Conflicting results of prenatal FISH with different probes for Down's Syndrome critical regions associated with mosaicism for a de novo del(21)(q22) characterised by molecular karyotyping: Case report

For the rapid detection of common aneuploidies either PCR or Fluorescence in situ hybridisation (FISH) on uncultured amniotic fluid cells are widely used. There are different commercial suppliers providing FISH assays for the detection of trisomies affecting the Down's syndrome critical regions (DSCR) in 21q22. We present a case in which rapid FISH screening with different commercial probes for the DSCR yielded conflicting results. Chromosome analysis revealed a deletion of one chromosome 21 in q22 which explained the findings. Prenatally an additional small supernumerary marker chromosome (sSMC) was discovered as well, which could not be characterised. Postnatal chromosome analysis in lymphocytes of the infant revealed complex mosaicism with four cell lines. By arrayCGH the sSMC was provisionally described as derivative chromosome 21 which was confirmed by targeted FISH experiments

DNA-Methylation Profiling of Fetal Tissues Reveals Marked Epigenetic Differences between Chorionic and Amniotic Samples

Epigenetic mechanisms including DNA methylation are supposed to play a key role in fetal development. Here we have investigated fetal DNA-methylation levels of 27,578 CpG loci in 47 chorionic villi (CVS) and 16 amniotic cell (AC) samples. Methylation levels differed significantly between karyotypically normal AC and CVS for 2,014 genes. AC showed more extreme DNA-methylation levels of these genes than CVS and the differentially methylated genes are significantly enriched for processes characteristic for the different cell types sampled. Furthermore, we identified 404 genes differentially methylated in CVS with trisomy 21. These genes were significantly enriched for high CG dinucleotid (CpG) content and developmental processes associated with Down syndrome. Our study points to major tissue-specific differences of fetal DNA-methylation and gives rise to the hypothesis that part of the Down syndrome phenotype is epigenetically programmed in the first trimester of pregnancy

DNA methylation patterns in CVS samples with chromosomal imbalances.

<p>PCA (A) and Hierarchic cluster analysis (B) of DNA methylation values of 464 CpG loci differentially methylated between chorionic villi biopsy samples without chromosomal imbalances (red spheres or red squares, respectively) and samples with trisomy 21 (47,XY+21; blue spheres or blue squares, respectively) (FDR<0.01). While in cases with trisomy 21 numerous epigenetic alterations could be identified, in cases with trisomy 18 (47,XX+21/47,XY+21, yellow spheres or yellow squares) only 15 differentially methylated loci have been found (C and D). Green color in the heatmap indicates low DNA-methylation, black intermediate and red high DNA-methylation values.</p

DNA methylation differs between AC and CVS samples.

<p>(A) Principal component analysis (PCA) of 22,234 CpG loci which passed quality tests and entered the analysis. Unsupervised analysis separated AC (green spheres) from CVS samples (red spheres). (B) Differential methylation analysis to separate AC from CVS samples (t-test, FDR<1×10⁻¹⁵). (C) Hierarchic cluster analysis of the 2418 CpG loci shown in (B). The upper panel on top of the heatmap indicates the sample type (green: AC, red:CVS) while the second panel indicates the fetuses’ sex (pink: female, cyan: male). Green color in the heatmap indicates low DNA-methylation, black intermediate and red high DNA-methylation values. Samples with known chromosomal aberrations and samples from ICSI were excluded. AC and CVS differ significantly in their DNA methylation pattern.</p

Hierarchic cluster analysis of DNA methylation values of 767 X-chromosomal CpG loci present on the array.

<p>The upper panel on top of the heatmap indicates the sample type (green: AC, red: CVS) while the second panel indicates the fetuses’ sex (pink: female, cyan: male). Samples with known chromosomal aberrations and samples from ICSI were excluded. Three CVS samples from male fetuses show a DNA methylation pattern similar to female samples (arrow). A blue bar at the right site indicates genes methylated specifically in male fetuses, an orange bar genes methylated in female samples and a black bar genes with tissue- specific DNA methylation.</p