Quantitative methylation analysis of developmentally important imprinted genes in human stillbirths from ART and spontaneous conception

Abstract

OBJECTIVE: To evaluate the epigenetic risk related to assisted reproductive technology (ART), the methylation status of H19, KvDMR1, PEG1, and SNPRN genes in human stillbirths were assessed. DESIGN: An experimental study. MATERIALSANDMETHODS: After obtaining the approval of the Institutional Ethics Committee, 43 muscle samples (MS) were obtained: stillbirths from ART pregnancy (n1⁄413), natural pregnancy (n1⁄417), and induced aborted fetuses from natural pregnancy (n1⁄413). These patients had no known anatomic or genetic abnormalities. The DNAmethylation patterns of H19, KvDMR1, PEG1, and SNRPN were analyzed using pyrosequencing. RESULTS: Clear hypomethylation (0%) or hypermethylation (100%) was not detected in the four imprinted genes. The percentage methylation of KvDMR1 in stillbirths from natural pregnancy was significantly higher than that in the other two groups (P 0.05). Box plot analysis showed that 5 of 120 (4.2%) analyzed DMR methylation values (for the four studied genes) represented outliers in stillbirths, but none outliers were found in normal pregnancies. ROC curve analysis showed a positive correlation between themethylation percentage of KvDMR1 and PEG1 and pregnancy loss in second trimester (P 0.05). There’s no evidence for an increased rate of potentially abnormal methylation values in the ART group. CONCLUSION: Imprinting errors of imprinted genes may contribute to pregnancy loss in the second half of pregnancy, but ART procedures might not increase the occurrence of aberrant methylation patterns of imprinted genes. Supported by: The National Natural Science Foundation of China (81170574), National Key Basic Research Development Plan of China (973 Program) (2007CB948104).