Fetal tissue sampling. The San Francisco experience with 190 pregnancies.

Prenatal diagnosis of genetic defects was done using fetal blood sampling in 167 at-risk pregnancies, by fetal skin biopsy in 15 pregnancies, and by fetal liver biopsy in 8 pregnancies. Fetal blood sampling was done by fetoscopy through January 1985 and by sonographically directed percutaneous umbilical blood sampling since then. In our series, cytogenetics has become the major indication for fetal blood sampling, increasing from 6% of the cases with fetoscopy to 48% with umbilical blood sampling. Fetoscopy provided pure fetal blood in 61% of cases while umbilical blood sampling provided pure fetal blood 97% of the time. The corrected risk of fetal demise after percutaneous umbilical fetal blood sampling was 2% and after fetoscopy was 4%.     

Method of Breech Management Incorporating Use of Fetal Blood Sampling

We have developed a method of breech management based on the use of fetal blood sampling. Twenty-five cases were studied throughout labour, and fetal blood samples taken throughout the first and second stages. These showed that the Apgar score at one minute correlated closely with the fetal pH just before delivery. We believe that cord compression is an important variable factor which can be assessed only by fetal blood sampling. If this shows that fetal anoxia is becoming severe then immediate delivery is mandatory.     

Antenatal diagnosis of thalassaemia major.

Haemoglobin synthesis was studied in fetal blood samples obtained at 17 to 20 weeks'' gestation in 22 women at risk of carrying a fetus with homozygous beta-thalassaemia. A presumptive diagnosis of homozygous beta-thalassaemia was made in four cases, and the pregnancy was terminated. An inconclusive answer was obtained in one case, and the patient also chose to have her pregnancy terminated. Two fetuses were lost as a result of the procedure. Of the remaining 15 pregnancies, 13 proceeded to term and two to 36 weeks; in each case a normal infant or one heterozygous for beta-thalassaemia was delivered. Current efforts should be directed towards improving the blood sampling technology so that fetal blood sampling can be used widely in those countries where thalassaemia is a major problem.     

Prenatal asphyxia,hyperlacticaemia, hypoglycaemia,and erythroblastosis in growth retarded fetuses.

The umbilical venous oxygen and carbon dioxide tensions, pH, lactate and glucose concentrations, nucleated red cell (erythroblast) count, and haemoglobin concentration were measured in 38 cases of intrauterine growth retardation in which fetal blood sampling was performed by cordocentesis. The oxygen tension was below the normal mean for gestational age in 33 cases; in 14 it was below the lower limit of the 95% confidence interval for normal pregnancies. The severity of fetal hypoxia correlated significantly with fetal hypercapnia, acidosis, hyperlacticaemia, hypoglycaemia, and erythroblastosis. These findings indicate that "birth asphyxia" is not necessarily due to the process of birth.     

Perinatal diagnosis of passive ITP: use of percutaneous umbilical blood sampling (PUBS)

Fetal blood samples can be obtained in utero by direct sampling of the umbilical cord vessels, using an ultrasound guided technique termed percutaneous umbilical sampling (PUBS). This procedure is being used more frequently in high risk pregnancies to obtain direct fetal laboratory data. In specialized centers, with trained personnel, the technique can be used with a high degree of safety and efficiency. Direct access to the fetal circulation can also allow an accurate determination of the fetal platelet count in cases of suspected fetal thrombocytopenia. The technique may be used to plan appropriate clinical management of maternal ITP as well as to diagnose the presence of fetal alloimmune thrombocytopenia. A logical strategy for obstetric management and evaluation of fetal risk can be planned. The procedure also has the potential to allow direct fetal treatment as has been the case in the management of severe fetal anemia.     

Fetal blood sampling in baboons (Papio spp.): Important procedural aspects and literature review

Background  The baboons ( Papio cynocephalus ) have similarities with human placentation and fetal development. Fetal blood sampling allows investigators to assess fetal condition at a specific point in gestation as well as transplacental transfer of medications. Unfortunately, assessing fetal status during gestation has been difficult and fetal instrumentation associated with high rate of pregnancy loss. Our objectives are to describe the technique of ultrasound guided cordocentesis (UGC) in baboons, report post-procedural outcomes, and review existing publications.
Methods  This is a procedural paper describing the technique of UGC in baboons. After confirming pregnancy and gestational age via ultrasound, animals participating in approved research protocols that required fetal assessment underwent UGC.
Results  We successfully performed UGC in four animals (five samples) using this technique. Animals were sampled in the second and third trimesters with fetal blood sampling achieved by sampling a free cord loop, placental cord insertion site or the intrahepatic umbilical vein. All procedures were without complication and these animals delivered at term.
Conclusions  Ultrasound guided fetal umbilical cord venipuncture is a useful and safe technique to sample the fetal circulation with minimal risk to the fetus or mother. We believe this technique could be used for repeated fetal venous blood sampling in the baboons.     

