In utero exposure to chlordecone affects histone modifications and activates LINE-1 in cord blood.

Environmental factors can induce detrimental consequences into adulthood life. In this study, we examined the epigenetic effects induced by in utero chlordecone (CD) exposure on human male cord blood as well as in blood-derived Ke-37 cell line. Genome-wide analysis of histone H3K4me3 distribution revealed that genes related to chromosome segregation, chromatin organization, and cell cycle have altered occupancy in their promoters. The affected regions were enriched in ESR1, SP family, and IKZF1 binding motifs. We also observed a global reduction in H3K9me3, markedly in repeated sequences of the genome. Decrease in H3K9me3 after CD exposure correlates with decreased methylation in LINE-1 promoters and telomere length extension. These observations on human cord blood were assessed in the Ke-37 human cell line. H3K4me3 and the expression of genes related to immune response, DNA repair, and chromatin organization, which were affected in human cord blood were also altered in CD-exposed Ke-37 cells. Our data suggest that developmental exposure to CD leads to profound changes in histone modification patterns and affects the processes controlled by them in human cord blood.

Good practices in collecting umbilical cord and placental blood / Boas práticas para a coleta de sangue de cordão umbilical e placentário / Buenas prácticas para la recolección de sangre del cordón umbilical y de la placenta

Abstract Objective: to identify the factors related to the quality of umbilical cord and placental blood specimens, and define best practices for their collection in a government bank of umbilical cord and placental blood. Method: this was a descriptive study, quantitative approach, performed at a government umbilical cord and placental blood bank, in two steps: 1) verification of the obstetric, neonatal and operational factors, using a specific tool for gathering data as non-participant observers; 2) definition of best practices by grouping non-conformities observed before, during and after blood collection. The data was analyzed using descriptive statistics and the following statistical software: Statistica(r) and R(r). Results: while there was a correlation with obstetrical and neonatal factors, there was a larger correlation with operational factors, resulting in the need to adjust the professional practices of the nursing staff and obstetrical team involved in collecting this type of blood. Based on these non-conformities we defined best practices for nurses before, during and after blood collection. Conclusion: the best practices defined in this study are an important management tool for the work of nurses in obtaining blood specimens of high cell quality.

Resumo Objetivos: identificar fatores relacionados à qualidade das amostras do sangue de cordão umbilical e placentário e definir boas práticas para sua coleta em um banco público de sangue de cordão umbilical e placentário. Método: pesquisa descritiva, abordagem quantitativa, realizada em um banco público de sangue de cordão umbilical e placentário, desenvolvida em duas etapas: 1) verificação dos fatores obstétricos, neonatais e operacionais, obtidos por coleta em instrumento próprio e observação não participante; 2) definição das boas práticas, por meio do agrupamento de não-conformidades observadas antes, durante e após a coleta do sangue. Os dados foram analisados por meio da estatística descritiva, utilizando-se dos softwares Statistica(r) e R(r). Resultados: houve correlação da influência dos fatores obstétricos e neonatais em menor escala quando comparados aos fatores operacionais, resultando na necessidade de readequar a prática profissional do enfermeiro e da equipe obstétrica envolvida no processo da coleta desse tipo de sangue. A partir das não-conformidades foram definidas boas práticas para o enfermeiro antes, durante e após a coleta. Conclusão: as boas práticas definidas neste estudo são importante ferramenta gerencial para o trabalho do enfermeiro na obtenção de amostras de sangue com alta qualidade celular.

Resumen Objetivos: identificar factores relacionados a la calidad de las muestras de la sangre del cordón umbilical y de la placenta y definir buenas prácticas para su recolección en un banco público de sangre del cordón umbilical y de la placenta. Método: investigación - descriptiva, abordaje cuantitativo, realizada en un banco público de sangre - del cordón umbilical y de la placenta, desarrollada en dos etapas: 1) verificación de los factores obstétricos, neonatales y operacionales, obtenidos por recolección con instrumento propio y observación no participante; 2) definición de las buenas prácticas, por medio del agrupamiento de no-conformidades observadas antes, durante y después de la recolección de la sangre. Los datos fueron analizados por medio de la estadística descriptiva, utilizando los softwares Statistica(r) y R(r). Resultados: hubo correlación de la influencia de los factores obstétricos y neonatales en menor escala cuando comparados a los factores operacionales, resultando en la necesidad de readecuar la práctica profesional del enfermero y del equipo obstétrico que participa en el proceso de la recolección de ese tipo de sangre. A partir de las no-conformidades fueron definidas buenas prácticas para el enfermero antes, durante y después de la recolección. Conclusión: las buenas prácticas definidas en este estudio son importante herramienta administrativa para el trabajo del enfermero en la obtención de muestras de sangre con alta calidad celular.

