Establishment of a novel human embryonic stem cell-derived trophoblastic spheroid implantation model

postprin

Human chorionic gonadotropin stimulates spheroid attachment on fallopian tube epithelial cells through the mitogen-activated protein kinase pathway and down-regulation of olfactomedin-1

OBJECTIVE: To study the effect of human chorionic gonadotropin (hCG) on olfactomedin-1 (Olfm1) expression and spheroid attachment in human fallopian tube epithelial cells in vitro. DESIGN: Experimental study. SETTING: Reproductive biology laboratory. PATIENT(S): Healthy non-pregnant women. INTERVENTION(S): No patient interventions. MAIN OUTCOME MEASURE(S): Luteinizing hormone/chorionic gonadotropin receptor (LHCGR) and Olfm1 expression in fallopian tube epithelium cell line (OE-E6/E7 cells). OE-E6/E7 cells treated with hCG, U0126 extracellular signal-regulated kinase (ERK) inhibitor, or XAV939 Wnt/β-catenin inhibitor were analyzed by Western blotting, real-time polymerase chain reaction, and in vitro spheroid attachment assay. RESULT(S): Human chorionic gonadotropin increased spheroid attachment on OE-E6/E7 cells through down-regulation of Olfm1 and activation of Wnt and mitogen-activated protein kinase (MAPK) signaling pathways. U0126 down-regulated both MAPK and Wnt/β-catenin signaling pathways and up-regulated Olfm1 expression. XAV939 down-regulated only the Wnt/β-catenin signaling pathway but up-regulated Olfm1 expression. CONCLUSION(S): Human chorionic gonadotropin activated both ERK and Wnt/β-catenin signaling pathways and enhanced spheroid attachment on fallopian tube epithelial cells through down-regulation of Olfm1 expression.postprin

A Novel, Stable, Estradiol-Stimulating, Osteogenic Yam Protein with Potential for the Treatment of Menopausal Syndrome

A novel protein, designated as DOI, isolated from the Chinese yam (Dioscorea opposita Thunb.) could be the first protein drug for the treatment of menopausal syndrome and an alternative to hormone replacement therapy (HRT), which is known to have undesirable side effects. DOI is an acid- and thermo-stable protein with a distinctive N-terminal sequence Gly-Ile-Gly-Lys-Ile-Thr-Thr-Tyr-Trp-Gly-Gln-Tyr-Ser-Asp-Glu-Pro-Ser-Leu-Thr-Glu. DOI was found to stimulate estradiol biosynthesis in rat ovarian granulosa cells; induce estradiol and progesterone secretion in 16- to 18-month-old female Sprague Dawley rats by upregulating expressions of follicle-stimulating hormone receptor and ovarian aromatase; counteract the progression of osteoporosis and augment bone mineral density; and improve cognitive functioning by upregulating protein expressions of brain-derived neurotrophic factor and TrkB receptors in the prefrontal cortex. Furthermore, DOI did not stimulate the proliferation of breast cancer and ovarian cancer cells, which suggest it could be a more efficacious and safer alternative to HRT.link_to_OA_fulltex

Trichinella inflammatory myopathy: host or parasite strategy?

The parasitic nematode Trichinella has a special relation with muscle, because of its unique intracellular localization in the skeletal muscle cell, completely devoted in morphology and biochemistry to become the parasite protective niche, otherwise called the nurse cell. The long-lasting muscle infection of Trichinella exhibits a strong interplay with the host immune response, mainly characterized by a Th2 phenotype

Human chorionic gonadotropin enhances receptivity of fallopian tube through down-regulation of olfactomedin-1

