Aged Oocytes Research Articles

Utility of Office Hysteroscopy in Diagnosing Retained Products of Conception After Early Pregnancy Loss After In Vitro Fertilization.

To evaluate the utility of office hysteroscopy in diagnosing and treating retained products of conception in patients with infertility who experience early pregnancy loss (EPL) after in vitro fertilization (IVF). We evaluated a retrospective cohort of 597 pregnancies that ended in EPL in patients aged 18-45 years who conceived through fresh or frozen embryo transfer at an academic fertility practice between January 2016 and December 2021. All patients underwent office hysteroscopy after expectant, medical, or surgical management of the EPL. The primary outcome was presence of retained products of conception at the time of office hysteroscopy. Secondary outcomes included incidence of vaginal bleeding, presence of intrauterine adhesions, treatment for retained products of conception, and duration of time from EPL diagnosis to resolution. Log-binomial regression and Poisson regression were performed, adjusting for potential confounders including oocyte age, patient age, body mass index, prior EPL count, number of prior dilation and curettage procedures, leiomyomas, uterine anomalies, and vaginal bleeding. Of the 597 EPLs included, 129 patients (21.6%) had retained products of conception diagnosed at the time of office hysteroscopy. The majority of individuals with EPL were managed surgically (n=427, 71.5%), in lieu of expectant management (n=140, 23.5%) or medical management (n=30, 5.0%). The presence of retained products of conception was significantly associated with vaginal bleeding (relative risk [RR] 1.72, 95% CI 1.34-2.21). Of the 41 patients with normal pelvic ultrasonogram results before office hysteroscopy, 10 (24.4%) had retained products of conception detected at the time of office hysteroscopy. When stratified by EPL management method, retained products of conception were significantly more likely to be present in individuals with EPL who were managed medically (adjusted RR 2.66, 95% CI 1.90-3.73) when compared with those managed surgically. Intrauterine adhesions were significantly less likely to be detected in individuals with EPL who underwent expectant management when compared with those managed surgically (RR 0.14, 95% CI 0.04-0.44). Of the 127 individuals with EPL who were diagnosed with retained products of conception at the time of office hysteroscopy, 30 (23.6%) had retained products of conception dislodged during the office hysteroscopy, 34 (26.8%) chose expectant or medical management, and 63 (49.6%) chose surgical management. The mean number of days from EPL diagnosis to resolution of pregnancy was significantly higher in patients who elected for expectant management (31 days; RR 1.18, 95% CI 1.02-1.37) or medical management (41 days; RR 1.54, 95% CI 1.25-1.90) when compared with surgical management (27 days). In patients with EPL after IVF, office hysteroscopy detected retained products of conception in 24.4% of those with normal pelvic ultrasonogram results. Due to the efficacy of office hysteroscopy in diagnosing and treating retained products of conception, these data support considering office hysteroscopy as an adjunct to ultrasonography in patients with infertility who experience EPL after IVF.

Evaluation of pre-implantation genetic testing for aneuploidy outcomes in patients without infertility undergoing in vitro fertilization compared to infertile controls.

To evaluate pre-implantation genetic testing for aneuploidy (PGT-A) outcomes in patients without infertility compared to infertile patients. We performed a retrospective cohort study of all patients without an infertility diagnosis ("fertile" patients) who utilized PGT-A at a large university-affiliated fertility center between 2016 and 2021. Fertile patients were 1-to-3 matched to infertile controls by age and number of oocytes retrieved. The primary outcome was blastocyst aneuploidy rate. Secondary outcomes included ovarian reserve markers, laboratory outcomes, and other PGT-A outcomes [rates of euploidy, mosaicism, and potentially transferrable (euploid + mosaic) embryos]. 283 fertile and 849 infertile patients were included. Median age, anti-Mullerian hormone, and day 2 estradiol levels were equivalent among groups; day 2 follicle-stimulating hormone levels were higher in fertile patients (6.9 vs. 6.5IU/mL, p < 0.01). The aneuploidy rate was similar among fertile and infertile patients (33.7% vs. 31.8%, p = 0.11); the euploidy rate was higher (50.8% vs. 47.0%, p < 0.01), and the mosaicism rate was lower in fertile patients (13.3% vs. 19.2%, p < 0.01). The rate of transferrable embryos was similar among groups (64.0% vs. 66.3%, p = 0.07), as was the percentage of patients yielding ≥ 1 euploid embryo (90.1% vs. 87.3%, p = 0.25). When controlling for significant covariates, multiple linear regression showed that aneuploidy rate was equivalent in both cohorts. Aneuploidy rate was similar in fertile and infertile patients. Fertile patients had slightly higher euploidy and lower mosaicism than infertile patients. Still, compared to fertile patients, infertile patients had equivalent rates of transferrable embryos and were just as likely to yield ≥ 1 euploid embryo.

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

Abstract 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 &amp;lt; 0.001; blastocyst formation rate: P &amp;lt; 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 &amp;lt; 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 INTEREST(S) This study was supported by grants from the National Key &amp; 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.

