Evaluation of screening performance of first-trimester competing-risks prediction model for small-for-gestational age in Asian population.

OBJECTIVE: To examine the external validity of the Fetal Medicine Foundation (FMF) competing-risks model for the prediction of small-for-gestational age (SGA) at 11-14 weeks' gestation in an Asian population. METHODS: This was a secondary analysis of a multicenter prospective cohort study in 10 120 women with a singleton pregnancy undergoing routine assessment at 11-14 weeks' gestation. We applied the FMF competing-risks model for the first-trimester prediction of SGA, combining maternal characteristics and medical history with measurements of mean arterial pressure (MAP), uterine artery pulsatility index (UtA-PI) and serum placental growth factor (PlGF) concentration. We calculated risks for different cut-offs of birth-weight percentile (< 10th , < 5th or < 3rd percentile) and gestational age at delivery (< 37 weeks (preterm SGA) or SGA at any gestational age). Predictive performance was examined in terms of discrimination and calibration. RESULTS: The predictive performance of the competing-risks model for SGA was similar to that reported in the original FMF study. Specifically, the combination of maternal factors with MAP, UtA-PI and PlGF yielded the best performance for the prediction of preterm SGA with birth weight < 10th percentile (SGA < 10th ) and preterm SGA with birth weight < 5th percentile (SGA < 5th ), with areas under the receiver-operating-characteristics curve (AUCs) of 0.765 (95% CI, 0.720-0.809) and 0.789 (95% CI, 0.736-0.841), respectively. Combining maternal factors with MAP and PlGF yielded the best model for predicting preterm SGA with birth weight < 3rd percentile (SGA < 3rd ) (AUC, 0.797 (95% CI, 0.744-0.850)). After excluding cases with pre-eclampsia, the combination of maternal factors with MAP, UtA-PI and PlGF yielded the best performance for the prediction of preterm SGA < 10th and preterm SGA < 5th , with AUCs of 0.743 (95% CI, 0.691-0.795) and 0.762 (95% CI, 0.700-0.824), respectively. However, the best model for predicting preterm SGA < 3rd without pre-eclampsia was the combination of maternal factors and PlGF (AUC, 0.786 (95% CI, 0.723-0.849)). The FMF competing-risks model including maternal factors, MAP, UtA-PI and PlGF achieved detection rates of 42.2%, 47.3% and 48.1%, at a fixed false-positive rate of 10%, for the prediction of preterm SGA < 10th , preterm SGA < 5th and preterm SGA < 3rd , respectively. The calibration of the model was satisfactory. CONCLUSION: The screening performance of the FMF first-trimester competing-risks model for SGA in a large, independent cohort of Asian women is comparable with that reported in the original FMF study in a mixed European population. © 2023 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

Evaluating healthcare response to COVID-19 across Southeast Asia: A post-Pandemic Reflection and Way Forward.

This paper provides a comprehensive analysis of Southeast Asian countries' responses to the COVID-19 pandemic, particularly focusing on Malaysia, Singapore, Thailand, the Philippines, Indonesia, and Myanmar. The primary objective is to explore how the pandemic has evolved in these nations, how the respective healthcare delivery systems responded, and the current COVID-19 status within each country. It presents epidemiological trends and governmental strategies adopted in combating the pandemic. The paper also outlines lessons learned and future challenges, highlighting key areas like global health diplomacy, the need for collaboration, clear government agency communication, and a stance against social discrimination. It culminates in an assessment of the postpandemic landscape, discussing the transformation of public health policies and the socio-economic implications of pandemic management.

Artificial placenta technology: History, potential and perception.

As presently conceptualised, the artificial placenta (AP) is an experimental life support platform for extremely preterm infants (i.e. 400-600 g; 21-23+6 weeks of gestation) born at the border of viability. It is based around the oxygenation of the periviable fetus using gas-exchangers connected to the fetal vasculature. In this system, the lung remains fluid-filled and the fetus remains in a quiescent state. The AP has been in development for some sixty years. Over this time, animal experimental models have evolved iteratively from employing external pump-driven systems used to support comparatively mature fetuses (generally goats or sheep) to platforms driven by the fetal heart and used successfully to maintain extremely premature fetuses weighing around 600 g. Simultaneously, sizable advances in neonatal and obstetric care mean that the nature of a potential candidate patient for this therapy, and thus the threshold success level for justifying its adoption, have both changed markedly since this approach was first conceived. Five landmark breakthroughs have occurred over the developmental history of the AP: i) the first human studies reported in the 1950's; ii) foundation animal studies reported in the 1960's; iii) the first extended use of AP technology combined with fetal pulmonary resuscitation reported in the 1990s; iv) the development of AP systems powered by the fetal heart reported in the 2000's; and v) the adaption of this technology to maintain extremely preterm fetuses (i.e. 500-600 g body weight) reported in the 2010's. Using this framework, the present paper will provide a review of the developmental history of this long-running experimental system and up-to-date assessment of the published field today. With the apparent acceleration of AP technology towards clinical application, there has been an increase in the attention paid to the field, along with some inaccurate commentary regarding its potential application and merits. Additionally, this paper will address several misrepresentations regarding the potential application of AP technology that serve to distract from the significant potential of this approach to greatly improve outcomes for extremely preterm infants born at or close to the present border of viability.

