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.

Identification of candidate surface antigens for non-invasive prenatal diagnosis by comparative global gene expression on human fetal mesenchymal stem cells.

Transplacental passage of circulating first-trimester fetal mesenchymal stem cells (fMSC) raises the prospect of harvesting fetal cells in maternal blood. Despite high sensitivity in model systems, negative selection and culture strategies yield fMSC only rarely in post-termination maternal blood. The different adhesion molecule profile of fMSC to competitor maternal cell types suggests that improved positive selection strategies may facilitate non-invasive prenatal diagnosis. We aimed to identify surface antigens specific to fMSC and not maternal peripheral blood lymphocytes (PBL), using genome-wide analysis of actively expressed transcripts. Maternal PBL and fMSC cultured from first-trimester blood, liver and bone marrow were assessed for global gene expression by Affymetrix U133Plus2.0 arrays. Data were analysed using Affymetrix GCOS01.2. Transcripts present in all fMSC (n = 9) but absent in all PBL samples (n = 3) were selected for further analysis of cell-surface membrane molecules by RT-PCR and immunocytochemistry. Of 1544 genes expressed in fMSC and not maternal PBL, filtering for cell-surface molecules yielded 159 genes. Of these, 29 had a mean expression ratio of >300 (P < 0.001), which represented 18 unique genes, and their positive expression in all fMSC samples was confirmed by RT-PCR. Candidates for non-invasive prenatal diagnosis were chosen for further analysis by immunocytochemistry. Surface expression of OSMR and COL1 proteins on all fMSC, but no maternal PBL was confirmed. Identification of novel surface antigens on circulating human fMSC and not maternal PBL facilitates positive selection strategies for isolating fMSC for non-invasive prenatal diagnosis.

Systematic analysis of research underfunding in maternal and perinatal health.

BACKGROUND: Little published evidence supports the widely held contention that research in pregnancy is underfunded compared with other disease areas. OBJECTIVES: To assess absolute and relative government and charitable funding for maternal and perinatal research in the UK and internationally. SEARCH STRATEGY, SELECTION CRITERIA, DATA COLLECTION, AND ANALYSIS: Major research funding bodies and alliances were identified from an Internet search and discussions with opinion leaders/senior investigators. Websites and annual reports were reviewed for details of strategy, research spend, grants awarded, and allocation to maternal and/or perinatal disease using generic and disease-specific search terms. MAIN RESULTS: Within the imprecision in the data sets, < or =1% of health research spend in the UK was on maternal/perinatal health. Other countries fared better with 1-4% investment, although nonexclusive categorisation may render this an overestimate. In low-resource settings, government funders focused on infectious disease but not maternal and perinatal health despite high relative disease burden, while global philanthropy concentrated on service provision rather than research. Although research expenditure has been deemed as appropriate for 'reproductive health' disease burden in the UK, there are no data on the equity of maternal/perinatal research spend against disease burden, which globally may justify a manyfold increase. AUTHOR'S CONCLUSIONS: This systematic review of research expenditure and priorities from national and international funding bodies suggests relative underinvestment in maternal/perinatal health. Contributing factors include the low political priority given to women's health, the challenging nature of clinical research in pregnancy, and research capacity dearth as a consequence of chronic underinvestment.

Transfusional fetal complications after single intrauterine death in monochorionic multiple pregnancy are reduced but not prevented by vascular occlusion.

