Serum and plasma determination of angiogenic and anti-angiogenic factors yield different results: The need for standardization in clinical practice

Objective.?The importance of an anti-angiogenic state as a mechanism of disease in preeclampsia is now recognized. Assays for the determination of concentrations of soluble vascular endothelial growth factor receptor (sVEGFR)-1, sVEGFR-2, placental growth factor (PlGF) and soluble endoglin (sEng) have been developed for research and clinical laboratories. A key question is whether these factors should be measured in plasma or serum. The purpose of this study was to determine if there are differences in the concentrations of these analytes between plasma and serum in normal pregnancy and in preeclampsia.

Methods.?Samples of maternal blood were obtained by venipuncture and collected in EDTA (lavender top) and serum collection tubes (red top). A standard laboratory procedure was implemented for the centrifugation, aliquoting and storage of samples. Plasma and serum from 70 women with normal pregnancies and 34 patients with preeclampsia were assayed for sVEGFR-1, sVEGFR-2, PlGF and sEng by ELISA. Nonparametric paired tests were used for analyses.

Results.?A significant difference between plasma and serum concentration was observed for sVEGFR-1 and sVEGFR-2 in normal pregnancy, and for sVEGFR-1, sVEGFR-2, PlGF and sEng in women with preeclampsia.

Conclusion.?The concentrations of sVEGFR-1, sVEGFR-2, PlGF and sEng when measured in maternal plasma and in serum are different. Therefore, the matrix used for the assay (serum versus plasma) needs to be considered when selecting thresholds for predictive studies and interpreting the growing body of literature on this subject.     

The use of angiogenic biomarkers in maternal blood to identify which SGA fetuses will require a preterm delivery and mothers who will develop pre-eclampsia

Objective: To determine (1) whether maternal plasma concentrations of angiogenic and anti-angiogenic factors can predict which mothers diagnosed with “suspected small for gestational age fetuses (sSGA)” will develop pre-eclampsia (PE) or require an indicated early preterm delivery (≤?34 weeks of gestation); and (2) whether risk assessment performance is improved using these proteins in addition to clinical factors and Doppler parameters.

Methods: This prospective cohort study included women with singleton pregnancies diagnosed with sSGA (estimated fetal weight <10th percentile) between 24 and 34 weeks of gestation (n?=?314). Plasma concentrations of soluble vascular endothelial growth factor receptor-1 (sVEGFR-1), soluble endoglin (sEng) and placental growth factor (PlGF) were determined in maternal blood obtained at the time of diagnosis. Doppler velocimetry of the umbilical (Umb) and uterine (UT) arteries was performed. The outcomes were (1) subsequent development of PE; and (2) indicated preterm delivery at ≤34 weeks of gestation (excluding deliveries as a result of spontaneous preterm labor, preterm pre-labor rupture of membranes or chorioamnionitis).

Results: (1) The prevalence of PE and indicated preterm delivery was 9.2% (n?=?29/314) and 7.3% (n?=?23/314), respectively; (2) the area under the receiver operating characteristic curve (AUC) for the identification of patients who developed PE and/or required indicated preterm delivery was greater than 80% for the UT artery pulsatility index (PI) z-score and each biochemical marker (including their ratios) except sVEGFR-1 MoM; (3) using cutoffs at a false positive rate of 15%, women with abnormal plasma concentrations of angiogenic/anti-angiogenic factors were 7–13 times more likely to develop PE, and 12–22 times more likely to require preterm delivery than those with normal plasma MoM concentrations of these factors; (4) sEng, PlGF, PIGF/sEng and PIGF/sVEGFR-1 ratios MoM, each contributed significant information about the risk of PE beyond that provided by clinical factors and/or Doppler parameters: women who had low MoM values for these biomarkers were at 5–9 times greater risk of developing PE than women who had normal values, adjusting for clinical factors and Doppler parameters (adjusted odds ratio for PlGF: 9.1, PlGF/sEng: 5.6); (5) the concentrations of sVEGFR-1 and PlGF/sVEGFR-1 ratio MoM, each contributed significant information about the risk of indicated preterm delivery beyond that provided by clinical factors and/or Doppler parameters: women who had abnormal values were at 8–9 times greater risk for indicated preterm delivery, adjusting for clinical factors and Doppler parameters; and (6) for a two-stage risk assessment (Umb artery Doppler followed by Ut artery Doppler plus biochemical markers), among women who had normal Umb artery Doppler velocimetry (n?=?279), 21 (7.5%) developed PE and 11 (52%) of these women were identified by an abnormal UT artery Doppler mean PI z-score (>2SD): a combination of PlGF/sEng ratio MoM concentration and abnormal UT artery Doppler velocimetry increased the sensitivity of abnormal UT artery Doppler velocimetry to 76% (16/21) at a fixed false-positive rate of 10% (p?=?0.06).

Conclusion: Angiogenic and anti-angiogenic factors measured in maternal blood between 24 and 34 weeks of gestation can identify the majority of mothers diagnosed with “suspected SGA” who subsequently developed PE or those who later required preterm delivery ≤34 weeks of gestation. Moreover, incorporation of these biochemical markers significantly improves risk assessment performance for these outcomes beyond that of clinical factors and uterine and umbilical artery Doppler velocimetry.     

