Preterm Birth Is Associated With Immune Dysregulation Which Persists in Infants Exposed to Histologic Chorioamnionitis.

Introduction: Preterm infants are at increased risk of exposure to histologic chorioamnionitis (HCA) when compared to term-born controls, and this is associated with several neonatal morbidities involving brain, lungs and gut. Preterm infants could benefit from immunomodulatory therapies in the perinatal period, but development of rational treatment strategies requires improved characterization of the perinatal response to HCA. We had two objectives: The first, to characterize the umbilical cord blood immune profile in preterm infants compared to term-born controls; the second, to investigate the postnatal immune response in preterm infants exposed to HCA versus those who were not. Population: For objective one 59 term infants [mean gestational age (GA) 39+4 (37+3 to 42+0)] and 55 preterm infants [mean GA29+0(23+3 to 32+0)] with umbilical cord samples available were included; for objective two we studied 96 preterm infants [mean GA29+1(23+2 to 32+0)] for whom placental histology and postnatal blood samples were available. Methods: Placental histopathology was used to identify reaction patterns indicative of HCA, and a customized immunoassay of 24 inflammatory markers and trophic proteins selected to reflect the perinatal immune response was performed on umbilical cord blood in term and preterm participants and postnatal day 5 blood in the preterm group. Results: The umbilical cord blood immune profile classified gestational age category with 86% accuracy (95% CI 0.78-0.92), p-value=1.242x10-14. Pro-inflammatory proteins IL-6, MCP-1 and CRP were elevated in the cord blood of preterm infants whilst BDNF, C3, C9, IL-18, MMP-9 and RANTES were decreased, compared to infants born at term. In preterm infants, exposure to HCA was associated with elevations in 8 immune proteins on postnatal day 5 (BDNF, C3, C5a, C9, IL-8, MCP-1, MIP-1ß and MMP-9) when compared to preterm infants who were not exposed. Conclusion: Preterm birth is associated with a distinct immune profile in umbilical cord blood and preterm infants exposed to HCA with evidence of a fetal inflammatory response have specific alterations in immune function that are apparent on day 5 of postnatal life.

Acute chorioamnionitis and intra-amniotic inflammation are more severe according to outside-in neutrophil migration within the same chorio-decidua.

OBJECTIVE: No information exists about whether acute histologic chorioamnionitis (acute-HCA) is more advanced and severe, and intra-amniotic inflammation is more frequent and intense according to outside in neutrophil migration within the same chorio-decidua. The objective of current study is to examine this issue. MATERIALS AND METHODS: We included 106 singleton preterm-births (gestational age at delivery: 20-34 weeks) due to either preterm-labor or preterm-PROM in the context of acute chorio-deciduitis. Study-population was divided into 3 groups according to outside-in neutrophil migration within chorio-decidua as follows: 1) group-1: 'inflammation restricted to the decidua' (n = 22); 2) group-2: 'inflammation restricted to the MT of chorion and the decidua' (n = 31); 3) group-3: 'inflammation in the CT of chorion' (n = 53). We examined the frequency of inflammation in each placental compartment beyond chorio-decidua (i.e., amnion, umbilical cord, and chorionic-plate), and total grade (1-8) of acute-HCA. Moreover, the frequency of intra-amniotic infection (defined as positive amniotic-fluid culture for aerobic and anaerobic bacteria and genital mycoplasmas) and intra-amniotic inflammation (defined as amniotic fluid WBC ≥ 19 cells/mm3), and an intra-amniotic inflammatory response gauged by amnioticfluid WBC count (cells/mm3) were examined in 50 amniotic fluid samples within 7 days of birth. RESULTS: Amnionitis, funisitis and chorionic plate inflammation were more frequent (each for P < 0.01) and median total grade of acute-HCA was increased (P < 0.001) according to outside-in neutrophil migration within chorio-decidua (group-1vs.group-2vs.group-3). Moreover, intra-amniotic infection and inflammation were more frequent (each-for P < 0.05) and median amniotic-fluid WBC count was increased (P < 0.01) according-to outside-in neutrophil-migration within chorio-decidua (group-1 vs. group-2 vs. group-3). CONCLUSION: Acute-HCA is more advanced and severe, and intra-amniotic inflammation is more frequent and intense according to outside in neutrophil migration within the same chorio-decidua. This finding suggests that what is now acute chorio-deciduitis should be subdivided.

