Profiling the DNA methylation patterns of imprinted genes in abnormal semen samples by next-generation bisulfite sequencing



Changes in DNA methylation modifications have been associated with male infertility. With the development of assisted reproductive technologies (ARTs), abnormal DNA methylation in sperm, especially in imprinted genes, may impact the health of offspring and requires an in-depth study.


In this study, we collected abnormal human semen samples, including asthenospermic, oligospermic, oligoasthenospermic and deformed sperm, and investigated the methylation of imprinted genes by reduced representation bisulfite sequencing (RRBS) and bisulfite amplicon sequencing on the Illumina platform.


The differentially methylated regions (DMRs) of imprinted genes, including H19, GNAS, MEG8 and SNRPN, were different in the abnormal semen groups. MEG8 DMR methylation in the asthenospermic group was significantly increased. Furthermore, higher methylation levels of MEG8, GNAS and SNRPN DMR in the oligospermic and oligoasthenospermic groups and a decrease in the H19 DMR methylation level in the oligospermic group were observed. However, the methylation levels of these regions varied greatly among the different semen samples and among individual sperm within the same semen sample. The SNP rs2525883 genotype in the H19 DMR affected DNA methylation. Moreover, DNA methylation levels differed in the abnormal semen groups in the non-imprinted genomic regions, including repetitive sequence DNA transposons and long/short interspersed nuclear elements (LINEs and SINEs).


Our study established that imprinted gene DMRs, such as H19, GNAS, SNRPN and MEG8, were differentially methylated in the abnormal semen groups with obvious inter- and intra-sample heterogeneities. These results suggest that special attention needs to be paid to possible epigenetic risks during reproduction.

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This work was supported by the National Natural Science Foundation of China (grant numbers 81872103 and 81971917) and the Science and Technology Climbing Fund of SIPPR (grant numbers PD2017-2 and PD2017-4).

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All authors contributed to the study conception and design. The study was designed and supervised by Qihan Wu and Xin Wang. Sample collection and semen parameter analyses were performed by Yuhua Sun, Xing Feng and Xin Wang. Material preparation, data collection and analysis were performed by Wanhong He, Sufen Zhang, Minjie Xu, Jianfeng Dai, Xiaohua Ni and Qihan Wu. The first draft of the manuscript was written by Qihan Wu, and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

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Correspondence to Xin Wang or Qihan Wu.

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This study was approved by the Medical Ethics Committee of Shanghai Institute of Planned Parenthood Research. All procedures performed in studies involving human participants were in accordance with the ethical standards of NHFPC Ethical Review of Human Biomedical Research (2010) and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

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He, W., Sun, Υ., Zhang, S. et al. Profiling the DNA methylation patterns of imprinted genes in abnormal semen samples by next-generation bisulfite sequencing. J Assist Reprod Genet (2020).

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  • DNA methylation
  • Imprinted genes
  • Spermatozoa