A prospective cohort study of the value of maternal plasma concentrations of angiogenic and anti-angiogenic factors in early pregnancy and midtrimester in the identification of patients destined to develop preeclampsia

Objective.?Changes in the maternal plasma concentrations of angiogenic (placental growth factor (PlGF) and vascular endothelial growth factor (VEGF)) and anti-angiogenic factors (sEng and vascular endothelial growth factor receptor-1 (sVEGFR-1)) precede the clinical presentation of preeclampsia. This study was conducted to examine the role of maternal plasma PlGF, sEng, and sVEGFR-1 concentrations in early pregnancy and midtrimester in the identification of patients destined to develop preeclampsia.

Methods. This longitudinal cohort study included 1622 consecutive singleton pregnant women. Plasma samples were obtained in early pregnancy (6–15 weeks) and midtrimester (20–25 weeks). Maternal plasma PlGF, sEng, and sVEGFR-1 concentrations were determined using sensitive and specific immunoassays. The primary outcome was the development of preeclampsia. Secondary outcomes included term, preterm, and early-onset preeclampsia. Receiving operating characteristic curves, sensitivity, specificity, positive and negative likelihood ratios, and multivariable logistic regression were applied. A p-value of <0.05 was considered significant.

Results.?(1) The prevalence of preeclampsia, term, preterm, (<37 weeks) and early-onset preeclampsia (<34 weeks) was 3.8 (62/1622), 2.5 (40/1622), 1.4 (22/1622) and 0.6% (9/1622), respectively; (2) Higher likelihood ratios were provided by ratios of midtrimester plasma concentrations of PlGF, sEng, and sVEGFR-1 than single analytes; (3) Individual angiogenic and anti-angiogenic factors did not perform well in the identification of preeclampsia as a whole; in particular, they perform poorly in the prediction of term preeclampsia; (4) In contrast, a combination of these analytes such as the PlGF/sEng ratio, its delta and slope had the best predictive performance with a sensitivity of 100%, a specificity of 98–99%, and likelihood ratios for a positive test of 57.6, 55.6 and 89.6, respectively, for predicting early-onset preeclampsia.

Conclusions.?(1) The PlGF/sEng ratio and its delta and slope had an excellent predictive performance for the prediction of early-onset preeclampsia, with very high likelihood ratios for a positive test result and very low likelihood ratios for a negative test result; and (2) Although the positive likelihood ratios are high and the positive predictive values low, the number of patients needed to be closely followed is 4:1 for the PlGF/sEng ratio and 3:1 for the slope of PlGF/sEng.     

An imbalance between angiogenic and anti-angiogenic factors precedes fetal death in a subset of patients: results of a longitudinal study

Objective.?Women with a fetal death at the time of diagnosis have higher maternal plasma concentrations of the anti-angiogenic factor, soluble vascular endothelial growth factor receptor (sVEGFR)-1, than women with a normal pregnancy. An important question is whether these changes are the cause or consequence of fetal death. To address this issue, we conducted a longitudinal study and measured the maternal plasma concentrations of selective angiogenic and anti-angiogenic factors before the diagnosis of a fetal death. The anti-angiogenic factors studied were sVEGFR-1 and soluble endoglin (sEng), and the angiogenic factor, placental growth factor (PlGF).

Methods.?This retrospective longitudinal nested case–control study included 143 singleton pregnancies in the following groups: (1) patients with uncomplicated pregnancies who delivered a term infant with an appropriate weight for gestational age (n?=?124); and (2) patients who had a fetal death (n?=?19). Blood samples were collected at each prenatal visit, scheduled at 4-week intervals from the first trimester until delivery. Plasma concentrations of sVEGFR-1, sEng, and PlGF were determined by specific and sensitive ELISA. A linear mixed-effects model was used for analysis.

Results.?(1) The average profiles of analyte concentrations as a function of gestational age for sVEGFR-1, sEng and PlGF were different between women destined to have a fetal death and those with a normal pregnancy after adjusting for covariates (p?<?0.05); (2) Plasma sVEGFR-1 concentrations in patients destined to have a fetal death were significantly lower between 7 and 11 weeks of gestation and became significantly higher than those of women with a normal pregnancy between 20 and 37 weeks of gestation (p?<?0.05); (3) Similarly, plasma sEng concentrations of women destined to have a fetal death were lower at 7 weeks of gestation (p?=?0.04) and became higher than those of controls between 20 and 40 weeks of gestation (p?<?0.05); (4) In contrast, plasma PlGF concentrations were higher among patients destined to develop a fetal death between 7 and 14 weeks of gestation and became significantly lower than those in the control group between 22 and 39 weeks of gestation (p?<?0.05); (5) The ratio of PlGF/(sVEGFR-1?×?sEng) was significantly higher in women destined to have a fetal death between 7 and 13 weeks of gestation (94–781%) and significantly lower (44–75%) than those in normal pregnant women between 20 and 40 weeks of gestation (p?<?0.05); (6) Similar results were obtained when patients with a fetal death were stratified into those who were diagnosed before or after 37 weeks of gestation.

Conclusions.?Fetal death is characterised by higher maternal plasma concentrations of PlGF during the first trimester compared to normal pregnancy. This profile changes into an anti-angiogenic one during the second and third trimesters.     

Unexplained fetal death is associated with increased concentrations of anti-angiogenic factors in amniotic fluid

Objective.?Angiogenesis is critical for successful pregnancy. An anti-angiogenic state has been implicated in preeclampsia, fetal growth restriction and fetal death. Increased maternal plasma concentrations of the anti-angiogenic factor, soluble vascular endothelial growth factor receptor (sVEGFR)-1, have been reported in women with preeclampsia and in those with fetal death. Recent observations indicate that an excess of sVEGFR-1 and soluble endoglin (sEng) is also present in the amniotic fluid of patients with preeclampsia. The aim of this study was to determine whether fetal death is associated with changes in amniotic fluid concentrations of sVEGFR-1 and sEng, two powerful anti-angiogenic factors.

