Plasma Soluble Fms-like Tyrosine Kinase-1 Research Articles

Plasma soluble fms-like tyrosine kinase-1, placental growth factor, and vascular endothelial growth factor system gene variants as predictors of survival in heart failure.

Soluble fms-like tyrosine kinase-1 (sFlt-1) and placental growth factor (PlGF), components of the vascular endothelial growth factor (VEGF) system, play key roles in angiogenesis. Reports of elevated plasma levels of sFlt-1 and PlGF in coronary heart disease and heart failure (HF) led us to investigate their utility, and VEGF system gene single nucleotide polymorphisms (SNPs), as prognostic biomarkers in HF. ELISA assays for sFlt-1, PlGF and N-terminal pro-B-type natriuretic peptide (NT-proBNP) were performed on baseline plasma samples from the PEOPLE cohort (n = 890), a study of outcomes among patients after an episode of acute decompensated HF. Eight SNPs potentially associated with sFlt-1 or PlGF levels were genotyped. sFlt-1 and PlGF were assayed in 201 subjects from the Canterbury Healthy Volunteers Study (CHVS) matched to PEOPLE participants. All-cause death was the major endpoint for clinical outcome considered. In PEOPLE participants, mean plasma levels for both sFlt-1 (125 ± 2.01 pg/ml) and PlGF (17.5 ± 0.21 pg/ml) were higher (both p < 0.044) than in the CHVS cohort (81.2 ± 1.31 pg/ml and 15.5 ± 0.32 pg/ml, respectively). sFlt-1 was higher in HF with reduced ejection fraction compared to HF with preserved ejection fraction (p = 0.005). The PGF gene SNP rs2268616 was univariately associated with death (p = 0.016), and was also associated with PlGF levels, as was rs2268614 genotype. Cox proportional hazards modelling (n = 695, 246 deaths) showed plasma sFlt-1, but not PlGF, predicted survival (hazard ratio 6.44, 95% confidence interval 2.57-16.1; p < 0.001) in PEOPLE, independent of age, NT-proBNP, ischaemic aetiology, diabetic status and beta-blocker therapy. Plasma sFlt-1 concentrations have potential as an independent predictor of survival and may be complementary to established prognostic biomarkers in HF.

Metabolomic biomarkers in midtrimester maternal plasma can accurately predict the development of preeclampsia

Early identification of patients at risk of developing preeclampsia (PE) would allow providers to tailor their prenatal management and adopt preventive strategies, such as low-dose aspirin. Nevertheless, no mid-trimester biomarkers have as yet been proven useful for prediction of PE. This study investigates the ability of metabolomic biomarkers in mid-trimester maternal plasma to predict PE. A case–control study was conducted including 33 pregnant women with mid-trimester maternal plasma (gestational age [GA], 16–24 weeks) who subsequently developed PE and 66 GA-matched controls with normal outcomes (mid-trimester cohort). Plasma samples were comprehensively profiled for primary metabolic and lipidomic signatures based on gas chromatography time-of-flight mass spectrometry (GC-TOF MS) and liquid chromatography Orbitrap mass spectrometry (LC-Orbitrap MS). A potential biomarker panel was computed based on binary logistic regression and evaluated using receiver operating characteristic (ROC) analysis. To evaluate whether this panel can be also used in late pregnancy, a retrospective cohort study was conducted using plasma collected from women who delivered in the late preterm period because of PE (n = 13) or other causes (n = 21) (at-delivery cohort). Metabolomic biomarkers were compared according to the indication for delivery. Performance of the metabolomic panel to identify patients with PE was compared also to a commonly used standard, the plasma soluble fms-like tyrosine kinase-1/placental growth factor (sFlt-1/PlGF) ratio. In the mid-trimester cohort, a total of 329 metabolites were identified and semi-quantified in maternal plasma using GC-TOF MS and LC-Orbitrap-MS. Binary logistic regression analysis proposed a mid-trimester biomarker panel for the prediction of PE with five metabolites (SM C28:1, SM C30:1, LysoPC C19:0, LysoPE C20:0, propane-1,3-diol). This metabolomic model predicted PE better than PlGF (AUC [95% CI]: 0.868 [0.844–0.891] vs 0.604 [0.485–0.723]) and sFlt-1/PlGF ratio. Analysis of plasma from the at-delivery cohort confirmed the ability of this biomarker panel to distinguish PE from non-PE, with comparable discrimination power to that of the sFlt-1/PlGF ratio. In conclusion, an integrative metabolomic biomarker panel in mid-trimester maternal plasma can accurately predict the development of PE and showed good discriminatory power in patients with PE at delivery.

