Showing papers by "Roberto Romero published in 2017"

Embryo implantation evolved from an ancestral inflammatory attachment reaction

Abstract: The molecular changes that support implantation in eutherian mammals are necessary to establish pregnancy. In marsupials, pregnancy is relatively short, and although a placenta does form, it is present for only a few days before parturition. However, morphological changes in the uterus of marsupials at term mimic those that occur during implantation in humans and mice. We investigated the molecular similarity between term pregnancy in the marsupials and implantation in eutherian mammals using the gray short-tailed opossum (Monodelphis domestica) as a model. Transcriptomic analysis shows that term pregnancy in the opossum is characterized by an inflammatory response consistent with implantation in humans and mice. This immune response is temporally correlated with the loss of the eggshell, and we used immunohistochemistry to report that this reaction occurs at the materno-fetal interface. We demonstrate that key markers of implantation, including Heparin binding EGF-like growth factor and Mucin 1, exhibit expression and localization profiles consistent with the pattern observed during implantation in eutherian mammals. Finally, we show that there are transcriptome-wide similarities between the opossum attachment reaction and implantation in rabbits and humans. Our data suggest that the implantation reaction that occurs in eutherians is derived from an attachment reaction in the ancestral therian mammal which, in the opossum, leads directly to parturition. Finally, we argue that the ability to shift from an inflammatory attachment reaction to a noninflammatory period of pregnancy was a key innovation in eutherian mammals that allowed an extended period of intimate placentation.

Weak functional connectivity in the human fetal brain prior to preterm birth.

Abstract: It has been suggested that neurological problems more frequent in those born preterm are expressed prior to birth, but owing to technical limitations, this has been difficult to test in humans. We applied novel fetal resting-state functional MRI to measure brain function in 32 human fetuses in utero and found that systems-level neural functional connectivity was diminished in fetuses that would subsequently be born preterm. Neural connectivity was reduced in a left-hemisphere pre-language region, and the degree to which connectivity of this left language region extended to right-hemisphere homologs was positively associated with the time elapsed between fMRI assessment and delivery. These results provide the first evidence that altered functional connectivity in the preterm brain is identifiable before birth. They suggest that neurodevelopmental disorders associated with preterm birth may result from neurological insults that begin in utero.

A Role for the Inflammasome in Spontaneous Preterm Labor With Acute Histologic Chorioamnionitis.

Abstract: Inflammasomes are cytosolic multiprotein complexes that orchestrate inflammation in response to pathogens and endogenous danger signals. Herein, we determined whether the chorioamniotic membranes from women in spontaneous preterm labor with acute histologic chorioamnionitis (1) express major inflammasome components; (2) express caspase (CASP)-1 and CASP-4 as well as their active forms; (3) exhibit apoptosis-associated speck-like protein containing a CARD (ASC)/CASP-1 complex formation; and (4) release the mature forms of interleukin (IL)-1β and IL-18. We utilized quantitative reverse transcription polymerase chain reaction, enzyme-linked immunosorbent assay, immunoblotting, and immunohistochemistry to determine the messenger RNA (mRNA) and protein expression of major inflammasome components, nucleotide-binding oligomerization domain (NOD) proteins, and the pro- and mature/active forms of CASP-1, CASP-4, IL-1β, and IL-18. The ASC/CASP-1 complex formation was determined using an in situ proximity ligation assay. When comparing the chorioamniotic membranes from women in spontaneous preterm labor with acute histologic chorioamnionitis to those without this placental lesion, we found that (1) the mRNA of NLR family pyrin domain-containing protein ( NLRP) 1, NLRP3, NLR family CARD domain-containing protein 4 ( NLRC4), and NOD2 were higher; (2) the NLRP3 protein was increased; (3) the mRNA and active form (p10) of CASP-1 were greater; (4) the mRNA and active form of CASP-4 were increased; (5) the mRNA and mature form of IL-1β were higher; (6) the mature form of IL-18 was elevated; and (7) ASC/CASP-1 complex formation was increased. In conclusion, spontaneous preterm labor with acute histologic chorioamnionitis is characterized by an upregulation of NLRP3 and the active form of CASP-4, as well as increased ASC/CASP-1 complex formation, which may participate in the activation of CASP-1 and the maturation of IL-1β and IL-18 in the chorioamniotic membranes. These findings provide the first evidence that supports a role for the inflammasome in the pathological inflammation implicated in spontaneous preterm labor with acute histologic chorioamnionitis.

The prediction of late-onset preeclampsia: Results from a longitudinal proteomics study.