Study design.?This cross-sectional study included patients with fetal death (n?=?35) and controls (n?=?129). Fetal death was subdivided according to clinical circumstances into: (1) unexplained (n?=?25); (2) preeclampsia and/or placental abruption (n?=?5); and (3) chromosomal/congenital anomalies (n?=?5). The control group consisted of patients with preterm labor (PTL) who delivered at term (n?=?92) and women at term not in labor (n?=?37). AF concentrations of sVEGFR-1 and sEng were determined by ELISA. Non-parametric statistics and logistic regression analysis were applied.

Results.?(1) Patients with a fetal death had higher median amniotic fluid concentrations of sVEGFR-1 and sEng than women in the control group (p?<?0.001 for each); (2) these results remained significant among different subgroups of stillbirth (p?<?0.05 for each); and (3) amniotic fluid concentrations of sVEGFR-1 and those of sEng above the third quartile were associated with a significant risk of unexplained preterm fetal death (adjusted OR?=?10.8; 95%CI 1.3–89.2 and adjusted OR 87; 95% CI 2.3–3323, respectively).

Conclusion.?Patients with an unexplained fetal death at diagnosis are characterized by an increase in the amniotic fluid concentrations of sVEGFR-1 and sEng. These observations indicate that an excess of anti-angiogenic factors in the amniotic cavity is associated with unexplained fetal death especially in preterm gestations.     

The change in concentrations of angiogenic and anti-angiogenic factors in maternal plasma between the first and second trimesters in risk assessment for the subsequent development of preeclampsia and small-for-gestational age

Introduction. An imbalance between angiogenic and anti-angiogenic factors has been proposed as central to the pathophysiology of preeclampsia (PE). Indeed, patients with PE and those delivering small-for-gestational age (SGA) neonates have higher plasma concentrations of soluble vascular endothelial growth factor receptor-1 (sVEGFR-1) and the soluble form of endoglin (s-Eng), as well as lower plasma concentrations of vascular endothelial growth factor (VEGF) and placental growth factor (PlGF) than do patients with normal pregnancies. Of note, this imbalance has been observed before the clinical presentation of PE or the delivery of an SGA neonate. The objective of this study was to determine if changes in the profile of angiogenic and anti-angiogenic factors in maternal plasma between the first and second trimesters are associated with a high risk for the subsequent development of PE and/or delivery of an SGA neonate.

Methods. This longitudinal case–control study included 402 singleton pregnancies in the following groups: (1) normal pregnancies with appropriate for gestational age (AGA) neonates (n = 201); (2) patients who delivered an SGA neonate (n = 145); and (3) patients who developed PE (n = 56). Maternal plasma samples were obtained at the time of each prenatal visit, scheduled at 4-week intervals from the first or early second trimester until delivery. In this study, we included two samples per patient: (1) first sample obtained between 6 and 15 weeks of gestation (‘first trimester’ sample), and (2) second sample obtained between 20 and 25 weeks of gestation (‘second trimester’ sample). Plasma concentrations of s-Eng, sVEGFR-1, and PlGF were determined by specific and sensitive immunoassays. Changes in the maternal plasma concentrations of these angiogenesis-related factors were compared among normal patients and those destined to develop PE or deliver an SGA neonate while adjusting for maternal age, nulliparity, and body mass index. General linear models and polytomous logistic regression models were used to relate the analyte concentrations, ratios, and product to the subsequent development of PE and SGA.

Results. (1) An increase in the maternal plasma concentration of s-Eng between the first and second trimesters conferred risk for the development of preterm PE and SGA (OR 14.9, 95% CI 4.9–45.0 and OR 2.9, 95% CI 1.5–5.6, respectively). (2) An increase in the maternal plasma concentration of sVEGFR-1 between the first and second trimester conferred risk for the development of preterm PE (OR 3.9, 95% CI 1.2–12.6). (3) A subnormal increase in maternal plasma PlGF concentration between the first and the second trimester was a risk factor for the subsequent development of preterm and term PE (OR 4.3, 95% CI 1.2–15.5 and OR 2.7, 95% CI 1.2–5.9, respectively). (4) In addition, the combination of the three analytes into a pro-angiogenic versus anti-angiogenic ratio (PlGF/(s-Eng × VEGFR-1)) conferred risk for the subsequent development of preterm PE (OR 3.7, 95% CI 1.1–12.1). (5) Importantly, patients with a high change in the s-Eng × sVEGFR-1 product had an OR of 10.4 (95% CI 3.2–33.8) for the development of preterm PE and 1.6 (95% CI 1.0–2.6) for the development of SGA.

Conclusions. Changes in the maternal plasma concentrations of s-Eng, sVEGFR-1, PlGF or their ratios between the first and second trimesters of pregnancy confer an increased risk to deliver an SGA neonate and/or develop PE.     

