Immature Oocytes Research Articles

Pro-cumulin addition in a biphasic in vitro oocyte maturation system modulates human oocyte and cumulus cell transcriptomes.

Biphasic in vitro oocyte maturation (IVM) can be offered as a patient-friendly alternative to conventional ovarian stimulation in in vitro fertilization (IVF) patients predicted to be hyper-responsive to ovarian stimulation. However, cumulative live birth rates after IVM per cycle are lower than after conventional ovarian stimulation for IVF. In different animal species, supplementation of IVM media with oocyte-secreted factors (OSFs) improves oocyte developmental competence through the expression of pro-ovulatory genes in cumulus cells. Whether the addition of OSFs in human biphasic IVM culture impacts the transcriptome of oocytes and cumulus cells retrieved from small antral follicles in minimally stimulated non-hCG-triggered IVM cycles remains to be elucidated. To answer this, human cumulus oocyte complexes (COCs) that were fully surrounded by cumulus cells or partially denuded at the time of retrieval were cultured in a biphasic IVM system either without or with the addition of pro-cumulin, a GDF9: BMP15 heterodimer. Oocytes and their accompanying cumulus cells were collected separately, and single-cell RNA-seq libraries were generated. The transcriptomic profile of cumulus cells revealed that pro-cumulin upregulated the expression of genes involved in cumulus cell expansion and proliferation while downregulating steroidogenesis, luteinization and apoptosis pathways. Moreover, pro-cumulin modulated the immature oocyte transcriptome during the prematuration step, including regulating translation, apoptosis and mitochondria remodeling pathways in the growing germinal vesicle (GV) oocytes. The addition of pro-cumulin also restored the transcriptomic profile of matured metaphase II (MII) oocytes that were partially denuded at collection. These results suggest that cumulus cell and oocyte transcriptome regulation by pro-cumulin may increase the number of developmentally competent oocytes after biphasic IVM treatment. Future studies should assess the effects of pro-cumulin addition in human biphasic IVM at the proteomic level and the embryological outcomes, particularly its potential to enhance outcomes of oocytes that are partially denuded at COC collection.

Impact of follicular size categories on oocyte quality at trigger day undergoing luteal phase stimulation protocol.

To investigate an optimal strategy by assessing the effectiveness of varying follicular sizes on trigger day during luteal phase stimulation protocol and provide evidence for personalized protocol adjustment. This was a retrospective study including a total of 661 patients who had started their in vitro fertilization cycle with a luteal phase stimulation (LPS) protocol during 2015-2023. We classified patients into groups according to the size of the dominant proportion of follicles on the human chorionic gonadotropin (hCG) trigger day: large, medium, and small. The metaphase II (MII) oocyte rate, immature oocyte rate, two pronuclei (2PN) fertilization rate, and available embryo rate were compared between groups. General linear model (GLM) analysis was performed for inter-group comparison of the oocyte and embryo quality. There was a statistically significant difference in the immature oocyte rate between the three groups (p = 0.005), with the Large group having the lowest immature oocyte rate and the Small group having the highest immature oocyte rate, and there was no difference in MII oocyte rate among the three groups. As for the 2PN fertilization rate, there was a statistical difference among the three groups, with the highest 2PN fertilization rate in the Large group and the lowest in the Small group. However, there was no statistically significant difference in the available embryo rate among the three groups. The GLM analysis suggested that different follicle sizes on the trigger day influenced the immature oocyte rate and the 2PN fertilization rate. But among the young women, applying different follicle sizes on the trigger day of the LPS regimen did not influence the oocyte and embryo quality, while in older women it affected the 2PN fertilization rate. Triggering when there is a high proportion of large follicles results in lower immature oocyte rate and higher 2PN fertilization rate when applying LPS protocol. However, age is a factor to be considered and the timing of triggering needs to be decided in clinical application with consideration of age and the actual situation.

Oocyte maturation defect in women undergoing IVF: contributing factors and effects on mature sibling oocyte outcomes.

