Model of transfection in human endometrial epithelium cells with hsa-miR-191-5p Model of transfection in human endometrial epithelium cells with hsa-miR-191-5p

Background: Human endometrial cells are important in blastocyst recognition and implantation. We have recently shown that miR-191-5p secreted into culture medium by human embryos cultured and transferred to woman on the fifth day of development was associated with the percentage of pregnant vs. non-pregnant patients. Little is known about the regulation and expression of endometrial miRNAs induced by embryonic miRNAs in endometrial tissue. Therefore, in the present work we explored the viability and transfection of RL95-2 endometrial cell line with agomiR-191. Results: The main results obtained in this study were: First, transfection of RL95-2 cell line with 100nM of lipofectamine in combination with 15, 30, and 60 nM of agomiR-191 for 3, 6 and 24 hours does not affect the viability of RL95-2 cells. Second, we observed expression of miR-191 with 60 pmol of agomiR-191 in a time dependent transfection. Conclusion:: Stimulation of RL95-2 endometrial cell line with lipofectamine does not modify their viability. The transfected RL95-2 endometrial cells showed increased the expression of miR-191.

Brain-Derived Neurotrophic Factor Regulates Ishikawa Cell Proliferation through the TrkB-ERK1/2 Signaling Pathway

(1) Background: Endometrial regulation is a necessary condition for maintaining normal uterine physiology, which is driven by many growth factors. Growth factors produced in the endometrium are thought to be related to the proliferation of endometrial cells induced by estradiol-17β (E2). In this study, we found that E2 can induce the secretion of brain-derived neurotrophic factor (BDNF) in Ishikawa cells (the cells of an endometrial cell line). Furthermore, Ishikawa cells were used in exploring the regulatory role of BDNF in endometrial cells and to clarify the potential mechanism. (2) Methods: Ishikawa cells were treated with different concentrations of BDNF (100, 200, 300, 400, and 500 ng/mL). The mRNA expression levels of various proliferation-related genes were detected through quantitative reverse transcription polymerase chain reaction, and the expression of various proliferation-related genes was detected by knocking out BDNF or inhibiting the binding of BDNF to its receptor TrkB. The expression levels of various proliferation-related genes were detected by performing Western blotting on the TrkB-ERK1/2 signaling pathway. (3) Results: Exogenous BDNF promoted the growth of the Ishikawa cells, but the knocking down of BDNF or the inhibition of TrkB reduced their growth. Meanwhile, BDNF enhanced cell viability and increased the expression of proliferation-related genes, including cyclin D1 and cyclin E2. More importantly, the BDNF-induced proliferation of the Ishikawa cells involved the ERK1/2 signaling pathway. (4) Conclusions: The stimulating effect of exogenous E2 on the expression of BDNF in the uterus and the action of BDNF promoted the proliferation of the Ishikawa cells through the TrkB-ERK1/2 signal pathway.

HHV-6A Infection of Endometrial Epithelial Cells Affects miRNA Expression and Trophoblast Cell Attachment

We recently reported that human herpesvirus 6 (HHV-6) infection is frequently present in endometrial tissue of women with unexplained infertility, and that virus infection induces a profound remodulation of miRNA expression in human cells of different origin. Since specific miRNA patterns have been associated with specific pregnancy outcomes, we aimed to analyze the impact of HHV-6A infection on miRNAs expression and trophoblast receptivity in human endometrial cells. To this purpose, a human endometrial cell line (HEC-1A) was infected with HHV-6A and analyzed for alterations in the expression of miRNAs and for permissiveness to the attachment of a human choriocarcinoma trophoblast cell line (JEG-3). The results showed that HHV-6A infection of endometrial cells up-modulates miR22 (26-fold), miR15 (19.5-fold), and miR196-5p (12.1 fold), that are correlated with implant failure, and down-modulates miR18 (11.4 fold), miR101-3p (4.6 fold), miR181-5p (4.9 fold), miR92 (3.3 fold), and miR1207-5p (3.9 fold), characterized by a low expression in preeclampsia. Moreover, HHV-6A-infected endometrial cells infected resulted less permissive to the attachment of trophoblast cells. In conclusion, collected data suggest that HHV-6A infection could modify miRNA expression pattern and control of trophoblast cell adhesion of endometrial cells, undermining a correct trophoblast cell attachment on endometrial cells.