Pravastatin to prevent recurrent fetal death in massive perivillous fibrin deposition of the placenta (MPFD)

Massive perivillous fibrin deposition of the placenta (MPFD) or maternal floor infarction (MFI) is a serious condition associated with recurrent complications including fetal death and severe fetal growth restriction. There is no method to evaluate the risk of adverse outcome in subsequent pregnancies, or effective prevention. Recent observations suggest that MFI is characterized by an imbalance in angiogenic/anti-angiogenic factors in early pregnancy; therefore, determination of these biomarkers may identify the patient at risk for recurrence. We report the case of a pregnant woman with a history of four consecutive pregnancy losses, the last of which was affected by MFI. Abnormalities of the anti-angiogenic factor, sVEGFR-1, and soluble endoglin (sEng) were detected early in the index pregnancy, and treatment with pravastatin corrected the abnormalities. Treatment resulted in a live birth infant at 34 weeks of gestation who had normal biometric parameters and developmental milestones at the age of 2. This is the first reported successful use of pravastatin to reverse an angiogenic/anti-angiogenic imbalance and prevent fetal death.     

A subset of patients destined to develop spontaneous preterm labor has an abnormal angiogenic/anti-angiogenic profile in maternal plasma: Evidence in support of pathophysiologic heterogeneity of preterm labor derived from a longitudinal study

Objective.?An imbalance between angiogenic and anti-angiogenic factors in maternal blood has been observed in several obstetrical syndromes including preeclampsia, pregnancies with fetal growth restriction and fetal death. Vascular lesions have been identified in a subset of patients with spontaneous preterm labor (PTL). It is possible that PTL may be one of the manifestations of an anti-angiogenic state. The aim of this study was to determine if patients prior to the clinical diagnosis of PTL leading to preterm delivery had plasma concentrations of angiogenic and anti-angiogenic factors different from normal pregnant women.

Study Design.?This longitudinal nested case–control study included normal pregnant women (n?=?208) and patients with PTL leading to preterm delivery (n?=?52). Maternal blood samples were collected at 6 gestational age intervals from 6 to36.9 weeks of gestation. The end point (time of diagnosis) of the study, ‘True PTL’, was defined as patients presenting with PTL and delivered within 1 day. Plasma concentrations of sVEGFR-1, sVEGFR-2, sEng and PlGF were determined by ELISA. Analysis was performed with both cross-sectional and longitudinal (mixed effects model) approaches.

Results. (1) Plasma sEng concentration in patients destined to develop PTL was higher than that in normal pregnant women from 15–20 weeks of gestation. The difference became statistical significant at 28 weeks of gestation, or approximately 5–10 weeks prior to the diagnosis of ‘true PTL’. (2) Backward analysis suggests that plasma concentrations of PlGF and sVEGFR-2 were lower, and those of sVEGFR-1 were higher in patients with PTL than in normal pregnant women less than 5 weeks prior to the diagnosis of ‘true PTL’; and (3) Plasma concentrations of sEng and sVEGFR-1 were higher and those of PlGF and sVEGFR-2 were lower in patients diagnosed with PTL and delivery within 1 day than in normal pregnant women who delivered at term.

Conclusion.?The changes in sEng are demonstrable several weeks prior to the onset of preterm parturition. In contrast, the changes in the other angiogenic proteins are present close to the onset of PTL and delivery. This observation supports the view that an imbalance of angiogenic factors participates in the pathophysiology of spontaneous preterm parturition.     

Endocan,a putative endothelial cell marker,is elevated in preeclampsia,decreased in acute pyelonephritis,and unchanged in other obstetrical syndromes

Objective: Endocan, a dermatan sulphate proteoglycan produced by endothelial cells, is considered a biomarker for endothelial cell activation/dysfunction. Preeclampsia is characterized by systemic vascular inflammation, and endothelial cell activation/dysfunction. Therefore, the objectives of this study were to determine whether: (1) plasma endocan concentrations in preeclampsia differ from those in uncomplicated pregnancies; (2) changes in plasma endocan concentration relate to the severity of preeclampsia, and whether these changes are specific or observed in other obstetrical syndromes such as small-for-gestational age (SGA), fetal death (FD), preterm labor (PTL) or preterm prelabor rupture of membranes (PROM); (3) a correlation exists between plasma concentration of endocan and angiogenic (placental growth factor or PlGF)/anti-angiogenic factors (soluble vascular endothelial growth factor receptor or sVEGFR-1, and soluble endoglin or sEng) among pregnancies complicated by preeclampsia; and (4) plasma endocan concentrations in patients with preeclampsia and acute pyelonephritis (both conditions in which there is endothelial cell activation) differ.

Method: This cross-sectional study included the following groups: (1) uncomplicated pregnancy (n?=?130); (2) preeclampsia (n?=?102); (3) pregnant women without preeclampsia who delivered an SGA neonate (n?=?51); (4) FD (n?=?49); (5) acute pyelonephritis (AP; n?=?35); (6) spontaneous PTL (n?=?75); and (7) preterm PROM (n?=?64). Plasma endocan concentrations were determined in all groups, and PIGF, sEng and VEGFR-1 plasma concentrations were measured by ELISA in the preeclampsia group.

Results: (1) Women with preeclampsia had a significantly higher median plasma endocan concentration than those with uncomplicated pregnancies (p?=?0.004); (2) among women with preeclampsia, the median plasma endocan concentration did not differ significantly according to disease severity (p?=?0.1), abnormal uterine artery Doppler velocimetry (p?=?0.7) or whether diagnosis was made before or after 34 weeks gestational age (p?=?0.3); (3) plasma endocan concentration in women with preeclampsia correlated positively with plasma anti-angiogenic factor concentrations [sVEGFR-1: Spearman rho 0.34, p?=?0.001 and sEng: Spearman rho 0.30, p?=?0.003]; (4) pregnancies complicated by acute pyelonephritis with bacteremia had a lower median plasma endocan concentration than pregnancies complicated by acute pyelonephritis without bacteremia (p?=?0.004), as well as uncomplicated pregnancies (p?=?0.001); and (5) there was no significant difference in the median plasma endocan concentration between uncomplicated pregnancies and those complicated by FD, delivery of an SGA neonate, PTL or preterm PROM (other members of the “great obstetrical syndromes”; each p?>?0.05).

