Objective: To examine the impact of laser-assisted hatching (LAH) technique on perinatal outcomes in blastocyst culture of low-grade cleavage embryos, so as to provide insights into improving the utilization of low-grade cleavage embryos and embryo quality. Methods: A total of 369 single live births after transfer of thawed blastocysts following in vitro fertilization or intracytoplasmic sperm injection at Zhejiang Provincial People's Hospital were selected as subjects, and they were divided into 51 conventional culture blastocysts and 318 LAH blastocysts based on whether LAH was performed on day 4 of blastocyst culture. Gestational age, birth weight, birth defects and maternal perinatal information were collected, and the prevalence of premature birth, birth weight and birth defects were analyzed after propensity score matching (PSM). Results: After PSM, 98 matched cases were included in the study. There were no statistically significant differences in maternal age, body mass index, type of infertility and blastocyst age (P>0.05), indicating effective matching. The birth weight of offspring in the LAH group was lower than that in the conventional culture group [(3 261.08±432.24) g vs. (3 464.18±444.46) g; P<0.05]. Conclusion The birth weight of offspring can be reduced by using LAH during blastocyst culture of low-grade cleavage embryos.

Objective:To investigate the effect of dydrogesterone (DYD) on the ultrastructure of the endometrium in the window of implantation.Methods:We prospectively studied endometrium tissues of patients who underwent in vitro fertilization and embryo transfer (IVF-ET) from July 2020 to July 2022 at the Center for Reproductive Medicine of Zhejiang Provincial People's Hospital. The ultrastructure of mid-luteal endometrium was examined by scanning electron microscopy and transmission electron microscopy. Firstly, we used self-control study to compare the ultrastructure of endometrium between the natural cycle group and the DYD 40 mg/d group in 20 patients with male factor infertility. Secondly, we used self-control study to compare the ultrastructure of the endometrium between the DYD 40 mg/d group and the DYD 80 mg/d group in 43 patients with recurrent implantation failure (RIF). Results:There were no significant differences in endometrial thickness, ultrasound grading, positivity rate of nucleolar channel system (NCS), pinopode score and secretor status score, in addition to number and morphology score of mitochondria, between the natural cycle group and the DYD 40 mg/d group (all P>0.05). In RIF patients, the DYD 80 mg/d group rendered significantly higher pinopode score (-46.26±19.97), NCS positivity rate [65.12% (28/43)] and mitochondrial morphology score [0 (0, 1)] than those in the DYD 40 mg/d group [-67.62±15.94, P<0.001; 39.53% (17/43), P=0.043; 0 (0, 0), P=0.023], while other characteristics were not significantly different between the two groups (all P>0.05). Conclusion:The ultrastructure of endometrium in DYD supplementation cycle was similar to that in the natural cycle. Increasing the dosage of DYD significantly improved pinopode maturity, NCS positivity rate and mitochondrial morphology of the endometrium in RIF patients.

Objective:To investigate the effect of dydrogesterone (DYD) on the ultrastructure of the endometrium in the window of implantation.Methods:We prospectively studied endometrium tissues of patients who underwent in vitro fertilization and embryo transfer (IVF-ET) from July 2020 to July 2022 at the Center for Reproductive Medicine of Zhejiang Provincial People's Hospital. The ultrastructure of mid-luteal endometrium was examined by scanning electron microscopy and transmission electron microscopy. Firstly, we used self-control study to compare the ultrastructure of endometrium between the natural cycle group and the DYD 40 mg/d group in 20 patients with male factor infertility. Secondly, we used self-control study to compare the ultrastructure of the endometrium between the DYD 40 mg/d group and the DYD 80 mg/d group in 43 patients with recurrent implantation failure (RIF). Results:There were no significant differences in endometrial thickness, ultrasound grading, positivity rate of nucleolar channel system (NCS), pinopode score and secretor status score, in addition to number and morphology score of mitochondria, between the natural cycle group and the DYD 40 mg/d group (all P>0.05). In RIF patients, the DYD 80 mg/d group rendered significantly higher pinopode score (-46.26±19.97), NCS positivity rate [65.12% (28/43)] and mitochondrial morphology score [0 (0, 1)] than those in the DYD 40 mg/d group [-67.62±15.94, P<0.001; 39.53% (17/43), P=0.043; 0 (0, 0), P=0.023], while other characteristics were not significantly different between the two groups (all P>0.05). Conclusion:The ultrastructure of endometrium in DYD supplementation cycle was similar to that in the natural cycle. Increasing the dosage of DYD significantly improved pinopode maturity, NCS positivity rate and mitochondrial morphology of the endometrium in RIF patients.