The association between the number of oocytes retrieved and cumulative live birth rate in different female age strata

To evaluate the association between the number of oocytes retrieved and cumulative live birth rate (CLBR) in different female age strata. 17,931 women undergoing their first IVF/ICSI-ET cycle in the Sir Run Run Shaw Hospital of Zhejiang University were grouped by age (A: ≤ 35 years; B: ≥ 36 years) as well as the number of oocytes retrieved (a: ≤ 5; b:6–9; c:10–14; d: ≥ 15). Multivariate regression analysis was performed to assess the OR of CLBR for the variable ‘age’ and ‘number of oocytes retrieved’. The group ≥ 36 years exhibited lower cumulative pregnancy rates (CPRs) and cumulative live birth rates (CLBRs), which are proportional to the number of oocytes retrieved but opposite to increasing age. Multivariate logistic regression analysis revealed that the age and number of oocytes retrieved remain significant independent predictive factors (P < 0.001). Age and number of oocytes retrieved are two independent factors affecting the CLBR. The discrepancy of the minimum number of oocytes retrieved for patients with different ages to achieve ideal CLBR is instructive for clinical practice. The practice of controlling the stimulation dose is feasible for patients ≤ 35 years who can achieve over 60% CLBR once the number of oocytes obtained is more than 6. However, additional stimulation cycles and accumulation of embryos are necessary for elderly group especially those ≥ 38 years old who need more than 14 oocytes to obtain higher live birth rate.

Melatonin ameliorates histone modification disorders in mammalian aged oocytes by neutralizing the alkylation of HDAC1

Aging-associated histone modification changes in oocytes have been sporadically reported, but the underlying mechanisms remain elusive. Here, we systematically characterize multiple histone modifications in oocytes during aging. We find that maternal and postovulatory aging markedly alter the status of histone modifications, specifically H4K12ac and H3K4me3, in both mouse and porcine oocytes. Meanwhile, we identify a substantial reduction in HDAC1 (histone deacetylase 1) protein in aged oocytes, which contributes to the changes in H4K12ac and H3K4me3. Moreover, by employing methylglyoxal (MG) and site-directed mutagenesis, we demonstrate that the elevated reactive carbonyl species (RCS) level induces HDAC1 degradation, likely through attacking the cysteine residues, thereby influences histone modification state. Importantly, supplementation of melatonin not only prevents the loss of HDAC1 protein, but also partially corrects the H4K12ac and H3K4me3 status in aged oocytes. To sum up, this study established the link between redox disequilibrium and histone modification alterations during mammalian oocyte aging.

Control of mitochondrial integrity influences oocyte quality during reproductive aging.

Reduced quality in oocytes from women of advanced maternal age (AMA) is associated with dysfunctional mitochondria. The objective of this study was to investigate the mechanisms controlling mitochondrial quality during maternal aging in mouse and human oocytes. We first evaluated the expression of proteins involved in the mitochondrial unfolded protein response (UPRmt) and mitophagy in in vivo matured metaphase II (MII) oocytes collected from young and aged mice. Expression of UPRmt proteins, HSPD1 and LONP1, and mitophagy proteins, total-PRKN and phosphorylated-PRKN, was significantly decreased in aged compared to young oocytes. Treatment of aged oocytes during in vitro maturation with the mitochondrially targeted antioxidant mitoquinone (MQ) specifically restored total-PRKN and phosphorylated-PRKN expression to levels seen in young oocytes. We next investigated whether maturing young oocytes under a high-oxygen environment would mimic the effects observed in oocytes from aged females. Phosphorylated-PRKN expression in oxidatively stressed young oocytes was reduced compared to that in oocytes matured under normal oxygen levels, and the mitochondrial DNA (mtDNA) copy number was increased. Treating oxidatively challenged young oocytes with MQ restored the phosphorylated-PRKN expression and mtDNA copy numbers. Treatment of oxidatively challenged oocytes with MQ also increased the co-localization of mitochondria and lysosomes, suggesting increased mitophagy. These data correlated with the developmental potential of the oocytes, as blastocyst development and hatching of oxidatively stressed oocytes were reduced, while treatment with MQ resulted in a significant increase in blastocyst development and hatching, and in the percentage of inner cell mass. Consistent with our results in mice, MII oocytes from women of AMA exhibited a significant decrease in phosphorylated-PKRN and total-PRKN compared to those of young women. Our findings suggest that the protein machinery to control the health of the mitochondria via UPRmt and mitophagy may be compromised in oocytes from aged females, which may result in inefficient clearance of dysfunctional mitochondria and reduced oocyte quality.

