Early embryo morphokinetics is a better predictor of post-ICSI live birth than embryo morphology: speed is more important than beauty at the cleavage stage.

Given the importance of embryo developmental competence assessment in reproductive medicine and biology, the aim of this study was to compare the performance of fertilization and cleavage morphokinetics with embryo morphology to predict post-ICSI live birth. Data from embryos cultured in a time-lapse microscopy (TLM) incubator and with known live birth outcomes (LB: embryos achieving live birth, n = 168; NLB: embryos not achieving live birth, n = 1633) were used to generate receiver operating characteristic (ROC) curves based on morphokinetic or morphological scores, and the respective areas under the curve (AUC) were compared. The association between live birth and 12 combinations of four morphokinetic quality degrees (A-D) with three morphological quality degrees (A-C) was assessed using multivariate analysis. Morphokinetic parameters from tPNa to t8 were reached earlier in LB compared with NLB embryos. The ROC curve analysis indicated that morphokinetic information is more accurate than conventional morphology to predict live birth [AUC = 0.64 (95% CI 0.58-0.70) versus AUC = 0.58 (95% CI 0.51-0.65)]. The multivariate analysis was in line with AUCs, revealing that embryos with poor morphokinetics, independently of their morphology, provide lower live birth rates (P < 0.001). A considerable percentage of embryos with top morphology presented poor morphokinetics (20.10%), accompanied by a severely reduced live birth rate in comparison with embryos with top morphology and morphokinetics (P < 0.001). In conclusion, TLM-derived early morphokinetic parameters were more predictive of live-birth achievement following ICSI than conventional morphology.

Maternal age affects the relationship of basal FSH and anti-Müllerian hormone concentrations with post-ICSI/IVF live birth.

RESEARCH QUESTION: Does the association of basal FSH and anti-Müllerian hormone (AMH) concentrations with post-IVF/intracytoplasmic sperm injection (ICSI) live birth change with maternal age? DESIGN: A total of 2003 IVF/ICSI patients were stratified according to basal FSH/AMH in concordant favourable (CF; AMH >1 ng/ml and FSH ≤10 IU/l), concordant unfavourable (CU; AMH ≤1 ng/ml and FSH >10 IU/l), discordant with favourable AMH (DFA) and discordant with favourable FSH (DFF) groups, as well as according to age in pre-advanced maternal age (pre-AMA; <35), AMA-1 (≥35, ≤37), AMA-2 (>37, ≤40) and AMA-3 (>40). IVF/ICSI outcomes were compared among CF, CU, DFA and DFF groups, and the association of basal FSH and AMH concentrations with live birth was tested by univariate and multivariate analysis in total, pre-AMA and AMA groups, separately. RESULTS: Different outcome patterns were observed in discordant AMH/FSH groups from different age categories; favourable basal FSH concentrations were associated with higher delivery rates in pre-AMA patients, but with lower delivery rates in AMA groups. Within pre-AMA patients, DFF patients presented higher delivery rates but lower oocyte yield compared with DFA patients. In the univariate analysis, favourable AMH (P < 0.02) and oocyte yield (P < 0.002) were positively associated with live birth in all AMA groups. The multivariate analysis revealed that favourable basal FSH, but not AMH or oocyte yield, is associated with live birth in pre-AMA patients independently of other variables (P = 0.012). CONCLUSIONS: The relationship of basal FSH and AMH with IVF/ICSI success changes with maternal age; basal FSH better reflects clinical outcomes probably determined by oocyte quality in pre-AMA patients, while AMH better suits AMA patients.

Faster fertilization and cleavage kinetics reflect competence to achieve a live birth after intracytoplasmic sperm injection, but this association fades with maternal age.

