencies of the N363S and ER22/23EK polymorphisms between healthy pregnant women and those with severe preeclampsia. However, the allelic and carrier frequencies of the BclI polymorphism were significantly higher in women with HELLP syndrome compared to healthy pregnant women (p=0.004; Odds ratio, 2.89) and to those with severe preeclampsia (p=0.013; Odds ratio, 2.56). CONCLUSION: Our observations suggest that among pregnant women, the BclI polymorphism is associated with the development of HELLP syndrome, but not that of severe preeclampsia. Since preeclampsia and HELLP syndrome develop exclusively in human, it seems particularly interesting that alignment analysis of DNA sequences obtained from databases indicated the absence of the BclI site in 6 animal vertebral species.

Circulating anti-heat-shock-protein antibodies in normal pregnancy and preeclampsia.

It has been previously reported that circulating anti-heat-shock-protein (Hsp) antibody levels are elevated in cardiovascular disorders. The aim of the present study was to determine circulating antihuman Hsp60, antimycobacterial Hsp65, and antihuman Hsp70 antibody levels in healthy pregnant women and preeclamptic patients and to investigate their relationship to the clinical characteristics of the study subjects, as well as to the markers of inflammation (C-reactive protein (CRP)), endothelial activation (von Willebrand factor antigen), or endothelial injury (fibronectin), oxidative stress (malondialdehyde) and to serum Hsp70 levels. Ninety-three preeclamptic patients and 127 normotensive healthy pregnant women were involved in this case control study. Serum anti-Hsp60, anti-Hsp65, anti-Hsp70, and Hsp70 levels were measured with enzyme-linked immunosorbent assay (ELISA). Serum CRP levels were determined by an autoanalyzer using the manufacturer's kit. Plasma von Willebrand factor antigen levels were quantified by ELISA, while plasma fibronectin concentration by nephelometry. Plasma malondialdehyde levels were measured by the thiobarbituric-acid-based colorimetric assay. For statistical analyses, nonparametric methods were applied. Anti-Hsp60, anti-Hsp65, and anti-Hsp70 antibodies were detected in all of our serum samples. There were no significant differences in serum anti-Hsp60, anti-Hsp65, and anti-Hsp70 antibody levels between the control and preeclamptic groups. Serum levels of Hsp70 and CRP, as well as plasma levels of VWF antigen, fibronectin, and malondialdehyde, were significantly higher in preeclamptic patients than in normotensive healthy pregnant women. Serum anti-Hsp60 antibody levels showed significant correlations with serum anti-Hsp65 antibody levels both in the control and the preeclamptic groups (Spearman R = 0.55 and 0.59; p < 0.001, respectively). However, no other relationship was found between clinical features (maternal age, smoking status, parity, body mass index, gestational age at blood draw, systolic and diastolic blood pressure, gestational age at delivery, and fetal birth weight) and measured laboratory parameters of the study subjects and serum anti-Hsp antibody levels in either study group. In conclusion, anti-Hsp60 and anti-Hsp70 antibodies as naturally occurring autoantibodies are present in the peripheral circulation of healthy pregnant women. Nevertheless, humoral immunity against heat shock proteins was not associated with preeclampsia. Further studies are warranted to explore the role of heat shock proteins and immune reactivity to them in the immunobiology of normal pregnancy and preeclampsia.

Increased plasma von Willebrand factor antigen levels but normal von Willebrand factor cleaving protease (ADAMTS13) activity in preeclampsia.

