Replication stress impairs chromosome segregation and preimplantation development in human embryos.

Human cleavage-stage embryos frequently acquire chromosomal aneuploidies during mitosis due to unknown mechanisms. Here, we show that S phase at the 1-cell stage shows replication fork stalling, low fork speed, and DNA synthesis extending into G2 phase. DNA damage foci consistent with collapsed replication forks, DSBs, and incomplete replication form in G2 in an ATR- and MRE11-dependent manner, followed by spontaneous chromosome breakage and segmental aneuploidies. Entry into mitosis with incomplete replication results in chromosome breakage, whole and segmental chromosome errors, micronucleation, chromosome fragmentation, and poor embryo quality. Sites of spontaneous chromosome breakage are concordant with sites of DNA synthesis in G2 phase, locating to gene-poor regions with long neural genes, which are transcriptionally silent at this stage of development. Thus, DNA replication stress in mammalian preimplantation embryos predisposes gene-poor regions to fragility, and in particular in the human embryo, to the formation of aneuploidies, impairing developmental potential.

Early detection of cryostorage tank failure using a weight-based monitoring system.

PURPOSE: To study the relationship between liquid nitrogen loss and temperature in cryostorage dewars and develop an early-warning alarm for impending tank failure. METHODS: Cryostorage dewars were placed on custom-engineered scales, and weight and temperature data were continuously monitored in the setting of slow, medium, and fast rate-loss of LN2 to simulate three scenarios of tank failure. RESULTS: LN2 Tank weights and temperatures were continuously monitored and recorded, with a calculated alarm trigger set at 10% weight loss and temperature of - 185 °C. With an intact tank, a 10% loss in LN2 occurred in 4.2-4.9 days. Warming to - 185 °C occurred in 37.8-43.7 days, over 30 days after the weight-based alarm was triggered. Full evaporation of LN2 required ~ 36.8 days. For the medium rate-loss simulation, a 10% loss in LN2 occurred in 0.8 h. Warming to - 185 °C occurred in 3.7-4.8 h, approximately 3 h after the weight-based alarm was triggered. For the fast rate-loss simulation, a 10% weight loss occurred within 15 s, and tanks were depleted in under 3 min. Tank temperatures began to rise immediately and at a relatively constant rate of 43.9 °C/h and 51.6 °C/h. Temperature alarms would have sounded within 0.37 and 0.06 h after the breech. CONCLUSIONS: This study demonstrates that a weight-based alarm system can detect tank failures prior to a temperature-based system. Weight-based monitoring could serve as a redundant safety mechanism for added protection of cryopreserved reproductive tissues.

Endocrine causes of recurrent pregnancy loss.

OBJECTIVE: To review the available data on endocrine disorders and recurrent pregnancy loss. FINDINGS: Our group found that most endocrine disorders do not seem to be correlated with a diagnosis of recurrent pregnancy loss (RPL). The exception to this is testing for thyroid stimulating hormone and thyroid antibodies, which is recommended due to a strong correlation with recurrent pregnancy loss and positive anti-thyroid peroxidase antibodies. CONCLUSION: The available literature supports testing thyroid function and antibodies in women with RPL. Testing for other endocrine disorders is only warranted if otherwise clinically indicated, independent from a history of recurrent pregnancy loss.

Minimizing mosaicism: assessing the impact of fertilization method on rate of mosaicism after next-generation sequencing (NGS) preimplantation genetic testing for aneuploidy (PGT-A).

