Methylation status of imprinted genes 1 DLK1 GTL2 MEST PEG1 ZAC PLAGL1 and LINE 1 elements in spermatozoa of normozoospermic men unlike H19 ICR is not associated with idiopathic recurrent spontaneous miscarriages

Spermatozoa of normozoospermic men in idiopathic recurrrent spontaneous miscarriage do not reveal aberrations in DNA methylation at intergenic differentially methylated regions, MEST, ZAC, and LINE-1 CpG islands.

Mandar Ankolkar, M.Sc., Vinita Salvi, M.D., Himangi Warke, M.D., Babu Rao Vundinti, Ph.D., N. H. Balasinor, Ph.D.

Volume 99, Issue 6, Pages 1668-1673.e2, May 2013


To study methylation aberrations 1 in spermatozoa at developmentally important imprinted regions to ascertain their role in early embryo loss in idiopathic Recurrent Spontaneous Miscarriages (RSM).

Case-Control Study.

Academic research setting at National Institute for Research in Reproductive Health, Parel, Mumbai.

Male partners of couples with a history of RSM and male partners of couples with proven fertility (control group).


Main Outcome Measure(s):
DNA methylation levels at imprinting control Regions of DLK1- GTL2, MEST(PEG1), ZAC(PLAGL1) by EpiTYPER MassARRAY and global methylation levels as measured by LINE-1 methylation and anti 5-methyl cytosine antibody in spermatozoa of 23 from Control group and 23 men from RSM group.

We did not observe any aberration in the total methylation levels in any of the imprinted genes or global methylation analyzed.

Our results indicate that paternal methylation aberrations at imprinting control regions of DLK1-GTL2, MEST (PEG1), and ZAC (PLAGL1) and global methylation levels are not associated with idiopathic RSM and may not be good epigenetic markers (unlike the H-19 imprinting control region) for diagnosis of idiopathic RSM.

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  • Jose Medrano


    First, I´d like to congratulate the authors for such a good work. I think that in this type of clinical studies it is important that researchers report their findings even if they didn´t see significant differences among compared groups.

    I´d also like to ask the authors what do they think about the use of the methylated DNA quantification kit (Sigma) that they used compared with the Infinium HumanMethylation450 BeadChip (Illumina). Since in my group we are also interested in the study of global methylation patterns, author´s comments would be more than welcome.


    • N.H. Balasinor

      Thank you very much for your interest and appreciation for our work. The global methylation estimation (Sigma) is an antibody based method, i.e. ELISA, which uses anti 5-methyl cytosine antibody. Whereas, the Illumina Human 450 bead chip is a sequence based method that will give global methylation at a much higher resolution. However, it is an expensive method, perhaps time consuming as compared to the ELISA and gives much more data than required. The sigma kit can be used in cases where a quick inexpensive and a rough estimate of the global methylation level is required.
      Hope we have satisfactorily obliged to your query. Plz feel free to ask if there is anything else.

      • Jose Medrano

        Thank you so much for the response. We´ll take care of your comments before deciding which methodology to use in our research.

        Best regards.

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