Steroid hormone regulation of EMP2 expression and localization in the endometrium

<p>Abstract</p> <p>Background</p> <p>The tetraspan protein epithelial membrane protein-2 (EMP2), which mediates surface display of diverse proteins, is required for endometrial competence in blastocyst implantation, and is uniquely correlated with poor survival from endometrial adenocarcinoma tumors. Because EMP2 is differentially expressed in the various stages of the murine and human estrous cycle, we tested the hypothesis that the steroid hormones progesterone and estrogen influence EMP2 expression and localization.</p> <p>Methods</p> <p>Frozen human proliferative and secretory endometrium were collected and analyzed for EMP2 expression using SDS-PAGE/Western blot analysis. The response of EMP2 to progesterone and estradiol was determined using a combination of real-time PCR, SDS-PAGE/Western blot analysis, and confocal immunofluorescence in the human endometrial carcinoma cell line RL95-2. To confirm the in vitro results, ovariectomized mice were treated with progesterone or estradiol, and EMP2 expression was analyzed using immunohistochemistry.</p> <p>Results</p> <p>Within normal human endometrium, EMP2 expression is upregulated in the secretory phase relative to the proliferative phase. To understand the role of steroid hormones on EMP2 expression, we utilized RL95-2 cells, which express both estrogen and progesterone receptors. In RL95-2 cells, both estradiol and progesterone induced EMP2 mRNA expression, but only progesterone induced EMP2 protein expression. To compare steroid hormone regulation of EMP2 between humans and mice, we analyzed EMP2 expression in ovarectomized mice. Similar to results observed in humans, progesterone upregulated endometrial EMP2 expression and induced EMP2 translocation to the plasma membrane. Estradiol did not promote translocation to the cell surface, but moderately induced EMP2 expression in cytoplasmic compartments in vivo.</p> <p>Conclusion</p> <p>These findings suggest that targeting of EMP2 to specific locations under the influence of these steroid hormones may be important for integrating the molecular responses required for implantation competence.</p

Superovulation with human chorionic gonadotropin (hCG) trigger and gonadotropin releasing hormone agonist (GnRHa) trigger differentially alter essential angiogenic factors in the endometrium in a mouse ART model†.

Gonadotropin-releasing hormone agonists (GnRHa) are used as an alternative to human chorionic gonadotropin (hCG) to trigger ovulation and decrease the risk of ovarian hyperstimulation syndrome. GnRHa is less potent at inducing ovarian vascular endothelial growth factor (VEGF), but may also affect endometrial angiogenesis and early placental development. In this study, we explore the effect of superovulation on endometrial angiogenesis during critical periods of gestation in a mouse model. We assigned female mice to three groups: natural mating or mating following injection with equine chorionic gonadotropin and trigger with GnRHa or hCG trigger. Females were killed prior to implantation (E3.5), post-implantation (E7.5), and at midgestation (E10.5), and maternal serum, uterus, and ovaries were collected. During peri-implantation, endometrial Vegfr1 and Vegfr2 mRNA were significantly increased in the GnRHa trigger group (P &lt; 0.02) relative to the hCG group. Vegfr1 is highly expressed in the endometrial lining and secretory glands immediately prior to implantation. At E7.5, the ectoplacental cone expression of Vegfa and its receptor, Vegfr2, was significantly higher in the hCG trigger group compared to the GnRHa group (P &lt; 0.05). Soluble VEGFR1 and free VEGFA were much higher in the serum of mice exposed to the hCG trigger compared to GnRHa group. At midgestation, there was significantly more local Vegfa expression in the placenta of mice triggered with hCG. GnRHa and hCG triggers differentially disrupt the endometrial expression of key angiogenic factors during critical periods of mouse gestation. These results may have significant implications for placental development and neonatal outcomes following human in vitro fertilization

Progesterone induces surface expression of EMP2 in mouse endometrium in vivo

Mice were injected with progesterone or sesame oil (control) daily for up to 5 days. Tissue was harvested at days 1, 2, and 4 of treatment or 3 days following completion of treatment, processed for immunohistochemistry, and then stained for EMP2 expression (A-D) or a vehicle control (E). (A) 24 hours of progesterone treatment; (B) 48 hours of treatment; (C) 4 days of treatment; (D) 3 days following treatment; (E) representative control treatment at 4 days. Magnification: (left) 100×; (right) 400×. The experiment was repeated at least 3 times with similar results; representative images are shown.<p><b>Copyright information:</b></p><p>Taken from "Steroid hormone regulation of EMP2 expression and localization in the endometrium"</p><p>Reproductive biology and endocrinology : RB&E 2008;6():15-15.</p><p>Published online 9 Apr 2008</p><p>PMCID:PMC2329639.</p><p></p

(A) Normal human endometrium specimens from subjects in proliferative and secretory phases were immunoblotted using EMP2 antisera

β-actin was used as a loading control. (B) Statistical analysis of EMP2 expression levels in proliferative (n = 3: subphase unspecified) and secretory (n = 3: one post-ovulatory day 3, one late, one unspecified) endometrial specimens from six independent patients. Data were analyzed using a 2-tailed, unpaired student t-test with a 95% confidence interval. *Changes in expression between the proliferative and secretory endometrium were significant, p = 0.038.<p><b>Copyright information:</b></p><p>Taken from "Steroid hormone regulation of EMP2 expression and localization in the endometrium"</p><p>http://www.rbej.com/content/6/1/15</p><p>Reproductive biology and endocrinology : RB&E 2008;6():15-15.</p><p>Published online 9 Apr 2008</p><p>PMCID:PMC2329639.</p><p></p