OBJECTIVE: To assess the relationship of early developmental kinetics with competence to provide a live birth and the impact of maternal age in this context. DESIGN: Retrospective cohort study including 4,915 embryos, of which 1,390 were transferred and provided a clinical outcome paired with morphokinetic data; 168 of them resulted in a live birth (LB), and 1,222 did not (NLB). Early morphokinetic parameters were compared between LB and NLB embryos from patients stratified into two age groups (<37 and ≥37 years), and between embryos at the same competence group from patients aged <37 and ≥37 years. The association of morphokinetic parameters with live birth was tested by univariate and multivariate analyses. SETTING: Fertility clinic. PATIENT(S): The study population included 1,066 patients undergoing autologous intracytoplasmic sperm injection cycles with fresh single (SET), double (DET) or triple (TET) embryo transfers on day 2 or 3. Of them, 669 patients produced NLB embryos and 134 produced LB embryos. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Fertilization and cleavage morphokinetic parameters and live birth. RESULT(S): In the total patient population, all morphokinetic parameters were achieved earlier in LB compared with NLB embryos. The same was observed in patients aged <37 years (P<.015), but not ≥37 years. Except for the t8 (time at which an 8-blastomere embryo was identified), all morphokinetic parameters were reached earlier in LB embryos from patients aged <37 years compared with LB embryos from patients aged ≥37 years. Univariate analysis revealed that earlier occurrence of all morphokinetic parameters was associated with live birth, although only earlier t2 (time at which two separate and distinct cells were identified) was associated with live birth independently from maternal age in the multivariate analysis. CONCLUSION(S): Despite its retrospective nature and performance in a single IVF center, this study presents novel data indicating that embryos competent to provide a live birth display overall faster early developmental kinetics compared with embryos that do not achieve a live birth after transfer, a difference that, however, narrows as maternal age advances. The findings suggest that fertilization and cleavage morphokinetic parameters may constitute valuable references for embryo selection strategies aiming to improve live birth rates, specifically before advanced maternal age while holding limited usefulness in advanced maternal age.

GnRH agonists to sustain the luteal phase in antagonist IVF cycles: a randomized prospective trial.

BACKGROUND: The addition of a GnRH analogue to the luteal phase in in vitro fertilization programs has been seldom proposed due to the presence of GnRH receptors in the endometrium. The aim of the study was to evaluate the effect of triptorelin addition in short antagonist cycles, compared to cycles where the only supplementation was progesterone. METHODS: The primary objective of this study was the study of the effect of Triptorelin addiction during the luteal phase on the live birth rate. Secondary objectives of efficacy were pregnancy rates and implantation rates, as well as safety in terms of OHSS risks. The study was a prospective, randomized, open study, performed in two independent Centers from July 2013 to October 2015. Patients were divided into three groups: a) Regular antagonist protocol, with only luteal progesterone; b) Antagonist protocol with luteal triptorelin as multiple injections, c) Antagonist protocol with luteal triptorelin as single bolus. Descriptive statistics were obtained for all the parameters. Mean and standard deviation were used for all quantitative parameters. Differences between percentages were studied using Chi-square test generalized to the comparison of several proportions. RESULTS: A total number of 1344 patients completed the study, 786 under the age of 35 years, and 558 over 35 years. It was observed an increase of positive HCG results, Clinical pregnancy rates and Delivery rates when triptorelin was added in the luteal phase, irrespective whether as a single bolus or five injections. This increase was statistically significant both for pregnancy rates and delivery rates. The statistic difference between pregnancies and deliveries obtained with or without luteal triptorelin reached p < 0,01. No increase of OHSS risk was observed. CONCLUSIONS: From this large study it appears that the concept of luteal phase supplementation should be revisited. From our study it appears that triptorelin addition to the luteal phase of antagonist cycles, either as a single bolus or using multiple injections, is a good tool to optimize ART results. TRIAL REGISTRATION: The study was approved by the Ethics Committee of Provincia di Bergamo (n 1203/2013).

Retrospective analysis of treatments with recombinant FSH and recombinant LH versus human menopausal gonadotropin in women with reduced ovarian reserve.

PURPOSES: The aim of this study is to determine whether a clinical advantage is gained with use of LH in combination with FSH or as a component of human menopausal gonadotropin (hMG) to achieve optimal ovarian stimulation. METHODS: In this study, we compared retrospectively two regimens, r-FSH/r-LH and hMG, for the treatment of women with reduced ovarian reserve, identified as subjects with antral follicle count (AFC) < 11 and AMH ≤ 1.1 ng/ml. RESULTS: Overall, the clinical pregnancy per started cycle was higher in the r-FSH/r-LH group (12.5 vs. 8.1%, P < 0.02), while implantation (11.1 vs. 9.5%) and miscarriage rates (29.9 vs. 35.9%) were comparable. Data were further analysed performing separate comparisons in subpopulations with different ranges of AFC, i.e. < 4, 4-6 and 7-10. Major differences between the two regimens were observed in women with AFC < 4. In this subpopulation, not only was the clinical pregnancies per started cycle higher in the r-FSH/r-LH group (10.2 vs. 1.5%, P < 0.01), but also implantation was significantly higher (13.0 vs. 2.8%, P < 0.02). CONCLUSIONS: A r-FSH/r-LH regimen appears to be beneficial for the treatment of women with extremely poor ovarian reserve. It should be considered however that, being retrospective, this study is affected by obvious limitations, such as post-treatment patient selection criteria and absence of randomisation.