Placental transfer of synthetic and endogenous estrogen conjugates in the rhesus monkey (Macaca mulatta)

The placental transfer of conjugated and nonconjugated estrogens was compared in the pregnant rhesus monkey. Placement of catheters in the maternal and fetal circulation allowed for the sampling of blood after the administration of radiolabeled naturally occurring and synthetic estrogens to mother or fetus. In all cases, nonconjugated-estrogen placental transfer was greater than conjugated-estrogen transfer. Comparison of the conjugated estrogens suggested that diethylstilbestrol-monoglucuronide (DESG) was transferred less efficiently than the estrone-sulfate (E, S). High-performance liquid chromatog-raphy (HPLC) of selected plasma samples revealed that 50–90% of the E, S observed in the maternal circulation was cleaved after fetal administration. In contrast, HPLC of maternal or fetal plasma samples after DESG administration revealed only intact DESG. These results emphasize differences in the placental transfer of the synthetic and naturally occurring estrogen hormones.     

Fetal-maternal transfer of [H3]-6 beta-hydroxycortisol in the pregnant ewe

Distribution and fetomaternal transfer of 6 beta-hydroxycortisol (6 beta-OHF) was studied using serial sampling following injection of tritium labelled 6 beta-OHF into various fluid compartments in the chronically cannulated unaesthesized pregnant ewe. There was a rapid transfer of 6 beta-OHF from the fetal circulation into amniotic fluid and maternal blood. In contrast, the maternal----fetal transfer of this steroid metabolite was considerably less. The sequence of appearance of 6 beta-OHF in fetal blood and amniotic fluid following injection into maternal blood suggests that this steroid is first transferred across the placenta to fetal blood before gaining entry into the amniotic fluid space. The half-lives of 6 beta-OHF after initial equilibration in maternal plasma, fetal plasma and amniotic fluid were 2.0 h, 5.1 h and 8.9 h respectively. The amniotic sac appears to contain a relatively static pool of 6 beta-OHF and may act as a "trap" for 6 beta-OHF in the ovine conceptus.     

Plasma placental RNA allelic ratio permits noninvasive prenatal chromosomal aneuploidy detection

Current methods for prenatal diagnosis of chromosomal aneuploidies involve the invasive sampling of fetal materials using procedures such as amniocentesis or chorionic villus sampling and constitute a finite risk to the fetus. Here, we outline a strategy for fetal chromosome dosage assessment that can be performed noninvasively through analysis of placental expressed mRNA in maternal plasma. We achieved noninvasive prenatal diagnosis of fetal trisomy 21 by determining the ratio between alleles of a single-nucleotide polymorphism (SNP) in PLAC4 mRNA, which is transcribed from chromosome 21 and expressed by the placenta, in maternal plasma. PLAC4 mRNA in maternal plasma was fetal derived and cleared after delivery. The allelic ratios in maternal plasma correlated with those in the placenta. Fetal trisomy 21 was detected noninvasively in 90% of cases and excluded in 96.5% of controls.     

Fetal distress and the condition of newborn infants.

In a prospective audit of the obstetric management of 1210 consecutive deliveries the association was investigated between the need for operative delivery for fetal distress during labour and the condition of the newborn infant. Operative delivery was performed for only 11.5% of the newborn infants with severe acidosis at birth (umbilical artery pH less than 7.12, base deficit greater than 12 mmol (mEq)/1), 24.1% of those with an Apgar score less than 7 at one minute, and 15.8% of those with both severe acidosis and a one minute Apgar score less than 7. Most of the infants delivered operatively were in a vigorous condition at birth and did not have severe acidosis. Fetal blood sampling was done in 4.0% of labours. As none of the fetal blood values were less than 7.20 and only three of the infants sampled in utero suffered severe acidosis at birth, fetal blood sampling would have had to be performed much more often to provide a useful guide to metabolic state at birth. While the large majority of "at risk" fetuses had continuous fetal heart rate monitoring in labour, this had not been provided in 48.7% of the labours of infants with severe acidosis, 38.7% of infants with a one minute Apgar score less than 7, and 47.4% of infants with both severe acidosis and a one minute Apgar score less than 7. Continuous fetal heart rate monitoring was associated with a much higher incidence of operative delivery for fetal distress than was intermittent fetal heart rate auscultation. These results suggest an urgent need to review present methods for assessing the intrapartum condition of the fetus, making the diagnosis of fetal distress, and assessing the condition of the infant at birth.     