Abstract: Background Late-onset preeclampsia is the most prevalent phenotype of this syndrome; nevertheless, only a few biomarkers for its early diagnosis have been reported. We sought to correct this deficiency using a high through-put proteomic platform. Methods A case-control longitudinal study was conducted, including 90 patients with normal pregnancies and 76 patients with late-onset preeclampsia (diagnosed at ≥34 weeks of gestation). Maternal plasma samples were collected throughout gestation (normal pregnancy: 2-6 samples per patient, median of 2; late-onset preeclampsia: 2-6, median of 5). The abundance of 1,125 proteins was measured using an aptamers-based proteomics technique. Protein abundance in normal pregnancies was modeled using linear mixed-effects models to estimate mean abundance as a function of gestational age. Data was then expressed as multiples of-the-mean (MoM) values in normal pregnancies. Multi-marker prediction models were built using data from one of five gestational age intervals (8-16, 16.1-22, 22.1-28, 28.1-32, 32.1-36 weeks of gestation). The predictive performance of the best combination of proteins was compared to placental growth factor (PIGF) using bootstrap. Results 1) At 8-16 weeks of gestation, the best prediction model included only one protein, matrix metalloproteinase 7 (MMP-7), that had a sensitivity of 69% at a false positive rate (FPR) of 20% (AUC = 0.76); 2) at 16.1-22 weeks of gestation, MMP-7 was the single best predictor of late-onset preeclampsia with a sensitivity of 70% at a FPR of 20% (AUC = 0.82); 3) after 22 weeks of gestation, PlGF was the best predictor of late-onset preeclampsia, identifying 1/3 to 1/2 of the patients destined to develop this syndrome (FPR = 20%); 4) 36 proteins were associated with late-onset preeclampsia in at least one interval of gestation (after adjustment for covariates); 5) several biological processes, such as positive regulation of vascular endothelial growth factor receptor signaling pathway, were perturbed; and 6) from 22.1 weeks of gestation onward, the set of proteins most predictive of severe preeclampsia was different from the set most predictive of the mild form of this syndrome. Conclusions Elevated MMP-7 early in gestation (8-22 weeks) and low PlGF later in gestation (after 22 weeks) are the strongest predictors for the subsequent development of late-onset preeclampsia, suggesting that the optimal identification of patients at risk may involve a two-step diagnostic process.

Clinical chorioamnionitis at term VII: the amniotic fluid cellular immune response.

Abstract: OBJECTIVES 1) To characterize the cellular composition of the amniotic fluid of patients diagnosed with clinical chorioamnionitis at term, as a function of the presence or absence of microorganisms determined by cultivation techniques, and 2) to characterize the cytokine production by white blood cells present in the amniotic fluid using flow cytometry-based techniques. MATERIALS AND METHODS Amniotic fluid samples from 20 women who had the diagnosis of clinical chorioamnionitis at term were analyzed using cultivation techniques (for aerobic and anaerobic bacteria as well as genital Mycoplasmas). Amniotic fluid IL-6 concentrations were determined by an enzyme-linked immunosorbent assay. Amniotic fluid leukocytes were visualized by using hematoxylin and eosin staining and immunofluorescence. Immunophenotyping of surface markers and cytokines was performed in amniotic fluid leukocytes using flow cytometry. RESULTS 1) Neutrophils (CD45+CD15+ cells) were the most common leukocyte subset found in the amniotic fluid, followed by monocytes (CD45+CD14+ cells); other white blood cells (such as lymphocytes and natural killer cells) were scarce in the amniotic fluid; 2) the absolute counts of neutrophils and monocytes were significantly higher in patients with microorganisms found in the amniotic fluid than in those without detectable microorganisms, using cultivation techniques; 3) there was a significant correlation between the absolute counts of neutrophils and monocytes determined by flow cytometry (Spearman's correlation=0.97; P<0.001); 4) there was a significant correlation between the absolute white blood cell count determined with a hemocytometer chamber and by flow cytometric analysis (Spearman's correlation=0.88; P<0.001); and 5) the profile of cytokine expression differed between monocytes and neutrophils; while neutrophils predominantly produced TNF-α and MIP-1β, monocytes expressed higher levels of IL-1β and IL-1α. CONCLUSION Flow cytometry analysis of the amniotic fluid of patients with intra-amniotic infection and clinical chorioamnionitis at term demonstrated that neutrophils and monocytes are the most common cells participating in the inflammatory process. We have characterized, for the first time, the differential cytokine expression by these cells in this important complication of pregnancy.