Chorioamnionitis induces hepatic inflammation and time-dependent changes of the enterohepatic circulation in the ovine fetus.

Chorioamnionitis, inflammation of fetal membranes, is an important cause of preterm birth and a risk factor for the development of adverse neonatal outcomes including sepsis and intestinal pathologies. Intestinal bile acids (BAs) accumulation and hepatic cytokine production are involved in adverse intestinal outcomes. These findings triggered us to study the liver and enterohepatic circulation (EHC) following intra-amniotic (IA) lipopolysaccharide (LPS) exposure. An ovine chorioamnionitis model was used in which circulatory cytokines and outcomes of the liver and EHC of preterm lambs were longitudinally assessed following IA administration of 10 mg LPS at 5, 12 or 24h or 2, 4, 8 or 15d before preterm birth. Hepatic inflammation was observed, characterized by increased hepatic cytokine mRNA levels (5h - 2d post IA LPS exposure) and increased erythropoietic clusters (at 8 and 15 days post IA LPS exposure). Besides, 12h after IA LPS exposure, plasma BA levels were increased, whereas gene expression levels of several hepatic BA transporters were decreased. Initial EHC alterations normalized over time. Concluding, IA LPS exposure induces significant time-dependent changes in the fetal liver and EHC. These chorioamnionitis induced changes have potential postnatal consequences and the duration of IA LPS exposure might be essential herein.

Biomarkers for a histological chorioamnionitis diagnosis in pregnant women with or without group B streptococcus infection: a case-control study.

BACKGROUND: Chorioamnionitis may cause serious perinatal and neonatal adverse outcomes, and group B streptococcus (GBS) is one of the most common bacteria isolated from human chorioamnionitis. The present study analyzed the impact of GBS infection and histological chorioamnionitis (HCA) on pregnancy outcomes and the diagnostic value of various biomarkers. METHODS: Pregnant women were grouped according to GBS infection and HCA detection. Perinatal and neonatal adverse outcomes were recorded with a follow-up period of 6 weeks. The white blood cell count (WBC), neutrophil ratio, and C-reactive protein (CRP) level from peripheral blood and soluble intercellular adhesion molecule-1 (sICAM-1), interleukin 8 (IL-8), and tumor necrosis factor α (TNF-α) levels from cord blood were assessed. RESULTS: A total of 371 pregnant women were included. Pregnant women with GBS infection or HCA had a higher risk of pathological jaundice and premature rupture of membranes and higher levels of sICAM-1, IL-8, and TNF-α in umbilical cord blood. Univariate and multivariate regression analysis revealed that sICMA-1, IL-8, TNF-α, WBC, and CRP were significantly related to an increased HCA risk. For all included pregnant women, TNF-α had the largest receiver operating characteristic (ROC) area (area: 0.841; 95% CI: 0.778-0.904) of the biomarkers analyzed. TNF-α still had the largest area under the ROC curve (area: 0.898; 95% CI: 0.814-0.982) for non-GBS-infected pregnant women, who also exhibited a higher neutrophil ratio (area: 0.815; 95% CI: 0.645-0.985) and WBC (area: 0.849; 95% CI: 0.72-0.978), but all biomarkers had lower value in the diagnosis of HCA in GBS-infected pregnant women. CONCLUSION: GBS infection and HCA correlated with several perinatal and neonatal adverse outcomes. TNF-α in cord blood and WBCs in peripheral blood had diagnostic value for HCA in non-GBS-infected pregnant women but not GBS-infected pregnant women.

Evaluation of leucine-rich alpha-2 glycoprotein as a biomarker of fetal infection.

