Proteomic analysis of extracellular vesicles secreted by primary human epithelial endometrial cells reveals key proteins related to embryo implantation

Background Successful implantation is dependent on coordination between maternal endometrium and embryo, and the role of EVs in the required cross-talk cell-to-cell has been recently established. In this regard, it has been reported that EVs secreted by the maternal endometrium can be internalized by human trophoblastic cells transferring their contents and enhancing their adhesive and invasive capacity. This is the first study to comprehensively evaluate three EV isolation methods on human endometrial epithelial cells in culture and to describe the proteomic content of EVs secreted by pHEECs from fertile women. Methods Ishikawa cells and pHEECs were in vitro cultured and hormonally treated; subsequently, conditioned medium was collected and EVs isolated. Ishikawa cells were used for the comparison of EVs isolation methods ultracentrifugation, ExoQuick-TC and Norgen Cell Culture Media Exosome Purification Kit (n = 3 replicates/isolation method). pHEECs were isolated from endometrial biopsies (n = 8/replicate; 3 replicates) collected from healthy oocyte donors with confirmed fertility, and protein content of EVs isolated by the most efficient methodology was analysed using liquid chromatography–tandem mass spectrometry. EV concentration and size were analyzed by nanoparticle tracking analysis, EV morphology visualized by transmission electron microscopy and protein marker expression was determined by Western blotting. Results Ultracentrifugation was the most efficient methodology for EV isolation from medium of endometrial epithelial cells. EVs secreted by pHEECs and isolated by ultracentrifugation were heterogeneous in size and expressed EV protein markers HSP70, TSG101, CD9, and CD81. Proteomic analysis identified 218 proteins contained in these EVs enriched in biological processes involved in embryo implantation, including cell adhesion, differentiation, communication, migration, extracellular matrix organization, vasculature development, and reproductive processes. From these proteins, 82 were selected based on their functional relevance in implantation success as possible implantation biomarkers. Conclusions EV protein cargos are implicated in biological processes related to endometrial receptivity, embryo implantation, and early embryo development, supporting the concept of a communication system between the embryo and the maternal endometrium via EVs. Identified proteins may define new biomarkers of endometrial receptivity and implantation success.

Pre-implantation mouse embryo movement under hormonally altered conditions

Precise regulation of embryo movement is crucial to successful implantation, but the role of ovarian hormones in this process is not understood. We ascertain the effects of altered hormonal environment on embryo movement using two delayed implantation models: Natural lactational Diapause (ND), a naturally occurring alternate model of pregnancy, and Artificially induced Diapause (AD), a laboratory version of ND. Our previous work suggests that embryos in a natural pregnancy (NP) first display unidirectional clustered embryo movement, followed by bidirectional scattering and spacing movement. In contrast, in the ND model, embryos are present as clusters near the oviductal-uterine junction for ~24-hours longer than NP, followed by locations consistent with a unidirectional scattering and spacing movement. Intriguingly, the AD model closely resembles embryo location in NP and not ND. Further, unlike the popular paradigm of reduced estrogen (E2) levels in diapause E2 levels are comparable across NP, ND, and AD, while progesterone (P4) levels are reduced in ND and highly increased in AD when compared to NP. Exogenous administration of E2 or P4 modifies the unidirectional clustered embryo movement, while E2 treatment causes a reduction in P4 and affects the bidirectional phase of embryo movement. Taken together, our data suggest embryo movement can be modulated by both P4 and E2. Understanding natural hormonal adaptation in diapause provides an opportunity to determine key players regulating embryo movement and implantation success. This knowledge can be leveraged to understand pregnancy survival and implantation success in hormonally altered conditions in the clinic.