CXCL10 and IL‐6: Markers of two different forms of intra‐amniotic inflammation in preterm labor

Abstract: Problem To determine whether amniotic fluid (AF) CXCL10 concentration is associated with histologic chronic chorioamnionitis in patients with preterm labor (PTL) and preterm prelabor rupture of the membranes (PROM) Method of Study This study included 168 women who had an episode of PTL or preterm PROM AF interleukin (IL)-6 and CXCL10 concentrations were determined by immunoassay Results (i) Increased AF CXCL10 concentration was associated with chronic (OR: 48; 95% CI: 17-14), but not acute chorioamnionitis; (ii) increased AF IL-6 concentration was associated with acute (OR: 42; 95% CI: 13-137) but not chronic chorioamnionitis; and (iii) an increase in AF CXCL10 concentration was associated with placental lesions consistent with maternal anti-fetal rejection (OR: 37; 95% CI: 13-104) (iv) All patients with elevated AF CXCL10 and IL-6 delivered preterm Conclusion Increased AF CXCL10 concentration is associated with chronic chorioamnionitis or maternal anti-fetal rejection, whereas increased AF IL-6 concentration is associated with acute histologic chorioamnionitis

Amniotic fluid neutrophils can phagocytize bacteria: A mechanism for microbial killing in the amniotic cavity.

Abstract: Problem Neutrophils are capable of performing phagocytosis, a primary mechanism for microbial killing. Intra-amniotic infection is characterized by an influx of neutrophils into the amniotic cavity. Herein, we investigated whether amniotic fluid neutrophils could phagocytize bacteria found in the amniotic cavity of women with intra-amniotic infection. Methods Amniotic fluid neutrophils from women with intra-amniotic infection were visualized by transmission electron microscopy (n=6). The phagocytic activity of amniotic fluid neutrophils from women with intra-amniotic infection and/or inflammation (n=10) or peripheral neutrophils from healthy individuals (controls, n=3) was tested using ex vivo phagocytosis assays coupled with live imaging. Phagocytosis by amniotic fluid neutrophils was also visualized by confocal microscopy (n=10) as well as scanning and transmission electron microscopy (n=5). Results (i) Intra-amniotic infection-related bacteria including cocci (eg Streptococcus agalactiae), bacilli (eg Bacteriodes fragilis and Prevotella spp.), and small bacteria without a cell wall (eg Ureaplasma urealyticum) were found inside of amniotic fluid neutrophils; (ii) peripheral neutrophils (controls) rapidly phagocytized S. agalactiae, U. urealyticum, Gardnerella vaginalis, and Escherichia coli; (iii) amniotic fluid neutrophils rapidly phagocytized S. agalactiae and G. vaginalis; and (iv) amniotic fluid neutrophils slowly phagocytized U. urealyticum and E. coli; yet, the process of phagocytosis of the genital mycoplasma was lengthier. Conclusion Amniotic fluid neutrophils can phagocytize bacteria found in the amniotic cavity of women with intra-amniotic infection, namely S. agalactiae, U. urealyticum, G. vaginalis, and E. coli. Yet, differences in the rapidity of phagocytosis were observed among the studied microorganisms. These findings provide a host defense mechanism whereby amniotic fluid neutrophils can kill microbes invading the amniotic cavity.

Neutrophil Extracellular Traps in the Amniotic Cavity of Women with Intra-Amniotic Infection: A New Mechanism of Host Defense:

Abstract: Neutrophil extracellular traps (NETs) control microbial infections through their antimicrobial activities attributed to DNA, histones, granules, and cytoplasmic proteins (eg, elastase). Intra-amniotic infection is characterized by the influx of neu-trophils into the amniotic cavity; therefore, the aim of this study was to determine whether amniotic fluid neutrophils form NETs in this inflammatory process. Amniotic fluid samples from women with intra-amniotic infection (n = 15) were stained for bacteria detection using fluorescent dyes. Amniotic fluid neutrophils were purified by filtration. As controls, neutrophils from maternal blood samples (n = 3) were isolated by density gradients. Isolated neutrophils were plated onto glass cover slips for culture with and without 100 nM of phorbol-12-myristate-13-acetate (PMA). NET formation was assessed by 4’,6-diamidino-2-phenylindole (DAPI) staining and scanning electron microscopy. Different stages of NET formation were visualized using antibodies against elastase and histone H3, in combination with DAPI staining, by confocal microscopy. Finally, maternal or neonatal neutrophils were added to amniotic fluid samples from women without intra-amniotic infection (n = 4), and NET formation was evaluated by DAPI staining. (1) NETs were present in the amniotic fluid of women with intra-amniotic infection; (2) all of the amniotic fluid samples had detectable live and dead bacteria associated with the presence of NETs; (3) in contrast to neu-trophils from the maternal circulation, amniotic fluid neutrophils did not require PMA stimulation to form NETs; (4) different stages of NET formation were observed by co-localizing elastase, histone H3, and DNA in amniotic fluid neutrophils; and (5) neither maternal nor neonatal neutrophils form NETs in the amniotic fluid of women without intra-amniotic infection. NETs are detectable in the amniotic fluid of women with intra-amniotic infection.

