Effect of small follicles on clinical pregnancy and multiple pregnancy rates in intrauterine insemination: a cohort study.

STUDY QUESTION: What is the effect of small follicles on clinical pregnancy and multiple pregnancy rates in women undergoing IUI with ovarian stimulation (IUI-OS)? SUMMARY ANSWER: The presence of ≥2 small follicles with a diameter of 10-12 or 12-14 mm was associated with an increased chance of clinical pregnancy and the presence of any 12-14 mm or larger follicles, but not smaller follicles, was statistically significantly associated with an increased risk for multiple pregnancy. WHAT IS KNOWN ALREADY: IUI-OS is widely used as the first-line treatment for unexplained or mild male factor infertility. However, IUI is associated with the risk of multiple pregnancy. While the positive association between the number of follicles ≥14 mm and the chance of pregnancy and the risk of multiple pregnancy is known, the impact of smaller follicles is uncertain. STUDY DESIGN, SIZE, DURATION: This was a retrospective cohort study that included women undergoing IUI cycles from January 2007 to May 2021 in one assisted reproduction center. PARTICIPANTS/MATERIALS, SETTING, METHODS: We studied the impact of the number and size of follicles on trigger day on clinical pregnancy and multiple pregnancy rates. Generalized estimation equation regression models were used to compute unadjusted and adjusted odds ratios and 95% CI in all women and only women who achieved clinical pregnancy separately. The chance of clinical pregnancy and multiple pregnancy for different numbers of small follicles in cycles with one >18-mm follicle was calculated using marginal effects estimate. MAIN RESULTS AND THE ROLE OF CHANCE: This cohort included 12 933 IUI cycles in 7504 women. The overall clinical pregnancy rate was 16.1% (2081/12 933), with a multiple pregnancy rate of 10.5% (218/2081). In the adjusted analysis, the chance of clinical pregnancy increased significantly with the increase in the number of follicles with the diameter of 14-16, 16-18, and 18-20 mm. As for 10-12 mm [adjusted odds ratio (aOR) 1.22, 95% CI 1.02-1.46] and 12-14 mm (aOR 1.29, 95% CI 1.07-1.56) follicles, only groups with ≥2 follicles of those sizes showed significantly increased chance of clinical pregnancy. In cycles that led to pregnancy, follicles with the diameter of 12-14 mm were associated with an increased risk of multiple pregnancy (aOR 1.73, 95% CI 1.19-2.53 for one such follicle; aOR 2.27, 95% CI 1.44-3.56 for ≥2 such follicles), while 10- to 12-mm follicles were not significantly associated with multiple pregnancy (aOR 1.18, 95% CI 0.72-1.95 for ≥2 such follicles). The associations of multiple pregnancy were similar when including all cycles. LIMITATIONS, REASONS FOR CAUTION: This was a retrospective observational study from a single center. The records of follicle diameter in our center were of a 2-mm interval which limited our ability to analyze the size of follicle as a continuous variable. Also, the number of cycles with a high number of small follicles was still limited which impeded more detailed analysis on the ≥2 follicles subgroup. Similarly, the value of some parts of the marginal probability estimation for multiple pregnancy versus pregnancy according to size and number of follicles was also limited by the low sample size of certain combinations. WIDER IMPLICATIONS OF THE FINDINGS: Follicles larger than 10 mm, especially those ≥12 mm, may need to be clearly recorded during transvaginal ultrasound surveillance and their potential effects on both pregnancy and multiple pregnancy can be discussed with couples undergoing IUI. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Natural Science Foundation of China (Grant numbers 82201912, 82371651, and 82071615) and Shanghai Sailing Program (21YF1423200). B.W.M. is supported by an NHMRC Investigator grant (GNT1176437). B.W.M. reports consultancy for ObsEva and Merck and travel support from Merck. B.W.M. has received research funding from Ferring and Merck. The authors declare no other competing interests. TRIAL REGISTRATION NUMBER: N/A.

Significant decrease of maternal mitochondria carryover using optimized spindle-chromosomal complex transfer.