The epidemiology of COVID-19 in ten Southeast Asian countries.

INTRODUCTION: Periodic benchmarking of the epidemiology of COVID-19 in the Association of Southeast Asian Nations (ASEAN) countries is critical for the continuous understanding of the transmission and control of COVID-19 in the region. The incidence, mortality, testing and vaccination rates within the ASEAN region from 1 January 2020 to 15 October 2021 is analysed in this paper. METHODS: COVID-19 data on cases, deaths, testing, and vaccinations were extracted from the Our World in Data (OWID) COVID-19 data repository for all the ten ASEAN countries. Comparative time-trends of the epidemiology of COVID-19 using the incidence rate, cumulative case fatality rate (CFR), delay-adjusted case fatality rate, cumulative mortality rate (MR), test positivity rate (TPR), cumulative testing rate (TR) and vaccination rate was carried out. RESULTS: Over the study period, a total of 12,720,661 cases and 271,475 deaths was reported within the ASEAN region. Trends of daily per capita cases were observed to peak between July and September 2021 for the ASEAN region. The cumulative case fatality rate (CFR) in Brunei, Cambodia, Indonesia, Laos, Malaysia, Myanmar, Philippines, Singapore, Thailand, and Vietnam, was of 0.9% (N=68), 2.2% (N=2,610), 3.5% (N=142,889), 0.1% (N=36), 1.2% (N=27,700), 4.0% (N=18,297), 1.6% (N=40,424), 0.1% (N=215), 1.7% (N=18,123), and 2.6% (N=21,043), respectively. CFR was consistently highest between January-June 2020. The cumulative mortality rate (MR) was 9.5, 13.7, 51.4, 0.2, 80.3, 32.4, 34.5, 1.6, 23.9 and 19.7 per 100,000 population, respectively. The cumulative test positivity rate (TPR) was 8.4%, 16.9%, 4.6%, 7.5%, 11.1%, 12.9%, 0.5%, 11.7%, and 3.6%, with the cumulative testing rate (TR) at 25.0, 90.1, 27.4, 917.7, 75.8, 177.8, 3303.3, 195.2, and 224.9 tests per 1,000 population in Cambodia, Indonesia, Laos, Malaysia, Myanmar, Philippines, Singapore, Thailand, and Vietnam, respectively. The percentage of population that completed vaccinations (VR) was 44.5%, 65.3%, 18.5%, 28.2%, 61.8%, 6.8%, 19.2%, 76.8%, 22.7%, and 10% in Brunei, Cambodia, Indonesia, Laos, Malaysia, Myanmar, Philippines, Singapore, Thailand, and Vietnam, respectively. CONCLUSION: In 2020, most countries in ASEAN had higher case fatality rates but lower mortalities per population when compared to the third quarter of 2021 where higher mortalities per population were observed. Low testing rates have been one of the factors leading to high test positivity rates. Slow initiation of vaccination programs was found to be the key factor leading to high incidence and case fatality rate in most countries in ASEAN. Effective public health measures were able to interrupt the transmission of this novel virus to some extent. Increasing preparedness capacity within the ASEAN region is critical to ensure that any future similar outbreaks can be dealt with collectively.

Battling COVID-19 pandemic waves in six South-East Asian countries: A real-time consensus review.

