Biomarkers In Amniotic Fluid Research Articles

Amniotic fluid proteomic analysis identifies IL1RL1, APOE, and NECTIN4 as new biomarkers for preterm birth

BackgroundDespite extensive research, the identification of effective biomarkers for early prediction of preterm birth (PTB) continues to be a challenging endeavor. This study aims to identify amniotic fluid (AF) protein biomarkers useful for the early diagnosis of PTB.MethodsWe initially identified the protein expression profiles in the AF of women with PTB (n = 22) and full-term birth (FTB, n = 22), from the First People’s Hospital of Yunnan Province who underwent amniocentesis from November 2019 to February 2020, using mass spectrometry employing the data-independent acquisition (DIA) technique, and then analyzed differentially expressed proteins (DEPs). Subsequently, the least absolute shrinkage and selection operator (LASSO) and random forest analysis were employed to further screen the key proteins for PTB biomarker identification. The receiver operating characteristic (ROC) analysis, calibration plots, and decision curve analyses (DCA) were utilized to assess the discrimination and calibration of the key biomarkers.ResultsA total of 25 DEPs were identified between the PTB and FTB groups, comprising 13 up-regulated and 12 down-regulated proteins. Three key protein biomarkers for early PTB diagnosis were identified: IL1RL1 (interleukin-1 receptor-like 1), APOE (apolipoprotein E), and NECTIN4 (nectin cell adhesion molecule 4). The results of the ROC analysis showed that the area under the curve (AUC) of the three proteins combined as a biomarker for early diagnosis of PTB was 0.913 (95% CI: 0.823-1.000), with a sensitivity of 0.864 and a specificity of 0.955, both superior to those of the individual biomarkers. Bootstrap internal validation revealed a concordance index (C-index) of 0.878, with a sensitivity of 0.812 and a specificity of 0.773, indicating the robust predictive performance of these biomarkers.ConclusionsWe identified three previously unexplored yet potentially useful protein biomarkers in AF for early PTB diagnosis: IL1RL1, APOE, and NECTIN4.

Amniotic fluid biomarkers of fetal stress response to cardiac dysfunction in twin-twin transfusion syndrome

Recipient twin (RT) cardiovascular changes in twin-twin transfusion syndrome (TTTS) are associated with increased mortality. This relationship is likely related to a cascade of pathophysiologic changes in the RT progressing to multisystem organ failure. We sought to evaluate biomarkers of inflammation, stress, and tissue injury within amniotic fluid (AF) of RTs undergoing selective fetoscopic laser photocoagulation (SFLP) for TTTS and examine their relationship with fetal echocardiographic findings. AF samples from RTs were obtained from 39 monochorionic-diamniotic pregnancies complicated by TTTS. Those without preoperative fetal echocardiograms or evidence of selective intrauterine growth restriction were excluded from analysis with 24 remaining samples. 15 commonly utilized clinical biomarkers for inflammation, stress, and tissue injury were assessed using a flow-cytometry based multiplex immunoassay. Fetal echocardiograms were performed prior to SFLP with myocardial performance index (MPI) utilized as a measure of cardiac dysfunction. Pearson’s correlation was used for data analysis. RT Left ventricular MPI (LV-MPI) ranged from 0.28-0.74, with 11/24 RTs demonstrating abnormal MPI for GA (Z-score > 2). RT AF cortisol (r = 0.4527, p = 0.020), IL-6 (r = 0.44, p = 0.025), CRP (r = 0.431, p = 0.028), and myoglobin (r = 0.401, p = 0.042) levels correlated with recipient left ventricular MPI (LV-MPI). AF cortisol strongly correlated with AF myoglobin (r = 0.576, p < 0.001). RT with abnormal LV-MPI exhibited significantly higher AF cortisol levels compared to those with normal LV-MPI (p = 0.029). RT amniotic fluid demonstrates markers of inflammation, stress and cardiac injury in recipient twins with cardiac dysfunction. These markers may be indicators of disease progression in TTTS secondary to cardiac dysfunction. The presence of these markers implicate stress and the immune response in the pathophysiologic response to cardiac dysfunction in TTTS.

