hCG Priming Research Articles

Androgen and inhibin B levels during ovarian stimulation before and after 8 weeks of low-dose hCG priming in women with low ovarian reserve.

Does 8 weeks of daily low-dose hCG administration affect androgen or inhibin B levels in serum and/or follicular fluid (FF) during the subsequent IVF/ICSI cycle in women with low ovarian reserve? Androgen levels in serum and FF, and inhibin B levels in serum, decreased following 8 weeks of hCG administration. Recently, we showed that 8 weeks of low-dose hCG priming, in between two IVF/ICSI treatments in women with poor ovarian responder (anti-Müllerian hormone (AMH) <6.29 pmol/l), resulted in more follicles of 2-5 mm and less of 6-10-mm diameter at the start of stimulation and more retrieved oocytes at oocyte retrieval. The duration of stimulation and total FSH consumption was increased in the IVF/ICSI cycle after priming. Hypothetically, hCG priming stimulates intraovarian androgen synthesis causing upregulation of FSH receptors (FSHR) on granulosa cells. It was therefore unexpected that antral follicles were smaller and the stimulation time longer after hCG priming. This might indicate a different mechanism of action than previously suggested. Blood samples were drawn on stimulation day 1, stimulation days 5-6, trigger day, day of oocyte retrieval, and oocyte retrieval + 5 days in the IVF/ICSI cycles before and after hCG priming (the control and study cycles, respectively). FF was collected from the first aspirated follicle on both sides during oocyte retrieval in both cycles. The study was conducted as a prospective, paired, non-blinded, single-center study conducted between January 2021 and July 2021 at a tertiary care center. The 20 participants underwent two identical IVF/ICSI treatments: a control cycle including elective freezing of all blastocysts and a study cycle with fresh blastocyst transfer. The control and study cycles were separated by 8 weeks (two menstrual cycles) of hCG priming by daily injections of 260 IU recombinant hCG. Women aged 18-40 years with cycle lengths of 23-35 days and AMH <6.29 pmol/l were included. Control and study IVF/ICSI cycles were performed in a fixed GnRH-antagonist protocol. Inhibin B was lower on stimulation day 1 after hCG priming (P = 0.05). Dehydroepiandrosterone sulfate (DHEAS) was significantly lower on stimulation day 1 (P = 0.03), and DHEAS and androstenedione were significantly lower on stimulation days 5-6 after priming (P = 0.02 and P = 0.02) The testosterone level in FF was significantly lower in the study cycle (P = 0.008), while the concentrations of inhibin B and androstenedione in the FF did not differ between the study and control cycles. A lower serum inhibin B in the study cycle corresponds with the antral follicles being significantly smaller after priming, and this probably led to a longer stimulation time in the study cycle. This contradicts the theory that hCG priming increases the intraovarian androgen level, which in turn causes more FSHR on developing (antral up to preovulatory) follicles. However, based on this study, we cannot rule out that an increased intra-follicular androgen level was present at initiation of the ovarian stimulation, without elevating the androgen level in serum and that an increased androgen level may have rescued some small antral follicles that would have otherwise undergone atresia by the end of the previous menstrual cycle. We retrieved significantly more oocytes in the Study cycle, and the production of estradiol per follicle ≥10-mm diameter on trigger day was comparable in the study and control cycles, suggesting that the rescued follicles were competent in terms of producing oocytes and steroid hormones. The sample size was small, and the study was not randomized. Our study design did not allow for the measurement and comparison of androgen levels or FSHR expression in small antral follicles before and immediately after the hCG-priming period. The results make us question the mechanism of action behind hCG priming prior to IVF. It is important to design a study with the puncture of small antral follicles before and immediately after priming to investigate the proposed hypothesis. Improved cycle outcomes, i.e. more retrieved oocytes, must be confirmed in a larger, preferably randomized study. This study was funded by an unrestricted grant from Gedeon Richter awarded to the institution. A.P. reports personal consulting fees from PregLem SA, Novo Nordisk A/S, Ferring Pharmaceuticals A/S, Gedeon Richter Nordics AB, Cryos International, and Merck A/S outside the submitted work and payment or honoraria for lectures from Gedeon Richter Nordics AB, Ferring Pharmaceuticals A/S, Merck A/S, and Theramex and Organon & Co and payment for participation in an advisory board for Preglem. Grants to the institution have been provided by Gedeon Richter Nordics AB, Ferring Pharmaceuticals A/S, and Merck A/S, and equipment and travel support has been given to the institution by Gedeon Richter Nordics AB. The remaining authors have no conflicts of interest to declare. ClinicalTrials.gov Identifier: NCT04643925.

P-682 Androgen and Inhibin B levels during ovarian stimulation before and after eight weeks of low dose hCG priming in women with low ovarian reserve