PURPOSE: Advances in preimplantation genetic testing (PGT) have led to practice changes in assisted reproductive technologies (ART), enabling fertility centers to transfer single embryos while maintaining excellent ongoing pregnancy rates, reducing miscarriage rates, and dramatically reducing ART-associated multiple pregnancies. The introduction of next-generation sequencing (NGS) allows PGT laboratories to assess for embryo mosaicism-although the true incidence and reproductive potential of predicted mosaic embryos are controversial. Due to concern for genetic contamination from other spermatozoa, most reference laboratories require use of intracytoplasmic sperm injection (ICSI) for single gene preimplantation genetic diagnosis (PGT-M). However, in PGT for aneuploidy (PGT-A), conventional insemination (IVF) is typically permissible. The purpose of this study was to evaluate rates of euploid, aneuploid, and mosaic in trophectoderm biopsy samples from embryos in IVF versus ICSI PGT-A cycles. Secondary aims were to assess sex ratio, and subtypes of aneuploidy and mosaicism in IVF versus ICSI PGT-A cycles. METHODS: We performed a retrospective review of women undergoing PGT-A at a single academic fertility center from July 1, 2015, to September 1, 2017. In all cycles, PGT-A was performed via trophectoderm biopsy on day 5 or 6 and analyzed using NGS at a single reference lab. We collected and compared patient demographics, fertility testing, cycle characteristics, and PGT-A outcomes between IVF and ICSI cycles. RESULTS: Three hundred two PGT-A cycles were included for analysis: 75 IVF and 227 ICSI cycles, resulting in 251 IVF and 724 ICSI biopsied blastocysts. Mean oocyte age of included cycles was 38.6 years (IVF) and 38.5 years (ICSI), p = 0.85. Baseline characteristics of IVF and ICSI PGT-A cycles were similar with the exception of semen parameters: IVF cycles had higher sperm concentration and total motility compared to ICSI cycles. PGT-A outcomes did not differ between IVF and ICSI cycles: euploid 27.9% (IVF) versus 30% (ICSI); aneuploid 45.4% (IVF) versus 43.1% (ICSI); no result 4.4% (IVF) versus 6.2% (ICSI). Though not significant, we identified a trend toward higher rate of mosaicism in IVF (25.9%) versus ICSI (20.9%). Among mosaic embryos, a lower percentage of simple mosaic embryos resulted from IVF (53.8%) versus ICSI (70.2%). Among aneuploid embryos, a non-significant higher percentage of complex aneuploidy resulted from IVF (16.3%) versus ICSI (9%). IVF resulted in a non-significant higher proportion of cycles with no transferrable embryos (42.7%) versus ICSI (36.6%). Numerical and sex chromosome involvement in mosaicism and aneuploidy were similar between IVF and ICSI cycles. CONCLUSION: IVF and ICSI NGS PGT-A have similar rates of euploid, aneuploid, and no result embryos, though IVF may result in higher rates of mosaicism and demonstrates differences in proportions of mosaic and aneuploid subtypes compared to ICSI. ICSI may be preferable to conventional insemination to minimize the rate of mosaic results in NGS PGT-A cycles.

Cyst aspiration or GnRH antagonist administration for ovarian cysts detected at the start of fresh in vitro fertilization cycles.

The primary objective of this study is to investigate the effect of transvaginal ultrasonogram (TVUS)-guided cyst aspiration or gonadotropin releasing hormone antagonist (GnRH-ant) administration for the management of solitary ovarian cysts detected at the start of in vitro fertilization (IVF) cycles on the outcomes of the same cycles. This is a single-center, retrospective, cohort study of patients who had TVUS-guided cyst aspiration or GnRH-ant treatment for ovarian cysts detected at the start of IVF during a 5-year period. Four hundred and three patients met inclusion criteria: 41 (10.2%) underwent cyst aspiration and 362 (89.2%) were treated with GnRH-ant. There was no difference in the demographics or baseline IVF cycle characteristics of the two groups. Patients treated with GnRH-ant had a longer duration of ovarian stimulation (10.8 ± 3.45 days versus 9.05 ± 4.06 days, p = 0.003) and required higher gonadotropin doses (3887.7 ± 1097.8 IU versus 3293.7 ± 990.5 IU; p = 0.01) compared with the cyst aspiration group. There was no difference in the clinical pregnancy (43.9% versus 41.4%), spontaneous miscarriage (9.76% versus 8.01%) and live birth (34.1% versus 33.4%) rates between the groups. Our findings suggest that cyst aspiration is comparable to GnRH-ant administration for the management of solitary ovarian cysts detected at the start of IVF cycles.

Does the time interval between hysteroscopic polypectomy and start of in vitro fertilization affect outcomes?