Prenatal screening of cytogenetic anomalies - a Western Indian experience.

BACKGROUND: Children born with congenital anomalies present a very high rate of perinatal death and neonatal mortality. Cytogenetic analysis is a convincing investigation along with clinical suspicion and biochemical screening tests. The current study was designed to characterize the prevalence and types of chromosomal abnormalities in high risk prenatal samples using different cytogenetic techniques. METHODS: This study was conducted on a total of 1,728 prenatal samples (1,324 amniotic fluids, 366 chorionic villi and 38 cord blood samples) from 1994 to 2014 at Institute of Human Genetics, Ahmedabad, India. Conventional karyotyping was conducted with GTG-banding. Molecular approaches were used (fluorescence in situ hybridization = FISH and/ or array-comparative genomic hybridization = aCGH) when indicated to detect karyotypic abnormalities. RESULTS: Abnormal karyotypes were detected in 125/1,728 (7.2%) cases. Trisomy 21 was the most common abnormality detected in 46 (2.7%) followed by trisomy 18 in 11 (0.6%) and trisomy 13 in 2 (0.1%) samples. Besides, structural abnormalities such as reciprocal and Robertsonian translocation were detected in 20 [1.2%] cases. Turner syndrome was diagnosed in seven (0.4%) cases; in six (0.34%) cases there was an inversion in the Y-chromosome. Heteromorphic variants were diagnosed in 22 (1.3%) cases. Finally, small supernumerary marker chromosomes (sSMC) were found in six (0.34%) cases. CONCLUSION: Conventional GTG-banding along with molecular cytogenetic techniques is useful in detecting genomic alterations and rearrangements. Comprehensive characterization of chromosomal rearrangements like sSMC has the potential to save potentially healthy fetuses from being terminated.

Prenatal diagnosis in severe von Willebrand disease families from India using combination of phenotypic and genotypic assays.

OBJECTIVE: This study aimed to offer genetic diagnosis to affected type 3 severe von Willebrand disease families. METHOD: Thirteen families were referred for prenatal diagnosis during the first and second trimesters of pregnancy. Prenatal diagnosis was offered by chorionic villus sampling between 11 and 12 weeks and by cordocentesis between 18 and 19.5 weeks of gestation. Phenotypic analysis included FVIII:C and von Willebrand factor antigen assays. A combination of molecular biological techniques which included PCR-restriction fragment length polymorphism technique using intron 40 variable number tandem repeat (VNTR) markers, conformation sensitive gel electrophoresis, direct DNA sequencing, and multiple ligation probe amplification (MLPA) were used to offer genotyic diagnosis in the remaining families. RESULTS: Diagnosis was offered by intron 40 VNTR analysis in eight families. In one family, the diagnosis was given by direct mutation detection technique, whereas in another diagnosis was given by MLPA technique as the index case showed the presence of large deletion within von Willebrand factor. In three families, diagnosis was offered by cordocentesis on the basis of phenotypic assays, further confirmed by genotyping. CONCLUSION: Both first and second-trimester prenatal diagnoses could be successfully offered using a combination of phenotypic and genotypic techniques to all severe von Willebrand disease families.

Invasive & non-invasive approaches for prenatal diagnosis of haemoglobinopathies: experiences from India.