Optimal WovenEndo Bridge (WEB) Device Size Selection Using Automated Volumetric Software

Introduction: Selecting the appropriate Woven EndoBridge (WEB) device sizing for the treatment of wide-neck bifurcation aneurysms (WNBAs) remains challenging. The aim of this study was to evaluate different volumetric-based imaging methodologies to predict an accurate WEB device size selection to result in a successful implantation. Methods: All consecutive patients treated with WEB devices for intracranial aneurysms from January 2019 to June 2020 were included. Aneurysm dimensions to calculate aneurysm volumes were measured using three different modalities: automated three-dimensional (3D) digital subtraction angiography (DSA), manual 3D DSA, and two-dimensional (2D) DSA. The device–aneurysm volume (DAV) ratio was defined as device volume divided by the aneurysm volume. WEB volumes and the DAV ratios were used for assessing the device implantation success and follow-up angiographic outcomes at six months. Pearson correlation, Wilcoxon Rank Sum test, and flow density approximations were used for estimating the WEB volumes and the imaging modality volumes for successful implantation. Results: A total of 41 patients with 43 aneurysms were included in the study. WEB device and aneurysm volume correlation coefficient was highest for 3D automatic (r = 0.943), followed by 3D manual (r = 0.919), and 2D DSA (r = 0.882) measurements. Measured median volumes were significantly different for 3D automatic and 2D DSA (p = 0.017). The highest rate of successful implantation (87.5%) was between 0.6 and 0.8 DAV ratio. Conclusion: Pre-procedural assessment of DAV ratios may increase WEB device implantation success. Our results suggest that volumetric measurements, especially using automated 3D volumes of the aneurysms, can assist in accurate WEB device size selection.

Influence of Prior Major Bleeding on the Safety and Effectiveness of Left Atrial Appendage Closure in Non-Valvular Atrial Fibrillation Patients: A Single-Center Study

Purpose: Left atrial appendage closure (LAAC) may present an alternative to anticoagulation in patients with non-valvular atrial fibrillation (NVAF). However, evidence regarding benefit of LAAC in patients with prior major bleeding is limited. We evaluate whether a previous bleeding factor influences the safety and effectiveness of LAAC. Methods: A total of 377 consecutive patients scheduled for LAAC were categorized into a bleeding group (n=137) and non-bleeding group (n=240). The implantation success and prevalence of severe peri-procedural complications, and efficacy/ safety endpoints during follow-up were investigated. Results: The bleeding group had more patients ≥75 years old (P=0.044), higher CHA2DS2-VASc (P=0.029) and HAS-BLED scores (P=0.001) than the non-bleeding group. Implantation success and severe peri-procedural complications were similar. During an average 2 years’ follow-up, major bleeding events (P=0.917), all-cause death (P=0.313), co-primary efficacy events (P=0.063), and the cumulative survival ratio (P=0.828) were comparable. However, the incidence of thromboembolism was lower in the bleeding group (P=0.031). The observed thromboembolism rate was reduced by 86.1% (P<0.005) and 43.9% (P<0.05) and the observed bleeding rate was decreased by 39.0% (P<0.05) and 34.8% (P<0.05) in the bleeding and non-bleeding groups, respectively. The extent of relative risk reduction in thromboembolism was significantly higher in the bleeding group (86.1% vs. 43.9%, P<0.01). Conclusion: LAAC may provide a safe and effective alternative to long-term antithrombotic therapy in NVAF patients with or without prior bleeding. The efficacy of LAAC in reducing thromboembolism instead of major bleeding seems to be higher in patients with versus without prior bleeding.