Amniotic fluid biomarkers predict the severity of congenital cytomegalovirus infection.

BACKGROUNDCytomegalovirus (CMV) is the most common intrauterine infection, leading to infant brain damage. Prognostic assessment of CMV-infected fetuses has remained an ongoing challenge in prenatal care, in the absence of established prenatal biomarkers of congenital CMV (cCMV) infection severity. We aimed to identify prognostic biomarkers of cCMV-related fetal brain injury.METHODSWe performed global proteome analysis of mid-gestation amniotic fluid samples, comparing amniotic fluid of fetuses with severe cCMV with that of asymptomatic CMV-infected fetuses. The levels of selected differentially excreted proteins were further determined by specific immunoassays.RESULTSUsing unbiased proteome analysis in a discovery cohort, we identified amniotic fluid proteins related to inflammation and neurological disease pathways, which demonstrated distinct abundance in fetuses with severe cCMV. Amniotic fluid levels of 2 of these proteins — the immunomodulatory proteins retinoic acid receptor responder 2 (chemerin) and galectin-3–binding protein (Gal-3BP) — were highly predictive of the severity of cCMV in an independent validation cohort, differentiating between fetuses with severe (n = 17) and asymptomatic (n = 26) cCMV, with 100%–93.8% positive predictive value, and 92.9%–92.6% negative predictive value (for chemerin and Gal-3BP, respectively). CONCLUSIONAnalysis of chemerin and Gal-3BP levels in mid-gestation amniotic fluids could be used in the clinical setting to profoundly improve the prognostic assessment of CMV-infected fetuses.FUNDINGIsrael Science Foundation (530/18 and IPMP 3432/19); Research Fund – Hadassah Medical Organization.

A UPLC-MS/MS method for the determination of oxidative stress biomarkers in amniotic fluid

Oxidative stress in the fetal period is associated with preterm birth as well as short and long-term adverse clinical outcomes. Here, an Ultra-Performance Liquid Chromatography–tandem Mass Spectrometry (UPLC-MS/MS) method for the simultaneous quantification of biomarkers of oxidative stress-derived damage to proteins and DNA in amniotic fluid (AF) samples is presented. Appropriate accuracy and precision levels, as well as sensitivity with limits of detection in the low nanomolar (<2 nM) range were achieved. The analytical method was applied to a set of AF samples and reference ranges of the biomarker panel are presented. Median concentrations of biomarkers of protein oxidation (ortho-, 3-chloro-, and 3-nitrotyrosine) and their precursors (para-tyrosine and phenylalanine) ranged between 0.6 and 3 nM and 23 and 30 μM, respectively, while levels of a biomarker of DNA-oxidation (8-hydroxydeoxyguanosine, 8OHdG) and its precursor (2′-deoxyguanosine) were found to be 0.18 and 3 nM, respectively. Detection frequencies of all metabolites were 100% with exception of 3-chlorotyrosine (3Cl-Tyr) and 8OHdG, that were only detected in 8% of samples. The developed method may be applied in research studies focusing on oxidative stress-related complications during pregnancy.

VP23.09: Oxidative stress biomarkers in amniotic fluid in fetuses with isolated congenital heart defect

VP23.09: Oxidative stress biomarkers in amniotic fluid in fetuses with isolated congenital heart defect

Antibody microarray analysis of the amniotic fluid proteome for predicting the outcome of rescue cerclage in patients with cervical insufficiency.