The thalassaemias and sickle cell disease are the commonest monogenic disorders in India. There are an estimated 7500 - 12,000 babies with ß-thalassaemia major born every year in the country. While the overall prevalence of carriers in different States varies from 1.5 to 4 per cent, recent work has shown considerable variations in frequencies even within States. Thus, micromapping would help to determine the true burden of the disease. Although screening in antenatal clinics is being done at many centres, only 15-20 per cent of pregnant women register in antenatal clinics in public hospitals in the first trimester of pregnancy. There are only a handful of centres in major cities in this vast country where prenatal diagnosis is done. There is considerable molecular heterogeneity with 64 mutations identified, of which 6 to 7 common mutations account for 80-90 per cent of mutant alleles. First trimester foetal diagnosis is done by chorionic villus sampling (CVS) and DNA analysis using reverse dot blot hybridization, amplification refractory mutation system (ARMS) and DNA sequencing. Second trimester diagnosis is done by cordocentesis and foetal blood analysis on HPLC at a few centres. Our experience on prenatal diagnosis of haemoglobinopathies in 2221 pregnancies has shown that >90 per cent of couples were referred for prenatal diagnosis of ß-thalassaemia after having one or more affected children while about 35 per cent of couples were referred for prenatal diagnosis of sickle cell disorders prospectively. There is a clear need for more data from India on non-invasive approaches for prenatal diagnosis.

Feasibility of ultrasound-guided percutaneous samplings in the pregnant baboon: a model for studies on transplacental transport.

OBJECTIVE: A study was conducted to test the feasibility of cordocenteses and amniocenteses at different gestational ages in pregnant baboons. STUDY DESIGN: Experiments were performed in 10 pregnant baboons at a median gestational age of 131 (range 92-169) days. At different time intervals, percutaneous samplings of amniotic fluid and fetal blood were performed under ultrasound guidance. Simultaneously, maternal blood samples were drawn. RESULTS: With a median fetal weight of 431 g (range 111-690 g), 29 of 30 cordocenteses (96.6%) and all 30 amniocenteses and maternal samplings (100%) were successful in obtaining the required quantities for analysis. One cordocentesis was abandoned because of insufficient visualization of the umbilical cord due to a placental haemorrhage. CONCLUSION: Percutaneous amniocentesis and cordocentesis can be performed with a high success rate in the pregnant baboon model. In combination with a 100% success rate in obtaining simultaneous maternal blood samples, this method is able to provide data on transplacental transport.

Four-dimensional ultrasound guidance of prenatal invasive procedures.

OBJECTIVE: Technological advances in ultrasonography have revolutionized prenatal diagnosis and treatment. Here we evaluate the effectiveness of using four-dimensional (4D) ultrasonography to guide prenatal invasive procedures. PATIENTS AND METHODS: Prenatal invasive procedures using 4D ultrasound were recorded prospectively in 93 cases: 10 amnioinfusions, 50 amniocenteses, 8 chorionic villus samplings (CVS) and 25 cordocenteses. The needle target site was first identified using the two-dimensional (2D) mode, and was then confirmed using the three-dimensional (3D) mode. The needle was inserted under 4D ultrasound guidance. After selecting the needle target site, the true position of the needle was determined in three planes ('real-time 3D targeting'). RESULTS: Using 4D ultrasound guidance, most procedures were performed within 5 min and with a 100% success rate, even in cases involving severe oligohydramnios (amniocentesis), thin placentas (CVS) or narrow umbilical veins (cordocentesis). Moreover, there were no serious complications during or after any procedure. CONCLUSIONS: 4D ultrasonography can be used to guide various prenatal invasive procedures offering clear information in all three planes. It is likely that such imaging will reduce the time taken to complete the procedures and reduce the risks associated with them. Published by John Wiley & Sons, Ltd.

Four-dimensional ultrasonographic guidance for invasive obstetric procedures.