Epifriedelanol delays the aging of porcine oocytes matured invitro

Oocyte aging directly affects the subsequent embryonic development. Epifriedelanol is the active ingredient of Aster tataricus L.F. extract, and it possesses potential anti-cancer, anti-inflammatory and antioxidant properties. In addition, epifriedelanol can slow the aging of human skin fibroblasts. To explore the effect of epifriedelanol on the aging of porcine oocytes matured in vitro, the aging model was first established, epifriedelanol was added to in vitro maturation (IVM) medium to investigate its anti-aging effects by observing oocyte maturation and embryonic development potential, and analyzing aging-related gene expression, reactive oxygen species and mitochondrial membrane potential levels. It was found that typical aging of porcine oocytes appeared from 66 h during in vitro maturation. Compared with the 44 h group, a larger perivitelline space, increased abnormality of microtubulin formation, and significantly lower blastocyst rate were observed in the 66 h and 72 h groups. Compared with the 0 μg/mL group, the first polar body extrusion, cleavage and blastocyst rates were significantly improved (P < 0.05) in 10 μg/mL group. The expression of oocyte developmental potential-related, SIRT family-related, antioxidant and anti-apoptotic-related genes was significantly up-regulated (P < 0.05), p53 and pro-apoptotic genes were significantly down-regulated (P < 0.05). In addition, the reactive oxygen species level was significantly decreased (P < 0.01), the mitochondrial membrane potential was significantly elevated (P < 0.01) in 10 μg/mL group. In conclusion, epifriedelanol delays the aging of porcine oocytes cultured in vitro by up-regulating SIRT family gene expression, enhancing the antioxidant and anti-apoptotic capacity of oocytes.

MiR-425-5p suppression of Crebzf regulates oocyte aging via chromatin modification

Female infertility due to declining oocyte quality with age remains a significant challenge for patients and physicians, despite extensive research efforts. Recent studies suggest that microRNAs (miRNAs), which respond to various stressors in the aging process, may provide a promising solution. With the approval of small RNA drugs for clinical use, miRNA-based treatment of oocyte aging appears to be a viable option. Through high-throughput sequencing, miR-425-5p was identified as the only miRNA elevated under natural aging and oxidative stress. Microinjection of inhibitors to inhibit miR-425-5p effectively improved compromised phenotypes of old oocytes in vitro. Further investigation revealed that Crebzf acts as a mediator of miR-425-5p's age-related functions in old oocytes. In vivo treatment with miR-425-5p antagomirs significantly improved impaired oocyte development in reproductively old females by targeting Crebzf. Single-cell RNA sequencing revealed that Crebzf plays a vital role in regulating mRNAs targeting histone H3, trimethylated lysine 4 (H3K4me3), a crucial marker for transcriptional silencing. Overexpression of miR-425-5p could hinder oocyte maturation by downregulating Crebzf expression and disrupting transcriptional regulation. Our findings provide new insights into the potential of miR-425-5p antagomirs as a treatment for female infertility and highlight an elegant mechanism by which miR-425-5p inhibition of Crebzf inhibits a developmental switch in GV oocytes by regulating a group of histone methyltransferase mRNAs.

Luteolin regulates the distribution and function of organelles by controlling SIRT1 activity during postovulatory oocyte aging.

The quality of oocytes determines their development competence, which will be rapidly lost if the oocytes are not fertilized at the proper time after ovulation. SIRT1, one of the sirtuin family members, has been proven to protect the quality of oocytes during postovulatory oocyte aging. However, evidence of the effect of SIRT1 on the activity of organelles including the mitochondria, the endoplasmic reticulum (ER), the Golgi apparatus, and the lysosomes in postovulatory aging oocyte is lacking. In this study, we investigated the distribution and function of organelles in postovulatory aged oocytes and discovered abnormalities. Luteolin, which is a natural flavonoid contained in vegetables and fruits, is an activator of SIRT1. When the oocytes were treated with luteolin, the abnormal distribution of mitochondria, ER, and Golgi complex were restored during postovulatory oocyte aging. The ER stress protein GRP78 and the lysosome protein LAMP1 increased, while the mitochondrial membrane potential and the Golgi complex protein GOLPH3 decreased in aged oocytes, and these were restored by luteolin treatment. EX-527, an inhibitor of SIRT1, disrupted the luteolin-mediated normal distribution and function of mitochondria, ER, Golgi apparatus, and lysosomes. In conclusion, we demonstrate that luteolin regulates the distribution and function of mitochondria, ER, Golgi apparatus, and lysosomes during postovulatory oocyte aging by activating SIRT1.

Fisetin Delays Postovulatory Oocyte Aging by Regulating Oxidative Stress and Mitochondrial Function through Sirt1 Pathway.

The quality of oocytes determines the development potential of an embryo and is dependent on their timely fertilization after ovulation. Postovulatory oocyte aging is an inevitable factor during some assisted reproduction technology procedures, which results in poor fertilization rates and impairs embryo development. We found that fisetin, a bioactive flavonol contained in fruits and vegetables, delayed postovulatory oocyte aging in mice. Fisetin improved the development of aged oocytes after fertilization and inhibited the Sirt1 reduction in aged oocytes. Fisetin increased the GSH level and Sod2 transcription level to inhibit ROS accumulation in aged oocytes. Meanwhile, fisetin attenuated aging-induced spindle abnormalities, mitochondrial dysfunction, and apoptosis. At the molecular level, fisetin decreased aging-induced aberrant expression of H3K9me3. In addition, fisetin increased the expression levels of the mitochondrial transcription factor Tfam and the mitochondrial genes Co2 and Atp8 by upregulating Sirt1 in aged oocytes. Finally, inhibition of Sirt1 reversed the anti-aging effects of fisetin. Taken together, fisetin delayed postovulatory oocyte aging by upregulating Sirt1.