OBJECTIVE: To investigate whether the time interval between hysteroscopic polypectomy and the start of IVF-ET cycles affect IVF cycle outcomes. DESIGN: Retrospective cohort. SETTING: Academic center. PATIENT(S): All patients diagnosed with endometrial polyps undergoing hysteroscopic polypectomy before fresh IVF-ET. INTERVENTION(S): Hysteroscopic polypectomy. MAIN OUTCOME MEASURE(S): Patients were divided into three groups based on the time interval between hysteroscopic polypectomy and the start of a fresh IVF-ET cycle. Group 1 consisted of patients who underwent IVF-ET after their next menses, group 2 after two or three menstrual cycles, and group 3 after more than three menstrual cycles. Demographics, baseline IVF characteristics, controlled ovarian stimulation response, and pregnancy outcomes after ET were compared among the groups. RESULT(S): A total of 487 patients met inclusion criteria: 241 in group 1 (49.5%), 172 in group 2 (35.3%), and 74 in group 3 (15.2%). There were no differences in the baseline characteristics of the three groups. Ovarian stimulation outcomes, specifically total stimulation days, total gonadotropins administered, and number of oocytes retrieved, were similar between groups. There were no differences in the mean number of embryos transferred. The overall pregnancy outcomes were similar for groups 1, 2, and 3: implantation rate (42.4%, 41.2%, and 42.1%, respectively), clinical pregnancy rate (48.5%, 48.3%, and 48.6%), spontaneous miscarriage rate (4.56%, 4.65%, and 4.05%), and live birth rate (44.0, 43.6%, and 44.6%). CONCLUSION(S): Because waiting for two or more menstrual cycles after hysteroscopic polypectomy does not necessarily yield superior outcomes, patients can undergo ovarian stimulation after their next menses without affecting IVF-ET outcomes.

Comparison of human papillomavirus infection and cervical cytology in women using copper-containing and levonorgestrel-containing intrauterine devices.

OBJECTIVE: To investigate whether there is a difference in cervical cytology and high-risk human papillomavirus (HPV) infection clearance between levonorgestrel- and copper-containing intrauterine device (IUD) users. METHODS: The electronic medical record system was searched by the appropriate procedural code for IUD insertion for all patients undergoing IUD placement during the study period (January 31, 2005 to January 31, 2012). Patients who received treatment for cervical dysplasia, had their IUDs removed, or conceived during the study period were excluded. High-risk HPV and cervical cytology results immediately preceding and after IUD placement were obtained. RESULTS: One hundred fifty patients had a copper-containing and 152 patients a levonorgestrel-containing IUD placed. The groups were comparable in terms of age, body mass index, duration of follow-up, and percentage of smokers. Sixty-six patients were high-risk HPV-positive before IUD insertion (30 in copper compared with 36 in the levonorgestrel IUD group, P=.4), and the groups had similar follow-up times (364.1±26.3 compared with 357.2±29.7 days, respectively, between the IUD placement and a repeat Pap test with high-risk HPV cotesting, P=.2). Of those, 21 (70%; 95% confidence interval [CI] 53.6-86.4%) cleared the infection after copper-containing IUD placement compared with 15 (42%; 95% CI 25.6-57.8%) in the levonorgestrel group (P=.04). There were only two (1.7%) new high-risk HPV infections in the copper compared with eight (6.9%) in the levonorgestrel group (P=.056). CONCLUSION: Our data suggest that levonorgestrel-containing IUD could be associated with decreased high-risk HPV infection clearance and possibly increased acquisition compared with the copper-containing IUD. LEVEL OF EVIDENCE: II.

Geographic disparities amongst patients with gynecologic malignancies at an urban NCI-designated cancer center.

OBJECTIVE: Women with gynecologic malignancies require specialized care. We hypothesize that a geographic disparity exists amongst patients with gynecologic malignancies and that longer distance and time traveled negatively impact completion of adjuvant therapy. METHODS: Patients with incident gynecologic malignancies at a single, urban NCI-designated cancer center were identified. Distances from the patient's home to the treating facility were calculated in miles and minutes. Demographic variables were evaluated for their impact on treatment outcomes using Chi-squared, ANOVA and Kruskal-Wallis analyses. RESULTS: One hundred and fifty consecutive patients were identified. The median distance traveled to the hospital was 16.9miles with a median travel time of 28min. The distance and time traveled were significantly different between insurance groups, with the uninsured traveling the furthest for care by distance (p=0.04) and time (p=0.03). Race, tumor site, medical comorbidities and median income at zip code were not associated with travel distance or time to the hospital. The majority of patients (87%) completed recommended initial treatment. Treatment completion was related to distance traveled with those patients living at the distance extremes (<10miles or >50miles) least likely to complete care (p<0.01). The presence of medical comorbidities (p<0.01) but not insurance status was correlated to treatment completion. CONCLUSIONS: Geographic disparities exist in women with gynecologic malignancies receiving treatment at an NCI-designated cancer center. Approaches to decreasing these disparities may include improved support for cancer patients needing assistance with travel and additional social work and psychosocial support to patients with medical co-morbidities.