OBJECTIVE: To document co-twin death/pregnancy loss and brain injury after single intrauterine death (sIUD) in monochorionic pregnancies. DESIGN: A total of 135 pregnancies with sIUD were reviewed for co-twin IUD, miscarriage and abnormal antenatal and postnatal neuro-imaging. SETTING: A tertiary referral fetal medicine unit from 2000 to 2007. POPULATION OR SAMPLE: All cases referred with a single fetal death in monochorionic pregnancy, including those where sIUD was spontaneous or occurred after fetoscopic laser treatment, or resulted from selective termination by cord occlusion with bipolar diathermy or intrafetal vascular ablation with interstitial laser. METHODS: Clinical details and ultrasound findings of the study population were retrieved from ultrasound and institutional databases. Delivery and neonatal outcome data were obtained from discharge summaries supplemented by individual chart review. MAIN OUTCOME MEASURES: Co-twin death or pregnancy loss and neurologic injury assessed on antenatal ultrasound and MR-imaging. RESULTS: A total of 81 sIUDs resulted from vascular occlusive feticide (diathermy or interstitial laser), 22 followed placental laser and 32 were spontaneous. In 22 pregnancies (16.8%), the co-twin died in utero and eight pregnancies miscarried (6.1%). Antenatal magnetic resonance (MR) imaging in 76/91 (83.5%) continuing pregnancies detected antenatal brain injury in five (6.6%). Three infants (two not scanned antenatally) had abnormalities detected postnatally. Brain abnormality was detected less often after procedure related (2.6%, 2/77) than spontaneous sIUD (22.2%, 6/27, P = 0.003) and after early compared with late gestation sIUD (3.6%, 4/111 versus 20.0%, 4/20; P = 0.02). CONCLUSIONS: We confirm substantial co-twin loss (22.9%) after monochorionic sIUD, but a low risk of antenatally acquired MRI-identified brain injury, suggesting this risk has been overestimated. Procedures restricting inter-twin transfusion reduce, but do not negate risk of brain injury.

The placenta contributes to activation of the renin angiotensin system in twin-twin transfusion syndrome.

The renin-angiotensin system (RAS) in twin-twin transfusion syndrome (TTTS) is up-regulated in the donor fetus's kidneys, but down-regulated in the recipient's. Ultrasonographic and echocardiographic features suggest that the recipient is also exposed to RAS components. In this study we investigated the role and origin of RAS components in the recipient fetus. Monochorionic diamniotic (MCDA) pregnancies were recruited from a tertiary fetal medicine service. Cord blood was collected from MCDA twins (TTTS and control non-TTTS) at delivery for renin and angiotensin II immunoassays. Placental tissue was flash-frozen for mRNA and protein expression or formalin-fixed for immunohistochemistry. Archival placenta and kidney samples were used for immunohistochemistry and in-situ hybridization. Plasma renin levels were elevated (p<0.05) in recipients (median 201 pg/ml, range 54-315 pg/ml) and donors (125 pg/ml, 25-296) with TTTS compared to controls (2.5 pg/ml, 1.1-1.5 pg/ml). The same was found with angiotensin II with high levels in both recipients (300.5 pg/ml, 86.1-488 pg/ml) and donors (239 pg/ml, 76.6-422) compared to controls (169.5 pg/ml, 89-220 pg/ml, p<0.05). Renin mRNA expression, and protein appeared qualitatively higher in the placental territory of the recipient compared to that of the donor and non-TTTS controls. We conclude that both fetuses in TTTS are exposed to high levels of RAS components; these appear to be produced from different sites, namely the kidney of the donor, and the placenta of the recipient. Given the markedly different phenotypes in the genetically identical fetuses with TTTS, we suggest that the source of RAS components may influence their clinical manifestations.

The natural caesarean: a woman-centred technique.

Although much effort has gone into promoting early skin-to-skin contact and parental involvement at vaginal birth, caesarean birth remains entrenched in surgical and resuscitative rituals, which delay parental contact, impair maternal satisfaction and reduce breastfeeding. We describe a 'natural' approach that mimics the situation at vaginal birth by allowing (i) the parents to watch the birth of their child as active participants (ii) slow delivery with physiological autoresuscitation and (iii) the baby to be transferred directly onto the mother's chest for early skin-to-skin. Studies are required into methods of reforming caesarean section, the most common operation worldwide.

Longitudinal blood flow in shared (arteriovenous anastomoses) and non-shared cotyledons in monochorionic placentae.