Conclusion: Median maternal plasma endocan concentrations were higher preeclampsia and lower in acute pyelonephritis with bacteremia than in uncomplicated pregnancy. No significant difference was observed in the median plasma endocan concentration between other great obstetrical syndromes and uncomplicated pregnancies. The difference in the direction of change of endocan in preeclampsia and acute pyelonephritis with bacteremia may be consistent with the view that both disease entities differ in pathogenic mechanisms, despite their associations with systemic vascular inflammation and endothelial cell activation/dysfunction.     

Association of first-trimester angiogenic factors with placental histological findings in late-onset preeclampsia

ObjectiveTo explore in women with late-onset preeclampsia (PE) the association between maternal levels of angiogenic/antiangiogenic factors in the first trimester of pregnancy and histological findings attributable to placental underperfusion (PUP).MethodsA nested case-control cohort study was conducted in 73 women with pregnancies complicated by late-onset PE (>34 weeks at delivery) matched with controls. First trimester uterine artery Doppler (UtA); maternal levels of placental growth factor (PlGF) and soluble fms-like tyrosine kinase-1 (sFlt-1) were retrieved. Placentas were histologically evaluated using a hierarchical and standardized classification system. One-way ANOVA with linear polynomial contrast or linear-by-linear association test was performed to test the hypothesis of a linear association across study groups (controls, PE without PUP and PE with PUP).ResultsIn 54 (74%) placentas, 89 placental histological findings qualifying for PUP were found. Across study groups, significant values were observed in maternal levels of decreased PlGF (MoM values: 1.53, 1.41 and 1.37; p < 0.001), increased sFlt-1 (MoM values: 3.11, 3.11 and 3.22; p = 0.002), increased sFlt-1/PlGF ratio (MoM values: 2.3, 2.3 and 2.44; p < 0.001), abnormal UtA Doppler (MoM values: 1, 1.26 and 1.32; p < 0.001), and worse perinatal outcomes in terms of gestational age at delivery, cesarean section for not reassuring fetal status, birth weight and neonatal acidosis.DiscussionIn late-onset PE an imbalance of circulating angiogenic and anti-angiogenic factors already present at 8–10 weeks of pregnancy was associated with histological findings reflecting placental insufficiency. An early first trimester screening by angiogenic factors might help to identify patients with placental involvement among late-onset PE cases.ConclusionIn late-onset preeclampsia, first-trimester uterine Doppler and circulating levels of angiogenic/antiangiogenic factors are associated with placental underperfusion.     

Maternal plasma concentrations of sST2 and angiogenic/anti-angiogenic factors in preeclampsia

Abstract

Objectives: Angiogenic/anti-angiogenic factors have emerged as one of the promising biomarkers for the prediction of preeclampsia. Since not all patients with preeclampsia can be identified by these analytes, the search for additional biomarkers continues. The soluble form of ST2 (sST2), a protein capable of binding to interleukin (IL)-33 and thus contributing to a Th1-biased immune response, has been reported to be elevated in maternal plasma of women with preeclampsia. The aims of this study were to examine: (1) differences in maternal plasma concentrations of sST2 and IL-33 between women diagnosed with preeclampsia and those having uncomplicated pregnancies; (2) the relationship between sST2, umbilical and uterine artery Doppler velocimetry, and the severity of preeclampsia; and (3) the performance of sST2 and angiogenic/anti-angiogenic factors in identifying patients with preeclampsia at the time of diagnosis.

Methods: This cross-sectional study included women with preeclampsia (n?=?106) and women with an uncomplicated pregnancy (n?=?131). Plasma concentrations of sST2, IL-33, soluble vascular endothelial growth factor receptor (sVEGFR)-1, soluble endoglin (sEng) and placental growth factor (PlGF) were determined by enzyme linked immune sorbent assay. Area under the receiver operating characteristic curve (AUC) for the identification of preeclampsia was examined for each analyte.

Results: (1) Patients with preeclampsia had a higher mean plasma concentrations of sST2 than those with an uncomplicated pregnancy (p?<?0.0001), while no significant difference in the mean plasma concentration of IL-33 between the two groups was observed; (2) the magnitude of this difference was greater in early-onset, compared to late-onset disease, and in severe compared to mild preeclampsia; (3) sST2 plasma concentrations did not correlate with the results of uterine or umbilical artery Doppler velocimetry (p?=?0.7 and p?=?1, respectively) among women with preeclampsia; (4) sST2 correlated positively with plasma concentrations of sVEGFR1-1 and sEng (Spearman’s Rho?=?0.72 and 0.63; each p?<?0.0001), and negatively with PlGF (Spearman’s Rho?=??0.56, p?<?0.0001); and (5) while the AUC achieved by sST2 and angiogenic/anti-angiogenic factors in identifying women with preeclampsia at the time of diagnosis were non-significantly different prior to term (<37 weeks of gestation), thereafter the AUC achieved by sST2 was significantly less than that achieved by angiogenic/anti-angiogenic factors.