This study aimed to identify demographic and clinical factors associated with low maturation rates and to investigate if the rate of immature oocytes impacts the outcomes of mature sibling oocytes. Women undergoing their first IVF-ICSI cycle between 2018 and 2022 at a fertility clinic were included. Cycles were classified into five groups according to the proportion of Metaphase II stage oocytes (MII): Null (0% MII, n = 46), Poor (1-25% MII, n = 44), Low (26-50% MII, n = 453), Acceptable (51-75% MII, n = 1641), and Optimal (76-100% MII, n = 2642). Demographic characteristics and clinical outcomes were compared between the five groups. In patients with a Null/Poor maturation rate, subsequent cycle outcomes were also evaluated. A total of 4826 cycles were included in the study; 69,909 oocytes were recovered, and 53,065 were MIIs (75.9%). The Null group was older, had lower levels of anti-Müllerian hormone (AMH), needed more gonadotropins and days of stimulation, had higher follicle stimulating hormone (FSH) levels on day 3, and had less follicles > 15mm on the day of trigger. When the outcomes of mature oocytes were compared, fertilization, usable blastocyst, aneuploidy, and life birth rates were comparable among groups. A binary logistic regression model using number of oocytes, paternal age, and trigger type with live birth rate endpoint found no differences between the categories and the base line Poor category. When patients whose maturation rate was Null/Poor, 42 (47.0%) carried out a second cycle; the maturation rate increased (56.9 ± 31.5 vs. 11.6 ± 11.2%, P < 0.0001). Our data suggest that poor responders are more likely to have low rates of oocyte maturation. The proportion of immature oocytes does not impact the outcomes of mature sibling oocytes. In patients with Null/Poor maturation in their first cycle, the subsequent cycle is often associated with improved maturation rates.

Enhancing oocyte competence in patients with polycystic ovarian syndrome: A microfluidic study on the effects of follicular fluid meiosis-activating sterol in in vitro oocyte maturation.

To evaluate the efficacy of a microfluidic culture system supplemented with follicular fluid meiosis-activating sterol (FF-MAS) on the maturation of immature oocytes in patients with polycystic ovarian syndrome (PCOS). A total of 438 germinal vesicle oocytes from 163 PCOS patients were included. Oocytes were divided into five groups: (1) cultured in static drops without FF-MAS, (2) cultured in static drops with FF-MAS, (3) cultured in a microfluidic device without FF-MAS, (4) cultured in a microfluidic device with FF-MAS for the first 2 h, and (5) cultured in a microfluidic device with FF-MAS for 24 h. Maturation, fertilization, and embryo development rates were assessed, alongside gene expression analysis using quantitative real-time PCR. Ultrastructural analysis was performed using transmission electron microscopy (TEM). The maturation rates for oocytes in groups 1 to 5 were 36%, 48%, 42%, 64%, and 65%, respectively, with groups 4 and 5 showing significant increases (p < 0.05). Fertilization rates were higher in groups 4 and 5 (p < 0.05). High-quality embryo formation rates were significantly higher in groups 3, 4, and 5. Gene expression analysis indicated enhanced expression of BRCA1 and TP53 in the dynamic culture groups. Ultrastructural analysis showed improved mitochondrial integrity and cytoplasmic maturation in the dynamic groups. The dynamic microfluidic culture system with FF-MAS supplementation significantly enhances oocyte maturation and embryo quality in PCOS patients by closely mimicking the natural in vivo environment. This approach shows promise for optimizing in vitro oocyte maturation and improving fertility outcomes.

Cryopreservation of ovarian cortex as fertility preservation in transgender men: A case report and literature review.

Transgender people have been experiencing significant advancements in their social visibility, although they continue to face frequent discrimination and exclusion. Among the issues encompassing transgender individuals' health care, the right to reproductive and sexual health have gained traction in the study landscape, necessitating an exploration of fertility preservation options for these patients. This report sheds light on the process of cryopreserving gametes and ovarian tissue after total hysterectomy with bilateral salpingo-oophorectomy for gender reassignment in a transgender man in hormonal masculinization. The procedure resulted in the cryopreservation of three immature oocytes and ovarian tissue fragments. The objective of this study is to contribute to the promotion of knowledge and the sharing of experiences, aiming to gradually enhance the attention on reproductive health among transgender individuals.

