Indicated vs universal third-trimester ultrasound examination in low-risk pregnancies: a pre-post-intervention study.

BACKGROUND: In low-risk pregnancies, a third-trimester ultrasound examination is indicated if fundal height measurement and gestational age discrepancy are observed. Despite potential improvement in the detection of ultrasound abnormality, prior trials to date on universal third-trimester ultrasound examination in low-risk pregnancies, compared with indicated ultrasound examination, have not demonstrated improvement in neonatal or maternal adverse outcomes. OBJECTIVE: The primary objective was to determine if universal third-trimester ultrasound examination in low-risk pregnancies could attenuate composite neonatal adverse outcomes. The secondary objectives were to compare changes in composite maternal adverse outcomes and detection of abnormalities of fetal growth (fetal growth restriction or large for gestational age) or amniotic fluid (oligohydramnios or polyhydramnios). STUDY DESIGN: Our pre-post intervention study at 9 locations included low-risk pregnancies, those without indication for ultrasound examination in the third trimester. Compared with indicated ultrasound in the preimplementation period, in the postimplementation period, all patients were scheduled for ultrasound examination at 36.0-37.6 weeks. In both periods, clinicians intervened on the basis of abnormalities identified. Composite neonatal adverse outcomes included any of: Apgar score ≤5 at 5 minutes, cord pH <7.00, birth trauma (bone fracture or brachial plexus palsy), intubation for >24 hours, hypoxic-ischemic encephalopathy, seizure, sepsis (bacteremia proven with blood culture), meconium aspiration syndrome, intraventricular hemorrhage grade III or IV, periventricular leukomalacia, necrotizing enterocolitis, stillbirth after 36 weeks, or neonatal death within 28 days of birth. Composite maternal adverse outcomes included any of the following: chorioamnionitis, wound infection, estimated blood loss >1000 mL, blood transfusion, deep venous thrombus or pulmonary embolism, admission to intensive care unit, or death. Using Bayesian statistics, we calculated a sample size of 600 individuals in each arm to detect >75% probability of any reduction in primary outcome (80% power; 50% hypothesized risk reduction). RESULTS: During the preintervention phase, 747 individuals were identified during the initial ultrasound examination, and among them, 568 (76.0%) met the inclusion criteria at 36.0-37.6 weeks; during the postintervention period, the corresponding numbers were 770 and 661 (85.8%). The rate of identified abnormalities of fetal growth or amniotic fluid increased from between the pre-post intervention period (7.1% vs 22.2%; P<.0001; number needed to diagnose, 7; 95% confidence interval, 5-9). The primary outcome occurred in 15 of 568 (2.6%) individuals in the preintervention and 12 of 661 (1.8%) in the postintervention group (83% probability of risk reduction; posterior relative risk, 0.69 [95% credible interval, 0.34-1.42]). The composite maternal adverse outcomes occurred in 8.6% in the preintervention and 6.5% in the postintervention group (90% probability of risk; posterior relative risk, 0.74 [95% credible interval, 0.49-1.15]). The number needed to treat to reduce composite neonatal adverse outcomes was 121 (95% confidence interval, 40-200). In addition, the number to reduce composite maternal adverse outcomes was 46 (95% confidence interval, 19-74), whereas the number to prevent cesarean delivery was 18 (95% confidence interval, 9-31). CONCLUSION: Among low-risk pregnancies, compared with routine care with indicated ultrasound examination, implementation of a universal third-trimester ultrasound examination at 36.0-37.6 weeks attenuated composite neonatal and maternal adverse outcomes.

Mesenchymal Stem Cells Suppress Inflammatory Cytokines in Lipopolysaccharide Exposed Preterm and Term Human Pregnant Myometrial Cells.

