Otília Brandão

Gene expression pattern of IGF2, PHLDA2, PEG10 and CDKN1C imprinted genes in spontaneous miscarriages or fetal deaths.

Genomic imprinting is defined as an epigenetic modification that leads to parent-of-origin specific monoallelic expression. Some current research on the fetal control growth has been focused on the study of genes that display imprinted expression in utero. Four imprinted genes, two paternally expressed (IGF2 and PEG10) and two maternally expressed (PHLDA2 and CDKN1C), are well known to play a role in fetal growth and placental development. Pregnancy loss in the general reproductive population is a very common occurrence and other genetic causes beyond chromosomal abnormalities could be involved in spontaneous miscarriages or fetal deaths, such as alteration of expression in imprinted genes particularly those related to fetal or placental growth. Quantitative Real Time PCR was performed to evaluate gene expressions patterns of the four mentioned genes in spontaneous miscarriages or fetal deaths from 38 women. Expression levels of PHLDA2 gene were upregulated in the first trimester pregnancy cases and all four imprinted genes studied were upregulated in the second trimester of pregnancy cases comparing with controls. In third trimester PEG10 was downregulated in fetal samples group. This is the first study presenting data from human imprinted genes expression in spontaneous miscarriages or fetal deaths cases from the three trimesters of pregnancy.

An efficient protocol for the detection of chromosomal abnormalities in spontaneous miscarriages or foetal deaths

OBJECTIVE Characterization of chromosomal abnormalities in 232 spontaneous miscarriages or foetal deaths using both classical and molecular cytogenetics. STUDY DESIGN Chromosomal abnormalities are responsible for 40-50% of all early pregnancy losses. Conventional cytogenetics is associated with 10-40% of culture failure. Comparative genomic hybridization (CGH) is a DNA-based technique that screens chromosome imbalances in the whole genome and may overcome this problem, although additional methods are required to distinguish between different ploidies, mosaicisms and maternal cell contamination. For a full characterization of chromosomal aberrations in 232 spontaneous miscarriages or foetal deaths we applied a sequential protocol that uses conventional cytogenetics, plus CGH and touch fluorescence in situ hybridization (Touch FISH). RESULTS Successful karyotyping was obtained in 173/232 (74.6%) of the cases, 66/173 (38.2%) of which had an abnormal chromosomal complement. CGH and Touch FISH analyses revealed another 19 abnormal cases in the 63 failures of culture. Overall there were 85/233 (36.6%) cases with an abnormal chromosomal complement, with examples from all three trimesters. Comparing cases, with or without chromosomal abnormalities, no statistical differences were found between women with one or recurrent miscarriages. On the contrary, significant differences were found comparing mean maternal ages or mean gestational ages, in cases with or without chromosomes abnormalities. CONCLUSION Adopting this sequential protocol, chromosomal complement information was available even in cases with culture failure.

Early Antenatal Diagnosis of Cardiac Defects Using Transvaginal Doppler Ultrasound: New Perspectives?

Objective: Cardiac defects are the most prevalent congenital anomalies. Screening policies have adopted an 18- to 22-week ultrasound scan to detect such anomalies. However, diagnosis may be feasible early in pregnancy using transvaginal Doppler ultrasound. Methods: Transvaginal ultrasound, including nuchal translucency (NT) measurement, is routinely performed at 10–13 weeks of gestation at our department. Complementary arterial and venous blood flow Doppler evaluation is also performed in cases of increased NT as a part of an ongoing research project. Results: Ultrasound revealed complete atrioventricular septal defects in 2 trisomic fetuses (trisomy 18 and 21) at 12 and 13 weeks of gestation, respectively, with increased NT and abnormal venous blood flow velocity. Conclusions: Incrased NT has been associated with major chromosomal anomalies and is being increasingly related to cardiac defects. Considering that venous blood flow patterns may provide additional clues to the cardiac function, it may be useful as a complementary tool for the earlier diagnosis of structural cardiac anomalies.

Aneuploidies detection in miscarriages and fetal deaths using multiplex ligation-dependent probe amplification: an alternative for speeding up results?

OBJECTIVE The aim of this prospective study was to apply the MLPA technique to products of miscarriages and fetal deaths in order to detect the more frequent chromosome aneuploidies and compare the results to conventional karyotyping. STUDY DESIGN Multiplex ligation-dependent probe amplification (MLPA) is a relatively new molecular technique for targeted detection of common chromosomal aneuploidies, namely trisomy 13, 18, 21 and sex chromosomal abnormalities. The reliability and high accuracy of this technique constitute an alternative for rapid results in large scale testing. In this study, a total of 489 DNA samples from fetal tissue were used for aneuploidy detection of chromosomes 13, 18, 21, X and Y using a commercial MLPA kit (SALSA P095) and were simultaneously subjected to conventional karyotyping. RESULTS MLPA was the only result available in 33% of the cases. A cytogenetic result was obtained in only 328/489 samples. MLPA detected 7.8% of chromosome aneuploidies. Among the total samples karyotyped, MLPA failed to detect some aneuploidies and the false-negative rate was 0.82%. As expected, ploidy changes and reciprocal translocations were not detected by this technique, but MLPA gave a conclusive result even in cases of mosaicism. CONCLUSION The present data confirm that MLPA is a rapid, simple and reliable method for detection of chromosome 13, 18, 21, X and Y abnormalities in fetal tissue.

Nuchal translucency and ductus venosus blood flow as early sonographic markers of thanatophoric dysplasia. A case report.