Role of Testosterone in Serum and Follicular Fluid in Sub Fertile Women Undergo Intracytoplasmic Sperm Injection

Background: Testosterone is the most common and regarded as the main hormone in the testicles among androgen (Tremblay, J. J. (2015) Testosterone synthesis is initiated by luteinizing hormone and cAMP signaling, promoting the transport of cholesterol to the inner mitochondrial membrane with the involvement of the steroidogenic acute regulatory protein (StAR). Cholesterol is then converted into pregnenolone, which further transforms into progesterone and is eventually metabolized into testosterone. This steroidogenesis pathway is similar in both thecae cells and interstitial cells of the ovaries (Cipolla-Neto, J., et al., 2022). Low-quality oocytes with a propensity for lower cleavage rates following fertilization have been associated with elevated androgen levels, more especially testosterone, in follicular fluid (FF) (Costa, L. O. B., et al., 2004). When FF contained immature oocytes as opposed to mature oocytes, testosterone levels were found to be noticeably higher. Pregnancy-associated follicles were found to have a higher estrogen-to-testosterone (E2/T) ratio reported. These results raise the possibility that an early follicular atresia linked to a low estrogen/androgen ratio in FF may reduce the viability of the oocytes and reduce the likelihood of fertilization and pregnancy purpose: to detect the follicular fluid and blood concentration of the testosterone hormone on subfertility in women undergoing Intracytoplasmic Sperm Injection (ICSI) for fertility treatment. Methods: This case-control study was carried out on eighty women, from these, forty females were sub-fertile patients defined as the cases, other forty females as a control included fertile females came to fertility center due to males factor of subfertility. A detailed medical history and physical examination were recorded. At the same day of ovum pickup blood and follicular fluid were collected, for hormonal analysis of testosterone. Results: Show significantly higher levels of serum and follicular testosterone in sub-fertile women (cases) than control group (p &lt; 0.05), and significant positive correlations between serum and follicular fluid. Additional materials: notes the number of references, tables, graphs, exhibits, test instruments, appendixes, or other supplemental materials in the paper. Also, the abstract must be written in a single paragraph in English, max 250 words. Conclusion: Testosterone hormone level in blood and follicular fluid were significantly higher in cases group than control and there is a positive correlations between serum and follicular fluid of testosterone.

190 Evaluation of the toxicity of different concentrations of dimethyl sulfoxide and ethylene glycol cryoprotectants on immature cattle oocytes selected by Brilliant cresyl blue stain and subsequent to polar body extrusion

190 Evaluation of the toxicity of different concentrations of dimethyl sulfoxide and ethylene glycol cryoprotectants on immature cattle oocytes selected by Brilliant cresyl blue stain and subsequent to polar body extrusion

Expression of Chitinase-like 3 Protein 1 (CHI3L1) in Buffalo Cumulus-oocyte Complexes during in vitro Maturation

Background: Buffaloes (Bubalus bubalis) are a vital livestock species in South Asia, significantly contributing to milk production and agricultural sustainability. However, reproductive inefficiency remains a significant challenge, adversely impacting productivity. In vitro maturation (IVM) of oocytes is a critical technique for improving reproductive efficiency in buffaloes. Chitinase-3-like Protein 1 (CHI3L1), known for its roles in tissue remodelling and inflammation, has emerged as a potential regulator in reproductive processes. This study investigates the expression of CHI3L1 in buffalo cumulus-oocyte complexes (COCs) during IVM to explore its role in oocyte maturation. Methods: Buffalo ovaries were collected post-slaughter and COCs were retrieved and cultured in TCM 199 medium supplemented with FSH, LH and estradiol. The COCs were classified into immature and mature groups based on morphological criteria after 24 hours of culture. Total RNA was extracted from both groups and CHI3L1 expression was quantified using quantitative real-time PCR (qRT-PCR). Western blot analysis was performed to assess CHI3L1 protein levels in both immature and mature oocytes. Result: The study found a significant upregulation of CHI3L1 expression in mature COCs, with a 3-fold increase compared to immature COCs (p less than 0.05). Western blot analysis confirmed the higher intensity of CHI3L1 protein in mature oocytes, correlating with the qRT-PCR results. These findings suggest that CHI3L1 plays a crucial role in the maturation of buffalo oocytes, potentially through its involvement in tissue remodelling and inflammatory responses within the COCs.

Anti-Centromere Protein A Antibody Disrupts the Competence of Mouse Oocytes Matured In Vitro.