The Effect of Embryo Transfer Process Duration on Implantation Success

OBJECTIVE: To evaluate the effect of the embryo transfer duration of standard and simple embryo transfer method. STUDY DESIGN: This study was a retrospective cohort study conducted at a tertiary ART Centre, between June 2018- September 2018. Day 5 fresh embryo transferred patients aged between 18 - 40, BMI <35 kg/m2 without uterine pathology were enrolled in the study. Patients were divided into two groups. Group-1 consisted of patients who had successful implantation and Group-2 consisted of patients who did not have implantation. Groups were compared according to their embryo transfer durations. Ninety-two patients were enrolled in the study. Also, sub-steps of as; cleaning of the cervical mucus and placing the outer catheter in the cervix, loading the embryo to the catheter, the period between embryo loading and embryo transfer, and following that, time spent for retracting the outer catheter evaluated. RESULTS: Between Group-1 and Group-2, there was no significant difference for the period of cervical cleaning and placing the outer catheter into the cervix (Respectively; 63 sec vs. 76 sec; p=0.18), the period of embryo loading (Respectively; 69sec vs. 71sec; p=0.46), the period between embryo loading and embryo transfer (Respectively; 10 sec vs. 10 sec; p=0.74, retracing the outer catheter (Respectively; 25.5sec vs. 24sec; p=0.42 and the total period of embryo transfer (182sec vs. 182.5 sec; p=0.55). CONCLUSION: The embryo transfer duration is not related to implantation rates. The duration of the embryo transfer process steps is not a distinguishing factor if a good-quality embryo transfer is done.

Male-derived copulatory plugs enhance implantation success in female Mus musculus†

Among a wide diversity of sexually reproducing species, male ejaculates coagulate to form what has been termed a copulatory plug. A number of functions have been attributed to copulatory plugs, including, but not limited to, the inhibition of female remating and the promotion of ejaculate movement. Here we demonstrate that copulatory plugs also influence the likelihood of implantation, which occurs roughly four days after copulation in mice. Using a bead transfer method to control for differences in ejaculate retention and fertilization rates, we show that implantation rates significantly drop among females mated to genetically engineered males incapable of forming plugs (because they lack functional TGM4, the main enzyme responsible for its formation). Surprisingly, this result does not correlate with differences in circulating progesterone levels among females, an important hormone involved in implantation. In this paper we discuss three models that connect male-derived copulatory plugs to implantation success, including the hypothesis that plugs contribute to a threshold amount of stimulation required for females to become receptive to implantation.

P1480French experience of SICD implantation

OnBehalf GPUR Introduction The subcutaneous ICD prevents complications of transvenous leads. Its implantation needs a defibrillation test. Performing this test increases the time of procedures.. Purpose The aim of our study is to describe the experience of subcutaneous ICD of Assitance Publique des Hopitaux de Paris (AP-HP). Materials and Methods In a retrospective cohort we included patients who were involved in subcutaneous ICD treatment at the 5 hospitals of AP-HP from December 2012 to April 2018.Analysis used the Kaplan-Meier method and the Mann-Whitney U test. Results 162 patients were implanted. 76% of implantations were for secondary prevention before 2017, but only 49% after 2017. 126 (77%) tests were successful at first shock, 14 (8.6%) needed a second shock or more, 9 (5.5%) did not have a test. The shock impedance was significantly higher when the shock was not effective at the first test, 82 ohms CI 95% [68; 112] versus 66 ohms CI 95% [64; 70], p &lt;0.05. 6 patients died (3.7%). Late follow up showed 23 (10%) appropriate shocks and 24 inappropriate shocks (11%) mainly due to T wave oversensing (37.5%). Conclusions This is the first french experience of SICD implantation. Success of defibrillation test was lower than expected. Caracteristics of implantation General anesthesia (%) 162 (100) Procedure time (min) Mean ± SD (median)Range 73± 24 (60)20-165 Cameron Health (%) Generator SQ-RX ® 1010 12 (7.4) Boston Scientific (%) Generator EMBLEM ® A209 100 (61.7) Generator EMBLEM ® A219 44 (27.2) VF time (sec) Mean ± SD (median)Range 16.5± 3.4(15)10-37 Shock Impedance (ohm) Mean ± SD (median) Range 72 ± 12.7 (68) 42-130 Success at 1st shock (%) 126 (77.7) Success at 2nd shock or more (%) 14 (8.6) No test performed (%) 12 (7.4) Implantation caracteristics and defibrillation testing. Impedance was significantly higher in patients without success at first shock : 82 ohms CI 95% [68 ;112] vs 66 ohms CI 95% [64 ;70], p &lt; 0.05. Abstract Figure. Survival without inappropriate shock