Multiple collapses of blastocysts after full blastocyst formation is an independent risk factor for aneuploidy - a study based on AI and manual validation.

BACKGROUND: The occurrence of blastocyst collapse may become an indicator of preimplantation embryo quality assessment. It has been reported that collapsing blastocysts can lead to higher rates of aneuploidy and poorer clinical outcomes, but more large-scale studies are needed to explore this relationship. This study explored the characteristics of blastocyst collapse identified and quantified by artificial intelligence and explored the associations between blastocyst collapse and embryo ploidy, morphological quality, and clinical outcomes. METHODS: This observational study included data from 3288 biopsied blastocysts in 1071 time-lapse preimplantation genetic testing cycles performed between January 2019 and February 2023 at a single academic fertility center. All transferred blastocysts are euploid blastocysts. The artificial intelligence recognized blastocyst collapse in time-lapse microscopy videos and then registered the collapsing times, and the start time, the recovery duration, the shrinkage percentage of each collapse. The effects of blastocyst collapse and embryo ploidy, pregnancy, live birth, miscarriage, and embryo quality were studied using available data from 1196 euploid embryos and 1300 aneuploid embryos. RESULTS: 5.6% of blastocysts collapsed at least once only before the full blastocyst formation (tB), 19.4% collapsed at least once only after tB, and 3.1% collapsed both before and after tB. Multiple collapses of blastocysts after tB (times ≥ 2) are associated with higher aneuploid rates (54.6%, P > 0.05; 70.5%, P < 0.001; 72.5%, P = 0.004; and 71.4%, P = 0.049 in blastocysts collapsed 1, 2, 3 or ≥ 4 times), which remained significant after adjustment for confounders (OR = 2.597, 95% CI 1.464-4.607, P = 0.001). Analysis of the aneuploid embryos showed a higher ratio of collapses and multiple collapses after tB in monosomies and embryos with subchromosomal deletion of segmental nature (P < 0.001). Blastocyst collapse was associated with delayed embryonic development and declined blastocyst quality. There is no significant difference in pregnancy and live birth rates between collapsing and non-collapsing blastocysts. CONCLUSIONS: Blastocyst collapse is common during blastocyst development. This study underlined that multiple blastocyst collapses after tB may be an independent risk factor for aneuploidy which should be taken into account by clinicians and embryologists when selecting blastocysts for transfer.

The density of the inner cell mass is a new indicator of the quality of a human blastocyst: a valid supplement to the Gardner scoring system.

STUDY QUESTION: Can the density of the inner cell mass (ICM) be a new indicator of the quality of the human blastocyst? SUMMARY ANSWER: The densification index (DI) developed in this study can quantify ICM density and provide positive guidance for ploidy, pregnancy, and live birth. WHAT IS KNOWN ALREADY: In evaluating the quality of ICM, reproductive care clinics still use size indicators without further evaluation. The main disadvantage of this current method is that the evaluation of blastocyst ICM is relatively rough and cannot meet the needs of clinical embryologists, especially when multiple blastocysts have the same ICM score, which makes them difficult to evaluate further. STUDY DESIGN, SIZE, DURATION: This observational study included data from 2272 blastocysts in 1991 frozen-thawed embryo transfer (FET) cycles between January 2018 to November 2021 and 1105 blastocysts in 430 preimplantation genetic testing cycles between January 2019 and February 2023. PARTICIPANTS/MATERIALS, SETTING, METHODS: FET, ICSI, blastocyst culture, trophectoderm biopsy, time-lapse (TL) monitoring, and next-generation sequencing were performed. After preliminary sample size selection, the 11 focal plane images captured by the TL system were normalized and the spatial frequency was used to construct the DI of the ICM. MAIN RESULTS AND THE ROLE OF CHANCE: This study successfully constructed a quantitative indicator DI that can reflect the degree of ICM density in terms of fusion and texture features. The higher the DI value, the better the density of the blastocyst ICM, and the higher the chances that the blastocyst was euploid (P < 0.001) and that pregnancy (P < 0.001) and live birth (P = 0.005) were reached. In blastocysts with ICM graded B and blastocysts graded 4BB, DI was also positively associated with ploidy, pregnancy, and live birth (P < 0.05). ROC analysis showed that combining the Gardner scoring system with DI can more effectively predict pregnancy and live births, when compared to using the Gardner scoring system alone. LIMITATIONS, REASONS FOR CAUTION: Accurate calculation of the DI value places high demands on image quality, requiring manual selection of the clearest focal plane and exposure control. Images with the ICM not completely within the field of view cannot be used. The association between the density of ICM and chromosomal mosaicism was not evaluated. The associations between the density of ICM and different assisted reproductive technologies and different culture conditions in embryo laboratories were also not evaluated. Prospective studies are needed to further investigate the impact of ICM density on clinical outcomes. WIDER IMPLICATIONS OF THE FINDINGS: ICM density assessment is a new direction in blastocyst assessment. This study explores new ways of assessing blastocyst ICM density and develops quantitative indicators and a corresponding qualitative evaluation scheme for ICM density. The DI of the blastocyst ICM developed in this study is easy to calculate and requires only TL equipment and image processing, providing positive guidance for clinical outcomes. The qualitative evaluation scheme of ICM density can assist embryologists without TL equipment to manually evaluate ICM density. ICM density is a simple indicator that can be used in practice and is a good complement to the blastocyst scoring systems currently used in most centers. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Key Research & Development Program of China (2021YFC2700603). The authors report no financial or commercial conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.