This study aimed to determine the association between umbilical cord leucine-rich alpha-2 glycoprotein (LRG) and fetal infection and investigate the underlying mechanism of LRG elevation in fetuses. We retrospectively reviewed the medical records of patients who delivered at Osaka University Hospital between 2012 and 2017 and selected those with histologically confirmed chorioamnionitis (CAM), which is a common pregnancy complication that may cause neonatal infection. The participants were divided into two groups: CAM with fetal infection (CAM-f[+] group, n = 14) and CAM without fetal infection (CAM-f[-] group, n = 31). Fetal infection was defined by the histological evidence of funisitis. We also selected 50 cases without clinical signs of CAM to serve as the control. LRG concentrations in sera obtained from the umbilical cord were unaffected by gestational age at delivery, neonatal birth weight, nor the presence of noninfectious obstetric complications (all, p > 0.05). Meanwhile, the LRG levels (median, Interquartile range [IQR]) were significantly higher in the CAM-f(+) group (10.37 [5.21-13.7] µg/ml) than in the CAM-f(-) (3.61 [2.71-4.65] µg/ml) or control group (3.39 [2.81-3.93] µg/ml; p < 0.01). The area under the receiver operating characteristic (ROC) curve of LRG for recognizing fetal infection was 0.92 (optimal cutoff, 5.08 µg/ml; sensitivity, 86%; specificity, 88%). In a mouse CAM model established by lipopolysaccharide administration, the fetal LRG protein in sera and LRG mRNA in the liver were significantly higher than those in phosphate-buffered saline (PBS)-administered control mice (p < 0.01). In vitro experiments using a fetal liver-derived cell line (WRL68) showed that the expression of LRG mRNA was significantly increased after interleukin (IL)-6, IL-1ß, and tumor necrosis factor- alpha (TNF-α) stimulation (p < 0.01); the induction was considerably stronger following IL-6 and TNF-α stimulation (p < 0.01). In conclusion, LRG is an effective biomarker of fetal infection, and fetal hepatocytes stimulated with inflammatory cytokines may be the primary source of LRG production in utero.

Application of a Prediction Model Based on the Laboratory Index Score in Prelabor Rupture of Membranes with Histologic Chorioamnionitis During Late Pregnancy.

BACKGROUND This study aimed to establish a prediction model based on the maternal laboratory index score (Lab-score) for histologic chorioamnionitis (HCA) in patients with prelabor rupture of membranes (PROM) during late pregnancy. MATERIAL AND METHODS Sixty-nine cases of pregnant women with PROM were retrospectively analyzed. The general information and laboratory indicators were compared between the HCA (n=22) and non-HCA (n=47) groups. A multivariate logistic regression method was used to establish the prediction model. We plotted the receiver operating characteristic curve and calculated the area under the curve (AUC). The clinical effectiveness of each model was compared by decision curve analysis. RESULTS Only C-reactive protein (CRP) in the laboratory index predicted HCA, but its diagnostic efficacy was not ideal (AUC=0.651). Then, we added CRP to the platelet/white blood cell count ratio and triglyceride level to construct the Lab-score. Based on the Lab-score, important clinical parameters, including body mass index, diastolic blood pressure, and preterm birth, were introduced to construct a complex joint prediction model. The AUC of this model was significantly larger than that of CRP (0.828 vs. 0.651, P=0.035), but not significantly different from that of Lab-score (0.828 vs. 0.724, P=0.120). Considering the purpose of HCA screening, the net benefit of the complex model was better than that of Lab-score and CRP. CONCLUSIONS The complex model based on Lab-score is useful in the clinical screening of high-risk populations with PROM and HCA during late pregnancy.

Histopathology of the fetal inflammatory response to intra-amniotic pathogens.

