ArticleLiterature Review

Oocyte senescence: A firm link to age-related female subfertility

Gynecological Endocrinology

March 2008
24(2):59-63

DOI:10.1080/09513590701733504

Source
PubMed

Authors:

Carla Tatone

University of L'Aquila

In women optimal fertility is maintained until 30 years of age and then decreases sharply. Therefore, the trend to delay childbearing until the fourth decade of life has contributed highly to the decline in total fertility rate observed in Western countries in recent decades. Much evidence supports the idea that age-related subfertility is mostly related to oocyte senescence. Based on the finding that maternal age negatively affects the storage of oocyte transcripts involved in major cellular functions, the present paper reviews the nuclear and cytoplasmic failure of the aged oocyte which can exert a negative influence on its developmental competence. Recognizing the potential role of oocyte-based technologies for improving clinical outcome for women with age-related etiologies of infertility, the importance of basic research aimed to increase knowledge of the aged oocyte and its microenvironment is also highlighted in order to set up new therapeutic strategies.

Prognostic Value of Oocyte Quality in Assisted Reproductive Technology Outcomes: A Systematic Review

Article

Mar 2021

Objective The aim of this systematic review was to survey and assess modern methodologies used to test oocyte quality that have prognostic value in predicting assisted reproductive technology (ART) outcomes. Evidence Review Following PRISMA guidelines, we surveyed the English-language literature between January 1st, 2010 and December 31st, 2019 using PubMed, Scopus, and Embase databases. Two reviewers screened for texts focusing on oocyte quality markers that predict ART outcomes, including fertilization, embryo quality, implantation, pregnancy, continued pregnancy, or live birth rate. Manuscripts that did not mention oocytes, or focused on non-human subjects, oocyte aging, oocyte maturation, embryo quality, interventions, or specific clinical diagnoses (endometriosis and polycystic ovarian syndrome) were deemed outside the scope of this analysis and excluded. Results Twenty-six relevant texts were identified, including 19 prospective and 7 retrospective studies (n=2,210 patients). We identified three general approaches for oocyte quality assessment: morphological evaluation (11 papers), genomics and proteomics (13 papers), and artificial intelligence (2 papers). Morphologic assessment did not show a consistent pattern of predictive value in IVF outcomes (7 papers in favor of its predictive value, 4 against). A considerable proportion of genomic and proteomic articles identified promising biomarkers that may predict pregnancy and live birth (12 in favor, 1 against). Machine learning is a rapidly growing frontier which minimizes subjectivity while potentially improving predictive ability (2 in favor). Conclusions Although there remains a lack of consensus on optimal methods to predict reproductive success, machine learning and genomics demonstrate promise in improving understanding of oocyte quality assessment and prognostication.

Polimorfismo CAG do gene AR e comprimento do telômero em mulheres com falência ovariana prematura

Poster

Oct 2014

Skewed X-chromosome inactivation and shorter telomeres associate with idiopathic premature ovarian insufficiency

Article

Aug 2018
FERTIL STERIL

Objective: To analyze whether telomere length, X-chromosome inactivation (XCI), and androgen receptor (AR) GAG polymorphism are related to idiopathic premature ovarian insufficiency (POI). Design: Case-control study. Setting: University hospital. Patient(s): A total of 121 women, including 46 nonsyndromic POI and 75 controls. Intervention(s): None. Main outcome measure(s): Age, weight, height, body mass index (BMI), systolic and diastolic arterial pressure, E2, androstenedione, T, and C-reactive protein were assessed. Telomere length was estimated by quantitative real-time polymerase chain reaction, XCI was measured using the Human Androgen Receptor and X-linked retinitis pigmentosa 2 (RP2) methylation assays. AR and FMR1 polymorphism was assessed by quantitative fluorescent polymerase chain reaction and sequencing. Result(s): Premature ovarian insufficiency women had a higher mean age, weighed less, and exhibited lower C-reactive protein, E2, and androstenedione levels. The AR polymorphism did not differ between the groups. Four patients had premutation (55-200 CGG repeats), and none displayed a full mutation in the FMR1 gene. However, patients with POI showed shorter telomere length and higher frequency of skewed XCI. Extreme skewing (≥90%) was observed in 15% of women with POI, and shorter telomeres correlated with XCI skewing in both groups. Conclusion(s): Skewed XCI and shortened telomere length were associated with idiopathic POI, despite no alterations in the AR and FMR1 genes. Additionally, there is a tendency for women with short telomeres to exhibit skewed XCI.

Potential risks to offspring of intrauterine exposure to maternal age-related obstetric complications

Article

Full-text available

Aug 2016

Several hypotheses have been proposed to explain the negative effects of delayed motherhood on an offspring’s morbidity later in life. However, these hypotheses are not supported by clinical and epidemiological evidence. Because advanced maternal age is associated with increased risk of obstetric complications, the aim of the present study was to ascertain whether the negative effects on offspring of intrauterine exposure to maternal age-related obstetric complications may explain the reported negative effects of delayed motherhood on offspring. To this end, a literature search was performed to identify relevant publications up to March 2016 on PubMed; references cited in relevant articles were also searched. There was a direct correlation between the risks to offspring conferred by intrauterine exposure to at least one of the obstetric complications present at the time of delivery in women aged ≥35 years and the risks to offspring of delayed motherhood. This correlation was not observed when comparing the risks to offspring of delayed motherhood and the risks associated with maternal transmission of defective mitochondria, chromosomal anomalies or DNA double-strand breaks. Most of the effects on offspring of intrauterine exposure to maternal age-related obstetric complications may be induced by epigenetic DNA reprogramming during critical periods of embryo or fetal development. Women wanting to enrol in a fertility preservation program to offset age-related declines in fertility should be informed not only about their chances of pregnancy and the percentage of live births, but also about the risks to themselves and their prospective offspring of delaying motherhood.

Effects of reproductive aging and postovulatory aging on the maintenance of biological competence after oocyte vitrification: Insights from the mouse model

Article

Jun 2011

Oxidative Stress and Human Ovarian Response—From Somatic Ovarian Cells to Oocytes Damage: A Clinical Comprehensive Narrative Review

Article

Full-text available

Jul 2022

Basic scientific research on human reproduction and oxidative damage has been extensively performed; however, a more clinical view is still lacking. As a result, exhaustive data on the influence of oxidative stress on human ovarian response and, consequently, on fertility are still lacking. This narrative review aims at summarizing the role of oxidative stress in different conditions associated to female infertility and to list some of the main antioxidant agents. A systematic literature search was performed in May 2022 to retrieve studies regarding the oxidative stress and the human ovarian response from somatic ovarian cells to oocytes damage. Only human studies were included and the authors focused their review, in particular, on clinical implications in order to define a new research perspective on the assessment of any eventual strategy to preserve women’s fertility. Thereby, the authors evaluated the contribution of DNA repair pathways in improving women’s fertility by reducing the DNA damage associated with aging or diseases, such as endometriosis or polycystic ovary syndrome, and eventually, in prolonging the reproductive lifespan after cancer treatment.