Melatonin restores the declining maturation quality and early embryonic development of oocytes in aged mice

With increase in women's age, the reproductive capability of female mammals decreases dramatically caused by age-related oxidative stress, coinciding with the decline in the ovarian reserve, and the quality and quantity of oocytes, which is the main determinant of female fertility. Melatonin, as an effective antioxidant and antiaging substance, is secreted by the pineal gland and been found in the follicular fluid as well, which has been turned out to enable to protect oocytes from oxidative stress during ovulation. However, the beneficial effects of melatonin on meiotic maturation in vitro and early embryo development of aged oocytes are still not fully understood. Thus, the aim of this study is to explore the potential mechanism of melatonin to improve the oocytes maturation and early embryonic development. The results suggested that oocyte quality decreased with age, whereas 10−6 M melatonin supplementation can significantly prompt the maturation quality of oocytes, the rate of fertilization and the formation rate of blastocyst. Mechanistic investigation indicated that melatonin supplementation not only restored the function of mitochondria by reducing reactive oxygen species (ROS) generation and early apoptosis, but also increased the level of ATP and total GSH through enhancing the mRNA expression levels of SIRT1, SIRT3, GPX4, SOD1 and SOD2. In conclusion, melatonin could alleviate the impairment of age-related oxidative stress to meiotic maturation and early embryonic development of oocytes. This study may provide a potential remediation strategy to improve the quality of oocytes from aged women and the efficiency of assisted reproductive technologies.

Aging-related aneuploidy is associated with mitochondrial imbalance and failure of spindle assembly

Despite aging is closely linked to increased aneuploidy in the oocytes, the mechanism of how aging affects aneuploidy remains largely elusive. Here, we applied single-cell parallel methylation and transcriptome sequencing (scM&T-seq) data from the aging mouse oocyte model to decode the genomic landscape of oocyte aging. We found a decline in oocyte quality in aging mice, as manifested by a significantly lower rate of first polar body exclusion (P < 0.05), and dramatically increasing aneuploidy rate (P < 0.01). Simultaneously, scM&T data suggested that a large number of differential expression genes (DEGs) and differential methylation regions (DMRs) were obtained. Next, we identified strong association of spindle assembly and mitochondrial transmembrane transport during oocyte aging. Moreover, we verified the DEGs related to spindle assembly (such as Naip1, Aspm, Racgap1, Zfp207) by real-time quantitative PCR (RT-qPCR) and checked the mitochondrial dysfunction by JC-1 staining. Pearson correlation analysis found that receptors for mitochondrial function were strongly positively correlated with abnormal spindle assembly (P < 0.05). In conclusion, these results suggested that the mitochondrial dysfunction and abnormal spindle assembly of aging oocytes ultimately may lead to increased oocyte aneuploidy.

Promising anti-ovarian aging herbal formulation He's Yangchao promotes in vitro maturation of oocytes from advanced maternal age mice

Ethnopharmacological relevanceMarveled at the discovery of artemisinin, the world's expectations for traditional Chinese medicine are rising. He's Yangchao formula (HSYC) is a traditional Chinese herbal formula with the effects of tonifying kidney and essence, and reconciling yin and yang. It has been clinically proven to have anti-ovarian aging effects. Age is the primary cause of diminished ovarian reserve and assisted reproductive failure in women, whether HSYC has the potential to improve in vitro maturation of oocytes from advanced maternal age (AMA) mice has yet to be determined. Aim of the studyThis study aims to evaluate the efficacy and possible mechanism of HSYC in promoting in vitro maturation of oocytes from AMA mice. Materials and methodsThe GV oocytes were obtained from young and aged mice. The GV oocytes from young mice were cultured in drops of M16 medium, and the GV oocytes from AMA mice were randomly divided four groups: Vehicle group (cultured in 90% M16 medium +10% blank serum), Low HSYC group (cultured in 90% M16 medium + 10% Low HSYC-medicated serum), High-HSYC group (cultured in 90% M16 medium +10% High HSYC-medicated serum), and Quercetin group (cultured in M16 medium supplemented with 10 μM quercetin). The rates of first polar body extrusion, reactive oxygen species (ROS), intracellular calcium, and mitochondrial membrane potential levels in each groups were observed. In addition, expression levels of mitochondrial function, autophagy, DNA damage, and antioxidant-related proteins were assessed. ResultsSupplementation of HSYC in vitro alleviated age-associated meiotic progression defects in maternally aged oocytes. Importantly, HSYC supplementation eliminated the age-related ROS accumulation to suppress DNA damage and autophagy during the in vitro maturation of maternally aged oocytes. Meanwhile, the mitochondrial function was improved after HSYC treatment, as manifested by higher mitochondrial membrane potential and lower Ca2+ levels. Furthermore, we found that HSYC supplementation during in vitro maturation of maternally aged oocytes upregulated the expression level of SIRT3, a crucial protein in regulating mitochondrial function. Consistently, the expression levels of the SOD2, PCG1α, and TFAM were increased, while the SOD2 acetylation level was decreased, which further proved its antioxidant function. ConclusionsHSYC supplementation promotes in vitro maturation of oocytes from AMA mice mainly via improving mitochondrial function and alleviating oxidative stress. The mechanism may be related to the regulation of SIRT3-dependent deacetylation of the SOD2 pathway.