INTRODUCTION: COVID-19 has caused unprecedented public health concerns, triggering an escalated burden to health systems worldwide. The pandemic has altered people's living norms, yet coherently escalating countries' socioeconomic instability. This real-time consensus review aims to describe the epidemiological trends of COVID-19 pandemic across six South-East Asian nations, and countryspecific experiences on pandemic preparedness, responses and interventions. METHODS: Consensus-driven approach between authors from the six selected countries was applied. Country specific policy documents, official government media statements, mainstream news portals, global statistics databases and latest published literature available between January-October 2020 were utilised for information retrieval. Situational and epidemiological trend analyses were conducted. Country-specific interventions and challenges were described. Based on evidence appraised, a descriptive framework was considered through a consensus. The authors subsequently outlined the lessons learned, challenges ahead and interventions that needs to be in place to control the pandemic. RESULTS: The total number of people infected with COVID-19 between 1 January and 16 November 2020 had reached 48,520 in Malaysia, 58,124 in Singapore, 3,875 in Thailand, 470,648 in Indonesia, 409,574 in Philippines and 70,161 in Myanmar. The total number of people infected with COVID- 19 in the six countries from January to 31 October 2020 were 936,866 cases and the mortality rate was 2.42%. Indonesia had 410,088 cases with a mortality rate of 3.38%, Philippines had 380,729 cases with a mortality rate of 1.90%, Myanmar had 52,706 cases with a mortality rate of 2.34%, Thailand had 3,780 cases with a mortality rate of 1.56%, Malaysia had 31,548 cases with a mortality rate of 0.79%, and Singapore had 58,015 cases with a mortality rate of 0.05% over the 10- month period. Each country response varied depending on its real-time situations based on the number of active cases and economic situation of the country. CONCLUSION: The number of COVID-19 cases in these countries waxed and waned over the 10-month period, the number of cases may be coming down in one country, and vice versa in another. Each country, if acting alone, will not be able to control this pandemic. Sharing of information and resources across nations is the key to successful control of the pandemic. There is a need to reflect on how the pandemic affects individuals, families and the community as a whole. There are many people who cannot afford to be isolated from their families and daily wage workers who cannot afford to miss work. Are we as a medical community, only empathising with our patients or are we doing our utmost to uphold them during this time of crisis? Are there any other avenues which can curb the epidemic while reducing its impact on the health and socio-economic condition of the individual, community and the nation?

First-trimester pre-eclampsia biomarker profiles in Asian population: multicenter cohort study.

OBJECTIVES: To (i) evaluate the applicability of the European-derived biomarker multiples of the median (MoM) formulae for risk assessment of preterm pre-eclampsia (PE) in seven Asian populations, spanning the east, southeast and south regions of the continent, (ii) perform quality-assurance (QA) assessment of the biomarker measurements and (iii) establish criteria for prospective ongoing QA assessment of biomarker measurements. METHODS: This was a prospective, non-intervention, multicenter study in 4023 singleton pregnancies, at 11 to 13 + 6 weeks' gestation, in 11 recruiting centers in China, Hong Kong, India, Japan, Singapore, Taiwan and Thailand. Women were screened for preterm PE between December 2016 and June 2018 and gave written informed consent to participate in the study. Maternal and pregnancy characteristics were recorded and mean arterial pressure (MAP), mean uterine artery pulsatility index (UtA-PI) and maternal serum placental growth factor (PlGF) were measured in accordance with The Fetal Medicine Foundation (FMF) standardized measurement protocols. MAP, UtA-PI and PlGF were transformed into MoMs using the published FMF formulae, derived from a largely Caucasian population in Europe, which adjust for gestational age and covariates that affect directly the biomarker levels. Variations in biomarker MoM values and their dispersion (SD) and cumulative sum tests over time were evaluated in order to identify systematic deviations in biomarker measurements from the expected distributions. RESULTS: In the total screened population, the median (95% CI) MoM values of MAP, UtA-PI and PlGF were 0.961 (0.956-0.965), 1.018 (0.996-1.030) and 0.891 (0.861-0.909), respectively. Women in this largely Asian cohort had approximately 4% and 11% lower MAP and PlGF MoM levels, respectively, compared with those expected from normal median formulae, based on a largely Caucasian population, whilst UtA-PI MoM values were similar. UtA-PI and PlGF MoMs were beyond the 0.4 to 2.5 MoM range (truncation limits) in 16 (0.4%) and 256 (6.4%) pregnancies, respectively. QA assessment tools indicated that women in all centers had consistently lower MAP MoM values than expected, but were within 10% of the expected value. UtA-PI MoM values were within 10% of the expected value at all sites except one. Most PlGF MoM values were systematically 10% lower than the expected value, except for those derived from a South Asian population, which were 37% higher. CONCLUSIONS: Owing to the anthropometric differences in Asian compared with Caucasian women, significant differences in biomarker MoM values for PE screening, particularly MAP and PlGF MoMs, were noted in Asian populations compared with the expected values based on European-derived formulae. If reliable and consistent patient-specific risks for preterm PE are to be reported, adjustment for additional factors or development of Asian-specific formulae for the calculation of biomarker MoMs is required. We have also demonstrated the importance and need for regular quality assessment of biomarker values. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

Diagnostic accuracy of haptoglobin within ovarian cyst fluid as a potential point-of-care test for epithelial ovarian cancer: an observational study.