The activity of ADAMTS13, the von Willebrand factor (VWF) cleaving protease is low in several conditions, including HELLP (haemolysis, elevated liver enzymes, and low platelet count) syndrome. As HELLP syndrome develops in most cases on the basis of preeclampsia, our aim was to determine whether plasma ADAMTS13 activity is decreased in preeclampsia. Sixty-seven preeclamptic patients, 70 healthy pregnant women and 59 healthy non-pregnant women were involved in this case-control study. Plasma ADAMTS13 activity was determined with the FRETS-VWF73 assay, while VWF antigen (VWF:Ag) levels with an enzyme-linked immunosorbent assay. The multimeric pattern of VWF was analyzed by SDS-agarose gel electrophoresis. There was no significant difference in plasma ADAMTS13 activity between the preeclamptic and the healthy pregnant and non-pregnant groups (median [25-75 percentile]: 98.8 [76.5-112.8] %, 96.3 [85.6-116.2] % and 91.6 [78.5-104.4] %, respectively; p > 0.05). However, plasma VWF:Ag levels were significantly higher in preeclamptic patients than in healthy pregnant and non-pregnant women (187.1 [145.6-243.1] % versus 129.3 [105.1-182.8] % and 70.0 [60.2-87.3] %, respectively; p < 0.001). The multimeric pattern of VWF was normal in each group. Primiparas had lower plasma ADAMTS13 activity than multi-paras (92.6 [75.8-110.6] % versus 104.2 [92.1-120.8] %; p = 0.011). No other relationship was found between clinical characteristics, laboratory parameters and plasma ADAMTS13 activity in either study group. In conclusion, plasma ADAMTS13 activity is normal in preeclampsia despite the increased VWF:Ag levels. However, further studies are needed to determine whether a decrease in plasma ADAMTS13 activity could predispose preeclamptic patients to develop HELLP syndrome.

Association between estrogen receptor alpha (ESR1) gene polymorphisms and severe preeclampsia.

Associations have been reported between estrogen receptor alpha (ESR1) gene polymorphisms and various pathological conditions, including cardiovascular diseases. Our aim was to investigate whether two polymorphisms of the ESR1 gene (ESR1 c.454 -397T>C: PvuII restriction site and c.454 -351A>G: XbaI restriction site) are associated with preeclampsia. In a case-control study, we analyzed blood samples from 119 severely preeclamptic patients and 103 normotensive, healthy pregnant women using the polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) method. All of the women were Caucasian. There was no association between severe preeclampsia and the PvuII and XbaI ESR1 gene polymorphisms separately. However, with the simultaneous carriage of both polymorphisms, the TT/AA genotype combination was significantly more frequent in severely preeclamptic patients than in healthy control subjects (24.4% vs. 9.7%, p=0.003), whereas the TT/AG combination was significantly less frequent in the severely preeclamptic group than in the control group (5.0% vs. 18.4%, p=0.002). According to the haplotype estimation, the homozygous T-A haplotype carriers had an increased risk of severe preeclampsia independent of maternal age, prepregnancy BMI, primiparity and smoking status (adjusted odds ratio [OR]: 4.36, 95% confidence interval [CI]: 1.65-11.53). The GG genotype of the XbaI polymorphism was associated with a lower risk of fetal growth restriction in patients with severe preeclampsia (OR: 0.23, 95% CI: 0.07-0.73). In conclusion, the homozygous T-A haplotype carriers of ESR1 PvuII and XbaI polymorphisms showed an increased risk of severe preeclampsia. In addition, the GG genotype of the XbaI polymorphism decreased the risk of fetal growth restriction in severely preeclamptic patients.

Family history of early-onset cardiovascular disorders is associated with a higher risk of severe preeclampsia.

AIM: The aim was to evaluate familial early-onset cardiovascular disorders as potential risk factors for severe preeclampsia. STUDY DESIGN: A case-control study was carried out by interviewing 162 primiparous severely preeclamptic women and 521 primiparous healthy control patients after delivery to determine the frequency of cardiovascular disorders (chronic hypertension, myocardial infarction, stroke) developed before the age of 50 among their parents. The chi2-test was utilized to estimate odds ratios (OR) and 95% confidence intervals (95% CI). The association was adjusted for pre-pregnancy body mass index, maternal age, and smoking habits before pregnancy using logistic regression analysis. RESULTS: Maternal and paternal early-onset chronic hypertension (adjusted OR: 3.84, 95% CI: 2.25-6.54; and adjusted OR: 3.26, 95% CI: 1.76-6.05) as well as paternal early-onset myocardial infarction (adjusted OR: 3.33; 95% CI: 1.51-7.32) were independent risk factors for severe preeclampsia. Early-onset stroke affected only the fathers of severely preeclamptic patients. Among the severely preeclamptic patients a positive family history of cardiovascular disorders developed before the age of 50 increased the risk of early-onset preeclampsia (developing before the 32nd gestational week) by 5.05-fold (95% CI: 3.08-8.31) compared with the control group. CONCLUSION: Our results suggest that the presence of familial early-onset cardiovascular disorders is a predisposing factor for severe preeclampsia.