Abstract Study question Does eight weeks of low dose human chorionic gonadotropin (hCG)-priming affect androgen or Inhibin B levels in serum and follicular fluid during IVF/ICSI treatment? Summary answer Inhibin B was reduced on stimulation day 1 in the IVF/ICSI cycle following hCG-priming. Androgen levels in serum and follicular fluid was not increased. What is known already In our recently published pilot study, eight weeks of low-dose hCG-priming was used to stimulate the intraovarian androgen synthesis in women with AMH &amp;lt;6.29 pmol/L. Priming resulted in more retrieved oocytes and more follicles sized 2-5 mm and less sized 6-10 mm at the start of stimulation. Stimulation time and FSH consumption was increased after priming. HCG priming potentially stimulate the intraovarian androgen synthesis causing upregulation of FSH receptors on granulosa cells, and therefore it was unexpected that the antral follicles were smaller and stimulation time longer following priming. This might indicate a different mechanism of action than previously thought. Study design, size, duration A prospective, paired, non-blinded, single-center study including 20 women conducted between January 2021 and July 2021 at a tertiary referral hospital. Participants underwent two identical IVF treatments: a Control cycle including elective freezing of all blastocysts and a Study cycle with fresh blastocyst transfer. The Control and Study cycles were separated by eight weeks (two menstrual cycles) of hCG-priming by daily injections of 260 IE hCG (details in our previously published paper). Participants/materials, setting, methods Women aged 18-40 years with cycle lengths of 23-35 days and AMH &amp;lt;6.29 pmol/L. Control and Study IVF cycles were performed in a fixed GnRH-antagonist protocol. Blood samples were taken on stimulation day 1, stimulation day 5-6, trigger day, day of oocyte pick up (OPU), and trigger day + 7 days in the Control and Study cycles. Follicular fluid was collected from the first aspirated follicle on both sides during OPU in both cycles. Main results and the role of chance Inhibin B serum levels were significantly lower on stimulation day 1 after hCG-priming, but no other significant differences in serum Inhibin B or androgen levels were seen. The concentrations of Inhibin B and androstenedione in the follicular fluid from the Study and Control cycle did not vary but testosterone was significantly lower in the Study cycle. A lower Inhibin B in the Study cycle corresponds with the antral follicles being significantly smaller after priming, and this probably led to a longer stimulation time in the Study cycle. This contradicts the theory that androgen priming causes more FSH receptors on developing (antral up to preovulatory) follicles. Instead, we hypothesize that androgen priming rescued some small antral follicles that would have otherwise undergone atresia by the end of the previous menstrual cycle. The mechanism of action was likely an increase in androgen levels and FSH receptor expression within these small follicles, however, not enough to elevate androgen levels in serum. We retrieved significantly more oocytes in the Study cycle and the production of estradiol per follicle ≥10 mm on trigger day was comparable in the Study and Control cycle, suggesting that the rescued follicles were competent in producing oocytes and steroid hormones. Limitations, reasons for caution The sample size was small, and androgen and Inhibin B levels were not the primary outcome. We have not demonstrated a higher androgen level or FSH receptor expression in small antral follicles following hCG-priming. Wider implications of the findings The results make us question the mechanism of action behind hCG-priming prior to IVF. It is important to design a study with retrieval of small antral follicles immediately after priming to investigate the proposed hypothesis and to confirm improved cycle outcomes i.e., more retrieved oocytes in a larger study population. Trial registration number NCT04643925

P-626 Time interval between ovulation triggering and oocyte retrieval and ART outcomes: an updated meta-analysis

Abstract Study question To explore whether prolonged hCG-ovum pickup(hCG-opu) interval improves assisted reproductive technology outcomes. Summary answer The clinical pregnancy rates can be increased by prolonging the hCG-opu interval, which would help develop more reasonable time schedules for fertility centers and patients. What is known already hCG-OPU interval greatly affects ART success, which allows to obtain the maximum number of competent mature oocytes while avoiding spontaneous ovulation. There is also no consensus regarding the optimal time between hCG-OPU, which some studies had shown that ideal ART outcomes could be obtained when oocyte retrieval was done more than 36 h -39 h after hCG priming, while others have concluded that an interval of more than 36 hours does not improve ART outcomes. Study design, size, duration CENTRAL, CNKI, Cochrane Systematic Reviews, EMBASE, MEDLINE, PUBMED, and Web of Science up to February 7, 2022 were searched for studies reporting associations between hCG-ovum pickup intervals and assisted reproductive technology outcomes. We searched for prospective or retrospective cohort studies measuring the hCG-OPU interval, using 36 hours as the cutoff value between short and long intervals. A combination of the following key search terms was used: “oocyte retrieval,” “human chorionic gonadotropin,” and “interval.” Participants/materials, setting, methods Studies analyzing the relationship between the hCG-OPU interval and ART outcomes were considered eligible for abstract screening. Intervention types included short (≤36 h) and long (&amp;gt;36 h) hCG-ovum pickup intervals in assisted reproductive technology cycles. All outcomes were based upon only fresh embryo transfers. Data were pooled using random-effects models. Heterogeneity was assessed using the I 2 statistic. Main results and the role of chance 12 studies were included in the meta-analysis, including five retrospective cohort studies, one prospective cohort study, and six randomized or quasi-randomized controlled trials. The short and long interval groups had similar oocyte maturation rates, fertilization rate and high-quality embryo rate (OR, 0.69; 95% CI, 0.45–1.06; I2 = 91.1%, OR, 0.88; 95% CI, 0.77–1.0; I2 = 44.4% and OR, 1.05; 95% CI, 0.95–1.17; I2 = 8.6%, respectively). The clinical pregnancy rates in the long retrieval group were significantly higher than in the short retrieval group (OR, 0.66; 95% CI, 0.45–0.95; I2 = 35.4%). The groups had similar miscarriage and live birth rates (OR, 1.92; 95% CI, 0.66–5.60; I2 = 0.0% and OR, 0.50; 95% CI, 0.24–1.04; I2 = 0.0%, respectively). Limitations, reasons for caution Using studies without patient data represents a potential source of bias. The hCG-OPU intervals were selected based on clinical relevance, resulting in statistical heterogeneity. Despite sensitivity analysis, such heterogeneity sources call for caution when interpreting the results. Wider implications of the findings The clinical pregnancy rates can be increased by prolonging the hCG-OPU (&amp;gt;36h), which would support the search for better clinical outcomes. The finding should be validated. And if validated by multicenter, randomized, controlled trials, long interval could be implemented in ART programs, resulting in better clinical pregnancy outcomes. Trial registration number not applicable

Eight weeks of androgen priming by daily low-dose hCG injections before ICSI treatment in women with low ovarian reserve.

