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.

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.

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.

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.

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.

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.

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.

Interleukin-6 (IL-6) and receptor (IL6-R) gene haplotypes associate with amniotic fluid protein concentrations in preterm birth.

Spontaneous preterm birth (PTB-gestational age <37 weeks) occurs in approximately 450 000 births annually in the United States and is one of the leading causes of neonatal morbidity and mortality. Risk of PTB is affected by complex gene-environment interactions that are not well understood. We examined the PTB candidate gene, Interleukin 6 (IL-6) and its receptor (IL6-R) in both Caucasian (145 PTB and 194 term maternal; 140 PTB and 179 term fetal) and African-American (76 PTB and 191 term maternal; 66 PTB and 183 term fetal) DNA. Eight single nucleotide polymorphisms (SNPs) in IL-6 and 22 SNPs in IL6R were examined for association with IL-6 amniotic fluid (AF) concentrations, as concentration of IL-6 is a hypothesized risk factor. In addition, IL-6 and IL6-R SNPs were analyzed for associations with PTB. Haplotype associations were tested by sliding windows. No strong single marker effects were observed in Caucasians; however, in African-American maternal IL-6R marker rs4553185 associated with PTB (allele P = 4.49 x 10(-3) and genotype P = 0.01). The strongest haplotype associations were observed in IL-6R with IL-6 cytokine concentration as outcome: Caucasian fetal (rs4601580-rs4845618) P = 1.6 x 10(-3) and African-American maternal (rs4601580-rs4845618-rs6687726-rs7549338) P = 2.30 x 10(-3). Significant results converged on three regions in the two genes: in IL-6 markers rs1800797, rs1800796 and rs1800795; in IL-6R markers rs4075015, rs4601580, rs4645618, rs6687726 and rs7549338 and markers rs4845623, rs4537545 and rs4845625. In conclusion, our results suggest that IL-6 AF concentration, in situations of PTB, result from variation in IL-6 and more importantly IL-6R.

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.

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.

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.

Racial disparity in pathophysiologic pathways of preterm birth based on genetic variants.

OBJECTIVE: To study pathophysiologic pathways in spontaneous preterm birth and possibly the racial disparity associating with maternal and fetal genetic variations, using bioinformatics tools. METHODS: A large scale candidate gene association study was performed on 1442 SNPs in 130 genes in a case (preterm birth < 36 weeks) control study (term birth > 37 weeks). Both maternal and fetal DNA from Caucasians (172 cases and 198 controls) and 279 African-Americans (82 cases and 197 controls) were used. A single locus association (genotypic) analysis followed by hierarchical clustering was performed, where clustering was based on p values for significant associations within each race. Using Ingenuity Pathway Analysis (IPA) software, known pathophysiologic pathways in both races were determined. RESULTS: From all SNPs entered into the analysis, the IPA mapped genes to specific disease functions. Gene variants in Caucasians were implicated in disease functions shared with other known disorders; specifically, dermatopathy, inflammation, and hematological disorders. This may reflect abnormal cervical ripening and decidual hemorrhage. In African-Americans inflammatory pathways were the most prevalent. In Caucasians, maternal gene variants showed the most prominent role in disease functions, whereas in African Americans it was fetal variants. The IPA software was used to generate molecular interaction maps that differed between races and also between maternal and fetal genetic variants. CONCLUSION: Differences at the genetic level revealed distinct disease functions and operational pathways in African Americans and Caucasians in spontaneous preterm birth. Differences in maternal and fetal contributions in pregnancy outcome are also different between African Americans and Caucasians. These results present a set of explicit testable hypotheses regarding genetic associations with preterm birth in African Americans and Caucasians.

Distinct pathophysiologic pathways induced by in vitro infection and cigarette smoke in normal human fetal membranes.

OBJECTIVE: The purpose of this study was to document distinct pathways that are initiated by lipopolysaccharide and cigarette smoke stimulation of normal term fetal membranes. STUDY DESIGN: Fetal membranes from nonsmoking women at term, not in labor, from cesarean deliveries were placed in an organ explant system and stimulated with cigarette smoke extracts (CSEs), lipopolysaccharide, or lipopolysaccharide + CSE. Media were assayed for an interleukin (IL)-1beta, -1 receptor antagonist, -6, -8, -10, tumor necrosis factor alpha, soluble tumor necrosis factor receptors 1 and 2, and matrix metalloproteinases 1, 2, 3, 8, 9, and 12. Tissue homogenates were assayed for apoptotic markers (p53, caspase 3 activity, and cleaved poly [ADP-ribose] polymerase-1). RESULTS: Lipopolysaccharide stimulation resulted in higher cytokine and matrix metalloproteinase concentrations, whereas it was lower after CSE and CSE + lipopolysaccharide stimulations, compared with control specimens. Apoptotic factors were several folds higher after CSE or CSE + lipopolysaccharide stimulation, compared with control specimens or lipopolysaccharide stimulations. CONCLUSION: Cigarette smoke showed immunoinhibitory properties that potentially were mediated by apoptosis and lipopolysaccharide-induced proinflammatory response. This study demonstrated 2 independent pathophysiologic pathways that may alter pregnancy outcome.