Anticentromere autoantibodies are associated with refractory IVF/ET failure, but causality is unclear. Experimental models are needed. Immature oocytes collected from 23-day-old mice were matured in vitro for 18h in a culture medium containing an anti-human centromere protein A (CENP-A) polyclonal antibody, and those oocytes' maturity and chromosome/spindle structure were assessed. Antibody exposure did not affect the germinal vesicle breakdown ratio but reduced the first polar body formation ratio by 13% at the highest concentration (70.0 µg/mL). Metaphase II (MII) oocytes were stained for chromosomes/spindles and grouped into aligned/barrel-like (AB), scattered/weakly-stained (SW), and condensed/absent (CA). Antibody exposure decreased AB and increased SW and CA in a dose-dependent manner. The AB/SW/CA percentages were 86/14/0, 86/14/0, 35/65/0, and 0/0/100 in the 0, 17.5, 35.0, and 70.0 µg/mL antibody groups, respectively (underlined values represent p < 0.05 compared with 0 µg/mL). Next, metaphase II oocytes were subjected to intracytoplasmic sperm injection, and the number of pronucleus/pronuclei (PN) was counted 6 h later. Antibody exposure decreased two pronuclei oocytes and increased non-two pronuclei oocytes dose-dependently. The percentages of 0/1/2/3 pronuclei oocytes were 43/0/57/0, 37/0/21/42, 16/28/48/8, and 91/4/4/0 in the 0, 17.5, 35.0, and 70.0µg/mL groups, respectively. Anti-CENP-A antibody impaired a linear alignment of chromosomes at metaphase II and enhanced one or three PN formation after ICSI, which are similar to findings reported for infertile women with anticentromere autoantibodies.

In Vitro Maturation of Oocytes Obtained from Ovarian Cortex Among Postpubertal Hematological Cancer Patients Undergoing Fertility Preservation.

Purpose: In vitro maturation (IVM) of oocytes obtained from ovarian tissue during ovarian tissue cryopreservation (OTC) is a technique for fertility preservation in patients with cancer obviating the need to postpone chemotherapy initiation. Little is known about IVM outcomes in hematological malignancies, especially post-chemotherapy. The purpose of this study was to evaluate the effect of cytotoxic treatment on the potential to retrieve immature oocytes and mature them in vitro and examine the association between serum inflammatory markers and these results. Methods: In this retrospective study, we evaluated inflammation markers, including B symptoms and IVM outcomes of 78 chemotherapy-naive and exposed patients diagnosed with Hodgkin lymphoma (HL), non-Hodgkin lymphoma (NHL), acute lymphoblastic leukemia (ALL), or acute myeloid leukemia (AML). Results: The mean number of oocytes found was 7.2 ± 7.2. The average number of oocytes matured by IVM was 2.8 ± 3.5, and a mean IVM rate was 32.1 ± 27.7%. All patients in the ALL and AML groups had previous exposure to chemotherapy before OTC, compared with 50.0% (7/14) and 31.9% (15/47) in the NHL and HL groups, respectively. Among patients with lymphoma, chemotherapy exposure was associated with the reduced number of retrieved oocytes (9.8 ± 7.7 vs. 5.3 ± 5.7 oocytes, p = 0.049) in the HL group but not with the number of mature oocytes or IVM rate. B symptoms were not associated with IVM outcomes. Lymphocyte count (ß = 1.584; p = 0.038) and lactate dehydrogenase (ß = 0.009; p = 0.043) were the only significant parameters associated with the number of matured oocytes in a linear regression model. Conclusion: IVM is a promising assisted reproductive technology, which holds great potential for patients in need of urgent fertility preservation or those who cannot receive hormonal stimulation. Our results demonstrate the feasibility of the technique even in the presence of B symptoms and elevated inflammation markers and in patients with previous exposure to chemotherapy.

Prostaglandin F2A Disturbs Oogenesis by Causing Meiotic Spindle Damage

Abstract According to recent data, prostaglandin F2 alpha can have a negative influence on meiosis during oogenesis. Previously, we have found that this prostaglandin may accelerate in a dosage-dependent way the postovulatory aging in ovulated mature oocytes, compromising the integrity of their meiotic spindles. Aim. The study aimed to investigate the effects of prostaglandin F2α on the course and outcome of oocyte meiosis in a mouse model. Materials and Methods. Mouse oocytes were matured in vitro in the presence of prostaglandin F2α in a concentration of 100 ng/ml. Their meiotic stage, spindle morphology and chromosome arrangement were assessed by immunofluorescent labeling of tubulin and fluorescent staining of DNA. Results. We obtained a higher percentage of immature oocytes in metaphase I after the treatment than in untreated control oocytes. In addition, there were specific morphological changes in the meiotic spindles of oocytes exposed to prostaglandin F2α associated with a reduced number of fibers. Conclusion. It is probable that prostaglandin F2α has an impact on the microtubule dynamics of the meiotic spindle that can prevent the transition of maturing oocytes to the second meiotic division, most likely by triggering the spindle assembly checkpoint.