Does 8 weeks of continuous low-dose hCG administration increase the proportion of antral follicles that reach the preovulatory state during ovarian stimulation (OS) in women with low ovarian reserve? The proportion of antral follicles (2-10 mm) that reached the preovulatory state did not increase. The administration of androgens prior to OS might upregulate FSH receptor (FSHR) expression on granulosa cells, making follicles more responsive to exogenous FSH stimulation during OS. LH and hCG stimulate the local follicular androgen synthesis in theca cells and may be used as an endogenous androgen priming method. Exogenous priming by testosterone and dehydroepiandrosterone (DHEA) have been shown to increase the number of retrieved oocytes and live birth rate but the studies are small, and their use is associated with side effects. A prospective, paired, non-blinded single-center study including 20 women serving as their own controls conducted between January 2021 and July 2021 at The University Hospital Copenhagen Rigshospitalet, Denmark. Participants underwent two identical consecutive IVF/ICSI treatments, a Control cycle and a Study cycle, separated by ∼8 weeks (two menstrual cycles) of daily injections of 260 IU recombinant hCG (rhCG). A freeze-all strategy was applied in the Control cycle. Both IVF/ICSI cycles were performed in a fixed GnRH antagonist protocol using a daily dose of 300 IU recombinant FSH (rFSH) and GnRH antagonist 0.25 mg from stimulation days 5-6. Follicular output rate, defined as the number of follicles >16 mm on hCG trigger day divided by the antral follicle count (2-10 mm) at baseline, did not increase after 8 weeks of hCG priming (P = 0.8). The mean number of oocytes retrieved was significantly higher after the hCG priming being 4.7 (2.8) vs 3.2 (1.7) in the Study and Control cycle, respectively (P = 0.01). The duration of stimulation was longer in the Study versus the Control cycle (P = 0.05), despite the use of identical hCG trigger criterion and similar diameters of the three biggest follicles on hCG trigger day in the two cycles (P = 0.9). The sample size was small, and the number of oocytes retrieved was not the primary endpoint. Larger studies are needed to confirm this finding. Long-term, low-dose rhCG administration may increase the number of oocytes retrieved during IVF/ICSI in women with low ovarian reserve, but more research is needed before firm conclusions can be drawn. This study was funded by an unrestricted grant from Gedeon Richter. A.P. reports personal consulting fees from PregLem SA, Novo Nordisk A/S, Ferring Pharmaceuticals A/S, Gedeon Richter Nordics AB, Cryos International, and Merck A/S outside the submitted work and payment or honoraria for lectures from Gedeon Richter Nordics AB, Ferring Pharmaceuticals A/S, Merck A/S, and Theramex and Organon & Co. Grants to the institution have been provided by Gedeon Richter Nordics AB, Ferring Pharmaceuticals A/S, and Merck A/S and receipt of equipment by the institution from Gedeon Richter Nordics AB is reported. The remaining authors have no conflicts of interest to declare. ClinicalTrials.gov Identifier: NCT04643925.

Pregnancy outcomes in infertile patients undergoing intracytoplasmic sperm injection and two interval programs of hCG-to-oocyte aspiration

This study aimed to determine the outcomes of intracytoplasmic sperm injection (ICSI) if oocyte retrieval was done 32-34 hours or 34:05-36 hours after human chorionic gonadotropin (hCG) injection. A randomized sample involving 186 patients with tubal failure was divided into groups A (96 patients) and B (90 patients). Intracytoplasmic sperm injection was performed on all patients according to described protocols. The number of oocytes retrieved, oocyte cumulus complex quality, number of fertilized eggs, and pregnancy rates were compared between groups. The total of oocytes retrieved in group B was significantly higher than in group A but not significant (P=0.068). The oocyte maturation rate was also significantly higher in the long interval group B than in the short interval group A (P=0.039). There was a significant difference between the two groups in terms of fertilization rate (0.040), and the pregnancy rate in group B was higher than that in group A, but it was not significant (P=0.055). The prolonged interval could also increase the pregnancy rate, but it was not significant. It seems that if the interval between hCG priming and oocyte retrieval is prolonged, the percentage of the number of oocytes retrieved, mature oocytes (MII), and fertilized oocytes increases.

P-626 Eight weeks of androgen priming by low dose hCG before IVF/ICSI in women with low ovarian reserve

Abstract Study question Does eight weeks of low dose hCG priming improve the outcome of ovarian stimulation for IVF/ICSI in women with low ovarian reserve. Summary answer Significantly more oocytes were retrieved following eight weeks of low dose hCG priming, when compared to an identical IVF/ICSI treatment immediately before the priming period. What is known already Administration of androgens prior to ovarian stimulation for IVF might upregulate the FSH receptor level on granulosa cells making the follicles more responsive to exogenous FSH. LH and hCG stimulate the follicular androgen synthesis, acting through LH receptors on the theca cells, enabling their use as an intra-ovarian androgen priming method. Exogenous testosterone and DHEA priming have been shown to increase the number of retrieved oocytes and the live birth rate, but systemic androgen administration is associated with side effects. Long term ‘intra-ovarian’ priming with hCG without systemic effects in women with low ovarian reserve has not previously been investigated. Study design, size, duration A prospective, paired, non-blinded study including 20 women serving as their own controls conducted between January 2021 and July 2021 at The University Hospital Copenhagen Rigshospitalet, Denmark. Participants underwent two identical consecutive IVF/ICSI treatments, a Control cycle including elective freezing of all blastocysts and a Study cycle with fresh blastocyst transfer. The Control and Study cycle were separated by approximately eight weeks (two menstrual cycles) of androgen priming by daily injections of 260 IE hCG. Participants/materials, setting, methods Inclusion criteria were age 18-40 years, regular menstrual cycle 23-35 days and AMH &amp;lt;6.29 pmol/L. Control and Study IVF/ICSI cycles were performed in a fixed GnRH-antagonist protocol using a daily dose of 300 IU rFSH and GnRH antagonist 0.25 mg from stimulation day 5-6. Primary endpoint was the follicular output rate defined as the pre-ovulatory follicle count (&amp;gt;16 mm) on hCG trigger day divided by the antral follicle count (2-10 mm) on cycle day 2-3. Main results and the role of chance 20 women with a mean (SD) age of 36.8 (3.2) years and an AMH level of 3.7 (1.5) pmol/L were included and completed the study. The number of oocytes retrieved was significantly higher in the Study compared to Control Cycle: 4.7 (2.8) vs. 3.2 (1.7), (P = 0.007), and the number of blastocysts was 1.2 (1.6) vs. 0.6 (0.7) in the Study compared to Control cycle, (P = 0.11). The follicular output rate was 0.4 (0.4) in the Study cycle initiated immediately after eight weeks of hCG priming, which was similar to the follicular output rate of 0.38 (0.2) in the Control cycle performed just before initiating the priming period (P = 0.81). The number of follicles above 10 mm on trigger day was 7.3 (5.0) in the Study and 5.3 (2.9) in the Control cycle (P = 0.018). Although the duration of stimulation seemed longer in the Study compared to Control cycle, 10.1 (1.5) and 8.9 (2.7) days respectively (P = 0.05), the same trigger criterion was defined and used in the two cycles confirmed by a similar mean diameter of the two biggest follicles on trigger day: 18.5 (1.1) vs. 18.3 (1.1) mm in the Study and Control cycle, respectively (P = 0.91). Limitations, reasons for caution The sample size calculation was based on the follicular output rate, and the study was not powered to look at oocyte retrieval rates. Bigger studies are needed to confirm this finding. Wider implications of the findings Although no increase in the follicular output rate was demonstrated, significantly more oocytes were retrieved after eight weeks of hCG priming, which is an important finding in a population of women with low ovarian reserve. Benefits of long term low dose hCG priming must be confirmed in a larger study. Trial registration number NCT04643925