Morphokinetics of embryos developed from oocytes matured in vitro.

PURPOSE: In in vitro maturation (IVM) cycles primed with human chorionic gonadotropin (hCG), both immature and mature oocytes are retrieved from antral follicles sized 8-12 mm. Using time-lapse microscopy, we compared the morphokinetic behavior of embryos developed from oocytes matured in vivo and in vitro, testing the hypothesis that IVM affects preimplantation development. Furthermore, we extended the morphokinetic analysis of these embryos by a comparison with embryos obtained in stimulated assisted reproduction technology (ART) cycles. METHODS: In IVM cycles primed with follicle-stimulating hormone (FSH)/hCG, prior to sperm microinjection, oocytes surrounded by an expanded cumulus at retrieval and presumably mature (EC-MII) were incubated for 6 h, while immature oocytes enclosed in a compact cumulus (CC) were matured in vitro for 30 h. The morphokinetics of embryos selected for transfer or cryopreservation, derived from EC-MII and CC oocytes, were comparatively and retrospectively analyzed in terms of cleavage times (t2, t3, t4, t5, and t8) and intervals (cc2, cc3, s2, s3). For further comparison, the morphokinetics of embryos selected for transfer or cryopreservation (ICSI) or giving rise to ongoing pregnancies (model) in stimulated ART cycles was also assessed. RESULTS: The morphokinetic behavior of EC-MII and CC embryos was entirely comparable, as suggested by the absence of statistical differences in the averages of all cleavage times and intervals. Almost all cleavage and interval times were also similar between EC-MII, CC, ICSI, and model groups, with the exception of t4 and s2, which were delayed and longer, respectively, in embryos generated in IVM cycles (EC-MII and CC). CONCLUSIONS: These findings do not support the hypothesis that maturation in vitro affects embryo morphokinetics, while they suggest only marginal differences in the morphokinetics of embryos developed from oocytes matured in vivo and in vitro in IVM cycles and embryos developed from mature oocytes recovered in stimulated cycles.

Clinical outcomes from mature oocytes derived from preovulatory and antral follicles: reflections on follicle physiology and oocyte competence.

PURPOSE: The aim of this retrospective study was to compare the competence of oocytes obtained from preovulatory and antral follicles. METHODS: Mature oocytes from preovulatory follicles were retrieved from women selected for standard IVF treatment (Group A). Mature oocytes from antral follicles were recovered from women undergoing hCG-primed in vitro maturation (IVM) treatment (Group B). Patients groups were matched for age, BMI, FSH, AMH and antral follicle count (AFC) values. In vivo matured oocytes from both groups were microinjected and resulting embryos were culture and selected on day 3 for embryo transfer. RESULTS: Oocyte pick-ups (OPU) were 315 and 204 in Groups A and B, respectively. Fertilization rates were comparable (72.8 and 75.9 %, respectively; P = 0.137). In Group A, in which the average number of embryos transferred was higher, clinical pregnancy rates per OPU (37.5 %) and embryo transfer (38.4 %) were superior in comparison to Group B (27.0 %, P = 0.013; 29.4 %, P = 0.041; respectively). On the other hand, implantation rates (Group A, 23.7 %; Group B, 20.8 %) and proportions of babies born per transferred embryo (Group A, 19.5 %; Group B, 16.9 %) were similar (P = 0.528 and 0.332, respectively). CONCLUSIONS: Overall, this suggests that oocyte competence is already achieved at the antral stage of follicle development.

Oocyte in vitro maturation in normo-ovulatory women.

