Extracellular Vesicles-mediated recombinant IL-10 protects against ascending infection-associated preterm birth by reducing fetal inflammatory response.

Background: Fetal inflammatory response mediated by the influx of immune cells and activation of pro-inflammatory transcription factor NF-κB in feto-maternal uterine tissues is the major determinant of infection-associated preterm birth (PTB, live births < 37 weeks of gestation). Objective: To reduce the incidence of PTB by minimizing inflammation, extracellular vesicles (EVs) were electroporetically engineered to contain anti-inflammatory cytokine interleukin (IL)-10 (eIL-10), and their efficacy was tested in an ascending model of infection (vaginal administration of E. coli) induced PTB in mouse models. Study design: EVs (size: 30-170 nm) derived from HEK293T cells were electroporated with recombinant IL-10 at 500 volts and 125 Ω, and 6 pulses to generate eIL-10. eIL-10 structural characters (electron microscopy, nanoparticle tracking analysis, ExoView [size and cargo content] and functional properties (co-treatment of macrophage cells with LPS and eIL-10) were assessed. To test efficacy, CD1 mice were vaginally inoculated with E. coli (1010CFU) and subsequently treated with either PBS, eIL-10 (500ng) or Gentamicin (10mg/kg) or a combination of eIL-10+gentamicin. Fetal inflammatory response in maternal and fetal tissues after the infection or treatment were conducted by suspension Cytometer Time of Flight (CyTOF) using a transgenic mouse model that express red fluorescent TdTomato (mT+) in fetal cells. Results: Engineered EVs were structurally and functionally stable and showed reduced proinflammatory cytokine production from LPS challenged macrophage cells in vitro. Maternal administration of eIL-10 (10 µg/kg body weight) crossed feto-maternal barriers to delay E. coli-induced PTB to deliver live pups at term. Delay in PTB was associated with reduced feto-maternal uterine inflammation (immune cell infiltration and histologic chorioamnionitis, NF-κB activation, and proinflammatory cytokine production). Conclusions: eIL-10 administration was safe, stable, specific, delayed PTB by over 72 hrs and delivered live pups. The delivery of drugs using EVs overcomes the limitations of in-utero fetal interventions. Protecting IL-10 in EVs eliminates the need for the amniotic administration of recombinant IL-10 for its efficacy.

Testing of drugs using human feto-maternal interface organ-on-chips provide insights into pharmacokinetics and efficacy.

Objectives: To improve preclinical drug testing during pregnancy, we developed multiple microfluidic organ-on-chip (OOC) devices that represent the structure, functions, and responses of the two feto-maternal interfaces (FMis) in humans (fetal membrane [FMi-OOC] and placenta [PLA-OOC]). This study utilized feto-maternal interface OOCs to test the kinetics and efficacy of drugs during pregnancy. Study design: The FMi-OOC contained amnion epithelial, mesenchymal, chorion trophoblast, and decidual cells. The PLA-OOC contained cytotrophoblasts (BeWo), syncytiotrophoblasts (BeWo + forskolin), and human umbilical vein endothelial cell lines. Therapeutic concentrations of either pravastatin or rosuvastatin (200 ng mL-1), a model drug for these experiments, were applied to either decidua (in FMi-OOC) and syncytiotrophoblasts (in PLA-OOC) chambers under normal and oxidative stress conditions (induced by cigarette smoke extract [CSE 1 : 25]) to evaluate maternal drug exposure during normal pregnancy or oxidative stress (OS) associated pathologies, respectively. We determined statin pharmacokinetics and metabolism (LC-MS/MS), drug-induced cytotoxicity (LDH assay), and efficacy to reduce OS-induced inflammation (multiplex cytokine assay). Results: Both OOCs mimicked two distinct human feto-maternal interfaces. The drugs tested permeated the maternal-fetal cell layers of the FMi-OOC and PLA-OOC within 4 hours and generated cell and time-specific statin metabolites from various cell types without causing any cytotoxicity. OS-induced pro-inflammatory cytokines were effectively reduced by statins by increasing anti-inflammatory cytokine response across the FMi-OOC and PLA-OOC. Conclusion: Two distinct feto-maternal interface OOCs were developed, tested, and validated for their utility to conduct preclinical trials during pregnancy. We demonstrated that the placenta and fetal membranes-decidual interface both are able to transport and metabolize drugs and that the safety and efficacy of a drug can be determined using the anatomical structures recreated on OOCs.