OBJECTIVE: To investigate haptoglobin within ovarian cyst fluid (OCF) as a diagnostic biomarker for epithelial ovarian cancer (EOC) and develop an in vitro diagnostic point-of-care device test (IVDPCT) for use in the operating theatre. DESIGN: Retrospective and prospective cohort study. SETTING: South-East Asia. POPULATION: Women with suspicious ovarian cysts. METHODS: Proteomic, immunohistochemical and ELISA methods measured haptoglobin in OCF to differentiate benign and EOCs. Diagnostic performance of haptoglobin was compared with CA125, risk malignancy indices (RMI) and frozen section. Blinded validation of the IVDPCT was performed. MAIN OUTCOME MEASURES: Prediction of malignancy. RESULTS: Haptoglobin concentration measured by ELISA was 0.70 ± 0.09 mg/ml in patients with benign cysts (n = 87), 6.22 ± 0.53 mg/ml in early stage-EOC (n = 17), and 6.57 ± 0.65 mg/ml in late stage-EOC (n = 20). Haptoglobin in EOCs was significantly higher than in benign cysts (P < 0.0001). Haptoglobin using rapid colorimetric assay (RCA) on a training set had a sensitivity of 97.3% and a specificity 92.0%, comparable to ELISA and frozen sections. The haptoglobin AUROC curve was 0.999 (95% CI 0.997-1.000) compared with 0.895 (95% CI 0.814-0.977, P < 0.05) for CA125. Haptoglobin performed significantly better than all the RMIs (P < 0.01). Blinded validation studies showed a minor drop in average diagnostic performance (sensitivity 85.2% and specificity 90.5%) compared with the training set. However, when compared with frozen section, haptoglobin was no worse in diagnostic accuracy for malignancy. CONCLUSION: Haptoglobin was identified as a biomarker for the detection of EOC with potential as a point-of-care diagnostic tool. TWEETABLE ABSTRACT: Haptoglobin within ovarian cyst fluid: a biomarker for epithelial ovarian cancer and point-of-care diagnostics.

Non-invasive prenatal testing for fetal inheritance of maternal ß-thalassaemia mutations using targeted sequencing and relative mutation dosage: a feasibility study.

OBJECTIVE: To evaluate whether targeted sequencing and relative mutation dosage can be used to diagnose correctly inheritance of maternal ß-thalassaemia mutations in cell-free DNA. DESIGN: Feasibility study using samples collected in a prenatal clinic. SETTING: South East Asia. POPULATION: Couples where both partners were known to be carriers of one of four common ß-thalassaemia mutations or an HbE mutation, and therefore at risk of carrying a fetus affected with ß-thalassaemia. METHODS: 49 samples previously identified as having inherited a paternal ß-thalassaemia mutation were amplified using nested polymerase chain reaction (PCR), and then sequencing. Relative mutation dosage was used to classify the fetus as having inherited the wild-type or mutant maternal allele. MAIN OUTCOME MEASURES: Classification of the fetus as 'unaffected' (if the maternal wild-type allele was inherited) or 'affected' with ß-thalassaemia (if the maternal mutant allele was inherited). RESULTS: A classification for inheritance of maternal allele was obtained in 48/49 samples (98.0%). A concordant call was made in 44/48 cases (91.7%): one false-positive and three false-negatives were obtained. Thus, we had an overall sensitivity of 87.5% [95% confidence interval (CI) 67.6-97.3%] and a specificity of 95.8% (95% CI 78.9-99.9%) for inheritance of maternal genotype. CONCLUSIONS: RMD for detection of inheritance of maternal ß-thalassaemia mutations has potential for clinical use. Our sequential approach could be applied to other single-gene disorders. TWEETABLE ABSTRACT: NIPT for ß-thalassaemia achieved using nested-PCR followed by relative mutation dosage.

Preterm prelabour rupture of membranes: a retrospective cohort study of association with adverse outcome in subsequent pregnancy.