A novel non-invasive embryo evaluation method (NICS-Timelapse) with enhanced predictive precision and clinical impact.

The selection of the finest possible embryo in in-vitro fertilization (IVF) was crucial and revolutionary, particularly when just one embryo is transplanted to lessen the possibility of multiple pregnancies. However, practical usefulness of currently used methodologies may be constrained. Here, we established a novel non-invasive embryo evaluation method that combines non-invasive chromosomal screening (NICS) and Timelapse system along with artificial intelligence algorithms. With an area under the curve (AUC) of 0.94 and an accuracy of 0.88, the NICS-Timelapse model was able to predict blastocyst euploidy. The performance of the model was further evaluated using 75 patients in various clinical settings. The clinical pregnancy and live birth rates of embryos predicted by the NICS-Timelapse model, showing that embryos with higher euploid probabilities were associated with higher clinical pregnancy and live birth rates. These results demonstrated the NICS-Timelapse model's significantly wider application in clinical IVF due to its excellent accuracy and noninvasiveness.

Anti-EGFR ScFv functionalized exosomes delivering LPCAT1 specific siRNAs for inhibition of lung cancer brain metastases.

Brain metastasis (BM) is one of the leading causes of cancer-related deaths in patients with advanced non-small cell lung cancer (NSCLC). However, limited treatments are available due to the presence of the blood-brain barrier (BBB). Upregulation of lysophosphatidylcholine acyltransferase 1 (LPCAT1) in NSCLC has been found to promote BM. Conversely, downregulating LPCAT1 significantly suppresses the proliferation and metastasis of lung cancer cells. In this study, we firstly confirmed significant upregulation of LPCAT1 in BM sites compared to primary lung cancer by analyzing scRNA dataset. We then designed a delivery system based on a single-chain variable fragment (scFv) targeting the epidermal growth factor receptor (EGFR) and exosomes derived from HEK293T cells to enhance cell-targeting capabilities and increase permeability. Next, we loaded LPCAT1 siRNA (siLPCAT1) into these engineered exosomes (exoscFv). This novel scFv-mounted exosome successfully crossed the BBB in an animal model and delivered siLPCAT1 to the BM site. Silencing LPCAT1 efficiently arrested tumor growth and inhibited malignant progression of BM in vivo without detectable toxicity. Overall, we provided a potential platform based on exosomes for RNA interference (RNAi) therapy in lung cancer BM.

Intermittent short-duration reoxygenation relieves high-altitude pulmonary hypertension via NOX4/H2O2/PPAR-γ axis.

High-altitude pulmonary hypertension (HAPH) is a severe and progressive disease that can lead to right heart failure. Intermittent short-duration reoxygenation at high altitude is effective in alleviating HAPH; however, the underlying mechanisms are unclear. In the present study, a simulated 5,000-m hypoxia rat model and hypoxic cultured pulmonary artery smooth muscle cells (PASMCs) were used to evaluate the effect and mechanisms of intermittent short-duration reoxygenation. The results showed that intermittent 3-h/per day reoxygenation (I3) effectively attenuated chronic hypoxia-induced pulmonary hypertension and reduced the content of H2O2 and the expression of NADPH oxidase 4 (NOX4) in lung tissues. In combination with I3, while the NOX inhibitor apocynin did not further alleviate HAPH, the mitochondrial antioxidant MitoQ did. Furthermore, in PASMCs, I3 attenuated hypoxia-induced PASMCs proliferation and reversed the activated HIF-1α/NOX4/PPAR-γ axis under hypoxia. Targeting this axis offset the protective effect of I3 on hypoxia-induced PASMCs proliferation. The present study is novel in revealing a new mechanism for preventing HAPH and provides insights into the optimization of intermittent short-duration reoxygenation.

Usable blastocysts developed from in-vitro-matured metaphase I oocytes in preimplantation genetic testing cycles.

