Time-lapse microscopy for embryos is a non-invasive technology used to characterize early embryo development. This study employs time-lapse microscopy and machine learning to elucidate changes in embryonic growth kinetics with maternal aging. We analyzed morphokinetic parameters of embryos from young and aged C57BL6/NJ mice via continuous imaging. Our findings show that aged embryos accelerated through cleavage stages (from 5-cells) to morula compared to younger counterparts, with no significant differences observed in later stages of blastulation. Unsupervised machine learning identified two distinct clusters comprising of embryos from aged or young donors. Moreover, in supervised learning, the extreme gradient boosting algorithm successfully predicted the age-related phenotype with 0.78 accuracy, 0.81 precision, and 0.83 recall following hyperparameter tuning. These results highlight two main scientific insights: maternal aging affects embryonic development pace, and artificial intelligence can differentiate between embryos from aged and young maternal mice by a non-invasive approach. Thus, machine learning can be used to identify morphokinetics phenotypes for further studies. This study has potential for future applications in selecting human embryos for embryo transfer, without or in complement with preimplantation genetic testing.

Age-related aneuploidy in human oocytes is a major factor contributing to decreased fertility and adverse reproductive outcomes. As females age, their oocytes are more prone to meiotic chromosome segregation errors, leading primarily to aneuploidy. Elevated aneuploidy rates have also been observed in oocytes from very young, prepubertal conceptions. A key barrier to developing effective treatments for age-related oocyte aneuploidy is our incomplete understanding of the molecular mechanisms involved. The challenge is becoming increasingly critical as more people choose to delay childbearing, a trend that has significant societal implications. In this review, we summarize current knowledge regarding the process of oocyte meiosis and folliculogenesis, highlighting the relationship between age and chromosomal aberrations in oocytes and embryos, and integrate proposed mechanisms of age-related meiotic disturbances across structural, protein, and genomic levels. Our goal is to spur new research directions and therapeutic avenues.

Age-related changes in the mitochondrial status of human cumulus cells (hCCs) impact oocyte quality; however, the relationship between hCC mitochondrial (dys)function and reproductive aging remains poorly understood. This study aimed to establish the interplay between hCC mitochondrial dysfunction and women\'s reproductive potential. In this investigation, 266 women were enrolled and categorized into two groups based on their age: a young group (<35 years old) and an advanced maternal age (AMA) group (≥35 years old). Comprehensive analysis of reproductive outcomes was conducted in our population. Various mitochondrial-related parameters were analyzed across distinct subsets. Specifically, mitochondrial membrane potential (∆Ψm) and mitochondrial mass were examined in 53 samples, mtDNA content in 25 samples, protein levels in 23 samples, bioenergetic profiles using an XF24 Extracellular Flux Analyzer in 6 samples, and levels of reactive oxygen species (ROS) and adenosine triphosphate (ATP) in 39 and 43 samples, respectively. In our study, the reproductive potential of AMA women sharply decreased, as expected. Additionally, an impairment in the mitochondrial function of hCCs in older women was observed; however, no differences were found in terms of mitochondrial content. Regarding oxidative phosphorylation, metabolic profiling of hCCs from AMA women indicated a decrease in respiratory capacity, which was correlated with an age-dependent decrease in the ATP synthase (ATP5A1) protein level. However, intracellular ROS and ATP levels did not differ between groups. In conclusion, our study indicates that age-related dysfunction in hCCs is associated with impaired mitochondrial function, and, although further studies are required, ATP synthase could be relevant in this impairment.

Aneuploid human eggs (oocytes) are a major cause of infertility, miscarriage, and chromosomal disorders. Such aneuploidies increase greatly as women age, with defective linkages between sister chromatids (cohesion) in meiosis as a common cause. We found that loss of a specific pool of the cohesin protector protein, shugoshin 2 (SGO2), may contribute to this phenomenon. Our data indicate that SGO2 preserves sister chromatid cohesion in meiosis by protecting a \"cohesin bridge\" between sister chromatids. In human oocytes, SGO2 localizes to both sub-centromere cups and the pericentromeric bridge, which spans the sister chromatid junction. SGO2 normally colocalizes with cohesin; however, in meiosis II oocytes from older women, SGO2 is frequently lost from the pericentromeric bridge and sister chromatid cohesion is weakened. MPS1 and BUB1 kinase activities maintain SGO2 at sub-centromeres and the pericentromeric bridge. Removal of SGO2 throughout meiosis I by MPS1 inhibition reduces cohesion protection, increasing the incidence of single chromatids at meiosis II. Therefore, SGO2 deficiency in human oocytes can exacerbate the effects of maternal age by rendering residual cohesin at pericentromeres vulnerable to loss in anaphase I. Our data show that impaired SGO2 localization weakens cohesion integrity and may contribute to the increased incidence of aneuploidy observed in human oocytes with advanced maternal age.