Znf706 regulates germ plasm assembly and primordial germ cell development in zebrafish

The cell fate of primordial germ cells (PGCs) in zebrafish is pre-determined by maternally deposited germ plasm, which is packaged into ribonucleoprotein complex in oocytes and inherited into PGCs-fated cells in embryos. However, the maternal factors regulating the assembly of germ plasm and PGC development remain poorly understood. In this study, we report that the maternal transcription factor Znf706 regulates the assembly of germ plasm factors into a granule-like structure localized perinuclearly in PGCs during migration. Maternal and zygotic mutants of znf706 (MZznf706) exhibit deficient germ plasm scattering at the early embryonic stage, decreased PGC numbers with some mislocation during PGC migration, and a lower female ratio in adulthood. Notably, the implementation of Znf706 CUT&Tag and RNA-seq on immature oocytes uncovers that Znf706 in stage I oocytes may promote transcription of several mitochondrial genes in addition to other functions. Hence, we propose that Znf706 is implicated in germ plasm assembly and PGC development in zebrafish.

PRP Influences Maturation and Fertilisation of Immature Mouse Oocytes.

In vitro maturation (IVM) of immature oocytes is a valuable method to enhance the rate of mature oocytes available for fertilisation. In the current study, platelet-rich plasma (PRP) was employed in IVM medium of immature oocytes. Harvested germinal vesicle stage oocytes with cumulus cells from female mature BALB/c mice divided into two groups of control and experiment. In the experimental group, GV oocytes matured in the IVM medium supplemented with 5% PRP, while in the control group, GV oocytes matured in the IVM medium without PRP. The percentage of GV, MI, MII and degenerated oocytes, zona pellucida thickness, perivitelline space size, diameter of mature oocytes, gene expression of apoptosis-related factors and subsequent development of matured oocytes were assessed. The PRP group displayed significantly improved outcomes in various parameters, including a higher proportion of MII and fertilised oocytes, cleavage and blastocyst embryos, compared to the control group. Moreover, the thickness of the zona pellucida was significantly lower in the PRP group than in the control group (p < 0.05). Furthermore, the PRP group demonstrated a significant decrease in the expression of transcripts associated with apoptosis (Bax and caspase-3); however, in the PRP group, a substantial increase in the expression of Bcl2l1, an apoptosis inhibitor, was observed when compared to the control group (p < 0.05). In conclusion, addition of PRP to the IVM culture media significantly increased oocyte maturation rate, leading to improved fertilisation and subsequent embryonic development. This enhancement highlights the positive influence of PRP on overall invitro maturation efficiency and early embryonic stages.

New Insights on In Vitro Maturation of Oocytes for Fertility Preservation

In the last decade, the evolution of oncofertility has sparked a resurgence of interest in in vitro maturation (IVM) due to its suitability in certain oncological scenarios where controlled ovarian hyperstimulation may not be feasible. The retrieval of immature cumulus–oocyte complexes from small antral follicles, regardless of the menstrual cycle phase, presents a swift opportunity to vitrify mature oocytes or embryos post-IVM in urgent situations or when stimulation is not advisable. Harvesting immature cumulus–oocyte complexes and immature oocytes can be achieved transvaginally or directly in the laboratory from extracorporeal ovarian tissue. Although IVM has transitioned from an experimental status due to safety validations, it relies on the intricate process of oocyte maturation. Despite successful live births resulting from IVM in fertility preservation contexts, the comparatively lower developmental competence of in vitro matured oocytes highlights the necessity to enhance IVM culture systems. Recent advancements in IVM systems hold promise in bolstering oocyte competence post-IVM, thereby narrowing the gap between IVM and outcomes from ovarian stimulation. Additionally, for optimizing the chances of conception in cancer survivors, the combination of IVM and ovarian tissue cryopreservation stands as the favored choice when ovarian stimulation is unfeasible.