OBJECTIVE: To assess the association of first pregnancy preterm prelabour rupture of membranes (PPROM) with adverse maternal and perinatal outcomes in the next pregnancy. DESIGN: Retrospective cohort study. SETTING: Grampian, Scotland, UK. POPULATION: Women with first deliveries recorded in the Aberdeen Maternity Neonatal Databank, 1986-2005. METHODS: Women identified from the AMND database (n = 37 776) were classified into exposed (PPROM in first pregnancy; n = 1979) and unexposed (no PPROM in first pregnancy; n = 35 797) cohorts. Each cohort (exposed n = 1174; unexposed n = 20 860) was followed up until December 2012 for next singleton pregnancy. MAIN OUTCOME MEASURES: Second pregnancy, miscarriage, pregnancy-induced hypertension (PIH), pre-eclampsia (PE), antepartum haemorrhage (APH) and postpartum haemorrhage, repeat PPROM, type of labour, mode of delivery, preterm delivery, low birth weight (LBW), admission to neonatal unit, neonatal infections and death. RESULTS: PPROM in the first singleton pregnancy was associated with an equal likelihood of second pregnancy but with a significantly increased risk of adverse outcomes in the next singleton pregnancy: PPROM [odds ratio (OR) (95% confidence interval (CI)): 6.6 (5.4-7.9)], PE [2.4 (1.7-3.5)], instrumental [2.2 (1.7-2.8)] and caesarean delivery [1.8 (1.5-2.3)], PIH [1.5 (1.2-1.9)] and APH [1.3 (1.1-1.6)] in the mother, and neonatal infection [5.4 (1.4-20.3)], death [2.6 (1.0-6.7)], admission to neonatal unit [2.4 (2.0-2.9)], preterm delivery [2.3 (1.8-2.9)] and LBW [1.44 (1.1-1.9)]. Even in women without a recurrent PPROM, there was still a significant increase in PIH [1.4 (1.1-1.8)], PE [2.3 (1.6-3.5)], instrumental [2.2 (1.7-2.9)] and caesarean delivery [1.9 (1.5-2.4)], and neonatal unit admission [1.6 (1.3-2.0)]. CONCLUSIONS: PPROM in the first pregnancy is associated with significant adverse maternal and perinatal outcomes in the next pregnancy, but not reduced likelihood of second pregnancy. TWEETABLE ABSTRACT: PPROM in the first singleton pregnancy increases risk of adverse maternal and perinatal outcomes in the next singleton pregnancy.

FZD7 drives in vitro aggressiveness in Stem-A subtype of ovarian cancer via regulation of non-canonical Wnt/PCP pathway.

Ovarian cancer (OC) can be classified into five biologically distinct molecular subgroups: epithelial-A (Epi-A), Epi-B, mesenchymal (Mes), Stem-A and Stem-B. Among them, Stem-A expresses genes relating to stemness and is correlated with poor clinical prognosis. In this study, we show that frizzled family receptor 7 (FZD7), a receptor for Wnt signalling, is overexpressed in the Stem-A subgroup. To elucidate the functional roles of FZD7, we used an RNA interference gene knockdown approach in three Stem-A cell lines: CH1, PA1 and OV-17R. Si-FZD7 OC cells showed reduced cell proliferation with an increase in the G0/G1 sub-population, with no effect on apoptosis. The cells also displayed a distinctive morphologic change by colony compaction to become more epithelial-like and polarised with smaller internuclear distances and increased z-axis height. Immunofluorescence (IF) staining patterns of pan-cadherin and ß-catenin suggested an increase in cadherin-based cell-cell adhesion in si-FZD7 cells. We also observed a significant rearrangement in the actin cytoskeleton and an increase in tensile contractility in si-FZD7 OC cells, as evident by the loss of stress fibres and the redistribution of phospho-myosin light chain (pMLC) from the sites of cell-cell contacts to the periphery of cell colonies. Furthermore, there was reciprocal regulation of RhoA (Ras homolog family member A) and Rac1 (Ras-related C3 botulinum toxin substrate 1 (Rho family, small GTP-binding protein Rac1)) activities upon FZD7 knockdown, with a significant reduction in RhoA activity and a concomitant upregulation in Rac1 activity. These changes in pMLC and RhoA, as well as the increased TopFlash reporter activities in si-FZD7 cells, suggested involvement of the non-canonical Wnt/planar cell polarity (PCP) pathway. Selected PCP pathway genes (cadherin EGF LAG seven-pass G-type receptor 3 (CELSR3), prickle homolog 4 (Drosophila) (PRICKLE4), dishevelled-associated activator of morphogenesis 1 (DAAM1), profilin 2 (PFN2), protocadherin 9 (PCDH9), protocadherin α1 (PCDHA1), protocadherin ß17 pseudogene (PCDHB17), protocadherin ß3 (PCDHB3), sprouty homolog 1 (SPRY1) and protein tyrosine kinase 7 (PTK7)) were found to be more highly expressed in Stem-A than non Stem-A subgroup of OC. Taken together, our results suggest that FZD7 might drive aggressiveness in Stem-A OC by regulating cell proliferation, cell cycle progression, maintenance of the Mes phenotype and cell migration via casein kinase 1ɛ-mediated non-canonical Wnt/PCP pathway.