ZONA PELLUCIDA DYSMORPHOLOGY IS ASSOCIATED WITH LOWER FERTILIZATION RATES IN ICSI

Although cytoplasmic dysmorphology of oocytes affects clinical outcomes, the relationship between the dysmorphology of the zona pellucida (ZP) observed during ICSI and clinical outcomes is still unclear. In this study, we aimed to clarify the clinical implications of dysmorphologies of ZP. We studied 3758 non-donor cycles who underwent oocyte retrieval at our private fertility center between January 2018 and September 2020. ZP dysmorphologies included ellipsoid ZP shape, partially overlapping ZP, and misshapen ZP. We defined the 383 cycles in which one or more ZP dysmorphological oocytes were observed in the ICSI procedure as the dys-ZP cycles, whereas 3375 cycles with normal-ZP were defined as the normal-ZP cycles for control. The patient characteristics were compared between the two groups to investigate the causes of ZP dysmorphologies. Furthermore, we compared fertilization rates, embryonic development and clinical outcomes for 550 dys-ZP oocytes and 10,228 normal-ZP oocytes. Fisher's exact test and Mann-Whitney U tests were used for statistical analysis. P<0.05 was considered statistically significant. There were no differences in maternal age or BMI between the two groups. The AMH level (2.41±2.39 vs 2.07±2.72 ng/ml), peak estradiol level at the time of hCG priming (1301.0±874.9 vs 1014.6±785.7 pg/ml), total dose of gonadotropins (2006.4±846.5 vs 1784.3±904.8 IU/ml), and the number of retrieved oocytes (8.69±5.94 vs 6.40±5.25) were significantly higher in the dys-ZP cycle than in the normal-ZP cycle. The incidence of ZP dysmorphologies was significantly lower in low ovarian stimulation (12.3%, 47/382 vs 17.2%, 580/3379). Fertilization rate was significantly lower with dys-ZP oocytes than with normal-ZP oocytes (72.9%, 401/550 vs 77.2%, 7893/10228). Embryo development and clinical outcomes did not differ between these two groups. In seventeen patients with observed ZP dysmorphology in multiple oocyte retrieval cycles (3 or more times), the normal fertilization rate was low (66.1%, 381/585 vs 77.2%, 7893/10228) and more than 3PN rate, degeneration rate, and non-fertilization rate were high (6.8%, 40/585 vs 3.2%, 327/10228; 8.9%, 52/585 vs 5.0%, 507/10226; 14.7%, 86/585 vs 11.7%, 1194/10228) even in oocytes without ZP dysmorphology, compared to other cycles. It was assumed that the AMH level, the dosage of the gonadotropins, and higher estradiol level may be positively correlated with the incidence of ZP dysmorphology. Since ZP dysmorphology can be associated with lower fertilization rates, embryologists should carefully evaluate ZP morphology during ICSI.

HCG Priming Before Ovary Collection Increasing The Oocyte Quality In The Domestic Cat

Oocyte competence is a determining factor that influences the embryo development. Embryos produced in vitro have a reduced developmental competence than embryos produced in vivo. Therefore, human Chorionic Gonadotropin (hCG) injection was carried out to improve the quality of the oocytes. The objective of this study was to evaluate the effect of ovarian stimulation with hCG before ovary collection on oocyte quality in the domestic cat. Oocyte donors were either 1) treated with a single dose of 200 IU hCG four days before ovary collection (hCG group), or, 2) no treatment before ovary collection (control group). The oocytes were collected by the slicing method. Immature cumulus oophorus complexes (COCs) from both groups were pooled and matured in vitro for 24-26 hours. Then mature oocytes were fertilized with epididymal sperm and cultured in vitro for seven days. The results study showed that the number of the dominant follicle (DF) and the number of COCs in the hCG group was higher than the control group in right and left ovaries (p&lt;0.05). The morulae and blastocyst rates from cleavage embryos were 88% and 75%, respectively. These results demonstrate that hCG priming of oocytes donors before ovary collection improve oocyte quality.

Natural-cycle in vitro fertilization (IVF) combined with in vitro maturation in infertile patients with polycystic ovarian syndrome (PCOS) requiring IVF

ObjectiveIn this study, we report an experience of 59 natural-cycle IVF combined with in vitro oocyte maturation (IVF/M) cycles in patients with PCOS requiring IVF recruited based on limitations to afford a conventional IVF treatment in a 9-years period. Results of IVF/M were compared with 164 cycles of IVF in PCOS patients. Material and methodsIn IVF/M cycles only hCG priming was used before oocyte recovery, with in vitro maturation of immature oocytes in a commercial medium. In conventional IVF group, recombinant FSH (rFSH) and GnRH agonist/antagonist for ovarian stimulation were used. In both groups, fertilization was achieved by intracytoplasmic sperm injection (ICSI) of mature oocytes and fresh embryos transferred at day 2 or day 3. ResultsIn all IVF/M cycles oocytes and transferable quality embryos were obtained, only in 6 IVF/M cycles mature oocytes were obtained at oocyte capture day. Clinical pregnancy rate per cycle was 39.0% vs 53.6% (p = 0.0682) and delivery rate per cycle was 30.5% vs 42.6% (p = 0.1209) in IVF/M and conventional IVF respectively. Patients with ovarian hyperstimulation syndrome (OHSS) were 0% in IVF/M vs 6.7% in conventional IVF (p = 0.0399). ConclusionOur experience in a private clinic in Mexico suggests that IVF/M can be a useful initial strategy to treat PCOS patients requiring IVF with comparable delivery rates to conventional IVF and a decreased risk of ovary hyperstimulation. IVF/M may be indicated to patients with limited resources paying without insurance for their infertility treatment.