Little is known about the biomarkers that can identify patient candidates suitable for rescue cerclage procedure. The purpose of the study was to identify novel biomarkers in amniotic fluid (AF) that can predict the outcome of rescue cerclage in patients with cervical insufficiency by using an antibody microarray. This case–control study was conducted using AF samples collected from singleton pregnant women who underwent rescue cerclage following a diagnosis of cervical insufficiency (19–25 weeks). Patients were divided into case (n=20) and control (n=20) groups based on the occurrence of spontaneous preterm delivery (SPTD) at <34 weeks of gestation after cerclage placement. The AF proteomes were analyzed using an antibody microarray for biomarker discovery work. Ten candidate biomarkers of interest were validated by enzyme-linked immunosorbent assay (ELISA). Thirty-one molecules studied showed significant intergroup differences (≥two-fold change in signal intensity). Validation by ELISA confirmed significantly higher levels of a proliferation-inducing ligand (APRIL), S100 calcium-binding protein A8/A9 complex (S100 A8/A9), tissue inhibitors of metalloproteinase-1 (TIMP-1), macrophage inflammatory protein-1α (MIP-1α), and interleukin-8 (IL-8) in women who had SPTD at <34 weeks. Of these, AF S100 A8/A9 and TIMP-1 levels were independent of other potentially confounding factors (e.g., cervical dilatation). S100 A8/A9 had the highest area under the curve (AUC) at 0.857. Using protein–antibody microarray technology, we identified differentially expressed proteins (DEPs) and several novel biomarkers (APRIL, IL-8, MIP-1α, S100 A8/A9, and TIMP-1) in AF from women who had SPTB at <34 weeks after cerclage for cervical insufficiency. These data can provide an insight into the molecular mechanisms underlying SPTD after rescue cerclage in patients with cervical insufficiency.

Analysis of Biomarkers for Congenital Heart Disease Based on Maternal Amniotic Fluid Metabolomics.

Congenital heart disease (CHD) is the most common birth defect. The prenatal diagnosis of fetal CHD is completely dependent on ultrasound testing, but only ~40% of CHD can be detected. The purpose of this study is to find good biomarkers in amniotic fluid (AF) to detect CHD in the second trimester, so as to better manage this group of people and reduce the harm of CHD to the fetus. Metabolites analysis were performed in two separate sets. The discovery set consisted of 18 CHD fetal maternal AF samples and 35 control samples, and the validation set consisted of 53 CHD fetal maternal AF samples and 114 control samples. Untargeted metabolite profiles were analyzed by gas chromatography/time-of-flight-mass spectrometry (GC-TOF/MS). Orthogonal partial least square discrimination analysis (OPLS-DA) demonstrated that CHD and control samples had significantly different metabolic profiles. Two metabolites, uric acid and proline, were significantly elevated in CHD and verified in two data sets. Uric acid was associated with CHD [odds ratio (OR): 7.69 (95% CI: 1.18–50.13) in the discovery set and 3.24 (95% CI:1.62–6.48) in the validation set]. Additionally, uric acid showed moderate predictive power; the area under curve (AUC) was 0.890 in the discovery set and 0.741 in the validation set. The sensitivity and specificity of uric acid to detect CHD was, respectively, 94.4 and 74.3% in the discovery set and 67.9 and 71.9% in the validation set. The identification of uric acid as a biomarker for CHD has the potential to stimulate research on the pathological mechanism of CHD and the development of a diagnostic test for clinical applications.

Maternal Plasma and Amniotic Fluid LBP, Pentraxin 3, Resistin, and IGFBP-3: Biomarkers of Microbial Invasion of Amniotic Cavity and/or Intra-amniotic Inflammation in Women with Preterm Premature Rupture of Membranes.