Obstetric endorsement of the utility of placental histologic examination remains infrequent, especially from obstetricians who do not have a placental pathologist as part of their own local clinical care team. Placental pathologic examinations are viewed as useless if they do not provide answers to urgent clinical questions. Increasingly, however, it is appreciated that while placental analysis should be considered with regard to its longer term value; results can assess lifelong risks of a wide range of diseases that have been tied to prenatal exposures (e.g., [1]), including distinguishing sex-specific differences in those risks. (e.g., [2]) This review will focus solely on acute fetal (?) inflammation, more specifically, the fetal neutrophil responses in umbilical cord, chorionic plate vessels and to some degree, the fetal system as a whole. This histologic fetal inflammatory response is often the most readily accessible aspect of "FIR" piece of FIRS (the fetal inflammatory response syndrome). Some researchers have defined FIRS by a combination of both cytokine (especially IL-6) levels and the histopathologic FIR (Musilova et al., 2018) [3]. As we and others have noted, many histology based FIR cases, even those associated with neurodevelopmental outcomes such as cerebral palsy, are clinically silent.(e.g., [4]) Current clinical diagnostic criteria may have high specificity as they are very good at identifying non-FIR cases. However, that high specificity is coupled with very low specificity, identifying only 10% of FIR (Doty et al., 2018 Jul) [5]. Our aim is to provide a conceptual framework for the readers of the journal to better understand how to answer the following questions: What is a neutrophil and how is it important in FIR? What is the differential diagnosis for histologic FIR? How long has there been FIR? What secondary processes may have been recruited (and when) to contribute to the final pathology and pathophysiology of the given pregnancy?

Expression of S100A Alarmins in Cord Blood Monocytes Is Highly Associated With Chorioamnionitis and Fetal Inflammation in Preterm Infants.

Background: Preterm infants exposed to chorioamnionitis and with a fetal inflammatory response are at risk for neonatal morbidity and adverse outcome. Alarmins S100A8, S100A9, and S100A12 are expressed by myeloid cells and have been associated with inflammatory activation and monocyte modulation. Aim: To study S100A alarmin expression in cord blood monocytes from term healthy and preterm infants and relate results to clinical findings, inflammatory biomarkers and alarmin protein levels, as well as pathways identified by differentially regulated monocyte genes. Methods: Cord blood CD14+ monocytes were isolated from healthy term (n = 10) and preterm infants (<30 weeks gestational age, n = 33) by MACS technology. Monocyte RNA was sequenced and gene expression was analyzed by Principal Component Analysis and hierarchical clustering. Pathways were identified by Ingenuity Pathway Analysis. Inflammatory proteins were measured by Multiplex ELISA, and plasma S100A proteins by mass spectrometry. Histological chorioamnionitis (HCA) and fetal inflammatory response syndrome (FIRS) were diagnosed by placenta histological examination. Results: S100A8, S100A9, and S100A12 gene expression was significantly increased and with a wider range in preterm vs. term infants. High S100A8 and S100A9 gene expression (n = 17) within the preterm group was strongly associated with spontaneous onset of delivery, HCA, FIRS and elevated inflammatory proteins in cord blood, while low expression (n = 16) was associated with impaired fetal growth and physician-initiated delivery. S100A8 and S100A9 protein levels were significantly lower in preterm vs. term infants, but within the preterm group high S100A gene expression, spontaneous onset of labor, HCA and FIRS were associated with elevated protein levels. One thousand nine hundred genes were differentially expressed in preterm infants with high vs. low S100A alarmin expression. Analysis of 124 genes differentially expressed in S100A high as well as FIRS and HCA groups identified 18 common pathways and S100A alarmins represented major hubs in network analyses. Conclusion: High expression of S100A alarmins in cord blood monocytes identifies a distinct clinical risk group of preterm infants exposed to chorioamnionitis and with a fetal inflammatory response. Gene and pathway analyses suggest that high S100A alarmin expression also affects monocyte function. The connection with monocyte phenotype and inflammation-stimulated S100A expression in other cell types (e.g., neutrophils) warrants further investigation.

Early pregnancy immune profile and preterm birth classified according to uteroplacental lesions.