Objective The objective of this study was to determine the cytokine response in human pregnant preterm and term myometrial cells exposed to lipopolysaccharide (LPS) and cocultured with mesenchymal stem cells (MSCs). Study Design Myometrium was obtained at cesarean delivery in term and preterm patients. Human myometrial cells were exposed to 5 µg/mL LPS for 4 hours followed by 1 µg/mL LPS for 24 hours and were cocultured with MSCs for 24 hours. Culture supernatants were collected at 24 hours and expression of cytokines, including interleukin-1ß (IL-1ß), IL-6, IL-8, tumor necrosis factor-α (TNF-α), transforming growth factor-ß (TGF-ß), and IL-10, was quantified by enzyme-linked immunosorbent assay. Results There was significantly increased expression of the proinflammatory cytokines IL-1ß, IL-6, IL-8, and TNF-α in preterm myometrial cells treated with LPS compared with untreated preterm myometrial cells. Coculture with MSCs significantly suppressed the proinflammatory cytokine levels in LPS-treated preterm versus treated term myometrial cells. Moreover, MSC cocultured preterm myometrial cells expressed increased levels of the anti-inflammatory cytokines TGF-ß and IL-10 compared with treated term myometrial cells. Conclusion MSCs ameliorate LPS-mediated inflammation in preterm human myometrial cells compared with term myometrial cells. Immunomodulatory effects of MSCs mediated through anti-inflammatory cytokine regulation suggest a potential cell-based therapy for preterm birth.

Mesenchymal Stem Cells Attenuate Lipopolysaccharide-Induced Inflammatory Response in Human Uterine Smooth Muscle Cells.

Objective The aim of this study was to determine if mesenchymal stem cells (MSCs) would suppress the inflammatory response in human uterine cells in an in vitro lipopolysaccharide (LPS)-based preterm birth (PTB) model. Study Design Cocultures of human uterine smooth muscle cells (HUtSMCs) and MSCs were exposed to 5 µg/mL LPS for 4 hours and further challenged with 1 µg/mL LPS for a subsequent 24 hours. Key elements of the parturition cascade regulated by toll-like receptors (TLRs) through activation of mitogen-activated protein kinases (MAPKs) were quantified in culture supernatant as biomarkers of MSC modulation. Results Coculture with MSCs significantly attenuated TLR-4, p-JNK, and p- extracellular signal-regulated kinase 1/2 (ERK1/2) protein levels compared with HUtSMCs monoculture ( p = 0.05). In addition, coculture was associated with significant inhibition of proinflammatory cytokines interleukin (IL)-6 and IL-8 ( p = 0.0001) and increased production of anti-inflammatory cytokines IL-10 and transforming growth factor (TGF)-ß1 ( p = 0.0001). Conclusion MSCs appear to play a role in significantly attenuating LPS-mediated inflammation via alteration of down-stream MAPKs. MSCs may represent a novel, cell-based therapy in women with increased risk of inflammatory-mediated preterm birth.

Auditory brainstem response (ABR) abnormalities across the life span of rats prenatally exposed to alcohol.