An EMT spectrum defines an anoikis-resistant and spheroidogenic intermediate mesenchymal state that is sensitive to e-cadherin restoration by a src-kinase inhibitor, saracatinib (AZD0530).

The phenotypic transformation of well-differentiated epithelial carcinoma into a mesenchymal-like state provides cancer cells with the ability to disseminate locally and to metastasise. Different degrees of epithelial-mesenchymal transition (EMT) have been found to occur in carcinomas from breast, colon and ovarian carcinoma (OC), among others. Numerous studies have focused on bona fide epithelial and mesenchymal states but rarely on intermediate states. In this study, we describe a model system for appraising the spectrum of EMT using 43 well-characterised OC cell lines. Phenotypic EMT characterisation reveals four subgroups: Epithelial, Intermediate E, Intermediate M and Mesenchymal, which represent different epithelial-mesenchymal compositions along the EMT spectrum. In cell-based EMT-related functional studies, OC cells harbouring an Intermediate M phenotype are characterised by high N-cadherin and ZEB1 expression and low E-cadherin and ERBB3/HER3 expression and are more anoikis-resistant and spheroidogenic. A specific Src-kinase inhibitor, Saracatinib (AZD0530), restores E-cadherin expression in Intermediate M cells in in vitro and in vivo models and abrogates spheroidogenesis. We show how a 33-gene EMT Signature can sub-classify an OC cohort into four EMT States correlating with progression-free survival (PFS). We conclude that the characterisation of intermediate EMT states provides a new approach to better define EMT. The concept of the EMT Spectrum allows the utilisation of EMT genes as predictive markers and the design and application of therapeutic targets for reversing EMT in a selective subgroup of patients.

Lipidomic analysis of human placental syncytiotrophoblast microvesicles in adverse pregnancy outcomes.

PROBLEM: Syncytiotrophoblast microvesicles (STBM) are shed from placenta into the maternal circulation. STBM circulate in increased amounts in adverse pregnancies, e.g., preeclampsia and recurrent miscarriages (RM). Recently dysregulation of lipid metabolites has been proposed to be associated with their pathogenesis. Lipid composition of STBM in healthy and adverse pregnancies remains unknown. OBJECTIVE: To determine lipid composition of STBM and whether STBM lipid composition differs in pathologic and normal pregnancies. STUDY DESIGN: Patients with Preeclampsia (n = 6) or history of RM (n = 9) (>2 consecutive losses <20 weeks) and gestational age-matched normal pregnant controls (same number as cases) were recruited. STBM were prepared from placental explant culture supernatant. Lipid profiling of STBM was performed by mass spectrometry in combination with liquid chromatography. We quantified ∼200 lipids in STBM including (i) glycerophospholipids (phosphatidylcholine, PC; phosphatidylethanolamine, PE; phosphatidylinositol, PI; phosphatidylglycerol, PG; phosphatidylserine, PS; phosphatidic acid, PA); (ii) sphingolipids (sphingomyelin, SM; ceramide, Cer; Glucosylceramide, GluCer; ganglioside mannoside 3, GM3); (iii) free cholesterol and cholesteryl esters, CE. RESULTS: The major lipid classes in STBM were SM, Chol, PS, PC and PI, along with PA and GM3 enrichments. SM/PC ratio showed a unique reversal (3:1) compared to that normally found in human cells or plasma. Level of total PS was significantly upregulated (p < 0.005) in preeclampsia patients, while PI (p < 0.0005), PA (p < 0.005), and GM3 (p < 0.05) were significantly downregulated. Similar trends were obtained in RM. CONCLUSIONS: Differential lipid expression of STBM in preeclampsia or RM includes those that are implicated in immune response, coagulation, oxidative stress, and apoptosis.

Systemic delivery of scAAV9 in fetal macaques facilitates neuronal transduction of the central and peripheral nervous systems.

Correction of perinatally lethal neurogenetic diseases requires efficient transduction of several cell types within the relatively inaccessible CNS. Intravenous AAV9 delivery in mouse has achieved development stage-specific transduction of neuronal cell types, with superior neuron-targeting efficiency demonstrated in prenatal compared with postnatal recipients. Because of the clinical relevance of the non-human primate (NHP) model, we investigated the ability of AAV9 to transduce the NHP CNS following intrauterine gene therapy (IUGT). We injected two macaque fetuses at 0.9 G with 1 × 10(13) vg scAAV9-CMV-eGFP through the intrahepatic continuation of the umbilical vein. Robust green fluorescent protein (GFP) expression was observed for up to 14 weeks in the majority of neurons (including nestin-positive cells), motor neurons and oligodendrocytes throughout the CNS, with a significantly lower rate of transduction in astrocytes. Photoreceptors and neuronal cell bodies in the plexiform and ganglionic retinal layers were also transduced. In the peripheral nervous system (PNS), widespread transduction of neurons was observed. Tissues harvested at 14 weeks showed substantially lower vector copy number and GFP levels, although the percentage of GFP-expressing cells remained stable. Thus, AAV9-IUGT in late gestation efficiently transduces both the CNS and PNS with neuronal predilection, of translational relevance to hereditary disorders characterized by perinatal onset of neuropathology.

