Epidemiology of stillbirths based on different gestational thresholds at a tertiary hospital.

INTRODUCTION: The stillbirth rate (SBR) is an important public health indicator. We studied the distribution of maternal and fetal characteristics and time trends of the SBR at KK Women's and Children's Hospital (KKH), Singapore, from 2004 to 2016 based on various definitions of stillbirth. METHODS: Data was obtained from the Data Warehouse and Stillbirth Reporting System of KKH from 2004 to 2016. SBRs were calculated based on three definitions (fetal deaths at ≥ 20 weeks, 24 weeks or 28 weeks of gestation per 1,000 total births) and were described with maternal and fetal characteristics, and by year. RESULTS: From 2004 to 2016, the SBR declined by 44.7%, 25.5% and 18.9% based on Definitions I, II and III, respectively. The SBR at KKH in 2016 was 5.2 (Definition I), 4.1 (Definition II) and 3.0 (Definition III) per 1,000 total births. The SBR was significantly higher in women aged ≥ 35 years, nulliparas and female fetuses. The number of live births at 24-27+6 weeks of gestation was more than four times higher than that of stillbirths (822 vs. 176). There were 104 (12.7%) neonatal deaths during this gestation period, giving a high survival rate of 87.3%. CONCLUSION: The SBR in KKH is relatively lower than that in other developed countries. There is a need to consider revising our hospital and national definitions of the stillbirth lower boundary from 28 weeks to 24 weeks of gestation. This would allow us to make better comparisons with other developed countries, in line with improvements in healthcare.

Single-Cell RNA Sequencing of the Cynomolgus Macaque Testis Reveals Conserved Transcriptional Profiles during Mammalian Spermatogenesis.

Spermatogenesis is highly orchestrated and involves the differentiation of diploid spermatogonia into haploid sperm. The process is driven by spermatogonial stem cells (SSCs). SSCs undergo mitotic self-renewal, whereas sub-populations undergo differentiation and later gain competence to initiate meiosis. Here, we describe a high-resolution single-cell RNA-seq atlas of cells derived from Cynomolgus macaque testis. We identify gene signatures that define spermatogonial populations and explore self-renewal versus differentiation dynamics. We detail transcriptional changes occurring over the entire process of spermatogenesis and highlight the concerted activity of DNA damage response (DDR) pathway genes, which have dual roles in maintaining genomic integrity and effecting meiotic sex chromosome inactivation (MSCI). We show remarkable similarities and differences in gene expression during spermatogenesis with two other eutherian mammals, i.e., mouse and humans. Sex chromosome expression in the male germline in all three species demonstrates conserved features of MSCI but divergent multicopy and ampliconic gene content.

Angiogenic factors during pregnancy in Asian women with elevated blood pressure in early pregnancy and the risk of preeclampsia: a longitudinal cohort study.

OBJECTIVE: It remains unclear what roles placenta-originated angiogenic factors play in the pathogenesis of preeclampsia among hypertensive women. We compared maternal soluble fms-like tyrosine kinase 1 (sFlt-1) and placental growth factor (PlGF) levels throughout pregnancy in women with normal blood pressure (BP), elevated BP and hypertension in early pregnancy and their risks of developing preeclampsia. DESIGN: A prospective cohort study. SETTING: KK Women's and Children's Hospital, Singapore. PARTICIPANTS: 923 women with singleton pregnancy <14 weeks of gestation were included in the prospective Neonatal and Obstetrics Risks Assessment cohort between September 2010 and October 2014. Systolic, diastolic, mean arterial blood pressure (MAP) were measured at 11-14 weeks. PRIMARY AND SECONDARY OUTCOMES: Maternal serum sFlt-1, PlGF and sFlt-1/PlGF ratio were tested at 11-14, 18-22, 28-32 and 34 weeks onwards of gestation. Preeclampsia was main pregnancy outcome. RESULTS: Women were divided based on their BP in early pregnancy: normal (n=750), elevated BP (n=98) and hypertension (n=75). Maternal sFlt-1 levels and sFlt-1/PlGF ratios were higher in hypertensive women throughout pregnancy, but maternal PlGF levels were not significantly lower. Rise in maternal systolic, diastolic BP and MAP at 11-14 weeks were significantly associated with higher sFlt-1/PlGF ratios during pregnancy. A 10 mm Hg increase in MAP was associated with a 5.6-fold increase in risk of preterm preeclampsia and a 3.3-fold increase in risk of term preeclampsia, respectively. CONCLUSION: Women with elevated BP in early pregnancy already had a higher sFlt-1/PlGF ratio in early gestation and throughout pregnancy, and an increased risk of preeclampsia. In contrast, PlGF levels in these women remained normal.