Per- and polyfluoroalkyl substances (PFAS) exposure in women seeking in vitro fertilization-embryo transfer treatment (IVF-ET) in China: Blood-follicular transfer and associations with IVF-ET outcomes

Article

May 2022
SCI TOTAL ENVIRON

As follicular fluid constitutes a critical microenvironment for the development of oocytes, investigation of environmental contaminants in follicular fluid may facilitate a better understanding of the influence of environmental exposure on reproductive health. In the present study, we aimed to investigate per- and polyfluoroalkyl substances (PFAS) exposure in women receiving in vitro fertilization-embryo transfer (IVF-ET) treatment, determine the blood-follicle transfer efficiencies (BFTE) of PFAS, and explore potential associations between PFAS exposure and selected IVF-ET outcomes. Our results revealed that n-PFOA was the most abundant PFAS in both serum and follicular fluid (FF) (median = 5.85 and 5.56 ng/mL, respectively), followed by n-PFOS (4.95 and 4.28 ng/mL), 6:2 Cl-PFESA (2.18 and 2.10 ng/mL), PFNA (1.37 and 1.37 ng/mL), PFUdA (0.33 and 0.97 ng/mL), PFDA (0.37 and 0.66 ng/mL), PFHxS (0.42 and 0.39 ng/mL), and PFHpS (0.11 and 0.10 ng/mL). The median BFTE ranged from 0.65 to 0.92 for individual PFAS, indicating a relatively high tendency of PFAS to cross the blood-follicle barrier (BFB). An inverted V-shaped trend was observed between the median BFTE and the number of fluorinated carbon atoms or the log Kow (octanol-water partition coefficient) for individual PFAS, suggesting the influence by physicochemical properties and molecular structures. Although our data did not find any clear pattern in the link between blood or follicular fluid concentrations of PFAS and selected IVF-ET outcomes, our study raises the need for better characterization of exposure to environmental chemicals in follicular fluid together with its potential influence on reproductive health.

Integrative proteome analysis implicates aberrant RNA splicing in impaired developmental potential of aged mouse oocytes

Article

Full-text available

Sep 2021
AGING CELL

Aging has many effects on the female reproductive system, among which decreased oocyte quality and impaired embryo developmental potential are the most important factors affecting female fertility. However, the mechanisms underlying oocyte aging are not yet fully understood. Here, we selected normal reproductively aging female mice and constructed a protein expression profile of metaphase II (MII) oocytes from three age groups. A total of 187 differentially expressed (DE) proteins were identified, and bioinformatics analyses showed that these DE proteins were highly enriched in RNA splicing. Next, RNA-seq was performed on 2-cell embryos from these three age groups, and splicing analysis showed that a large number of splicing events and genes were discovered at this stage. Differentially spliced genes (DSGs) in the two reproductively aging groups versus the younger group were enriched in biological processes related to DNA damage repair/response. Binding motif analysis suggested that PUF60 might be one of the core splicing factors causing a decline in DNA repair capacity in the subsequent development of oocytes from reproductively aging mice, and changing the splicing pattern of its potential downstream DSG Cdk9 could partially mimic phenotypes in the reproductively aging groups. Taken together, our study suggested that the abnormal expression of splicing regulation proteins in aged MII oocytes would affect the splicing of nascent RNA after zygotic genome activation in 2-cell embryos, leading to the production of abnormally spliced transcripts of some key genes associated with DNA damage repair/response, thus affecting the developmental potential of aged oocytes.

Rabbit as an Aging Model in Reproduction: Advanced Maternal Age Alters GLO1 Expression in the Endometrium at the Time of Implantation

Article

Full-text available

Oct 2020

Advanced maternal age is associated with adverse pregnancy outcomes and the decline of female fertility in mammals. A potential reason for reduced fertility is metabolic changes due to protein modifications by advanced glycation end products. To elucidate the aging process in female reproduction, we analysed a key enzyme for detoxification of reactive dicarbonyls, the glyoxalase 1 (GLO1), in reproductive organs and blastocysts of young and old rabbits at the preimplantation stage. At day 6 post coitum, uterine, oviductal, ovarian tissue and blastocysts from young (16–20 weeks) and old rabbits (>108 weeks) were characterised for GLO1 expression. GLO1 amounts, enzymatic activity and localisation were quantified by qPCR, Simple Western, activity assay and immunohistochemistry. The GLO1 enzyme was present and active in all reproductive tract organs in a cell-type-specific pattern. Ovarian follicle and uterine epithelial cells expressed GLO1 to a high extent. In tertiary follicles, GLO1 expression increased, whereas it decreased in the endometrium of old rabbits at day 6 of pregnancy. In blastocysts of old animals, GLO1 expression remained unchanged. In early pregnancy, advanced maternal age leads to modified GLO1 expression in ovarian follicles and the endometrium, indicating an altered metabolic stress response at the preimplantation stage in older females.

IVF outcomes of women with discrepancies between age and serum anti-Müllerian hormone levels

Article

Full-text available

Jul 2019
REPROD BIOL ENDOCRIN

Background: To determine the effects of age and the serum anti-Müllerian hormone (AMH) level on in vitro fertilization (IVF) outcomes, especially among young women with low serum AMH levels and older women with high AMH levels. Methods: This study was a cohort study in which a total of 9431 women aged 20-51 years who were undergoing their first IVF cycles were recruited. Ovarian response parameters included the number of retrieved oocytes, the number of 2 pronuclear zygotes (2PN), and the frequency of good-quality embryos (GQE). Pregnancy outcomes included the clinical pregnancy rate (CPR), live birth rate (LBR), miscarriage rate (MR), and cumulative CPR and LBR (CCPR and CLBR). Results: Among women under 35 years of age, the ovarian response, CPR, CCPR, LBR and CLBR (p < 0.01) were significantly lower in the low-AMH group than in the average-AMH and high-AMH groups. In women above 35 years of age, the ovarian response, CPR, CCPR and CLBR (p < 0.01) were significantly higher in the average-AMH and low-AMH groups. The LBR in the older high-AMH group was significantly higher (37.45% vs 20.34%, p < 0.01) than that in the older low-AMH group, but there was no difference (37.45% vs 32.46%, p = 0.11) compared with the older average-AMH group. When there was a discrepancy between age and the AMH level, the young low-AMH group showed a poorer ovarian response but a better CPR (58.01% vs 49.44%, p < 0.01) and LBR (48.52% vs 37.45%, p < 0.01) than the older high-AMH group. However, the CCPR (65.37% vs 66.11%, p = 0.75) and CLBR (56.35% vs 52.89%, p = 0.15) between the two groups were comparable. The conservative CLBR in the two discrepancy groups increased until the third embryo transfer and reached a plateau thereafter. Conclusion(s): Even with a relatively low AMH level, young women still had better pregnancy outcomes following IVF than older women. However, increasing the AMH level improves the cumulative outcomes of the older group to a comparable level through a notable and superior ovarian response.