Conference Theme: Reproductive Health: Nano to GlobalTubal ectopic pregnancy (TEP) occurs in 2 to 5 % of all pregnancy. In humans, TEP may lead to tubal rupture and consequently hemorrhage and maternity death. Various factors, including tubal infection, tubal damage, smoking and in vitro fertilization are associated with TEP. Previous microarray studies suggested that Olfactomedin-1 (Olfm1) is down-regulated in receptive endometrium during the window of implantation. In vitro studies demonstrated that down-regulation of Olfm1 would increase attachment of spheroids (embryo surrogate) onto human endometrial epithelial cells, while recombinant Olfm1 reduces spheroids attachment in vitro. Clinical study found that intrauterine injection of hCG could enhance implantation rate in IVF/ICSI. Our preliminary result showed that Olfm1 was also expressed in epithelial cells of Fallopian tube. Interestingly, results from our laboratory demonstrated that decreased epithelial Olfm1 expressions were found in Fallopian tubes from women with TEP. Furthermore, our unpublished data suggested that human chorionic gonadotropin (hCG), a heterodimeric peptide hormone secreted by human pre-implantation embryo, down-regulated Olfm1 expression in Fallopian tube epithelial cells (OE-E6/E7). Therefore, we hypothesize that embryonic hCG and possibly sex steroid hormone down-regulates Oflm1 in the Fallopian tube leading to tubal implantation and hence causing TEP. This study aimed to find out the role of hCG in establishment of TEP. Immunohistochemistry of estrogen receptor alpha (ER alpha), progesterone receptor (PR), glucocorticoid receptor (GR), hCG receptor and Olfm1 was compared in sections of Fallopian tubes taken from women in the follicular (n=6) and luteal (n=5) phases . JAR spheroid-Fallopian epithelial cell (OE-E6/E7) coculture model was used to evaluate the attachment rate. No significant difference (p>0.05) was observed in ER alpha, PR and GR expressions in both follicular and luteal phase samples; while hCG receptor and Olfm1 were up- and down-regulated respectively in luteal phase Fallopian tubes compared to those in follicular phase. In the co-culture model, hCG (25IU/ml) down-regulated Olfm1 expression but stimulates JAR spheroids attachment onto OE-E6/E7 cells when the OE-E6/E7 cells but not the JAR cells were treated with hCG. Moreover, only hCG and its beta-subunit could enhance attachment in vitro. Knockdown of hCG receptor in the OE-E6/E7 cells by siRNA reduced; while knockdown of Olfm1 increased spheroids attachment in vitro. Taken together, Olfm1 showed a cyclic expression pattern in Fallopian epithelial cells, which is the same as endometrium epithelial cells. hCG down-regulated Olfm1 and enhanced spheroids attachment onto Fallopian epithelial cells. The molecular mechanism by which hCG down-regulates Olfm1 warrants further investigation. (The work is partly supported by CRCG grant to KFL

The endocrine disruptor perfluorooctane sulfonate (PFOS) modulates embryo attachment and trophoblasts invasion through Wnt/b-catenin inhibition

No. P-07Conference Theme: Past, Present and Future of Obstetrics and Gynaecolog

Role of hCG on attachment of trophoblast onto endometrial epithelial cells through activation of ERK and Wnt/beta-catenin signaling pathway

Symposium on Osteoporosis and Oral Presentatio

In-vitro study on the effect of ulipristal acetate on human embryo implantation using a trophoblastic spheroid and endometrial cell co-culture model

Poster PresentationPoster awardsConference theme: Challenges in Sexual and Reproductive HealthObjectives: Ulipristal acetate (UPA), a selective progesterone receptor modulator, has been introduced for use in emergency contraception. The main mechanism of action is inhibiting or delaying ovulation. Whether UPA can have secondary action by inhibiting implantation is still uncertain. The present study examined the effect of UPA on human embryo implantation using an in-vitro human trophoblastic spheroid and endometrial cell co-culture model. Method: We studied the effect of UPA on implantation using a trophoblastic spheroidsendometrial cell attachment assay. The JAr (human choriocarcinoma) and Ishikawa (human endometrial adenocarcinoma) cell lines were treated with graded concentrations of UPA (0, 0.04, 0.4 and 4μM) for 24 hours. We took the peak serum drug level after oral administration of UPA 30 mg, i.e. 0.4 μM, as the pharmacological concentration, and our experimental range covered tentimes below and above this. After treatment, the JAr cells were trypsinized and gently shaken at 106rpm overnight to form spheroids of 100-150mm size, which were used as the embryo surrogate. A confluent monolayer of the Ishikawa cells was used as the endometrium surrogate, onto which the spheroids were seeded and cultured for 1 hour at 37oC under 5% CO2. The coculture was then shaked at 140rpm for 10 minutes to remove any unattached spheroids. The number of attached JAr spheroid was then counted under light microscope. Attachment rate was defined as the ratio of the number of attached spheroids to the total number seeded. The experiment was also repeated using cultured primary human endometrial cells (aspirated 7 days after the LH surge) as the endometrium surrogate, which was co-cultured with trophoblastic spheroids for 3 hours after treating the respective cells with 4μM UPA. The results were pooled from 19 and 7 independent repeats for the Ishikawa and primary endometrial cell experiments respectively. Results: In the Ishikawa experiments, there was no significant difference in the trophoblastic spheroid attachment rate after treatment with UPA at 0 (93.0%), 0.04 (93.6%), 0.4 (93.4%) and 4 (91.4%) μM concentrations (p > 0.05). In the primary endometrial cell experiments, again no significant difference was observed in trophoblastic spheroid attachment rate between the treatment group (UPA 4μM, 42.1%) compared to the control (without UPA treatment, 48.3%, p > 0.05). Significant suppression of spheroid attachment rate (p < 0.001) was observed in the positive controls which were set up with methotrexate 5μM treatment. Conclusions: UPA at pharmacological concentration used for emergency contraception may not have inhibitory effect on embryo implantation.link_to_OA_fulltex

The Endocrine Disruptor Perflurooctane Sulfonate (PFOS) modulates spheroids attachment onto endometrial cells through Wnt/β -catenin Inhibition

Poster Presentation: Theme 2: Cancer & Cell Biology: no. 2.2