OBJECTIVES: In this study we aimed to quantitate monochorionic twin placental blood flow in vivo through arterio-venous anastomoses (AVA) and corresponding vessels within normal cotyledons. METHODS: The topography of chorionic plate vasculature was mapped using colour Doppler in ten monochorionic diamniotic twin (MCDA) pregnancies. Cotyledonary flow was derived by insonation of chorionic veins draining normal (n=10) and paired control shared cotyledons (n=10). Venous volume flow was calculated from five determinations of vessel diameter and three of time average mean velocity (TAMV). Measurements were repeated every 2-4 weeks from 18 until 32 weeks' gestation. RESULTS: Blood flow through non-shared and shared cotyledons increased with gestation (p<0.0001). Median flow at 28 weeks through shared cotyledons was 16 ml/min (15-21) (median, interquartile range), lower than in shared cotyledons (31, 25-35) (p<0.001), as was median volume flow across gestation calculated as area under the curve (shared cotyledons 126 (122-167), control cotyledons 269 (214-274), p=0.01). However, velocity was similar, with the difference due to smaller vein diameters draining shared compared to normal cotyledons (mean 3.6mm (SD 0.8) vs. 4.5mm (0.8), p=0.004). Ex vivo quantitation of insonated cotyledons and of all cotyledons confirmed the difference in vein diameter in the placentae studied. CONCLUSIONS: Blood flow through shared cotyledons was lower across gestation than through paired normal cotyledons in the placenta studied due to the smaller diameter of the AVA vessels. The size of AVAs rather than simply their presence and direction may contribute to determining transfusional imbalance in monochorionic twins.

Histomorphometric characterisation of shared and non-shared cotyledonary villus territories of monochorionic placentae in relation to pregnancy complications.

OBJECTIVE: Theoretical estimates and physiological inferences suggest that the structure of a shared cotyledon differs from a non-shared cotyledon. The aim of this study was to characterise the histomorphometry of terminal villi in shared and non-shared cotyledons in monochorionic placentae, both from uncomplicated twins and from those with twin-twin transfusion syndrome (TTTS) or discordant growth restriction (DeltaIUGR). METHODS: Forty-one monochorionic placentae from Caucasian non-smokers were obtained at caesarean section. Their vascular anatomy and placental territories were ascertained by dye injection. After fixation, full thickness histological blocks were obtained by systematic random sampling from each twin's territory and the shared cotyledons. Fifty randomly selected terminal villi were assessed for: (i) median villus diameter (ii) median villus capillary diameter (iii) median fetomaternal diffusion distance (iv) median no. of capillaries/villus (v) degree of vascularization (median percentage cross-sectional area of terminal villi occupied by capillaries) using a stage micrometer and image analysis programme. The histomorphometric findings were then correlated with birthweight discordance, placental territory discordance and DeltaAVAs (no. of AVAs from smaller twin (donor) to larger twin (recipient) minus no. of AVAs from larger to smaller twin). RESULTS: Histomorphometric variables were similar in shared and non-shared cotyledons of uncomplicated MCDA twins. However, the median diameter of terminal villi in shared cotyledons in DeltaIUGR and TTTS placentae was significantly smaller [51.2 microm (48.2-58.3), p<0.001 and 52.6 microm (53.1-50.4), p<0.001], and had a similar number of smaller capillaries, larger fetomaternal diffusion distance and reduced vascularization compared to non-shared IUGR and TTTS placentae. However, Deltadiameter (defined as the difference between median diameters of terminal villi in large minus small twins' territories) rose with increasing birthweight discordance (Pearson correlation coefficient=0.82, p<0.001). Multiple linear regression analysis revealed that Deltadiameter was influenced by placental territory discordance (p<0.001) and birthweight discordance (p<0.01): log10 Deltadiameter=1.38+(0.01 x birthweight discordance)+(0.56 x log10 placental territory discordance) (R2=0.82, p<0.001), but there was no significant relationship with DeltaAVA and AAA. In the TTTS group, Deltadiameter correlated significantly with DeltaAVA only: log10Deltadiameter=1.44+(0.02 x DeltaAVA) (R2=0.3, p<0.001). CONCLUSIONS: This is the first study to characterise the histomorphometry of shared and non-shared cotyledons in MC twins. The findings suggest that abnormal placentation, rather than placental vascular anatomy may be responsible for DeltaIUGR in MC twins, whereas TTTS arises from imbalance in interfetal transfusion with resultant differing terminal villus histomorphometric features in donor, recipient and shared cotyledons.

