Low-input lipidomics reveals lipid metabolism remodelling during early mammalian embryo development.

Lipids are indispensable for energy storage, membrane structure and cell signalling. However, dynamic changes in various categories of endogenous lipids in mammalian early embryonic development have not been systematically characterized. Here we comprehensively investigated the dynamic lipid landscape during mouse and human early embryo development. Lipid signatures of different developmental stages are distinct, particularly for the phospholipid classes. We highlight that the high degree of phospholipid unsaturation is a conserved feature as embryos develop to the blastocyst stage. Moreover, we show that lipid desaturases such as SCD1 are required for in vitro blastocyst development and blastocyst implantation. One of the mechanisms is through the regulation of unsaturated fatty-acid-mediated fluidity of the plasma membrane and apical proteins and the establishment of apical-basal polarity during development of the eight-cell embryo to the blastocyst. Overall, our study provides an invaluable resource about the remodelling of the endogenous lipidome in mammalian preimplantation embryo development and mechanistic insights into the regulation of embryogenesis and implantation by lipid unsaturation.

An innovative design for trophectoderm biopsy without laser pulses: a step-by-step demonstration.

OBJECTIVE: To present a novel trophectoderm biopsy method, independent of laser pulses, using innovatively designed micropipettes on blastocysts at different stages and show variable characteristics. DESIGN: A step-by-step demonstration of this method with narrated video. SETTING: In vitro laboratory fertilization. PATIENTS: Individuals whose embryos underwent preimplantation genetic testing. INTERVENTIONS: Trophectoderm biopsy is accomplished using micropipettes that contain a set of innovative designs. Both biopsy and holding pipettes are characterized by sharp, flat opening ends. The holding pipette is designed with an inclined plane on the outer wall surface of its opening end. It is used to help the biopsy pipette make contact with the holding pipette with increased stability, preventing slipping during the detachment of the trophectoderm cells. There is a narrow structure inside the biopsy pipette, which is designed to trap released fragments and prevent sample loss. A trophectoderm biopsy for fully expanded blastocysts starts from artificial shrinkage, followed by zona pellucida drilling. Then, 5-10 trophectoderm cells are aspirated into a biopsy pipette. The blastocyst is released from the holding pipette, the edge of the opening end of the biopsy pipette is tightly pressed onto the inclined plane of the holding pipette, and the biopsy pipette is flicked directly without laser pulses or pulling off the trophectoderm cells. The aspirated trophectoderm cells are subsequently detached by mechanical friction between the edges of the biopsy and holding pipettes. Apart from drilling the zona pellucida for fully expanded blastocysts, the remaining steps do not require lasers. For hatching (including peanut-shaped and 8-shaped) and hatched blastocysts, a trophectoderm biopsy is accomplished by aspirating the cells without securing the blastocyst with a holding pipette, followed by detachment using the direct flicking method. MAIN OUTCOME MEASURES: The biopsy time, sample loss rate, successful DNA amplification rate, and survival rate. RESULTS: The innovatively designed micropipettes facilitate the successful detachment of trophectoderm cells through a single direct flicking procedure. This eliminates thermal damage caused by laser pulses, notably simplifying operational steps and shortening the biopsy time. Significant differences were noted between the direct flicking and conventional methods, wherein laser pulses and pulling of trophectoderm cells are prerequisites for cell detachment. When comparing the average biopsy time of fully expanded blastocyst (61 ± 8s vs. 104 ± 9s, P<.05), peanut-shaped hatching blastocyst (35 ± 6s vs. 113 ± 13s, P<.05), 8-shaped hatching blastocyst (32 ± 4s vs. 59 ± 6s, P<.05), and hatched blastocyst (34 ± 4s vs. 67 ± 8s, P<.05), the direct flicking method shows a significantly decreased biopsy time. The narrow structure inside the biopsy pipette effectively prevents sample loss, showing a significantly reduced sample loss rate (0%) compared with the conventional biopsy method (18%) for trainees. Moreover, a satisfactory survival rate (100%) and successful DNA amplification rate (99.5%) were achieved using the direct flicking method. CONCLUSIONS: This innovative trophectoderm biopsy method, independent of laser pulses, has wide applicability and a satisfactory, stable performance. Moreover, the simplicity of the method makes it easy to master.

Reactive oxygen species and ovarian diseases: Antioxidant strategies.

Reactive oxygen species (ROS) are mainly produced in mitochondria and are involved in various physiological activities of the ovary through signaling and are critical for regulating the ovarian cycle. Notably, the imbalance between ROS generation and the antioxidant defense system contributes to the development of ovarian diseases. These contradictory effects have critical implications for potential antioxidant strategies that aim to scavenge excessive ROS. However, much remains to be learned about how ROS causes various ovarian diseases to the application of antioxidant therapy for ovarian diseases. Here, we review the mechanisms of ROS generation and maintenance of homeostasis in the ovary and its associated physiological effects. Additionally, we have highlighted the pathological mechanisms of ROS in ovarian diseases and potential antioxidant strategies for treatment.

Effects of immediate versus delayed frozen embryo transfer in high responder patients undergoing freeze-all cycles.