The association with age, human tissue kallikreins 6 and 10 and hemostatic markers for survival outcome from epithelial ovarian cancer.

PURPOSE: To assess known cancer biomarkers CA-125, human tissue kallikreins KLK6 and KLK10, hemostatic markers and age with 5-year survival outcome from epithelial ovarian cancer. METHODS: Forty-one benign cyst cohorts and 83 patients diagnosed with ovarian cancer were recruited. The following assays were performed: fibrinogen, vWF antigen, D: -dimer, ATIII activity, tPA, PAI-1, uPAR, KLK6, KLK10 and CA-125. Follow-up visits of cancer patients of more than 60 months were noted. Data between those who survived past 60 months and mortality from cancer were analyzed. RESULTS: Only 24 patients lived past 60 months, and 31 died (advanced stage n = 27). Those living past 60 months were significantly older and associated with similar pre-operative levels seen in benign cyst cohorts especially for KLK6, fibrinogen, vWF, AT levels despite upregulation of D: -dimer, CA-125 and KLK10. Ovarian cancer cohorts living past 60 months were younger than those who died within 12 months (n = 12). Mortality within 12 months was associated with older age, upregulation of KLK6, fibrinogen, D: -dimer, vWF, tPA antigen and reduced ATIII levels. Similarly, mortality within 36 months of disease showed older age with upregulation of CA-125, KLK6 D: -dimer vWF antigen and tPA antigen levels. Late stage cancer (III/IV) showed upregulated CA-125, KLK6, KLK10, D: -dimer and reduced AT compared to early stage cancer (I/II). The 5-year survival rate for early cancer was 80%, advanced 22.9% and overall 5-year survival rate was 43.6%. CONCLUSION: Older age together with the novel biomarkers studied and their association with adverse outcome from epithelial ovarian cancer was seen especially within 12 and 36 months of disease. Those who lived past 60 months of disease showed similar pre-operative levels seen in benign cyst cohorts despite elevated D: -dimer, CA125 and KLK 10. An enlarged study is needed to confirm these findings.

Development of quantitative-fluorescence polymerase chain reaction for the rapid prenatal diagnosis of common chromosomal aneuploidies in 1,000 samples in Singapore.

INTRODUCTION: We aimed to develop a rapid quantitative-fluorescence polymerase chain reaction (QF-PCR) to detect common foetal aneuploidies in the Singapore population within 48 hours of sample collection in order to alleviate parental anxiety. METHODS: DNA from 1,000 foetal samples (978 amniotic fluids, 14 chorion villi and eight foetal blood samples) was analysed using a QF-PCR of 19 microsatellite markers located on chromosomes 13, 18, 21, X and Y. A total of 523 samples were archived before the QF-PCR analysis (archived), while QF-PCR was performed and the results obtained within 48 hours of sample collection in the remaining 477 samples (live). The results were confirmed with their respective karyotypes. RESULTS: In total, 47 autosomal trisomies (T) were found: 30 among the archived (three T13, 12 T18, 15 T21) and 17 among the live (four T18, 13 T21) samples. The QF-PCR results were verified with their respective karyotypes. We achieved 100 percent sensitivity (lower 95 percent confidence interval [CI], 92.8 percent) and specificity (lower 95 percent CI, 99.5 percent), and the time taken from sample collection to the obtaining of results for the 477 live samples was less than 48 hours. CONCLUSION: Prenatal diagnostic results of common chromosomal abnormalities can be released within 48 hours of sample collection using QF-PCR. Parental anxiety is alleviated and clinical management is enhanced with this short waiting time.

In vivo model to determine fetal-cell enrichment efficiency of novel noninvasive prenatal diagnosis methods.