Are women in Singapore gaining weight appropriately during pregnancy: a prospective cohort study.

BACKGROUND: We aimed to study gestational weight gain (GWG) in a Singaporean population and compare it with Institute of Medicine (IOM) 2009 GWG guidelines. METHODS: Nine hundred twenty-six women with low-risk singleton pregnancy were enrolled in a prospective cohort study from 2010 to 2014 in a Singapore tertiary maternity hospital. Seven hundred twenty-four patients had maternal weight information till term pregnancy and were included in analysis. Participants were categorized according to their first antenatal visit body mass index (BMI) as underweight, normal weight, overweight and obese. Total GWG for each BMI group was calculated. Multivariate logistic regression was performed to determine the predictors of total GWG above and below IOM guidelines. RESULTS: Obese women had a mean total GWG (9.1 kg) that exceeded the upper limit IOM guidelines (9 kg). In multivariate analysis of predictors of total GWG above IOM guidelines, being overweight (adjusted OR: 3.91 [95% CI, 2.60-5.88]; p < .0001) and obese (adjusted OR: 4.78 [95% CI, 2.80, 8.15]; p < .0001) significantly increased the risks of gaining weight above IOM guidelines during pregnancy, compared to being normal weight. CONCLUSIONS: Overweight and obesity are independent significant risk factors for gaining excessive gestational weight. Appropriate weight management for overweight and obese Singaporean women prior to and during pregnancy is important.

Serum sFlt-1/PlGF ratio has better diagnostic ability in early- compared to late-onset pre-eclampsia.

Background To establish gestational specific cutoffs for the soluble fms-like tyrosine kinase-1 (sFlt-1) and placental growth factor (PlGF) ratio as a diagnostic tool for pre-eclampsia (PE) in an Asian population. Methods 82 subjects (48 PE patients and 34 controls) were recruited. sFlt-1 and PlGF were analysed on the Roche Cobas e411 analyzer and their ratio was calculated. Diagnostic performance was evaluated using receiver-operating characteristics (ROC) curves. Optimal cutoffs for sFlt-1/PlGF ratio were determined for different gestation phases. Results The most optimal cut-off for the study group is 32 with a sensitivity and specificity of 85.1% and 100% and Youden Index (J) of 0.85. Applying this cutoff for early-onset PE (EO-PE), sensitivity increased to 95.8% while specificity remains at 100% (J=0.96). However, for late onset PE (LO-PE), sensitivity decreases to 73.9% while specificity remains at 100% (J=0.74). Two cutoffs were further determined for EO-PE and LO-PE - the first focusing on high sensitivity; the second focusing on high specificity. For EO-PE, cutoff <17 yielded sensitivity of 100% and specificity of 94.4% (J=0.94) while cutoff ≥32 yielded sensitivity of 95.8% and specificity of 100% (J=0.95). For LO-PE, cutoff <22 has a sensitivity of 82.6% and a specificity of 91.7% (J=0.74) while cutoff ≥32 yielded sensitivity of 73.9% and specificity of 100% (J=0.74). Conclusion While our study found an overall cutoff at 32 regardless of gestation age, it has limited diagnostic accuracy for LO-PE in our study. Multiple cutoffs focusing on either high sensitivity or high specificity enhance the performance of the sFlt-1/PlGF ratio as a diagnostic tool for PE and contribute to the identification of women at risk of PE in our Asian region.

Comparing two regimens of intravaginal misoprostol with intravaginal gemeprost for second-trimester pregnancy termination: a randomised controlled trial.