Multi-Omics Analysis Reveals Translational Landscapes and Regulations in Mouse and Human Oocyte Aging.

Abnormal resumption of meiosis and decreased oocyte quality are hallmarks of maternal aging. Transcriptional silencing makes translational control an urgent task during meiosis resumption in maternal aging. However, insights into aging-related translational characteristics and underlying mechanisms are limited. Here, using multi-omics analysis of oocytes, it is found that translatomics during aging is related to changes in the proteome and reveals decreased translational efficiency with aging phenotypes in mouse oocytes. Translational efficiency decrease is associated with the N6-methyladenosine (m6A) modification of transcripts. It is further clarified that m6A reader YTHDF3 is significantly decreased in aged oocytes, inhibiting oocyte meiotic maturation. YTHDF3 intervention perturbs the translatome of oocytes and suppress the translational efficiency of aging-associated maternal factors, such as Hells, to affect the oocyte maturation. Moreover, the translational landscape is profiled in human oocyte aging, and the similar translational changes of epigenetic modifications regulators between human and mice oocyte aging are observed. In particular, due to the translational silence of YTHDF3 in human oocytes, translation activity is not associated with m6A modification, but alternative splicing factor SRSF6. Together, the findings profile the specific translational landscapes during oocyte aging in mice and humans, and uncover non-conservative regulators on translation control in meiosis resumption and maternal aging.

Growth hormone reduces aneuploidy and improves oocytes quality by JAK2-MAPK3/1 pathway in aged mice

BackgroundThe global delay in women’s reproductive age has raised concerns about age-related infertility. The decline in oocyte quality is a limiting factor of female fertility, yet there are currently no strategies to preserve oocyte quality in aged women. Here, we investigated the effects of growth hormone (GH) supplementation on aneuploidy of aged oocytes.MethodsFor the in vivo experiments, the aged mice (8-month-old) were intraperitoneally injected with GH daily for 8 weeks. For the in vitro experiments, germinal vesicle oocytes from aged mice were treated with GH during oocyte maturation. The impacts of GH on ovarian reserve before superovulation was evaluated. Oocytes were retrieved to assess oocyte quality, aneuploidy and developmental potential characteristics. Quantitative proteomics analysis was applied to investigate the potential targets of GH in aged oocytes.ResultsIn this study, we demonstrated that GH supplementation in vivo not only alleviated the decline in oocyte number caused by aging, but also improved the quality and developmental potential of aged oocytes. Strikingly, we discovered that GH supplementation reduced aneuploidy in aged oocytes. Mechanically, in addition to improving mitochondrial function, our proteomic analysis indicated that the MAPK3/1 pathway may be involved in the reduction in aneuploidy of aged oocytes, as confirmed both in vivo and in vitro. In addition, JAK2 may also act as a mediator in how GH regulates MAPK3/1.ConclusionsIn conclusion, our research reveals that GH supplementation protects oocytes against aging-related aneuploidy and enhances the quality of aged oocytes, which has clinical significance for aged women undergoing assisted reproduction technology.Graphical

O-206 Growth hormone supplementation reduces aneuploidy and improves the quality of aged oocytes by activating the JAK2-ERK1/2 pathway