Mutations in mitochondrial DNA (mtDNA) contribute to a variety of serious multi-organ human diseases, which are strictly inherited from the maternal germline. However, there is currently no curative treatment. Attention has been focused on preventing the transmission of mitochondrial diseases through mitochondrial replacement (MR) therapy, but levels of mutant mtDNA can often unexpectedly undergo significant changes known as mitochondrial genetic drift. Here, we proposed a novel strategy to perform spindle-chromosomal complex transfer (SCCT) with maximal residue removal (MRR) in metaphase II (MII) oocytes, thus hopefully eliminated the transmission of mtDNA diseases. With the MRR procedure, we initially investigated the proportions of mtDNA copy numbers in isolated karyoplasts to those of individual oocytes. Spindle-chromosomal morphology and copy number variation (CNV) analysis also confirmed the safety of this method. Then, we reconstructed oocytes by MRR-SCCT, which well developed to blastocysts with minimal mtDNA residue and normal chromosomal copy numbers. Meanwhile, we optimized the manipulation order between intracytoplasmic sperm injection (ICSI) and SCC transfer and concluded that ICSI-then-transfer was conducive to avoid premature activation of reconstructed oocytes in favor of normal fertilization. Offspring of mice generated by embryos transplantation in vivo and embryonic stem cells derivation further presented evidences for competitive development competence and stable mtDNA carryover without genetic drift. Importantly, we also successfully accomplished SCCT in human MII oocytes resulting in tiny mtDNA residue and excellent embryo development through MRR manipulation. Taken together, our preclinical mouse and human models of the MRR-SCCT strategy not only demonstrated efficient residue removal but also high compatibility with normal embryo development, thus could potentially be served as a feasible clinical treatment to prevent the transmission of inherited mtDNA diseases.

Earlier second polar body transfer and further mitochondrial carryover removal for potential mitochondrial replacement therapy.

The second polar body (PB2) transfer in assisted reproductive technology is regarded as the most promising mitochondrial replacement scheme for preventing the mitochondrial disease inheritance owing to its less mitochondrial carryover and stronger operability. However, the mitochondrial carryover was still detectable in the reconstructed oocyte in conventional second polar body transfer scheme. Moreover, the delayed operating time would increase the second polar body DNA damage. In this study, we established a spindle-protrusion-retained second polar body separation technique, which allowed us to perform earlier second polar body transfer to avoid DNA damage accumulation. We could also locate the fusion site after the transfer through the spindle protrusion. Then, we further eliminated the mitochondrial carryover in the reconstructed oocytes through a physically based residue removal method. The results showed that our scheme could produce a nearly normal proportion of normal-karyotype blastocysts with further reduced mitochondrial carryover, both in mice and humans. Additionally, we also obtained mouse embryonic stem cells and healthy live-born mice with almost undetectable mitochondrial carryover. These findings indicate that our improvement in the second polar body transfer is conducive to the development and further mitochondria carryover elimination of reconstructed embryos, which provides a valuable choice for future clinical applications of mitochondrial replacement.

The pregnancy outcomes of infertile women with polycystic ovary syndrome undergoing intrauterine insemination with different attempts of previous ovulation induction.

Background: Polycystic ovary syndrome (PCOS) is one of the most common reasons for infertility. The consensus of the treatment of infertile women with PCOS is ovulation induction (OI) for six to nine attempts before in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI). Nowadays, more attention was paid to a rising, noninvasive treatment, intrauterine insemination (IUI), as some experts claimed IUI could benefit PCOS patients with infertility. Our study means to investigate the outcomes of IUI for PCOS patients and if patients' previous OI cycles can be a predictive factor for IUI outcomes. Methods: A total of 1,086 PCOS patients was included and 1,868 IUI cycles were performed between January 2007 and July 2021 in the department of assisted reproduction in Shanghai Ninth People's Hospital. All included patients underwent IUI treatments with letrozole+human menopausal gonadotropin (LE+hMG) for ovarian stimulation. Results: The pregnancy outcomes were not associated with the attempts of failed OI cycles previously. Specifically, the clinical pregnancy rate was 21.14% for PCOS patients without previous OI cycles, 21.95% for PCOS patients with 1-2 previous OI cycles and 23.64% for PCOS patients with 3 or more previous OI cycles (p=0.507). The corresponding live birth rate was 16.64%, 18.06%, and 18.68%, respectively, of which the difference was not statistically significant (p=0.627). The cumulative rate per patient was 38.59% for clinical pregnancy and 31.03% for live birth, and approximately 98% of the pregnancies occurred in the first 3 cycles of IUI. Conclusion: PCOS women with different attempts of OI cycles had similar pregnancy outcomes after IUI, thus a history of repeated failures of OI treatments was not a predictive factor for the pregnancy outcomes in IUI cycles. Most pregnancies occurred in the first three cycles of IUI, so we strongly recommended three attempts of IUI for PCOS women before they switched to IVF/ICSI. Generally, IUI might be an assist for infertile women with PCOS before IVF/ICSI and might accelerate pregnancy for target women without invasive manipulations.

