Detection of Chromosome 21-encoded mRNA of Placental Origin in Maternal Plasma

Abstract

Background: mRNA of placental origin (i.e., human placental lactogen and ␤-human chorionic gonadotropin) has been demonstrated to be easily detectable in maternal plasma. We tested whether detection of chromosome 21-encoded mRNA of placental origin is possible in maternal plasma obtained during the first trimester. Methods: Plasma samples were obtained from pregnant women between weeks 9 -13 of pregnancy. RNA was isolated from 800 or 1600 L of plasma by silica-based affinity isolation and, after on-column DNase treatment, was subjected to two-step, one-tube reverse transcription-PCR with gene specific primers. Results: Three chromosome 21-encoded genes located within the Down syndrome critical region with overexpression in trisomy 21 placentas were screened for expression in early placental tissue to select their potential use for RNA based plasma screening. One of the chromosome 21-encoded genes (LOC90625) showed strong expression in first trimester placenta similar to CSH1 (human placental lactogen) and was selected for plasma analysis. The RNA isolation assay was validated with CSH1 mRNA, which could be detected in the plasma of all women tested in weeks 9 -13 of pregnancy. RNA from the chromosome 21-encoded, placentally expressed gene, LOC90625, was present in maternal first-trimester plasma and could be detected in 60% of maternal plasma samples when 800 L of plasma was used and in 100% of samples when 1600 L of plasma was used. Conclusion: The detection of chromosome 21-encoded mRNA of placental origin in maternal plasma during the first trimester may allow development of plasma-RNA-based strategies for prenatal prediction of Down

Noninvasive prenatal diagnosis of fetal chromosomal aneuploidy by massively parallel genomic sequencing of DNA in maternal plasma

“…Recently, a number of approaches have been developed. One strategy targets a fetal-specific subset of nucleic acids in maternal plasma, e.g., placental mRNA (9)(10)(11) and DNA molecules bearing a placental-specific DNA methylation signature (12)(13)(14). The fetal chromosomal dosage is then assessed by allelic ratio analysis of SNPs within the targeted molecules.…”

Non Invasive Prenatal Diagnosis of Down Syndrome

“…The main drawback of this RNA-SNP allelic ratio approach is that it relies upon the fetus inheriting two different SNP alleles in a region which is transcribed into m RNA and therefore only fetuses heterozygous for the analysed SNP can be successfully diagnosed. Another candidate gene for this purpose is LOC90625 within the Down syndrome critical region that is over expressed in trisomy 21 placentas even from the first www.intechopen.com trimester [Oudejans et al, 2003]. RNA from this chromosome was found to be present in 60-100% of maternal samples depending on the volume of plasma sample analysed leading to the conclusion that the detection of encoded m RNA could be used in NIPD.…”

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“…Several papers published in recent years have shown that cell-free fetal mRNA (cff RNA) also circulates in the maternal plasma, and is present in a relatively stable form within apoptotic bodies (Ng et al, , 2003aTsui et al, 2002;Oudejans et al, 2003). The mRNA transcripts of two genes expressed in the placenta, human placental lactogen (hPL) and human chorionic gonadotrophin (βhCG) have been detected and quantified throughout gestation in the maternal circulation .…”