BACKGROUND: Fetal alcohol syndrome (FAS) is a leading cause of neurodevelopmental impairments (NDIs) in developed countries. Sensory deficits can play a major role in NDI, yet few studies have investigated the effects of prenatal alcohol exposure on sensory function. In addition, there is a paucity of information on the lifelong effects of prenatal alcohol exposure. Thus, we sought to investigate the effects of prenatal alcohol exposure on auditory function across the life span in an animal model. Based on prior findings with prenatal alcohol exposure and other forms of adverse prenatal environments, we hypothesized that animals prenatally exposed to alcohol would show an age-dependent pattern of (i) hearing and neurological abnormalities as postweanling pups, (ii) a substantial dissipation of such abnormalities in young adulthood, and (iii) a resurgence of such abnormalities in middle-aged adulthood. METHODS: Pregnant rats were randomly assigned to an untreated control (CON), a pair-fed control (PFC), or an alcohol-treated (ALC) group. The ALC dams were gavaged with 6 mg/kg alcohol daily from gestation day (GD) 6 to 21. The PFC dams were gavaged daily from GD6 to GD21 with an isocaloric and isovolumetric water-based solution of maltose-dextrins and pair-fed to the ALC dams. The CON dams were the untreated group to which the ALC and CON groups were compared. Hearing and neurological functions in the offspring were assessed with the auditory brainstem response (ABR) at the postnatal ages of 22, 220, and 520 days. RESULTS: In accord with our hypothesis, ABR abnormalities were first observed in the postweanling pups, largely dissipated in young adulthood, and then resurged in middle-aged adulthood. This age-related pattern suggests that the ALC pups had a developmental delay that dissipated in young adulthood and an enhanced age-related deterioration that occurred in middle-aged adulthood. Such a pattern is consistent with the fetal programming hypothesis of adult-onset diseases (the Barker hypothesis). CONCLUSIONS: Our findings have important clinical implications for the assessment and management of (i) childhood hearing disorders and their comorbidities (i.e., speech-and-language, learning, and attention deficit disorders) and (ii) enhanced age-related hearing and neurological degeneration in middle-aged adulthood that can result from prenatal alcohol exposure. We recommend hearing evaluation be a part of any long-term follow-up for FAS patients and patients exposed to any adverse prenatal environment.

A molecular signature of an arrest of descent in human parturition.

OBJECTIVE: This study was undertaken to identify the molecular basis of an arrest of descent. STUDY DESIGN: Human myometrium was obtained from women in term labor (TL; n = 29) and arrest of descent (AODes; n = 21). Gene expression was characterized using Illumina HumanHT-12 microarrays. A moderated Student t test and false discovery rate adjustment were applied for analysis. Confirmatory quantitative reverse transcription-polymerase chain reaction and immunoblot were performed in an independent sample set. RESULTS: Four hundred genes were differentially expressed between women with an AODes compared with those with TL. Gene Ontology analysis indicated enrichment of biological processes and molecular functions related to inflammation and muscle function. Impacted pathways included inflammation and the actin cytoskeleton. Overexpression of hypoxia inducible factor-1a, interleukin -6, and prostaglandin-endoperoxide synthase 2 in AODes was confirmed. CONCLUSION: We have identified a stereotypic pattern of gene expression in the myometrium of women with an arrest of descent. This represents the first study examining the molecular basis of an arrest of descent using a genome-wide approach.

Characterization of the transcriptome of chorioamniotic membranes at the site of rupture in spontaneous labor at term.

OBJECTIVE: The purpose of this study was to compare the transcriptome between the site of membrane rupture and the chorioamniotic membranes away from the site of rupture. STUDY DESIGN: The transcriptome of amnion and chorion (n=20 each) from and distal to the site of rupture from women with spontaneous labor and vaginal delivery at term after spontaneous rupture of membranes was profiled with Illumina HumanHT-12 microarrays. Selected genes were validated with the use of quantitative reverse transcription-polymerase chain reaction. RESULTS: Six hundred seventy-seven genes were differentially expressed in the chorion between the rupture and nonrupture sites (false discovery rate<0.1; fold change>1.5). Quantitative reverse transcription-polymerase chain reaction confirmed the differential expression in 10 of 14 genes. Enriched biological processes included anatomic structure development, cell adhesion and signal transduction. Extracellular matrix-receptor interaction was the most impacted signaling pathway. CONCLUSION: The transcriptome of fetal membranes after spontaneous rupture of membranes in term labor is characterized by region- and tissue-specific differential expression of genes that are involved in signature pathways, which include extracellular matrix-receptor interactions.

Emergence of hormonal and redox regulation of galectin-1 in placental mammals: implication in maternal-fetal immune tolerance.