Clinical chorioamnionitis at term VIII: a rapid MMP-8 test for the identification of intra-amniotic inflammation.

Abstract: Objective Clinical chorioamnionitis is the most common infection/inflammatory process diagnosed in labor and delivery units worldwide. The condition is a syndrome that can be caused by (1) intra-amniotic infection, (2) intra-amniotic inflammation without demonstrable microorganisms (i.e. sterile intra-amniotic inflammation), and (3) maternal systemic inflammation that is not associated with intra-amniotic inflammation. The presence of intra-amniotic inflammation is a risk factor for adverse maternal and neonatal outcomes in a broad range of obstetrical syndromes that includes clinical chorioamnionitis at term. Although the diagnosis of intra-amniotic infection has relied on culture results, such information is not immediately available for patient management. Therefore, the diagnosis of intra-amniotic inflammation could be helpful as a proxy for intra-amniotic infection, while results of microbiologic studies are pending. A rapid test is now available for the diagnosis of intra-amniotic inflammation, based on the determination of neutrophil collagenase or matrix metalloproteinase-8 (MMP-8). The objectives of this study were (1) to evaluate the diagnostic indices of a rapid MMP-8 test for the identification of intra-amniotic inflammation/infection in patients with the diagnosis of clinical chorioamnionitis at term, and (2) to compare the diagnostic performance of a rapid MMP-8 test to that of a conventional enzyme-linked immunosorbent assay (ELISA) interleukin (IL)-6 test for patients with clinical chorioamnionitis at term. Materials and methods A retrospective cohort study was conducted. A transabdominal amniocentesis was performed in patients with clinical chorioamnionitis at term (n=44). Amniotic fluid was analyzed using cultivation techniques (for aerobic and anaerobic bacteria as well as genital Mycoplasmas) and broad-range polymerase chain reaction (PCR) coupled with electrospray ionization mass spectrometry (PCR/ESI-MS). Amniotic fluid IL-6 concentrations were determined by ELISA, and rapid MMP-8 results were determined by Yoon's MMP-8 Check®. Intra-amniotic inflammation was defined as an elevated amniotic fluid IL-6 concentration ≥2.6 ng/mL, and intra-amniotic infection was diagnosed by the presence of microorganisms in the amniotic fluid accompanied by intra-amniotic inflammation. The diagnostic indices of Yoon's MMP-8 Check® for the identification of intra-amniotic inflammation were calculated. In order to objectively compare Yoon's MMP-8 Check® with the ELISA IL-6 test for the identification of intra-amniotic inflammation, we used an amniotic fluid white blood cell (WBC) count ≥50 cells/mm3 to define intra-amniotic inflammation. Results (1) A positive rapid MMP-8 test had a sensitivity of 82.4% (28/34), specificity of 90% (9/10), positive predictive value of 96.6% (28/29), negative predictive value of 60% (9/15), positive likelihood ratio 8.2 (95% CI 1.3-53.2), and negative likelihood ratio 0.2 (95% CI 0.1-0.4) for the identification of intra-amniotic inflammation (prevalence 77.3%); (2) a positive rapid MMP-8 test had a sensitivity of 91.7% (22/24), specificity of 65% (13/20), positive predictive value of 75.9% (22/29), negative predictive value of 86.7% (13/15), positive likelihood ratio of 2.6 (95% CI 1.4-4.8), and negative likelihood ratio of 0.1 (95% CI 0.03-0.5) for the identification of intra-amniotic infection; (3) the rapid MMP-8 test had a significantly higher specificity than the ELISA IL-6 test in the identification of intra-amniotic inflammation as determined by an amniotic fluid WBC count ≥50 cells/mm3. The sensitivity and accuracy of the rapid MMP-8 test were comparable to those of the ELISA IL-6 test; and (4) importantly, the rapid MMP-8 test had 100% sensitivity and 100% negative predictive value in the identification of neonates affected with fetal inflammatory response syndrome (FIRS). Conclusion The rapid diagnosis of intra-amniotic inflammation is possible by analysis of amniotic fluid using a point-of-care test for MMP-8. Patients with a positive test are at risk of delivering a neonate affected with systemic inflammation, a risk factor for adverse neonatal outcome.