BackgroundWe sought to determine whether lipopolysaccharide binding protein (LBP), pentraxin 3, resistin, and insulin-like growth factor binding protein (IGFBP)-3 in plasma and amniotic fluid (AF) can predict microbial invasion of the amniotic cavity (MIAC), intra-amniotic inflammation (IAI), and microbial-associated IAI in women with preterm premature rupture of membranes (PPROM).MethodsThis was a retrospective cohort study involving 168 singleton pregnant women with PPROM. AF obtained via amniocentesis was cultured and assayed for interleukin (IL)-6 to define IAI and for IL-8 to compare with AF biomarkers. Plasma samples were collected at the time of amniocentesis, and C-reactive protein (CRP) levels in serum were compared with plasma biomarkers. The stored plasma and AF samples were assayed for LBP, pentraxin 3 (PTX3), resistin, and IGFBP-3 by ELISA.ResultsMultivariate logistic regression analysis revealed that: 1) elevated plasma and AF levels of LBP were independently associated with increased risks of MIAC, IAI, and microbial-associated IAI; 2) elevated AF, but not plasma, PTX3, and resistin levels were independently associated with increased risks of MIAC, IAI, and microbial-associated IAI; 3) decreased IGFBP-3 levels in the plasma were independently associated with only IAI, whereas those in the AF were associated with only microbial-associated IAI. Among the tested biomarkers, AF PTX3 and resistin had the highest predictive performance for MIAC, IAI, and microbial-associated IAI (area under the curves [AUC] = 0.85–0.95), which is similar to the performance of AF IL-8. The AUCs of the plasma LBP and IGFBP-3 were similar to that of serum CRP with respect to IAI.ConclusionMaternal plasma LBP and IGFBP-3 are potential biomarkers for the non-invasive identification of IAI in women with PPROM, with a similar accuracy to the serum CRP level. AF LBP, PTX3, resistin, and IGFBP-3 may be involved in the intra-amniotic inflammatory responses in PPROM complicated by MIAC.

Inflammatory and Angiogenic Mediators in Amniotic Fluid Are Associated With the Development of Retinopathy of Prematurity in Preterm Infants.

PurposeTo investigate whether elevated levels of inflammatory/angiogenic and growth mediators in amniotic fluid (AF) and the presence of intra-amniotic infection are associated with the occurrence and progression of retinopathy of prematurity (ROP) in preterm infants.MethodsThis retrospective cohort study included 175 premature singleton infants who were born between 23+0 and 32+0 weeks. AF obtained via amniocentesis was cultured, and endoglin, endostatin, insulin-like growth factor-binding protein (IGFBP)-2, IGFBP-3, IGFBP-4, IL-6, IL-8, matrix metalloproteinase-8, matrix metalloproteinase-9, and vascular endothelial growth factor receptor-1 levels were assayed by ELISA. The primary outcome measures included the occurrence of any stage ROP, severe ROP (stage ≥3), and vision-threatening type 1 ROP requiring treatment.ResultsMultiple logistic regression analyses revealed that there are significant associations between elevated AF endoglin levels and ROP occurrence; between elevated AF endoglin, endostatin, and IGFBP-2 levels and severe ROP; and between high AF endoglin, IL-6, and IL-8 levels and vision-threatening ROP requiring treatment, after adjusting for potential postnatal confounders. Using stepwise regression analyses, antenatal prediction models based on these AF biomarkers and prenatal factors were developed for the ROP outcomes, which had good discriminatory power (area under the curves, 0.731–0.863). However, we found that intra-amniotic infection is not associated with ROP occurrence and progression.ConclusionsElevated levels of inflammatory (IL-6 and IL-8) and angiogenic (endoglin and IGFBP-2) mediators in the AF, but not the presence of intra-amniotic infection, are independently associated with the occurrence and progression of ROP in preterm infants. These findings suggest that the pathophysiologic events that predispose preterm neonates to ROP may begin before delivery.

Proteomic Analysis of Amniotic Fluid Proteins for Predicting the Outcome of Emergency Cerclage in Women with Cervical Insufficiency.

We aimed to identify novel biomarkers in amniotic fluid (AF) that predict the outcome of emergency cerclage in women with cervical insufficiency. This retrospective cohort study included 40 singleton pregnant women who received emergency cerclage for cervical insufficiency (17-25weeks) and underwent amniocentesis. Label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to identify AF proteins in pooled samples (n = 16) using a nested case-control approach. The six candidate biomarkers of interest were validated by enzyme-linked immunosorbent assays (ELISA) in the final cohort (n = 40). The differentially expressed proteins (DEPs) were analyzed by pathway analysis software. The primary outcome measure was failure of emergency cerclage [defined as spontaneous preterm delivery (SPTD) at < 34weeks of gestation after cerclage placement]. Sixty-eight proteins were differentially expressed (P < 0.001) in AF from SPTD cases and near-term controls, of which 44 (64.7%) were upregulated and 24 (35.3%) were downregulated. Validation by ELISA confirmed that AF from women with cerclage failure contained significantly higher levels of myeloperoxidase, lactoferrin, glucose-6-phosphate isomerase, lipocalin-2, and lymphocyte cytosolic protein 1, the first four of which were independent of cervical dilatation at presentation. The five pathways with the most differentially regulated proteins were actin cytoskeleton signaling, acute phase response signaling, ILK signaling, glycolysis, and gluconeogenesis. Proteomic analyses of AF in this study identified DEPs and specific protein pathways related to poor prognosis after emergency cerclage for cervical insufficiency. Four novel independent biomarkers in AF for cerclage failure have been identified using proteomics.