RESEARCH QUESTION: Are blastocysts derived from in-vitro-matured metaphase I (MI) oocytes less likely to produce usable embryos for transfer compared with those derived from in-vivo-matured oocytes in cycles undergoing preimplantation genetic testing (PGT)? DESIGN: The primary outcome was usable blastocyst rate, which was compared between blastocysts derived from in-vitro-matured MI oocytes after ovarian stimulation and from in-vivo-matured oocytes. Logistic regression analysis using generalized estimating equations was used to control for confounders in the analysis of factors that may influence the chance of a blastocyst being usable and in the comparison of embryological outcomes. Student's t-test, Mann-Whitney U test, chi-squared tests or Fisher's exact tests were used to compare clinical and pregnancy outcomes. RESULTS: A total of 1810 injected metaphase II (MII) oocytes from 154 PGT cycles involving 154 couples were included in this study. A total of 1577 MII oocytes were in-vivo-matured and 233 were in-vitro-matured MI oocytes. The usable blastocyst rate was similar between the in-vitro-matured MI oocyte group and the in-vivo-matured oocyte group (adjusted RR 0.97, 95% CI 0.40 to 2.34). Three live births were achieved using usable blastocysts derived from in-vitro-matured MI oocytes. CONCLUSIONS: If in-vitro-matured MI oocytes can be fertilized and develop into blastocysts, their ability to provide usable embryos for transfer is similar compared with those developed from in-vivo-matured oocytes. These blastocysts could be considered valuable for women with few viable embryos in assisted reproductive technology cycles.

Cumulative live birth rate and neonatal outcomes after early rescue ICSI: a propensity score matching analysis.

STUDY QUESTION: Is early rescue ICSI (E-RICSI) an effective and safe technique compared to conventional ICSI? SUMMARY ANSWER: Despite the higher multi-pronucleus (PN) rate compared to conventional ICSI, E-RICSI did not add extra risks to clinical and neonatal outcomes. WHAT IS KNOWN ALREADY: Based on the finding that the second polar body was released in 80% of fertilized oocytes by 4 h after exposure to spermatozoa and in ∼90% of fertilized oocytes by 6 h, E-RICSI brings forward the timing of rescue ICSI to 6 h after initial insemination, and effectively prevents oocyte aging and embryo-uterus asynchrony. However, some researchers still voice concerns about the efficacy and safety of E-RICSI, and comparative studies are limited. STUDY DESIGN SIZE DURATION: A retrospective cohort study was conducted on patients who underwent conventional ICSI or E-RICSI treatment between January 2015 and December 2020 at a university-affiliated hospital. Using 1:1 propensity score matching, 1496 cases entered each group. PARTICIPANTS/MATERIALS SETTING METHODS: In total, 1496 couples undergoing conventional ICSI oocyte retrieval cycles and 1496 undergoing E-RICSI oocyte retrieval cycles were enrolled in this study, and basic clinical characteristics, embryologic data, clinical outcomes and neonatal data were compared between groups. The embryos in the E-RICSI group were divided into two subgroups: those fertilized by iIVF (IVF subgroup) and those fertilized by E-RICSI (E-RICSI subgroup); the embryologic data, clinical outcomes, and neonatal data for these subgroups were also compared with the conventional ICSI group. Logistic regression was used for statistical analysis with potential confounder adjustment. MAIN RESULTS AND THE ROLE OF CHANCE: The 2PN rate, blastocyst formation rate, and viable blastocyst formation rate of the E-RICSI group were significantly lower compared to the conventional ICSI group (2PN rate: P < 0.001; blastocyst formation rate: P < 0.001; viable blastocyst formation rate: P = 0.004), and the multi-PN rate in the E-RICSI group was significantly higher than the conventional ICSI group (P < 0.001). However, the number of 2PN embryos, normal cleavage embryo rate, Day 3 high-quality cleavage embryo rate, and high-quality blastocyst rate were similar between groups. When considering the IVF embryos and E-RCSI embryos in the E-RICSI group independently, the 2PN rate of the conventional ICSI group was significantly lower than E-RICSI subgroup but higher than the IVF subgroup, whereas the blastocyst formation rate and viable blastocyst formation rate were higher than E-RICSI embryos but comparable to IVF embryos. As for the clinical and neonatal outcomes, the implantation rate of the E-RICSI subgroup was significantly lower than the IVF subgroup but comparable to the conventional ICSI group, while the low birthweight (LBW) rate was significantly lower compared with the conventional ICSI group but similar with the IVF subgroup. No other differences were observed among the three groups for cumulative clinical pregnancy rate, cumulative live birth rate, and the pregnancy outcomes per transfer including clinical pregnancy, ectopic pregnancy, miscarriage, and live birth, either in fresh or frozen embryo transfer cycles. Furthermore, neonatal outcomes, including cesarean section, sex ratio, LBW, preterm birth, and macrosomia, were similar among groups. LIMITATIONS REASONS FOR CAUTION: This study is limited by the retrospective design, limited sample size, and short follow-up period. However, our study underlies the need for large-scale, multi-center randomized controlled trials with long-term follow-up. WIDER IMPLICATIONS OF THE FINDINGS: Short-term insemination (3 h) combined with E-RICSI may be a safe and effective method to prevent the occurrence of total fertilization failure, and patients with normal or borderline sperm could be encouraged to try IVF first. STUDY FUNDING/COMPETING INTERESTS: This study was supported by grants from the National Key & Development Program of China (No. 2021YFC2700603) and the National Natural Science Foundation of China (No. 81801443). The authors declare no conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.