The cytokine network in women with an asymptomatic short cervix and the risk of preterm delivery.

Abstract: Problem To characterize the amniotic fluid (AF) inflammatory-related protein (IRP) network in patients with a sonographic short cervix (SCx) and to determine its relation to early preterm delivery (ePTD). Method of study A retrospective cohort study included women with a SCx (≤25 mm; n=223) who had amniocentesis and were classified according to gestational age (GA) at diagnosis and delivery (ePTD <32 weeks of gestation). Results (i) In women with a SCx ≤ 22 1/7 weeks, the concentration of most IRPs increased as the cervix shortened; those with ePTD had a higher rate of increase in MIP-1α, MCP-1, and IL-6 concentrations than those delivering later; and (ii) the concentration of most IRPs and the correlation between several IRP pairs were higher in the ePTD group than for those delivering later. Conclusion Women with a SCx at 16-22 1/7 weeks have a unique AF cytokine network that correlates with cervical length at diagnosis and GA at delivery. This network may aid in predicting ePTD.

Neutrophil extracellular traps in acute chorioamnionitis: A mechanism of host defense.

Abstract: Problem Neutrophil extracellular traps (NETs) were recently described as a mechanism for microbial killing in the amniotic cavity of women with intra-amniotic infection. Such a clinical condition can result in acute chorioamnionitis, a placental lesion characterized by the infiltration of maternal neutrophils in the chorioamniotic membranes. Herein, we investigated whether these infiltrating neutrophils form NETs in the chorioamniotic membranes from women who underwent spontaneous term or preterm labor with acute chorioamnionitis. Method of study Chorioamniotic membrane samples were collected from women who underwent spontaneous term or preterm labor with acute chorioamnionitis (n=10 each). Controls included chorioamniotic membrane samples from women who delivered at term or preterm with or without labor in the absence of acute chorioamnionitis (n=10 each). NETs were visualized and semiquantified in the chorioamniotic membranes by using antibodies against neutrophil elastase and histone H3 in combination with DAPI staining. Results Neutrophil extracellular traps were abundant in the chorioamniotic membranes from women who underwent spontaneous term or preterm labor with acute chorioamnionitis. NETs were rarely found, or not visualized at all, in the chorioamniotic membranes from women who delivered at term or preterm with or without labor in the absence of acute chorioamnionitis. Conclusion Neutrophil extracellular traps are abundant in the chorioamniotic membranes from women who underwent spontaneous term or preterm labor with acute chorioamnionitis. These findings suggest that chorioamniotic neutrophils can form NETs as a mechanism of host defense against infection or danger signals.

A Role for the Inflammasome in Spontaneous Labor at Term with Acute Histologic Chorioamnionitis.

Abstract: Inflammasomes are cytosolic signaling platforms that regulate the activation of caspase (CASP)-1, which induces the maturation of interleukin (IL)-1β and IL-18. Herein, we determined whether the chorioamniotic membranes from women in spontaneous labor at term with acute histologic chorioamnionitis express major inflammasome components and whether these changes are associated with the activation of CASP-1 and CASP-4 and the release of mature IL-1β and IL-18. When comparing the chorioamniotic membranes from women in spontaneous labor at term with acute histologic chorioamnionitis to those without this placental lesion, we found that (1) the messenger RNA (mRNA) abundance of NLR family pyrin domain containing 3 ( NLRP3), NLR family CARD domain containing 4 ( NLRC4), absent in melanoma 2 ( AIM2), and nucleotide binding oligomerization domain 2 ( NOD2) was higher; (2) the NLRP3 and NLRC4 protein quantities were increased; (3) the mRNA and protein expressions of CASP-1 and its active forms were greater; (4) CASP-4 was increased at the mRNA level only; (5) the mRNA and protein expressions of IL-1β and its mature form were higher; and (6) a modest increase in the total protein concentration and abundance of the mature form of IL-18 was observed. In vitro incubation of the chorioamniotic membranes with the CASP-1 inhibitor, VX765, decreased the release of endotoxin-induced IL-1β and IL-18 (2-fold) but not IL-6 or tumor necrosis factor α. In conclusion, spontaneous labor at term with acute histologic chorioamnionitis is characterized by an upregulation of inflammasome components which, in turn, may participate in the activation of CASP-1 and lead to the release of mature IL-1β by the chorioamniotic membranes. These results support a role for the inflammasome in the mechanisms responsible for spontaneous labor at term with acute histologic chorioamnionitis.