Diversity in cytokine response to bacteria associated with preterm birth by fetal membranes.

OBJECTIVE: This study compared cytokine and prostaglandin (PG) responses by fetal membranes stimulated with 4 different bacterial species associated with preterm birth (PTB). STUDY DESIGN: Fetal membranes (n = 13 from normal term cesarean sections [not in labor]) in an organ explant system were stimulated with heat-killed Ureaplasma parvum, Gardanerella vaginalis, Escherichia coli, group B Streptococcus (GBS), or lipopolysaccharide (LPS). Cytokines (interleukin [IL]-1beta, IL-6, IL-8, IL-10, tumor necrosis factor [TNF]-alpha, and interferon-gamma) and PG (PGF(2alpha) and PGE(2)) concentrations were quantitated and compared. RESULTS: LPS and E coli increased all cytokine and PG productions compared with controls. Cytokine profiles were similar after G vaginalis and GBS stimulation. G vaginalis increased PGE(2), whereas GBS increased PGF(2alpha). U parvum demonstrated the mildest response with only IL-10 and TNF-alpha concentrations being higher with no detectible effect on PGs. CONCLUSION: Fetal membrane cytokine signatures of 4 different bacteria associated with PTB are distinct, suggesting that infection as a potential cause of PTB is not homogeneous in its presentation.

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.

Genetic regulation of amniotic fluid TNF-alpha and soluble TNF receptor concentrations affected by race and preterm birth.

Racial disparity in spontaneous preterm birth (PTB) between African Americans and Caucasians in the US is unexplained, but is probably related to differences in amniotic fluid (AF) inflammatory cytokine profiles. Therefore, this study analyzed the association of 34 single nucleotide polymorphisms (SNPs) in TNF-alpha and its receptor genes (TNFR1 and TNFR2) with AF TNF-alpha and soluble TNF receptor (R1 and R2) concentrations in PTB. Samples consisted of African American and Caucasian cases (PTB), and controls (term birth) for which both cytokine, and maternal and fetal genotype data were available. Analyses were performed with genotype, case, and maker-status interaction in the model for log transformed cytokine concentrations. In Caucasians, two interactions between genotype and pregnancy outcome associated with cytokine concentrations, whereas 14 gene variants in African Americans showed interactions with pregnancy outcome, and 13 showed association with genetic markers. In conclusion, cytokine concentrations in African American preterm births can be partially explained by interactions between pregnancy outcome, SNPs and infection. This does not appear to be the case in Caucasians. These findings may be important in understanding disparity in rates of PTB between the two populations.

Patterns of cytokine profiles differ with pregnancy outcome and ethnicity.

BACKGROUND: Preterm birth (PTB) is hypothesized to be an inflammatory response disease. However, no single factor alone is likely to explain PTB risk. It is more probable that coordinated networks of cytokines affect risk. METHODS: Therefore, we examined the relationships between amniotic fluid (AF) cytokines/chemokines and related biomarkers in PTB and normal term deliveries in African Americans and Caucasians. Data were obtained from African American (41 preterm labor and 91 term labor) and Caucasian (105 preterm labor and 100 term labor) pregnant mothers. Pro-inflammatory cytokines and related molecules interleukin (IL)-1, IL-6, IL-8, and tumor necrosis factor- (TNF)-alpha, TNF soluble receptors (sTNFR1 and sTNFR2), and anti-inflammatory cytokine IL-10 that were all previously associated with PTB were studied. Correlations between biomarkers were calculated; differences of correlation coefficients between AF from African American and Caucasian samples in preterm labor and term labor were measured. RESULTS: Multiple differences were observed between African American and Caucasian preterm and term birth groups. In term birth the strongest differences were between pro- and anti-inflammatory correlations, whereas in PTB differences were equally distributed between pro-inflammatory/anti-inflammatory and pro-inflammatory/pro-inflammatory correlations. Three correlation patterns differed significantly between AF from PTB African Americans with and without microbial invasion of the intra-amniotic cavity (MIAC); no differences were observed in Caucasians with MIAC. CONCLUSION: Correlation analyses of cytokine measurements suggest coordinated interplay during pregnancy; significant differences exist between African Americans and Caucasians. Such analyses can serve as a means of understanding risk factors in these populations.

Amniotic fluid interleukin-6 increase is an indicator of spontaneous preterm birth in white but not black Americans.