Characterisation of deep arterio-venous anastomoses within monochorionic placentae by vascular casting.

OBJECTIVE: To characterise arterio-venous anastomoses (AVA) in monochorionic (MC) placentae and determine (i) whether shared cotyledons lie beneath the co-termination of an artery from one twin and a vein to the contralateral twin and (ii) whether all AVA can be detected by visual inspection of the chorionic plate. METHODS: Vascular casts were made of 15 MC placentae. The number of typical AVAs suspected visually before digestion was compared with the number of AVAs identified after acid digestion. RESULTS: Thirty-three of 67 (49%) suspected typical AVAs were confirmed as typical after casting. There were five false positives and no false negatives. The remainder were classified as atypical AVAs, found in > or =90% of MC placentae. Type I (small vascular connections between two apparently normal cotyledons not seen before casting) and Type II (shared cotyledons arising within larger apparently normal cotyledons) atypical AVAs were found in 53% and 73% of placentae, respectively. CONCLUSIONS: Only half the shared cotyledons in MC placentae are characterised by co-termination of an artery and vein on the chorionic plate. We report the existence of deep anastomoses beneath the chorionic plate that cannot be visualised by chorionic plate inspection. These findings have implications for laser treatment of twin-twin transfusion syndrome.

Fetal plasma testosterone correlates positively with cortisol.

BACKGROUND: Fetal exposure to testosterone has been implicated in programming childhood behaviour, but little is known about the determinants of fetal testosterone concentrations. AIMS: To investigate the relation between fetal testosterone and maternal and fetal cortisol. METHODS: Clinically indicated blood samples taken from 44 human fetuses (mean gestational age 27 weeks, range 15-38), together with paired maternal samples, were analysed for testosterone and cortisol concentrations. RESULTS: Male fetuses had significantly higher concentrations of testosterone than females. Female but not male fetal concentrations rose significantly with gestational age. Fetal testosterone correlated positively with both fetal cortisol and maternal testosterone concentrations. Multiple regression showed that maternal testosterone and fetal cortisol were independently correlated with fetal plasma testosterone in both sexes. CONCLUSION: Unlike the norm in the adult, where testosterone production is often inhibited by cortisol, in the fetus there is a positive link between the two.

Transfer and metabolism of thyrotropin releasing hormone across the perfused human term placenta.

The transport and uptake of TRH was investigated in the maternal-fetal-placental unit of perfused human term placenta. The degradation of TRH in biological fluid was first determined by incubating [125I]TRH with 100 microL 50% maternal or cord sera with or without pretreatment with 200 microM of p-hydroxymercuriphenyl sulfonic acid (p-HMSA), a proline dipeptidase inhibitor. Transplacental transfer of TRH was then studied by adding 10 microCi of [125I]- or [3H]TRH to the maternal circulation of dually perfused isolated lobule of human term placenta with or without 200 microM p-HMSA. Creatinine was used as an internal marker. The rate of degradation of TRH (P < 0.001) and inhibition by p-HMSA were significantly higher in maternal than cord sera (P < 0.05). In the maternal circulation, TRH concentration declined rapidly from 100% at time 0 to 33.5 +/- 1.2% at 120 min. The fetal concentration increased from undetectable levels to a maximum of 1.8 +/- 0.3% at 120 min with a low feto-maternal ratio (0.08 +/- 0.02). Perfusion in the presence of p-HMSA, however, did not significantly change fetal concentration, or the maternal and fetal concentration-time integral levels of TRH. Chromatography of maternal, fetal, and placental homogenates showed that TRH was metabolized by the placenta into small molecular weight fragments predominantly released in the maternal circulation. These results suggest that human placenta acts as an enzymatic barrier to the free passage of TRH.