OBJECTIVE: To review the scientific literature concerning the application of oocyte in vitro maturation (IVM) as a treatment for normo-ovulatory women. DESIGN: Critical evaluation and comparison of the most relevant clinical studies. SETTING: Public IVF unit. PATIENT(S): Normo-ovulatory women. INTERVENTION(S): Oocyte IVM and cryopreservation. MAIN OUTCOME MEASURE(S): Maturation, fertilization, implantation, and pregnancy rates. RESULT(S): Maturation rates in vitro are suboptimal and influenced by several factors. IVM oocytes fertilize and undergo development in vitro with rates similar to in vivo matured control oocytes. In IVM cycles implantation and pregnancy rates are lower compared with controlled ovarian stimulation treatments, but accurate patient selection can improve IVM clinical outcome. CONCLUSION(S): In normal responders, IVM does not reach success rates similar to conventional ovarian stimulation regimens. However, it represents a milder approach to assisted reproduction treatment and an alternative intervention for specific conditions.

Clomiphene citrate versus high doses of gonadotropins for in vitro fertilisation in women with compromised ovarian reserve: a randomised controlled non-inferiority trial.

BACKGROUND: The aim of the present randomised controlled non-inferiority trial is to test whether in women with compromised ovarian reserve requiring in vitro fertilisation, a protocol of ovarian stimulation using exclusively clomiphene citrate performs similarly to a regimen with high doses of gonadotropins. METHODS: Women with day 3 serum FSH > 12 IU/ml on at least two occasions or previous poor response to hyper-stimulation were recruited at four Italian infertility units. Selected women were allocated to clomiphene citrate 150 mg/day from day 3 to day 7 of the cycle (n=145) or to a short protocol with GnRH agonist 0.1 mg and recombinant FSH 450 IU daily (n=146). They were randomised by means of a computer-generated list into two groups. The study was not blinded. The main outcome of the study was the delivery rate per started cycle. RESULTS: The study was interrupted after the scheduled two years of recruitment before reaching the sample size. 148 women were allocated to clomiphene citrate and 156 to the short protocol with high doses of gonadotropins; 124 and 125 participants were analysed in the groups, respectively. Women allocated to high doses of gonadotropins retrieved more oocytes and had a higher probability to perform embryo-transfer. However, the chances of success were similar. The delivery rate per started cycle in women receiving clomiphene citrate and high-dose gonadotropins was 3% (n=5) and 5% (n=7), respectively (p=0.77). The mean estimated cost per delivery in the two groups was 81,294 and 113,107 Euros, respectively. No side-effects or adverse events were observed. CONCLUSIONS: In women with compromised ovarian reserve selected for in vitro fertilisation, ovarian stimulation with clomiphene citrate or high-dose gonadotropins led to similar chances of pregnancy but the former is less expensive. TRIAL REGISTRATION: Trial registered on http://www.clinicaltrials.gov (NCT01389713).

Deep pelvic endometriosis negatively affects ovarian reserve and the number of oocytes retrieved for in vitro fertilization.

OBJECTIVE: Endometriosis-associated infertility results in reduced ovarian response, fewer oocytes available for fertilization, compromised oocyte quality and higher miscarriage rates. A consistent proportion of women with endometriosis require in vitro fertilization. We sought to clarify the impact of deep infiltrating pelvic disease on antral follicle count and ovarian response to follicle-stimulating hormone (FSH) stimulation in patients with severe endometriosis. DESIGN: Retrospective cohort study. SETTING: University hospital. POPULATION: Patients with severe endometriosis (stages III-IV; n=51) were divided into two groups regarding localization of endometriosis during surgical staging: ovarian (n=27) and both ovarian and deep infiltrating disease (n=24). METHODS: A total of 73 long-protocol ovulation induction cycles with recombinant FSH for an intracytoplasmic sperm injection program were given. On day 3 of the cycle, measurements of FSH and luteinizing hormone and an ultrasound evaluation of antral follicle count were performed. MAIN OUTCOME MEASURES: Number of oocytes collected at ovum pick up, number of mature oocytes, number of embryos transferred and clinical pregnancy rate. RESULTS: Ovarian reserve in terms of antral follicle count was damaged in both groups but, if adjusted for age, it was significantly lower in the ovarian and pelvic infiltrating group compared with patients having only ovarian endometriosis. Pelvic deep infiltrating disease significantly impacted on the number of oocytes collected at pick up when adjusted for age. CONCLUSIONS: Deep infiltrating pelvic disease can negatively affect ovarian reserve in terms of antral follicle count and number of oocytes retrieved. Mechanisms underlying this phenomenon need to be elucidated.