Vitrification is an important assisted reproductive technology, although it induces mitochondrial dysfunction in embryos. Herein, we aimed to investigate whether age-associated accumulation of advanced glycation end-products (AGEs) in oocytes impairs the recovery of embryos from cryopreservation-induced mitochondrial dysfunction/damage. Mouse eight-cell stage embryos developed in vitro were vitrified and warmed and incubated up to the blastocyst stage. AGE levels in oocytes were higher in both aged mice and AGE accumulation mouse models (MGO-mice) than those in young and control mice. In addition, the level of SIRT1 upregulation was lower for embryos of aged and MGO-mice than that for embryos of young and control mice. The highest mitochondrial DNA (mtDNA) content was detected in blastocysts derived from vitrified embryos of aged and MGO-mice. The spent culture medium of blastocysts derived from both aged and MGO-mice contained higher mtDNA content than that of the blastocysts derived from young and control mice. EX527 increased mtDNA content in the spent culture medium of vitrified embryos derived from young mice. In addition, p62 aggregate levels were higher in vitrified embryos of control mice than those in vitrified embryos of MGO-mice. The SIRT1 activator, resveratrol, increased p62 aggregation levels in vitrified embryos derived from young and aged mice, whereas vitrification did not affect p62 aggregation levels in embryos from aged mice. Therefore, age-associated AGE accumulation induces decreased responsive SIRT1 upregulation following vitrified-warmed treatment and impairs mitochondrial quality control activity in vitrified embryos.

Maternal aging has been reported to reduce oocyte quality and, in turn, lower the developmental potential of the resulting embryos. Here, we show that maternally aged oocytes display two strikingly different phenotypes: some have normal morphology, whereas others have significantly shrunk cytoplasm. The latter phenotype usually prevails in aged females. Our objective was to characterize both types of maternally aged oocytes and investigate the origins of this diversity. Importantly, our experiments indicate that shrunk maternally aged oocytes are severely compromised in terms of mitochondrial functionality as compared to their young or morphologically normal maternally aged counterparts: they display significantly decreased mitochondrial activity and lower amounts of ROS. In contrast, morphologically normal maternally aged oocytes had the same mitochondrial activity as young ones, while their ROS levels were higher. Surprisingly, the shrunk phenotype was completely absent in maternally aged oocytes that matured in vitro, suggesting that it is not caused inherently by maternal aging, but may be related to other factors, like postovulatory aging. Indeed, an additional culture of in vitro matured young and old oocytes (i.e., in vitro postovulatory aging) significantly decreased their mitochondrial activity and led to cytoplasm shrinkage. In vivo postovulatory aging had a similar effect on oocytes from both young and old females. Finally, we examined the developmental potential of oocytes obtained from aged females. Shrunk (i.e., most likely postovulatory aged) oocytes failed to become fertilized, whereas morphologically normal ones (i.e., most likely not subjected to postovulatory aging) underwent fertilization and subsequent cleavage divisions, although they achieved the 2-cell stage less frequently than morphologically normal oocytes from young females. Importantly, the quality of blastocysts as well as the live birth rate for morphologically normal oocytes from old and young females were similar. In summary, our data clearly indicate that two pools of oocytes present in oviducts of aged females differ significantly in their quality and developmental potential and that the more severely affected phenotype results most likely from a synergistic action of maternal and postovulatory aging.

Maternal aging is associated with a decline in oocyte quality, which leads to the decreased fertility. Therefore, developing approaches to reduce aging-induced deterioration of oocyte quality in older women is important. Near-infrared cell protector-61 (IR-61), a novel heptamethine cyanine dye, has the potential for antioxidant effects. In this study, we found that IR-61 can accumulate in the ovaries and improved ovarian function of naturally aged mice; it also increased the oocyte maturation rate and quality by maintaining the integrity of the spindle/chromosomal structure and reducing the aneuploidy rate. In addition, the embryonic developmental competence of aged oocytes was improved. Finally, RNA-sequencing analysis indicated that IR-61 might perform the beneficial effects on aged oocytes by regulating mitochondrial function, this was confirmed by immunofluorescence analysis of mitochondrial distribution and reactive oxygen species. Taken together, our findings demonstrate that IR-61 supplementation in vivo can increase oocyte quality and protect oocytes from aging-induced mitochondrial dysfunction, and thus could improve the fertility of older women and efficiency of assisted reproductive technology.

