Anna L. David

Umbilical vein testosterone in female infants born to mothers with polycystic ovary syndrome is elevated to male levels

The aetiology of polycystic ovary syndrome (PCOS) is poorly understood, but an intrauterine hyperandrogenic environment has been implicated. This study was designed to assess whether the female offspring of mothers with PCOS are exposed to raised levels of testosterone (T) in utero. In this case–control study, three groups of pregnant women were recruited from the labour ward: PCOS women with a female baby (n = 10, PCOS girls); control women with a female baby (n = 20, control girls) and control women with a male baby (n = 10, control boys). Maternal and umbilical vein (UV) blood was assayed for T levels. UV T in PCOS girls was significantly raised, compared with control girls (p < 0.012). The difference in UV T between PCOS girls and control boys was not significant (p < 0.254). This is the first demonstration of a hyperandrogenic in utero environment in PCOS pregnancies; UV T in female infants is raised to male levels.

Clinical applications of prenatal and postnatal therapy using stem cells retrieved from amniotic fluid

Purpose of review To review the potential of stem cells derived from amniotic fluid and applications in prenatal and postnatal therapy. Recent findings We have recently described that pluripotent stem cells can be isolated from amniotic fluid defined as amniotic fluid stem (AFS) cells by selection for expression of the membrane stem cell factor receptor c-Kit. AFS cells maintained for over 250 population doublings retained long telomeres and normal karyotype. Clonal human lines verified by retroviral marking were induced to differentiate into cell types representing each embryonic germ layer, including adipogenic, osteogenic, myogenic, endothelial, neuronal, and hepatic lineages. Rat AFS cells have been able to improve the repair of damaged smooth muscle in cryoinjury bladders. Furthermore, AFS cells could be differentiated toward cardiomyogenic lineages, when co-cultured with neonatal cardiomyocytes and have potential to generate hematopoietic lineages both in vitro and in vivo. These cells have been applied into fetal therapy, and widely used for tissue repair in animal models. Finally, we demonstrated a feasible way to do in-utero autologous AFS transplantation in sheep. Summary Stem cells derived from amniotic fluid are a relatively new source of cells that could have a therapeutic value in various diseases prenatally and/or postnatally.

The impact of cross‐border reproductive care or ‘fertility tourism’ on NHS maternity services

High order multiple pregnancies have substantial morbidity and mortality. Fertility treatment is commonly responsible for their conception and is available globally with variable regulation. We investigated cross‐border fertility treatment in these pregnancies in a UK fetal medicine unit, recording mode of conception, country of fertility treatment, reason for non‐UK treatment and fetal reduction. Over an 11‐year period, 109 women had a high order multiple pregnancy. Ninety‐four women (86%) conceived with fertility treatment of whom 24 (26%) had this performed overseas. Cross‐border fertility treatment poses an increasing challenge to obstetricians. National data on its occurrence is urgently needed.

Local delivery of VEGF adenovirus to the uterine artery increases vasorelaxation and uterine blood flow in the pregnant sheep

Impaired materno-placental perfusion causes two important obstetric complications, fetal growth restriction and preeclampsia. This study investigated whether adenoviral vector-mediated overexpression of vascular endothelial growth factor (VEGF) in the uterine arteries (UtAs) increases uterine artery blood flow (UBF). First-generation adenovirus vectors (5 × 1011 particles) containing the VEGF gene (Ad.VEGF-A or -D) or the β-galactosidase reporter gene (Ad.lacZ) were injected into the UtAs of pregnant sheep (n=6) at 88–102 days of gestation (term=145 days). UBF was measured using Doppler sonography before, and 4–7 days after injection. Mean UBF increased significantly from 233±156 (s.d.) ml min−1 to 753±415 ml min−1 following Ad.VEGF-A injection (P=0.005, n=5); Ad.lacZ infection had no significant effect. Organ bath experiments on uterine arterial sections 4–7 days after injection showed that, compared with Ad.lacZ vessels, Ad.VEGF-A-transduced vessels had a reduced contractile response to phenylephrine (Emax 148±10.9 vs Emax 228.2±27.5, P<0.05) but increased relaxation with bradykinin (pD2 (−log EC50) values 9.11±0.01 vs 8.65±0.11, P<0.05). Injection of Ad.VEGF-A into the UtAs increases UBF by enhancing vasodilatation. This may provide the basis for therapy in pregnancies complicated by uteroplacental insufficiency.