BACKGROUND: Frozen embryo transfer (FET) can greatly improve the pregnancy outcomes for high responder patients. However, it is not known whether the timing of FET is a risk factor on pregnancy outcomes in high responder patients undergoing freeze-all cycles. METHODS: A retrospective cohort study to compare the pregnancy outcomes of the immediate and delayed FET groups in high responder patients undergoing freeze-all cycles. The two groups were defined as that FET took place either within the first menstrual cycle following oocyte retrieval or afterwards. Propensity score matching was used to make the potential risk factors of the two groups comparable. Multivariable regression analysis was used to study the effect of the timing of FET on pregnancy outcomes in the entire cohort and propensity score-matched cohort, even in different controlled ovarian hyperstimulation protocol cohorts as subgroup analysis. RESULTS: We obtained 1130 patients in immediate FET group and 998 patients in delayed FET group, and the average age of the two groups were 30.30 and 30.63. We showed that the immediate FET group were equivalent to delayed FET group in the entire cohort [clinical pregnancy rate (CPR), 61.0% versus 63.4%, adjusted odd ratio (OR), 0.939, 95% confidence interval (CI), 0.781-1.129; spontaneous abortion rate (SAR), 10.1% versus 12.6%, adjusted OR, 0.831, 95% Cl (0.628-1.098); live birth rate (LBR), 49.9% versus 49.2%, adjusted OR, 1.056, 95% Cl (0.883-1.263)]. The same results were obtained by χ2 test in the propensity score-matched cohort (CPR, 60.5% versus 63.5%; SAR, 11.6% versus 12.3%; LBR, 48% versus 49.3%) (P > 0.05). Subgroup analysis indicated that pregnancy outcomes of immediate FET were no difference to delayed FET in gonadotropin-releasing hormone agonist (GnRH-a) protocol (P > 0.05). The SAR of the immediate FET group were lower than that of the delayed FET group in GnRH antagonist protocol (adjusted OR, 0.645, 95% CI, 0.430-0.966) (P < 0.05), no differences were observed in CPR and LBR (P > 0.05). CONCLUSIONS: The pregnancy outcomes of immediate FET were no difference to delayed FET in high responder population undergoing freeze-all cycles.

An effective method for trophectoderm biopsy using mechanical blunt dissection: a step-by-step demonstration.

OBJECTIVE: To present an effective approach to trophectoderm biopsy for blastocysts of different stages and characteristics by mechanical blunt dissection (MBD). DESIGN: Stepwise demonstration with still pictures and operational video clips to explain tips and tricks for trophectoderm biopsy. (This demonstration was approved by the Reproductive Study Ethics Committee at Shengjing Hospital of China Medical University.) SETTING: In vitro fertilization laboratory. PATIENT(S): Patients who underwent preimplantation genetic testing. INTERVENTION(S): The illustrated techniques of blastocyst trophectoderm biopsy using micromanipulation methods include artificial shrinkage, zona pellucida drilling, injecting media from the drilling, aspiration of trophectoderm cells into the biopsy pipette (outer diameter 27 µm for fully expanded blastocysts and peanut-shaped hatching blastocysts; outer diameter 20 µm for 8-shaped hatching and hatched blastocysts), detachment of the trophectoderm cells by laser pulse combined with MBD (performed using the rims of the biopsy and holding pipettes), and release of the biopsy fragment. MAIN OUTCOME MEASURE(S): Successful biopsy rate and survival after warming. RESULT(S): Our biopsy strategy does not involve assisted hatching on day-3 or day-4 embryos, which can leave the embryo undisturbed in culture up to the expanded blastocyst stage. Notably, this approach demonstrates several noteworthy advantages for sampling blastocysts of different stages and characteristics, and it maintains a desirable successful biopsy rate (95.4%, n = 1,872) and survival rate after warming (100%, n = 440). The MBD method may reduce thermal damage because fewer laser pulses are used, compared with the traditional laser-only biopsy techniques. For noncollapsed blastocysts after artificial shrinkage, the strategy of injecting medium from the zona pellucida drilling helps to separate the trophectoderm cells from the zona pellucida, thus facilitating the biopsy procedure. For peanut-shaped hatching blastocysts, this approach could provide better control over the aspiration of trophectoderm cells into the biopsy pipette. Especially if the inner cell mass is herniating from the zona pellucida, the trophectoderm biopsy can be performed away from the inner cell mass to avoid damaging it. In addition, the MBD approach combined with the biopsy pipette (outer diameter 20 µm) can effectively control the target number of trophectoderm cells, thus simplifying the process of obtaining a biopsy from a hatched blastocyst. CONCLUSION(S): Our biopsy approach demonstrates several noteworthy advantages. Considering its benefits and the simplicity of its execution, this systematic biopsy method for blastocysts of different stages and characteristic can be widely applied.

[Performance of computer-assisted imaging system in detection of squamous intraepithelial lesion of uterine cervix].

OBJECTIVE: To evaluate the performance of computer-assisted imaging system in the detection of cervical squamous intraepithelial lesion and quality-assurance. METHODS: Manual PAP screening (n = 140 580) and image-assisted screening (n = 32 885) were compared for the detection rates of squamous cell abnormalities, the atypical squamous cells (ASC) to squamous intraepithelial lesion (SIL) ratio, the positive rates of high risk human papillomavirus (HR-HPV) test in the case of atypical squamous cells of undetermined significance (ASC-US), and the correlation between cytopathology and histopathology. RESULTS: Compared with manual screening, computer-assisted imaging system showed increased overall positive detection by 0.32%, decreased detection of ASC by 0.21%, increased detection of low-grade squamous intraepithelial lesion (LSIL) by 0.22%, increased detection of high-grade squamous intraepithelial lesion or worse (HSIL) by 0.31%, and decreased ASC to SIL ratio from 2.59 to 1.60. Computer-assisted imaging system did not change the HR-HPV positive rate of the patients who were ASC-US, or the coincidence rate between cytopathology and histopathology. Moreover, the productivity of the laboratory operation increased 58.33%. CONCLUSION: Computer-assisted imaging system significantly increases the overall positive detection rate of cervical SIL, improves accuracy and work efficiency of screening, decreases the ASC/SIL rate, and strengths the quality-assurance of laboratory testing.