Journal
EPIGENETICS
Volume 6, Issue 3, Pages 368-379Publisher
TAYLOR & FRANCIS INC
DOI: 10.4161/epi.6.3.14196
Keywords
cytotrophoblast purification; placental fibroblast purification; DNA methylation; epigenetics; placenta; cell type-specific methylation
Funding
- Canadian Institutes of Health Research [MOP86758, MOP220231]
- POPH (Operational Program for Human Potential) [SFRH/BD/28642/2006]
- MCTES (Ministry of Science, Technology and Higher Education)
- Fundação para a Ciência e a Tecnologia [SFRH/BD/28642/2006] Funding Source: FCT
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Epigenetic processes, such as DNA methylation, are known to regulate tissue specific gene expression. We explored this concept in the placenta to define whether DNA methylation is cell-type specific. Cytotrophoblasts and fibroblasts were isolated from normal midtrimester placentas. Using immunocytochemistry, we demonstrated 95% purity for cytotrophoblasts and 60-70% for fibroblasts. We compared DNA methylation profiles from cytotrophoblasts, fibroblasts and whole placental villi using bisulfite modified genomic DNA hybridized to the Illumina Methylation27 array. Euclidean cluster analysis of the DNA methylation profiles showed two main clusters, one containing cytotrophoblasts and placenta, the other fibroblasts. Differential methylation analysis identified 442 autosomal CpG sites that differed between cytotrophoblasts and fibroblasts, 315 between placenta and fibroblasts and 61 between placenta and cytotrophoblasts. Three candidate methylation differences were validated by targeted pyrosequencing assays. Pyrosequencing assays were developed for CpG sites less methylated in cytotrophoblasts than fibroblasts mapping to the promoter region of the beta subunit of human chorionic gonadotropin 5 (CGB5), as well as two CpG sites mapping to each of two tumor suppressor genes. Our data suggest that epigenetic regulation of gene expression is likely to be a key factor in the functional specificity of cytotrophoblasts. These data are proof of principle for cell-type specific epigenetic regulation in placenta and demonstrate that the methylation profile of placenta is mainly driven by cytotrophoblasts.
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