DNA damage and repair in the female germline: contributions to ART

Article

Mar 2019
HUM REPROD UPDATE

Background: DNA integrity and stability are critical determinants of cell viability. This is especially true in the female germline, wherein DNA integrity underpins successful conception, embryonic development, pregnancy and the production of healthy offspring. However, DNA is not inert; rather, it is subject to assault from various environment factors resulting in chemical modification and/or strand breakage. If structural alterations result and are left unrepaired, they have the potential to cause mutations and propagate disease. In this regard, reduced genetic integrity of the female germline ranks among the leading causes of subfertility in humans. With an estimated 10% of couples in developed countries taking recourse to ART to achieve pregnancy, the need for ongoing research into the capacity of the oocyte to detect DNA damage and thereafter initiate cell cycle arrest, apoptosis or DNA repair is increasingly more pressing. Objective and rationale: This review documents our current knowledge of the quality control mechanisms utilised by the female germline to prevent and remediate DNA damage during their development from primordial follicles through to the formation of preimplantation embryos. Search methods: The PubMed database was searched using the keywords: primordial follicle, primary follicle, secondary follicle, tertiary follicle, germinal vesical, MI, MII oocyte, zygote, preimplantation embryo, DNA repair, double-strand break and DNA damage. These keywords were combined with other phrases relevant to the topic. Literature was restricted to peer-reviewed original articles in the English language (published 1979-2018) and references within these articles were also searched. Outcomes: In this review, we explore the quality control mechanisms utilised by the female germline to prevent, detect and remediate DNA damage. We follow the trajectory of development from the primordial follicle stage through to the preimplantation embryo, highlighting findings likely to have important implications for fertility management, age-related subfertility and premature ovarian failure. In addition, we survey the latest discoveries regarding DNA repair within the metaphase II (MII) oocyte and implicate maternal stores of endogenous DNA repair proteins and mRNA transcripts as a primary means by which they defend their genomic integrity. The collective evidence reviewed herein demonstrates that the MII oocyte can engage in the activation of major DNA damage repair pathway(s), therefore encouraging a reappraisal of the long-held paradigm that oocytes are largely refractory to DNA repair upon reaching this late stage of their development. It is also demonstrated that the zygote can exploit a number of protective strategies to mitigate the risk and/or effect the repair, of DNA damage sustained to either parental germline; affirming that DNA protection is largely a maternally driven trait but that some aspects of repair may rely on a collaborative effort between the male and female germlines. Wider implications: The present review highlights the vulnerability of the oocyte to DNA damage and presents a number of opportunities for research to bolster the stringency of the oocyte's endogenous defences, with implications extending to improved diagnostics and novel therapeutic applications to alleviate the burden of infertility.

Melatonin regulates the activities of ovary and delays the fertility decline in female animals via MT1/AMPK pathway

Article

Dec 2018

Female fertility irreversibly declines with aging and this, is primarily associated with the decreased quality and quantity of oocytes. To evaluate whether a long‐term of melatonin treatment would improve the fertility of aged mice, different concentrations of melatonin (10‐3, 10‐5, 10‐7 M) were supplemented into drinking water. Melatonin treatments improved the litter sizes of mice at the age of 24 weeks. Mice treated with 10‐5 M melatonin had the largest litter size among other concentrations. At this optimal concentration, melatonin not only significantly increased the total number of oocytes but also their quality, having more oocytes with normal morphology that could generate more blastocyst after in vitro fertilization in melatonin (10‐5 M) treated group than that in the controls. When these blastocysts were transferred to recipients, the litter size was also significantly larger in melatonin treated mice than that in controls. The increases in TAOC and SOD level, and decreases in MDA were detected in ovaries and uterus from melatonin treated mice compared to the controls. Melatonin reduced ROS level and maintained mitochondrial membrane potential in the oocytes cultured in vitro. Mechanistically studies revealed that the beneficial effects of melatonin on oocytes were mediated by MT1 receptor and AMPK pathway. Thereafter, MT1 knocking out (MT1‐KO) were generated and shown significantly reduced number of oocytes and litter size. The expression of SIRT1, C‐myc, and CHOP were down‐regulated in the ovary of MT1‐KO mice, but SIRT1 and p‐NF‐kB protein level were elevated in response to disturbed redox balance. The results have convincingly proven that melatonin administration delays ovary aging and improves fertility in mice via MT1/AMPK pathway. This article is protected by copyright. All rights reserved.

Sirtuins in gamete biology and reproductive physiology: Emerging roles and therapeutic potential in female and male infertility

Article

Feb 2018
HUM REPROD UPDATE

Background: Sirtuins (SIRT1-7) are a family of NAD+-dependent deacetylases that catalyze post-translational modifications of proteins. Together, they respond to metabolic challenges, inflammatory signals or hypoxic/oxidative stress, and are associated with aging and longevity. The role of Sirtuins in the regulation of fertility emerged in 2003 when a defective reproductive phenotype was observed in SIRT1-null mice. Although studies on Sirtuins in reproductive biology have been increasing in the last years, a recent comprehensive update on this issue is still lacking. Objective and rationale: This review is aimed to provide knowledge on the activation mechanism and cellular role of Sirtuins and to give an update of the rapid development of Sirtuin research in female and male reproduction under physiological and pathological conditions. The final goal is to assess whether strategies aimed to improve Sirtuin expression or activity could have therapeutic potential for infertility associated with polycystic ovarian syndrome (PCOS), endometriosis, diabetes, xenobiotic stress and aging. Search methods: The MEDLINE database was examined for peer-reviewed original articles. The following keywords were searched: 'Sirtuin', 'ovary', 'oocyte', 'ovarian follicle', 'embryo', 'endometrium', 'sperm' and 'testis'. These keywords were combined with other search phrases relevant to the topic. Outcomes: Our knowledge of Sirtuins in reproductive functions has grown exponentially over the last few years. The majority of the work carried out so far has focused on SIRT1 with a prevalence of studies on female reproduction. Numerous studies have provided evidence that down-regulation of SIRT1 is associated with physiological or pathological reduction of ovarian reserve. SIRT1 has also been shown to regulate proliferation and apoptosis in granulosa cells whereas SIRT3 was found to promote luteinisation. Biochemical modulation of Sirtuin activity has led to discoveries of the roles of SIRT1, SIRT2, SIRT3 and SIRT6 in improving the competence of oocytes grown or matured in vitro in humans and animal models. Recently, SIRT1, SIRT2 and SIRT3 have emerged as protectors of oocyte against postovulatory aging. Transgenic models provide strong evidence that SIRT1 is involved in spermatogenesis by influencing specific functions of male germ cell, Sertoli cells and Leydig cells. When our attention moves to post-fertilization events, maternally derived SIRT3 appears crucial in the protecting early embryos against stress conditions. Finally, increasing SIRT1 activity may have the potential to ameliorate fertility in PCOS, diabetes, endometriosis, xenobiotic stress and aging. Overall, these effects have been ascribed to Sirtuin-mediated regulation of energy homoeostasis, mitochondrial biogenesis, chromatin remodelling and protection against oxidative stress. Wider implications: The present review provides challenges and opportunities to stimulate research and exploit Sirtuin-based signalling as diagnostic tools and potential targets for therapeutic applications in reproductive medicine.