AIM: To compare the efficacy and safety of intravaginal misoprostol 200 µg, 400 µg and gemeprost regimens for second-trimester termination of pregnancy (TOP). METHODS: A three- armed randomi sed controlled trial (Clinical Trial Certificate 1100015) where 116 women undergoing second-trimester TOP were given intravaginal misoprostol 200 µ g (n=37), misoprostol 400 µg (n=40) or gemeprost 1 mg (n=39) at 4- hour intervals until abortion occurred with a maximum of five doses. RESULTS: The misoprostol 400 µg group had the highest incidence of successful abortions (92.5%) compared to the misoprostol 200 µg (70.3%; p=0.017) and gemeprost 1 mg (74.4%; p=0.037) within 48 hours. There was no significant difference in abortion rate between misoprostol 200 µg and gemeprost. The misoprostol 400 µg group had the highest incidence of fever (70.0%) compared to misoprostol 200 µg (24.3%; p<0.001) and gemeprost 1 mg (46.2%; p=0.041). The gemeprost group had the highest incidence of diarrhoea (38.5%) compared to misoprostol 400 µg (10.0%; p=0.004) and misoprostol 200 µg (8.1%; p=0.003) groups. CONCLUSIONS: Intravaginal misoprostol 400 µ g at 4- hour intervals was the most effective regimen but was associated with a high incidence of fever. Misoprostol 200 µg demonstrated similar effectiveness as gemeprost and had lower incidence of diarrhoea. Gemeprost should not be first line for medical therapy given the cost, storage requirements and lower efficacy.

Gestational age-specific reference intervals for serum thyroid hormone levels in a multi-ethnic population.

BACKGROUND: Thyroid disorders are common during pregnancy. To date, a limited number of studies have reported differences in serum thyroid hormone concentrations between different ethnic groups. We sought to establish gestational age-specific reference intervals for serum levels of thyroid hormones in a multi-ethnic population and investigate whether separate reference intervals should be used for different ethnic groups. METHODS: A total of 926 pregnant women from multiple ethnic groups attended four separate study visits spanning the three trimesters. Venous blood samples were taken at 9 to 14 weeks, 18 to 22 weeks, 28 to 32 weeks, and 34 to 39 weeks of gestation. Serum concentrations of thyroid-stimulating hormone (TSH), free thyroxine (T4), free triiodothyronine (T3), total T4, total T3, thyroid peroxidase antibody and thyroglobulin antibody were measured using Abbott Architect immunoassays. A total of 562 women with singleton pregnancies were found to be negative for both thyroid autoantibodies at all four study visits and thus included in the reference sample group for the establishment of reference intervals (2.5th to 97.5th percentiles). RESULTS: Reference intervals for serum thyroid hormones at 9-14 weeks of gestation derived from the combined group of pregnant women are as follows: TSH, 0.01-2.39 mIU/L; free T4, 11.4-19.5 pmol/L; free T3, 4.23-6.69 pmol/L; total T4, 77.8-182.4 nmol/L; total T3, 1.39-2.97 nmol/L. No differences in the five thyroid parameters' reference intervals are detectable among the ethnic groups except that at study visit 3 (28-32 weeks of gestation), the upper reference limit of total T3 in Malays (3.20 nmol/L; 90% CI, 2.99-3.76 nmol/L) is slightly higher than that in Chinese (2.86 nmol/L; 90% CI, 2.70-2.98 nmol/L). CONCLUSIONS: The findings from this study on a multi-ethnic cohort highlight the importance of establishing locally derived and gestational age-specific reference intervals for the five thyroid hormone parameters.

Extracellular vesicles yield predictive pre-eclampsia biomarkers.