Autologous transplantation of amniotic fluid-derived mesenchymal stem cells into sheep fetuses

Long-term engraftment and phenotype correction has been difficult to achieve in humans after in utero stem cell transplantation mainly because of allogeneic rejection. Autologous cells could be obtained during gestation from the amniotic fluid with minimal risk for the fetus and the mother. Using a sheep model, we explored the possibility of using amniotic fluid mesenchymal stem cells (AFMSCs) for autologous in utero stem cell/gene therapy. We collected amniotic fluid (AF) under ultrasound-guided amniocentesis in early gestation pregnant sheep (n = 9, 58 days of gestation, term = 145 days). AFMSCs were isolated and expanded in all sampled fetal sheep. Those cells were transduced using an HIV vector encoding enhanced green fluorescent protein (GFP) with 63.2% (range 38.3–96.2%) transduction efficiency rate. After expansion, transduced AFMSCs were injected into the peritoneal cavity of each donor fetal sheep at 76 days under ultrasound guidance. One ewe miscarried twin fetuses after amniocentesis. Intraperitoneal injection was successful in the remaining 7 fetal sheep giving a 78% survival for the full procedure. Tissues were sampled at postmortem examination 2 weeks later. PCR analysis detected GFP-positive cells in fetal tissues including liver, heart, placenta, membrane, umbilical cord, adrenal gland, and muscle. GFP protein was detected in these tissues by Western blotting and further confirmed by cytofluorimetric and immunofluorescence analyses. This is the first demonstration of autologous stem cell transplantation in the fetus using AFMSCs. Autologous cells derived from AF showed widespread organ migration and could offer an alternative way to ameliorate prenatal congenital disease.

Ultrasound-guided percutaneous delivery of adenoviral vectors encoding the beta-galactosidase and human factor IX genes to early gestation fetal sheep in utero.

In utero gene therapy may provide treatment of genetic diseases before significant organ damage, allow permanent genetic correction by reaching stem cell populations, and provide immune tolerance against the therapeutic transgenes and vectors. We have used percutaneous ultrasound-guided injection as a minimally invasive fetal procedure. First-generation adenoviruses encoding the nuclear localizing beta-galactosidase reporter gene or the human factor IX (hFIX) gene, or colloidal carbon were delivered via the umbilical vein (UV, n = 4), heart (intracardiac [IC], n = 2), liver parenchyma (intrahepatic [HE], n = 11), peritoneal cavity (intraperitoneal [IP], n = 14), skeletal musculature ([intramuscular [IM], n = 11), or the amniotic cavity (intraamniotic [IA], n = 14) to early-gestation fetal sheep (0.3 gestation = day 33-61). Postmortem analysis was performed at 2, 9, or 28 days after injection. Although fetal survival was between 77% and 91% for IP, HE, IA, and IM routes, no fetuses survived UV or IC procedures. The hFIX levels reaching 1900 and 401 ng/ml (IP), 30 ng/ml (HE), 66.5 and 39 ng/ml (IA), and 83 and 65.5 ng/ml (IM), respectively, were determined 2 days after injection and decreased at birth to 16.5 ng/ml (IP), 7 ng/ml (HE), 4.5 ng/ml (IA), and 4 and 0 ng/ml (IM). Polymerase chain reaction (PCR) and immunohistochemistry showed broadest hFIX transgene spread and highest localised beta-galactosidase expression, respectively, after IP administration. Antibodies were observed against vector but not against hFIX.

Fetal intracranial haemorrhages caused by fetal and neonatal alloimmune thrombocytopenia: an observational cohort study of 43 cases from an international multicentre registry