Rich Get Richer: Extraversion Statistically Predicts Reduced Internet Addiction through Less Online Anonymity Preference and Extraversion Compensation.

Internet addiction may arise from multiple factors and personality tendencies have been previously implicated. Prior studies have found that extraversion may be a protective factor mitigating against internet addiction, yielding a "rich-get-richer" effect. However, few studies have explored how extraversion may influence internet addiction from the perspective of online-offline integration. Drawing on a sample of 428 college students, the current study examined a serial mediation model exploring the underlying mechanisms of how extraversion may statistically predict internet addiction through online-offline integration and antecedent factors. The serial mediation model analyses indicated that extraverted internet users exhibited a weaker preference for online anonymity and less online extraversion compensation, thus formulating a higher level of online-offline integration than introverted individuals, which, in turn, appeared to reduce the risk of internet addiction. In contrast, with regard to specific components of online-offline integration, introverted internet users preferred online anonymity, which reduced their relationship integration and increased their likelihood of internet addiction; similarly, the introverted individuals were also more likely to exhibit an extraversion compensation effect. That is, they were more extraverted on the internet than in general; hence, they had a lower level of self-identity integration, resulting in a greater likelihood of experiencing internet addiction. These results highlight the importance of online-offline integration that may account for personality variations in social and psychological outcomes related to internet use, and suggest a role for online anonymity preference and extraversion compensation in influencing specific components of integration.

Therapeutic role of granulocyte macrophage colony-stimulating factor (GM-CSF) in patients with persistent thin endometrium: A prospective and randomized study.

OBJECTIVE: To assess the effect of granulocyte macrophage colony-stimulating factor (GM-CSF) on unresponsive thin (<7 mm) endometrium in women undergoing frozen-thawed embryo transfer. METHODS: A single-center, randomized, prospective study enrolled 304 women with thin unresponsive endometrium from Shanghai Ninth People's Hospital between March 2017 and May 2018. Of them, 161 patients received an intrauterine infusion of GM-CSF and 143 patients served as controls. After hysteroscopy, a gel with or without GM-CSF was administered to fill the uterine cavity completely or up to 5 mL only. The primary outcome was confirmed pregnancy and secondary outcomes included endometrial thickness and implantation rate. RESULTS: Patients who were administered GM-CSF had a significantly higher chemical pregnancy rate (35.3% vs 20.0%; P=0.009) and clinical pregnancy rate (28.6% vs 13.3%; P=0.005) compared with patients in the control group. Patients treated with GM-CSF had significantly higher endometrial thickness compared with controls (7.83 ± 1.45 mm vs 7.37 ± 0.70 mm, P=0.003). CONCLUSION: GM-CSF therapy can effectively increase endometrial thickness and improve the clinical pregnancy rate in patients with persistent thin endometrium. The therapeutic role of GM-CSF for infertile women under in vitro fertilization and embryo transfer (IVF-ET) treatment can be further explored. CHINESE CLINICAL TRIAL REGISTER: ChiCTR-IPR-17011242.

Cross-linked hyaluronan gel to improve pregnancy rate of women patients with moderate to severe intrauterine adhesion treated with IVF: a randomized controlled trial.