Cutting Edge: Fetal/Placental Type I IFN Can Affect Maternal Survival and Fetal Viral Load during Viral Infection.

Abstract: Pregnant women have greater mortality and complications associated with viral infections compared with the general population, but the reason for the increased susceptibility is not well defined. Placenta type I IFN is an important immune modulator and protects the pregnancy. We hypothesized that loss of placental IFN affects the regulation of the maternal immune system, resulting in the differential response to infections observed in pregnancy. Pregnant mice lacking the IFN-α/β receptor (IFNAR) became viremic and had higher mortality compared with nonpregnant animals. Notably, an embryo with functional IFN signaling alone was sufficient to rescue the pregnant IFNAR-/- dam from virus-associated demise. Placental IFN was also an important regulator of viral replication in placental tissue and significantly affected viral transmission to the fetus. These findings highlight the role of fetal/placental IFN in the modulation of viral infection in the mother and fetus.

Mutations in fetal genes involved in innate immunity and host defense against microbes increase risk of preterm premature rupture of membranes (PPROM).

Abstract: Background Twin studies have revealed a significant contribution of the fetal genome to risk of preterm birth. Preterm premature rupture of membranes (PPROM) is the leading identifiable cause of preterm delivery. Infection and inflammation of the fetal membranes is commonly found associated with PPROM. Methods We carried out whole exome sequencing (WES) of genomic DNA from neonates born of African-American mothers whose pregnancies were complicated by PPROM (76) or were normal term pregnancies (N = 43) to identify mutations in 35 candidate genes involved in innate immunity and host defenses against microbes. Targeted genotyping of mutations in the candidates discovered by WES was conducted on an additional 188 PPROM cases and 175 controls. Results We identified rare heterozygous nonsense and frameshift mutations in several of the candidate genes, including CARD6, CARD8, DEFB1, FUT2, MBL2, NLP10, NLRP12, and NOD2. We discovered that some mutations (CARD6, DEFB1, FUT2, MBL2, NLRP10, NOD2) were present only in PPROM cases. Conclusions We conclude that rare damaging mutations in innate immunity and host defense genes, the majority being heterozygous, are more frequent in neonates born of pregnancies complicated by PPROM. These findings suggest that the risk of preterm birth in African-Americans may be conferred by mutations in multiple genes encoding proteins involved in dampening the innate immune response or protecting the host against microbial infection and microbial products.

Inflammasome assembly in the chorioamniotic membranes during spontaneous labor at term

Abstract: Problem Inflammasome activation requires two steps: priming and assembly of the multimeric complex. The second step includes assembly of the sensor molecule and adaptor protein ASC (an apoptosis-associated speck-like protein containing a CARD), which results in ASC speck formation and the recruitment of caspase (CASP)-1. Herein, we investigated whether there is inflammasome assembly in the chorioamniotic membranes and choriodecidual leukocytes from women who underwent spontaneous labor at term. Method of Study Using in situ proximity ligation assays, ASC/CASP-1 complexes were determined in the chorioamniotic membranes from women who delivered at term without labor or underwent spontaneous labor at term with or without acute histologic chorioamnionitis (n=10-11 each). Also, ASC speck formation was determined by flow cytometry in the choriodecidual leukocytes isolated from women who delivered at term with or without spontaneous labor (n=9-12 each). Results (i) ASC/CASP-1 complexes were detected in the chorioamniotic membranes; (ii) ASC/CASP-1 complexes were greater in the chorioamniotic membranes from women who underwent spontaneous labor at term than in those without labor; (iii) ASC/CASP-1 complexes were even more abundant in the chorioamniotic membranes from women who underwent spontaneous labor at term with acute histologic chorioamnionitis than in those without this placental lesion; (iv) ASC speck formation was detected in the choriodecidual leukocytes; and (v) ASC speck formation was greater in the choriodecidual leukocytes isolated from women who underwent spontaneous labor at term than in those without labor. Conclusion There is inflammasome assembly in the chorioamniotic membranes and choriodecidual leukocytes during spontaneous labor at term.

In vivo activation of invariant natural killer T cells induces systemic and local alterations in T-cell subsets prior to preterm birth.