Abstract Study question Does growth hormone (GH) supplementation improves the quality of oocytes in aged mice via reducing aneuploidy? Summary answer Daily injections of GH for 8 weeks reduces aneuploidy by activating JAK2-ERK1/2 pathway and improves the quality of oocyte in aged mice. What is known already Aging-induced low-quality oocytes, which are mainly caused by aneuploidy, decrease female fertility. However, few treatments are available to improve the poor quality of oocytes in aged women. Age-related decline in GH levels may be related to the insufficient reproductive potential in aged women. Several studies have reported that GH improves assisted reproductive technology outcomes in poor ovarian response patients, although its role is still controversial. In animal models, GH supplementation improved the oocyte quality by enhancing mitochondrial function. However, whether GH reduces the aneuploidy rate in aged oocytes, and if so the underlying mechanisms, are unclear. Study design, size, duration The aged (8-month-old) C57BL/6J female mice were used in the study. For the in vivo experiments, the aged mice were intraperitoneally injected with GH (GenSci, Changchun, China; 1.6 mg/kg body weight) or an equivalent volume of normal saline for 8 weeks; For the in vitro experiments, germinal vesicle (GV) oocytes from aged mice were treated with GH (200 ng/mL) or PBS in M16 medium. Participants/materials, setting, methods We counted oocytes by immunohistochemistry to assess ovarian reserve. The expression of growth hormone receptor and mitochondrial genes were measured by quantitative real-time PCR. Time–lapse incubator was utilized to record the developmental potential of early embryos. Immunofluorescence was performed to assess parameters of oocyte quality (mitochondrial functions, spindle/chromosome defects, and DNA damage). Chromosome spread and DNA sequencing were used to analyze aneuploidy rate. We performed quantitative proteomics analysis to identify the potential targets of GH. Main results and the role of chance Firstly, we found that GH promoted GHR expression in aged oocytes (30.64±1.70 vs 21.68±1.08, P &amp;lt; 0.001). GH ameliorated aging-induced decline in ovarian reserve of aged mice, with increased ovarian index (0.040%±0.0031 vs 0.023%±0.0019, P &amp;lt; 0.01), and number of antral follicles (8.67±1.2 vs 3.33±0.88, P &amp;lt; 0.05) compared to the controls. In addition, GH improved the quality of aged oocytes, representing restored mitochondrial functions (P &amp;lt; 0.05), decreased DNA damage (23.37±1.76 vs 38.35±2.52, P &amp;lt; 0.001) . As expected, GH promoted the fertilization rate (39.9%±1.10 vs 14.4%±0.62, P &amp;lt; 0.001) and early embryo development (P &amp;lt; 0.01) of aged oocytes. Of note, GH recovered the spindle/chromosome defects (42.6%±2.12 vs 63.9%±1.83, P &amp;lt; 0.01, and reduced aneuploidy rate (20.9%±1.61 vs 39.3%±1.84, P &amp;lt; 0.01) in aged oocytes. GH activated ERK1/2 expression to decrease aneuploidy rate in aged oocytes. In addition, JAK2 might be involved in the regulation of GH on ERK1/2 expression in aged oocytes. Altogether, GH restored age-related decline in oocyte number, and decreased the occurrence rate of aneuploidy by activating JAK2-ERK1/2 pathway, thereby improving the quality of aged oocytes. Limitations, reasons for caution The molecular action mechanism of GH to regulate aneuploidy remains to be determined. Besides, future work should be extended to human oocyte to determine whether this mechanism is conserved between mice and humans Wider implications of the findings Our work expounds a theoretical basis for application of GH to improve the fertility of aged women. Besides, the results also feed new ideas for the prevention and treatment of oocyte quality decline in assisted reproductive technology. Trial registration number not applicable

P-791 Supplementation of mitochondria from endometrial mesenchymal stem cells (EnMSCs) improves oocyte quality in aged mice

Abstract Study question To improve the quality of aging oocytes by supplementing the mitochondria from endometrial mesenchymal stem cells (EnMSCs) via microinjection. Summary answer Supplementation with EnMSC-derived mitochondria can improve the quality of oocytes and promote embryo development in aging mice. What is known already Maternal aging is one of the major causes of reduced ovarian reserve and low oocyte quality in elderly women. Decreased oocyte quality is the main cause of age-related infertility. Mitochondria are multifunctional energy stations that determine the oocyte quality. The mitochondria in aging oocytes display functional impairments with mtDNA damage, which leads to reduced competence and developmental potential of oocytes. To improve oocyte quality, mitochondrial supplementation is carried out as a potential therapeutic approach. However, the selection of suitable cells as the source of mitochondria remains controversial. Study design, size, duration Study design: Eight-month-old ICR mice were purchased and then were further raised to ten months of age for use in the experiments. EnMSCs were stably cultivated and oocytes were extracted from ten-month-old ICR mice. Study size: About 300 GV oocytes and 134 MII oocytes were counted . 80 fertilized oocytes were transplanted into the fallopian tubes of surrogate mothers. Study duraton: About one half a year. Participants/materials, setting, methods Participants:ten-month-old ICR mice. Setting and methods: We cultivated EnMSCs from aged mice and extracted mitochondria from EnMSCs. And then GV oocytes were supplemented with mitochondria via microinjection. To further assess the effects of mitochondrial supplementation on embryo development, MII oocytes from aging mice were fertilized by ICSI combining EnMSCs mitochondria microinjection.To test whether mitochondrial supplementation could improve in vivo embryo development in aging mice, embryo transplantation was performed after mitochondrial microinjection combined with ICSI. Main results and the role of chance In this study, we found that the mitochondria derived from EnMSCs could significantly improve the quality of aging oocytes.Supplementation with EnMSC mitochondria significantly increased the blastocyst ratio of MII oocytes from aging mice after intracytoplasmic sperm injection (ICSI). We also found that the birth rate of mitochondria-injected aging oocytes was significantly increased after embryo transplantation. Limitations, reasons for caution Microinjection into oocytes was invasive and some oocytes died after microinjection. The ability of this method to improve oocyte quality was limited because the MII rate of oocytes after mitochondria supplementation was 35.65±1.75% while the rate of oocytes from 6–8 weeks of mice in a previous study was nearly 80%. Wider implications of the findings For clinical treatment, the EnMSCs can be obtained from the patient's own menstrual blood as the sources of mitochondrial supplementation. A noninvasive procedure for cell extraction ensures safety, and the findings of this work might provide a valuable strategy for patients with declining oocyte quality due to aging. Trial registration number Not applicable

P-636 Euploid blastocysts obtained in advanced maternal age women either in donor or autologous egg cycles showed similar live birth rates: a retrospective cohort study

