Ultrasonographic measurement of the transcerebellar diameter for gestational age estimation in the third trimester.

BACKGROUND: Ultrasonographic estimation of gestational age is done routinely by using fetal biometric parameters, such as the biparietal diameter (BPD), femur length (FL), abdominal circumference (AC), and head circumference (HC). Gestational dating by ultrasound is reliable in the 1st trimester, but discrepancies increase to more than 3 weeks in the 3rd trimester. AIMS AND OBJECTIVES: To study the accuracy of gestational age estimation by the transcerebellar diameter (TCD) in the 3rd trimester of singleton pregnancies whose gestational age is known. MATERIALS AND METHODS: A prospective study was carried out in the Department of Obstetrics and Gynecology at NDMC Medical College and Hindu Rao Hospital, Delhi, India, from October 2017 to May 2019 after obtaining clearance from the ethical committee. The study included 100 women aged 18-35 years with singleton pregnancies in the 3rd trimester with a gestational age ≥ 28 weeks; the women provided informed consent and filled in an F-form to rule out any sex determination of the baby. Only those women who had regular menstrual cycles 6 months before conception, who were sure of their last menstrual period (LMP), who had a gestational dating scan done up to 14 weeks, and for whom congenital malformation of the fetus had been ruled out by mid-trimester ultrasound were included. Gestational age was obtained for the measured BPD, HC, AC, FL, and TCD. A paired t-test was used for the comparison between the period of gestation (POG) by the 1st ultrasonography (USG) and that by the AC, BPD, FL, HC, and TCD. RESULTS: The estimation of gestational age by the BPD becomes gradually less reliable after 32 weeks of pregnancy, that by the HC and FL after 36 weeks, and that by the AC after 28 weeks. Regression analysis was used to find a correlation of the various parameters, such as the POG by the 1st USG, BPD, HC, AC, FL, and TCD, with the POG by LMP. The correlation coefficient (r), coefficient of determination (R2), and p values were calculated. Of all the sonographic parameters for the estimation of gestational age, the TCD showed the highest correlation (r = 0.979; p < 0.0001) and the AC the lowest correlation. CONCLUSION: The mean difference between the estimate of gestational age by the TCD and that by the 1st trimester USG increased from ± 1 day at weeks 28-32 to ± 1-2 days at weeks 32-36 and further increased to ± 6 days after 36 weeks of pregnancy, which was statistically significant. This indicated that the TCD could estimate gestational age within 6 days of near-term pregnancy. Thus, the TCD was a reliable parameter for the estimation of gestational age within 6 days of near-term pregnancy along with routine biometry and an alternative parameter for the evaluation of gestational age when the LMP is not known.

Fetal-specific hypermethylated RASSF1A quantification in pregnancy.

OBJECTIVE: To quantify cell free fetal DNA (cffDNA) with fetal specific epigenetic marker, hypermethylated RASSF1A, in maternal plasma of normal pregnant women from 20 weeks of gestation and to assess its relationship with maternal age, height, pre-pregnancy weight and body mass index (BMI). METHODS: Hundred normal pregnant women within the gestational age of 21-40 weeks were randomly selected and grouped into five (n = 20). Group 1: 21-24, Group 2: 25-28, Group 3: 29-32, Group 4: 33-36 and Group 5: 37-40 weeks. Maternal plasma DNA was extracted, digested with methylation-sensitive restriction enzyme, BstUI and the fetal specific DNA (cffDNA) was quantified by Real-time polymerase chain reaction (qRT-PCR). RESULTS: The mean hypermethylated RASSF1A concentrations in different gestational groups were Group 1: 30.1 ± 14.9, Group 2: 52.6 ± 22.18, Group 3: 93.2 ± 19.08, Group 4: 172.8 ± 26.81 and Group 5: 337.8 ± 52.9 copies/ml. Pearson's correlation analysis showed highly significant positive correlation between cffDNA and gestational age (r = 0.899, p < 0.001). BMI was also found to be positively related to cffDNA (r = 0.217, p < 0.05). However, it did not show any correlation with maternal age, height and pre-pregnancy weight. CONCLUSIONS: The gestational age-dependent increase of hypermethylated RASSF1A; the fetal specific epigenetic marker in maternal plasma was demonstrated, in an Indian study group of normal pregnant women. Findings would form the basis of future studies involving pregnancy complications that would aid in the early diagnosis of placental pathologies with hypermethylated RASSF1A.

Evaluation of fetal hypermethylated RASSF1A in pre-eclampsia and its relationship with placental protein-13, pregnancy associated plasma protein-A and urine protein.

OBJECTIVES: Cell free fetal DNA (cffDNA) and its hypermethylated RASSF1A gene signify a recent advancement in non-invasive prenatal diagnosis of feto-placental anomalies like pre-eclampsia. The study uses hypermethylated RASSF1A gene to quantify cffDNA and to assess its relationship with placental and urine proteins in pre-eclampsia cases. DESIGN AND METHODS: DNA was isolated from plasma samples of clinically diagnosed cases of pre-eclampsia (n=103) and normal pregnancy (n=616) from 21weeks of gestation. Through methylation sensitive enzyme (BstUI) digestion; followed by real time-polymerase chain reaction (RT-PCR), quantification of hypermethylated RASSF1A was done. Immunoassays determined: placental protein-13 (pp-13) and pregnancy associated plasma protein A (PAPP-A) and pyrogallol red molybdate assay for 24h urine protein. RESULTS: Highly significant differences between control and pre-eclampsia cases for hypermethylated RASSF1A concentrations were found; Group I: 33±7.35 vs 74.46±16.71, Group II: 53.75±16.65 vs 244.22±35.68, Group III: 93.25±19.08 vs 412.31±80.18, Group IV: 144.30±18.13 vs 1056.89±153.78, Group V: 307.55±40.76 vs 2763.76±259.76copies/ml. Multivariate Pearson's correlation analysis of hypermethylated RASSF1A with pp-13, PAPP-A and urine proteins showed positive and very highly significant (P<0.001) associations. CONCLUSIONS: Diagnostic potential of fetal specific, hypermethylated RASSF1A was evaluated. Its positive relationship with placental and urine proteins submit the case for considering it as a reliable marker for pre-eclampsia.