INTRODUCTION: Preterm birth is a heterogeneous phenotype, with placental abnormalities underlying many cases. The etiology of preterm births that occur in the absence of placental abnormalities, however, remain enigmatic and we considered that early pregnancy biomarkers may provide clues. METHODS: Women from a hospital-based cohort (2008-2012, n = 397) were randomly selected within 6 strata of term and preterm birth with and without placental decidual vasculopathy (arteriopathy), intrauterine inflammation/infection (acute chorioamnionitis), or no lesions. Lipids and inflammatory markers were analyzed in first trimester samples (12.5 ± 0.6 weeks) and related to outcome groups (referent, term births with no lesions). Factor analysis then clustered analytes and related these to preterm birth groups, adjusted for covariates and stratified by pre-pregnancy obesity. RESULTS: Three biomarker patterns were identified. Immune activation cytokines (33% of the variance) were associated with preterm birth with no lesions (aOR 1.5, 95%CI 1.1-2.1), particularly among obese women. In contrast, inflammatory chemokines (9% of variance) were associated with term and preterm vasculopathy among non-obese women (aOR 2.6 [1.3, 4.7] and 2.0 [1.1, 3.0], respectively). DISCUSSION: The early pregnancy maternal immune profile is related to preterm births classified according to placental lesions, and these associations vary according to obesity status.

Cyclic AMP in human preterm infant blood is associated with increased TLR-mediated production of acute-phase and anti-inflammatory cytokines in vitro.

BACKGROUND: Preterm infants are at high risk of infection and have distinct pathogen recognition responses. Suggested mechanisms include soluble mediators that enhance cellular levels of cAMP. The aim of this study was to assess the relationship between blood cAMP concentrations and TLR-mediated cytokine production in infants during the first month of life. METHODS: Cord and serial peripheral blood samples (days of life 1-28) were obtained from a cohort of very preterm (<30 weeks' gestational age) and term human infants. Whole-blood concentrations of cAMP and FSL-1 and LPS in vitro stimulated cytokine concentrations were measured by ELISA and multiplex bead assay. RESULTS: cAMP concentrations were higher in cord than in peripheral blood, higher in cord blood of female preterm infants, and lower at Days 1 and 7 in infants exposed to chorioamnionitis, even after adjusting for leukocyte counts. TLR2 and TLR4-mediated TNF-α, IL-1ß, IL-6, IL-12p70, and IL-10 production in vitro increased over the first month of life in preterm infants and were positively correlated with leukocyte-adjusted cAMP levels and reduced by exposure to chorioamnionitis. CONCLUSIONS: The ontogeny of blood cAMP concentrations and associations with chorioamnionitis and TLR-mediated production of cytokines suggest that this secondary messenger helps shape distinct neonatal pathogen responses in early life.

A rat model of placental inflammation explains the unexplained elevated maternal serum alpha-fetoprotein associated with adverse pregnancy outcomes.

AIM: It has been reported in numerous studies that elevated maternal serum alpha-fetoprotein (MS-AFP) is associated with adverse pregnancy outcomes (APO), such as pre-eclampsia, stillbirth, preterm birth and fetal growth restriction. However, the mechanism linking elevated MS-AFP and APO is obscure. In this study, we tried to explore the mechanism by using pregnant rats. METHODS: Lipopolysaccharide (LPS) was used to induce placental inflammation in pregnant rats. Maternal serum and placental inflammatory cytokines and placental morphology were used to assess the level of placental inflammation. The incidences of APO and the levels of MS-AFP were evaluated. The expressions of alpha-fetoprotein (AFP) in the related organs and fetal serum AFP levels were detected. RESULTS: Compared to saline-treated pregnant rats, LPS led to elevated maternal serum and placental inflammatory cytokines and a higher rate of placental inflammation. Lipopolysaccharide resulted in the features of APO and at the same time elevated MS-AFP. Maternal serum alpha-fetoprotein levels were significantly correlated to the evaluation parameters of APO. Lipopolysaccharide did not increase the expressions of AFP in fetal liver, maternal liver and placenta, but reduced the fetal serum AFP levels. CONCLUSION: The phenomenon that elevated MS-AFP is associated with APO, which has been reported in human pregnancies, is observed in our rat model. Placental inflammation can be the potential cause linking the two manifestations together. Although the source of elevated MS-AFP is not identified, fetal blood circulation is suspected. Our study may provide an animal model for the future studies on this subject.