Evaluation of blood vessel injury, oxidative stress and circulating inflammatory factors in an L-NAME-induced preeclampsia-like rat model.

Preeclampsia is a pregnancy-specific disease characterized by hypertension as well as proteinuria after the 20th week of pregnancy. Animal models are effective tools for studying the pathogenesis, diagnostic criteria and treatment methods of preeclampsia. The present study sought to establish and evaluate a preeclampsia-like Sprague Dawley (SD) rat model using N-nitro-L-arginine methyl ester (L-NAME). Rats were randomly assigned to 7 groups (n=10 in each): Control rats and rats treated with low-dose L-NAME (40 mg/kg body weight/day) starting from gestational day (GD) 9, medium-L-NAME (75 mg/kg body weight/day) starting from GD 9 (9D ML group), high-dose L-NAME (125 mg/kg body weight/day) starting from GD 9, low-dose L-NAME starting from GD 10, medium-dose L-NAME starting from GD 10 and high-dose L-NAME starting from GD 10. Blood pressure (BP), 24-h proteinuria, fetal intrauterine growth, histopathological changes, the plasma soluble fms-like tyrosine kinase-1 (sFlt-1)/placental growth factor (PLGF) ratio and cytokine levels were evaluated. Elevated BP, increased urinary albumin excretion, severe endotheliosis, mesangial expansion and increased sFlt-1/PLGF ratios were observed in the experimental groups compared with the control group (P<0.05), particularly in the 9D ML group. The results of the present study may optimize the conditions of the previously established L-NAME-induced preeclampsia SD rat model and aid further study into the pathogenesis of preeclampsia.

Causative Effects of Genetically Determined High Maternal/Fetal Endothelin-1 on Preeclampsia-Like Conditions in Mice.

Endothelin-1 (ET-1) is implicated in the pathophysiology of preeclampsia. An association between an EDN1 gene polymorphism with high ET-1 and preeclampsia was reported in humans, but their cause and effect relationships have not been defined. We examined the pregnancy effects in mice with a modified Edn1 allele that increases mRNA stability and thus ET-1 production. Heterozygous Edn1H/+ females showed no obvious abnormalities before pregnancy, but when mated with wild-type (WT) males developed a full spectrum of preeclampsia-like phenotypes, including increased systolic blood pressure, proteinuria, glomerular endotheliosis, and intrauterine fetal growth restriction. At 7.5 days post-coitus, the embryos from Edn1H/+ dams, regardless of their Edn1 genotype, lagged 12 hours in development compared with embryos from WT dams, had disoriented ectoplacental cones, and retained high E-cadherin expression. In contrast, WT females mated with Edn1H/+ males, which also carried half of the fetuses with Edn1H/+ genotype, showed a mild systolic blood pressure increase only. These WT dams had 2× higher plasma soluble fms-like tyrosine kinase-1 than WT dams mated with WT males. In human first trimester trophoblast cells, pharmacological doses of ET-1 increased the cellular sFlt1 transcripts and protein secretion via both type A and B ET-1 receptors. Our data demonstrate that high maternal ET-1 production causes preeclampsia-like phenotypes during pregnancy, affecting both initial stage of trophoblast differentiation/invasion and maternal peripheral vasculature during late gestation. High fetal ET-1 production, however, could cause increased soluble fms-like tyrosine kinase-1 in the maternal circulation and contribute to blood pressure elevation.