Liposomal thyroxine: a noninvasive model for transplacental fetal therapy.

Drugs that cross the placenta sparingly are currently given directly to the fetus by invasive procedures. We investigated whether anionic small unilamellar (SUV) liposomes of different lipid compositions enhanced the transfer and uptake of T4 in an in vitro model of perfused human term placenta. T4-encapsulated anionic liposomes were prepared using lecithin (F-SUV) or distearoyl phosphatidylcholine (S-SUV) with cholesterol and dicetylcholine. The size distribution, encapsulation efficiency, and stability were determined in blood-based media. The transfer kinetics of free and liposomally encapsulated T4 were studied in a dually perfused isolated lobule of human term placenta, with creatinine and liposomal carboxyfluorescein as marker substances. Concentrations of T4 and rT3 were measured by RIA. T4 crossed the placenta sparingly (1.9 +/- 0.5%) because it was metabolized to rT3 (9.2 +/- 1.3%). Transplacental transfer of T4 was significantly increased by F-SUV (15.8 +/- 2.1%; P < 0.001) and S-SUV liposomes (7.1 +/- 1.2%; P < 0.001), with a concomitant decrease in fetal rT3 levels (P < 0.001). Placental uptake of F-SUV (13.5 +/- 2.0%; P < 0.001) was greater than that of S-SUV liposomes (6.7 +/- 0.8%; P < 0.001). Our data suggest that anionic liposomes increase transplacental transfer of T4. If confirmed in vivo, liposomes may provide an alternative noninvasive method of drug delivery to the fetus.

Transport and metabolism of thyrotrophin-releasing hormone across the fetal membrane.

To determine the transfer and metabolism of TRH by human fetal membranes, the bidirectional transport and uptake of TRH was investigated by adding 125I-labeled TRH (100,000 cpm) or commercial TRH either to the maternal or the fetal compartment of an in vitro model of cultured human fetal membranes obtained from term and preterm placenta. Transmembrane transfer was also studied in the presence of 200 microM p-hydroxy-mercuriphenyl-sulphonic acid (p-HMSA), a dipeptidase enzyme inhibitor. Creatinine and heparin were used as an internal markers. Metabolites of TRH were separated from intact molecules by gel filtration on Sephadex G-10. The structural integrity of the membrane was confirmed by electron microscopy. The transmembrane transfer of radiolabeled and commercial TRH were comparable across both preterm and term placenta. When transport was studied from the maternal to fetal side, the maternal concentration of TRH declined rapidly from 100% at time 0 to 19.31 +/- 2.26% at 8 h with a concomitant increase in the fetal concentration from undetectable to a maximum of 2.56 +/- 0.38% with a fetomaternal ratio of 0.16 +/- 0.01. Transfer of TRH from the fetal to maternal compartment was similar to that of maternal to fetal. Chromatography of maternal and fetal media showed that TRH was metabolized by the membrane into small molecular weight fragments. Treatment of the membrane with p-HMSA increased TRH transport from the maternal to fetal compartment to 18.12 +/- 0.91 (P < 0.001) with an fetomaternal ratio of 0.35 +/- 0.02 (P < 0.001). Although transmembrane transfer of TRH from the fetal to maternal side was also increased by p-HMSA, levels achieved were less than that from maternal to fetal (12.26 +/- 1.50%; P < 0.05). These results suggest that the human fetal membrane acts as an enzymatic barrier to the bidirectional transfer of TRH from 24 weeks gestation.

Fetal hypothalamic-pituitary-adrenal stress responses to invasive procedures are independent of maternal responses.