OBJECTIVE: This pilot study evaluated the use of 4-dimensional (4D) real-time ultrasonographic needle guidance for amniocentesis, chorionic villus sampling (CVS), cordocentesis, and intrauterine transfusions. METHODS: Ninety-nine consecutive procedures were performed with 4D (real-time) multiplanar ultrasonographic imaging. Amniocentesis was done freehand in 3 orthogonal planes of view. Chorionic villus sampling, cordocentesis, and intrauterine transfusions were accomplished with a needle guide and 2 projected orthogonal planes. RESULTS: Needle tip visualization in the A, B, and C orthogonal planes during amniocentesis was noted in 93%, 63%, and 69% of cases, respectively. When a needle guide was used during CVS and cordocentesis, the needle tip was always seen in the 2 projected orthogonal planes, and no lateralization occurred. Four intrauterine transfusions were done with the 4D technique. The only procedural complication in any patient was bradycardia from vessel spasm during an intrauterine transfusion, requiring a cesarean delivery. There were no statistical differences (P > .05) between the numbers of needle insertions required in the 4D group compared with a historical control group in which 2-dimensional ultrasonographic needle guidance was used. CONCLUSIONS: In this feasibility study, a real-time 4D needle guidance technique was successfully used to perform amniocentesis, CVS, cordocentesis, and intrauterine transfusion. This appeared to contribute to the accuracy of needle placement by eliminating the lateralization phenomenon when a fixed needle guide attachment was used (for CVS and cordocentesis). Needle tip visualization was seen in each orthogonal plane in most freehand 4D amniocentesis cases. Future developments in 4D ultrasonographic technology may refine the utility of this technique for invasive obstetric procedures.

La transfusión intrauterina por cordocentesis: Una alternativa terapéutica con expectativa de vida / Intrauterine transfusion by cordocentesis: A therapuetic alternative with life expectancy

La enfermedad hemolítica por anti D es una grave problemática de nuestro medio que es reconocido como causa de muerte perinatal. Conociendo que las pacientes con enfermedad hemolítica severa, las posibilidades de viabilidad fetal son nulas a pocas semanas de gestación donde otros tratamientos no fueron suficientes, nuestro trabajo demuestra que la transfusión intrauterina por cordocentesis (TIUPC) es una técnica que logra alta expectativa de vida ya que permite no solo reabsorber el hidrops fetal alcanzando embarazos a término con fetos viables, sino también evolución de los recién nacidos sin necesidad de internaciones prolongadas, con menor costo, en relación a los gastos ocasionados por el manejo multidisciplinario que requerían los neonatos sin este tratamiento.

Ultrasound-guided procedures for prenatal diagnosis and therapy.

Ultrasonography has expanded the capabilities of perinatologists to examine,test, and treat the fetus. Amniocentesis and CVS are safe and widely available procedures, which can be used to diagnose a multitude of abnormalities through karyotype analysis and molecular studies. CVS allows earlier diagnosis, but both procedures can provide highly accurate results in the first half of pregnancy. Cordocentesis has fewer indications, but allows direct laboratory testing of fetal blood. Fetocentesis and fetal biopsy are reserved for limited indications, but can play a crucial role in the diagnosis of some conditions, which cannot be assessed less invasively. Fetal transfusion is an important tool in the treatment of isoimmunization, some other forms of fetal anemia, and alloimmune thrombocytopenia. Amnioreduction is a commonly used procedure for the treatment of polyhydramnios and TTTS. Multifetal reduction and selective termination offer previously unavailable options to patients carrying multiple gestations. Fetal shunts can reduce perinatal morbidity and mortality in cases of bladder outlet obstruction and hydrothorax. The limited experience with cord ligation procedures and balloon valvuloplasty suggests these relatively new procedures may serve a greater role in the future as techniques are improved. By providing guidance for all of these procedures, real-time ultrasonography has revolutionized prenatal diagnosis and therapy; it will continue to be a crucial component in evaluating and treating complicated pregnancies.

Teaching ultrasound-guided invasive procedures in fetal medicine: learning curves with and without an electronic guidance system.

OBJECTIVE: To compare the learning curves of inexperienced junior obstetrics/gynecology registrars for ultrasound-guided invasive procedures on a training model, with and without an electronic guidance system. STUDY DESIGN: Four junior registrars performed their first 100 procedures on a training model with a new electronic guidance system, and four other junior registrars performed their first 100 procedures on the same training model without using the guidance system. All procedures were performed using a free-hand technique. We evaluated the quality of the procedure, which we defined as the time spent with the entire needle clearly visualized on the screen over the total duration of the procedure. We constructed learning curves for the eight junior registrars for comparative analysis. RESULTS: Quality of the procedure increased over time for all trainees. The learning curves were significantly steeper for trainees using the electronic guidance system. Trainees using the electronic guidance system performed better in the middle of their learning curve (procedures 25-75). All trainees reached the same level of quality by the end of their 100 procedures. CONCLUSIONS: The automated electronic guidance system helps faster learning but, after 100 procedures on a training model, both groups reached the same level of quality.