Objective To characterise pregnancies where the fetus or neonate was diagnosed with fetal and neonatal alloimmune thrombocytopenia (FNAIT) and suffered from intracranial haemorrhage (ICH), with special focus on time of bleeding onset. Design Observational cohort study of all recorded cases of ICH caused by FNAIT from the international No IntraCranial Haemorrhage (NOICH) registry during the period 2001–2010. Setting 13 tertiary referral centres from nine countries across the world. Participants 37 mothers and 43 children of FNAIT pregnancies complicated by fetal or neonatal ICH identified from the NOICH registry was included if FNAIT diagnosis and ICH was confirmed. Primary and secondary outcome measures Gestational age at onset of ICH, type of ICH and clinical outcome of ICH were the primary outcome measures. General maternal and neonatal characteristics of pregnancies complicated by fetal/neonatal ICH were secondary outcome measures. Results From a total of 592 FNAIT cases in the registry, 43 confirmed cases of ICH due to FNAIT were included in the study. The majority of bleedings (23/43, 54%) occurred before 28 gestational weeks and often affected the first born child (27/43, 63%). One-third (35%) of the children died within 4 days after delivery. 23 (53%) children survived with severe neurological disabilities and only 5 (12%) were alive and well at time of discharge. Antenatal treatment was not given in most (91%) cases of fetal/neonatal ICH. Conclusions ICH caused by FNAIT often occurs during second trimester and the clinical outcome is poor. In order to prevent ICH caused by FNAIT, at-risk pregnancies must be identified and prevention and/or interventions should start early in the second trimester.

Gastroschisis: sonographic diagnosis, associations, management and outcome

Gastroschisis is a defect in the abdominal wall, typically on the right side of a normally inserted umbilical cord through which bowel and other abdominal contents herniate. Classically, no membrane covers the herniated abdominal contents, which distinguishes the defect from exomphalos, an important differential diagnosis. Gastroschisis is usually diagnosed prenatally using ultrasound examination. The prevalence is increasing worldwide from approximately 0.1 per 10 000 total births in the 1970s to over 5 in the early 2000s. The reasons for this are unknown, but factors such as maternal smoking, recreational drugs and young maternal age are strongly associated with the defect. The increasing prevalence is causing concern because the cost of treating gastroschisis is high. Neonatal morbidity depends on significant complicating factors such as bowel atresia or necrosis and prolonged post‐operative ileus. Foetuses with gastroschisis are more likely to be born premature and with intra‐uterine growth restriction, both of which contribute to the morbidity. Gastroschisis requires early surgery after birth, often followed by prolonged neonatal care. However, advances in surgical and post‐operative care in the last decade have meant that currently 90% of affected neonates survive, with few long‐term problems. Copyright

Association of isolated short femur in the mid‐trimester fetus with perinatal outcome

To evaluate the prevalence of fetal isolated short femur in a cohort of women screened for Down syndrome by the integrated test, and to compare the outcome of fetuses with isolated short femur in the mid‐trimester with that of fetuses with normal femur length (controls).

Somatic activating mutations in Pik3ca cause sporadic venous malformations in mice and humans

Mutant Pik3ca gives rise to venous malformations. PI3K-ing the best treatment Venous malformations are a type of congenital vascular anomalies composed of dilated blood vessels, which can cause a variety of complications such as pain, disfigurement, and bleeding. The available treatments for these malformations are invasive and not particularly effective. Now, Castel et al. and Castillo et al. have both identified mutations in the phosphatidylinositol 3-kinase (PI3K) pathway as a cause of venous malformations, studied these in numerous mouse models, and demonstrated that they can be effectively treated by inhibiting PI3K activity, paving the way for future clinical trials. Venous malformations (VMs) are painful and deforming vascular lesions composed of dilated vascular channels, which are present from birth. Mutations in the TEK gene, encoding the tyrosine kinase receptor TIE2, are found in about half of sporadic (nonfamilial) VMs, and the causes of the remaining cases are unknown. Sclerotherapy, widely accepted as first-line treatment, is not fully efficient, and targeted therapy for this disease remains underexplored. We have generated a mouse model that faithfully mirrors human VM through mosaic expression of Pik3caH1047R, a constitutively active mutant of the p110α isoform of phosphatidylinositol 3-kinase (PI3K), in the embryonic mesoderm. Endothelial expression of Pik3caH1047R resulted in endothelial cell (EC) hyperproliferation, reduction in pericyte coverage of blood vessels, and decreased expression of arteriovenous specification markers. PI3K pathway inhibition with rapamycin normalized EC hyperproliferation and pericyte coverage in postnatal retinas and stimulated VM regression in vivo. In line with the mouse data, we also report the presence of activating PIK3CA mutations in human VMs, mutually exclusive with TEK mutations. Our data demonstrate a causal relationship between activating Pik3ca mutations and the genesis of VMs, provide a genetic model that faithfully mirrors the normal etiology and development of this human disease, and establish the basis for the use of PI3K-targeted therapies in VMs.