Detection of the placental epigenetic signature of the maspin gene in maternal plasma.

The discovery of fetal DNA in the plasma of pregnant women has opened up new approaches for noninvasive prenatal diagnosis and monitoring. Up to now, the lack of a fetal DNA marker that can be universally detected in maternal plasma has limited the clinical application of this technology. We hypothesized that epigenetic differences between the placenta and maternal blood cells could be used for developing such a marker. By using bisulfite DNA sequencing, the methylation status of the maspin gene promoter in placental tissues and paired maternal blood cells from pregnant women was analyzed. The maspin gene promoter was found to be hypomethylated in placental tissues and densely methylated in maternal blood cells. Genotyping of a single nucleotide polymorphism within the unmethylated maspin sequences in maternal plasma demonstrated that these sequences were derived from the fetus. By using real-time quantitative methylation-specific PCR, unmethylated maspin sequences were detected in maternal plasma in all three trimesters of pregnancy and were cleared within 24 h after delivery. The maternal plasma concentration of unmethylated maspin sequences was elevated by a median of 5.7 times in preeclamptic pregnancies compared with nonpreeclamptic pregnancies. Hypomethylated maspin DNA is the first universal marker for fetal DNA in maternal plasma, thus allowing the measurement of fetal DNA concentrations in pregnancy-associated disorders, irrespective of fetal gender and genetic polymorphisms. Differential DNA methylation between the placenta and maternal blood cells may be exploited to develop further markers for noninvasive prenatal assessment.

[1]  D. Bianchi,et al.  Detection of male and female fetal DNA in maternal plasma by multiplex fluorescent polymerase chain reaction amplification of short tandem repeats , 2000, Human Genetics.

[2]  P. Johnson,et al.  Presence of filterable and nonfilterable mRNA in the plasma of cancer patients and healthy individuals. , 2002, Clinical chemistry.

[3]  L. Poon,et al.  Clinical Chemistry 47:9 1607–1613 (2001) Molecular Diagnostics and Genetics Effects of Blood-Processing Protocols on Fetal and Total DNA Quantification in Maternal Plasma , 2001 .

[4]  Y. Lo,et al.  Increased maternal plasma fetal DNA concentrations in women who eventually develop preeclampsia. , 2001, Clinical chemistry.

[5]  N. Niikawa,et al.  Detection of cell free placental DNA in maternal plasma: direct evidence from three cases of confined placental mosaicism , 2004, Journal of Medical Genetics.

[6]  M. Hendrix,et al.  The tumour suppressor gene maspin is differentially regulated in cytotrophoblasts during human placental development. , 2002, Placenta.

[7]  R. Chiu,et al.  mRNA of placental origin is readily detectable in maternal plasma , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[8]  L. E. McDonald,et al.  A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[9]  J. Herman,et al.  Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[10]  R. Chiu,et al.  Prenatal exclusion of β thalassaemia major by examination of maternal plasma , 2002, The Lancet.

[11]  A. Benachi,et al.  New strategy for prenatal diagnosis of X-linked disorders. , 2002, The New England journal of medicine.

[12]  L. Poon,et al.  Differential DNA methylation between fetus and mother as a strategy for detecting fetal DNA in maternal plasma. , 2002, Clinical chemistry.

[13]  J. Costello,et al.  Methylation matters: a new spin on maspin , 2002, Nature Genetics.

[14]  C R Cantor,et al.  Chip-based genotyping by mass spectrometry. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[15]  Hong Duan,et al.  Role for DNA methylation in the control of cell type–specific maspin expression , 2002, Nature Genetics.

[16]  T K Lau,et al.  Quantitative analysis of fetal DNA in maternal plasma and serum: implications for noninvasive prenatal diagnosis. , 1998, American journal of human genetics.

[17]  J. M. Costa,et al.  Circulating cell-free fetal DNA in maternal serum appears to originate from cyto- and syncytio-trophoblastic cells. Case report. , 2004, Human reproduction.

[18]  I. Sargent,et al.  Quantitative abnormalities of fetal DNA in maternal serum in preeclampsia. , 1999, Clinical chemistry.

[19]  Y. Lo,et al.  Quantitative analysis of aberrant p16 methylation using real-time quantitative methylation-specific polymerase chain reaction. , 1999, Cancer research.

[20]  J. Herman,et al.  Gene silencing in cancer in association with promoter hypermethylation. , 2003, The New England journal of medicine.

[21]  I. Sargent,et al.  Presence of fetal DNA in maternal plasma and serum , 1997, The Lancet.

[22]  W. Holzgreve,et al.  Fetal DNA in maternal plasma is elevated in pregnancies with aneuploid fetuses , 2000, Prenatal diagnosis.

[23]  N. M. Hjelm,et al.  Maternal plasma fetal DNA as a marker for preterm labour , 1998, The Lancet.

[24]  C. Lam,et al.  Predominant hematopoietic origin of cell-free DNA in plasma and serum after sex-mismatched bone marrow transplantation. , 2002, Clinical chemistry.

[25]  E. Schisterman,et al.  Two-stage elevation of cell-free fetal DNA in maternal sera before onset of preeclampsia. , 2004, American journal of obstetrics and gynecology.

[26]  N M Hjelm,et al.  Prenatal diagnosis of fetal RhD status by molecular analysis of maternal plasma. , 1998, The New England journal of medicine.

[27]  Y. Lo,et al.  Increased fetal DNA concentrations in the plasma of pregnant women carrying fetuses with trisomy 21. , 1999, Clinical chemistry.