Implementation of a clinical guideline to decrease laboratory tests in newborns evaluated for early onset sepsis.

BACKGROUND: Creation of a clinical guideline to reduce the number of complete blood counts (CBCs) obtained on healthy term infants for early onset sepsis (EOS) evaluation secondary to maternal chorioamnionitis. METHODS: A clinical guideline was introduced at four neonatal intensive care units (NICU) to reduce laboratory tests during EOS evaluation. Measures include frequency and timing of CBCs, culture negative sepsis, length of stay, and readmission rate. RESULTS: Mean number of CBCs per patient significantly decreased (2.31±0.62 versus 1.52±0.65) without increasing trends for patients with culture negative sepsis, length of stay, or re-admission. CONCLUSION: The clinical guideline demonstrated a significant reduction in the number of CBCs obtained in well-appearing infants admitted to the NICU secondary to maternal chorioamnionitis.

Predictive value of procalcitonin, C-reactive protein, and white blood cells for chorioamnionitis among women with preterm premature rupture of membranes.

OBJECTIVE: To determine the predictive value of procalcitonin, C-reactive protein (CRP), and white blood cells (WBC) for chorioamnionitis among women with preterm premature rupture of membranes (PPROM). METHODS: A prospective cross-sectional study of all women with singleton pregnancy and PPROM admitted to a referral hospital in Shiraz, Iran, from 2016 to 2018. All women were hospitalized until delivery. The incidence of chorioamnionitis was recorded. Maternal serum CRP, procalcitonin, and WBC were measured on the day of admission and the day before termination of pregnancy. The diagnostic accuracy of each test was evaluated by receiver operator characteristic (ROC) curve analysis. RESULTS: Overall, 75 women with PPROM were included in the study. After termination of pregnancy, 34 (45.3%) were diagnosed with clinical chorioamnionitis. Those with chorioamnionitis had significantly higher serum levels of CRP both on admission (P=0.004) and before termination of pregnancy (P<0.001). The area under the curve for last CRP was 0.78 (95% confidence interval, 0.57-0.84), indicating moderate accuracy. Procalcitonin and WBC had low accuracy to predict chorioamnionitis. CONCLUSION: Among CRP, procalcitonin, and WBC, maternal serum CRP was found to be the most accurate predictor of chorioamnionitis among women with PPROM.

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

INTRODUCTION: This study was performed to determine whether the combination of maternal blood and amniotic fluid biomarkers can improve the predictive accuracy of histologic chorioamnionitis (HC). METHODS: This 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. RESULTS: The 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). DISCUSSION: We 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.

Small RNA (sRNA) expression in the chorioallantois, endometrium and serum of mares following experimental induction of placentitis.

Intrauterine infection and inflammation remain a major cause of preterm labour in women and mares, with little known about small RNA (sRNA) expression in tissue or circulation. To better characterise placental inflammation (placentitis), we examined sRNA expression in the endometrium, chorioallantois and serum of mares with and without placentitis. Disease was induced in 10 mares via intracervical inoculation of Streptococcus equi ssp. zooepidemicus, either with moderate or high levels of inoculum; three uninoculated gestationally matched mares were used as controls. Matched chorioallantois and endometrium were sampled in two locations: Region 1, gross inflammation near cervical star with placental separation and Region 2, gross inflammation without placental separation. In Region 1, 26 sRNAs were altered in chorioallantois, while 20 were altered in endometrium. Within Region 2, changes were more subdued in both chorioallantois (10 sRNAs) and endometrium (two sRNAs). Within serum, we identified nine significantly altered sRNAs. In summary, we have characterised the expression of sRNA in the chorioallantois, the endometrium and the serum of mares with experimentally induced placentitis using next-generation sequencing, identifying significant changes within each tissue examined. These data should provide valuable information about the physiology of placental inflammation to clinicians and researchers alike.

Maternal peripheral blood platelet-to-white blood cell ratio and platelet count as potential diagnostic markers of histological chorioamnionitis-related spontaneous preterm birth.