Abstract Study question Do euploid blastocysts have the same implantation and live birth rate in autologous and donor egg cycles in patients with advanced maternal age (AMA)? Summary answer Euploid blastocysts transferred in AMA patients showed that implantation and live birth rates (LBR) were not significantly different between reproductively younger and older women. What is known already Aneuploidy rates increase steadily with age, reaching &amp;gt;80% in women &amp;gt;42 years old. The high prevalence of aneuploidies may provide an explanation for the low LBR achieved by assisted reproductive treatments, especially for women with AMA. Similarly, among oocyte donors, age is critical to achieving reproductive success independently of the recipients’ age. In donor cycles, recipients using oocytes from older donors had a statistically significantly lower LBR than younger donors. Currently, it still remains unclear whether the LBR differs between young donors versus old infertile patients in the context of euploid blastocyst transfers. Study design, size, duration Multicenter retrospective study with the preimplantation genetic test for aneuploidies (PGT-A) in both autologous and donor oocytes cycles between January 2018 and January 2021 was conducted at three different institutions (IVI-RMA, GeneraLife and Humanitas Fertility Center). 556 patients were included and matched 1:1 for the age of recipients, euploid blastocyst morphological quality, and day of development. In the frozen embryo transfers, endometrial preparation was done using hormonal replacement therapy (HRT) or natural cycles (NC). Participants/materials, setting, methods AMA patients older than 39 years underwent IVF. 556 patients who underwent autologous or donor oocyte cycles with at least one euploid blastocyst transferred were matched for oocyte age, blastocyst morphological quality and day of development. Blastocyst ploidy was assessed by aCGH or NGS analysis. Patients with uterine pathologies reported in the dataset were excluded. The primary outcome was LBR per first euploid blastocyst transfer. Outcomes were adjusted for confounders via logistic regression analyses. Main results and the role of chance 278 single euploid blastocysts tested by PGT-A were transferred in both autologous and donor oocyte cycles. No statistical differences were reported in the maternal age at transfer in the two groups (41.91 ± 2.03 and 41.75 ± 1.98, p = 0.55), endometrial thickness [(8.67, 95%CI: 8.45-8.88) and (8.36, 95%CI: 8.05-8.67)] and type of endometrial preparation (HRT or NC). Implantation rate and pregnancy rate were similar without statical difference in autologous or donor cycles, respectively [57.91%, (95%CI: 51.87-63.78) versus 56.83%, (95%CI: 50.79-62.74); p = 0.99)] and N = 158/278 (56.83%, 95%CI: 50.79-62.74) versus N = 161/278 (57.91%, 95%CI: 51.87-63.78), p = 0.86. The rate of ectopic pregnancy was not statistically different in both arms. Even the miscarriage rate was comparable in autologous N = 46/158, 16.55% (95%CI:12.38-21.45) versus donor group N = 45/161, 16.19%, (95%CI:12.06-21.05); p = 0.98. The LBR was not statistically different in autologous N = 111/278 (39.93%, 95%CI:34.13-45.95) and donor group 114/278 (41.01%, 95%CI:35.17-47.04), p = 0.86. No statistical difference was reported for gestational age and birthweight at delivery. The logistic regression analyzed as putative confounders: oocyte age, number of inseminated MII-oocytes, zygotes and euploid blastocysts, sperm parameters, endometrial preparation and endometrial thickness showed no statistical differences. Limitations, reasons for caution Despite the matched design, the retrospective nature is the major limitation of this study. Also, patients received PGT-A using NGS and CGH arrays technology. The aCGH analysis is less sensitive than more recent NGS technology. Lastly, the prevalence of adenomyosis could be underestimated due to retrospective data collection. Wider implications of the findings The dramatic decline in IVF treatment success is primarily caused by aneuploidies. Euploid blastocysts show LBR largely independent of oocyte age. Our study suggests no uterine effect of aging. The reproductive decline in women is mainly related to meiotic failure, therefore future insights should develop strategies to overcome chromosomal aneuploidies. Trial registration number NA

O-059 ntermetabolites of cholesterol synthesis in granulosa cell and oocyte aging