Galectin-1 is an anti-inflammatory lectin with pleiotropic regulatory functions at the crossroads of innate and adaptive immunity. It is expressed in immune privileged sites and is implicated in establishing maternal-fetal immune tolerance, which is essential for successful pregnancy in eutherian mammals. Here, we show conserved placental localization of galectin-1 in primates and its predominant expression in maternal decidua. Phylogenetic footprinting and shadowing unveil conserved cis motifs, including an estrogen responsive element in the 5' promoter of LGALS1, that were gained during the emergence of placental mammals and could account for sex steroid regulation of LGALS1 expression, thus providing additional evidence for the role of galectin-1 in immune-endocrine cross-talk. Maximum parsimony and maximum likelihood analyses of 27 publicly available vertebrate and seven newly sequenced primate LGALS1 coding sequences reveal that intense purifying selection has been acting on residues in the carbohydrate recognition domain and dimerization interface that are involved in immune functions. Parsimony- and codon model-based phylogenetic analysis of coding sequences show that amino acid replacements occurred in early mammalian evolution on key residues, including gain of cysteines, which regulate immune functions by redox status-mediated conformational changes that disable sugar binding and dimerization, and that the acquired immunoregulatory functions of galectin-1 then became highly conserved in eutherian lineages, suggesting the emergence of hormonal and redox regulation of galectin-1 in placental mammals may be implicated in maternal-fetal immune tolerance.

Chorioamnionitis and increased galectin-1 expression in PPROM --an anti-inflammatory response in the fetal membranes?

PROBLEM: Galectin-1 can regulate immune responses upon infection and inflammation. We determined galectin-1 expression in the chorioamniotic membranes and its changes during histological chorioamnionitis. METHOD OF STUDY: Chorioamniotic membranes were obtained from women with normal pregnancy (n = 5) and from patients with pre-term pre-labor rupture of the membranes (PPROM) with (n = 8) and without histological chorioamnionitis (n = 8). Galectin-1 mRNA and protein were localized by in situ hybridization and immunohistochemistry. Galectin-1 mRNA expression was also determined by quantitative reverse transcriptase polymerase chain reaction. RESULTS: Galectin-1 mRNA and protein were detected in the amniotic epithelium, chorioamniotic fibroblasts/myofibroblasts and macrophages, chorionic trophoblasts, and decidual stromal cells. In patients with PPROM, galectin-1 mRNA expression in the fetal membranes was higher (2.07-fold, P = 0.002) in those with chorioamnionitis than in those without. Moreover, chorioamionitis was associated with a strong galectin-1 immunostaining in amniotic epithelium, chorioamniotic mesodermal cells, and apoptotic bodies. CONCLUSION: Chorioamnionitis is associated with an increased galectin-1 mRNA expression and strong immunoreactivity of the chorioamniotic membranes; thus, galectin-1 may be involved in the regulation of the inflammatory responses to chorioamniotic infection.

Severe preeclampsia is characterized by increased placental expression of galectin-1.

OBJECTIVE: Galectin-1 is a major anti-inflammatory protein expressed by the placenta and immune cells that can bias the character of inflammatory responses toward the Th2 type. Galectin-1 is expressed in immune privileged sites, it can facilitate immune tolerance and tumor immune escape, and it has been successfully used for the suppression of experimental autoimmune diseases as well as graft-versus-host disease in murine models. We propose that an abnormal immune response in some pregnancy complications may be associated with changes in placental expression of galectin-1. To test this hypothesis, we studied placental galectin-1 mRNA and protein expression and localization in women with preeclampsia (PE) and in those who delivered a small-for-gestational age (SGA) neonate. STUDY DESIGN: This cross-sectional study included pregnant women matched for gestational age at delivery in the following groups: (1) severe PE (n = 10), (2) severe PE complicated with SGA (n = 10), (3) SGA without PE (n = 10), and (4) controls (n = 10). Galectin-1 mRNA and protein were localized in placentas by in situ hybridization and immunofluorescence microscopy. Galectin-1 mRNA expression was determined by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR), and galectin-1 protein content by Western blot. Non-parametric statistics were used for analysis. RESULT: (1) In normal term placentas, galectin-1 mRNA or immunofluorescence signals were detected in the trophoblasts, villous stromal cells, Hofbauer cells, endothelial cells of the villous blood vessels, and the villous stroma. (2) Placental galectin-1 mRNA expression was significantly higher in severe PE (with or without SGA) than in controls (1.47-fold, p = 0.004; 1.44-fold, p = 0.003, respectively) and in SGA (1.68-fold, p = 0.001; 1.64-fold, p = 0.001, respectively). (3) Trophoblasts in placentas of patients with severe PE had the most intense galectin-1 immunostaining. CONCLUSIONS: (1) We report for the first time the placental expression and localization of galectin-1 mRNA and demonstrate that the protein is abundantly present in third trimester human placentas. (2) Placental galectin-1 expression is higher in severe PE than in normal pregnancy regardless of the presence of SGA. (3) However, it is not altered in SGA without PE. We propose that the increased placental expression of galectin-1 in patients with severe PE may represent a fetal response to an exaggerated systemic maternal inflammation; thus, galectin-1 may be implicated in maternal-fetal immune tolerance in humans.