Quantitative Proteomics by SWATH-MS of Maternal Plasma Exosomes Determine Pathways Associated With Term and Preterm Birth.

Exosomes are membrane-bound nanovesicles that transport molecular signals between cells. This study determined changes in maternal plasma exosome proteomics contents in term and preterm births. Maternal plasma (MP) samples were collected from group 1: term not in labor (TNIL, n = 13); group 2: term in labor (TL, n = 11); group 3: preterm premature rupture of membranes (pPROM, n = 8); and group 4: preterm birth (PTB, n = 13). Exosomes isolated from plasma by differential density centrifugation followed by size exclusion chromatography were characterized by morphology (electron microscopy), quantity and size (nanoparticle tracking analysis), and markers (western blot). A quantitative, information-independent acquisition [sequential windowed acquisition of all theoretical mass spectra (SWATH-MS)] approach was used to determine the protein profile in exosomes. Ingenuity Pathway Analysis determined pathways associated with the protein profile identified in exosomes. MP exosomes were spherical, had a mean diameter of 120 nm, and were positive for exosomal proteins CD63 and TSG101 irrespective of pregnancy status. No distinct changes in exosome quantities were seen in maternal circulation across the groups. SWATH-MS identified 72 statistically significant proteins across the groups studied. Bioinformatics analysis showed the proteins within the exosomes in TNIL, TL, pPROM, and PTB target pathways mainly associated with inflammatory and metabolic signals. Exosomal data suggest that homeostatic imbalances, specifically inflammatory and endocrine signaling, might disrupt pregnancy maintenance resulting in labor-related changes both at term and preterm. Reflection of physiologic changes in exosomes is suggestive of its usefulness as biomarkers and cellular function indicators.

Amniotic Fluid Exosome Proteomic Profile Exhibits Unique Pathways of Term and Preterm Labor.

Our objective was to determine the amniotic fluid-derived exosomal proteomic profile in patients who had spontaneous preterm birth (PTB) or preterm premature rupture of membranes (pPROM) compared with those who delivered at term. A cross-sectional study of a retrospective cohort was used to quantify and determine the protein content of exosomes present in amniotic fluid, in PTB or pPROM, and normal term labor (TL) or term not in labor (TNIL) pregnancies. Exosomes were isolated by differential centrifugation and quantified using nanocrystals (Qdot) coupled to CD63 and placental alkaline phosphatase (PLAP) by fluorescence nanoparticle tracking analysis. The exosomal proteomic profile was identified by liquid chromatography-tandem mass spectrometry, and a small ion library was constructed to quantify the proteomic data by Sequential Window Acquisition of All Theoretical analysis. Ingenuity Pathway Analysis determined canonical pathways and biofunctions associated with dysregulated proteins. Amniotic fluid exosomes have similar shape and quantity regardless of the conditions; however, the PLAP/CD63 ratios for TL, PTB, and pPROM were significantly higher (∼3.8-, ∼4.4-, and ∼3.5-fold, respectively) compared with TNIL. The PLAP/CD63 ratio was also significantly higher (∼1.3-fold) in PTB compared with pPROM. Biological functions primarily indicated nonspecific inflammatory response regardless of condition, but unique profiles were also identified in cases (PTB and pPROM) compared with term. Amniotic fluid exosomes provide information specific to normal and abnormal parturition. Inflammatory marker enrichment and its uniqueness in term and preterm pregnancies support the value of exosomes in determining underlying physiology associated with term and preterm parturition.