Paired fetal and maternal samples were obtained, at fetal blood sampling and intrauterine transfusion, to study hypothalamic-pituitary-adrenal stress responses. This confirmed that the fetus mounts an hypothalamic-pituitary-adrenal stress response to transfusion via the intrahepatic vein, which involves piercing the fetal trunk, but not to transfusion via the placental cord insertion [mean cortisol response via intrahepatic vein delta = 52.6 nmol/L, 95% CI (25.3-79.9), P = 0.001; mean beta-endorphin response delta =106 pg/mL, 95% CI (45.3-167), P = 0.002]. Baseline maternal fetal ratios were 13 [95% CI (10.7-15.2)] for cortisol and 0.8 [95% CI (0.5-1.0)] for beta-endorphin. The novel findings were: 1) that the fetal responses were independent of those of the mother, which did not change during transfusion at either site; 2) that there was a correlation between baseline fetal and maternal cortisol levels (r = 0.58, n = 51, P < 0.0001) but not between baseline fetal and maternal ss-endorphin levels, suggesting cortisol transfer across the placenta, rather than joint control by placental CRH; and 3) that fetal beta-endorphin responses were apparent from 18 weeks gestation and independent of gestational age, whereas fetal cortisol responses were apparent from 20 weeks gestation and were dependent on gestational age (y = -91.4 + 5.08x, r = 0.51; n = 16; P = 0.04; CI for slope, 0.16-10.0), consistent with the maturation of the fetal pituitary before the fetal adrenal.

Twin-to-twin transfusion syndrome results from dynamic asymmetrical reduction in placental anastomoses: a hypothesis.

Although placental vascular anastomoses between the fetoplacental circulations are ubiquitous in monochorionic twin pregnancies, the factors regulating their formation and maintenance are not understood. Increasing evidence implicates asymmetric anastomotic patterns in the aetiology of severe twin-to-twin transfusion syndrome (TTTS). The authors propose that anastomoses between placental circulations in monochorionic twins occur in a random manner at the embryological stage of connection of embryonic and extra-embryonic circulations. Placental expansion is then associated with random disruption of anastomoses and regression of their associated villus districts. TTTS develops as discordant loss of anastomoses results in asymmetrical flow resistance. Pregnancies with fetal growth concordance but discordant nuchal translucency at 10-14 weeks are at increased risk of developing subsequent severe TTTS because these clinical features indicate significant pressure differentials in the presence of a placentoplacental circulation, consistent with the presence of numerous, asymmetric anastomoses. However, since the anastomotic pattern is dynamic in the first half of pregnancy this hypothesis predicts that it will not be possible to devise a clinical test at 12 weeks that will predict with certainty the outcome of monochorionic twin pregnancies in relation to TTTS because this depends on random subsequent events.

Pseudo-arterio-arterial anastomoses in twin-twin transfusion syndrome.

OBJECTIVE: It has recently been claimed that fetoscopic recognition of a haemodynamic equator within an arterio-arterial anastomosis (AAA) suggests minimal net intertwin flow. This was based on blood from one fetus being dark and from the other bright red, the boundary between them reciprocating with the fetal heart beats. However, bright red indicates that the blood had passed through a cotyledon and been freshly oxygenated, which should be impossible in an AAA. We applied a computer model of chorionic vessels to determine a configuration that reproduced this phenomenon. METHODS: A previously published TTTS model was extended to provide placental detail in a segment containing four cotyledons of each placenta supplied by three generations of placental arteries and veins. RESULTS: Reciprocating flow is not unique to AAAs. It also occurs in the chorionic arteries of any cotyledon deprived of its venous outflow, in a similar manner to that in which reverse end-diastolic flow occurs in umbilical arteries when whole placental resistance is high. If venous return from the common chorionic vein in the recipient (draining the venous end of an AVA) is blocked as might happen after laser, there can be bidirectional flow from one umbilical artery insertion, through two cotyledons to the other insertion. We define this phenomenon as a pseudo-AAA (PAAA). The inclusion of two cotyledons in this path means that its resistance cannot match the low flow resistance of a true AAA, and transmission of the contralateral pulsatile pattern is absorbed in the cotyledons. Thus, PAAA Doppler patterns differ from true AAA patterns in that two sets of systolic peaks, one forward and one reverse, can be discerned in true AAAs but only one in PAAAs. CONCLUSIONS: We demonstrate how venous occlusion of an arterio-venous anastomosis may produce a pseudo-AAA colour equator at endoscopy. However, visual observation of reciprocating flow is not sufficient to define a vessel as a true AAA which instead requires ultrasonical identification of two systolic patterns.