Abstract: Preterm birth, the leading cause of neonatal morbidity and mortality worldwide, is frequently preceded by spontaneous preterm labour, a syndrome of multiple aetiologies. Pathological inflammation is causally linked to spontaneous preterm labour. Indeed, direct activation of invariant natural killer T (iNKT) cells via α-galactosylceramide induces preterm labour/birth largely by initiating systemic and local (i.e. decidua and myometrium) innate immune responses. Herein, we investigated whether iNKT-cell activation altered local and systemic T-cell subsets. Administration of α-galactosylceramide induced an expansion of activated CD1d-restricted iNKT cells in the decidua and a reduction in the number of: (1) total T cells (conventional CD4+ and CD8+ T cells) through the down-regulation of the CD3ɛ molecule in the peripheral circulation, spleen, uterine-draining lymph nodes (ULNs), decidua and/or myometrium; (2) CD4+ regulatory T cells in the spleen, ULNs and decidua; (3) T helper type 17 (Th17) cells in the ULNs but an increase in the number of decidual Th17 cells; (4) CD8+ regulatory T cells in the spleen and ULNs; and (5) CD4+ and CD8+ forkhead box protein 3 negative (Foxp3- ) responder T cells in the spleen and ULNs. As treatment with rosiglitazone prevents iNKT-cell activation-induced preterm labour/birth, we also explored whether the administration of this peroxisome proliferator-activated receptor gamma (PPARγ) agonist would restore the number of T cells. Treating α-galactosylceramide-injected mice with rosiglitazone partially restored the number of T cells in the spleen but not in the decidua. In summary, iNKT-cell activation altered the systemic and local T-cell subsets prior to preterm labour/birth; however, treatment with rosiglitazone partially reversed such effects.

Fetal death: An extreme manifestation of maternal anti-fetal rejection

Abstract: Objective The aim of this study was to determine the association between chronic placental inflammation and amniotic fluid (AF) markers of maternal anti-fetal rejection as well as the presence of microorganisms in the AF fluid of patients with fetal death. Study design This cohort study included 40 patients with fetal death whose placentas were examined for chronic inflammatory lesions and whose AF chemokine ligand (CXCL)10 and interleukin (IL)-6 concentrations were determined by immunoassays. AF was processed for bacteria, mycoplasmas and viruses using cultivation and molecular microbiologic techniques (i.e. PCR-ESI/MS). Results (1) The most prevalent placental findings were maternal vascular underperfusion (63.2%, 24/38), followed by chronic inflammatory lesions (57.9%, 22/38); (2) chronic chorioamnionitis (18/38) was three times more frequent than villitis of unknown etiology (6/38); (3) an elevated AF CXCL10 concentration (above the 95th centile) was present in 60% of the cases, and a receiver operating characteristics (ROC)-derived cut-off of 2.9 ng/mL had a sensitivity of 73% and a specificity of 75% in the identification of chronic placental inflammatory lesions; (4) only five cases had microbial invasion of the amniotic cavity, and the presence of microorganisms did not correlate with chronic placental inflammation. Conclusion In women with unexplained fetal death, there is an association between elevated AF CXCL10 and chronic placental inflammatory lesions. Therefore, we conclude that a subset of patients with fetal death may have endured a breakdown of maternal-fetal tolerance, which cannot be attributed to microorganisms in the amniotic cavity.

Proposed Diagnostic Criteria for the Case Definition of Amniotic Fluid Embolism in Research Studies

Abstract: Baylor College of Medicine, Houston, TX; Perinatology Research Branch of the Division of Obstetrics and Maternal-Fetal Medicine, Division of Intramural Research, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Detroit, MI, and Bethesda, MD; Columbia University, New York, NY; Centers for Disease Control and Prevention, Atlanta, GA; Banner Health, Phoenix, AZ; University of Texas Medical Branch, Galveston, TX Copyright r 2017 Wolters Kluwer Health, Inc. All rights reserved. DOI: 10.1097/01.aoa.0000521205.50664.87

Neonatal uterine bleeding as a biomarker for reproductive disorders during adolescence: a worldwide call for systematic registration by nurse midwife.

Abstract: Neonatal uterine bleeding (NUB) occurs in approximately 5% of newborns and is generally considered to be of little clinical significance. However, the real clinical importance of this condition and its long-term implications remain to be determined. The reason why NUB is rare despite high circulating levels of progesterone can be attributed to a progesterone resistance present in a majority of neonates. Recent work indicates that NUB represents a significant biomarker for events that can occur later-on during adolescence. Indeed, clinical studies have shown that "neonatal menstruation" constitutes a sign of fetal distress during late pregnancy, reflecting a stage of endometrium development that may subsequently have an impact on the reproductive life of the adolescent and the young adult. Via retrograde flow, NUB can cause endometrial stem/progenitor cells to arrive into the pelvic cavity and survive there, dormant underneath the peritoneal surface, until menarche activates them. Indeed, there is both clinical and epidemiological evidence of a link between NUB and adolescent endometriosis. In addition, if progesterone resistance persists till the onset of menarche, in case of an early teen pregnancy, it can result in a disorder of deep placentation. Therefore, we propose that NUB should be carefully recorded so that prospective studies can examine its links with reproductive disorders in adolescence and beyond.