PURPOSE: To evaluate whether the cross-linked hyaluronan (cHA) gel can improve the clinical pregnancy rate of patients with moderate to severe intrauterine adhesion (IUA) who underwent operative hysteroscopy followed by embryo transfer. METHODS: Women with moderate to severe IUA desiring to undergo embryo transfer were recruited in this randomized controlled trial. The patients were randomized on the day of receiving hysteroscopy. The control group received standard hysteroscopy, while cHA gel was applied to the treatment group at the end of hysteroscopy and 5-7 days after operation. All patients were expected to undergo in vitro fertilization (IVF)/intracytoplasmic sperm injection and frozen-thawed embryo transfer (FET). RESULTS: A total of 306 patients were enrolled in this study, of which 202 were assigned to the treatment group and 104 to the control group. Both the clinical pregnancy rate (26.3% [49/186] vs. 15.3% [13/85], P = 0.045), the implantation rate (17.7% [57/322] vs. 9.8% [15/153], P = 0.025), and the endometrial thickness on the day of embryo transfer (7.97 ± 1.37 vs. 7.50 ± 0.60 mm, P < 0.001) were significantly higher in the treatment group compared to the control group. In addition, histological assessment of the paired endometrial tissues collected before and after operation revealed a relatively higher number of tubular glands after operation (15.1 ± 13.2 vs. 28.8 ± 30.4, P = 0.166). CONCLUSIONS: To conclude, the application of cHA gel in patients with moderate to severe IUA during hysteroscopy can improve the quality of endometrium and uterine receptivity and consequently enhance the clinical pregnancy rate after IVF/CSI and FET.

Progestin vs. Gonadotropin-Releasing Hormone Antagonist for the Prevention of Premature Luteinizing Hormone Surges in Poor Responders Undergoing in vitro Fertilization Treatment: A Randomized Controlled Trial.

Objective: Progestin was recently used as an alternative of gonadotropin-releasing hormone (GnRH) analog for preventing premature luteinizing hormone (LH) surge with the aid of vitrification techniques, however, limited data were available about the potential of progestin in poor responders undergoing in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) treatment. We performed a randomized parallel controlled trial to investigate the difference of progestin and GnRH antagonist in poor responders. Methods: A total of 340 poor responders who met with Bologna criteria were randomly allocated into the progestin-primed ovarian stimulation (PPOS) group and GnRH antagonist group. Fresh embryo transfer was preferred in the GnRH antagonist group and freeze-all was performed in the PPOS group. The primary outcome was the incidence of premature LH surge, secondary outcomes were the number of retrieved oocytes, the number of viable embryos and the pregnancy outcomes. Results: The results showed that the incidence of premature LH surge in PPOS group was lower than that in antagonist group (0 vs. 5.88%, P < 0.05). In PPOS group, the average numbers of oocytes and viable embryos were comparable to those in GnRH antagonist group (3.7 ± 2.6 vs. 3.4 ± 2.4; 1.6 ± 1.7 vs. 1.4 ± 1.3, P > 0.05), the live birth rate was similar between the two groups (21.8 vs. 18.2%, RR 1.25 (95% confidence interval 0.73, 2.13), P > 0.05). Conclusions: The study demonstrated that PPOS had a more robust control for preventing premature LH rise than GnRH antagonist in poor responders, but PPOS in combination with freeze-all did not significantly increase the probability of pregnancy than GnRH antagonist protocol for poor responders.

Optimal Ovulation Trigger-Oocyte Pickup Interval in Progestin-Primed Ovarian Stimulation Protocol: A Retrospective Study Using Propensity Score Matching.

Background: To investigate the optimal ovulation trigger-oocyte pickup (OPU) interval of a progestin-primed ovarian stimulation (PPOS) protocol. Method: Patients with normal ovarian reserve in their first PPOS OPU cycle were enrolled in this retrospective cohort study between July 2013 and April 2018. This retrospective cohort study included two parts. In part I, we studied the regression trend of mature oocyte rate, implantation rate, and live birth rate within the whole ovulation trigger-OPU interval of 7,258 patients. To homogenize some clinical characters that were key regulators of OPU time, in part II, we used propensity score matching to auto-select patients among trigger-OPU interval group 1 (35.6-36.4 h), group 2 (36.4-37.1 h), and group 3 (37.1-37.8 h) and analyzed clinical outcomes. Results: Study part I showed that the whole ovulation trigger-OPU interval (33-39.5 h) of PPOS protocol had a trend of a high mature oocyte rate (>80%), increasing implantation rate, and high live birth rate. Propensity score matching of patients with homogeneous clinical characteristics further indicated that the trigger-OPU interval within groups 2 and 3 (36.4-37.8 h) had significantly higher mature oocyte rates (84.54% vs. 84.60% vs. 82.34%, P = 0.002) and implantation rates (34.17% vs. 34.37% vs. 29.61%, P < 0.05) than group 1. The same tend was observed in the live birth rate. Conclusions: The ovulation trigger-OPU interval of 36.4-37.8 h is optimal for most patients using a PPOS protocol.

