Expression And Significance Of DNMT3A In Ectopic Endometrial Cells From Women With Adenomyosis

Objective:To test the expression of DNMT3A in ectopic endometrium and ectopic endometrial stromal cells from women with adenomyosis compared with endometrium and endometrial stromal cells from women without adenomyosis; to construct transgenic cells with DNMT3A overexpression lentivirus, to investigate the relationship between DNMT3A and the pathogenesis of adenomyosis.Methods:Semi-quantitative RT-PCR and Western-blot were used to detect the expression of DNMT3A mRNA and protein in ectopic endometrium from women with adenomyosis(n=20) and endometrium from women without adenomyosis (n=20); the ectopic endometrial stromal cells and nomal endometrial stromal cells were isolated, purified, cultured, and then analyzed by reverse transcriptase PCR (RT-PCR) and Western blot, respectively. The DNMT3A overexpression lentivirus were constructed by biotechnology company, then infecting the ectopic endometrial stromal cells, Western-blot was used to detect Flag-tagged proteins to determine the infection efficiency.Result:We found that mRNA and protein levels of DNMT3A were strikingly lower in ectopic endometrium compared with nomal endometrium. The relative abundance of DNMT3A mRNA to β-actin mRNA in ectopic endometrium and nomal endometrium were0.865±0.44and1.498±0.96, respectively. The relative abundance of DNMT3A protein to P-actin protein in ectopic endometrium and nomal endometrium were0.145±0.076and0.726±0.326, respectively (P<0.05), these results presented statistical significance. Comparing with nomal endometrial stromal cells, the fold changes of DNMT3A mRNA and protein in ectopic endometrial stromal cells were0.57and0.38, respectively. The DNMT3A-Flag fusion protein was detected in the transgenic cells, which means successful infection.Conclusion:The aberrantly low expression of DNMT3A in ectopic endometrium and ectopic endometrial stromal cells compared with nomal endometrium and endometrial stromal cells may be associated with the pathogenesis of adenomyosis, implying that adenomyosis might be an epigenetic disease. Transgenic cells with DNMT3A overexpression were constructed successfully, providing the bases to continue to study the roles of DNMT3A in stromal cells of adenomyosis.