Reduced auditory acuity in rat pups from excess and deficient omega-3 fatty acid consumption by the mother.

Consumption of the nutrients omega-3 fatty acids (omega-3 FA) during pregnancy and lactation is considered beneficial to fetal and infant development. It may also reduce the incidence and severity of preterm births by prolonging gestational length. However several recent human and animal studies have reported that over-supplementation with omega-3 FA, especially in the form of fish oil, can have adverse effects on fetal and infant development and the auditory brainstem response (ABR). Our goal was to assess further the effects of omega-3 FA excess and deficiency during pregnancy and lactation on the offspring's auditory acuity as evidenced by their ABR thresholds. Female Wistar rats were given diets that were either deficient, adequate (control) or excess in omega-3 FA from day 1 of pregnancy through lactation. The offspring were ABR-tested at the postnatal age of 24 days. The rat pups in the Excess treatment condition had significantly elevated (worse) ABR thresholds, postnatal growth restriction, and a trend for increased postnatal mortality in comparison to the Control group. The Deficient group was intermediate. In conclusion, excess or deficient amounts of omega-3 FA during pregnancy and lactation in the laboratory rat adversely affected the offspring's auditory acuity. Postnatal thriving was also adversely affected. Consuming or administering large or inadequate amounts of omega-3 FA during pregnancy and lactation seems inadvisable because of the potential for adverse effects on infant development.

Sildenafil citrate and fetal outcome in pregnant rats.

OBJECTIVE: To determine the effects of sildenafil citrate on fetal growth in maternal rats exposed to hypoxia. METHODS: Timed pregnant rats were randomized to either hypoxia or control on gestational days (GD) 18-20, and received either sildenafil (45 mg/kg) orally every 12 h on GD 18-21 or an equal volume of sterile water. Fetal pups were retrieved by laparotomy on GD 21. Pup weight and length were evaluated and cGMP measured in maternal and fetal blood. RESULTS: In the non-hypoxic rats, sildenafil exposure was associated with a decrease in size(4.75 +/- 0.43 vs. 5.11 +/- 0.34 g, p = 0.00). In contrast, in the hypoxic rat model, sildenafil exposure was associated with increased size of the offspring (5.48 +/- 0.45 vs. 5.16 +/- 0.36 g, p = 0.016). Maternal cGMP levels were increased in the presence of both sildenafil and hypoxia (23.0 +/- 10.5 vs. 15.6 +/- 2.7 pmol/ml, p = 0.001). CONCLUSION: Exposure to sildenafil in a non-hypoxic setting results in a decrease in fetal size. Sildenafil in the presence of a stimulus, hypoxia, will lead to increased fetal size. These results suggest that sildenafil may have some influence on fetal growth. How these effects occur and by what mechanism remain to be determined.

Prolonged in utero meconium exposure impairs spatial learning in the adult rat. Central Prize Award.