Chorioamniotic membrane senescence: a signal for parturition?

OBJECTIVE: Senescence is an important biological phenomenon involved in both physiologic and pathologic processes. We propose that chorioamniotic membrane senescence is a mechanism associated with human parturition. The present study was conducted to explore the association between senescence and normal term parturition by examining the morphologic and biochemical evidences in chorioamniotic membranes. STUDY DESIGN: Chorioamniotic membranes were collected from normal term deliveries; group 1: term labor and group 2: term, not in labor. Senescence-related morphologic changes were determined by transmission electron microscopy and biochemical changes were studied by senescence-associated (SA) ß-galactosidase staining. Amniotic fluid samples collected from both term labor and term not in labor were analyzed for 14 SA secretory phenotype (SASP) markers. RESULTS: Morphologic evidence of cellular senescence (enlarged cells and organelles) and a higher number of SA ß-galactosidase-stained amnion and chorion cells were observed in chorioamniotic membranes obtained from women in labor at term, when compared to term not in labor. The concentration of proinflammatory SASP markers (granulocyte macrophage colony-stimulating factor, interleukin-6 and -8) was significantly higher in the amniotic fluid of women in labor at term than women not in labor. In contrast, SASP factors that protect against cell death (eotaxin-1, soluble Fas ligand, osteoprotegerin, and intercellular adhesion molecule-1) were significantly lower in the amniotic fluid samples from term labor. CONCLUSION: Morphologic and biochemical features of senescence were more frequent in chorioamniotic membranes from women who experienced term labor. Senescence of chorioamniotic membranes were also associated with amniotic fluid SASP markers.

Histological evidence of oxidative stress and premature senescence in preterm premature rupture of the human fetal membranes recapitulated in vitro.

Preterm prelabor rupture of the membranes (pPROM) may lead to preterm births (PTBs). We investigated premature senescence of fetal membranes in women with pPROM and spontaneous PTB with intact membranes (<34 weeks) and the inducibility fetal membrane senescence phenotype by oxidative stress in vitro. IHC was performed for p53, p21, and phospho (p)-p38 mitogen-activated protein kinase (MAPK) as markers of senescence phenotype in pPROM, PTBs, and term births. Term fetal membranes were exposed to cigarette smoke extract to induce oxidative stress. Western blots documented p-p53 and p-p38 MAPK. Transmission electron microscopy assessed cellular morphologic features in clinical and cigarette smoke extract-treated membranes. A total of 80% of pPROM cells and >60% of term cells were positive for all three senescence phenotype markers, and concentrations were higher than in PTBs (P < 0.05). p53 staining was comparable in membranes from PTB and term birth pregnancies, whereas only <30% and <45% of cells were positive for p21 and p38 MAPK, respectively. In vitro cigarette smoke extract exposure increased p-p38 MAPK without any detectable change in p-p53 MAPK. Enlargement of organelles consistent with senescence phenotype was evident in pPROM and term membranes in vivo and after cigarette smoke extract treatment in vitro but was less apparent in PTBs. Histologic and biochemical resemblance of pPROM and term membranes suggests premature senescence of the membranes is a mechanistic feature in pPROM, and this can be phenocopied in an in vitro model.

Amniotic fluid metabolomic analysis in spontaneous preterm birth.