Thanatophoric dysplasia (TD) is the most frequent form of lethal osteochondrodysplasias. Prenatal diagnosis is commonly accomplished in the second-trimester scan, but occasionally TD may not be clearly distinguished from the other osteochondrodysplasias, with consequent important prognostic implications. However, in order to confirm the diagnosis, complementary radiological, pathological and molecular studies are mandatory. We present a case of TD diagnosed in the late first trimester with the contribution of nuchal translucency combined with ductus venosus blood flow assessment.

Diagnosis and cause of death in a neonatal intensive care unit – How important is autopsy?

Objectives. To characterize mortality in a tertiary referral Neonatal Intensive Care Unit (NICU) in Portugal and evaluate the concordance between ante-mortem and post-mortem diagnoses. Methods. Retrospective review of the clinical and pathological records of infants who died in five consecutive years was done. Pathological findings and clinical diagnoses were compared and classified according to general concordance and to modified Goldman classification. Results. During the referred period, 1938 patients were admitted to the NICU, with a mortality rate of 5.7% (110 patients). The median of age at death was 10.5 days and the most frequent causes of death were congenital malformations and prematurity with its complications. Autopsy was performed in 53 patients resulting in a 48.2% overall autopsy rate. There was complete agreement between pathological and clinical diagnoses in 18 cases (34%) and additional findings were identified in 22 cases; in 13 cases (24.5%), the diagnosis was revised or established by pathology. Five autopsies revealed information relevant for genetic counseling. Conclusion. Despite the high agreement rate between clinical and pathological diagnoses, autopsy frequently added important data, including several cases in which it established the diagnosis or provided information relevant for parental counseling regarding future pregnancies.

Complete mole in a dichorionic twin pregnancy after intracytoplasmic sperm injection

A dichorionic twin pregnancy with complete hydatidiform mole and coexistent fetus is a rare and challenging situation, whose pathogenesis has not been yet fully understood. We present a case of a 39-year-old woman who underwent intracytoplasmic sperm injection with two embryos transfer. The 12-week gestation ultrasound examination revealed normal fetus and placenta with features of hydatidiform mole, leading to pregnancy termination. Autopsy and histological examinations diagnosed a complete mole coexisting with a normal fetus, and the genetic analysis showed a diploid fetus with biparental genome and molar tissue with paternal diploidy. This case highlighted that complete molar pregnancies may still occur even though pregnancy is achieved after intracytoplasmic sperm injection. A review of the literature was performed by collecting data from the few similar reported cases and by commenting on the pathogenesis of this rare condition.

A Novel Mutation in FOXF1 Gene Associated with Alveolar Capillary Dysplasia with Misalignment of Pulmonary Veins, Intestinal Malrotation and Annular Pancreas

Alveolar capillary dysplasia with misalignment of pulmonary veins (ACD/MPV) is a rare, fatal, neonatal developmental lung disorder, which usually presents as persistent pulmonary hypertension unresponsive to treatment. The authors report the case of a neonate with persistent pulmonary hypertension, associated with duodenal stenosis secondary to annular pancreas and intestinal malrotation. Support treatment, inhaled nitric oxide, oral sildenafil and nebulized iloprost were used with no clinical improvement. The neonate presented an overwhelming course, with hypoxemia refractory to treatment. At autopsy lung histology showed the characteristic features of ACD/MPV. DNA sequence analysis revealed a heterozygous nonsense mutation c.539C>A;p.S180X, in the first exon of FOXF1. FOXF1 has been identified as one of the genes responsible for ACD/MPV associated with multiple congenital malformations. This clinical case is the first report of a heterozygous nonsense mutation c.539C>A;p.S180X in the first exon of FOXF1, in a patient with ACD/MPV associated with annular pancreas and intestinal malrotation.

Revisiting Acrania: Same Phenotype, Different Aetiologies

Objective: To evaluate the contribution of prenatal and postmortem examinations in establishing the aetiology of acrania. Methods: Retrospective evaluation of 14 cases of acrania managed through elective termination of pregnancy. Results: The median maternal age was 30 years (range 18–40) and median gestational age at diagnosis was 13 weeks (range 12–15). One mother had epilepsy and was taking anticonvulsants and another had uncontrolled type II diabetes mellitus. Only 3 women were using folic acid at conception. Chromosomal abnormalities were detected in 3 of 8 cases analyzed. Unilateral anopthalmia, cervical rachischisis, midline facial and limb defects coexisted with acrania in 4 cases. Acrania with craniofacial dysmorphism and asymmetrical finger amputation were observed in a case of amniotic band syndrome. A previous history of anencephaly was documented in 1 case. Conclusion: Acrania is a characteristic phenotypic expression of a variety of different aetiologies. Investigation with cytogenetic studies and postmortem are essential to provide a definitive answer. This will provide a better understanding of the underlying aetiology and help establish the recurrence risk for future pregnancies.

Phenotypic Expression in the First Case of Complete Trisomy 12: Combination of Prenatal Ultrasound and Necropsic Examination

The authors report the first case of a complete trisomy 12 detected antenatally at 16 weeks of gestation. The ultrasonographic features (craniofacial abnormalities) correlated well with postmortem findings. The absence of the hypophysis gland, olfactory pathways and both adrenal glands, as well as developmental anomalies of the central nervous system, were additional findings at autopsy. The recognition of complete trisomy 12 phenotype can be helpful in the pre- and postnatal diagnosis of future similar cases and parental counseling.