Conclusions: Preeclampsia is associated with increased maternal plasma concentrations of sST2. The findings that sST2 concentrations do not correlate with uterine or umbilical artery Doppler velocimetry in women with preeclampsia suggest that elevated maternal plasma sST2 concentrations in preeclampsia are not related to the increased impedance to flow in the utero-placental circulation. The performance of sST2 in identifying preeclampsia at the time of diagnosis prior to 37 weeks of gestation was comparable to that of angiogenic/anti-angiogenic factors. It remains to be elucidated if an elevation of maternal plasma sST2 concentrations in pregnancy is specific to preeclampsia.     

相关细胞因子在预测宫内感染中的意义

宫内感染对妊娠的影响及对母儿造成的危害日益受到关注,其可导致早产及围生儿死亡、新生儿呼吸窘迫综合征等不良妊娠结局;但目前以临床诊断为主,使许多亚临床感染的孕产妇得不到及时诊治,而宫内感染一旦出现临床症状时病情往往已经危重,近年,预测宫内感染的相关研究成为热点。研究表明,白细胞介素10、粒细胞集落刺激因子、白细胞介素6、基质金属蛋白酶9、高迁移率族蛋白1、人类β防御素等细胞因子对宫内感染有一定的预测价值,联合蛋白组学方法可能实现宫内感染的早期诊断、早期治疗。综述宫内感染预测的相关细胞因子的研究近况。     

Mid-pregnancy levels of angiogenic markers as indicators of pathways to preterm delivery

Objective: To determine whether mid-pregnancy levels of angiogenic markers were associated with increased risk of preterm delivery (PTD). Methods: We studied a subcohort from the Pregnancy Outcomes and Community Health Study for whom mid-pregnancy angiogenic markers (soluble fms-like tyrosine kinase-1 [sFlt-1], soluble endoglin [sEng] and placental growth factor [PlGF]) and covariate data were available (N = 1301). Angiogenic marker levels were grouped as high/not high (sFlt-1 and sEng), low/not low (PlGF) and high/intermediate/low (sFlt-1). Associations between levels of angiogenic markers and PTD/PTD subtype were determined for women who were nonsmokers during pregnancy (N = 933). Results: Low PlGF and high sEng were associated with medically-indicated PTD and PTD <35 weeks, largely due to preeclampsia (PE). Excluding PE and small-for-gestational-age infants, low sFlt-1 was positively associated with medically-indicated PTD. Conclusions: Among nonsmokers, mid-pregnancy levels of angiogenic markers may mark multiple pathways leading to PTD, only one attributable to PE.     

Angiogenic biomarkers for prediction of early preeclampsia onset in high-risk women

Abstract

Objective: Chronic hypertension, pregestational diabetes mellitus, history of prior preeclampsia and obese nulliparity are maternal conditions associated with increased preeclampsia risk. Whether altered maternal angiogenic factor levels allow for prediction of pending disease is unclear. Our objective was to evaluate angiogenic factors for early preeclampsia prediction in high-risk women.

Methods: Serial serum specimens were collected from 157 women at high preeclampsia risk and 50 low-risk controls between 23 and 36 weeks gestation in 3 windows (23–27.6, 28–31.6, and 32–35.6 weeks) in a two-center observational cohort. Soluble fms-like tyrosine kinase-1 (sFlt1), placental growth factor (PlGF) and soluble endoglin (sEng) were measured by ELISA.

Results: Multivariate parsimonious logistic regression analyses using backward elimination for prediction of early-preeclampsia (diagnosed?<?34 weeks) found the best-fitting model included the predictors (1) sFlt1 measured in the second window (28–31.6 weeks) with AUC 0.85, sensitivity 67% and specificity 96% and (2) sFlt1 measured in the first window (23–27.6 weeks) and sEng change between first and second window with AUC 0.91, sensitivity 86% and specificity 96%.

Conclusions: Two-stage sampling screening protocol utilizing sFlt1 and sEng is promising for prediction of preeclampsia diagnosed before 34 weeks. Larger studies are needed to confirm these findings.     

A role of the anti-angiogenic factor sVEGFR-1 in the ‘mirror syndrome’ (Ballantyne's syndrome)

Background.?‘Mirror syndrome’ (Ballantyne's syndrome) refers to the association of fetal hydrops with placentomegaly and severe maternal edema. Preeclampsia occurs in approximately 50% of these cases. Soluble vascular endothelial growth factor receptor-1 (sVEGFR-1), an anti-angiogenic factor, has been implicated in the pathophysiology of preeclampsia (PE).

Objective.?The objective of this study was to determine if the maternal plasma concentration of sVEGFR-1 is elevated in patients with mirror syndrome.

Study design.?This case-control study included patients with uncomplicated pregnancies (n = 40) and those with mirror syndrome (n = 4) matched for gestational age. Mirror syndrome was defined as fetal hydrops and severe maternal edema. Maternal plasma sVEGFR-1 concentrations were determined using specific enzyme-linked immunosorbent assays. Immunohistochemistry of sVEGFR-1 on villous trophoblasts was also performed in samples from one patient with mirror syndrome and compared with those from a patient with spontaneous preterm delivery matched for gestational age. Non-parametric statistics were used for analysis (p < 0.05).