OBJECTIVE: To identify metabolic changes associated with early spontaneous preterm birth (PTB; <34 weeks) and term births, using high-throughput metabolomics of amniotic fluid (AF) in African American population. METHOD: In this study, AF samples retrieved from spontaneous PTB (<34 weeks [n = 25]) and normal term birth (n = 25) by transvaginal amniocentesis at the time of labor prior to delivery were subjected to metabolomics analysis. Equal volumes of samples were subjected to a standard solvent extraction method and analyzed using gas chromatography/mass spectrometry (MS) and liquid chromatography/MS/MS. Biochemicals were identified through matching of ion features to a library of biochemical standards. After log transformation and imputation of minimum observed values for each compound, t test, correlation tests, and false discovery rate corrections were used to identify differentially regulated metabolites. Data were controlled for clinical/demographic variables and medication during pregnancy. RESULTS: Of 348 metabolites measured in AF samples, 121 metabolites had a gestational age effect and 116 differed significantly between PTB and term births. A majority of significantly altered metabolites could be classified into 3 categories, namely, (1) liver function, (2) fatty acid and coenzyme A (CoA) metabolism, and (3) histidine metabolism. The signature of altered liver function was apparent in many cytochrome P450-related pathways including bile acids, steroids, xanthines, heme, and phase II detoxification of xenobiotics with the largest fold change seen with pantothenol, a CoA synthesis inhibitor that was 8-fold more abundant in PTB. CONCLUSION: Global metabolic profiling of AF revealed alteration in hepatic metabolites involving xenobiotic detoxification and CoA metabolism in PTB. Maternal and/or fetal hepatic function differences may be developmentally related and its contribution PTB as a cause or effect of PTB is still unclear.

1,25-Dihydroxyvitamin D deficiency is associated with preterm birth in African American and Caucasian women.

Vitamin (vit) D deficiency and preterm birth (PTB) are more prevalent among African American (AA) women compared to caucasian (Cau) women. Because vit D is important in regulating cell-mediated immune responses, vit D insufficiency or deficiency during pregnancy may enhance inflammation in pregnant women and increase the risk of PTB. In this study, circulatory levels of 25-hydroxy (OH) and 1,25-dihydroxy (OH)2 vit D were measured using chemiluminescence and radioimmunoassay techniques, respectively, in AA (n = 108) and Cau (n = 84) women who delivered at term and preterm. The results from this study suggest that the serum levels of the 25-(OH) vit D concentrations tend to decrease (P = .06) in the Cau women who delivered at preterm compared to those delivering at term. However, the 25-(OH) vit D levels in Cau and AA between term and preterm deliveries were not significantly different. The serum levels of 1,25-(OH)2 vit D were found to be significantly lower in AA women compared to Cau women (P < .02) at term, and in the Cau (P < .01) and AA (P < .04) women delivering at preterm compared to those delivering at term. One-way analysis of variance demonstrated that 1,25-(OH)2 vit D levels were significantly lower in participants delivering at preterm (<34 weeks and between 34 and 37 weeks) compared to those delivering at term (>37 weeks).These results suggest that low levels of serum 1,25-(OH)2 vit D are associated with PTB, and vit D can potentially be used as a novel diagnostic marker in the detection of PTB.

Fetal DNA Methylation Associates with Early Spontaneous Preterm Birth and Gestational Age.

Spontaneous preterm birth (PTB, <37 weeks gestation) is a major public health concern, and children born preterm have a higher risk of morbidity and mortality throughout their lives. Recent studies suggest that fetal DNA methylation of several genes varies across a range of gestational ages (GA), but it is not yet clear if fetal epigenetic changes associate with PTB. The objective of this study is to interrogate methylation patterns across the genome in fetal leukocyte DNA from African Americans with early PTB (24(1/7)-34(0/7) weeks; N = 22) or term births (39(0/7)-40(6/7)weeks; N = 28) and to evaluate the association of each CpG site with PTB and GA. DNA methylation was assessed across the genome with the HumanMethylation450 BeadChip. For each individual sample and CpG site, the proportion of DNA methylation was estimated. The associations between methylation and PTB or GA were evaluated by fitting a separate linear model for each CpG site, adjusting for relevant covariates. Overall, 29 CpG sites associated with PTB (FDR<.05; 5.7×10(-10)

Group B Streptococcus colonization and higher maternal IL-1ß concentrations are associated with early term births.

