Lisa Leth Maroun

Quality assessment of a placental perfusion protocol

Validation of in vitro test systems using the modular approach with steps addressing reliability and relevance is an important aim when developing in vitro tests in e.g. reproductive toxicology. The ex vivo human placental perfusion system may be used for such validation, here presenting the placental perfusion model in Copenhagen including control substances. The positive control substance antipyrine shows no difference in transport regardless of perfusion media used or of terms of delivery (n=59, p<0.05). Negative control studies with FITC marked dextran correspond with leakage criteria (<3 ml h(-1) from the fetal reservoir) when adding 2 (n=7) and 20mg (n=9) FITC-dextran/100 ml fetal perfusion media. Success rate of the Copenhagen placental perfusions is provided in this study, including considerations and quality control parameters. Three checkpoints suggested to determine success rate revealed that 15% of the cannulated placentae received in one year (n=202) were successfully perfused.

Kinetics of silica nanoparticles in the human placenta

Abstract The potential medical applications of nanoparticles (NPs) warrant their investigation in terms of biodistribution and safety during pregnancy. The transport of silica NPs across the placenta was investigated using two models of maternal–foetal transfer in human placenta, namely, the BeWo b30 choriocarcinoma cell line and the ex vivo perfused human placenta. Nanotoxicity in BeWo cells was examined by the MTT assay which demonstrated decreased cell viability at concentrations >100 µg/mL. In the placental perfusion experiments, antipyrine crossed the placenta rapidly, with a foetal:maternal ratio of 0.97 ± 0.10 after 2 h. In contrast, the percentage of silica NPs reaching the foetal perfusate after 6 h was limited to 4.2 ± 4.9% and 4.6 ± 2.4% for 25 and 50 nm NPs, respectively. The transport of silica NPs across the BeWo cells was also limited, with an apparent permeability of only 1.54 × 10−6 ± 1.56 × 10−6 cm/s. Using confocal microscopy, there was visual confirmation of particle accumulation in both BeWo cells and in perfused placental tissue. Despite the low transfer of silica NPs to the foetal compartment, questions regarding biocompatibility could limit the application of unmodified silica NPs in biomedical imaging or therapy.

Neural tube defects and associated anomalies in a fetal and perinatal autopsy series

Neural tube defects (NTDs) are congenital malformations of the central nervous system (CNS) secondary to abnormal closure of the neural tube during embryonic development. This study provides information on NTD rate, distribution, associated morphologic anomalies and organ weights in a Danish fetal and perinatal autopsy series during a 16 year period. The data were extracted from the autopsy reports of a consecutive series of 1984 fetal and perinatal autopsies from the Copenhagen area performed in the period 1989–2004. Registered parameters included: The location and morphology of the NTD, associated morphological anomalies, and organ weights. Organ weights were evaluated according to new fetal autopsy standards and grouped as low, normal or high. Ninety‐seven NTD cases were found (4.9%): Spina bifida (38 cases), cephalocele (17 cases) and anencephaly (42 cases). 63% of NTD cases had associated morphologic anomalies. Among the most common were hydrocephalus, NTD in another region, and anomalies in the urogenital system. 58% of the NTD cases had abnormal weight of one or more organs. Most notable was low adrenal weight not only in anencephalic fetuses but also in cases with cephalocele, suggesting a possible association.

Prenatal detection of congenital heart disease in a low risk population undergoing first and second trimester screening

The prenatal detection rate of congenital heart disease (CHD) is low compared with other fetal malformations. Our aim was to evaluate the prenatal detection of CHD in Eastern Denmark.

Fetal Cor Triatriatum Dexter: A Report of Two Cases Associated with Nuchal Edema in Early Second Trimester

Two early-2nd-trimester fetuses were aborted as a result of nuchal edema and suspected severe heart malformation. At autopsy one fetus demonstrated nuchal edema, mild hydronephrosis, and cor triatriatum dexter associated with ventricular septal defect and tubular hypoplasia of the aortic arch. The other fetus demonstrated severe nuchal edema, and cor triatriatum dexter was the only malformation. Cor triatriatum dexter is a rare cardiac malformation characterized by division of the right atrium into 2 compartments by a usually fenestrated membrane representing remnants of the right valve of the embryonic sinus venosus. This malformation has been diagnosed in adults and children by echocardiography, surgery, or autopsy but has not previously been published in fetuses.

Correlation between prenatal diagnosis by ultrasound and fetal autopsy findings in second‐trimester abortions

Abstract  We evaluated the correlation between prenatal diagnosis by ultrasound and autopsy findings, based on 52 second‐trimester pregnancies terminated due to fetal malformations or chromosome aberrations diagnosed at a gestational age of 12–25 weeks. In 24 pregnancies, there was full agreement between ultrasound and autopsy. In 23 fetuses, the main diagnosis was confirmed and additional or more specific findings were observed on autopsy. In five fetuses, there were considerable differences. Discrepancies between ultrasound and autopsy findings were mainly anomalies undetectable by ultrasound and thus expected; however, about one‐third of the discrepancies were not expected, representing findings that were ‘missed’ at ultrasound. The main ultrasound diagnoses were confirmed in the majority of the pregnancies, but the additional information obtained at autopsy in more than half of the fetuses clearly shows the value and benefit of postmortem fetal examination following termination of a pregnancy.

