How Placenta Promotes the Successful Reproduction in High-Altitude Populations: A Transcriptome Comparison between Adaptation and Acclimatization.

As the best adapted high altitude population, Tibetans feature a relatively high offspring survival rate. Genome-wide studies have identified hundreds of candidate SNPs related to high altitude adaptation of Tibetans, although most of them have unknown functional relevance. To explore the mechanisms behind successful reproduction at high altitudes, we compared the placental transcriptomes of Tibetans, sea level Hans (SLHan), and Han immigrants (ImHan). Among the three populations, placentas from ImHan showed a hyperactive gene expression pattern. Their increased activation demonstrates a hypoxic stress response similar to sea level individuals experiencing hypoxic conditions. Unlike ImHan, Tibetan placentas were characterized by the significant up-regulation of placenta-specific genes, and the activation of autophagy and the tricarboxylic acid (TCA) cycle. Certain conserved hypoxia response functions, including the antioxidant system and angiogenesis, were activated in both ImHan and Tibetans, but mediated by different genes. The coherence of specific transcriptome features linked to possible genetic contribution was observed in Tibetans. Furthermore, we identified a novel Tibetan-specific EPAS1 isoform with a partial deletion at exon six, which may be involved in the adaption to hypoxia through the EPAS1-centred gene network in the placenta. Overall, our results show that the placenta grants successful pregnancies in Tibetans by strengthening the natural functions of the placenta itself. On the other hand, the placenta of ImHan was in an inhabiting time-dependent acclimatization process representing a common hypoxic stress response pattern.

Hormonal regulation of endometrial olfactomedin expression and its suppressive effect on spheroid attachment onto endometrial epithelial cells.

BACKGROUND: Olfactomedin (Olfm) is a member of a diverse group of extracellular matrix proteins important for neuronal growth. Recent microarray studies identified Olfm as one of the down-regulated transcripts in receptive endometrium at the time of embryo attachment and implantation. However, the underlying molecular mechanisms that govern Olfm expression and its effect on embryo attachment and implantation remain unknown. METHODS: The expression of Olfm in the human endometrium was investigated by real-time PCR, western blotting and immunohistochemistry on human endometrial biopsies from natural and ovarian stimulated cycles. To investigate the function of Olfm in trophoblast-endometrial cell attachment, an in vitro spheroid-endometrial cell co-culture study was performed. RESULTS: Human endometrial Olfactomedin-1 and -2(Olfm-1 and -2) transcripts decreased significantly from the proliferative to the secretory phases of the menstrual cycle. Olfm protein was strongly expressed in the luminal and glandular epithelium and moderately in the stromal cells of human endometria. Ovarian stimulation significantly decreased (P < 0.05) the expression of endometrial Olfm-1 and -2 transcripts in patients receiving IVF treatment when compared with those in the natural cycle. Importantly, recombinant Olfm-1 suppressed JAr spheroid attachment onto Ishikawa cells and this was not associated with changes of ß-catenin and E-cadherin expression in trophoblast and endometrial cells. CONCLUSIONS: Decreased expression of Olfm during the receptive phase of the endometrium may allow successful trophoblast attachment for implantation.

Excessive ovarian stimulation up-regulates the Wnt-signaling molecule DKK1 in human endometrium and may affect implantation: an in vitro co-culture study.

