Dataset Information

Quantitative proteomic profiling of Cervicovaginal fluid from pregnant women with term and preterm birth.

ABSTRACT:

Background

Preterm birth (PTB) is one of major causes of perinatal mortality and neonatal morbidity, but knowledge of its complex etiology is still limited. Here we present cervicovaginal fluid (CVF) protein profiles of pregnant women who subsequently delivered at spontaneous preterm or term, aiming to identify differentially expressed CVF proteins in PTB and term birth.

Methods

The CVF proteome of women who sequentially delivered at preterm and term was analyzed using isobaric tags for relative and absolute quantitation (iTRAQ) coupled with two-dimensional nanoflow liquid chromatography-tandem mass spectrometry (2D-nLC-MS/MS). We compared the CVF proteome of PTB (n?=?5) and control subjects (term birth, n?=?7) using pooled control CVF (term birth, n?=?20) as spike-in standard.

Results

We identified 1294 CVF proteins, of which 605 were newly identified proteins. Of 990 proteins quantified in both PTB and term birth, 52 proteins were significantly up/down-regulated in PTB compared to term birth. The differentially expressed proteins were functionally associated to immune response, endopeptidase inhibitors and structural constituent of cytoskeleton. Finally, we confirm the down-regulation of SERPINB7 (a serine-type protease inhibitor) in PTB compared to control by Western blot.

Conclusions

Taken together, our study provide quantitative CVF proteome profiles of pregnant women who ultimately delivered at preterm and term. These promising results could help to improve the understanding of PTB etiology and to discover biomarkers for asymptomatic PTB.

SUBMITTER: Kim YE

PROVIDER: S-EPMC7885372 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Publications

Quantitative proteomic profiling of Cervicovaginal fluid from pregnant women with term and preterm birth.

Kim Young Eun YE Kim Kwonseong K Oh Han Bin HB Lee Sung Ki SK Kang Dukjin D

Proteome science 20210215 1

<h4>Background</h4>Preterm birth (PTB) is one of major causes of perinatal mortality and neonatal morbidity, but knowledge of its complex etiology is still limited. Here we present cervicovaginal fluid (CVF) protein profiles of pregnant women who subsequently delivered at spontaneous preterm or term, aiming to identify differentially expressed CVF proteins in PTB and term birth.<h4>Methods</h4>The CVF proteome of women who sequentially delivered at preterm and term was analyzed using isobaric ta ...[more]

PMID: 33588889

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Project description:Changes in vaginal microbiota that is associated with preterm birth (PTB) leave specific metabolite fingerprints that can be detected in the cervicovaginal fluid (CVF) using metabolomics techniques. In this study, we characterize and validate the CVF metabolite profile of pregnant women presenting with symptoms of threatened preterm labor (PTL) by both 1H-nuclear magnetic resonance spectroscopy (NMR) and enzyme-based spectrophotometry. We also determine their predictive capacity for PTB, singly, and in combination, with current clinical screening tools - cervicovaginal fetal fibronectin (FFN) and ultrasound cervical length (CL). CVF was obtained by high-vaginal swabs from 82 pregnant women with intact fetal membranes presenting between 24 and 36?weeks gestation with symptoms of threatened, but not established, PTL. Dissolved CVF samples were scanned with a 400?MHz NMR spectrometer. Acetate and other metabolites were identified in the NMR spectrum, integrated for peak area, and normalized to the total spectrum integral. To confirm and validate our observations, acetate concentrations (AceConc) were also determined from a randomly-selected subset of the same samples (n?=?57), by spectrophotometric absorption of NADH using an acetic acid assay kit. CVF FFN level, transvaginal ultrasound CL, and vaginal pH were also ascertained. Acetate normalized integral and AceConc were significantly higher in the women who delivered preterm compared to their term counterparts (P?=?0.002 and P?=?0.006, respectively). The 1H-NMR-derived acetate integrals were strongly correlated with the AceConc estimated by spectrophotometry (r?=?0.69; P?<?0.0001). Both methods were equally predictive of PTB <37?weeks (acetate integral: AUC?=?0.75, 95% CI?=?0.60-0.91; AceConc: AUC?=?0.74, 95% CI?=?0.57-0.90, optimal predictive cutoff of >0.53?g/l), and of delivery within 2?weeks of the index assessment (acetate integral: AUC?=?0.77, 95% CI?=?0.58-0.96; AceConc: AUC?=?0.68, 95% CI?=?0.5-0.9). The predictive accuracy of CVF acetate was similar to CL and FFN. The combination of CVF acetate, FFN, and ultrasound CL in a binary logistic regression model improved the prediction of PTB compared to the three markers individually, but CVF acetate offered no predictive improvement over ultrasound CL combined with CVF FFN. Elevated CVF acetate in women with symptoms of PTL appears predictive of preterm delivery, as well as delivery within 2?weeks of presentation. An assay of acetate in CVF may prove of clinical utility for predicting PTB.

