Correlation of DNA methylation status of imprinted gene H19 ICR with oligozoospermia and asthenozoospermia / 中华男科学杂志

<p><b>OBJECTIVE</b>To study the correlation of the DNA methylation status of the imprinted gene H19 imprinting control region (ICR) with oligozoospermia and asthenozoospermia.</p><p><b>METHODS</b>We 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.</p><p><b>RESULTS</b>The 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).</p><p><b>CONCLUSION</b>The reduced DNA methylation of the H19 ICR is negatively correlated with sperm concentration but not associated with sperm motility.</p>

ICSI causes abnormal H3K9 methylation in the male pronuclei and growth retardation of mouse embryos / 中华男科学杂志

<p><b>OBJECTIVE</b>To evaluate the safety of intracytoplasmic sperm injection (ICSI) in the mouse model.</p><p><b>METHODS</b>We simulated clinical ICSI technology and comprehensively evaluated it by parthenogenetic activation, immunofluorescence, embryo transplantation, examination of early implantation, and measurement of the crown-rump length (CRL).</p><p><b>RESULTS</b>ICSI significantly reduced the ability of preimplantation embryo development of the mouse, especially after the 8-cell stage (P < 0.01). The fluorescence of H3K9 dimethylation was abnormal at the male pronuclei of the embryos derived from ICSI. Further examination of the development of the transferred ICSI embryos indicated no significant difference in the rate of early implantation at E5. 5 days as compared with normal fertilization (P = 0.6), but the percentage of "normal embryos" was decreased significantly at E9.5 days (P < 0.01). Obvious growth retardation phenotype was observed even in the normal ICSI embryos at E9.5 days.</p><p><b>CONCLUSION</b>ICSI might result in growth retardation of embryos by affecting H3K9 dimethylation in the male pronuclei.</p>