The Amniotic Fluid Proteome Differs Significantly between Donor and Recipient Fetuses in Pregnancies Complicated by Twin-to-Twin Transfusion Syndrome.

BackgroundTwin-to-twin transfusion syndrome (TTTS) is a serious complication of monochorionic twin pregnancies. It results from disproportionate blood supply to each fetus caused by abnormal vascular anastomosis within the placenta. Amniotic fluid (AF) is an indicator reflecting the various conditions of the fetus, and an imbalance in AF volume is essential for the antenatal diagnosis of TTTS by ultrasound. In this study, two different mass spectrometry quantitative approaches were performed to identify differentially expressed proteins (DEPs) within matched pairs of AF samples.MethodsWe characterized the AF proteome in pooled AF samples collected from donor and recipient twin pairs (n = 5 each) with TTTS by a global proteomics profiling approach and then preformed the statistical analysis to determine the DEPs between the two groups. Next, we carried out a targeted proteomic approach (multiple reaction monitoring) with DEPs to achieve high-confident TTTS-associated AF proteins.ResultsA total of 103 AF proteins that were significantly altered in their abundances between donor and recipient fetuses. The majority of upregulated proteins identified in the recipient twins (including carbonic anhydrase 1, fibrinogen alpha chain, aminopeptidase N, alpha-fetoprotein, fibrinogen gamma chain, and basement membrane-specific heparan sulfate proteoglycan core protein) have been associated with cardiac or dermatologic disease, which is often seen in recipient twins as a result of volume overload. In contrast, proteins significantly upregulated in AF collected from donor twins (including IgGFc-binding protein, apolipoprotein C-I, complement C1q subcomponent subunit B, apolipoprotein C-III, apolipoprotein A-II, decorin, alpha-2-macroglobulin, apolipoprotein A-I, and fibronectin) were those previously shown to be associated with inflammation, ischemic cardiovascular complications or renal disease.ConclusionIn this study, we identified proteomic biomarkers in AF collected from donor and recipient twins in pregnancies complicated by TTTS that appear to reflect underlying functional and pathophysiological challenges faced by each of the fetuses.

Cardiovascular Biomarkers in Amniotic Fluid, Umbilical Arterial Blood, Umbilical Venous Blood, and Maternal Blood at Delivery, and Their Reference Values for Full-Term, Singleton, Cesarean Deliveries.