Project description:OBJECTIVES:Perturbation of the choriodecidual space before the onset of spontaneous preterm birth (sPTB) could lead to a concomitant rise in both cervicovaginal fluid (CVF) cytokine and fetal fibronectin (FFN), and assessing the concentrations of both markers could improve the prediction of sPTB (delivery before 37 completed weeks of gestation). Therefore, we prospectively determined mid-trimester changes in CVF cytokine and FFN concentrations, and their predictive capacity for sPTB in asymptomatic pregnant women. STUDY DESIGN:CVF collected at 20+0-22+6 weeks (n?=?47: Preterm-delivered?=?22, Term-delivered?=?25) and 26+0-28+6 weeks (n?=?50: Preterm-delivered?=?17, Term-delivered?=?33) from 63 asymptomatic pregnant women at risk of sPTB were examined. Cytokine and FFN concentrations were determined by multiplexed bead-based immunoassay and 10Q Rapid analysis (Hologic, MA, USA) respectively. The 20+0-22+6/26+0-28+6 weeks ratios of cytokines and FFN concentrations were compared between preterm- and term-delivered women using Receiver Operating Characteristics curves to predict sPTB. Also, bacterial 16S rDNA from 64 samples (20+0-22+6 weeks n?=?36, 26+0-28+6 weeks n?=?28) was amplified by polymerase chain reaction to determine associations between vaginal microflora, cytokine and FFN concentrations. RESULTS:Changes in RANTES and IL-1? concentrations between 20+0-22+6 and 26+0-28+6 weeks, expressed as a ratios, were predictive of sPTB, RANTES (AUC?=?0.82, CI?=?0.62-0.94) more so than IL-1? (AUC?=?0.71, CI?=?0.53-0.85) and FFN (not predictive). Combining these markers (AUC?=?0.83, CI?=?0.63-0.95) showed similar predictive capacity as RANTES alone. FFN concentrations at 26+0-28+6 weeks correlated with IL-1? (r?=?0.4, P?=?0.002) and RANTES (r?=?0.3, P?=?0.03). In addition, there was increased prevalence of vaginal anaerobes including Bacteroides, Fusobacterium and Mobiluncus between gestational time points in women who experienced sPTB compared to the term women (P?=?0.0006). CONCLUSIONS:CVF RANTES and IL-1? in mid-trimester of pregnancy correlate with quantitative FFN. The levels of CVF RANTES and IL-1? decline significantly in women who deliver at term unlike women who deliver preterm. This observation suggests that sPTB may be characterised by sustained choriodecidual inflammation and may have clinical value in serial screening for sPTB if confirmed by larger studies.

Project description:BackgroundPreterm birth is a leading cause of death in children under the age of five. The risk of preterm birth is increased by maternal HIV infection as well as by certain antiretroviral regimens, leading to a disproportionate burden on low- and medium-income settings where HIV is most prevalent. Despite decades of research, the mechanisms underlying spontaneous preterm birth, particularly in resource limited areas with high HIV infection rates, are still poorly understood and accurate prediction and therapeutic intervention remain elusive.ObjectivesMetabolomics was utilized to identify profiles of preterm birth among pregnant women living with HIV on two different antiretroviral therapy (ART) regimens.MethodsThis pilot study comprised 100 mother-infant dyads prior to antiretroviral initiation, on zidovudine monotherapy or on protease inhibitor-based antiretroviral therapy. Pregnancies that resulted in preterm births were matched 1:1 with controls by gestational age at time of sample collection. Maternal plasma and blood spots at 23-35 weeks gestation and infant dried blood spots at birth, were assayed using an untargeted metabolomics method. Linear regression and random forests classification models were used to identify shared and treatment-specific markers of preterm birth.ResultsClassification models for preterm birth achieved accuracies of 95.5%, 95.7%, and 80.7% in the untreated, zidovudine monotherapy, and protease inhibitor-based treatment groups, respectively. Urate, methionine sulfone, cortisone, and 17α-hydroxypregnanolone glucuronide were identified as shared markers of preterm birth. Other compounds including hippurate and N-acetyl-1-methylhistidine were found to be significantly altered in a treatment-specific context.ConclusionThis study identified previously known as well as novel metabolomic features of preterm birth in pregnant women living with HIV. Validation of these models in a larger, independent cohort is necessary to ascertain whether they can be utilized to predict preterm birth during a stage of gestation that allows for therapeutic intervention or more effective resource allocation.

Project description:Spontaneous preterm birth (PTB) before 37 completed weeks of gestation resulting from preterm labor (PTL) is a leading contributor of perinatal morbidity and mortality. Early identification of at-risk women by reliable screening tests could alleviate this health issue; however, conventional methods such as obstetric history and clinical risk factors, uterine activity monitoring, biochemical markers, and cervical sonography for screening women at risk for PTB have proven unsuccessful in lowering the rate of PTB. Cervicovaginal fluid (CVF) might prove to be a useful, readily available biological fluid for identifying diagnostic PTB biomarkers. Human columnar epithelial endocervical-1 (End1) and vaginal (Vk2) cell secretomes were employed to generate a stable isotope labeled proteome (SILAP) standard to facilitate characterization and relative quantification of proteins present in CVF. The SILAP standard was prepared using stable isotope labeling by amino acids in cell culture (SILAC) of End1 and Vk2 through seven passages. The labeled secreted proteins from both cell lines were combined and characterized by liquid-chromatography-tandem mass spectrometry (LC-MS/MS). In total, 1211 proteins were identified in the End1-Vk2 SILAP standard, with 236 proteins being consistently identified in each of the replicates analyzed. Individual proteins were found to contain <0.5% of the endogenous unlabeled forms. Identified proteins were screened to provide a set of 15 candidates that have either previously been identified as potential PTB biomarkers or could be linked mechanistically to PTB. Stable isotope dilution LC-multiple reaction monitoring (MRM/MS) assays were then developed for conducting relative quantification of the 15 candidate biomarkers in human CVF samples from term and PTB cases. Three proteins were significantly elevated in PTB cases (desmoplakin isoform 1, stratifin, and thrombospondin 1 precursor), providing a foundation for further validation in larger patient cohorts.

Dataset Information

Quantitative proteomic profiling of Cervicovaginal fluid from pregnant women with term and preterm birth.

Background

Methods

Results

Conclusions

Publications

Quantitative proteomic profiling of Cervicovaginal fluid from pregnant women with term and preterm birth.

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