Live birth rate after human chorionic gonadotropin priming in vitro maturation in women with polycystic ovary syndrome

BackgroundIn vitro maturation (IVM) has some advantages over conventional in vitro fertilization (IVF), particularly in polycystic ovary syndrome (PCOS) where the risk of ovarian hyperstimulation is high. We studied the live birth rate in a large series of PCOS women undergoing human chorionic gonadotropin (hCG)-priming IVM.MethodsThis retrospective study included women with PCOS aged 18–42 years undergoing IVM with hCG priming. We reported live birth rate after the first embryo transfer and cumulative live birth rate from embryos obtained in the IVM cycle. We also performed logistic regression to assess which factors predicted number of oocytes and live birth.ResultsWe included 921 women (age 28.9±3.5 years, body mass index 21.8±3.1 kg/m2, infertility duration 3.7±2.6 years, 81% primary infertility, 88% first IVF attempt, 94% ovulation induction failure). Live birth rate after the first embryo transfer was 31.7%, with a cumulative live birth rate from the cycle of 33.7%. High anti-Müllerian hormone levels predicted a high number of oocytes and a high oocyte maturation rate while the opposite was the case when luteinizing hormone levels were high.ConclusionsIn women with PCOS, hCG priming IVM was feasible and resulted in acceptable live birth rates.

Ultrasound imaging improves hormone therapy strategies for induction of ovulation and in vitro fertilization in the endangered dusky gopher frog (Lithobates sevosa).

Establishing captive breeding populations of amphibians is an important conservation strategy to safeguard against ongoing declines of wild populations and provide broodstock for reintroduction programs. The endangered dusky gopher frog (DGF) has never naturally reproduced in captivity and requires breeding intervention to sustain the population. Methods for inducing ovulation in female DGFs using hormone therapies have not been evaluated. To address this need, we tested four exogenous hormone treatments to induce ovulation in DGFs (n = 11/treatment), including: treatment (A) gonadotropin-releasing hormone agonist (GnRHa); (B) GnRHa with dopamine antagonist metoclopramide hydrochloride; (C) GnRHa and human chorionic gonadotropin (hCG) and (D) GnRHa with hCG following two low hCG priming doses. Treatments B, C and D resulted in a significantly greater (P < 0.0125) number of ovulating females compared to the control (no hormone); Treatment A was not different from control. For ovulating females, the number of eggs, relative fecundity and cleavage rates of eggs were compared between the four hormone treatments and initial ultrasound grade. Between treatments, there was no difference in number of eggs or relative fecundity; however, Treatments A and D resulted in higher (P < 0.05) cleavage rates than Treatment C, but were not different from Treatment B. Ultrasound imaging was used to assess the ovarian state of DGF females prior to and following hormone therapy. A grading scale (Grades 1-5) was developed to characterize ovarian states. Ultrasound grade was found to be a significant (P = 0.002) predictor for ovulation following hormone treatment, with only high-grade females (Grades 3-4) ovulating in response to hormones. Ultrasound grade did not influence egg numbers or cleavage rate (P > 0.05). Results demonstrate multiple hormone therapies are available for stimulating ovulation in female DGFs and ultrasonography is a valuable tool to inform hormone therapy. Ultimately, these reproductive technologies are critical to enhance breeding and reintroduction efforts for the DGF.

Cumulus cells surrounding oocytes with high developmental competence exhibit down-regulation of phosphoinositol 1,3 kinase/protein kinase B (PI3K/AKT) signalling genes involved in proliferation and survival.

STUDY QUESTIONIs the phosphoinositol 1,3-kinase/protein kinase B (PI3K/AKT) pathway expression profile in cumulus cells (CCs) a potential marker of oocyte competence and predictive of pregnancy outcome?SUMMARY ANSWEREleven genes (AKT1, ARHGEF7, BCL2L1, CCND1, E2F1, HRAS, KCNH2, PIK3C2A, SHC1, SOS1 and SPP1) in the PI3K/AKT pathway were significantly down-regulated in CCs from oocytes that went on to produce a pregnancy compared to CCs associated with a negative outcome.WHAT IS KNOWN ALREADYThe PI3K/AKT pathway plays a pivotal role in the interdependence and continuous feedback between the oocyte and CCs.STUDY DESIGN SIZE, DURATIONThe expression analysis of 92 transcripts in the PI3K/AKT pathway in CCs from patients with negative or positive pregnancy outcome, after single embryo transfer, was performed. Mouse CCs target gene expression was conducted to associate the expression profile of PI3K/AKT pathway to oocyte developmental profile.PARTICIPANTS/MATERIALS, SETTING, METHODSFifty-five good prognosis IVF patients who had been referred to IVF or intracytoplasmic sperm injection treatment for male-factor infertility or tubal disease were enroled. CCs from single cumulus-oocyte complexes (COCs) from 16 patients who underwent a single embryo transfer were analyzed. Twenty-five CD-1 mice were used to assess gene expression in CCs associated with oocytes with different competence in relation to hCG priming. A total 220 human COCs were collected. The RNA extracted from CCs of 16 selected patients was used to analyze PI3K/AKT pathway gene expression employing a 96-well custom TaqMan Array. Expression data of CCs associated to positive IVF outcome were compared to data from negative outcome samples. Mice were sacrificed after 9, 12, 15, 21 and 24 h post-hCG administration to obtain CCs from MII oocytes with different developmental competence. Akt1, Bcl2l2 and Shc1 expression were tested in the collected mouse CCs. In addition, the expression of upstream regulator ESR1, the gene encoding for the oestrogen receptor ERβ, and the downstream effectors of the pathway FOXO1, FOXO3 and FOXO4 was evaluated in human and mouse samples.MAIN RESULTS AND THE ROLE OF CHANCETranscripts involved in the PI3K Signaling Pathway were selectively modulated according to the IVF/ICSI outcome of the oocyte. Eleven transcripts in this pathway were significantly down-regulated in all samples of CCs from oocytes with positive when compared those with a negative outcome. These outcomes were confirmed in mouse CCs associated with oocytes at different maturation stages. Expression data revealed that the down-regulation of ESR1 could be related to oocyte competence and is likely to be the driver of expression changes highlighted in the PI3K/AKT pathway.LIMITATIONS REASONS FOR CAUTIONSmall sample size and retrospective design.WIDER IMPLICATIONS OF THE FINDINGSThe CCs expression profile of PI3K/AKT signaling genes, disclosed a specific CCs gene signature related to oocyte competence. It could be speculated that CCs associated with competent oocytes have completed their role in sustaining oocyte development and are influencing their fate in response to metabolic and hormonal changes by de-activating anti-apoptotic signals.STUDY FUNDING/COMPETING INTEREST(S)Supported by Merck Serono an affiliate of Merck KGaA, Darmstadt, Germany (research grant for the laboratory session; Merck KGaA reviewed the manuscript for medical accuracy only before journal submission. The authors are fully responsible for the content of this manuscript, and the views and opinions described in the publication reflect solely those of the authors). The authors declare no conflict of interest.