Association between maternal Group B Streptococcus (GBS) colonization diagnosed between 35 and 37 weeks of gestation and early term birth (between 37 and 39 weeks) and maternal-fetal inflammatory response associated with this condition were tested. In this cohort study of women delivering at term at Centennial Women's Hospital in Nashville, TN, GBS status and other clinical and demographic data were obtained from medical records. Exposed women were those testing positive for GBS (GBS positive [n = 490]) and the unexposed tested negative for GBS (GBS negative [n = 1,127]). To determine the inflammatory response associated with GBS, a cross sectional study, maternal and fetal plasma biomarkers (IL-1ß, IL-2, IL-6, IL-8, and TNF-α) were measured in the same cohort. T-tests and logistic regression determined association between GBS status, biomarker concentrations and early term birth. Gestational age was reduced to 271.1 (95% CI 270.4, 271.1) for cases compared to 274.7 (95% CI 274.4, 275.1) days for controls (p < 0.0001). The odds of early term birth was increased by threefold in cases (OR 3.28; 95% CI 2.60-4.15; p < 0.0001). The mean birth weight in cases (3285.3 g) (95% CI 3242.6, 3327.9) was lower than the controls, 3373.8 g (95% CI 3348.9, 3398.7) (p = 0.0004). Maternal IL-1ß was greater in cases (22.8 ng/ml; range 5.2-157.7 ng/ml) compared to controls (5.7; range 2.4-69.5 ng/ml; p < 0.0001). IL-1ß was higher in fetal plasma in cases vs. controls (20.33 vs. 8.18 ng/ml; p = 0.01). A 10 ng/ml increase in maternal IL-1ß was associated with increased risk for GBS infection (OR: 1.628, CI: 1.163-2.278; p = 0.0045). GBS colonization shortened gestational age at term and IL-1ß concentration in maternal plasma is an indicator of GBS status.

Ethnic disparity in amniotic fluid levels of hyaluronan, histone H2B and superoxide dismutase in spontaneous preterm birth.

AIMS: To document racial disparity in an immune modulator, hyaluronan (HA), an antimicrobial, histone H2B (H2B), and an antioxidant, superoxide dismutase (SOD) in amniotic fluid (AF) from African-American (AA) and Caucasian (C) subjects with spontaneous preterm birth (PTB). METHODS: In a case (PTB) control (term) study, AF samples were analyzed for HA, H2B and SOD by ELISA. Differences in analyte concentrations between races were documented and a secondary analysis based on histologic chorioamnionitis (HC) was also performed. RESULTS: No differences in the median HA, H2B and SOD were seen between cases and controls. AA cases had lower HA but higher H2B and SOD than controls. Analyte concentrations were not different between C cases and controls. AA samples at term had lower H2B and SOD compared to C samples at term. Cases with HC showed higher H2B and SOD. CONCLUSION: We report ethnic disparity in AF antimicrobial, immune mediator and antioxidant factors. Dysregulated AF production of HA, H2B and SOD was associated with PTB in AA, not in C, suggesting an overwhelming inflammatory response in AA PTB, whereas inflammation is likely a secondary phenomenon in C PTB.

Amniotic fluid and maternal race influence responsiveness of fetal membranes to bacteria.

Spontaneous preterm birth (PTB) and preterm prelabor rupture of membranes (pPROM) occur more frequently in African-American women than in other racial groups. This may be due to an enhanced inflammatory response to pathogens associated with the condition. It is also possible that amniotic fluid (AF) has different immunomodulatory properties in African-American women that increase their risk of PTB and pPROM. To test this, we cultured fetal membranes from European-American and African-American women with sterile medium (control), Escherichia coli, Gardnerella vaginalis, Group B streptococci (GBS), Polyporphorans gingivalis, Mycoplasma hominis, Ureaplasma urealyticum or Ureaplasma parvum in the presence and absence of 50% autologous AF. Cytokine concentrations were quantified in the conditioned medium. All bacterial species increased IL-8 production. IL-1ß and TNF-α production were stimulated by LPS, E. coli, and G. vaginalis compared with control, but responses to Group B streptococci and P. gingivalis were limited to IL-1ß and TNF-α respectively. Genital mycoplasmas stimulated TNF-α and IL-10 but had no effect on IL-1ß production. African-Americans had twice the IL-1ß response to E. coli as European-Americans (P=0.031). Conversely, European-Americans produced more IL-8 in response to LPS than African-Americans (P=0.026). AF had both pro- and anti-inflammatory properties that varied between races and pathogens. These results suggest that the host response to fetal membrane infections is complex and not generalizable. Interventions to prevent PTB and pPROM may need to be customized based on a patient's race, type of bacterial infection and factors in her AF.