Ultrasound-guided blood sampling of rabbit fetuses.

A rabbit animal model for hemolytic disease of the newborn has been previously described. However, evaluating the effects of this disease was limited to histologic and hematologic examinations of liveborn kitlings. To assess the feasibility of in utero blood sampling, we performed ultrasound-guided cardiac sampling of 50 fetuses in 16 New Zealand White does on days 26 and 27 of gestation. The overall rate of successful sampling was 80%. The procedure-related mortality declined to 35% by the third phase of the study. The mean (+/- SD) hematocrit (%) and reticulocyte values (#/100 RBCs) on day 26 were 26.3 +/- 3.3 and 35.6 +/- 5.1, respectively; values on day 27 were 31.3 +/- 4.9 and 27.5 +/- 7.6. The results of this study suggest that hematologic data can be obtained from rabbit fetuses in the majority of cases with only moderate fetal loss.     

Blood flow in ductus venosus in early uncomplicated pregnancy

The ductus venosus is the very important part of fetal venous circulation. It plays a central role in return of venous blood from the placenta. This unique shunt carries well-oxygenated blood from the umbilical vein through the inferior atrial inlet on its way across the foramen ovale. Using Doppler ultrasound, it is possible to assess the blood flow in fetal vessels including ductus venosus. It is observed, in animal and human studies, that the typical waveform for blood flow in ductus venosus in early pregnancy can be different depending on numerous conditions e.g. fetal karyotype. This study is performed to assess the physiologic parameters of blood flow in ductus venosus in uncomplicated early pregnancy. Material and Methods: 404 women were examined between 11+0 and 13+6 weeks (+ days) of gestation by ultrasound. Fetal crown-rump length (CRL) was measured to assess the gestational age. The assessment of risk of fetal abnormalities was based on nuchal translucency (NT) measurement. The ductus venosus blood flow with color and spectral Doppler was obtained in all patients. The following features were assessed: pulsatility index (PI), and direction of flow (positive/negative) during atrial contraction (wave A). All cases were followed up to 22 weeks of gestation when the control scan was performed. Results and conclusions: 30 cases were excluded from the uncomplicated group due to: high risk of fetal abnormalities, fetal loss, confirmed fetal abnormalities and utero-placental pathology. 374 women were considered as uncomplicated pregnancy. In both uncomplicated and complicated groups the mean values for pulsatility index (PI) were established. The mean PI value in uncomplicated pregnancies was: 0.91 (SD +/- 0.32). No significant differences between groups were noticed. In 370 cases of uncomplicated pregnancy the A wave direction was positive but in 1.1% of cases the reverse flow in atrial contraction was observed.     

Effect of cooling and heating on the regional distribution of blood flow in fetal sheep

This is a study on the effect of cooling and heating amniotic fluid on blood flow to fetal tissues and organs. In 8 unanaesthetized, chronically-catheterised fetal sheep (129-137 days gestation) cold or warm water was passed through tubing encircling the fetus in utero and blood flow was measured using the radionuclide-labelled 15 mu spheres. Following cooling for 30 min, amniotic fluid temperature fell 9.6 degrees C to 29.9 +/- 2.1 degrees C (SEM) fetal arterial temperature fell 2.37 degrees C to 37.30 +/- 0.36, and maternal arterial temperature fell 0.53 degrees C to 38.58 +/- 0.16. Blood flow through the fetal skin fell 60% (P less than 0.01) to 13.6 ml/min per 100 g tissue. Blood flow to the brown fat increased 186% (P less than 0.05) to 99.6 ml/min per 100 g. Following warming for 20 min, fetal temperature rose to 40.43 +/- 0.19 degrees C, and skin blood flow did not change significantly relative to initial control period but rose 200% above that during cooling (P less than 0.01). During both cooling and heating, blood flow to the adrenals rose significantly (P less than 0.05) whereas flow to the carcass, brain, kidneys, and placenta was not altered detectably. Continuous sampling of blood from the inferior vena cava during microsphere injection failed to detect any evidence of arterio-venous shunting through the skin at any temperature studied. Overall, the blood flow responses are consistent with a thermoregulatory role for the skin and brown fat in the near-term fetal sheep.     