Predictive factors for live birth after in vitro maturation of oocytes in women with polycystic ovary syndrome.

In vitro maturation (IVM) of human oocytes can be an alternative treatment option to conventional in vitro fertilization. Women with polycystic ovary syndrome (PCOS) are considered the classical candidates for IVM because of the associated ovarian morphology and because IVM diminishes the risk of developing ovarian hyperstimulation syndrome. The objective of this study was to identify predictive factors for live birth in a cohort of women with PCOS who underwent IVM. This retrospective study included 159 patients with PCOS who had IVM cycles in which single or double embryo transfer was performed. The IVM protocol included three days of gonadotropin ovarian stimulation and hCG priming when the leading follicle size was 10-12mm. Collected cumulus-oocyte complexes were cultured for 24h for maturation. Intracytoplasmic sperm injection (ICSI) was used for fertilization. Embryo transfer was performed two days after fertilization. Demographic and clinical parameters were analyzed with logistic regression to identify predictors for live birth. The women's mean age was 27.4years, the mean number of retrieved oocytes was 14, and the live birth rate was 34.6%. The logistic regression revealed the following significant factors for live birth: infertility duration (OR 0.9; 95% CI, 0.82-0.98), number of collected oocytes (OR 1.56; 95% CI, 1.01-3.2), embryo cell number (OR 2.1; 95% CI, 1.4-3.5), and embryo grade (OR 1.84; 95% CI, 1.13-4.2). Infertility duration, oocyte number, embryo cell number, and embryo grade were the most significant predictors for live birth after IVM in PCOS patients. These prognostic factors can be used when planning treatment or counselling patients.

Role of Low-dose Human Chorionic Gonadotropin in Follicular Phase for Thin Endometrium in Frozen Embryo Replacement Cycles in in vitro Fertilization/Intracytoplasmic Sperm Injection Patients: A Pilot Study

ABSTRACTIntroductionOne of the most challenging problems inin vitro fertilization(IVF) is patient with thin endometrium. The objective of the study was to ascertain whether daily human chorionic gonadotropin (hCG) for 7 days with estrogen in hormone replacement frozen embryo transfer (FET) cycles during follicular phase can increase the endometrial thickness (ET) and reduce the cancellation of cycles.Materials and methodsTwenty-five infertile patients with resistant thin endometrium who had antagonist protocol and planned for frozen embryo replacement were recruited. These patients had prior attempts to thicken their endometrium which had failed. All the patients received estrogen daily from D2/3 of cycle. On day 8 or 9 of estrogen administration, 200 IU of hCG was given daily for 7 days. After 7 days on hCG priming (D14/15), ET was measured and progesterone was started accordingly. Identification of an intrauterine gestational sac with fetal heart beat by transvaginal ultrasonography constituted clinical pregnancy.ResultsMean ET increased significantly from 5.84 to 7.61 mm (p &lt; 0.01). About 72% of patients had more than 20% improvement in their ET after hCG priming. About 76% achieved an ET more than 7 mm. Overall, 50% became pregnant. The ongoing pregnancy rate was 40%.ConclusionA total of 200 IU hCG endometrial priming for 7 days in the proliferative phase of hormone replacement cycles for FET is a highly promising approach to thicken thin endometrium with failed prior attempts.How to cite this articleNigam S, Shankar K, Varma TR. Role of Low-dose Human Chorionic Gonadotropin in Follicular Phase for Thin Endometrium in Frozen Embryo Replacement Cycles inin vitro Fertilization/Intracytoplasmic Sperm Injection Patients: A Pilot Study. Int J Infertil Fetal Med 2017;8(3):101-105.

Human chorionic gonadotrophin priming for fertility treatment with in vitro maturation.