CD-1 mouse fertility rapidly declines and is accompanied with early pregnancy loss under conventional housing conditions

Article

Dec 2017
THERIOGENOLOGY

CD-1 mice are commonly employed as a research model for defining mechanisms controlling early mammalian development and for understanding environmental impacts on mammalian fertility. CD-1 female mice were kept four to eight months under conventional animal care housing, and were fed ad libitum with normal laboratory mouse chow. Female weight, mating success, oocyte morphology, blastocyst development in vivo and in vitro, and RT-qPCR analysis of trophectoderm cell markers (Cdx2, Slc2a1, and Atp1a1 transcript abundance, and CDX2 localization) were assessed and contrasted with outcomes from four-week-old control CD-1 mice. Embryo development in vivo in four to eight-month-old mice was significantly reduced compared to four-week-old controls. Oocytes and blastocysts from four to eight-month-old CD-1 mice displayed high levels of fragmentation and degradation, significantly reduced embryo cell counts, decreased Cdx2 transcript abundance, and number of CDX2 positive cells in morulae. We have discovered that female CD-1 mice housed under conventional conditions display a rapid loss of fecundity as they age over a few months. Paradoxically, embryo loss can be avoided by placing early embryos collected from four to eight-month-old mice into culture to support development to the blastocyst stage. We conclude that oocyte quality rapidly declines in CD-1 female mice housed under conventional animal care conditions. Thus, four to eight-month-old female CD-1 mice represent a very distinct research model from that of younger mice and this older research animal model may be preferred for understanding environmental and physiological influences limiting fertility in women.

H2O2-induced mild stress in relation with in vitro ovine oocyte developmental competence: Implications for blastocyst apoptosis and related genes expression

Article

May 2017
CELL MOL BIOL

In this study, in vitro maturation was performed in presence of various concentrations (0, 10, 100, or 1000 μM) of H2O2. The intracellular glutathione (GSH) level, fertilization, cleavage, and blastocyst rates, total cell number, and apoptotic cell number and expression of Bax, Bcl-2, and p53 genes in blastocyst-stage embryos were studied. At 10 μM H2O2 concentration, a higher GSH level was detected in comparison to the other groups while oocytes exposed to 1000 μM H2O2 had the lowest GSH level. Treatment of oocytes with 1000 μM H2O2 decreased the rate of two pronuclei formation as compared with other groups. A higher rate of blastocyst formation was seen in 100 μM H2O2 group as compared with the control group. However, exogenous H2O2 in maturation medium did not affect total cell numbers and apoptotic cell ratio at the blastocyst stage. Moreover, mRNA transcript abundance of Bax, Bcl-2, and p53 genes was similar between blastocysts derived from H2O2-induced oocytes and control blastocysts. Treatment of oocytes with H2O2 at mild level during in vitro maturation had a positive effect on GSH level and this, in turn, may lead to improvement in preimplantation embryonic development.

Impact of vitrification on the mitochondrial activity and redox homeostasis of human oocyte

Article

Full-text available

May 2016

STUDY QUESTION Do the extreme conditions of vitrification affect mitochondrial health and reactive oxygen species (ROS) levels of human oocytes? SUMMARY ANSWER Vitrification of discarded human oocytes shifts the intracellular redox potential towards oxidation but does not alter the mitochondrial potential or intracellular ROS levels. WHAT IS KNOWN ALREADY Recent studies have reflected increased ROS levels in warmed young oocytes and have highlighted the temporal dynamic loss of mitochondrial potential that could, therefore, lead to a decrease in ATP production, impairing embryo development. Mitochondrial function can also be evaluated in vivo by the FAD/NAD(P)H autofluorescence ratio, which reflects the respiratory chain activity and is considered as a marker of the intracellular redox state. STUDY DESIGN, SIZE, DURATION A total of 629 discarded Metaphase II (MII) oocytes collected from June 2013 to April 2014 were included in this control (fresh oocytes, n= 270) versus treatment (vitrified oocytes, n= 359) study. PARTICIPANTS/MATERIALS, SETTING, METHODS Discarded MII oocytes were donated to research by young (36 years old) women who underwent ovarian stimulation for IVF at a university-affiliated private fertility clinic. Redox state was assessed by measuring the FAD/NAD(P)H autofluorescence ratio, while ROS and mitochondrial activity were reported by in vivo labelling with carboxy-H2DCFDA and JC-1, respectively. MAIN RESULTS AND THE ROLE OF CHANCE Young and aged oocytes showed high and similar survival rates (81.8 versus 83.1%, not significant). Confocal microscopy revealed that the FAD/NAD(P)H ratio was significantly higher in vitrified oocytes than in fresh oocytes, suggesting a significant shift towards the oxidized state in oocytes after vitrification, regardless of the maternal age. Mitochondrial distribution was not affected by vitrification. Furthermore, it was not possible to resolve any difference in mitochondrial potential using JC-1 potentiometric dye or in reactive oxygen species (ROS) production (assessed with H2-DCFDA staining) between fresh and vitrified oocytes. Therefore, measurement of intracellular redox potential by autofluorescence imaging may be a more sensitive method to assess oxidative stress or mitochondrial demise in human oocytes because it showed a higher resolving power than JC-1 staining and displayed less variability than H2-DCFDA staining. LIMITATIONS, REASONS FOR CAUTION Owing to sample availability, MII discarded oocytes (in vitro matured oocytes and unfertilized oocytes 20 h after ICSI) were included in the study. These discarded oocytes do not necessarily reflect the physiological condition of the MII human oocyte. WIDER IMPLICATIONS OF THE FINDINGS Although vitrified oocytes yield comparable clinical outcomes compared with fresh oocytes, lower cleavage and blastocyst rates can be observed during in vitro culture. Data here obtained suggest that the redox state of human oocytes could be affected by vitrification. Therefore, the importance of adding protective antioxidant molecules to the vitrification solution and to the post-warming culture medium to improve embryo cleavage deserves some research. STUDY FUNDING/COMPETING INTEREST(S) This research project was supported by the Valencian Government (Val+i+D program, M.N.-C.), INCLIVA Foundation for health research (G.S.-A.) and by the University of L'Aquila and Regione Abruzzo (‘Reti per l'Alta Formazione’ – P.O.F.S.E. Abruzzo 2007–2013 G.D.E.). No conflicts of interest were declared.

Vitrification impact on human oocyte mitochondrial activity and redox homeostasis

Conference Paper

Full-text available

Jul 2014

Decoding the Proteome of In-Vitro Fertilization Ovarian Follicular Fluid for Women Over 35 Years

Article

Jan 2016

Aim: The study of follicular fluid using proteomic techniques could provide a useful tool for understanding follicular fluid components and their effect on pregnancy outcome. The aim of the study is to identify and catalog follicular fluid proteins in women 35 years of age or older. Material and Method: Follicular fluid was collected from 21 couples, of which 11 couples achieved successful pregnancy and 10 couples failed to get pregnant. Samples were analyzed by multidimensional chromatography coupled with in-line nanospray ionization mass spectrometry on an LTQ XL ion trap mass spectrometer. We used the Biomarker Analysis Program from PDQuest software to identify protein constituents in pregnant and non-pregnant groups. Results: In total, 1024 protein specimens were identified. The proteins identified were consistent throughout the experiment and within each of the analyzed specimens. Discussion: A compiled listing of follicular fluid proteins could be a potential starting point for the identification and evaluation of important proteins involved in the development of oocytes; the results of our study may fill a noticeable knowledge-gap in the understanding of follicular fluid proteome. © 2016, Journal of Clinical and Analytical Medicine. All rights reserved.