OBJECTIVE: This study examined the differences in the inflammatory cytokine interleukin (IL)-6 and the immunoinhibitory cytokine IL-10 in the amniotic fluid of black and white women in spontaneous preterm birth. METHODS: In this study, 321 amniotic fluids from cases (preterm birth 36 or fewer weeks' gestation) and controls (normal term delivery longer than 37 weeks' gestation) were collected (147 cases [49 blacks and 98 whites] and 174 controls [85 blacks and 89 whites]) at the time of active labor. IL-6 and IL-10 concentrations were measured by immunoassays. Using normal-term delivery as controls, logistic regression models were used to estimate odds ratios (OR) and 95% confidence intervals (CIs) for preterm birth. RESULTS: A significant difference in IL-6 concentration was observed in white cases (cases: 3773 pg/mL; controls: 1682 pg/mL; P = .0003), compared with controls, but not in blacks (cases: 2042 pg/mL; controls: 2366 pg/mL; P = .6). In a combined multivariable analysis, when the highest and the lowest quartiles of IL-6 were compared in whites, the ORs (95% CI) for preterm birth across quartiles were 1.74 (0.62-4.88), 1.09 (0.39-3.02), and 5.68 (2.15-15.0). No such association was found in blacks. IL-10 concentration was not different between cases and controls in either race. CONCLUSIONS: Race-specific associations exist between IL-6 but not IL-10 concentration and preterm birth. Elevated IL-6 concentrations are associated with preterm birth in whites but not blacks.

Racial disparity in maternal-fetal genetic epistasis in spontaneous preterm birth.

OBJECTIVE: To understand the differences in genetic interactions among tumor necrosis factor-alpha, interleukin-6 and their receptor gene variants between black and white patients in spontaneous preterm birth. STUDY DESIGN: Maternal and fetal DNA (n = 1195) were collected from cases (preterm birth < 36 weeks' gestation; n = 448), controls (> 37 weeks' gestation; n = 747), and genotyped for single nucleotide polymorphisms in tumor necrosis factor-alpha, tumor necrosis factor receptor 1, and tumor necrosis factor receptor 2, interleukin-6, and interleukin-6 receptor loci. Multifactor dimensionality reduction analysis was used to test all single and multilocus combinations for the ability to predict pregnancy outcome. RESULTS: In white patients, multilocus interactions in maternal DNA between single nucleotide polymorphisms at -7227 (interleukin-6), 22,215 (interleuki-6 receptor) and -3448 (tumor necrosis factor-alpha) was predictive of approximately 59.1% (P < .02; odds ratio, 2.3 [95% confidence interval = 1.6-3.4]) of pregnancy outcome. In white fetal DNA and black maternal DNA, no significant interactive models were observed. In black patients, the best epistatic model was in fetal DNA between single nucleotide polymorphisms at 17,691 (tumor necrosis factor-receptor 1) and at -3448 (tumor necrosis factor-alpha) and was predictive of pregnancy outcome 68.3% of the time (P < .01; odds ratio, 5.0 [95% confidence interval = 2.6-9.6]). CONCLUSION: Analyses of multilocus interactions found/associated different models in black and white patients in both maternal and fetal DNA with preterm birth as outcome. Significant maternal-fetal interactions were not detected in either race.

Racial disparity in amniotic fluid concentrations of tumor necrosis factor (TNF)-alpha and soluble TNF receptors in spontaneous preterm birth.

OBJECTIVE: Preterm birth rate in the United States is higher in blacks than whites. It has been hypothesized that a differential inflammatory response may explain this disparity. The objective of this study is to examine the inflammatory cytokine, tumor necrosis factor (TNF)-alpha and soluble TNF receptor concentrations (sTNFR1 and sTNFR2) in the amniotic fluid of black and white women at delivery. STUDY DESIGN: Amniotic fluid samples were collected during active labor from 158 cases (preterm births, gestational age 22(0/7) weeks to 36(0/7) weeks, 52 black and 106 white) and 175 controls (term births, gestational age 37(0/7) weeks to 42(0/7) weeks, 87 black and 88 white) at Centennial Women's Hospital, Nashville, TN. Amniotic fluid TNF-alpha, sTNFR1, and sTNFR2 concentrations and the molar ratios of TNF-alpha to its receptors were compared between cases and controls within each racial group. RESULTS: Median TNF-alpha concentration was associated with preterm birth when whites and blacks were analyzed together, with cases having higher values (191.5 pg/mL) than controls (68.9 pg/mL; P < .001). There were no significant associations with sTNFR1 or sTNFR2 concentrations between cases (2409.4 and 2934.3 pg/mL, respectively) and controls (2759.9 and 3084.1 pg/mL, respectively) when the racial groups were analyzed together (P = .08, P = .4, respectively). Black cases associated with higher TNF-alpha concentrations (1287.0 pg/mL in cases and 67.3 pg/mL in controls; P < .001). In whites there was no association between TNF-alpha and preterm birth (P = .3). The molar ratio of TNF-alpha/total sTNFR (R1 plus R2) associated with higher TNF-alpha in black cases, compared with black controls (P < .001). There was no significant association between white cases and controls for ligand receptor ratios (P = .3). CONCLUSION: The TNF-alpha/sTNFR profile in pregnancy differs between racial groups, suggesting a difference in bioavailability of TNF-alpha. The larger molar ratio of TNF-alpha/sTNFR in black cases may be indicative of a TNF-alpha mediated pathological process of preterm birth in blacks but not in whites.