In vitro maturation (IVM) is a fertility treatment that involves the transvaginal retrieval of immature oocytes, and their subsequent maturation and fertilisation. Although the live birth rate is lower than conventional in vitro fertilisation (IVF) with ovarian stimulation, it is a useful treatment, as it avoids the risk of ovarian hyperstimulation syndrome (OHSS). Women with polycystic ovaries (PCO) or polycystic ovarian syndrome (PCOS) are at an increased risk of OHSS. Thus, IVM may be a more useful treatment in this patient group.Strategies to maximise the maturation rates of the immature oocytes are important. This review focuses on the administration of human chorionic gonadotrophin (hCG) prior to immature oocyte retrieval. To determine the effectiveness and safety of hCG priming in subfertile women who are undergoing IVM treatment in the context of assisted reproduction. We searched the following electronic databases up to 29 August 2016: Cochrane Gynaecology and Fertility Group Specialised Register of controlled trials, CENTRAL, MEDLINE, Embase, PsycINFO, and CINAHL. We also searched the trial registries ClinicalTrials.gov and WHO ICTPR to identify ongoing and registered trials. We sought recently published papers not yet indexed in the major databases, and reviewed the reference lists of reviews and retrieved studies as sources of potentially relevant studies. There were no language restrictions. We included randomised controlled trials (RCTs) that compared hCG priming with placebo or no priming in women undergoing IVM. We also included RCTs that compared different doses of hCG, or the timing of oocyte retrieval. The primary outcomes were live birth rate and miscarriage rate per woman randomised. Two review authors independently selected studies for inclusion, and with a third author, assessed risk of bias and extracted data. We contacted the original authors where data were missing. For dichotomous outcomes, we used the Mantel-Haenszel method to calculate odds ratios (OR). For continuous outcomes, we calculated the mean differences (MD) between treatment groups. We assessed statistical heterogeneity using the I² statistic. We assessed the overall quality of the evidence using GRADE methods. We included four studies, with a total of 522 women, in the review. One of these studies did not report outcomes per woman randomised, and so was not included in formal analysis. Three studies investigated 10,000 units hCG priming compared to no priming. One study investigated 20,000 units hCG compared to 10,000 units hCG priming. Three studies only included women with PCOS (N = 122), while this was an exclusion criteria in the fourth study (N = 400).We rated all four studies as having an unclear risk of bias in more than one of the seven domains assessed. The quality of the evidence was low, the main limitations being lack of blinding and imprecision.When 10,000 units hCG priming was compared to no priming, we found no evidence of a difference in the live birth rates per woman randomised (OR 0.65, 95% confidence intervals (CI) 0.24 to 1.74; one RCT; N = 82; low quality evidence); miscarriage rate (OR 0.60, 95% CI 0.21 to 1.72; two RCTs; N = 282; I² statistic = 21%; low quality evidence), or clinical pregnancy rate (OR 0.52, 95% CI 0.26 to 1.03; two RCTs, N = 282, I² statistic = 0%, low quality evidence). Though inconclusive, our findings suggested that hCG may be associated with a reduction in clinical pregnancy rates; 22% of women who received no priming achieved pregnancy, while between 7% and 23% of women who received hCG priming did so.The study comparing 20,000 units hCG with 10,000 units hCG did not report sufficient data to enable us to calculate odds ratios.No studies reported on adverse events (other than miscarriage) or drug reactions. This review found no conclusive evidence that hCG priming had an effect on live birth, pregnancy, or miscarriage rates in IVM. There was low quality evidence that suggested that hCG priming may reduce clinical pregnancy rates, however, these findings were limited by the small number of data included. As no data were available on adverse events (other than miscarriage) or on drug reactions, we could not adequately assess the safety of hCG priming. We need further evidence from well-designed RCTs before we can come to definitive conclusions about the role of hCG priming, and the optimal dose and timing.

What threshold values of antral follicle count and serum AMH levels should be considered for oocyte cryopreservation after in vitro maturation?

What threshold values of ultrasonographic antral follicle count (AFC) and serum anti-Müllerian hormone (AMH) levels should be considered for ensuring the cryopreservation of sufficient number of in vitro matured (IVM) oocytes, in cancer patients seeking fertility preservation (FP)? AFC and serum AMH values >20 follicles and 3.7 ng/ml, respectively, are required for obtaining at least 10 IVM oocytes for cryopreservation. IVM of cumulus oocyte complexes (COCs) followed by oocyte cryopreservation has emerged recently as an option for urgent FP. Recent data have reported that, in healthy patients, 8-20 cryopreserved oocytes after ovarian stimulation would maximize the chance of obtaining a live birth. Although both AFC and AMH have been reported as predictive factors of IVM success in infertile patients with polycystic ovary syndrome (PCOS), there is a dramatic lack of data regarding the values of these parameters in oncological patients as candidates for FP. From January 2009 to April 2015, we prospectively studied 340 cancer patients, aged 18-41 years, as candidates for oocyte cryopreservation following IVM. All patients had AFC and AMH measurements, 48-72 h before oocyte retrieval, regardless of the phase of the cycle. COCs were recovered under ultrasound guidance 36 h after hCG priming. Logistic regression allowed the determination of threshold values of AFC and AMH, for obtaining at least 8, 10 or 15 matures oocytes frozen after the IVM procedure. Similar analyses were performed for a final number of mature oocytes ≤2. Among the 340 cancer patients included, 300 were diagnosed with breast cancers, 14 had hematological malignancies and 26 underwent the procedure for others indications. Overall, the mean age of the population was 31.8 ± 4.5 years. Mean AFC and serum AMH levels were 21.7 ± 13.3 follicles and 4.4 ± 3.8 ng/ml, respectively. IVM was performed in equal proportions during the follicular or luteal phase of the cycle (49 and 51%, respectively). Statistical analysis showed that AFC and AMH values above 28 follicles and 3.9 ng/ml, 20 follicles and 3.7 ng/ml and 19 follicles and 3.5 ng/ml are required, respectively, for obtaining at least 15, 10 or 8 frozen IVM oocytes with a sensitivity ranging from 0.82 to 0.90. On the contrary, ≤2 IVM oocytes were cryopreserved when AFC and AMH were <19 follicles and 3.0 ng/ml, respectively. Although the potential of cryopreserved IVM oocytes from cancer patients remains unknown, data obtained from infertile PCOS women have shown a dramatically reduced competence of these oocytes when compared with that of oocytes recovered after ovarian stimulation. As a consequence, the optimal number of IVM oocytes frozen in candidates for FP is currently unpredictable. Cryopreservation of oocytes after IVM should be considered in the FP strategy when ovarian stimulation is unfeasible, in particular when markers of the follicular ovarian status are at a relatively high range. Further investigation is needed to objectively assess the real potential of these IVM oocytes after cryopreservation. Therefore, even when a good COCs yield is expected, we should systematically encourage IVM in combination with ovarian tissue cryopreservation. No external funding was obtained for the present study. The authors have no conflict of interest to declare. Not applicable.

MiR-125b regulates endometrial receptivity by targeting MMP26 in women undergoing IVF-ET with elevated progesterone on HCG priming day.

On the women undergoing IVF-ET with elevated progesterone on human chorionic gonadotrophin priming, the assisted reproductive technology outcome is poor. But, due to the unknown mechanism of this process, no effective method has been found to overcome this difficulty. Here, we investigated the roles of miR-125b and its target gene, MMP26, in endometrial receptivity (ER) in these women. The expression of miR-125b was significantly up-regulated in EECs in women with elevated progesterone during the window of implantation, and it showed a progesterone-dependent effect in vitro. Similarly, the expression of miR-125b was significantly up-regulated in the preimplantation period, and was down-regulated in the implantation period and the post-implantation period in mouse EECs. In addition, miR-125b showed a greater decrease at implantation sites than it did at interimplantation sites. The luciferase report assay demonstrated that MMP26 is a target gene of miR-125b. And the expression profile of MMP26 showed an inverse relationship with miR-125b in vivo and in vitro. Overexpression of miR-125b in human EECs inhibited cell migration and invasion. Gain-of-function of miR-125b induced a significant decrease in the number of implantation sites. In conclusion, these data shed new light on how miR-125b triggers ER decline through the regulation of MMP26 function.