Ovarian support cell in vitro maturation (OSC-IVM) results in healthy murine live births with no evidence of reprotoxicology in a multigenerational study

Research questionDoes application of human stem cell-derived ovarian support cells (OSCs) for in vitro maturation (IVM) have a safe reproductive toxicity profile? DesignA total of 70 B6/CBA 6–8-week-old stimulated female mice were used to collect immature oocytes (n=2,025) at the germinal vesicle stage. Oocytes were retrieved denuded (n=930) or with cumulus cells (COCs, n=1,095). These were randomly assigned to traditional IVM media (MediCult-IVMTM, Origio) or the same IVM media (MediCult-IVMTM, Origio) further supplemented with human OSCs (FertiloTM, Gameto Inc.). Metaphase II (M2) oocytes were inseminated by intracytoplasmic sperm injection and cultured to blastocyst stage. Vitrified blastocysts were transferred to pseudopregnant females, and F1 offspring were evaluated for various health and developmental metrics. Select F1 mice were outbred to produce the F2 generation, assessing live births, sex ratio, morphology, behavior, and health. ResultsIn mice OSC-IVM generally led to a decreased maturation rate compared to Traditional-IVM (p=0.0101). Subsequent embryo culture yielded similar cleavage rates between the four groups (p=0.7547). However, there was a significant distinction in the blastocyst formation with OSC-IVM showing higher blastocyst formation rates for both denuded oocytes (p=0.0408) and COCs (p=0.0063). Live births, sex ratio, development, and behavior in F1 and F2 generations revealed no adverse effects or significant differences, confirming the absence of long-term toxicity. ConclusionsThis study demonstrates the safety of the OSC-IVM system, with no adverse effects on embryo development or offspring health and fertility across generations.

Impact of heat stress on female reproduction in farm animals: challenges and possible remedies

Global warming poses significant challenges to agriculture, particularly within the livestock industry. Heat stress in animals refers to the physiological and metabolic distress experienced when environmental heat causes an animal’s body temperature to exceed its thermoneutral zone. Heat stress has a significant detrimental impact on reproductive outcomes, posing a direct threat to the profitability of livestock operations. Heat stress disrupts the normal hormonal balance essential for reproduction in female livestock, adversely affecting events of folliculogenesis and embryogenesis. As a result, the developmental competence of oocytes is compromised, which is crucial for successful fertilization and reproduction. Consequently, heat stress can lead to issues such as immature oocytes, atypical or sluggish embryonic development, silent estrus, and increased pregnancy loss causing a decline in reproductive rates and livestock owners face substantial financial losses. Despite using various cooling strategies, such as water sprinkler systems, cooling shades, and cooling devices, these measures are often insufficient to restore normal reproductive rates. Therefore, it is crucial to implement and explore novel strategies to enhance reproductive efficiency during periods of heat stress. Potential mitigation techniques include estrus synchronization, artificial insemination, hormonal alterations, embryo transfer, genetic engineering, sophisticated breeding strategies, the use of nutraceuticals, and changes in management practices. This review aims to consolidate recent findings on the adverse effects of heat stress on female reproduction, especially in tropical environments, and to evaluate potential strategies for improving summer fertility of livestock. Bangladesh Journal of Animal Science 53(3): 77-100, 2024

Robotic Manipulation of Cumulus–Oocyte Complexes for Cumulus Cell Removal

The removal of cumulus cells from cumulus–oocyte complexes is a critical step in clinical in vitro fertilization. Since the oocyte is partially occluded by the surrounding cumulus cells and individual cumulus cells are small in size, it is difficult for embryologists to assess the oocyte's maturity before cumulus cell removal and to completely remove all the cumulus cells manually . Furthermore, it is easy for the oocyte to become lost inside the micropipette during aspiration due to the inaccuracy of manual control. To deal with these difficulties, a robotic system was developed to completely remove cumulus cells from mature oocytes. In this study, an EPSANet50 network was developed to accurately assess the maturity of oocytes, avoiding the removal of cumulus cells around the immature oocyte. An adaptive controller was designed to accurately position oocytes at the target position, reducing the loss of oocytes inside the micropipette. An improved Yolov5s network was proposed to quantify the number and size of cumulus cells and assess the completeness of cumulus cell removal. The experimental results on mouse cumulus–oocyte complexes showed that the robotic system had a higher success rate (98.0 ± 1.8% vs. 85.3 ± 2.4%) and lower discard rate (4.1 ± 2.7% vs. 19.6 ± 3.5%) than the manual operation. Moreover, a higher amplification rate and lower non-specific rate were also achieved by the robotic system in the subsequent genetic testing procedure, indicating reduced genetic contamination from the cumulus cells.