The Edinburgh postnatal depression scale (EPDS) is commonly used in screening for major postpartum depression (PPD). We explored the clinical factors associated with score changes.
Mothers (n=1,287) who delivered a single live-born infant in Kokura Medical Center in Japan during 2018-2019 were analyzed. The EPDS-Japanese version was conducted at the first and fourth weeks after childbirth. Scores of ≥9 were considered to indicate an increased risk of PPD.
The scores improved during the four-week period (5.03±0.12 to 3.79±0.10). Primiparity, Cesarean section (CS), and a low Apgar score were identified as initial risk factors, however, primiparity remained in the multivariate analysis (aOR 2.02, 95% CI 1.37-2.97). Age ≥35 years was associated with worsened scores (aOR 1.88, 95%CI 1.01-3.51), but CS improved (aOR 0.38, 95%CI 0.21-0.70). Primiparity, CS, and neonatal respiratory support were the initial risk factors, while infant anomaly was a late risk factor in mothers whose infants were admitted to the neonatal intensive care unit (NICU) (aOR 3.35, 95%CI 1.31-8.56). In mothers of infants with an NICU stay of ≥4 weeks, infant anomaly was associated with worsened scores (aOR 6.61, 95%CI 1.11-39.3), while respiratory support was associated with improved scores (aOR 0.09, 95%CI 0.01-0.65). Twenty-six mothrs with worsened scores received psychiatric support; three developed PPD. Two of the three were ≥35 years of age, neither of their infants had anomalies.
Maternal aging and infant anomaly were risk factors for PPD. PPD occurred in mothers with worsened EPDS scores after mental care. Puerperants with worsening risk factors should be targeted to control PPD.

Human female fertility and reproductive lifespan decrease significantly with age, resulting in an extended post-reproductive period. The central dogma in human female reproduction contains two important aspects. One is the pool of oocytes in the human ovary (the ovarian reserve; approximately 106 at birth), which diminishes throughout life until menopause around the age of 50 (approximately 103 oocytes) in women. The second is the quality of oocytes, including the correctness of meiotic divisions, among other factors. Notably, the increased rate of sub- and infertility, aneuploidy, miscarriages, and birth defects are associated with advanced maternal age, especially in women above 35 years of age. This postponement is also relevant for human evolution; decades ago, the female aging-related fertility drop was not as important as it is today because women were having their children at a younger age. Spindle assembly is crucial for chromosome segregation during each cell division and oocyte maturation, making it an important event for euploidy. Consequently, aberrations in this segregation process, especially during the first meiotic division in human eggs, can lead to implantation failure or spontaneous abortion. Today, human reproductive medicine is also facing a high prevalence of aneuploidy, even in young females. However, the shift in the reproductive phase of humans and the strong increase in errors make the problem much more dramatic at later stages of the female reproductive phase. Aneuploidy in human eggs could be the result of the non-disjunction of entire chromosomes or sister chromatids during oocyte meiosis, but partial or segmental aneuploidies are also relevant. In this review, we intend to describe the relevance of the spindle apparatus during oocyte maturation for proper chromosome segregation in the context of maternal aging and the female reproductive lifespan.

Aneuploidy originating during meiosis in oocytes is the major cause of reduced fertility, implantation failure and miscarriage in women beyond their mid-thirties. Loss of chromosome cohesion, and defective microtubule dynamics and spindle assembly are, in turn, the major contributors to the error-prone nature of chromosome segregation in the oocytes of older women. However, the underlying molecular defects are not well understood. Altered function of MPS1 and AURKC have been shown to induce multipolar spindle phenotypes in murine oocytes and cancer cells, however, their role in reproductive aging associated oocyte aneuploidy is not known. Although age-related gamete and embryonic aneuploidy has been studied in female rodents, the horse may be a more appropriate animal model. Similar to women, aged mares suffer from reduced fertility and an increased incidence of oocyte aneuploidy. Moreover, mares show a long interval (decades) to reproductive senescence and, unlike rodents but similar to women, horse oocytes assemble the meiotic spindle in a slow and unstable manner, independent of microtubule organizing centers. In this study we found that oocytes from aged mares have lower expression of mRNA for Mps1, Spc25 and AurkC than oocytes from young mares while gene expression for other meiosis regulators did not differ. To assess the ability of horse oocytes to correctly form a bipolar spindle, in vitro matured MII oocytes were allowed to re-form their spindle after nocodazole-induced microtubule depolymerization. To investigate the importance of MPS1 and AURKC function in spindle (re)assembly, various concentrations of a MPS1 inhibitor (MPS1i, Compound 5) or an AURK inhibitor (AURKi, ZM447439) were included after nocodazole washout. MII oocytes from aged mares showed a higher incidence of spindle abnormalities after exposure to MPS1i. In contrast, Aurora kinase inhibition severely impaired microtubule organization and spindle formation in all oocytes, irrespective of mare age. In conclusion, gene expression for the kinases Mps1, Spc25, and AurkC is reduced in oocytes from aged mares. Moreover, spindle (re)assembly in aged mares\' oocytes is more unstable when Mps1 is inhibited. Overall, this suggests that compromised Mps1 activity predisposes to meiotic spindle instability in aged mare oocytes. This spindle instability could predispose to chromosome segregation errors.