In Vitro Maturation in Women with vs. without Polycystic Ovarian Syndrome: A Systematic Review and Meta-Analysis.

ObjectiveTo evaluate in vitro maturation (IVM) in sub-fertile women with polycystic ovarian syndrome (PCOS) undergoing in vitro fertilisation (IVF), by comparing outcomes with a control group of non-PCOS.Study designA search strategy was developed for PubMed and studies reporting rates of the following outcomes (live birth; clinical pregnancy; implantation; cycle cancellation; oocyte maturation; oocyte fertilization; miscarriage) between patients with PCOS, PCO and controls undergoing IVM were deemed eligible. The review was conducted in accordance to the PRISMA guidelines and included studies quality was assessed through the Newcastle-Ottawa Quality scale. ORs with their corresponding 95% CIs were calculated for the main analysis and subgroup analyses were performed for PCOS cases vs. controls and PCOS vs. PCO cases. Alternative analyses were performed for live birth and clinical pregnancy, based on cycles and on women. Subgroup analyses for FSH stimulation, hCG priming and type of procedure (IVF/ICSI) were undertaken for all meta-analyses encompassing at least four study arms. Random effects models were used to calculate pooled effect estimates.ResultsEleven studies were identified. A total of 268 PCOS patients (328 cycles), 100 PCO patients (110 cycles) and 440 controls (480 cycles) were included in the meta-analysis. A borderline trend towards higher birth rates among PCOS patients emerged (pooled OR = 1.74, 95%CI: 0.99–3.04) mainly reflected at the subgroup analysis vs. controls. Clinical pregnancy (pooled OR = 2.37, 95%CI: 1.53–3.68) and implantation rates (pooled OR = 1.73, 95%CI: 1.06–2.81) were higher, while cancellation rates lower (pooled OR = 0.18, 95%CI: 0.06-0.47) among PCOS vs. non-PCOS subjects; maturation and miscarriage rates did not differ between groups, while a borderline trend towards lower fertilization rates among PCOS patients was observed.ConclusionThe present meta-analysis provides preliminary evidence on the effectiveness of IVM as a treatment option when offered in sub-fertile PCOS women, as the latter present at least as high outcome rates as those in non-PCOS.

Human in vitro oocyte maturation is not associated with increased imprinting error rates at LIT1, SNRPN, PEG3 and GTL2

Does in vitro maturation (IVM) of cumulus-enclosed germinal vesicle (GV) stage oocytes retrieved from small antral follicles in minimally stimulated cycles without an ovulatory hCG dose induce imprinting errors at LIT1, SNRPN, PEG3 and GTL2 in human oocytes? There is no significant increase in imprinting mutations at LIT1, SNRPN, PEG3 and GTL2 after IVM of cumulus-enclosed GV oocytes from small antral follicles in minimally stimulated cycles without hCG priming. Animal models have generally demonstrated correct methylation imprint establishment for in vitro grown and matured oocytes. For human IVM, well-designed studies allowing conclusions on imprint establishment are currently not available. Immature oocyte-cumulus complexes from 2 to 9 mm follicles were retrieved in polycystic ovary syndrome (PCOS) subjects in minimally stimulated cycles without hCG priming and matured in vitro. In vivo grown oocytes were retrieved after conventional ovarian stimulation for IVF/ICSI or after ovulation induction. Imprinting error rates at three maternally methylated (LIT1, SNRPN and PEG3) and one paternally methylated (GTL2) imprinted genes were compared in 71 in vitro and 38 in vivo matured oocytes. The limiting dilution bisulfite sequencing technique was applied, allowing increased sensitivity based on multiplex PCR for the imprinted genes and the inclusion of non-imprinted marker genes for cumulus cell DNA contamination. In vitro as well as in vivo matured oocytes showed only a few abnormal alleles, consistent with epimutations. The abnormalities were more frequent in immature than in mature oocytes for both groups, although no significant difference was reached. There was no statistically significant increase in imprinting errors in IVM oocytes. This single cell methylation analysis was restricted to a number of well-selected imprinted genes. Genome-wide methylation analysis of single human oocytes is currently not possible. IVM is a patient-friendly alternative to conventional ovarian stimulation in PCOS patients and is associated with reduced gonadotrophin costs and avoidance of OHSS. The results of this study show for the first time that optimized human IVM procedures have no significant effects on the establishment of maternal DNA methylation patterns at LIT1, SNRPN, PEG3 and GTL2. This study was supported by research funds from Agentschap voor Innovatie door Wetenschap en Technologie (IWT-TBM 110680), Wetenschappelijk Fonds Willy Gepts (WFWG 2011) and German Research Foundation (HA 1374/12-2). There are no competing interests.

Changes in coagulation and fibrinolytic indices in women with polycystic ovarian syndrome undergoing controlled ovarian hyperstimulation.

Background. Polycystic ovarian syndrome (PCOS) women undergoing in vitro fertilization and embryo transfer (IVF-ET) treatment always attain a low cumulative pregnancy rate disaccording with the satisfactory number of oocytes. Objective. We aim to evaluate the status of coagulation and fibrinolytic system in PCOS patients undergoing controlled ovarian hyperstimulation (COH) process. Method. Of the 97 women, 30 patients with PCOS composed the study group; 67 women of child-bearing age with normal endocrine function composed the control group. All participants underwent GnRH agonist standard long protocol, and plasma HCY, FVIII, FX, and D-dimer levels as well as hormone parameters were measured at day of full downregulation, hCG priming, and embryos transfer. Results. On day of full downregulation, FX levels were significantly higher in PCOS group (P < 0.01). On hCG priming day, FX and estrogen levels in PCOS group were higher than in the control group and FVIII levels were significantly lower on day of embryos transfer whereas FX and E2 levels were significantly higher in PCOS group. Conclusion. Hypercoagulable state during peri-implantation phase would probably lead to poor microcirculation of endometrium and be one of the most important disadvantages of successful implantation and subsequent clinical pregnancy.