Color and power Doppler combined with Fetal Intelligent Navigation Echocardiography (FINE) to evaluate the fetal heart.

Abstract: Objective To evaluate the performance of color and bidirectional power Doppler ultrasound combined with Fetal Intelligent Navigation Echocardiography (FINE) in examining the fetal heart. Methods A prospective cohort study was conducted of fetuses in the second and third trimesters with a normal heart or with congenital heart disease (CHD). One or more spatiotemporal image correlation (STIC) volume datasets, combined with color or bidirectional power Doppler (S-flow) imaging, were acquired in the apical four-chamber view. Each successfully obtained STIC volume was evaluated by STICLoop™ to determine its appropriateness before applying the FINE method. Visualization rates for standard fetal echocardiography views using diagnostic planes and/or Virtual Intelligent Sonographer Assistance (VIS-Assistance®) were calculated for grayscale (removal of Doppler signal), color Doppler and S-flow Doppler. In four cases with CHD (one case each of tetralogy of Fallot, hypoplastic left heart and coarctation of the aorta, interrupted inferior vena cava with azygos vein continuation and asplenia, and coarctation of the aorta with tricuspid regurgitation and hydrops), the diagnostic potential of this new technology was presented. Results A total of 169 STIC volume datasets of the normal fetal heart (color Doppler, n = 78; S-flow Doppler, n = 91) were obtained from 37 patients. Only a single STIC volume of color Doppler and/or a single volume of S-flow Doppler per patient were analyzed using FINE. Therefore, 60 STIC volumes (color Doppler, n = 27; S-flow Doppler, n = 33) comprised the final study group. Median gestational age at sonographic examination was 23 (interquartile range, 21–27.5) weeks. Color Doppler FINE generated nine fetal echocardiography views (grayscale) using (1) diagnostic planes in 73–100% of cases, (2) VIS-Assistance in 100% of cases, and (3) a combination of diagnostic planes and/or VIS-Assistance in 100% of cases. The rate of generating successfully eight fetal echocardiography views with appropriate color and S-flow Doppler information was 89–100% and 91–100% of cases, respectively, using a combination of diagnostic planes and/or VIS-Assistance. However, the success rate for the ninth echocardiography view (i.e. superior and inferior venae cavae) was 33% and 30% for color and S-flow Doppler, respectively. In all four cases of CHD, color Doppler FINE demonstrated evidence of abnormal fetal cardiac anatomy and/or hemodynamic flow. Conclusions The FINE method applied to STIC volumes of normal fetal hearts acquired with color or bidirectional power Doppler information can generate successfully eight to nine standard fetal echocardiography views (via grayscale, color Doppler or power Doppler) in the second and third trimesters. In cases of CHD, color Doppler FINE demonstrates successfully abnormal anatomy and/or Doppler flow characteristics. Published 2017. This article is a U.S. Government work and is in the public domain in the USA. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of the International Society of Ultrasound in Obstetrics and Gynecology.

A Prospective Study of the Use of Fetal Intelligent Navigation Echocardiography (FINE) to Obtain Standard Fetal Echocardiography Views.

Abstract: Objective: To evaluate the performance of Fetal Intelligent Navigation Echocardiography (FINE) applied to spatiotemporal image correlation (STIC) volume datasets of the normal fetal heart in generating standard fetal echocardiography views. Methods: In this prospective cohort study of patients with normal fetal hearts (19-30 gestational weeks), one or more STIC volume datasets were obtained of the apical four-chamber view. Each STIC volume successfully obtained was evaluated by STICLoop™ to determine its appropriateness before applying the FINE method. Visualization rates for standard fetal echocardiography views using diagnostic planes and/or Virtual Intelligent Sonographer Assistance (VIS-Assistance®) were calculated. Results: One or more STIC volumes (total n = 463) were obtained from 246 patients. A single STIC volume per patient was analyzed using the FINE method. In normal cases, FINE was able to generate nine fetal echocardiography views using: (1) diagnostic planes in 76-100% of the cases, (2) VIS-Assistance® in 96-100% of the cases, and (3) a combination of diagnostic planes and/or VIS-Assistance® in 96-100% of the cases. Conclusion: FINE applied to STIC volumes can successfully generate nine standard fetal echocardiography views in 96-100% of cases in the 2nd and 3rd trimesters. This suggests that the technology can be used as a method of screening for congenital heart disease.