Effect of hysteroscopy before starting in-vitro fertilization for women with recurrent implantation failure: A meta-analysis and systematic review.

OBJECTIVE: To study if hysteroscopy (HSC) before starting an in-vitro fertilization (IVF) cycle improves IVF outcomes in women with recurrent implantation failure (RIF). METHODS: The Medline, Cochrane, EMBASE, and Google Scholar databases were searched using the following keywords until March 31, 2017: in-vitro fertilization; infertility; hysteroscopy; recurrence; embryo implantation; and pregnancy. Randomized controlled trials (RCTs), two-arm prospective studies, and retrospective studies were included. RESULTS: Three RCTs, 3 nonrandomized prospective studies, and 2 retrospective cohort studies were included. The eligible studies included 3932 women with RIF: 1841 in the HSC group and 2091 in the control group. The clinical pregnancy rate and implantation rate was significantly higher in the HSC group compared with the control group (for clinical pregnancy rate, pooled odds ratio [OR] = 1.64, 95% confidence intervals [CI]: 1.30-2.07, P < 0.001; for implantation rate, pooled OR = 1.22, 95% CI: 1.02-1.45, P = 0.025). The live birth rate (pooled OR = 1.30, 95% CI: 0.90-1.88, P = 0.168) and the miscarriage rate (pooled OR = 0.94, 95% CI: 0.66-1.35, P = 0.744) of the 2 groups were not statistically significantly. CONCLUSIONS: HSC improved the implantation rate and clinical pregnancy rates, but failed to improve live birth rate and did not affect the miscarriage rate in women with RIF undergoing IVF. Since HSC plays a significant role in pregnancy and birth outcomes of women with RIF, further studies are warranted.

The comprehensive mutational and phenotypic spectrum of TUBB8 in female infertility.

Human oocyte maturation is a precondition for fertilization and ensuing embryonic development. Previously, we identified TUBB8 variants as a genetic determinant of human oocyte maturation arrest and showed that these variants cause variable and mixed phenotypes in oocyte maturation and early embryo development. We also estimated that rare inherited or de novo variants in the TUBB8 gene accounted for 30% of individuals in a small cohort of patients affected by oocyte maturation arrest. In the present study, we recruited a further 87 patients from unrelated families diagnosed with oocyte maturation or early embryonic arrest and identified 30 patients carrying TUBB8 variants. The corresponding phenotypes not only include oocyte maturation arrest, failure of fertilization, and early embryonic arrest, but also extend to the new phenotype of failure of embryo implantation. These observations provide the most detailed mutational and phenotypic spectrum of TUBB8, further extend the spectrum of variants and dysfunctional oocyte and embryo phenotypes caused by TUBB8 variants, and confirm previous findings for a critical role of TUBB8 during oocyte maturation and early embryonic development. Thus, TUBB8 mutation screening might not only be a genetic diagnostic marker for patients with oocyte maturation arrest, but might also have clinical implications for evaluating the competence of patients' functional oocytes with first polar body (PB1).

Dual trigger of final oocyte maturation in poor ovarian responders undergoing IVF/ICSI cycles.