BACKGROUND: Histological chorioamnionitis (HCA) is one of the leading causes of spontaneous preterm birth, thus, to identify novel biomarkers for the early diagnosis of HCA is in a great need. OBJECTIVE: To investigate the diagnostic value of maternal peripheral blood platelet-to-white blood cell ratio (PLT/WBC) and platelet (PLT) counts in HCA-related preterm birth. METHODS: A total of 400 patients with preterm birth were enrolled in this study: non-HCA group (n = 193) and HCA group (n = 207), and 87 full-term pregnancies were enrolled as the control. The peripheral blood of the participators was collected, and the neutrophil count, WBC count, platelet count, and levels of C-reactive protein (CRP) and procalcitonin were recorded, and the platelet-to-white blood cell ratio (PLT/WBC) of the participators was calculated. Receiver operating characteristic (ROC) curve has been drawn to show the sensitivity and specificity of PLT/WBC and PLT count for the diagnosis of HCA-related spontaneous preterm birth patients. RESULTS: The neutrophil count, WBC count, and procalcitonin show no significant differences among the three groups, and the PLT count, PLT/WBC, and CRP (P < 0.05) were significantly increased in HCA group compared with non-HCA group; moreover, the area under the curve (AUC) of PLT/WBC, PLT, and CRP was 0.744 (95% confidence interval [CI], 0.6966-0.7922), 0.8095 (95% CI, 0.7676-0.8514), and 0.5730 (95% CI, 0.5173-0.6287), respectively. CONCLUSION: Platelet count and PLT/WBC may become a potential biomarker of HCA-related spontaneous preterm birth.

Antibody Microarray Analysis of Plasma Proteins for the Prediction of Histologic Chorioamnionitis in Women With Preterm Premature Rupture of Membranes.

We aimed to identify maternal blood biomarkers predictive of histologic chorioamnionitis (HCA) in the plasma of women with preterm premature rupture of membranes (PPROM) and to determine whether the combination of these biomarkers with conventional clinical variables can improve the prediction of HCA. This retrospective cohort study included 82 consecutive women with PPROM (23-34 gestational weeks) who delivered within 96 hours of blood sampling. A membrane-based human antibody microarray was used to analyze the plasma proteome. The validation of 5 candidate biomarkers of interest was performed by enzyme-linked immunosorbent assay (ELISA) in the final cohort (n = 82). Serum C-reactive protein (CRP) levels were measured at sampling. Seventy-nine molecules studied exhibited intergroup differences. Validation by ELISA confirmed higher levels of plasma matrix metalloproteinase-9 (MMP-9), interleukin-6 (IL-6), S100 A8/A9, and insulin-like growth factor-binding protein 1 (IGFBP-1), but not tissue inhibitor of metalloproteinase 1 (TIMP-1), in women with HCA than in women without HCA. Using a stepwise regression analysis, a combined prediction model was developed, which included the plasma MMP-9, serum CRP levels, and gestational age (area under the curve [AUC], 0.932). The AUC for this model was significantly greater than that for any single variable included in the predictive model. Protein-antibody microarray technology can be useful in identifying plasma-based predictors for HCA. This study suggests that plasma MMP-9, IL-6, IGFBP-1, and S100 A8/A9 are important noninvasive predictors for HCA in women with PPROM and that the best predictive model, which combined these biomarkers with conventional clinical factors, can significantly improve the predictability for HCA.

Increased TLR4 and TREM-1 expression on monocytes and neutrophils in preterm birth: further evidence of a proinflammatory state.