The outcome of antenatal ultrasound diagnosed anomalies of the kidney and urinary tract in a large Danish birth cohort

Objective Antenatal ultrasound diagnosed anomalies of the kidney and urinary tract (AUDAKUT) are reported in 0.3%–5% on prenatal ultrasound (US) and 0.3%–4.5% on postnatal US. The anterior-posterior diameter of the renal pelvis (APD) is an essential measurement. Series with low threshold values of APD prenatally and postnatally will include healthy infants. It is important to avoid follow-up of such infants. Interventions In 2006, new Danish guidelines for AUDAKUT were introduced. Aim of study Investigations of incidences and type of AUDAKUT based on Danish guidelines, including long-term follow-up. Design Cohort study. Setting Copenhagen University Hospital Hvidovre and Copenhagen University Hospital Rigshospitalet, Denmark. Patients Consecutive cases with AUDAKUT in the second and third trimesters, which were either terminated before 22 completed weeks of gestation or born in the 8-year period January 2006–December 2013. Patients were followed until June 2014. Results 50 193 live born children and 24 terminated fetuses (0.05%) were included. The prevalence of AUDAKUT was only 0.39% prenatally, 0.29% at first postnatal US and 0.22% at the end of follow-up, including terminated cases. The greater the prenatal and postnatal APD, the higher risk of febrile urinary tract infection (fUTI) and surgical intervention, and lower probability of resolution. 25% of the identified patients had fUTI and/or surgery. Conclusions We recommend threshold values of APD at least 10 mm in the third trimester and in general at least 12 mm at first postnatal US for intensive follow-up. In this largest to date unselected birth cohort of AUDAKUT, the incidences of clinically significant AUDAKUT were in the lowest range of those previously published.

Rare novel variants in the ZIC3 gene cause X-linked heterotaxy

Variants in the ZIC3 gene are rare, but have demonstrated their profound clinical significance in X-linked heterotaxy, affecting in particular male patients with abnormal arrangement of thoracic and visceral organs. Several reports have shown relevance of ZIC3 gene variants in both familial and sporadic cases and with a predominance of mutations detected in zinc-finger domains. No studies so far have assessed the functional consequences of ZIC3 variants in an in vivo model organism. A study population of 348 patients collected over more than 10 years with a large variety of congenital heart disease including heterotaxy was screened for variants in the ZIC3 gene. Functional effects of three variants were assessed both in vitro and in vivo in the zebrafish. We identified six novel pathogenic variants (1,7%), all in either male patients with heterotaxy (n=5) or a female patient with multiple male deaths due to heterotaxy in the family (n=1). All variants were located within the zinc-finger domains or leading to a truncation before these domains. Truncating variants showed abnormal trafficking of mutated ZIC3 proteins, whereas the missense variant showed normal trafficking. Overexpression of wild-type and mutated ZIC protein in zebrafish showed full non-functionality of the two frame-shift variants and partial activity of the missense variant compared with wild-type, further underscoring the pathogenic character of these variants. Concluding, we greatly expanded the number of causative variants in ZIC3 and delineated the functional effects of three variants using in vitro and in vivo model systems.

Pathologic Evaluation of Normal and Perfused Term Placental Tissue

This study reports for the 1st time the incidence and interobserver variation of morphologic findings in a series of 34 term placentas from pregnancies with normal outcome used for perfusion studies. Histologic evaluation of placental tissue is challenging, especially when it comes to defining “normal tissue” versus “pathologic lesions.” A scoring system for registration of abnormal morphologic findings was developed. Light microscopic examination was performed independently by 2 pathologists, and interobserver variation was analyzed. Findings in normal and perfused tissue were compared and selected findings were tested against success parameters from the perfusions. Finally, the criteria for frequent lesions with fair to poor interobserver variation in the nonperfused tissue were revised and reanalyzed. In the perfused tissue, the perfusion artefact “trophoblastic vacuolization,” which is believed to represent dilated transtrophoblastic channels, was reproducible and significantly correlated to the perfusion marker “fetal leakage.” In longer perfusions, microscopy of the perfused cotyledon revealed bacteria in the fetal vessels. This finding led to an adjustment in the perfusion protocol with addition of antibiotics to the medium. In the “normal” tissue, certain lesions were very frequent and showed only fair or poor interobserver agreement. Revised minimum criteria for these lesions were defined and found reproducible. This study has emphasized the value of pathologic examination as a supplement in placental perfusion models. Examination of the perfused cotyledon for trophoblastic vacuolization is recommended as an additional quality marker in perfusion models. The study also underlines the need for exact definitions of abnormality in frequent placental lesions.

Ghrelin in the fetal pancreas - a digital quantitation study.

Hasselby JP, Maroun LL, Federspiel BH, Vainer B. Ghrelin in the fetal pancreas – a digital quantitation study. APMIS 2011.