Short fetal leukocyte telomere length and preterm prelabor rupture of the membranes.

BACKGROUND: Rupture of the fetal membranes is a common harbinger of imminent labor and delivery. Telomere shortening is a surrogate for oxidative stress (OS) and senescence. Fetal leukocyte and placental membrane DNA telomere lengths were evaluated to determine their association with preterm prelabor rupture of the membranes (pPROM) or spontaneous preterm births with intact membranes (PTB), compared to term birth. METHODS: Telomere lengths were quantified in cord blood leukocytes (n = 133) from three major groups: 1) pPROM (n = 28), 2) PTB (n = 69) and 3) uncomplicated full term births (controls, n = 35), using real-time quantitative PCR. Placental membrane specimens (n = 18) were used to correlate fetal leukocyte and placental telomere lengths. Telomere length differences among the groups were analyzed by ANOVA. Pearson correlation coefficients determined relationships between leukocyte and placental membrane telomere lengths. RESULTS: In pregnancies with intact membranes, fetal leukocyte telomere length was inversely proportional to gestational age. The mean telomere length decreased as gestation progressed, with the shortest at term. pPROM had telomere lengths (9962 ± 3124 bp) that were significantly shorter than gestational age-matched PTB (11546 ± 4348 bp, p = 0.04), but comparable to term births (9011 ± 2497 bp, p = 0.31). Secondary analyses revealed no effects of race (African American vs. Caucasian) or intraamniotic infection on telomere length. A strong Pearson's correlation was noted between fetal leukocyte and placental membrane telomere lengths (ρ = 0.77; p<0.01). CONCLUSIONS: Fetal leukocyte telomere length is reduced in pPROM compared to PTB but is similar to term births. pPROM represents a placental membrane disease likely mediated by OS-induced senescence.

A single-nucleotide polymorphism in the fetal catechol-O-methyltransferase gene is associated with spontaneous preterm birth in African Americans.

Catechol-O-methyltransferase (COMT) activity has been reported to be higher in African Americans (AA) than Caucasians (Cau). COMT converts 2- and 4-hydroxy (OH) estrogens to 2- and 4-methoxyestrogens, respectively, and can increase estrogenic milieu locally in tissues. To assess whether the increased incidence of preterm birth (PTB) among AA women is associated with single-nucleotide polymorphism (SNP) in the COMT gene, we examined variations in maternal and fetal COMT genes and their association with pregnancy outcomes (term vs preterm pregnancies) using 4 functional SNPs: rs4633, rs4680, rs4818, and rs6269 in both AA and Cau. We analyzed samples from 267 AA women (191 term and 76 preterm pregnancies) and 339 Cau (194 term and 145 preterm pregnancies) in this study. The results showed a significant difference (P < .05) in allele and genotype frequencies between term and preterm AA and Cau women in 3 SNPs in both maternal and fetal DNA. The analysis revealed that in AA fetal COMT genes, SNP rs4818 is associated with PTB at the allele (C; P < .001), genotype (C/C; P < .01), and 2- (P < .03) and 3 (P < .04)-window haplotype levels. Multidimensionality reduction analysis also showed a significant (P < .01) association between rs4818 and PTB. In conclusion, our study demonstrated that a synonymous polymorphism, rs4818 in the fetal COMT gene, is associated with PTB in AA.