Vitamin D restores angiogenic balance and decreases tumor necrosis factor-α in a rat model of pre-eclampsia.

Deficiency of vitamin D is correlated with pre-eclampsia (PE), a hypertensive disorder of pregnancy, and is characterized by angiogenic imbalance and inflammation. The aim of this study was to investigate whether vitamin D supplementation can restore the angiogenic balance and ameliorate inflammation in a rat model of PE. PE was induced using l-nitroarginine methylester. Normal pregnant and PE-induced rats were supplemented with vitamin D on gestation days 14-19. Blood pressure was significantly increased in PE-induced rats compared with normal pregnant rats (P < 0.05), and vitamin D supplementation ameliorated this difference. In addition, rats from the PE group had lower vascular endothelial growth factor (VEGF; P < 0.01), and higher plasma-soluble FMS-like tyrosine kinase-1 (sFlt-1) and tumor necrosis factor-α (TNF-α; P < 0.01 for both) compared with the normal pregnant group. The vitamin D treatment group had significantly increased VEGF, and reduced sFlt-1 and TNF-α compared with the untreated PE group. Moreover, vitamin D supplementation was able to reduce the oxidative stress by lowering the plasma oxidative stress marker malondialdehyde. Vitamin D supplementation plays an important role in restoring angiogenic balance and reducing inflammation in pregnancy-induced hypertension.

Vitamin D supplementation improves pathophysiology in a rat model of preeclampsia.

Deficiency of vitamin D (VD) is associated with preeclampsia (PE), a hypertensive disorder of pregnancy characterized by proinflammatory immune activation. We sought to determine whether VD supplementation would reduce the pathophysiology and hypertension associated with the reduced uterine perfusion pressure (RUPP) rat model of PE. Normal pregnant (NP) and RUPP rats were supplemented with VD2 or VD3 (270 IU and 15 IU/day, respectively) on gestation days 14-18 and mean arterial pressures (MAPs) measured on day 19. MAP increased in RUPP to 123 ± 2 mmHg compared with 102 ± 3 mmHg in NP and decreased to 113 ± 3 mmHg with VD2 and 115 ± 3 mmHg with VD3 in RUPP rats. Circulating CD4+ T cells increased in RUPP to 7.90 ± 1.36% lymphocytes compared with 2.04 ± 0.67% in NP but was lowered to 0.90 ± 0.19% with VD2 and 4.26 ± 1.55% with VD3 in RUPP rats. AT1-AA, measured by chronotropic assay, decreased from 19.5 ± 0.4 bpm in RUPPs to 8.3 ± 0.5 bpm with VD2 and to 15.4 ± 0.7 bpm with VD3. Renal cortex endothelin-1 (ET-1) expression was increased in RUPP rats (11.6 ± 2.1-fold change from NP) and decreased with both VD2 (3.3 ± 1.1-fold) and VD3 (3.1 ± 0.6-fold) supplementation in RUPP rats. Plasma-soluble FMS-like tyrosine kinase-1 (sFlt-1) was also reduced to 74.2 ± 6.6 pg/ml in VD2-treated and 91.0 ± 16.1 pg/ml in VD3-treated RUPP rats compared with 132.7 ± 19.9 pg/ml in RUPP rats. VD treatment reduced CD4+ T cells, AT1-AA, ET-1, sFlt-1, and blood pressure in the RUPP rat model of PE and could be an avenue to improve treatment of hypertension in response to placental ischemia.