OBJECTIVE: To develop an in vivo model to determine fetal-cell enrichment efficiency of novel noninvasive prenatal diagnosis methods. METHODS: Efficiency of our three-step enrichment protocol was determined in vitro before fetal nucleated red blood cells (FNRBCs) were enriched from first-trimester maternal blood samples collected from the same patients pre- and postsurgical termination of pregnancy (TOP) (n = 10). FNRBCs enriched were identified using embryonic epsilon-globin immunocytochemistry and chromosomal fluorescence in situ hybridization. RESULTS: We recovered 37% of spiked FNRBCs (95% confidence interval (CI) 28.5-45.6; n = 8) in in vitro experiments. We show a consistent threefold increase in the number of epsilon + FNRBCs in maternal blood obtained immediately post-TOP (p = 0.005). A mathematical relationship was derived: observed number of pretermination primitive FNRBCs = 0.6 + 0.31 (coefficient between pretermination/post-termination primitive FNRBCs, 95% CI 0.12-0.49; p = 0.005) x observed number of post-termination primitive FNRBCs (R2 = 0.65). CONCLUSION: Our data demonstrate that maternal blood obtained immediately post-TOP would be a good in vivo model to determine the enrichment efficiency of novel protocols and methods for noninvasive prenatal diagnosis.

Refining quantitative fluorescent polymerase chain reaction for prenatal detection of X chromosomal anomalies in the major Southeast Asian populations.

INTRODUCTION: This study aimed to refine the current quantitative fluorescent polymerase chain reaction (QF-PCR) screen to detect X chromosome anomalies for prenatal diagnosis in the major Southeast-Asian populations. METHODS: 100 amniotic fluid samples from Chinese, Malay and Indian origins were subjected to QF-PCR using the X chromosome markers, HPRT, X22 and AMXY, along with the autosomal marker D21S1411. RESULTS: Out of the 100 samples tested by markers X22 and HPRT, eight samples were homozygous for both markers, of which seven were resolved by comparison with the autosomal marker D21S1411. CONCLUSION: 99 percent of samples could be tested for X chromosome copy numbers, increasing the stringency for detection of X chromosome anomalies by QF-PCR. All results were confirmed by cytogenetics.

FastFISH: technique for ultrarapid fluorescence in situ hybridization on uncultured amniocytes yielding results within 2 h of amniocentesis.

Rapid aneuploidy detection methods allow prenatal diagnosis results to be released within 48 h, but not on the same day as the invasive test. We aimed to develop a rapid fluorescence in situ hybridization (FISH) method (FastFISH) that releases accurate results on the same day as amniocentesis. FastFISH was optimized to be completed within 2 h of sample collection using CEP and LSI probes for chromosomes 13, 18, 21, X, Y and DiGeorge syndrome (DGS). The technique was tested on 100 consecutive amniotic fluid samples in a blinded study. It was also validated as a 1-day molecular genetic test on three representative fetal tissue samples: chorionic villus, amniotic fluid and fetal blood. In the blinded study, FastFISH results were ready within 2 h of sample collection. Of the 100 amniotic fluid samples, 49 male and 50 female fetuses were identified. One fetus was 47, XXY (Klinefelter syndrome). Three fetuses had trisomy 21. One fetus suspected of DGS by ultrasound was identified as normal. Results of FastFISH analyses in all 100 cases were concordant with their karyotypes (100% accuracy; lower 95% CI, 97.05%). In the 1-day test validation, all results were released on the same day and were concordant with their respective karyotypes. FastFISH allows results to be released on the same day as amniocentesis. It represents the necessary development for a 1-day prenatal diagnosis service.

The association between fibrinogen, von Willebrand Factor, antithrombin III, and D-dimer levels and survival outcome by 36 months from ovarian cancer.

Thirty-five patients diagnosed to have ovarian cancer (early FIGO stage I/II n = 11, advanced FIGO stage III/IV, n = 24) were evaluated for hemostatic parameters relating to survival outcome by 36 months of disease. Systemic plasminogen activators and inhibitors were evaluated and we found no significant association with survival outcome and eventually only fibrinogen, von Willebrand Factor (vWF), antithrombin III (ATIII), and D-dimer levels were determined for their association with disease outcome by 12 months, 24 months, and 36 months. Twenty-four patients succumbed to the disease by 36 months (early n = 2, advanced n = 22). The 11 surviving patients (advanced n = 3, including one deceased at 52 months) is still living past 36 months and 82 months at the time of analysis. Elevated fibrinogen, vWF, and D-dimer together with reduced ATIII levels were found to be associated with poor survival outcome by 12 months of disease. Moreover, elevated vWF and D-dimer with reduced ATIII levels was strongly implicated with poor survival outcome by 36 months from ovarian cancer. The overall survival rate at 36 months from ovarian cancer was 31.4%. It is therefore suggested that fibrinogen, vWF, ATIII, and D-dimer levels be used together as prognostic markers for disease outcome especially in patients with advanced ovarian cancer within 36 months of disease. An expanded study is required to confirm these findings.