Background: Several cardiovascular biomarkers have regulatory functions in perinatal physiology.Aim: This study aimed to analyze the feto-maternal distribution pattern of biomarkers in samples of amniotic fluid, umbilical arterial blood, umbilical venous blood, and maternal blood samples, and to establish reference values. Each linked sample set consisted of the combined samples obtained in an individual pregnancy.Study design: We performed a prospective, observational, cross-sectional, single-center study.Subjects: The sample cohort included 189 neonates who were born to 170 mothers. A total of 162/189 neonates were full term and 129/189 were delivered by elective cesarean section.Outcome measures: Midregional pro-adrenomedullin (MRproADM [nmol/L]), midregional pro-atrial natriuretic peptide (MRproANP [pmol/L]), brain natriuretic peptide (BNP [pg/mL]), N-terminal pro-brain natriuretic peptide (NTproBNP [pg/mL]), copeptin [pmol/L], and high-sensitive troponin I (hsTnI [pg/mL]) levels were measured.Results: In singleton, full-term, primary cesarean deliveries (n = 91), biomarker levels (median, [IQR]) at delivery were as follows. MRproADM levels in umbilical arterial blood/umbilical venous blood/amniotic fluid/maternal blood were 0.88 (0.20)/0.95 (0.18)/2.80 (1.18)/1.10 (0.54), respectively. MRproANP levels were 214.23 (91.38)/216.03 (86.15)/0.00 (3.82)/50.67 (26.81), respectively. BNP levels were 14.60 (25.18)/22.08 (18.91)/7.15 (6.01)/6.20 (18.23), respectively. NTproBNP levels were 765.48 (555.24)/816.45 (675.71)/72.03 (55.58)/44.40 (43.94), respectively. Copeptin levels were 46.17 (290.42)/5.54 (9.08)/9.97 (7.44)/4.61 (4.59), respectively. Levels of hsTnI were 6.20 (4.25)/5.60 (5.01)/0.45 (1.73)/2.50 (2.40), respectively.Conclusion: We determined reference values for biomarkers in term neonates delivered by primary cesarean section in amniotic fluid, umbilical arterial and venous blood, and maternal blood. Biomarkers in the fetal circulation appear to be of primary fetal origin, except for MRproADM.

The combination of maternal blood and amniotic fluid biomarkers improves the predictive accuracy of histologic chorioamnionitis

IntroductionThis study was performed to determine whether the combination of maternal blood and amniotic fluid biomarkers can improve the predictive accuracy of histologic chorioamnionitis (HC). MethodsThis retrospective study included 80 singleton pregnant women who were suspected to have intrauterine infection and underwent measurement of two maternal blood biomarkers [maternal white blood cell count (mWBC) and maternal C-reactive protein level (mCRP)] and three amniotic fluid biomarkers [amniotic white blood cell count (aCell), amniotic glucose level (aGlucose), and amniotic lactate dehydrogenase level (aLDH)]. We divided the patients into two groups based on the presence or absence of HC and assessed the predictors of HC using logistic regression models: Model 1, combination of mWBC and mCRP; Model 2, combination of Model 1 and aGlucose; and Model 3, combination of Model 2, aCell, and aLDH. ResultsThe multivariable analysis showed that aCell was the only significant predictor of HC [odds ratio, 1.24; 95% confidence interval (CI), 1.06–1.68] independent of mWBC, mCRP, aGlucose, and aLDH. The c-statistics were higher in Model 3 (0.803; 95% CI, 0.701–0.905) than Model 1 (0.634; 95% CI, 0.511–0.758) and Model 2 (0.785; 95% CI, 0.684–0.887). DiscussionWe found that the combination of maternal blood and amniotic fluid biomarkers can improve the predictive accuracy of HC. Therefore, our data provide relevant information to support counseling with regard to improving the predictive accuracy of HC in patients with suspected intrauterine infection.

The association of second trimester biomarkers in amniotic fluid and fetal outcome

Objective: To identify the level of amniotic fluid lactate (AFL), placental growth factor (PLGF), and vascular endothelial growth factor (VEGF) at second trimester amniocentesis, and to compare levels in normal pregnancies with pregnancies ending in a miscarriage, an intrauterine growth restricted fetus (IUGR) or decreased fetal movements.Study design: A prospective cohort study. Amniotic fluid was consecutively collected at amniocentesis in 106 pregnancies. Fetal wellbeing at delivery was evaluated from medical files and compared with the levels of AFL, VEGF, and PLGF at the time of amniocentesis.Results: The median level of AFL was 6.9 mmol/l, VEGF 0.088 pg/ml, and PLGF 0.208 pg/ml. The median levels of AFL in pregnancies ended in miscarriage were significantly higher (10.7 mmol/l) compared to those with a live new-born (6.9 mmol/L, p = .02). The levels of VEGF (p = .2) and PLGF (p = .7) were not affected. In pregnancies with an IUGR, the median level of AFL was higher compared to those with normal fetal growth (p = .003). No differences VEGF (p = .5), but significant lower PLGF were found in IUGR pregnancies (p = .03).Conclusions: Pregnancies ending in a miscarriage or with IUGR had significantly higher median values of AFL but lower values of PLGF in the amniotic fluid at the time of second trimester amniocentesis compared to normal pregnancies.