Abstract W MP28: Early Elevation of Plasma Soluble Fms-Like Tyrosine Kinase-1 is Associated With Poor Functional Outcome After Subarachnoid Hemorrhage

Background: Vasospasm (VSP) and brain injury following subarachnoid hemorrhage (SAH) are associated with tissue hypoxia and vascular endothelial growth factor (VEGF) release. Soluble Fms-like tyrosine kinase-1 (sFlt-1), a truncated soluble form of VEGF receptor-1, is an endogenous VEGF inhibitor released in response to hypoxia and nitric oxide deficiency. SFlt-1 is anti-angiogenic and mediates endothelial dysfunction. We hypothesize sFlt-1 elevation may be associated with VSP and poor SAH outcome. Methods: We prospectively enrolled consecutive SAH subjects, banked serial blood samples, and evaluated their modified Rankin scores (mRS) at 3 month intervals. Poor functional outcome was defined as mRS&gt;2. Angiographic VSP was defined as &gt;50% reduction in caliber of any vessel on post-SAH day 7 cerebral angiogram. In 63 SAH subjects, we compared plasma sFlt-1 by ELISA on post-SAH days 3 and 5 by VSP and outcome status using Wilcoxon rank sum or Student’s t-test depending on data distribution. Bonferroni correction was used for multiple comparisons. Logistic regression was used to adjust for confounders. Associations were measured using Pearson’s or Spearman’s correlation depending on data distribution. Results: Twenty-seven subjects (43%) had poor 3-month outcome and 31 (49%) developed VSP. Elevated sFlt-1 level on post-SAH day 3 was associated with poor 3-month outcome (p=0.02) while post-SAH day 5 sFlt-1 level showed no association. SFlt-1 levels were not associated with Hunt and Hess (HH) or Fisher grades or with VSP. SFlt-1 was inversely correlated to VEGF (p=0.04, r=0.26). Post-SAH day 3 sFlt-1 level was independently associated with poor SAH outcome after adjustment for HH grade, age, and VEGF level (p=0.03). VEGF was not associated with VSP or SAH outcome. Conclusion: Early elevation of plasma sFlt-1 on post-SAH day 3 is independently associated with poor 3-month SAH outcome after adjustment for clinical predictors of SAH outcome and for VEGF. Inverse correlation suggests possible negative feedback control between sFlt-1 and VEGF in SAH. Future studies are necessary to determine the source of sFlt-1 in SAH and its role in SAH-associated brain injury. Replication in a larger cohort is necessary to validate sFlt-1 as a potential biomarker for SAH outcome.

Cost and resource implications with serum angiogenic factor estimation in the triage of pre‐eclampsia

To analyse the economic and resource implications of using plasma soluble fms-like tyrosine kinase-1 s(Flt1) and placenta growth factor (PlGF) measurements in pre-eclampsia evaluation and management. Retrospective cost analysis of our prospective cohort study. Boston, Massachusetts (USA). Women (n=176) presenting to the hospital at <34weeks of gestation for evaluation of possible pre-eclampsia during 2009-10. Cases without complete cost or outcome data (n=9) and re-enrolments (n=18) were excluded. Modelled comparisons between the standard approach (combination of blood pressure, urinary protein excretion, alanine aminotransferase and platelet counts) and a novel approach (ratio of plasma sFlt1 and PlGF) using actual hospital data converted to 2012 US dollars in accordance with the Centers for Medicare and Medicaid Services. Direct 2-week costs and resource use by groups having true or false positive and negative test results for adverse outcomes according to approach. The improved specificity of the novel approach decreased the proportion of women falsely labelled as test-positive from 42.3% (34.4-50.2%) to 4.0% (0.85-7.15%) and increased the proportion correctly labelled as test-negative from 23.5% (16.7-30.3%) to 61.7% (53.9-69.5%). This could potentially reduce average per-patient costs by $1215. Substantial quantities of resources [47.2% (35.7-58.7%) of antenatal admissions and 72.5% (68.0-77.0%) of tests for fetal wellbeing] were unnecessarily used for women who were truly negative. A proportion of iatrogenic preterm deliveries among women with negative results was potentially avoidable representing further cost and resource savings. Clinical use of the plasma sFlt1 and PlGF ratio improves risk stratification among women presenting for pre-eclampsia evaluation and has the potential to reduce costs and resource use.