Circulating extracellular vesicles (EVs) such as cholera toxin B chain (CTB)- or annexin V (AV)-binding EVs were previously shown to be rich sources of biomarkers. Here we test if previously identified pre-eclampsia (PE) candidate biomarkers, TIMP-1 in CTB-EVs (CTB-TIMP) and PAI-1 in AV-EVs (AV-PAI) complement plasma PlGF in predicting PE in a low-risk obstetric population. Eight hundred and forty-three prospectively banked plasma samples collected at 28 + 0 to 32 + 0 gestation weeks in the Neonatal and Obstetrics Risk Assessment (NORA) cohort study were assayed by sandwich ELISAs for plasma PlGF, CTB-TIMP1 and AV-PAI1. Nineteen patients subsequently developed PE 7.3 (±2.9) weeks later at a mean gestational age of 36.1 ± 3.5 weeks. The biomarkers were assessed for their predictive accuracy for PE using stepwise multivariate logistic regression analysis with Firth correction and Areas under the curve (AUC). To achieve 100% sensitivity in predicting PE, the cut-off for plasma PlGF, CTB-TIMP1 & AV-PAI1 were set at <1235, ≤300 or >1300 and <10,550 pg/mL plasma, respectively. The corresponding AUCs, specificity and PPV at a 95% confidence interval were 0.92, 52.1% and 4.7%; 0.72, 44.5% and 4.0%; and 0.69, 21.5% and 2.9%, respectively. At 100% sensitivity, the three biomarkers had a combined AUC of 0.96, specificity of 78.6%, and PPV of 9.9%. This is the first large cohort validation of the utility of EV-associated analytes as disease biomarkers. Specifically, EV biomarkers enhanced the predictive robustness of an existing PE biomarker sufficiently to justify PE screening in a low-risk general obstetric population.

Incremental Cost-Effectiveness Analysis of Gestational Diabetes Mellitus Screening Strategies in Singapore.

The objective of this study was to conduct an incremental cost-effectiveness analysis from the payer's perspective in Singapore of 3 gestational diabetes mellitus screening strategies: universal, targeted, or no screening. A decision tree model assessed the primary outcome: incremental cost per quality-adjusted life year (QALY) gained. Probabilities, costs, and utilities were derived from the literature, the Growing Up in Singapore Towards healthy Outcomes (GUSTO) birth cohort study, and the KK Women's and Children's Hospital's database. Relative to targeted screening using risk factors, universal screening generates an incremental cost-effectiveness ratio (ICER) of $USD10,630/QALY gained. Sensitivity analyses show that disease prevalence rates and intervention effectiveness of glycemic management have the biggest impacts on the ICERs. Based on the model and best available data, universal screening is a cost-effective approach for reducing the complications of gestational diabetes mellitus in Singapore as compared with the targeted screening approach or no screening.

Plasma biomarker discovery in preeclampsia using a novel differential isolation technology for circulating extracellular vesicles.

OBJECTIVE: To circumvent the complex protein milieu of plasma and discover robust predictive biomarkers for preeclampsia (PE), we investigate if phospholipid-binding ligands can reduce the milieu complexity by extracting plasma extracellular vesicles for biomarker discovery. STUDY DESIGN: Cholera toxin B chain (CTB) and annexin V (AV) which respectively binds GM1 ganglioside and phosphatidylserine were used to isolate extracellular vesicles from plasma of PE patients and healthy pregnant women. The proteins in the vesicles were identified using enzyme-linked immunosorbent assay, antibody array, and mass spectrometry. RESULTS: CTB and AV were found to bind 2 distinct groups of extracellular vesicles. Antibody array and enzyme-linked immunosorbent assay revealed that PE patients had elevated levels of CD105, interleukin-6, placental growth factor, tissue inhibitor of metallopeptidase 1, and atrial natriuretic peptide in cholera toxin B- but not AV-vesicles, and elevated levels of plasminogen activator inhibitor-1, pro-calcitonin, S100b, tumor growth factor ß, vascular endothelial growth factor receptor 1, brain natriuretic peptide, and placental growth factor in both cholera toxin B- and AV-vesicles. CD9 level was elevated in cholera toxin B-vesicles but reduced in AV vesicles of PE patients. Proteome analysis revealed that in cholera toxin B-vesicles, 87 and 222 proteins were present only in PE patients and healthy pregnant women respectively while in AV-vesicles, 104 and 157 proteins were present only in PE and healthy pregnant women, respectively. CONCLUSION: This study demonstrated for the first time that CTB and AV bind unique extracellular vesicles, and their protein cargo reflects the disease state of the patient. The successful use of these 2 ligands to isolate circulating plasma extracellular vesicles for biomarker discovery in PE represents a novel technology for biomarker discovery that can be applied to other specialties.