Single-cell and bulk transcriptional profiling of mouse ovaries reveals novel genes and pathways associated with DNA damage response in oocytes

Immature oocytes enclosed in primordial follicles stored in female ovaries are under constant threat of DNA damage induced by endogenous and exogenous factors. Checkpoint kinase 2 (CHEK2) is a key mediator of the DNA damage response (DDR) in all cells. Genetic studies have shown that CHEK2 and its downstream targets, p53, and TAp63, regulate primordial follicle elimination in response to DNA damage. However, the mechanism leading to their demise is still poorly characterized. Single-cell and bulk RNA sequencing were used to determine the DDR in wild-type and Chek2-deficient ovaries. A low but oocyte-lethal dose of ionizing radiation induces ovarian DDR that is solely dependent on CHEK2. DNA damage activates multiple response pathways related to apoptosis, p53, interferon signaling, inflammation, cell adhesion, and intercellular communication. These pathways are differentially employed by different ovarian cell types, with oocytes disproportionately affected by radiation. Novel genes and pathways are induced by radiation specifically in oocytes, shedding light on their sensitivity to DNA damage, and implicating a coordinated response between oocytes and pregranulosa cells within the follicle. These findings provide a foundation for future studies on the specific mechanisms regulating oocyte survival in the context of aging, therapeutic and environmental genotoxic exposures.

Adolescent exposure to tris(1,3-dichloro-2-propyl) phosphate (TCPP) induces reproductive toxicity in zebrafish through hypothalamic-pituitary-gonadal axis disruption

Tris(1,3-dichloro-2-propyl) phosphate (TCPP), a prevalent organophosphorus flame retardant in aquatic environments, has raised significant concerns regarding its ecological risks. This study aims to explore the impacts of TCPP on the reproductive functions of zebrafish and delineate its gender-related toxic mechanisms. By assessing the effects on zebrafish of 10 mg/L TCPP exposure from 30 to 120 days post-fertilization (dpf), we thoroughly evaluated the reproductive capability and endocrine system alterations. Our findings indicated that TCPP exposure disrupted gender differentiation in zebrafish and markedly impaired their reproductive capacity, resulting in decreased egg laying and offspring development quality. Histological analyses of gonadal tissues showed an abnormal increase in immature oocytes in females and a reduction in mature sperm count and spermatogonial structure integrity in males, collectively leading to compromised embryo quality. Additionally, molecular docking results indicated that TCPP showed a strong affinity for estrogen receptors, and TCPP-treated zebrafish exhibited imbalanced sex hormones and increased estrogen receptor expression. Alterations in genes associated with the hypothalamic-pituitary-gonadal (HPG) axis and activation of the steroidogenesis pathway suggested that TCPP targets the HPG axis to regulate sex hormone homeostasis. Tamoxifen (TAM), as a competitive inhibitor of estrogen, exhibited a biphasic effect, as evidenced by the counteraction of TCPP-induced effects in both male and female zebrafish after TAM addition. Overall, our study underscored the gender-dependent reproductive toxicity of TCPP exposure in zebrafish, characterized by diminished reproductive capacity and hormonal disturbances, likely due to interference in the HPG axis and steroidogenesis pathways. These findings emphasize the critical need to consider gender differences in chemical risk assessments for ecosystems and highlight the importance of understanding the mechanisms underlying the effects of chemical pollutants on the reproductive health of aquatic species.

Human Umbilical Cord Mesenchymal Stem Cells Derived Exosomes Improved The Aged Mouse IVM Oocytes Quality.

During assisted reproductive technology (ART) treatment, the aged women, especially those over 35years old, have fewer mature oocytes and poorer quality of the oocytes comparing with the young women. In vitro maturation (IVM) technology facilitates the usage of immature oocytes, which is clinically important for the aged women. However, the maturation rate is low for the oocytes from the aged women. Human umbilical cord mesenchymal stem cells derived exosomes (HUCMSCs-exosomes), as important mediators of intercellular communication, have been widely used to restore ovarian function and improve female fertility. In this study, we isolated HUCMSCs-exosomes and collected the immature germinal vesicle oocytes from the naturally aged mouse model. And we added these HUCMSCs-exosomes to the conventional IVM culture system. The effects of HUCMSCs-exosomes on IVM oocytes were observed and analyzed from multiple aspects including maturation rate, spindle morphology, mitochondria function, and development potential. We found the quality of oocytes was improved by HUCMSCs-exosomes. Based on the results, we propose that HUCMSCs-exosomes may provide a novel and cell free strategy in the improvement of the IVM in elderly infertile women in the future.