Advanced Maternal Age and Assisted Reproductive Technologies in an Irish Population

Article

Full-text available

Sep 2015
Ir Med J

In recent decades the amount of women over 40 seeking assisted reproductive technology (ART) interventions in order to become pregnant has dramatically increased, both in Ireland and worldwide. This is due to an increase in the average age at which women are choosing to have their first child while additionally, many couples are choosing to have a second family later in life. However, as with natural conception, ART success rates decrease with maternal age. In the present study, we perform a 16 year retrospective analysis on our clinical data of women between 40 and 45 years of age, who have undergone ART at a tertiary referral ART clinic. The percentage of patients in this age group was analysed over time, in order to determine follicle recruitment, % oocyte yield, embryonic quality, positive hCG (pregnancy rate), clinical pregnancy rate and rate of preclinical pregnancy loss. Results from our clinic show that women greater than 43 years of age have a significantly reduced reproductive potential compared to women in the 40 to 42 years age group. Woman in the 43-45 age group showed reduced fertilization rates (53.73% versus 58.82%), reduced positive hCG rates (11.51% versus 19.03%) and clinical pregnancy rates (5.04% versus 12.52%) and increased rates of preclinical pregnancy loss (56.23% versus 34.23%), compared to women in the 40-42 age group. With the age at which couples are choosing to have children constantly increasing, novel ART treatment strategies need to be developed. http://imj.ie//ViewArticleDetails.aspx?ArticleID=15111

JOE150246

Data

Full-text available

Aug 2015

Article

Aug 2015
J ENDOCRINOL

Why in vitro fertilization (IVF) pregnancy rates decline sharply after age 43, is unknown. In this study we compared granulosa cell (GC) function in young oocyte donors (n=31, ages 21-29), middle-aged (n=64, ages 30-37) and older infertile patients (n=41, ages 43-47). Gene expressions related to gonadotropin activity, steroidogenesis, apoptosis and luteinization were examined by real-time PCR and western blot in GCs collected from follicular fluid. FSH receptor (FSHR), aromatase (CYP19A1) and 17β-hydroxysteroid dehydrogenase (HSD17B) expression were found down regulated with advancing age, while LH receptor (LHCGR), P450scc (CYP11A1) and progesterone receptor (PGR) were up regulated. Upon in vitro culture, GCs were found to exhibit lower proliferation and increased apoptosis with aging. While FSH supplementation stimulated GCs growth and prevented luteinization in vitro. These observations demonstrate age-related functional declines in GCs, consistent with premature luteinization. To avoid premature luteinization in women above age 43, we advanced oocyte retrieval by administering human chorionic gonadotropin (hCG) at maximal leading follicle size of 16mm (routine 19-21mm). Compared to normal cycles in women of similar age, earlier retrieved patients demonstrated only a marginal increase in oocyte prematurity, yet exhibited improved embryo numbers as well as quality and respectable clinical pregnancy rates. Premature follicular luteinization appears to contribute to rapidly declining IVF pregnancy chances after age 43, and can be avoided by earlier oocyte retrieval.

Discovery of porcine maternal factors related to nuclear reprogramming and early embryo development by proteomic analysis

Article

Full-text available

Jun 2015
PROTEOME SCI

Differentiated cell nuclei can be reprogrammed to a pluripotent state in several ways, including incubation with oocyte extracts, transfer into enucleated oocytes, and induced pluripotent stem cell technology. Nuclear transfer-mediated reprogramming has been proven to be the most efficient method. Maternal factors stored in oocytes have critical roles on nuclear reprogramming and early embryo development, but remain elusive. In this study, we showed most of porcine oocytes became nuclear matured at 33 h of IVM and the rate had no significant difference with oocytes at 42 h of IVM (p > 0.05). Moreover, the cleavage and blastocyst rates of SCNT and PA embryos derived from 42O were significantly higher than that of 33O (p < 0.05). But 33O could sustain IVF embryo development with higher cleavage and blastocyst rates comparing to 42O (p < 0.05). To clarify the development potential difference between 33O and 42O, 18 differentially expressed proteins were identified by proteomic analysis, and randomly selected proteins were confirmed by Western blot. Bioinformatic analysis of these proteins revealed that 33O highly synthesized proteins related to fertilization, and 42O was rich in nuclear reprogramming factors. These results present a unique insight into maternal factors related to nuclear reprogramming and early embryo development.

Hormone replacement therapy (HRT) in optimising fertility in premature ovarian insufficiency (POI) patients. 16th World Congress of Gynecological Endocrinology, Florence, Italy, 7.03.2014.

Conference Paper

Full-text available

Mar 2014

Svetlana Dragojevic Dikic

Endometriosis may be associated with mitochondrial dysfunction in cumulus cells from subjects undergoing in vitro fertilization-intracytoplasmic sperm injection, as reflected by decreased adenosine triphosphate production

Article

Dec 2014
FERTIL STERIL

To determine whether endometriosis is associated with mitochondrial dysfunction in cumulus (granulosa [GC]) cells of subjects undergoing IVF-intracytoplasmic sperm injection (ICSI). Prospective cohort study. An IVF clinic in a tertiary academic care center. Eleven women with endometriosis and 39 controls. None. Cumulus cell adenosine triphosphate (ATP) levels, mitochondrial DNA (mtDNA), and genomic DNA (gDNA) number. Cumulus cell ATP content was 65% lower in subjects with surgically proven endometriosis (median 312.5 attomoles/ng total DNA, interquartile range = 116.0-667.8) compared with controls (median 892.4 attomoles/ng total DNA, interquartile range = 403.0-1,412.2). There was no significant difference in mtDNA:gDNA ratio. There were no significant differences in age, body mass index (BMI), basal serum FSH level, total oocyte number, metaphase II (M2) oocyte number, metaphase I oocyte number, percentage of M2 oocytes, fertilization rate, implantation rate, or pregnancy rate (PR). Multivariate regression analysis showed significant positive correlations between ATP and [1] M2 oocyte number (r = 0.307) and [2] pregnancy (r = 0.332). There were also trends toward positive correlations between ATP and [3] age (r = 0.283), [4] total number of oocytes (r = 0.271), [5] percentage of M2 oocytes (r = 0.249), and [6] implantation rate (r = 0.293). There were no statistically significant correlations between mtDNA:gDNA ratio and any demographic factors or clinical outcomes measured. Surgically confirmed endometriosis may be associated with cumulus cell mitochondrial dysfunction in subjects undergoing IVF-ICSI for infertility, as reflected by decreased ATP production. Copyright © 2014 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Hormone replacement therapy and successful pregnancy in a patient with premature ovarian failure

Article

Nov 2009

Premature ovarian failure (POF), premature ovarian insufficiency, premature menopause or hypergonadotropic hypogonadism, a serious life-changing condition that affects young women, remains an enigma and the researcher's challenge. In the present article we described a case of singleton pregnancy in a 33-year-old patient, presenting with POF and treated with hormone replacement therapy. Twenty months later this therapy led to maturation of one follicle, recruitment and fertilisation of the residual oocyte and spontaneous pregnancy ensued. A normal infant was delivered by cesarean section.