Objective: Increased inflammation is considered as a risk factor and a promoter of preterm birth (PTB). Monocytes and neutrophils are the main sources of cytokines in the early inflammatory phase. So far, very few studies have indicated CD14/TLR4 and TREM-1 on the monocytes and neutrophils as important targets in PTB. Materials and methods: TLR4 and TREM-1 on CD14+ maternal and cord blood monocytes and neutrophils were detected using flow cytometry in 48 normal term women, 48 PTB with chorioamnionitis (CCA) women, and 40 PTB without CCA women. In the fetal membranes, mRNA and protein levels of the CD14/TLR4-TREM-1 signaling pathway, CD14, TLR4, NF-κBp65, and TREM-1 were analyzed by qRT-PCR and western blot. ELISA was further used to detect TLR4 and TREM-1 levels in maternal and cord serums. Results: Compared with the normal term and PTB without CCA women, we found that (1) TLR4 and TREM-1 levels on CD14+ maternal and cord blood monocytes and neutrophils in the PTB with CCA group were elevated (p < .001); (2) the protein and mRNA expressions of CA14, TLR4, NF-κBp65, and TREM-1 of the PTB with CCA group were upregulated (p < .001); (3) Maternal and cord serum concentrations of TLR4 and TREM-1 in the PTB with CCA group were greater (p < .001). Conclusions: The high levels of TLR4 and TREM-1 surface expression were observed on CD14+ maternal and cord blood monocyte and neutrophils, confirming their proinflammatory profiles in PTB with CCA. TLR4 and TREM-1 on monocyte and neutrophils might have a role in infection-related PTB.

Maternal Serum Interleukin-6: A Non-Invasive Predictor of Histological Chorioamnionitis in Women with Preterm-Prelabor Rupture of Membranes.

OBJECTIVE: To assess the added value of maternal serum levels of IL-6 in women with preterm-prelabor rupture of membranes (PPROM) as a non-invasive test for the prediction of histological chorioamnionitis (HCA). METHODS: This was a prospective cohort study of pregnant women between 20 + 0 and 36 + 6 weeks of gestation with a confirmed diagnosis of PPROM. Logistic regression models were created for the prediction of HCA and compared by assessing the improvement in their Naegelkerke R2 as a measure of goodness of fit. Predictive performance of all models was assessed by receiver operating characteristics curve (ROC) analysis and compared by the DeLong method. RESULTS: From 47 women with PPROM, 31 (66%) developed HCA. Maternal serum IL-6 ≥19.5 pg/dL was the best cut-off point for the prediction of HCA (OR = 15; 95% CI: 3.6-61; p < 0.01). A model comprising maternal characteristics and IL-6 ≥19.5 pg/dL showed an area under the curve of 0.85 (95% CI: 0.74-0.95), significantly improving the previous models of IL-6 ≥19.5 pg/dL (R2: 23.3 vs. 34.1%; p = 0.01) or maternal characteristics (R2: 8.4 vs. 34.1%; p < 0.01). CONCLUSIONS: A model comprising maternal serum levels of IL-6 plus maternal characteristics proves to be a good non-invasive predictor of HCA.

Maternal blood endotoxin activity in pregnancies complicated by preterm premature rupture of membranes.

Objective: To compare maternal blood endotoxin activity (EA) in women with preterm premature rupture of membranes (PPROM) with gestational age (GA) matched controls; to evaluate serial EA till birth in PPROM and its correlation with latency to delivery. Methods: We followed singleton preterm pregnancies from admission with PPROM until birth. Uncomplicated, GA-matched pregnancies served as controls. Demographics, birth and neonatal outcomes were collected. EA (EAA™) was assessed serially in PPROM and at study entry in controls. EA was compared using Mann Whitney and Wilcoxon tests, p value <.05 was considered significant. Results: We recruited 20 cases of PPROM and 20 controls. Demographics were similar between groups. Mean GA of PPROM was 29.0 ± 2.2 weeks and median latency was 7.5 (IQR 14.1) weeks. Median EA at admission following PPROM was significantly elevated over controls (0.43 (0.18) versus 0.36 (0.2); p < .02). Overall there was no difference in median EA at admission and in labor (0.43 (0.18) versus 0.33 (0.21); p = .2) following PPROM. However, on comparing cases with latency to delivery ≤7 days (n = 10) versus >7 days (n = 10), there was a significant drop in EA in the latter group (0.44 (0.2) versus 0.34 (0.2); p < .004). Conclusions: EA in PPROM represents a promising biomarker in predicting the clinical evolution of preterm birth.