BACKGROUND: High serum estradiol (E2) levels following ovarian stimulation lead to reduced implantation and pregnancy rates, yet the underlying mechanisms remain unknown. We investigated if aberrant expression of genes in the Wnt-signaling pathway may be involved. METHODS: Microarray and real-time PCR analysis were performed to analyze gene expression profiles of endometrial samples taken at day hCG + 7 in stimulated cycles, and days LH + 7 and LH + 10 in natural cycles. Expression of several Wnt-signaling transcripts, including Dickkopf homolog 1 (DKK1), DKK2 and secreted frizzled-related protein 4 (sFRP4), was analyzed throughout the menstrual cycle. JAr spheroid/Ishikawa endometrial cell co-culture experiments were established to study effects of DKK1 on spheroid attachment in vitro. RESULTS: We identified 351 differentially expressed genes. Endometrial samples taken at hCG + 7 had similar expression profiles to those at LH + 10. DKK1 transcripts were up-regulated and DKK2 and sFRP4 were down-regulated in the stimulated compared with LH + 7 group (all P < 0.05). DKK1 transcripts were low in proliferative phase (PS) and increased in late-secretory phase (LS, P < 0.05), although DKK2 peaked in mid-secretory phase (P < 0.05). sFRP4 transcripts were high in PS. Treatment of spheroid with recombinant human DKK-1 protein dose-dependently suppressed (P < 0.05 versus control) spheroids attachment onto endometrial cells (associated with decreased beta-catenin protein): this suppression was nullified by anti-DKK1 antibody. CONCLUSION: Gene expression patterns in stimulated cycles resembled those of LS in natural cycles, when the implantation window is about to close, suggesting high serum E2 and/or progesterone concentrations may advance endometrial development, altering the implantation window and possibly decreasing pregnancy rate. Aberrant expression of DKK1 might impair embryo attachment and implantation in vivo.

Aberrant expression of angiopoietins-1 and -2 and vascular endothelial growth factor-A in peri-implantation endometrium after gonadotrophin stimulation.

BACKGROUND: Ovarian stimulation affects normal endometrial development. The expression of angiopoietin-1 (Ang-1), angiopoietin-2 (Ang-2) and vascular endothelial growth factor-A (VEGF-A) and the vascular state in the peri-implantation endometrium in women with natural and gonadotrophin-stimulated cycles were compared. METHODS: The expression of these angiogenesis-associated molecules in endometrial biopsies, collected on Day 7 after human chorionic gonadotrophin injection or luteinizing hormone surge in stimulated or natural cycles respectively, or at mid-luteal phase of women undergoing diagnositic laparoscopy, were analysed. RESULTS: Women with gonadotrophin-stimulation had lower Ang-1, but higher Ang-2, mRNA and protein expression (P < 0.05), and increased concentrations of von Willebrand factor (vWF) and blood vessel density than those with natural cycles (P < 0.05). Although stimulated cycles had higher VEGF-A mRNA expression (P = 0.023), VEGF-A protein expression was similar between the groups. Lower Ang-1/Ang-2 but higher Ang-2/VEGF-A mRNA ratios (P = 0.025) were found after gonadotrophin-stimulation. The ratios were negatively (P < 0.001) and positively correlated (P < 0.001) with estradiol levels, respectively. Cyclical changes in Ang-1 and Ang-2, but not in VEGF-A expression were noted. CONCLUSIONS: The decreased Ang-1 concentration and Ang-1/Ang-2 ratio and the increased Ang-2 concentration, with the increased vWF concentration and blood vessel density, in stimulated cycles suggests advanced endometrial angiogenesis after gonadotrophin-stimulation.

Gene expression profiling of human peri-implantation endometria between natural and stimulated cycles.

OBJECTIVE: To investigate the effect of high serum E(2) levels in gonadotropin-stimulated cycles (hCG+7) on the gene expression patterns of human endometrium compared with natural cycles on the seventh day of LH surge (LH+7) and elucidate the underlying molecular changes that may be related to endometrial receptivity. DESIGN: Observational study. SETTING: University Hospital. PATIENTS(S): Infertile patients with normal menstrual cycles undergoing IVF treatment. INTERVENTION(S): Gonadotropin stimulation and endometrial biopsy. MAIN OUTCOME MEASURE(S): Gene expression by microarray and quantitative polymerase chain reaction (qPCR). RESULT(S): Endometrial samples from natural (n = 5) and stimulated (n = 8) cycles were collected. Patients in the stimulated cycles were classified as moderate (n = 4) or excessive (n = 4) responders if their serum E(2) levels on the day of administration of hCG were 20,000 pmol/L, respectively. The RNA transcripts were profiled by Affymetrix HG-U133A microarray. Clustering and principal component analysis demonstrated a significant difference (>or=2-fold) in the expression patterns of 411 genes among the three groups. Putative estrogen response elements or progesterone response elements were identified in the promoter regions of 49 differentially expressed genes of diverse biologic functions. The qPCR confirmed the microarray result in 47 endometrial samples. CONCLUSION(S): High serum E(2) and/or progesterone modulate the gene expression profiles of human endometrium and may affect endometrial receptivity.