In utero paternity testing utilizing fetal blood obtained by midtrimester fetoscopy.

To evaluate paternity in a case where both rape and conjugal coitus had occurred close to the time of conception, fetoscopy with fetal blood sampling was performed at 20 weeks gestation. Detailed blood group typing of the wife, husband, and fetus, and the presence of a very long Y chromosome in the last two, indicated a 99.9% chance that the fetus was fathered by the husband and only a 0.1% chance that it was fathered by "some other male Caucasian." The couple elected to continue the pregnancy. Neonatal testing verified the prenatal findings.     

Sampling pure fetal blood by fetoscopy in second trimester of pregnancy.

A technique for fetal blood-sampling in the second trimester of pregnancy (between 16 and 22 weeks'' gestation) combining fetoscopy with real-time ultrasound was used in 48 attempts at fetal blood-sampling. Specimens containing fetal red cells with or without amniotic fluid or maternal blood, and adequate for diagnosing haemoglobinopathies, were obtained in 45 of the 48 fetoscopies. Sampling was successful in all 18 patients with a posterior placenta, and in 27 of the 30 with an anterior placenta. In 22 of the last 27 consecutive fetoscopies pure fetal blood was taken; the placenta was anterior in 16 and posterior in six. Out of 17 cases sampled between 18 and 22 weeks'' gestation pure fetal blood was obtained in 16. The volume of the samples varied from 50 to 500 microliter. The ability to obtain pure fetal blood consistently even when the placenta is anterior will increase knowledge of fetal physiology and the scope of prenatal diagnosis.     

Fetal cells in maternal blood: recovery by charge flow separation

Fetal blood cells can be recovered from the maternal circulation by charge flow separation (CFS), a method that obviates the risks associated with amniocentesis and chorionic villus sampling. By CFS, we processed blood samples from 13 women carrying male fetuses, 2 carrying fetuses with trisomy 21, and 1 who had delivered a stillborn infant with trisomy 18. On average more than 2000 fetal nucleated red blood cells were recovered per 20-ml sample of maternal blood. Recovery of fetal cells was confirmed by fluorescence in situ hybridization with probes for chromosomes Y, 18 and 21. After culturing of CFS-processed cells, amplification by the polymerase chain reaction revealed Y-chromosomal DNA in clones from four of six women bearing male fetuses, but not in clones from three women bearing female fetuses. Received: 8 January 1996 / Revised: 22 March 1996     

Amino acid gradients across the intestinal circulation in fetal lambs.

Amino acids, including glutamine, glutamate and asparagine are major metabolic substrates for the adult enterocyte of several species. To determine whether circulating amino acids are utilized by the fetal intestine, we studied nine fetal sheep (mean gestational age 128 +/- 5 days; term: 147 days). Catheters were inserted into the descending aorta (DA) and the mesenteric vein (MV) to allow for simultaneous blood sampling across the intestine. Fetal blood gas, haemoglobin; O2 saturation and O2 tension were measured. Ammonia was determined by an enzymatic method and HPLC analysis was used to measure the content of all amino acids in DA (descending aorta) and MV (mesenteric vein). Intestinal blood flow measurements were obtained using the radionuclide microsphere method. Intestinal blood flow (81 +/- 28 ml/min/100g) and arterial pH (7.37 +/- 0.04) were within normal range for unstressed fetal lambs. Glutamine and glutamate were the only amino acids that were significantly taken up across the fetal intestinal circulation. The fetal intestine extracted approximately 21% of the delivered glutamine (6.8 +/- 4.5 mumol/min/100g), 7% of the delivered glutamate (1.3 +/- 1.1 mumol/min/100g) and 2.7% of the delivered oxygen (43.0 +/- 19.1 mumol/min/100g). These data suggest that glutamine and glutamate are major substrates for the intestine in unstressed fetal lambs.     