Ethnic disparity in spontaneous preterm birth and maternal pre-pregnancy body mass index.

PURPOSE: To investigate differences in pre-pregnancy BMI status in patients with spontaneous preterm birth (PTB) compared with term birth and assess the role of ethnicity as a risk modifier in BMI-associated PTB. METHODS: A case-control study involving self-reported African American and Caucasian women delivering singletons in Nashville, TN, USA, 2003-2009. Maternal pre-pregnancy BMI was recorded in 447 PTB-cases (African American = 145, Caucasian = 302) and 1315 term-birth controls (African American = 522; Caucasian = 793). Crude and adjusted odds ratio (OR and AOR) for PTB were calculated using normal BMI (18.5-24.9 kg/m(2)) as reference. Age, education, marital status, income, smoking, parity, previous PTB and pregnancy weight gain were included as covariates in logistic regression. RESULTS: No significant differences were noted in the OR for PTB among different BMI categories when women of different ethnicity were combined. Odds of PTB were greater in obese than in normal weight Caucasian women, even after adjusting for confounders (AOR = 1.84, 95%CI [1.15, 2.95]). Obese African American women had a decreased crude OR for PTB, although this was not significant after adjusting for confounders (AOR = 0.72, 95%CI [0.38, 1.40]). The odds for early PTB (<32 weeks) were decreased in obese compared with normal weight African American women (OR = 0.23, 95%CI [0.08, 0.70]), whereas they were increased in obese compared with normal weight Caucasian women (OR = 2.30, 95%CI [1.32, 4.00]). CONCLUSION: The risk for PTB in women with different pre-pregnancy BMI categories differs according to ethnicity.

Amniotic fluid eicosanoids in preterm and term births: effects of risk factors for spontaneous preterm labor.

OBJECTIVE: To evaluate amniotic fluid arachidonic acid metabolites using enzymatic and nonenzymatic (lipid peroxidation) pathways in spontaneous preterm birth and term births, and to estimate whether prostanoid concentrations correlate with risk factors (race, cigarette smoking, and microbial invasion of amniotic cavity) associated with preterm birth. METHODS: In a case-control study, amniotic fluid was collected at the time of labor or during cesarean delivery. Amniotic fluid samples were subjected to gas chromatography, negative ion chemical ionization, and mass spectrometry for prostaglandin (PG) E2, PGF2α, and PGD2 and for 6-keto-PGF1α (thromboxane 2 and F2-isoprostane). Primary analysis examined differences between prostanoid concentrations in preterm birth (n=133) compared with term births (n=189). Secondary stratified analyses (by race, cigarette smoking, and microbial invasion of amniotic cavity) compared eicosanoid concentrations in three epidemiological risk factors. RESULTS: Amniotic fluid F2-isoprostane, PGE2, and PGD2 were significantly higher at term than in preterm birth, whereas PGF2α was higher in preterm birth 6-keto-PGF1α and thromboxane 2 concentrations were not different. Data stratified by race (African American or white) showed no significant disparity among prostanoid concentrations. Regardless of gestational age status, F2-isoprostane was threefold higher in smokers, and other eicosanoids were also higher in smokers compared with nonsmokers. Preterm birth with microbial invasion of amniotic cavity had significantly higher F2-isoprostane compared with preterm birth without microbial invasion of amniotic cavity. CONCLUSION: Most amniotic fluid eicosanoid concentrations (F2-isoprostane, PGE2, and PGD2), are higher at term than in preterm births. The only amniotic fluid eicosanoid that is not higher at term is PGF2α.

An overview of racial disparities in preterm birth rates: caused by infection or inflammatory response?