87: Biomarkers in amniotic fluid at the time of fetal myelomeningocele surgery predict premature rupture of membranes

87: Biomarkers in amniotic fluid at the time of fetal myelomeningocele surgery predict premature rupture of membranes

Oxidative stress biomarkers in amniotic fluid of pregnant women with hypothyroidism

Purpose Hypothyroidism in pregnancy is the serious state that may lead to fetal morbidity and mortality. Oxidative stress biomarkers in the amniotic fluid can provide important information on the health, development and maturation of the fetus during pregnancy. In this study, we examined whether maternal hypothyroidism contributes to increased oxidative stress biomarkers in the amniotic fluid during the first trimester of pregnancy. Materials and methods The study was conducted on healthy pregnant women and pregnant women with hypothyroidism (gestational age: 16–18 weeks). Oxidative stress biomarkers, such as superoxide anion (O2 •−), hydrogen peroxide (H2O2), nitric oxide (NO), peroxynitrite (ONOO−), lipid peroxide (LPO), reduced glutathione (GSH) and oxidized glutathione (GSSG) were assayed in the amniotic fluid. Results The results of this study indicated that concentrations of O2 •− and NO are significantly higher, while the concentration of H2O2 is significantly lower in the amniotic fluid of pregnant women with hypothyroidism in comparison to healthy pregnant women. There were no differences in concentrations of LPO, GSH and GSSG among tested groups. Also, we found that amniotic fluid concentration of O2 •− is negatively correlated with the body weight and Apgar score values of the newborns. Conclusion These results suggest that pregnancy hypothyroidism is characterized by the amniotic fluid oxidative stress. Incorporation of the oxidative stress biomarkers measurement in the amniotic fluid may be of clinical importance in the management of pregnancy hypothyroidism.

Amniotic Fluid Infection in Preterm Pregnancies with Intact Membranes.

Introduction. Intra-amniotic infection (IAI) is a major cause of preterm labor and adverse neonatal outcome. We evaluated amniotic fluid (AF) proteolytic cascade forming biomarkers in relation to microbial invasion of the amniotic cavity (MIAC) and IAI in preterm pregnancies with intact membranes. Material and Methods. Amniocentesis was made to 73 women with singleton pregnancies; 27 with suspected IAI; and 46 controls. AF biomarkers were divided into three cascades: Cascade 1: matrix metalloproteinase-8 (MMP-8), MMP-9, myeloperoxidase (MPO), and interleukin-6; Cascade 2: neutrophil elastase (HNE), elafin, and MMP-9; Cascade 3: MMP-2, tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), MMP-8/TIMP-1 molar ratio, and C-reactive protein (CRP). MMP-8 was measured by an immunoenzymometric assay and the others were measured by ELISA. Standard biochemical methods, molecular microbiology, and culture techniques were used. Results. MMP-8, MMP-9, MPO, elafin, and TIMP-1 concentrations were higher in IAI suspected cases compared to controls and also in IAI suspected cases with MIAC compared to those without MIAC when adjusted by gestational age at amniocentesis. All biomarkers except elafin and MMP-2 had the sensitivity of 100% with thresholds based on ROC-curve. Odd ratios of biomarkers for MIAC were 1.2-38 and 95% confidential intervals 1.0-353.6. Conclusions. Neutrophil based AF biomarkers were associated with IAI and MIAC.