Endothelial function progressively deteriorates during normal pregnancy

Objective: To elucidate changes in endothelial function throughout the gestational period in normal pregnancy and its relationship with plasma soluble fms-like tyrosine kinase-1 (sFlt-1) levels. Methods: Endothelial function was evaluated by reactive hyperemia index (RHI) using Endo-PAT2000 and plasma sFlt-1 levels were measured simultaneously by ELISA. Results: RHI gradually deteriorated with increasing gestational age. Plasma sFlt-1 levels exhibited a gradual increase at late pregnancy and were inversely correlated with RHI. Conclusion: Maternal endothelial function gradually deteriorates with increasing gestational age and there is an inverse correlation between endothelial function and plasma sFlt-1 levels in normal pregnancy.

Pravastatin Attenuates Hypertension, Oxidative Stress, and Angiogenic Imbalance in Rat Model of Placental Ischemia-Induced Hypertension

Preeclampsia is a pregnancy-specific condition characterized by an imbalance of circulating angiogenic factors and new-onset hypertension. Although current treatment options are limited, recent studies suggest that pravastatin may improve angiogenic profile and reduce blood pressure in preeclampsia. We hypothesized pravastatin would restore angiogenic balance and reduce mean arterial pressure (MAP) in rats with reduced utero-placental perfusion pressure (RUPP)-induced hypertension. Pravastatin was administered intraperitoneally (1 mg/kg per day) in RUPP (RUPP+P) and normal pregnant rats (NP+P) from day 14 to 19 of pregnancy. On day 19, MAP was measured via catheter, conceptus data were recorded, and tissues collected. MAP was increased (P<0.05) in RUPP compared with NP dams, and pravastatin ameliorated this difference. Pravastatin attenuated decreased fetal weight and plasma vascular endothelial growth factor and the RUPP-induced increased soluble fms-like tyrosine kinase-1 when compared with NP dams. Pravastatin treatment did not improve angiogenic potential in RUPP serum and decreased (P<0.05) endothelial tube formation in NP rats. RUPP rats presented with indices of oxidative stress, such as increased placental catalase activity and plasma thiobarbituric acid reactive substances along with decreased plasma total antioxidant capacity compared with NP controls, and pravastatin attenuated these effects. MAP, fetal weight, plasma vascular endothelial growth factor, and plasma soluble fms-like tyrosine kinase-1 were unchanged in NP+P compared with NP controls. The present data indicate that treatment with pravastatin attenuates oxidative stress and lowers MAP in placental ischemia-induced hypertension, but may have negative effects on circulating angiogenic potential during pregnancy. Further studies are needed to determine whether there are long-term deleterious effects on maternal or fetal health after pravastatin treatment during pregnancy-induced hypertension or preeclampsia.

Low 2‐methoxyestradiol levels at the first trimester of pregnancy are associated with the development of pre‐eclampsia

To determine whether maternal plasma levels of 2-methoxyestradiol (2-ME) are decreased early in pregnancies that subsequently develop pre-eclampsia (PE) and whether this difference could be attributed to the presence of Val158Met catechol-O-methyltransferase (COMT) polymorphism in the placenta. Clinical characteristics and plasma samples were collected at 11 to 14 weeks prospectively in a cohort of patients. From them, 13 PE and 72 control pregnant women were chosen. Plasma soluble fms-like tyrosine kinase1 and placental growth factor levels were measured by electrochemiluminescence and 2-ME was measured by high-performance liquid chromatography with mass spectrometry/mass spectrometry detection. At delivery, placental tissue was collected and the Val158Met COMT polymorphism was determined by restriction fragment length polymorphism-PCR. At 11 to 14 weeks, patients who would develop PE have significantly lower plasma levels of 2-ME than controls [1.9 ± 2 standard error of the mean (SEM) vs 61.7 ± 27 pg/mL, P < 0.05]. The Val158Met polymorphism was more frequent in controls than in PE patients and the placental presence of COMT polymorphism was associated with a decreased risk of developing PE [PE: 23.1% vs control: 66.6%; χ(2) = 10.9, p = 0.0041]. Lower plasma concentrations of 2-ME during early pregnancy in patients who subsequently develop PE were found. Presence of placental Val158Met COMT polymorphism is associated with a decreased risk to develop PE, suggesting a protective role against PE.

