[Effects of obesity on global genome DNA methylation and gene imprinting in mouse spermatozoa].

OBJECTIVE: To investigate the influence of high fat diet-induced obesity (HFDIO) on the differentially methylated region (DMR) of the imprinted gene and global genome methylation of sperm DNA. METHODS: We performed bisulfite sequencing on the DMR of the imprinted gene and global genome methylation of sperm DNA in the mouse model of HFDIO. RESULTS: No statistically significant differences were found between the HFDIO model and normal control mice in MEG3-IG (93.73 vs 97.26%, P = 0.252), H19 (98.00 vs 97.83%, P = 0.920), IGF2 (97.34 vs 96.25%, P =0.166), IGF2R (1.43 vs 1.11%, P = 0.695), PEG3 (0.19 vs 0.38%, P = 0.537), MEST (0.23 vs 0.68%, P = 0.315), NNAT (0.31 vs 0.00%, P = 0.134), or SNRPN (1.88 vs 3.13%, P = 0.628). A total of 8 942 DMRs were detected across the sperm genome (P <0.05). Gene functional enrichment analysis indicated that the enriched terms with the largest numbers of genes were the metabolic process (n = 1 482), RNA synthesis (n = 779), and transcription (n = 767). CONCLUSIONS: The methylation level underwent no significant change in the DMRs of the imprinted genes from the mice with HFDIO, but the CG methylation of the genes involved in the metabolic process, RNA synthesis and transcription were significantly altered.

Preliminary investigation of the effects of an FCF inhibitor interference with Septin in the early stage embryos in mice.

The objective of the present study was to investigate the effects of for chlorfenuron (FCF) interference with the septin protein on early stage embryos in mice. The 1­cell embryos were collected and divided into an FCF interference group and a control group. The FCF interference group was cultured in FCF media and the control group was cultured in dimethyl sulphoxide media at 37˚C with 5% CO2 until the desired phase was achieved. Septin2 protein expression was detected using immunofluorescence and western blot analysis. Blastocyst α­tubulin was stained by immunofluorescence to observe the alterations in spindles and microtubules. The rate of early embryo development into blastocysts was significantly reduced following FCF treatment (P<0.05). In the control group, septin2 was observed with a confocal microscope; septin2 was expressed in embryos at all stages and mainly in the blastomeres from the 2­cell stage onwards, with the expression concentrated in the nuclei of the blastomeres as identified by strong fluorescence. In the FCF interference group, septin2 was weakly expressed in the nuclei of blastomeres at the 2­ and 4­cell stages, and in the granulated blastomeres at the 4­ and 8­cell stages. Expression was barely observed in and following the morula. Granulation was observed starting from the 4­ and 8­cell stages. Compared with the control group, the FCF interference group exhibited irregular microtubules, abnormal spindle morphology and disordered chromosome arrangement in the blastocysts. The septin2 protein was expressed throughout the early stage embryo from the 2­cell stage to the blastocyst and localized in the nuclei of blastomeres. When the septin protein experienced interference by the FCF inhibitor, septin2 protein expression was reduced, which simultaneously resulted in abnormal embryonic development, uneven cytoplasmic division, various sizes and a reduced number of blastomeres, granulation in the blastomeres, disordered blastocyst microtubule distribution, spindle shape alterations and an abnormality of chromosome arrangement.