Prediction of preterm delivery by second trimester inflammatory biomarkers in the amniotic fluid

ObjectiveTo search for a correlation between mid-pregnancy altered levels of inflammatory markers and preterm delivery. MethodsA prospective cohort series included 39 patients undergoing amniocentesis one additional milliliter of amniotic fluid (AF) was stored for later dosage of interleukin-6 (Il-6), matrix metalloproteinase-9 (MMP-9), glucose and C-reactive protein (CRP). Maternal serum CRP and glucose levels were also obtained. Exclusion criteria were multiple pregnancies, chorioamnionitis, group B streptococcus colonization, bacterial vaginosis and cases with proven aneuploidy. We searched for correlation between AF and plasmatic markers and also for a difference between patients with term and preterm delivery. Results33 participants were eligible and one third had preterm delivery. Levels of the plasmatic biomarkers did not correlate with the AF biomarkers except for plasmatic glucose and AF IL-6 levels (r=0.350; p=0.016). The levels of all AF biomarkers did not differ significantly between the pre-term and the term groups (p>0.05). The optimal screening cutoffs for identifying pregnancies at risk were different than the ones initially indicated. ConclusionMid-pregnancy amniotic fluid biomarker levels do not correlate with preterm delivery. Plasma CRP is not correlated with these markers. Cutoff levels suggested are sparse and heterogeneous. Larger studies are needed before advising routine measurement of these markers.

Evaluation of kidney injury biomarkers in rat amniotic fluid after gestational exposure to cadmium.

Cadmium is a well-characterized nephrotoxic agent that is also capable of accumulating and diffusing across the placenta; however, only a few studies have addressed its effects over fetal kidneys and none of them has used a panel of sensitive and specific biomarkers for the detection of kidney injury. The goal of this study was to determine cadmium renal effects in rat fetuses by the quantification of early kidney injury biomarkers. Pregnant Wistar rats were exposed by inhalation to an isotonic saline solution or to CdCl2 solution (DDel =1.48 mg Cd kg(-1) day(-1) ) during gestational days (GD) 8-20. On GD 21, dams were euthanized and samples obtained. Kidney injury biomarkers were quantified in amniotic fluid samples and fetal kidneys were microscopically evaluated to search for histological alterations. Our results showed that cadmium exposure significantly raised albumin, osteopontin, vascular endothelial growth factor and tissue inhibitor of metalloproteinases-1 levels in amniotic fluid, whereas it decreased creatinine. Clusterin, calbindin and IFN-inducible protein 10 did not show any change. Accordingly, histological findings showed tubular damage and precipitations in the renal pelvis. In conclusion, gestational exposure to cadmium induces structural alterations in fetal renal tissue that can be detected by some kidney injury biomarkers in amniotic fluid samples. Copyright © 2016 John Wiley & Sons, Ltd.

Two-Dimensional Differential Gel Electrophoresis to Identify Protein Biomarkers in Amniotic Fluid of Edwards Syndrome (Trisomy 18) Pregnancies

BackgroundEdwards syndrome (ES) is a severe chromosomal abnormality with a prevalence of about 0.8 in 10,000 infants born alive. The aims of this study were to identify candidate proteins associated with ES pregnancies from amniotic fluid supernatant (AFS) using proteomics, and to explore the role of biological networks in the pathophysiology of ES.MethodsAFS from six second trimester pregnancies with ES fetuses and six normal cases were included in this study. Fluorescence-based two-dimensional difference gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS) were used for comparative proteomic analysis. The identified proteins were further validated by Western blotting and the role of biological networks was analyzed.ResultsTwelve protein spots were differentially expressed by more than 1.5-fold in the AFS of the ES pregnancies. MALDI-TOF/MS identified one up-regulated protein: apolipoprotein A1 (ApoA1), and four under-regulated proteins: vitamin D binding protein (VDBP), alpha-1-antitrypsin (A1AT), insulin-like growth factor-binding protein 1 (IGFBP-1), and transthyretin (TTR). Western blot and densitometric analysis of ApoA1, A1AT, IGFBP-1, and TTR confirmed the alteration of these proteins in the amniotic fluid samples. Biological network analysis revealed that the proteins of the ES AFS were involved mainly in lipid and hormone metabolism, immune response, and cardiovascular disease.ConclusionsThese five proteins may be involved in the pathogenesis of ES. Further studies are needed to explore.