Natural Cycle Controls Research Articles

Serum progesterone concentrations on the day of oocyte retrieval above 9.23 ng/ml may predict ovarian hyperstimulation syndrome risk in in vitro fertilized patients.

The aim of our study was to evaluate serum progesterone levels on the day of oocyte retrieval as a promising biomarker inorder to evaluate the risk of ovarian hyperstimulation syndrome in a group with controlled ovarian hyperstimulation protocols using either gonadotropin antagonists or agonists (GnRH), compare with a natural cycle control group. Patients were divided into 3 groups (148 patients in total): control group in the natural cycle, patients treated with GnRH agonist and patients treated with GnRH antagonist. When we compared both controlled ovarian hyperstimulation (COH) protocol groups with the control group, we found statistically higher levels of progesterone in patients after COH (control versus long protocol group: 1.43 ± 1.28 ng/ml versus 8.95 ± 5.95 ng/ml; P < 0.001; control versus GnRH antagonist group: 1.43 ± 1.28 ng/ml versus 7.18 ± 5.13 ng/ml; P < 0.001). According to receiver operating characteristic (ROC) analysis, the level of serum progesterone on the day of oocyte retrieval, above which the risk of ovarian hyperstimulation syndrome (OHSS) is associated with a more than fourfold higher risk (OR 4.24; 95% CI 2.6 - 6.9) was found to be 9.23 ng/ml, with AUC: 0.896, P = 0.026 (95% CI 0.845 - 0.947). Progesterone level on the day of oocyte retrieval may be used as an additional sensitivity marker in treatment of early forms as well by freezing of embryos in prevention of late forms of OHSS.

Controlled ovulation of the dominant follicle using progestin in minimal stimulation in poor responders

BackgroundThe use of progestin (P) during ovarian stimulation is effective in blocking the luteinizing hormone (LH) surge in women with normal ovarian reserve, however, its effects have not been determined in poor responders. This study aimed to explore the follicular dynamics in P-primed minimal stimulation in poor responders.MethodsA total of 204 infertile women with diminished ovarian reserve were allocated into the medroxyprogesterone acetate (MPA) group or the natural-cycle control group in an alternating order. MPA (10 mg) was administered daily beginning from the early follicular phase and a low dose of hMG was added in the late follicular phase if the serum FSH level was lower than 8.0mIU/ml. When a dominant follicle reached maturity, triptorelin 100 μg and hCG 1000 IU were used for trigger, and oocytes were retrieved 34-36 h later.All viable embryos were cryopreserved for subsequent frozen embryo transfer. Natural cycle IVF was used as controls.ResultsCompared with the natural cycle group, the MPA group exhibited a larger pre-ovulatory follicle (18.7 ± 1.8 mm vs 17.2 ± 2.2 mm), a longer follicular phase (13.6 ± 3.6 days vs 12.3 ± 3.2 days), and higher peak oestradiol values (403.88 ± 167.16 vs 265.26 ± 122.16 pg/ml), while maintaining lower LH values (P < 0.05). The incidences of spontaneous LH surge and premature ovulation decreased significantly (1.0% vs 50%; 2% vs. 10.8%, respectively; P < 0.05). A greater number of oocytes and viable embryos were harvested from the MPA group than from the natural cycle group (P < 0.05). Moreover,the clinical pregnancy rate was slightly higher in the MPA group than in the natural cycle controls, but the difference was not significant (11.8% vs 5.9%, P > 0.05).ConclusionThis study supported the hypothesis that P-primed minimal stimulation achieved ovulation control of the dominant follicle and did not adversely affect the quality of oocytes in poor responders. Therefore, P-priming is a promising approach to overcome premature ovulation in minimal stimulation for poor responders.Trial registrationChiCTR-OCH-14004176. Registered on January 8, 2014.

GnRH analogues may increase endometrial Hoxa10 promoter methylation and affect endometrial receptivity.

The present study aimed to investigate whether gonadotropin-releasing hormone analogues (GnRH-as), including GnRH agonists and antagonists, affect endometrial homeobox (Hox) a10 DNA methylation during the implantation window in mice. GnRH analogue mouse models were used and were treated with either human menopausal gonadotropin (HMG) and a GnRH agonist or HMG and a GnRH antagonist. Uterus samples were collected 48 h after GnRH analogue treatment or ovulation. Bisulfite sequencing polymerase chain reaction (PCR), quantitative-PCR and western blot analysis were performed to assess Hoxa10 and integrin β3 expression. Scanning electron microscope analyses were conducted to analyze pinopode development. Compared with the natural cycle control mice, mice in the GnRH analogue groups were found to exhibit increased levels of methylation at the Hoxa10 promoter, decreased Hoxa10 mRNA and protein expression and disrupted pinopode development. These findings suggest that GnRH-as may be associated with altered Hoxa10 DNA methylation, thus GnRH-as may affect uterine Hoxa10 expression and endometrial receptivity.