Tumor necrosis factor α induces a model of preeclampsia in pregnant baboons (Papio hamadryas)

Preeclampsia is a common disease of pregnancy characterised by maternal hypertension and proteinuria. Abnormal placentation in early pregnancy and abnormal cytokine and anti-angiogenic factor expression are thought to contribute to the clinical syndrome of endothelial dysfunction evident in the second half of gestation. The mechanisms underlying both the placental pathology and its translation to the maternal clinical syndrome are not fully understood. A model of preeclampsia manifest by clinically evident endothelial dysfunction (increased blood pressure and proteinuria) was induced by administration of low-dose TNF-α for 2weeks at mid-gestation in pregnant baboons (Papio hamadryas). Blood pressure was monitored continuously and remotely by intra-arterial radiotelemetry. Following TNF-α infusion, there was an increase in systolic and diastolic blood pressure and development of proteinuria in pregnant treated animals, but not in pregnant saline controls nor in non-pregnant TNF-α treated animals. The treated pregnant animals also developed elevated plasma soluble FMS-like tyrosine kinase-1 (sFLT-1) and increased placental mRNA expression of sFLT-1 and soluble endoglin (sEng). These results clearly demonstrate that the cytokine TNF-α can induce the clinical and biochemical features of human preeclampsia. The results identify a link between cytokines, placental dysfunction and endothelial dysfunction resulting in a loss of maternal blood pressure control.

Maternal plasma soluble fms‐like tyrosine kinase‐1 and free vascular endothelial growth factor at 11 to 13 weeks of gestation in preeclampsia

To investigate the maternal plasma concentration of soluble fms-like tyrosine kinase-1 (sFlt-1) and free vascular endothelial growth factor (free-VEGF) at 11 to 13 weeks of gestation in patients destined to develop preeclampsia (PE) and to examine whether any possible differences in maternal plasma levels are related to uterine artery pulsatility index (PI) and maternal serum placental growth factor (PlGF). Plasma free-VEGF, plasma sFlt-1, serum PlGF and uterine artery PI were measured at 11 to 13 weeks in 90 cases that subsequently developed PE and in 180 unaffected controls. In the majority of cases of PE and controls the levels of free-VEGF were undetectable. In the pregnancies that developed PE, compared to unaffected controls, uterine artery PI was higher, serum PlGF was lower but there was no significant difference in levels of sFlt-1. Measurement of free-VEGF and sFlt-1 in maternal blood at 11 to 13 weeks of gestation is not useful in the prediction of pregnancies destined to develop PE.

Maternal plasma soluble fms‐like tyrosine kinase‐1 and placental growth factor levels as biochemical markers of gestational hypertension for Malaysian mothers

To establish baseline levels of maternal plasma soluble fms-like tyrosine kinase-1 (sFlt-1) and placental growth factor (PlGF) among normotensive Malaysian mothers and to compare the marker levels between normotensives and mothers with gestational hypertension (GH). Plasma sFlt-1 and PlGF were measured by enzyme-linked immunosorbent assay in an unmatched, case-control study. The results were subjected to normality testing and analyzed by Mann-Whitney U-tests. Among normotensive mothers, both sFlt-1and PlGF showed a general increase in levels from the 24th to 32nd weeks of pregnancy. PlGF levels in normotensive mothers with gestational diabetes mellitus were reduced compared to those without the disease, while levels of sFlt-1 were elevated. Mothers with GH had reduced levels of PlGF with increased levels of sFlt-1 when compared to normotensive mothers. Among the normotensive mothers followed up until delivery, the inversed pattern of reduced PlGF and increased sFlt-1 marker levels was found in 40% of those who developed GH later in pregnancy. Plasma levels of sFlt-1and PlGF in normotensive mothers may be influenced by gestational diabetes mellitus and GH. GH mothers show an inversed pattern of marker levels compared to normotensive mothers.