Previous studies show that a dual trigger ovulation regimen significantly improves number and maturity of retrieved oocytes for normal ovarian responders or patients with history of low oocyte yield. The current retrospective cohort study investigated whether dual trigger of final oocyte maturation may benefit IVF outcomes for poor ovarian responders fulfilling the Bologna criteria. Undertaken between May 2014 and August 2016, the study involved 1350 patients undergoing 1389 IVF/intracytoplasmic sperm injection treatment cycles. Patients triggered with 5000 IU human chorionic gonadotrophin (HCG) alone (328 cycles) were compared with those undergoing dual triggering with 5000 IU HCG plus 0.1 mg gonadotrophin-releasing hormone agonist (GnRHa) (386 cycles) and patients triggered with 10,000 IU HCG (363 cycles) were compared with those undergoing dual triggering with 10,000 IU HCG plus 0.1 mg GnRHa (312 cycles). The dual trigger groups showed significantly higher number of oocytes collected and number of mature oocytes compared with their respective HCG trigger group (P < 0.001). Oocyte retrieval rate and percentage of mature oocytes retrieved were also both significantly higher in the dual trigger groups (P < 0.001). Fertilization rate, number of viable embryos, implantation rate, clinical pregnancy rate and miscarriage rate were not significantly different between groups.

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

BACKGROUND: The 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. METHODS: A 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. RESULTS: Compared 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). CONCLUSION: This 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 REGISTRATION: ChiCTR-OCH-14004176 . Registered on January 8, 2014.

Short-term copper intrauterine device placement improves the implantation and pregnancy rates in women with repeated implantation failure.

OBJECTIVE: To study if hysteroscopy and short-term copper intrauterine device placement (Cu-IUD) improves the pregnancy rates of women with repeated implantation failure (RIF) undergoing frozen-thawed embryo transfer (FET). DESIGN: Retrospective study. SETTING: Medical university hospital. PATIENT(S): Infertile women with at least two implantation failures with the use of at least one good-quality embryo. INTERVENTION(S): All patients received operative hysteroscopy in the follicular cycle, and if endometrial polyps, polypoid endometrium, or intrauterine adhesions were found they were removed. In some patients, a Cu-IUD was inserted immediately after hysteroscopy and removed after two menstrual periods before embryo implantation. All patients underwent in vitro fertilization or intracytoplasmic sperm injection and FET. MAIN OUTCOME MEASURE(S): Clinical pregnancy and implantation rates. RESULT(S): A total of 440 women with a mean age of 33.42 ± 4.45 years (range 23-47 y) were included. There were 382 patients (554 cycles) in the IUD group and 58 patients (87 cycles) in the non-IUD group. The two groups were similar regarding age, body mass index, and infertility factors. The IUD group had a significantly higher implantation rate (29.29% vs. 16.56%), chemical pregnancy rate (53.25 vs. 41.38%), and clinical pregnancy rate (45.13% vs. 26.44%) than the non-IUD group. Multivariable regression analysis indicated that the odds of a chemical pregnancy was significantly increased with IUD usage. CONCLUSION(S): Cu-IUD placement for two menstrual cycles at the time of hysteroscopy can improve the implantation and pregnancy rates in women with RIF.

Three-Dimensional HyCoSy With Perfluoropropane-Albumin Microspheres as Contrast Agents and Normal Saline Injections Into the Pelvic Cavity for Morphological Assessment of the Fallopian Tube in Infertile Women.

OBJECTIVES: To apply the three-dimensional (3D) hysterosalpingo-contrast sonography (HyCoSy) with perfluoropropane-albumin microspheres as contrast agents and normal saline injections into the pelvic cavity for assessment of the tubal patency and adhesions of fimbrial parts. METHODS: Fifty-five infertile female patients were recruited to undergo 3D HyCoSy with normal saline injected into the pelvic cavity, in which the tubal patency was observed by visualizing the spillage of contrast agents from the fimbriae, and the fimbrial adhesion was confirmed by the finger-like projections of the fimbriae and their floating and moving status. RESULTS: Of the 55 patients, bilateral tubal patency was observed in 44 (80.0%), unilateral tubal patency and the other partial occlusion in 7 (12.7%), unilateral partial occlusion and the other complete occlusion in 3 (5.4%), and bilateral complete occlusion in 1 (1.8%). The fimbrial parts were observed in 105 fallopian tubes, among which 101 were seen with the finger-like fimbriae floated and moved in the pelvic cavity, whereas 4 tubes were not because of adhesion to the pelvic cavity (n = 3) or the ovary and intestine (n = 1). More than three visible, quite long, and distributed evenly finger-like projections were present for the patent fimbrial parts; however, fewer, flat, and not evenly distributed finger-like projections were present for the adhesive tubes. No serious complications occurred during or after this procedure. CONCLUSIONS: Combination of 3D HyCoSy with normal saline injected into the pelvic cavity may be a feasible and safe procedure to assess tubal patency and adhesions of the fimbrial parts.