Conventional IVF as a laboratory strategy to rescue fertility potential in severe poor responder patients: The impact of reproductive aging

Article

Full-text available

Aug 2013
GYNECOL ENDOCRINOL

Objective: To investigate whether laboratory strategies can improve in vitro fertilization (IVF) outcome in poor responder patients. We compared the effectiveness of conventional IVF and intra cytoplasmic sperm injection (ICSI) in assisted reproductive technologies cycles in which only one or two oocytes were retrieved at ovarian pick up, in the absence of male infertility. Design: Retrospective analysis of 425 cycles in 386 poor responder patients. Intervention(s): Standard stimulation protocol with gonadotropins and gonadotropin releasing hormone (GnRH) antagonist. Main outcome measure(s): Fertilization rate, cleavage rate, good-quality embryo rate, implantation rate, clinical pregnancy rate (PR) and miscarriage rate. Results: IVF was found to be more advantageous for implantation and PR, especially in patients under 35 years and in women aged between 35 and 38 years. No differences were noted in the other parameter evaluated. Patients aged over 38 years showed no difference using the two techniques. Conclusion: The employment of ICSI in the absence of a male factor can reduce reproductive outcome in poor responder. Probably because of aging-related defects overcoming the advantage of sperm selection, the choice of IVF technique is not relevant to reproductive success when oocyte quality is compromised by reproductive aging. Although further randomized trials are needed to confirm our results, we propose that, in absence of male infertility, conventional IVF might be the technique of choice in young patients, especially in those aged below 35 years.

The effect of aging on the metabolic function and structure of mitochondria in Hamster oocytes

Article

Full-text available

Apr 2009
FASEB J

In human oocytes there is a significant variation in the pregnancy potential. This low proportion of fruitful couplings appears to be influenced by changes in mitochondrial structure and function. Hamster ova resemble human oocytes in size and fertility success rate and so serve as a model for understanding the human oocyte. ATP from the oocytes of young and aging hamsters was measured by a luciferase/luciferin reaction. On average, individual hamster oocytes from young animals (91 eggs from 6 animals) contain 144 ± 9.1 fmoles of ATP (0.16 uM). There is a 21% (P = 0.01) decrease in ATP (114 ± 10.1 fmoles, 0.14 uM in concentration) in old hamsters (88 eggs from 6 animals) compared to young hamsters. RT‐ PCR was used to quantitate mtDNA in individual oocytes. This analysis indicated that there is also a decrease in mtDNA molecules per oocyte as the animal ages (11,466 ± 2,025 (young) vs 7,544 ± 2,507 (old), P= 0.07). The morphology of oocytes of young and old hamsters was examined by transmission electron microscopy. TEM indicated that mitochondrial morphology undergoes distinct changes as the hamster ages. In old hamsters, the cristae are decreased in number and length compared to those in young animals. In hamsters, the decrease in ATP levels seen in aged oocytes is correlated with a similar decrease in the number of mtDNA molecules. The consequence of these changes is a reduction of 25% in ATP concentration. Such a dramatic decrease in mitochondrial energy production could compromise normal oocyte biogenesis and contribute to the reduced pregnancy potential seen in aged mammalian oocytes.

Age-Associated Metabolic and Morphologic Changes in Mitochondria of Individual Mouse and Hamster Oocytes

Article

Full-text available

May 2013
PLOS ONE

Background In human oocytes, as in other mammalian ova, there is a significant variation in the pregnancy potential, with approximately 20% of oocyte-sperm meetings resulting in pregnancies. This frequency of successful fertilization decreases as the oocytes age. This low proportion of fruitful couplings appears to be influenced by changes in mitochondrial structure and function. In this study, we have examined mitochondrial biogenesis in both hamster (Mesocricetus auratus) and mouse (Mus musculus) ova as models for understanding the effects of aging on mitochondrial structure and energy production within the mammalian oocyte. Methodology/Principal Findings Individual metaphase II oocytes from a total of 25 young and old mice and hamsters were collected from ovarian follicles after hormone stimulation and prepared for biochemical or structural analysis. Adenosine triphosphate levels and mitochondrial DNA number were determined within individual oocytes from young and old animals. In aged hamsters, oocyte adenosine triphosphate levels and mitochondrial DNA molecules were reduced 35.4% and 51.8%, respectively. Reductions of 38.4% and 44% in adenosine triphosphate and mitochondrial genomes, respectively, were also seen in aged mouse oocytes. Transmission electron microscopic (TEM) analysis showed that aged rodent oocytes had significant alterations in mitochondrial and cytoplasmic lamellae structure. Conclusions/Significance In both mice and hamsters, decreased adenosine triphosphate in aged oocytes is correlated with a similar decrease in mtDNA molecules and number of mitochondria. Mitochondria in mice and hamsters undergo significant morphological change with aging including mitochondrial vacuolization, cristae alterations, and changes in cytoplasmic lamellae.

Effect of maternal age on the outcomes of in vitro fertilization and embryo transfer (IVF-ET)

Article

Full-text available

Aug 2012
Sci China Life Sci

This is a retrospective, observational study to evaluate the effect of maternal age on the outcomes of in vitro fertilization and embryo transfer (IVF-ET). 11830 IVF-ET cycles from 10268 women were included. Four groups of different maternal age periods were compared. The groups were 21-30 years old group (4549 cycles), 31-35 years old group (4424 cycles), 36-40 years old group (2429 cycles), and over 40 years old group (428 cycles). The mean starting dose of Gn and mean total dose of Gn in each cycle were significantly higher (P<0.01), while the mean retrieved oocyte number was significantly lower (P<0.01) in groups of higher maternal age period than those in each of the lower groups. The biochemical pregnancy rate and the clinical pregnancy rate were significantly lower (P<0.01), while the miscarriage rate was significantly higher (P<0.01) in groups of higher maternal age period than those in the lower groups. No difference was found in two-pronuclear zygotes (2PN) rate and good quality embryo rate among different groups. Birth defect rate was also comparable in the born babies in different groups. In the group with patients' age over 40 years old, the pregnancy rate was 26.87%, the clinical pregnancy rate was 19.39%, while the miscarriage rate after clinical pregnancy was 36.14%. To draw the conclusion, patients with higher maternal age had worse IVF outcomes. In women of fertile age, patients between 20 and 30 years old have the best IVF outcomes. Patients over 40 years old have poor IVF outcome and high miscarriage rate, which suggested the necessity of preimplantation genetic screening (PGS).