Effects of GnRH antagonist on endometrial protein profiles in the window of implantation.

GnRH antagonists can suppress luteinizing hormone and follicle-stimulating hormone (FSH), with less initial stimulatory effect and lower risk of ovarian hyperstimulation syndrome. The effects of GnRH antagonists on embryonic implantation remain controversial. To evaluate the effects of GnRH antagonists, endometrial tissues were biopsied from 12 women with intracytoplasmic sperm injection treatment, in which four subjects undergoing controlled ovulation stimulation with rFSH and GnRH antagonist, four subjects with a GnRH agonist long protocol, and four natural cycle controls. After iTRAQ quantification analysis, 24 proteins showed differential expression between natural cycle and agonist treatment group and 39 proteins between natural cycle and antagonist treatment group. A total of seven proteins demonstrated differential expression only in antagonist treatment group. Bioinformatic analysis implied these proteins can function in cell processes including angiogenesis, cell proliferation, apoptosis, cell migration, and immune response. Furthermore, GnRH antagonist suppressed the function of GNAS and ANPEP, which were important for endometrial functions. Immunohistochemical staining showed that ANPEP was mainly localized in the human endometrial stroma, while ACO2, CDC5L, and GNAS were mainly localized in the glands. This study could provide insights into the effect of GnRH antagonists on the endometrium, and help optimize the embryo implantation and improve the success rate for GnRH antagonist protocol.

GnRH antagonists may affect endometrial receptivity

HOXA10 is an essential regulator of endometrial receptivity. To determine the effect of GnRH antagonists on endometrial receptivity, we assessed endometrial HOXA10 expression in GnRH antagonist, GnRH agonist, and natural cycles. Prospective case-control study. University academic medical center. Nineteen subjects were included: 12 subjects underwent controlled ovarian hyperstimulation with recombinant FSH and used either a GnRH antagonist or a GnRH agonist; seven control subjects underwent natural cycles. Pipelle endometrial biopsies were obtained 11 days after hCG administration or spontaneous LH surge in untreated cycles, respectively. Immunohistochemistry was used to assess HOXA10 protein expression in endometrial glands and stroma. Endometrial HOXA10 protein expression. HOXA10 expression was significantly decreased in endometrial stromal cells in GnRH antagonist-treated cycles compared with GnRH agonist-treated cycles or natural cycle control subjects. There was no significant difference in glandular cell HOXA10 expression among the three groups. Use of GnRH antagonists may be associated with impaired HOXA10 expression in endometrial stromal cells and thus may affect endometrial receptivity.

Genomic and proteomic analyses of the endometrium during the window of implantation following laser capture microdissection: Comparison of natural and controlled ovarian hyperstimulation (COH) cycles

The process of embryo implantation is a condition of equilibrium in the up- and down-regulation of diverse endometrial genes/proteins under control of steroid hormones and other local factors. The objective of this study was to compare endometrial gene and protein expression during the window of implantation of natural and COH cycles following tissue compartmentalization using laser capture microdissection (LCM). Prospective, randomized and blinded research study. Endometrial biopsies were obtained during the putative window of implantation (day 21) from (a) COH cycles of oocyte donors (n = 10); and (b) natural cycle controls (n = 5). Oocyte donors were stimulated either with a GnRH agonist (long protocol) and recombinant FSH (rFSH) or with rFSH and a GnRH antagonist. For global gene expression profiling, cRNA was generated and hybridized to Affymetrix Hu95A microarrays. Prior to proteomic analysis, LCM was used to isolate specific endometrial cell types, i.e. epithelial and stromal cells. Proteomics was performed to the compartmentalized endometrium using surface enhanced laser desorption/ionization (SELDI)-time of flight spectrometry (TOF) technology. Protein lysates were loaded robotically on an IMAC 3 chip pretreated with CuSO4. Each resulting protein peak was determined and its intensity normalized for total ion current. Statistical analysis of the genomic data was performed using pair wise multiple group comparison (SAM software), while statistical analysis of the proteomic data was based on the Student’s t test and CART analysis using Ciphergen’s Biomarker Patterns Software. SAM analysis reveled that the endometrium of COH cycles differed in 18 genes in comparison to the timely matched natural cycles. Optineurin and procollagen type 3 endopeptidase were genes up regulated in the COH group, which were validated by real time RT-PCR. SELDI spectra revealed similar protein fingerprinting in the dissected endometrial epithelium in natural versus COH cycles stimulated with a GnRH antagonist. On the other hand, microdissected epithelium from GnRH agonist cycles depicted a different fingerprinting compared to natural and GnRH antagonist cycles. Furthermore, microdissected stromal compartments differed in the protein fingerprinting in the three studied groups. Several proteins, with masses ranging from 2.5 to 17 kD were differentially expressed among the three groups. Importantly, two unique 11 kD and 15 kD protein peaks were observed in the GnRH agonist cycles. Minimal differences in gene expression were found when comparing COH versus natural cycles. SELDI fingerprinting revealed a similar protein expression pattern in the epithelium of natural and GnRH antagonist cycles. However, differences were observed in GnRH agonist cycles. The stromal compartment demonstrated divergent protein fingerprinting amongst groups. These results provide a basic background on global gene and protein expression analysis of the endometrium during the window of implantation under natural and stimulated conditions.