[Rapid karyotyping in the 2nd and 3rd trimester: results and experiences]. / Schnelle Karyotypisierung im II. und III. Trimenon: Ergebnisse und Erfahrungen.

Rapid karyotyping in the second and third trimester is an increasing field of collaboration between women's hospitals and human genetics. Techniques available for rapid karyotyping are: 1. Amniocentesis; to obtain amniotic fluid cells for culturing and subsequent chromosome harvesting using the pipette method or the "in situ" technique. The average time between preparation of the amniotic fluid and the verbal notification of the analysed karyotype is 4.65 days for the pipette method and 5.97 days for the "in situ" technique. The major advantages are that amniocentesis can be handled safely by many gynaecologist, and the amniotic fluid samples can be posted easily to cytogenetic units familiar with rapid karyotyping. The main disadvantage is that currently only a few laboratories are able to handle the pipette method or the "in situ" technique for rapid karyotyping. 2. Fetal blood sampling (cordocentesis); and subsequent chromosome analysis on cultivated fetal lymphocytes leading to results within 2 to 4 days. The main advantage of this procedure is the reliability of the results obtained. Fetal blood sampling, however, is restricted to specialists; this may involve scheduling delays. 3. Placental biopsy; with subsequent direct preparation and long term culturing. In comparison to both other techniques this procedure is faster if direct preparation is used. Results can be obtained even on the same day. The main disadvantage, however, is the problem with the reliability of the direct preparation results. They must be confirmed by time-consuming long-term culturing. Data are presented on the likelihood of abnormal ultrasound findings being caused by chromosomal aberrations.(ABSTRACT TRUNCATED AT 250 WORDS)

In utero diagnosis of congenital infections by direct fetal sampling.

Counseling for infections during pregnancy has traditionally focused on the clinical and laboratory findings of infection in the mother and the estimated risk of fetal damage associated with possible transmission of infection to the child. Now, with the use of techniques for fetal sampling, it is possible to diagnose infections of the fetus in utero and to correlate that information with the occurrence of fetal damage. The techniques that are available for sampling include amniocentesis, cordocentesis and chorionic villus sampling. The laboratory tests include: a) isolation of the organism in appropriate laboratory systems; b) detection of the DNA or RNA of the organism directly or with amplification with techniques such as PCR; c) detection of the organism by fluorescence or in situ hybridization; and d) identification of IgM or IgA fetal antibody to the organism by ELISA or similar methods. In utero infections can be documented for agents such as rubella, cytomegalovirus, parvovirus, Toxoplasma gondii and human immunodeficiency virus type 1. Further information is needed concerning the sensitivities and specificities of these methods for identifying fetal infection and predicting fetal damage.

Guidelines for blood sampling and measurement of pH and blood gas values in obstetrics. Based upon a workshop held in Zurich, Switzerland, March 19, 1993 by an Ad Hoc Committee.

Guidelines for the clinical indications for measuring pH and blood gas values in fetal blood, the procedures of blood sampling and measurement and some reference values for the evaluation of the data are given. They cover: prenatal sampling of blood from the umbilical vessels in conjunction with cordocentesis, intra partum sampling of fetal capillary blood by skin puncture of the presenting part, post partum sampling of blood from a clamped section of the umbilical cord and general analytical techniques.

Gaining access to the embryonic-fetal circulation via first-trimester endoscopy: a step into the future.

OBJECTIVE: To assess the feasibility of gaining access to the embryonic-fetal circulation via first-trimester transcervical embryoscopy. METHODS: A fiberoptic endoscope with a 3.5-mm, wide-angle lens was passed transcervically through the chorion and into the exocoelomic cavity of women undergoing pregnancy termination. A 26-gauge heparinized needle was passed through the sideport of the endoscope and inserted into the blood vessels of the chorionic plate or umbilical cord to obtain blood samples. RESULTS: With a modified endoscope, we have been able to gain access into the embryonic-fetal circulation and obtain a small aliquot of blood in five of eight cases. CONCLUSIONS: Our experience establishes the feasibility of gaining access to the human embryonic-fetal circulation. This work is expected to serve as a basis for further studies of the diagnosis and treatment of congenital diseases in early pregnancy.