Epigenetic reprogramming during male germ cell differentiation plays a crucial role in regulating genome functions at critical stages of development. DNA methylation profiles are mainly established and maintained by DNMT family members, whereas TET dioxygenases are essential for active DNA demethylation in the paternal pronuclei and in ESCs.The present study focused on the expression of TET1 3 enzymes during human spermatogenesis and in mature spermatozoa. Furthermore, it should be clarified whether aberrant expression of TET1 3, DNMT1 and DNMT3A mRNAs are associated with aberrant imprinting of the H19 gene, male subfertility and poor outcome of ICSI treatment.During normal human spermatogenesis TET1 3 mRNAs are present in the cytoplasm of pachytene spermatocytes from stage I to stage V, while TET1 3 proteins are successively expressed: TET2 is expressed in the cytoplasm of late pachytene spermatocytes of stage V, then TET1 can be observed in the nuclei of step 1 round spermatids at stage I, and TET3 in the nuclei of step 3 round spermatids at stage III. 5hmC appears only in step 5 elongated spermatids. All three TETs are still detectable at the mRNA and protein level in mature sperm cells in considerable amounts.TET1 3, DNMT1 and DNMT3A mRNA levels in spermatozoa are significantly reduced in subfertile patients. In addition, they are closely associated with male age and routine semen parameters. TET2 and TET3 mRNA levels in mature sperm cells are significantly associated with pregnancy and fertilization rate after ICSI treatment, respectively. Aberrant methylation of TET3, but not TET1 2, CpG-promoter might down-regulate TET3-mRNA transcription. Finally, DNA methylation reprogramming enzymes, especially DNMT3A, might be associated with the methylation pattern of the imprinted gene H19 in human mature spermatozoa.Our data provide first hints that TETs enzymes and DNA-demethylation are essential for male germ cell differentiation. In addition, TET1 3, DNMT1 and DNMT3A mRNA levels in human mature spermatozoa seems to be important for male fertility and success of ART treatment.
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