Transcriptomic analysis of cell-free fetal RNA suggests a specific molecular phenotype in trisomy 18

Abstract

Background Trisomy 18 is a common human aneuploidy that is associated with significant perinatal mortality. Unlike the well-characterized “critical region” in trisomy 21 (21q22), there is no corresponding region on chromosome 18 associated with its pathogenesis. The high morbidity and mortality of affected individuals has limited extensive investigations. Objective To better understand the molecular mechanisms underlying the congenital anomalies observed in this condition, we investigated the in utero gene expression profile of second trimester fetuses affected with trisomy 18. Methods Total RNA was extracted from cell-free amniotic fluid supernatant from aneuploid fetuses and euploid controls matched for gestational age and hybridized to Affymetrix U133 Plus 2.0 arrays. Individual differentially-expressed transcripts were obtained by two-tailed t-tests. Over-represented functional pathways among these genes were identified with DAVID and Ingenuity® Pathways Analysis. Results Three hundred fifty-two probe sets representing 251 annotated genes were statistically significantly differentially expressed between trisomy 18 and controls. Only 7 genes (2.8% of the annotated total) were located on chromosome 18, including ROCK1, an up-regulated gene involved in valvuloseptal and endocardial cushion formation. Pathway analysis indicated disrupted function in ion transport, MHCII/T-cell mediated immunity, DNA repair, G-protein mediated signaling, kinases, and glycosylation. Significant down-regulation of genes involved in adrenal development was identified, which may explain both the abnormal maternal serum estriols and the pre and postnatal growth restriction in trisomy 18. Comparison of this gene set to one previously generated for trisomy 21 fetuses revealed only six overlapping differentially-regulated genes. Conclusion This study contributes novel information regarding functional developmental gene expression differences in fetuses with trisomy 18.

The Amniotic Fluid Transcriptome as a Guide to Understanding Fetal Disease

“…3). The RNA was extracted, amplified, labeled and hybridized to either the Affymetrix U133A expression microarray (Larrabee et al 2005) or the Affymetrix GeneChip Human Genome U133 Plus 2.0 expression microarray (Slonim et al 2009;Koide et al 2011;Hui et al 2012aHui et al ,b, 2013aEdlow et al 2014;Massingham et al 2014). Despite the availability of more sophisticated expression microarrays, the U133 Plus 2.0 was chosen because of the need for compatibility with an essential downstream in silico tool, the Connectivity Map (Lamb et al 2006).…”

“…In the studies of both DS and T18, differentially regulated genes included those that are thought to be expressed specifically from the central nervous system. This finding motivated a 2012 meta-analysis to mine previously published data (Slonim et al 2009;Koide et al 2011;Hui et al 2012a) with a specific focus on neural transcripts (Hui et al 2012b). Hui et al identified the 536 genes universally present in the cffRNA of 12 of 12 euploid samples, 746 genes universally present in the cffRNA of five of five fetuses with T18, and the 1184 genes universally present in the cffRNA of seven of seven fetuses with DS.…”

“…Mortality is most often linked to cardiac and renal malformations or to postnatal feeding difficulties resulting from central nervous system abnormalities. A 2011 study by Koide et al (2011) sought to compare the genome wide transcriptome of five female fetuses with T18 to six female euploid fetuses. Two hundred and fifty-one (251) genes showed statistically significant differential expression, seven of which were located on chromosome 18, and six of which overlapped with the 419 genes differentially regulated in DS (Table 3, Fig.…”

“…Pregnancy-specific beta-1-glycoprotein 3 223054_at DNAJB11 DnaJ (Hsp40) homolog, subfamily B, member 11 DS, Down syndrome (Slonim et al 2009); Ob, obesity (Edlow et al 2014); T18, Edwards syndrome (Koide et al 2011); TTTS, Twin -twin transfusion syndrome (Hui et al 2013b); Turner, Turner syndrome (Massingham et al 2014). pathways that resulted in well-characterized fetal morphological anomalies. In the studies of both DS and T18, differentially regulated genes included those that are thought to be expressed specifically from the central nervous system.…”

Other Body Fluids as Non-invasive Sources of Cell-Free DNA/RNA

“…Functional analysis of the trisomy 18 fetuses indicated abnormalities in ion transport, immunity, DNA repair and G protein signaling and dysregulation of adrenal development-associated pathways [22].…”