Abstract With the development of society, there has been a significant delay in female fertility and an increasing desire for childbearing among females of advanced maternal age in recent decades. Nevertheless, women over 35 years old have more risk of infertility and miscarriage compared to young women, due to decreased oocyte quality. These changes are mainly caused by meiotic defects, especially aneuploidy. Therefore, exploring the mechanisms of aging-related meiotic defects and aneuploidy in oocytes is of great significant for improving the pregnancy outcomes of aged women. Oocytes are enveloped by granulosa cells (GCs) to form follicles, which constitute the reproductive units in ovaries. GCs form a metabolic community with oocytes for oocyte growth and maturation. Numerous studies have reported that metabolic coupling between GCs and oocytes could affect the process of oocyte meiosis resumption, spindle assembly, and chromatin arrangement. Maternal aging is related to disruption of oocyte-GC interactions that leads to disorders of the oocyte meiotic process. Therefore, abnormal metabolic coupling between GCs and oocytes is an important reason for decreased quality in aged oocytes. To further clarify the metabolic coupling changes between GCs and oocytes during ovarian aging, we systematically characterized the dynamic changes in the overall transcriptomic landscapes of oocytes and GCs from young and aged mice throughout oocyte meiosis (during the growing oocyte [GO], full-grown oocyte [FGO], metaphase I [MI] oocyte [MIO], and metaphase II [MII] oocyte [MIIO] phases). Data revealed that the mevalonate (MVA) pathway, an essential metabolic pathway that uses acetyl-CoA to produce cholesterol and isoprenoids, was specifically highly expressed in GCs with the resumption of oocyte meiosis, and the expression of MVA pathway in GCs decreased with age. Atorvastatin-mediated inhibition of MVA metabolism in GCs decreased the first polar body extrusion (PBE) rate, and increased oocyte meiotic defects and aneuploidy in young cumulus-oocyte complexes (COCs). Further studies showed that the effect of the MVA pathway on oocyte meiosis was mainly regulated by protein isoprenoylation mediated by the inter-metabolites of cholesterol synthesis. Importantly, upregulation of the MVA pathway in aged GCs could ameliorate the depletion of ovarian reserve, decrease oocyte meiotic defects, and improve oocyte quality. In conclusion, metabolite in granulosa cell is an important factor determining the oocyte quality; MVA pathway in GCs is a critical regulator of meiotic maturation and euploidy in oocytes, and age-associated MVA pathway abnormalities contribute to oocyte meiotic defects and aneuploidy; Supplementation of inter-metabolites of cholesterol synthesis is a new therapeutic target for clinical intervention to improve oocyte quality. Trial registration number XXXX

O-208 Maternal spindle transfer restores the developmental competence of in vitro aged oocytes with diminished metabolic activity identified by hyperspectral imaging

Abstract Study question Can the developmental competence of poor-quality oocytes identified by hyperspectral imaging be restored by maternal spindle transfer? Summary answer Maternal spindle transfer can be efficiently assisted by hyperspectral imaging to identify and subsequently repair the developmental competence of poor-quality oocytes. What is known already Oocyte cytoplasmic dysfunctions (including, but not limited to the mitochondria) are a major cause for poor embryonic development. Given that maternal spindle transfer (MST) allows the replacement of the entire cytoplasm of an oocyte, it is a promising method for the enhancement of oocyte developmental competence. However, accurate diagnostic tools for oocyte quality assessment are lacking. Recently, a novel hyperspectral imaging approach has been shown to classify oocytes non-invasively based on their metabolic profile, which could assist in the evaluation of oocyte developmental potential prior to MST. Study design, size, duration Oocytes were obtained from young female mice and analyzed blindly by hyperspectral imaging either immediately after collection (fresh group, n = 122), after overnight culture (in-vitro aged group, n = 58) or on a group of MST oocytes (n = 36), where the meiotic spindle from in-vitro aged oocytes was transferred into enucleated fresh oocytes. Oocytes were inseminated by ICSI and cultured in-vitro individually. Correlations between the oocyte’s metabolic profiles and rate of development to the blastocyst stage were performed. Participants/materials, setting, methods Oocytes were collected from 6-10 weeks-old superovulated B6CBAF1 females. MST was performed in manipulation medium supplemented with cytochalasin B. After enucleation, karyoplasts were exposed to an inactivated Sendai virus solution to promote membrane fusion. ICSI was performed using a Piezo-Drill actuator. Hyperspectral imaging was done in the near-infra-red regime to avoid photoxicity. Image processing was done using dimensionality reduction, which was then feed to an Akaike Information Criterion algorithm to classify the different metabolic subpopulations. Main results and the role of chance Fresh oocytes showed metabolic profiles that significantly differed from the profiles of in-vitro aged oocytes, which were characterized by a pronounced diminished metabolic activity. While in the fresh group, 67% of the oocytes developed into expanded blastocysts, none of the in-vitro aged oocytes with low metabolism produced a blastocyst (p &amp;lt; 1x10-05). Interestingly, MST oocytes that were imaged 1h after nuclear reconstruction, showed a restored metabolic spectrum that resembled the profile from fresh non-manipulated controls. This restoration of the metabolic activity was concomitantly associated with an enhancement of developmental competence, with expanded blastocyst rates (63%) comparable to those of fresh non-manipulated controls (p = 0.668). Blastocyst developmental rates of fresh oocytes after hyperspectral imaging were comparable to oocytes from the same cohort that were not imaged and used as controls. Limitations, reasons for caution The metabolic profiles obtained from these experimental groups need to be correlated with implantation rates in mouse. The impaired embryo developmental pattern characteristic of poor-quality oocytes may not be exclusively attributed to metabolic deficiencies, so caution must be exerted when generalizing these results to oocytes with other cytoplasmic dysfunctions. Wider implications of the findings MST experiments confirm that the developmental decline of these oocytes is primarily attributed to abnormal function of cytoplasmic factors involved in metabolism, while the nuclear genome remains developmentally competent. MST can benefit from hyperspectral imaging to identify oocytes with impaired developmental potential within a cohort. Trial registration number not applicable