Objective:To establish and evaluate a follicle isolation method, combination of enzymatically digestion with strainer filtration, for synchronously collecting mouse follicles across different developmental stages.Methods:Pre-cutted ovarian tissue blocks were digested with enzymes mixed by Collagenase I and DNase I, and then filtered through three different pore size cell strainers to synchronously separate the follicles at developmental stages. The follicles obtained by turning over and washing the 100 μm strainer were recorded as group A (>100 μm), as same as the follicles obtained by turning over and washing the 40 μm strainer were recorded as group B (40-100 μm), and the follicles obtained by turning over and washing the 20 μm strainer were recorded as group C (20-40 μm). The quantity, morphology, viability and developmental potency were examined among these harvested follicles.Results:In terms of quantity, follicles in group C accounted for most, followed by follicles in group A, and the follicles in group B were the least. Morphologically, the basal membrane integrity rate of follicles in group C was higher than that of groups B and A ( P<0.001 and P<0.001, respectively). As for viability, the survival rate of follicles in group A was 70.59% (120/170), which was higher than that in group B (51.79%, 58/112) and group C (35.90%, 28/78) ( P=0.001 6 and P<0.001, respectively). In terms of developmental potency, after 96 h of in vitro culture, the diameter of follicles in group A increased from (113.64±9.57) μm to (150.95±45.90) μm ( P=0.002 4), and the diameter of follicles in group B increased from (88.12±9.12) μm to (120.61±18.00) μm ( P<0.001). In group C, monolayer-layer granulosa cells were attached to the follicles within 24 h of culture, and the connection structure between oocytes and granulosa cells was lost. The oocytes were completely exposed and failed to be cultured successfully. Conclusion:Combining enzymatically digestion with multiple strainers filtration is a rapid and effective method for follicle collecting, which is capable to isolate different developmental stage mouse follicles synchronously with morphological integrity, favorable viability and good developmental potency.

Objective:To establish and evaluate a follicle isolation method, combination of enzymatically digestion with strainer filtration, for synchronously collecting mouse follicles across different developmental stages.Methods:Pre-cutted ovarian tissue blocks were digested with enzymes mixed by Collagenase I and DNase I, and then filtered through three different pore size cell strainers to synchronously separate the follicles at developmental stages. The follicles obtained by turning over and washing the 100 μm strainer were recorded as group A (>100 μm), as same as the follicles obtained by turning over and washing the 40 μm strainer were recorded as group B (40-100 μm), and the follicles obtained by turning over and washing the 20 μm strainer were recorded as group C (20-40 μm). The quantity, morphology, viability and developmental potency were examined among these harvested follicles.Results:In terms of quantity, follicles in group C accounted for most, followed by follicles in group A, and the follicles in group B were the least. Morphologically, the basal membrane integrity rate of follicles in group C was higher than that of groups B and A ( P<0.001 and P<0.001, respectively). As for viability, the survival rate of follicles in group A was 70.59% (120/170), which was higher than that in group B (51.79%, 58/112) and group C (35.90%, 28/78) ( P=0.001 6 and P<0.001, respectively). In terms of developmental potency, after 96 h of in vitro culture, the diameter of follicles in group A increased from (113.64±9.57) μm to (150.95±45.90) μm ( P=0.002 4), and the diameter of follicles in group B increased from (88.12±9.12) μm to (120.61±18.00) μm ( P<0.001). In group C, monolayer-layer granulosa cells were attached to the follicles within 24 h of culture, and the connection structure between oocytes and granulosa cells was lost. The oocytes were completely exposed and failed to be cultured successfully. Conclusion:Combining enzymatically digestion with multiple strainers filtration is a rapid and effective method for follicle collecting, which is capable to isolate different developmental stage mouse follicles synchronously with morphological integrity, favorable viability and good developmental potency.