Results.?(1) The median maternal plasma concentration of sVEGFR-1 was significantly higher in patients with mirror syndrome than in the control group (median: 3974 pg/mL, range: 3083–10 780 vs. median: 824 pg/mL, range: 260–4712, respectively; p < 0.001). (2) All patients with mirror syndrome had sVEGFR-1 concentrations above the 95^th percentile for gestational age. Syncytiotrophoblast, especially syncytial knots, showed strong staining with antibodies against sVEGFR-1 in placental samples from the patient with mirror syndrome, but not in those from the patient with spontaneous preterm delivery.

Conclusion.?High maternal plasma concentrations of sVEGFR-1 were observed in mirror syndrome. We propose that this anti-angiogenic factor may participate in the pathophysiology of this syndrome. Thus, maternal plasma determination of sVEGFR-1 may help to identify the hydropic fetus that places the mother at risk for preeclampsia.     

Discordant clinical presentations of preeclampsia and intrauterine fetal growth restriction with similar pro- and anti-angiogenic profiles

Abstract

Objective: To evaluate the plasma levels of angiogenic factors in preeclampsia (PE) and intrauterine fetal growth restriction (IUGR) and their potential as biomarkers to distinguish normal from pathologic pregnancies.

Methods: Case control study included singleton pregnancies in four categories: (i) normal (n?=?29), (ii) PE (n?=?15), (iii) PE and IUGR (n?=?16) and (iv) IUGR (n?=?24). The classification of IUGR included umbilical artery Doppler resistance. Maternal plasma placental growth factor (PlGF), soluble fms-like tyrosine kinase-1 (sFlt-1), soluble kinase domain receptor (sKDR) and soluble endoglin (sEng) as well as fetal umbilical artery sFlt-1 levels were determined. Each individual marker and their ratios were assessed for their potential to distinguish normal pregnancy from pregnancies affected by PE and/or IUGR.

Results: We found (i) elevated plasma sFlt-1, sEng and reduced PlGF, sKDR in PE and IUGR; (ii) similar angiogenic profiles in PE and IUGR and (iii) sEng and sFlt-1*sEng/PlGF performed best as biomarkers in identifying pathologic pregnancies.

Conclusions: PE and IUGR have similar angiogenic profiles, suggesting that angiogenic marker profiles lack specificity in identifying PE and that other factors are required for the development of PE instead of IUGR. sEng should be included in a biomarker profile for predicting PE or IUGR.     

A longitudinal study of angiogenic (placental growth factor) and anti-angiogenic (soluble endoglin and soluble vascular endothelial growth factor receptor-1) factors in normal pregnancy and patients destined to develop preeclampsia and deliver a small for gestational age neonate.

INTRODUCTION: Accumulating evidence suggests that an imbalance between pro-angiogenic (i.e., vascular endothelial growth factor (VEGF) and placental growth factor (PlGF)) and anti-angiogenic factors (i.e., soluble VEGF receptor-1 (sVEGFR-1, also referred to as sFlt1)) is involved in the pathophysiology of preeclampsia (PE). Endoglin is a protein that regulates the pro-angiogenic effects of transforming growth factor beta, and its soluble form has recently been implicated in the pathophysiology of PE. The objective of this study was to determine if changes in maternal plasma concentration of these angiogenic and anti-angiogenic factors differ prior to development of disease among patients with normal pregnancies and those destined to develop PE (preterm and term) or to deliver a small for gestational age (SGA) neonate. METHODS: This longitudinal nested case-control study included 144 singleton pregnancies in the following groups: (1) patients with uncomplicated pregnancies who delivered appropriate for gestational age (AGA) neonates (n = 46); (2) patients who delivered an SGA neonate but did not develop PE (n = 56); and (3) patients who developed PE (n = 42). Longitudinal samples were collected at each prenatal visit, scheduled at 4-week intervals from the first or early second trimester until delivery. Plasma concentrations of soluble endoglin (s-Eng), sVEGFR-1, and PlGF were determined by specific and sensitive ELISA. RESULTS: (1) Patients destined to deliver an SGA neonate had higher plasma concentrations of s-Eng throughout gestation than those with normal pregnancies; (2) patients destined to develop preterm PE and term PE had significantly higher concentrations of s-Eng than those with normal pregnancies at 23 and 30 weeks, respectively (for preterm PE: p < 0.036 and for term PE: p = 0.002); (3) patients destined to develop PE (term or preterm) and those who delivered an SGA neonate had lower plasma concentrations of PlGF than those with a normal pregnancy throughout gestation, and the maternal plasma concentration of this analyte became detectable later among patients with pregnancy complications, compared to normal pregnant women; (4) there were no significant differences in the plasma concentrations of sVEGFR-1 between patients destined to deliver an SGA neonate and those with normal pregnancies; (5) patients destined to develop preterm and term PE had a significantly higher plasma concentration of sVEGFR-1 at 26 and 29 weeks of gestation than controls (p = 0.009 and p = 0.0199, respectively); and (6) there was no significant difference in the increment of sVEGFR-1 between control patients and those who delivered an SGA neonate (p = 0.147 at 25 weeks and p = 0.8285 at 40 weeks). CONCLUSIONS: (1) Changes in the maternal plasma concentration of s-Eng, sVEGFR-1, and PlGF precede the clinical presentation of PE, but only changes in s-Eng and PlGF precede the delivery of an SGA neonate; and (2) differences in the profile of angiogenic and anti-angiogenic response to intrauterine insults may determine whether a patient will deliver an SGA neonate, develop PE, or both.     