Infection has been hypothesized to be one of the factors associated with spontaneous preterm birth (PTB) and with the racial disparity in rates of PTB between African American and Caucasian women. However, recent findings refute the generalizability of the role of infection and inflammation. African Americans have an increased incidence of PTB in the setting of intraamniotic infection, periodontal disease, and bacterial vaginosis compared to Caucasians. Herein we report variability in infection- and inflammation-related factors based on race/ethnicity. For African American women, an imbalance in the host proinflammatory response seems to contribute to infection-associated PTB, as evidenced by a greater presence of inflammatory mediators with limited or reduced presence of immune balancing factors. This may be attributed to differences in the genetic variants associated with PTB between African Americans and Caucasians. We argue that infection may not be a cause of racial disparity but in association with other risk factors such as stress, nutritional deficiency, and differences in genetic variations in PTB, pathways and their complex interactions may produce differential inflammatory responses that may contribute to racial disparity.

The expression pattern of two novel cytokines (IL-24 and IL-29) in human fetal membranes.

OBJECTIVE: interleukin (IL)-24 and -29 are novel cytokines, produced by immune cells in response to microbial antigens. The functions of these cytokines in the reproductive system are unknown. We examined the expression pattern of IL-24 and IL-29 in human fetal membranes from preterm and term births and in in vitro in response to microbial antigens. METHODS: fetal membranes collected from cesarean sections at term (normal, not in labor) were placed in culture for 48 h. These membranes were then stimulated with bacterial lipopolysaccharide (LPS) or viral antigen poly-inosinic and cytidylic acid (polyIC) for an additional 24 h. Amniotic fluids (AF) and fetal membranes were also collected from preterm and term deliveries. IL-24 and IL-29 expressions were studied by RT-PCR. ELISA documented culture media and AF cytokine concentrations. RESULTS: IL-24 and IL-29 expressions were seen in cultured fetal membranes regardless of stimulation. Expressions were also found in preterm and term labor membranes, but not in non-labor tissues at term. IL-24 concentrations were higher after LPS stimulation whereas IL-29 concentrations were higher after polyIC-stimulation. AF analysis did not detect either of the cytokines either preterm or term. CONCLUSION: this is the first study to report IL-24 and IL-29 expressions in human fetal membranes. Higher concentrations of these cytokines in response to distinct infectious stimuli suggest different pathways for fetal immune response during infection.

Perinatal sepsis caused by Williamsia serinedens infection in a 31-year-old pregnant woman.

Williamsia serinedens has been isolated from soil but has not yet been implicated in human disease. We report the first case of perinatal sepsis caused by a dual-morphotype form of Williamsia serinedens in a 31-year-old pregnant woman hospitalized with preterm labor.

Association of genetic variants, ethnicity and preterm birth with amniotic fluid cytokine concentrations.

We examined the association of 166 single nucleotide polymorphisms (SNPs) in cytokines and cytokine related genes with cytokine concentrations (IL-1beta, IL-8, and IL-10) in the amniotic fluid (AF). These cytokines have been associated with spontaneous preterm birth (PTB) and their genetic regulation may play a role in disease risk. These associations were studied in both PTB and term births in African Americans and Caucasians; maternal and fetal genotypes were studied separately. Analyses modeled genotype, pregnancy status, and marker by pregnancy status (case/control) interaction with cytokine concentration as outcome. Our results indicate that AF cytokines (IL-1beta and IL-10) were associated with interactions between pregnancy status and both maternal and fetal SNPs, with the most significant interactions being observed for African Americans with IL-1beta concentration (maternal at IL1RAP rs1024941 p < 10(-3), fetal IL1RAP rs3773953 p < 10(-3)). AF IL-10 concentrations also showed evidence for association with SNPs in both ethnicities with the most significant interaction in Caucasian maternal samples (IL10 rs1800896 p < 10(-3)). Our data indicate that the genetic regulation of cytokine concentrations in PTB likely differs by ethnicity. AF cytokine concentrations were associated with interactions between genotype and PTB in African Americans, but less so in Caucasians.