Derivation of rate of arterio-arterial anastomotic transfusion between monochorionic twin fetuses by Doppler waveform analysis.

We aimed to determine rates of interfetal transfusion along arterio-arterial (AA) anastomoses in monochorionic twins in vivo from analysis of Doppler waveform patterns. Twenty-one monochorionic twin pregnancies in which an AA anastomosis was identified antenatally underwent serial Doppler velocimetry. Unidirectional AA anastomotic flow rates increased with increasing gestational age (log y = 8 x 10(-9)x - 5 x 10(-8); p = 0.0002). The mean net rate of flow through an AA anastomosis at 28 weeks gestation was 7.6 x 10(-8) l/s (SD = 4.9 x 10(-8) l/s). This flow was significantly related to the distribution of arterio-venous (AV) anastomoses (p = 0.009) and birthweight discordancy (p = 0.006). We derived estimates of flow along individual AV anastomoses by assuming that net AA countertransfusion is shared equally among uncompensated AV anastomoses, and speculate that the median AV flow rate at 28 weeks is in the order of 6 x 10(-8) l/s. In conclusion, this study demonstrates that flow rates along AA anastomoses can be quantified antenatally. These are the first estimates of flow rates in vivo along placental anastomoses. Although AA net flows are modest, chronic unbalanced counterflow of this magnitude in the absence of compensatory superficial anastomoses could lead to significant haemodynamic compromise.

Occlusion of arterio-arterial anastomosis manifesting as acute twin-twin transfusion syndrome.

In vivo, ex vivo and modelling studies suggest that arterio-arterial anastomoses (AAAs) protect against haemodynamic imbalance in monochorionic twins and thus the development of TTTS. We report the acute onset of severe TTTS at 34 weeks' gestation in a patient with an antenatally visualized AAA which was shown at injection studies to have been obliterated, presumably by thrombosis. Computer modelling with the relevant clinical data confirmed that occlusion of the AAA alone was sufficient to reproduce the clinical manifestations. A study of the vascular configuration of AAA in the fixed placenta suggested that its small diameter and turbulent flow may have contributed to its occlusion. This case report shows that the unmasking of unbalanced AVA configurations by occlusion of a protective AAA can manifest as TTTS.

Transmitted arterio-arterial anastomosis waveforms causing cyclically intermittent absent/reversed end-diastolic umbilical artery flow in monochorionic twins.

OBJECTIVES: To characterize the phenomenon of retrograde transmission of arterio-arterial anastomosis (AAA) interference patterns on umbilical artery (UA) waveform by (a) documenting the periodicity, (b) correlation with in vivo and in vitro demonstration of AAAs and (c) reproducing these patterns by computer modelling. METHODS: Monochorionic twins (MC) twins underwent placental and umbilical Doppler studies. AAAs were sought by pulse wave Doppler of their bi-directional interference pattern and confirmed by postnatal injection studies. The periodicity of transmitted patterns in the UA was determined. Determinants of the transmitted patterns were ascertained by computer modelling of physiological and fetal variables. RESULTS: Among 83 prospectively studied MC twin pregnancies; a transmitted pattern was observed in 6 (7 per cent) patients for 15-114 days. This was found in 20 per cent (6/30) of smaller MC twins discordant for growth restriction but in no appropriately grown twins. It was only observed in association with AAAs validated both in vivo and in ex vivo. Computer modelling demonstrated that this pattern could be reproduced by summating end diastolic flow with a high pulsatility index in the UA in the presence of a large AAA. Consistent with this, MC twins with a transmitted pattern had larger AAAs (median diameter 4.3 mm interquartile range 4.1-5.2) compared to MC twins discordant for intrauterine growth restriction (2.1 mm interquartile range 1.5 to 2.8) (P<0.05) without a transmitted pattern. Perinatal mortality was similar in the fetuses with and without transmitted patterns (0/12 vs. 2/48 P=0.7).