Flexibility in starting ovarian stimulation at different phases of the menstrual cycle for treatment of infertile women with the use of in vitro fertilization or intracytoplasmic sperm injection.

OBJECTIVE: To investigate flexibility in starting controlled ovarian stimulation at any phase of the menstrual cycle in infertile women undergoing treatment with assisted reproduction. DESIGN: Retrospective cohort study. SETTING: Academic tertiary-care medical center. PATIENT(S): At total of 150 infertile patients undergoing in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) treatment. Ninety of the women also underwent frozen embryo transfer (FET) procedures. INTERVENTION(S): Depending on the phase of the menstrual cycle when ovarian stimulation was started, three groups of patients were identified, namely: conventional group (ovarian stimulation started in the early follicular phase), late follicular phase group, and luteal phase group. When dominant follicles were observed, final oocyte maturation was triggered with the use of GnRH agonist and hCG. In all three groups, viable embryos were cryopreserved for subsequent transfer. PRIMARY OUTCOME: number of mature oocytes retrieved. SECONDARY OUTCOMES: fertilization rate, viable embryo rate per oocyte retrieved, cancellation rate, and clinical pregnancy outcomes from FET cycles. RESULTS(S): There were no differences in the mean number of mature oocytes retrieved in the conventional group, late follicular phase group, and luteal phase group (5.7 ± 3.6, 5.2 ± 3.7, and 5.2 ± 3.9, respectively). Similarly, no significant differences were observed in the viable embryo rate per oocyte retrieved (37.9%, 38.5%, and 43.6%), clinical pregnancy rates (41.5%, 45.5%, and 38.9%), and implantation rates (30.7%, 30.2%, and 27.1%) in the three groups. CONCLUSION(S): All three ovarian stimulation protocols were observed to be equally effective. These results demonstrate that ovarian stimulation can be commenced on any day of the menstrual cycle. CLINICAL TRIAL REGISTRATION NUMBER: ChiCTR-OPN-15007332.

Mutations in TUBB8 and Human Oocyte Meiotic Arrest.

Background Human reproduction depends on the fusion of a mature oocyte with a sperm cell to form a fertilized egg. The genetic events that lead to the arrest of human oocyte maturation are unknown. Methods We sequenced the exomes of five members of a four-generation family, three of whom had infertility due to oocyte meiosis I arrest. We performed Sanger sequencing of a candidate gene, TUBB8, in DNA samples from these members, additional family members, and members of 23 other affected families. The expression of TUBB8 and all other ß-tubulin isotypes was assessed in human oocytes, early embryos, sperm cells, and several somatic tissues by means of a quantitative reverse-transcriptase-polymerase-chain-reaction assay. We evaluated the effect of the TUBB8 mutations on the assembly of the heterodimer consisting of one α-tubulin polypeptide and one ß-tubulin polypeptide (α/ß-tubulin heterodimer) in vitro, on microtubule architecture in HeLa cells, on microtubule dynamics in yeast cells, and on spindle assembly in mouse and human oocytes. Results We identified seven mutations in the primate-specific gene TUBB8 that were responsible for oocyte meiosis I arrest in 7 of the 24 families. TUBB8 expression is unique to oocytes and the early embryo, in which this gene accounts for almost all the expressed ß-tubulin. The mutations affect chaperone-dependent folding and assembly of the α/ß-tubulin heterodimer, disrupt microtubule behavior on expression in cultured cells, alter microtubule dynamics in vivo, and cause catastrophic spindle-assembly defects and maturation arrest on expression in mouse and human oocytes. Conclusions TUBB8 mutations have dominant-negative effects that disrupt microtubule behavior and oocyte meiotic spindle assembly and maturation, causing female infertility. (Funded by the National Basic Research Program of China and others.).

Quantitative methylation level of the EPHX1 promoter in peripheral blood DNA is associated with polycystic ovary syndrome.