OBJECTIVE: The purpose of this study was to examine the effects of prolonged in utero meconium exposure on adult learning and memory, as measured by the Morris water maze. STUDY DESIGN: Timed pregnant Long-Evans rats were studied. On gestational day 20 (term, 21 days of gestation), laparotomy was performed, and each maternal animal received an injection of clear amniotic fluid or meconium-stained amniotic fluid into each gestational sac. The laparotomy incision was closed, and the animals received postoperative monitoring through delivery. On postnatal days 145 to 148, the offspring underwent Morris water maze testing. The mean (+/-SEM) for the latency time was reported for each day's trial and compared between groups. RESULTS: There were significant differences between meconium-stained amniotic fluid group and clear amniotic fluid group in the mean time to platform on day 1 (82.7 +/- 1.8 seconds vs 75.9 +/- 3.0 seconds; P=.04), day 2 (60.5 +/- 3.5 seconds vs 47. 8 +/- 4.6 seconds; P=.03), and day 3 (56.5 +/- 4.5 seconds vs 34.7 +/- 4.4 seconds; P=.001). However, there were no differences on days 4 and 5. There were also no differences between recall and response learning trials that were done after a 12-day retention period. CONCLUSION: In the absence of hypoxia or infection, prolonged in utero meconium exposure is associated with a delay of spatial learning in the adult rat.

Prenatal cocaine exposure influences the growth and life span of laboratory rats.

Laboratory rats prenatally exposed to alcohol, nicotine, amphetamine, undernutrition or hypoxia can exhibit shortened life span and other signs of enhanced age-related degeneration. We evaluated the possibility of similar effects following prenatal cocaine exposure. Pregnant rats received 20 or 40 mg/kg cocaine HCl subcutaneously (C20, C40), twice daily, from gestation days (GD) 7-20. Untreated control (UTC) and pair-fed control (PFC) groups were also used. The pregnant C40, C20, and PFC dams ate less food and gained less weight than the UTC dams did. The pregnant C40 and C20 dams drank more water than the UTC dams did, and the pregnant PFC dams drank less than the UTC dams did. The C40 and PFC offspring had delayed earflap openings. The C40 male and female offspring had lower birth weights than their cohorts in the other three groups. The C40 female and male offspring remained significantly underweight until postnatal day (PND) 28 and PND56, respectively. During young adulthood, the males and females in the C20, C40, and PFC groups had normal body weights. During old adulthood, however, the C20 and C40 males and the C20, C40, and PFC females developed reduced body weights as compared with their UTC cohorts. The C20 and C40 male offspring and the C20, C40, and PFC female offspring also had life spans that were 7-12% shorter than that of their UTC cohorts. Thus, groups that showed reduced body weights in old age also showed shorter life spans. These results provided converging evidence that prenatal cocaine exposure caused enhanced age-related degeneration. Observations on cardiac and other organ pathology were also made. Health implications for children born to cocaine-abusing women are discussed.

Timing of fetal nucleated red blood cell count elevation in response to acute hypoxia.

We determined the effect of an acute episode of severe hypoxia on peripheral nucleated red blood cell (RBC) counts in the fetal rat. Timed pregnant rats were randomized to a 2-hour exposure to hypoxia (placement in a chamber containing a gas mixture with 9% O2 +3% CO2 + balanced N2) or to a 2-hour exposure to a sham chamber containing room air. Two maternal animals per group underwent cesarean section immediately after the 2-hour period and then 4, 12, 24, 36, 48, and 60 h after exposure. Fetal nucleated RBC counts were compared between groups at each time interval. The nucleated RBC counts were not significantly different in the hypoxia group until 12 h (mean +/- SEM 158.0 +/- 22.4 RBC/10 high-power fields vs. 90.6 +/- 11.0; p = 0.03) and 24 h (133.2 +/- 16.0 vs. 84.1 +/- 9.0; p = 0.04) after exposure. There were no differences between groups 36, 48, and 60 h after exposure. In the near-term rat fetus, acute hypoxia was associated with a delayed but transient increase in peripheral nucleated RBC counts.