Robert Saura

Nutritional and genetic determinants of vitamin B and homocysteine metabolisms in neural tube defects: A multicenter case–control study†

Neural tube defects (NTDs) are severe congenital malformations due to failure of neural tube formation in early pregnancy. The proof that folic acid prevents NTDs raises the question of whether other parts of homocysteine (Hcy) metabolism may affect rates of NTDs. This French case‐control study covered: 77 women aged 17–42 years sampled prior to elective abortion for a severe NTDs (cases) and 61 women aged 20–43 years with a normal pregnancy. Plasma and erythrocyte folate, plasma B6, B12 and Hcy were tested as five polymorphisms MTHFR 677 C → T, MTHFR 1298 A → C, MTR 2756 A → G, MTTR 66 A → G and TCN2 776 C → G. Cases had significantly lower erythrocyte folate, plasma folate, B12 and B6 concentrations than the controls, and higher Hcy concentration. The odds ratio was 2.15 (95% CI: 1.00–4.59) for women with the MTRR 66 A → G allele and it was decreased for mothers carrying the MTHFR 1298 A → C allele. In multivariate analysis, only the erythrocyte folate concentration (P = 0.005) and plasma B6 concentration (P = 0.020) were predictors. Red cell folate is the main determinant of NTDs in France. Folic acid supplement or flour fortification would prevent most cases. Increased consumption of vitamins B12 and B6 could contribute to the prevention of NTDs. Genetic polymorphisms played only a small role. Until folic acid fortification becomes mandatory, all women of reproductive age should consume folic acid in a multivitamin that also contains B12 and B6.

Confined placental mosaicism and pregnancy outcome: a distinction needs to be made between types 2 and 3.

To study the influence of types 2 and 3 confined placental mosaicism (CPM) on pregnancy outcome.

Submicroscopic deletion of chromosome 16p13.3 in patients with Rubinstein-Taybi syndrome.

The Rubinstein-Taybi syndrome (RTS) is a well-defined entity characterized by growth and mental retardation, broad thumbs and halluces, and typical face. The RTS locus was assigned to 16p13.3, and interstitial submicroscopic deletions of this region (RT1 cosmid, D16S237) were initially identified in 25% of RTS patients. The gene for the human CREB binding protein, the transcriptional coactivator CBP, is included in the RT1 cosmid, and mutations in CBP have recently been identified in nondeleted RTS patients. We investigated 30 French patients with RTS. Among these patients, 3 had the RT1 microdeletion (frequency 10%). There is no obvious phenotypic difference between the patients with and without the RT1 deletion. The RT1 probe appears useful for confirmation of the diagnosis but is of little interest as a screening tool. By pooling data including the previous series and our current series, the cumulative frequency of the 16p13.3 microdeletion is 11.9% (19 in 159). This frequency of approximately 12% deleted patients appears more accurate than the 25% previously reported. Molecular investigations of CBP are in process in our series to clarify the cause of RTS.

Pregnancy Outcome and Prognosisin Fetuses with Increased First-TrimesterNuchal Translucency

Objective: One of the concerns of prenatal diagnosis is to find sensitive markers to screen for chromosome abnormalities, such as serum assays or nuchal translucency (NT). This study reports our experience with NT measurement during the first trimester of pregnancy. Materials: The study was performed prospectively on 252 fetuses with either NT ≧3 mm or cystic hygroma. Results: We observed 50 abnormal karyotypes, i.e. 19.8%. The incidence of chromosome abnormalities increased with increasing maternal age and increasing NT thickness. For the 202 fetuses with normal karyotypes, outcome was unfavourable in 32 cases: 23 elective terminations of pregnancy, 8 spontaneous abortions and 1 neonatal death. Outcome was favourable in 141 cases. Twenty-nine pregnancies were lost to follow-up. Conclusion: Measurement of NT at 12 weeks’ gestation seems to be a good marker for chromosome abnormalities. When the karyotype is normal, the pregnancy outcome remains correlated with the degree of NT thickness. The finding of NT >3 mm between 10 and 14 weeks’ gestation dictates rigorous ultrasound monitoring and caution when predicting pregnancy outcome.

Chromosome analysis of adenomatous polyps of the colon: Possible existence of two differently evolving cytogenetic groups

A chromosomal study of 42 colonic adenomatous polyps was performed using a technique of direct chromosome analysis derived from the prenatal procedure for diagnosing chromosomal alterations from chorionic villi sampling. Abnormal karyotypes were found in 22 cases. Trisomy 7, the most frequently found alteration, was found in 13 cases, followed by trisomy 13 (nine cases). Monosomy 18 was observed in two cases; in one of these, that of a polyp which had degenerated into an intra-mucosal adenocarcinoma, it was associated with 17p monosomy. Interestingly, these two types of alterations (trisomy 7 versus 18 and 17p monosomy) were not found together in the same lesion. This suggests that there could be two distinct chromosomal behaviors which might be related to the two cytogenetic groups described for colorectal adenocarcinoma. However, the respective frequencies of such cytogenetic groups varied inversely between adenomas and adenocarcinomas, thus suggesting that they evolve differently.

Donnai-Barrow syndrome: four additional patients.

In 1993, Donnai and Barrow reported a new syndrome in two sets of sibs and in an unrelated child, including diaphragmatic hernia, exomphalos, absent corpus callosum, hypertelorism, myopia, and sensorineural deafness. Since then, only four similar patients have been documented. We describe four additional patients, including two sibling pairs from healthy parents. This report firmly establishes this syndrome as a distinct clinical entity and provides further evidence for its previously postulated autosomal recessive inheritance.

Prenatal diagnosis using array-CGH: a French experience.