Developmental competence in oocytes and cumulus cells: Candidate genes and networks

Article

Full-text available

Feb 2012
SYST BIOL REPROD MEC

Common aspects of infertility can be seen across several species. In humans, dairy cows, and mares there is only a 25-35% chance of producing a live offspring after a single insemination, whether natural or artificial. Oocyte quality and subsequent embryo development can be affected by factors such as nutrition, hormonal regulation, and environmental influence. The objective of this study was to identify genes expressed in oocytes and/or cumulus cells, across a diverse range of species, which may be linked to the ability an oocyte has to develop following fertilization. Performing a meta-analysis on previously published microarray data on various models of oocyte and embryo quality allowed for the identification of 56 candidate genes associated with oocyte quality across several species, 4 of which were identified in the cumulus cells that surround the oocyte. Twenty-one potential biomarkers were associated with increased competence and 35 potential biomarkers were associated with decreased competence. The upregulation of Metap2, and the decrease of multiple genes linked to mRNA and protein synthesis in models of competence, highlights the importance of de novo protein synthesis and its regulation for successful oocyte maturation and subsequent development. The negative regulation of Wnt signaling has emerged in human, monkey, bovine, and mouse models of oocyte competence. Atrx expression was linked to decreased competence in both oocytes and cumulus cells. Biological networks and transcription factor regulation associated with increased and decreased competence were also identified. These genes could potentially act as biomarkers of oocyte quality or as pharmacological targets for manipulation in order to improve oocyte developmental potential.

Cytogenetic causes of male infertility

Article

Jelena Lissitsina

Infertiilsus esineb 10-15% peredest, olles umbes pooltel paaridel seotud mehepoolsete põhjustega, millest umbes 30% moodustavad geneetilised tegurid eeskätt kromosomaalsed haigused ja geenide mutatsioonid. Käesolev töö näitas Eestis oluliselt kõrgemat (13.4%) kromosoomianomaaliate esinemissagedust infertiilsetel azoo- ja oligozoospermiaga meestel võrreldes fertiilsete meestega. Peamiseks aberratsiooniks azoospermiaga meestel oli lisa-X kromosoom (Klinefelteri sündroom, 47,XXY), kuid oligozoospermiaga meestel mitmesugust tüüpi translokatsioonid. Polümorfsete kromosoomivariantide esinemissagedus oli infertiilsetel meestel sarnane kontrollgrupiga. Kuigi 9. ja Y kromosoomide mõned variandid esinesid infertiilsetel meestel kontrollgrupist sagedamini, ei olnud erinevused statistiliselt olulised. Kromosoomivariantide osatähtsus infertiilsuse tekkes ei ole veel selge, kuigi mõnel juhul võib variandi (eriti de novo tekkelise) esinemine põhjustada häireid meioosi normaalses kulgemises. Uurimuse tulemusena leidsime kaks uut infertiilsusega seotud kromosoomi regiooni. 1) Ühel patsiendil esines retsiprookne translokatsioon t(7;16) murrukohaga regioonis 16p13.3, kus paiknevad protamiini geenid. See võimaldas meil esmakordselt kromosoomiaberratsiooni kaudu kinnitada arvamust, et protamiini geenide ekspressiooni muutused võivad olla meeste infertiilsuse tekke põhjuseks. 2) Teisel patsiendil esines lisakromosoom - inv dup(22)(q11.1), mis võib olla üheks uueks põhjuseks hüpogonadotroopse hüpogonadismi ja infertiilsuse tekkes tingituna kas otsesest lisakromosoomi mõjust või 22. kromosoomi tsentromeerse/ peritsentromeerse regiooni järjestuse 4-kordsest doosist. Lisaks spermatogeneesi mõjutavatele spetsiifilistele geenidele võib kromosoomianomaaliatega meeste fenotüüpi (k.a. fertiilsust) mõjutada ka interaktsioon teiste geenidega. Uurides karüotüübiga 47,XXY meespatsiente, leidsime, et neil sageli esinev alfa-1 antitrüpsiini madal tase või puudulikkus võib olla üheks lisafaktoritest, mis osalevad infertiilsuse ja azoospermia tekkes. Infertility affects 10-15% of couples whereas in half of these couples male-related causes may be detected. Among the variety of reasons for male infertility, in about 30% of infertile males genetic factors should be considered, including chromosomal abnormalities and gene mutations. The present study showed significantly higher (13.4%) frequency of major chromosomal abnormalities in infertile men with azoo- and oligozoospermia compared with that in fertile men. The most frequent abnormality in azoospermic men was additional X chromosome (Klinefelter´s syndrome, 47,XXY), but in oligozoospermic men translocations between different chromosomes. Incidence of polymorphic chromosomal variants was similar both in infertile men and control group. Although some variants of chromosomes 9 and Y were more frequent in infertile men than in controls, the differences were not statistically significant. The role of chromosomal variants in male infertility is not fully clear, but it may be assumed that in some cases they (especially of de novo origin) disturb normal meiosis. The present study revealed two new chromosomal regions related to infertility. 1) One patient had autosomal reciprocal translocation t(7;16) with breakpoint at the region 16p13.3, where protamine genes have been localized. For the first time by this chromosomal abnormality we could confirm the proposition that alterations in the expression of protamine genes may be one of the causes of male infertility. 2) Another patient had a supernumerary chromosome - inv dup(22)(q11.1). This may be a new cause of hypogonadotropic hypogonadism and infertility either due to the additional chromosome inv dup(22)(q11.1) itself or four-hold dosage of centromeric/pericentromeric region sequence. In addition to the effect of specific genes, involved in spermatogenesis, interaction of other genes can modify the clinical features (including fertility) of patients with chromosomal abnormalities. Studies on 47,XXY patients showed that these men often have deficiency of alpha-1 antitrypsin or its low level. That may be one of the additional factors participating in the pathogenesis of azoospermia and infertility. Väitekirja elektrooniline versioon ei sisalda publikatsioone.

Enhancing the Assessment of Reprogenetic Technologies: The Case of Mitochondrial Replacement

Chapter

Oct 2016

Inmaculada de Melo-Martín

Reprogenetic technologies, which combine the power of reproductive techniques with the tools of genetic science and technology, promise prospective parents a remarkable degree of control to pick and choose the likely characteristics of their offspring. Prominent authors such as Agar, Buchanan, DeGrazia, Green, Harris, Robertson, Savulescu, and Silver have flocked to the banner of reprogenetics. For them, increased reproductive choice and reduced suffering through the elimination of genetic disease and disability are just the first step. They advocate use of these technologies to create beings who enjoy longer and healthier lives, possess greater intellectual capacities, and are capable of more refined emotional experiences. Indeed, Harris and Savulescu take reprogenetic technologies to be so valuable that their use is not only morally permissible but morally obligatory. Rethinking Reprogenetics challenges this mainstream view with a contextualized, gender-attentive philosophical perspective. It shows that one need not be a Luddite, a social conservative, or a religious zealot to be critical of reprogenetics. Pointing out the flawed nature of the arguments put forward by the technologies’ proponents, Rethinking Reprogenetics reveals the problematic nature of the assumptions underpinning current evaluations of these technologies and offers a framework for a more critical and skeptical assessment.