Angiogenic biomarkers for prediction of maternal and neonatal complications in suspected preeclampsia

Objective: To determine if maternal serum angiogenic factors predict maternal and neonatal complications in women presenting to an acute care setting with suspected preeclampsia. Study design: Maternal serum samples were prospectively collected from women with suspected preeclampsia at the time of initial presentation to hospital triage with signs or symptoms of preeclampsia. Soluble fms-like tyrosine kinase-1 (sFlt1), placental growth factor (PlGF), and soluble endoglin (sEng) were measured by ELISA. The primary outcome was a composite of maternal and neonatal complications. Results: Of 276 women with suspected preeclampsia, 78 developed maternal or neonatal complications. Among women presenting prior to 37 weeks gestation, sFlt1, PlGF, and sEng were significantly different in women who developed maternal and neonatal complications as compared to women without complications. Higher levels of sFlt1, sEng, and the sFlt1:PlGF ratio were associated with an increased odds of complications among women presenting prior to 37 weeks. A multivariable model combining the sFlt1:PlGF ratio with clinical variables was more predictive of complications (AUC 0.91, 95% CI 0.85–0.97) than a model using clinical variables alone (AUC 0.82, 95% CI 0.79–0.90). Conclusion: Angiogenic biomarkers associate with maternal and neonatal complications in women with suspected preeclampsia, and may be useful for risk stratification.     

Angiogenic imbalance as a contributor to the pathophysiology of preeclampsia among black African women

Objective: The pathogenesis of preeclampsia remains unclear despite extensive research. Altered angiogenic balance has been hypothesized to play a significant role in the clinical manifestations of this syndrome. However this imbalance has not been investigated extensively among black African women. The aim of this study was to investigate the maternal levels of the angiogenic factors soluble vascular endothelial growth factor receptor 1 (sFLT1) and placental growth factor (PlGF) among black African women with preeclampsia.

Methods: A case control study was conducted in the Mthatha hospital complex in South Africa including 51 women with preeclampsia and 82 women with uncomplicated pregnancies. Blood samples were drawn from participants and serum was used to assess sFLT1, and PlGF levels quantified using specific enzyme linked immunosorbent assays. Non- parametric statistics were used for analysis.

Results: Black African women with preeclampsia were found to have significantly lower levels of PlGF (90.3?±?8.9?pg/ml versus 172.8?±?20.2?pg/ml; p?p?p?Conclusion: The results support an altered angiogenic balance may contribute to the pathogenesis/pathophysiology of preeclampsia among black African women as reported in other populations.     

Fetal death: A condition with a dissociation in the concentrations of soluble vascular endothelial growth factor receptor-2 between the maternal and fetal compartments

Objective.?An anti-angiogenic state has been implicated in the pathophysiology of preeclampsia, fetal growth restriction and fetal death. Vascular endothelial growth factor (VEGF), an indispensible angiogenic factor for embryonic and placental development exerts its angiogenic properties through the VEGF receptor (VEGFR)-2. A soluble form of this protein (sVEGFR-2) has been recently detected in maternal blood. The aim of this study was to determine if fetal death was associated with changes in the concentrations of sVEGFR-2 in maternal plasma and amniotic fluid.

Study Design.?Maternal plasma was obtained from patients with fetal death (n?=?59) and normal pregnant women (n?=?134). Amniotic fluid was collected from 36 patients with fetal death and the control group consisting of patients who had an amniocentesis and delivered at term (n?=?160). Patients with fetal death were classified according to the clinical circumstances into the following groups: (1) unexplained; (2) preeclampsia and/or placental abruption; (3) chromosomal and/or congenital anomalies. Plasma and amniotic fluid concentrations of sVEGFR-2 were determined by ELISA. Non-parametric statistics and logistic regression analysis were applied.

Results.?(1) Patients with a fetal death had a significantly lower median plasma concentration of sVEGFR-2 than normal pregnant women (p?<?0.001). The median plasma concentration of sVEGFR-2 in patients with unexplained fetal death and in those with preeclampsia/abruption, but not that of those with congenital anomalies, was lower than that of normal pregnant women (p?=?0.006, p?<?0.001 and p?=?0.2, respectively); (2) the association between plasma sVEGFR-2 concentrations and preterm unexplained fetal death remained significant after adjusting for potential confounders (OR: 3.2; 95% CI: 1.4–7.3 per each quartile decrease in plasma sVEGFR-2 concentrations); (3) each subgroup of fetal death had a higher median amniotic fluid concentration of sVEGFR-2 than the control group (p?<?0.001 for each); (4) the association between amniotic fluid sVEGFR-2 concentrations and preterm unexplained fetal death remained significant after adjusting for potential confounders (OR: 15.6; 95% CI: 1.5–164.2 per each quartile increase in amniotic fluid sVEGFR-2 concentrations); (5) among women with fetal death, there was no relationship between maternal plasma and amniotic fluid concentrations of sVEGFR-2 (Spearman Rho: 0.02; p?=?0.9).

Conclusion.?Pregnancies with a fetal death, at the time of diagnosis, are characterized by a decrease in the maternal plasma concentration of sVEGFR-2, but an increase in the amniotic fluid concentration of this protein. Although a decrease in sVEGFR-2 concentration in maternal circulation depends upon the clinical circumstances of fetal death, an increase in sVEGFR-2 concentration in amniotic fluid seems to be a common feature of fetal death. It remains to be determined if the perturbation in sVEGFR-2 concentrations in maternal and fetal compartments observed herein preceded the death of a fetus.