Array-CGH or Chromosomal Microarray Analysis (CMA) is increasingly used in prenatal diagnosis throughout the world. However, routine practices are very different among centers and countries, regarding CMA indications, design and resolution of microarrays, notification and interpretation of Copy Number Alterations (CNA). We present our data and experience from our Fetal Medicine Center on 224 prospective prenatal diagnoses. Our approach is practical, and aims to propose a strategy to offer Chromosomal Microarray Analysis (CMA) to selected fetuses and to help to interpret CNA. We hope that this publication could encourage development of CMA in centers that have not started yet this activity in prenatal routine, and could contribute to edict guidelines in this field.

Monozygotic twins concordant for blood karyotype, but phenotypically discordant: a case of "mosaic chimerism".

We report on 23 years old discordant monozygotic (MZ) twins, one with minor anomalies and mental delay, the other one being normal. Both had 46,XX,dup(11)(p12p15)/46,XX mosaicism in blood, with a similar proportion of abnormal cells (respectively, 16% and 17%). However, interphase fluorescence in situ hybridization (FISH) analysis performed on buccal smear and urinary sediment using specific probes located at the duplicated region showed that mosaicism was only present in the abnormal twin, with 68% abnormal cells. We hypothesize that the postzygotic chromosomal rearrangement may have occurred early in one embryo after the twinning event, and the blood mosaicism observed in both twins would have resulted from blood exchanges via placental anastomoses. This hypothesis of chimerism is strongly supported by twin‐to‐twin transfusion syndrome observed during fetal life of our twins. This case and those previously reported lead us to suggest that blood is particularly unsuitable for cytogenetic investigations of twins.

Reduced placental telomere length during pregnancies complicated by intrauterine growth restriction.

Objectives Recent studies have shown that telomere length was significantly reduced in placentas collected at delivery from pregnancies complicated by intrauterine growth restriction secondary to placental insufficiency. Placental telomere length measurement during ongoing pregnancies complicated by intrauterine growth restriction has never been reported. This was the main objective of our study. Methods In our center, late chorionic villus samplings were performed between 18 and 37 weeks of amenorrhea in 24 subjects with severe intrauterine growth restriction (cases) and in 28 subjects with other indications for prenatal diagnosis (controls). Placental insufficiency was assessed by histo-pathological examination. Relative measurement of telomere length was carried out prospectively by quantitative Fluorescent In Situ Hybridization using fluorescent Peptide Nucleic Acid probes on interphase nuclei obtained from long-term cultured villi and with an automated epifluorescent microscope. A quantitative Polymerase Chain Reaction technique was performed to confirm the quantitative Fluorescent In Situ Hybridization results. The number of copies of gene loci encoding the RNA template (hTERC) and the catalytic subunit (hTERT) of the enzyme complex telomerase were also estimated in these placentas by Fluorescent In Situ Hybridization. Results Mean fluorescence intensity of telomere probes estimated by quantitative Fluorescent In Situ Hybridization was significantly less for cases compared to controls (p<0.001). This result indicated that mean telomere length was significantly reduced in placentas during pregnancies complicated by intrauterine growth restriction. Reduced telomere length was confirmed by the quantitative Polymerase Chain Reaction technique. No copy number variation of the hTERC and hTERT loci was noticed for cases, or for controls. Conclusion This study clearly demonstrates a reduction of placental telomere length in ongoing pregnancies (from 18 to 37 weeks of amenorrhea) complicated by severe intrauterine growth restriction secondary to placental insufficiency.

A freehand ultrasonographically guided technique in transabdominal chorionic villus sampling in more than 24 000 consecutive cases

We read with a great deal of interest the article by Calda and Brestak about chorionic villus vacu-sampling using the Vacutainer system (Calda and Brestak, 2009). We take this opportunity to reply to the authors as the published technique and the results obtained were not in agreement with our own practical experience. Since 1983, in our prenatal diagnostic center we have carried out chorionic villus sampling on more than 24 000 women using first the transcervical route and then the transabdominal route. The transcervical route was used from 1983 to 1988, but we then abandoned this method because of a very high rate of miscarriages linked with the procedure (∼4%). Since 1988, we have carried out transabdominal chorionic villus sampling exclusively, using the same technique between the 12th and the 37th week of amenorrhea. The technique we use is based on that described by Smidt-Jensen and Hahnemann (1984). It consists of an extra-amniotic transabdominal chorionic villus sampling puncture, monitored with ultrasound, using a 20-gauge needle rinsed with heparin serum then linked to a 20mL syringe by a plastic extension tube (Saura et al., 1992). The rate of miscarriage when using the 20-gauge needle in our medical fetal center is the same as for amniocentesis (Brun et al., 2003). Our first point is that the 20-gauge needle we use for chorionic villus sampling causes no more pain than amniocentesis, and therefore has the advantage of not requiring a local anesthetic prior to the chorionic villus puncture, which is necessary with an 18-gauge needle (Calda and Brestak, 2009; Vandenbossche et al., 2007). Next, we stress the importance of the quantity of chorionic villi sampled. It is the quantity of chorionic villi fragments that will determine the culture time, and thus ultimately, the advantage of using the chorionic villus sampling method. The smaller this quantity, the longer it takes to obtain conventional karyotyping of the culture. It may take up to 15–20 days, or even longer, if the quantity sampled is less than 5 mg, as reported by the authors. In this case, the advantage to be obtained from chorionic villus sampling is very much reduced and the benefits of choriocentesis over amniocentesis become debatable. Thus, the threshold amount of chorionic villi that is sufficient, and which the authors set at 5 mg, seems to us to be completely inadequate. We believe that a large amount of chorionic material should be sampled (more than 15–20 mg) mainly in order to obtain the conventional karyotype after 6–8 days culture. In our opinion, it is essential to obtain about 15–20 mg of chorionic villi in order to: