OBJECTIVETo study the correlation of the DNA methylation status of the imprinted gene H19 imprinting control region (ICR) with oligozoospermia and asthenozoospermia.

METHODSWe eliminated chromosomal abnormality as the cause of male infertility in the subjects by karyotype analysis and detection of Y-chromosome microdeletions, and identified 18 cases of single factor-induced oligozoospermia (sperm concentration < 15 x 10(6)/ml) and 20 cases of single factor-induced asthenozoospermia (progressively motile sperm <32%) by computer-aided sperm analysis (CASA). Then we extracted genome-wide sperm DNA, treated it with bisul- fite, subjected the target gene fragments to PCR amplification and sequencing. Lastly, we analyzed the DNA methylation status of the target genes with BIQ Analyzer and processed the data using SPSS17.0.

RESULTSThe DNA methylation level of the H19 ICR was increased significantly in the oligozoospermia patients ([9.19 +/- 2.45]%, P < 0.05), especially in the severe oligozoospermia males with sperm concentration < 3 x 10(6)/ml (P < 0.01), as compared with that of the 20 fertile control men ([0.30 +/- 0.06]%). However, no significant differences were found in the level ([0.30 +/- 0.07]%) and pattern of the DNA methylation of the H19 ICR (P = 0.62). Further analysis of the DNA methylation status of the CTCF-6 binding sites indicated that the DNA methylation degree was significant higher in the oligozoospermia men ([2.67 +/- 0.75]%) than in the fertile control ([0.05 +/- 0.03]%) or the asthenozoospermia group ([0.03 +/- 0.02]%), with no significant differences between the latter two (P = 0.35).

CONCLUSIONThe reduced DNA methylation of the H19 ICR is negatively correlated with sperm concentration but not associated with sperm motility.