Endometrial hormone receptors and proliferation index in the periovulatory phase of stimulated embryo transfer cycles in comparison with natural cycles and relation to clinical pregnancy outcome

Objective: To investigate the endometrial steroid receptors and proliferation index in GnRH analogue/hMG-stimulated cycles in comparison with natural cycles and their relation to clinical pregnancy outcome. Design: Prospective observational study. Setting: Tertiary referral center. Patient(s): Twenty-seven stimulated patients with GnRH agonist and hMG. Twenty normo-ovulatory patients were the natural cycle controls. Intervention(s): Endometrial aspiration biopsies: in stimulated cycles on the day of oocyte retrieval within the ET cycle (Day OPU) (n = 20) or 2 days later (Day OPU + 2) (n = 7); in natural cycles on the natural day of ovulation (Day NO) (n = 10) or on the day of ovulation + 2 (Day NO + 2) (n = 10). Main Outcome Measure(s): Comparison of endometrial maturation, estrogen (ER) and P receptor (PR), and proliferation index by immunohistochemistry in natural and stimulated cycles, correlation with pregnancy outcome in stimulated cycles. Result(s): Stimulated cycles Day OPU showed significantly advanced endometrial maturation compared to natural cycles Day NO; stromal ER and glandular and stromal PR staining was lower in stimulated than in natural cycles, but higher on Day OPU than on Day NO + 2; proliferation index was lower in all stimulated cycles. Steroid receptors and proliferation index in stimulated cycles were unrelated to clinical pregnancy occurrence. Conclusion(s): Compared to natural cycles, ovarian stimulation induced an imbalance in endometrial ER and PR and led to a profound antimitotic effect in the peri-ovulatory phase. These parameters were, however, not predictive of clinical pregnancy in cycles with ET.

Histochemical localization of endometrial insulin-like growth factor binding protein-1 and -3 during the luteal phase in controlled ovarian hyperstimulation cycles: a controlled study

Objective: To determine if controlled ovarian hyperstimulation (COH) affects the endometrial expression of IGFBP-1 and IGFBP-3. Design: Prospective, controlled study. Setting: Tertiary infertility clinic. Patient(s): Eighteen oocyte donors undergoing COH cycles and 17 natural cycle controls. Intervention(s): Controlled ovarian hyperstimulation, endometrial biopsies. Main Outcome Measure(s): Immunohistochemical scoring of endometrial IGFBP-1 and -3 expression, morphological endometrial dating, and serum estradiol (E 2), LH, and progesterone (P 4) concentrations. Result(s): No statistically significant difference was observed between natural and stimulated cycles in change in IGFBP-1 or -3 over standardized cycle days throughout the window of embryo implantation (days 17–24). The IGFBP-1 and -3 expression was zero or near zero for both the natural and COH cycles until day 12–13. Both IGFBPs showed increased production throughout the secretory phase. Advanced endometrial histology (≥1 day) in glands and stroma was noted in COH cycles. Significant positive correlations of E 2 and P 4 were noted with IGFBP-1 and -3 but not with advanced endometrial morphology in the COH cycles. Conclusion(s): The COH cycles have no significantly increased endometrial IGFBP-1 or -3 expression throughout the implantation phase of the luteal cycle compared with normal menstrual cycles. Both IGFBPs were absent in the proliferative phase and increased throughout the secretory portion of the embryo implantation window.