Myo-inositol in patients with polycystic ovary syndrome: a novel method for ovulation induction.

BACKGROUND: Polycystic ovary syndrome (PCOS) is often characterized by chronic oligo- or anovulation (usually manifested as oligo- or amenorrhea), and hyperandrogenism. In addition, 30-40% of PCOS women have impaired glucose tolerance, and a defect in the insulin signaling pathway (inositol-containing phosphoglycan mediators) seems to be implicated in the pathogenesis of insulin resistance. PCOS patients are subfertile as a consequence of such ovulatory disorders and often need drugs, such as clomiphene citrate or follicle-stimulating hormone, for ovulation induction, which increases the risk of multiple pregnancy and ovarian hyperstimulation syndrome. We hypothesized that the administration of an isoform of inositol (myo-inositol), belonging to the vitamin B complex, would improve the insulin-receptor activity, restoring normal ovulatory function. MATERIALS AND METHODS: Twenty-five PCOS women of childbearing age with oligo- or amenorrhea were enrolled in the study. Ovulatory disorder due to PCOS was apparently the only cause of infertility; no tubal defect or deficiency of male semen parameters was found. Myo-inositol combined with folic acid (Inofolic) 2 g twice a day was administered continuously. During an observation period of 6 months, ovulatory activity was monitored with ultrasound scan and hormonal profile, and the numbers of spontaneous menstrual cycles and eventually pregnancies were assessed. RESULTS: Twenty-two out of the 25 (88%) patients restored at least one spontaneous menstrual cycle during treatment, of whom 18 (72%) maintained normal ovulatory activity during the follow-up period. A total of 10 singleton pregnancies (40% of patients) were obtained. Nine clinical pregnancies were assessed with fetal heart beat at ultrasound scan. Two pregnancies evolved in spontaneous abortion. CONCLUSION: Myo-inositol is a simple and safe treatment that is capable of restoring spontaneous ovarian activity and consequently fertility in most patients with PCOS. This therapy did not cause multiple pregnancy.

Natural cycle as first approach in aged patients with elevated follicle-stimulating hormone undergoing intracytoplasmic sperm injection: a pilot study.

BACKGROUND: Poor ovarian response to standard in vitro fertilization-embryo transfer (IVF-ET) protocols or different regimens of treatment, as consequence of a diminished ovarian reserve, correlates strictly with patient age, elevated follicle-stimulating hormone (FSH) and reduced antral follicle count. The aim of the present pilot study was to evaluate the outcome of patients with poor prognostic features undergoing IVF-ET with natural cycles as a first approach and not as a consequence of a previous failure treatment. MATERIALS AND METHODS: Eighteen aged patients (mean +/- standard deviation 40.2 +/- 0.7 years, range 37-43 years) with elevated serum FSH and reduced antral follicle count underwent intracytoplasmic sperm injection (ICSI) after spontaneous ovulation. RESULTS: A total of 26 natural cycles with ICSI were analyzed. Pregnancy was observed in three patients, of which two were ongoing as assessed by fetal heart beat at ultrasound scan performed 4-5 weeks after ET. CONCLUSION: The overall pregnancy rates achieved (11.5% per cycle, 20.0% per ET) are comparable with those of conventional IVF-ET in aged patients, and not impaired by a single embryo transferred. Better embryo quality, as a consequence of natural selection of oocytes, better endometrium receptivity and monthly repeatability of the procedure, can balance the relatively low chance to perform ET.

Pathologic findings in hysteroscopy before in vitro fertilization-embryo transfer (IVF-ET).

BACKGROUND: The aim of this study was to evaluate hysteroscopy routinely performed prior to in vitro fertilization-embryo transfer (IVF-ET). METHODS: We analyzed in a prospective study 300 patients who underwent hysteroscopy before the first IVF-ET cycle. We analyzed then in a retrospective manner 300 patients who did not perform hysteroscopy. RESULTS: One-hundred-and-eighty (60%) hysteroscopies were normal but 120 (40%) revealed an unsuspected intrauterine abnormality. We did not find statistically significant differences between patients with normal or abnormal hysteroscopy in any characteristic. We found a statistically significant difference in pregnancy rate between women who performed hysteroscopy before IVF-ET cycle and in women who did not perform it. CONCLUSIONS: Hysteroscopy, as a routine examination, should be performed before the first IVF-ET cycle in all patients.