Steroid synthesis and metabolic pathways play important roles in the pathophysiology of PCOS, but until now there have been no studies on the methylation profiles of specific genes in steroid synthesis pathways that are known to be associated with PCOS. Here we used MassARRAY quantitative methylation analysis to determine the methylation levels of each CpG site or cluster in the promoters of EPHX1, SRD5A1, and CYP11A1 in 64 peripheral blood samples. We further examined the methylation level of EPHX1 in an independent cohort consisting of 116 people. Finally, we investigated the role of EPHX1 in steroidogenesis in the KGN cell line. For SRD5A1 and CYP11A1, there was no significant difference in methylation level between patients and controls. For EPHX1, however, the methylation levels of a few consecutive CpG sites and clusters were found to be significantly associated with PCOS. The methylation levels of a number of CpG clusters or sites were significantly lower in patients than in controls in the first cohort consisting of 64 people, such as clusters 13-14 (P<0.05), 15-16 (P<0.001), and 19-24 (P<0.001) and sites CpG_53 (P<0.01) and CpG_54 (P<0.05). Among differentiated methylation sites and clusters, the methylation levels of the CpG cluster 13-14 and CpG cluster 19-24 in PCOS patients were significantly lower than in controls in the second cohort of 116 people (P<0.05 for both). In addition, knockdown and overexpression experiments in KGN cells showed that EPHX1 can regulate estradiol concentrations, and this indicates a role for EPHX1 in steroidogenesis. Our study has demonstrated that methylation of the EPHX1 promoter might be associated with PCOS. This study provides direct evidence that methylation plays an important role in PCOS and demonstrates a novel role for EPHX1 in female reproduction.

Quantitative analysis of follistatin (FST) promoter methylation in peripheral blood of patients with polycystic ovary syndrome.

Epigenetic mechanisms may contribute to polycystic ovary syndrome (PCOS). To date, however, no studies have associated CpG methylation levels of any candidate gene with PCOS susceptibility. Follistatin (FST), an activin-binding protein, is expressed in numerous tissues and is shown to have linkage with PCOS. However, results from case-control association analyses between this gene and PCOS are inconsistent. Thus, this study investigated possible association of methylation levels in the promoter and 5'-untranscribed region (UTR) of the FST gene with PCOS incidence in peripheral blood leukocytes and endometrial tissue. Using mass array quantitative methylation analysis, first the 5'-UTR methylation in FST was analysed in 130 PCOS patients and 120 controls. The methylation level of the FST gene was further studied in endometrium from 24 controls and 24 PCOS patients. This study demonstrates that methylation levels of CpG sites in the FST promoter and 5'-UTR are not associated with PCOS. Nonetheless, this was the first study to quantitatively evaluate the methylation levels of a candidate gene in association with PCOS. Further studies should be performed to examine methylation in other candidate genes. Understanding the epigenetic mechanisms involved in PCOS may yield new insights into the pathophysiology of the disorder. Animal models demonstrate that epigenetic reprogramming may contribute to polycystic ovary syndrome (PCOS). To date, however, no studies have associated CpG methylation levels of any candidate gene with PCOS susceptibility. Follistatin (FST), an activin-binding protein, is expressed in numerous tissues and is a PCOS candidate gene. However, results from association analyses between this gene and PCOS are inconsistent. Thus, we investigated possible association of methylation levels in the promoter and 5'-UTR of the FST gene with PCOS incidence in peripheral blood leukocytes and endometrial tissue. Using mass array quantitative methylation analysis, we firstly analysed 5'-UTR methylation in 40 PCOS patients and 40 controls. We then validated results in a second sample consisting of 90 PCOS patients and 80 controls. The methylation level of the FST gene was further studied in endometrium from 24 controls and 24 PCOS patients. Finally, we quantitatively analysed FST expression in the endometrium using real-time PCR. Our study demonstrated that methylation levels of CpG sites in the FST promoter and 5'-UTR are not associated with PCOS. Nonetheless, as far as is known, this is the first study to quantitatively evaluate the methylation levels of a candidate gene in association with PCOS. Further studies should be performed to examine methylation in other candidate genes. Understanding the epigenetic mechanisms involved in PCOS may yield new insights into the pathophysiology of the disorder.