Current and emerging techniques of fetal cell separation from maternal blood

Intense research has been carried out in recent years into methods that aim to harvest fetal genetic material from maternal blood as substitutes to amniocentesis and chorionic villus sampling. Just over 30 years have past since the first fetal cells were separated from maternal blood using flow cytometry highlighting the prospect of non-invasive prenatal diagnosis of fetal abnormalities. The aim of this review paper is to describe the most commonly used cell separation methods with emphasis on the isolation of fetal cells from maternal blood. The most significant breakthroughs and advances in fetal cell separation are reviewed and critically analyzed. Although much has been accomplished using well established techniques, a rapid and inexpensive method to separate fetal cells with great accuracy, sensitivity and efficiency to maximize cell yield is still required. In the past decade MEMS (Micro Electro Mechanical Systems) technologies have enabled the miniaturization of many biological and medical laboratory processes. Lab-on-chip systems have been developed and encompass many modules capable of processing different biological samples. Such chips contain various integrated components such as separation channels, micropumps, mixers, reaction and detection chambers. This article will also explore new emerging MEMS based separation strategies, which hope to overcome the current limitations in fetal cell separation.     

ITP in pregnancy and the newborn: introduction

Idiopathic thrombocytopenic purpura (ITP) occurs more commonly in young women and is one of the commonest immune mediated disorders in pregnancy. It may exist as an incidental finding in an otherwise healthy pregnant woman or may be associated with symptomatic reduction in the platelet count and varying degrees of clinical hemorrhage. The condition termed incidental thrombocytopenia of pregnancy is invariably associated with a platelet count of greater than 100 x 10(9)/L and a very low incidence of fetal thrombocytopenia. Symptomatic thrombocytopenia is more commonly associated with low platelet counts in the fetus (estimated between 20%-40%). It has recently been suggested that the incidence of fetal thrombocytopenia is substantially lower than this figure. The management of ITP in pregnancy is complicated by the fact that fetal thrombocytopenia is difficult to diagnose and carries substantial risks during the delivery process with rare cases of fetal hemorrhage occurring spontaneously in utero. Unfortunately there are no laboratory studies that can be performed precisely in the mother that may predict the occurrence of fetal thrombocytopenia. Maternal management is usually directed towards treatment of maternal symptoms. Maternal treatment or response to treatment is inconsistently associated with predictable changes in the fetal platelet count. Obstetric management is aimed at reducing the risks of life threatening fetal hemorrhage occurring at the time of delivery, and fetal management is directed towards the obtaining of fetal platelet samples in order to plan an appropriate strategy for obstetrical delivery. Fetal blood samples are obtained either by a scalp vein puncture at the time of delivery or earlier in gestation by the use of the newer technique termed percutaneous umbilical blood sampling. Fetuses with platelet counts of less then 50 x 10(9)/L are generally delivered by cesarean section whereas those with counts greater than 50 x 10(9)/L are allowed to proceed with vaginal delivery assuming no obstetrical contraindications exist. The use of IVIgG therapy during pregnancy has theoretical implications on improving platelet counts in the mother in situations of severe hemorrhage, however cannot be considered to be appropriate treatment for the prevention of fetal thrombocytopenia, since the exogenous transport of IVIgG across the placenta appears to be inconsistent and unpredictable.(ABSTRACT TRUNCATED AT 400 WORDS)     

Nontrophoblastic placental tumors

The aim of the study was to investigate potential influence of placental tumors on fetal outcome. The study comprised 10 cases of placental tumors. The analysis included the sonographic assessment of the tumor, signs of fetal anemia, as well as signs of hemodynamic disturbances or heart failure, and intrauterine treatment. The fetal hemodynamic was examined on the basis of Doppler blood flow in the umbilical artery and vein, middle cerebral artery, and ductus venous. The evaluation of fetal heart included the measurement of heart size, blood flow through cardiac valves and the assessment of fetal heart function based on cardiovascular score. The fetal outcome was also assessed according to birthweight, gestational age at delivery, pH, Ap score at 5th minute, abnormal neurological development and the need of intrauterine therapy. Ten cases of placental tumors were prenatally detected from 1999 to 2011. Among them 7 cases of hypoechogenic, non-vascularized cysts were identified and these neither effected the hemodynamics nor complicated fetal outcome. The vascularized tumors (chorioangioma) were the cause of severe anemia and hemodynamic disturbances and these led to fetal cardiac heart failure. In all cases of vascularized tumors from 2-3 intrauterine transfusion were performed. Rich vascularized tumors (chorioangioma) may cause hemodynamic disturbances and fetal heart failure. This may require intrauterine treatment and may result in abnormal fetal outcome and neurological development.