Intraovarian Injection of Lyophilized Platelet-Rich Plasma in Patients with Poor Ovarian Response

Article

Jun 2024

Objective: To demonstrate the effect of intraovarian injection of lyophilized platelet-rich plasma (PRP) in poor ovarian responders. Methods: This retrospective cohort study evaluated data for 262 women who underwent intraovarian injection of lyophilized PRP at two private fertility clinics. Women treated from December 2021 to September 2022 and who met the Bologna criteria were included. Intraovarian injection of lyophilized PRP prepared from 50[Formula: see text]mL of autologous peripheral blood was followed by two centrifugations and activation with 2% CaCl 2 , in which cells were removed with a filter and stored at room temperature. The differences in the follicle-stimulating hormone and antral follicle count (AFC), and the number of retrieved oocytes, mature oocytes, fertilized embryos, cleavage embryos, and blastocysts were measured before and 3 months after a single injection. A biological pregnancy with embryos generated after injection was also examined. Results: Patients’ median age and anti-Mullerian hormone (AMH) levels were 44 years and 0.38[Formula: see text]ng/mL, respectively. The number of retrieved oocytes or mature oocytes and fertilized embryos or cleavage embryos was higher after injection. The number of cleavage embryos in women [Formula: see text] years significantly increased after injection. Of the 262 patients who underwent intraovarian injection of lyophilized PRP, 71 underwent embryo transfers using embryos obtained 3 months after injection, and 14 had biological pregnancies. Seven of these patients were in their 40s. Out of 14, one had a live birth, six are ongoing pregnancies, six had chemical abortions, and one had a miscarriage. Conclusions: Intraovarian injection of lyophilized PRP improved egg retrieval results, especially in patients older than 40 years. Furthermore, this treatment may contribute to biological pregnancy in patient with poor ovarian response but needs to be further investigated in more cases in the future.

Frozen-Thawed Embryo Transfer Using Autologous Oocytes Retrieved from Women Aged ≥45 Years: A Retrospective Study

Article

Aug 2023

Objective: To assess the pregnancy outcomes, including live birth rate and pregnancy loss for women aged 45–49 years at the time of oocyte retrieval, who underwent frozen autologous transfer of embryos. Methods: Among frozen-thawed embryo transfers (ET) performed at our clinic from September 2013 to March 2021, 1,114 cycles of women aged 45–49 years at the time of oocyte retrieval were examined retrospectively. Positive pregnancy test, clinical pregnancy, pregnancy loss, and live birth rates per transfer among each age group were analyzed using one-way analysis of variance or the Fisher’s exact test. Results: The mean age at the time of oocyte retrieval and ET was 45.8 and 46.5 years, respectively. Positive pregnancy test rate was 7.6%, of which 72.9% resulted in pregnancy loss. The clinical pregnancy and live birth rates were 5.1% and 2.0%, respectively, with a live birth rate of 3.0% for women with oocytes retrieved at 45 years of age. Multivariate analysis revealed that younger oocyte retrieval age was associated with biochemical and clinical pregnancy; however, no such association was observed for cases with live birth. The oocyte retrieval age of patients with live births was 45 years in 16 cases, 46 years in six cases, and 47 years in one case. In one case of live birth (46 years old), 21 trisomy was observed in the child; however, congenital abnormalities were not observed in other cases. Chorionic tests were performed on the fetuses in 12 of the 33 clinical abortions, of which 11 showed chromosomal abnormalities. Conclusions: Fertility treatment with autologous oocytes in advanced-age women has a low success rate. Although prognosis is poor above 45 years of age, these results can help in providing information to aid patients in their decisions regarding whether or not to pursue treatment.

Protective effect of rutin on ferroptosis‐induced oxidative stress in aging laying hens through Nrf2/HO‐1 signaling

Article

Nov 2022

Oxidative stress is a major cause of ovarian aging and follicular atresia. There is growing evidence that showed potential roles of rutin in antidiabetic, anti‐inflammatory, antitumor, antibacterial and antioxidant, although it is yet unclear what the underlying mechanism is. Here, we looked into the potential effects of rutin on oxidative stress in the prehierarchical small white follicles (SWFs) from 580‐day‐old (D580) laying chickens. According to the findings, aging D580 layer ferroptosis was much higher than it was for laying hens during the peak period (280‐day‐old, D280). In both naturally aged and d‐gal‐induced chicken SWFs, rutin treatment concurrently boosted cell proliferation and prevented apoptosis. In addition, rutin inhibited the increased ferroptosis in aging hens. Meanwhile, rutin markedly suppressed the elevated ferroptosis and descending antioxidant capacity of D280‐culture‐SWFs from chicken elicited by d‐gal. Rutin's activation of the Nrf2/HO‐1 pathway hastened the SWFs' verbal battle with oxidative damage and reduced ferroptosis. Furthermore, activation of the ferroptosis signal increased the oxidative damage in SWFs. In conclusion, rutin alleviated oxidative stress that was induced by ferroptosis in aging chicken SWFs through Nrf2/HO‐1 pathway. These findings point to a novel mechanism by which rutin protects SWFs from oxidative stress by suppressing ferroptosis, which is presumably a fresh approach to slowing ovarian aging in laying hens.

Ovarian stimulation and oocyte quality

Article

Jan 2022

Dietary tributyrin improves reproductive performance, antioxidant capacity, and ovary function of broiler breeders

Article

Full-text available

Aug 2021
POULTRY SCI

The objective of this experiment was to investigate the influence of dietary tributyrin on reproduction performance and ovary function of broiler breeders with different egg laying rate. 256 AA broiler breeders (48-wk-old) were allocated to four treatment in a 2 × 2 factorial arrangement with the main effects of tributyrin supplementation [0 and 1000 mg/kg tributyrin (TRI)] and two egg laying rate levels [average (AR, 81.01%±0.79%) and low (LR, 70.98%±0.95%)]. The results shown that the LR breeders presented higher egg weight, but lower egg laying rate, qualified egg rate and feed efficiency than the AR breeders (P(laying) < 0.05). Also, the superoxidase dismutase (SOD) activity in magnum was lower while malondialdehyde (MDA) was higher in ovary and magnum of LR breeders than that in the AR breeders (P(laying) < 0.05). Dietary supplementation with tributyrin significantly enhanced egg weight (P(TRI) < 0.05), increased albumen height as well as Haugh unit (HU) in AR breeders (P(interaction) < 0.05), and also had higher total antioxidant capacity (T-AOC) and lower MDA in ovary (P(TRI) < 0.05). The cell apoptosis rate and pro-apoptosis related gene expression (caspase 8, 9 and Bax) in the ovary of LR breeders was higher, while anti-apoptosis related gene (Bcl-2) expression were lower in LR breeders when compared with the AR breeders (P(laying) < 0.05). Dietary supplementation with tributyrin decreased the cell apoptosis rate and down-regulated caspase 9 expression in LR breeders (P(Interaction) < 0.05), up-regulated the Bcl-2 expression in both two breeders (P(TRI) < 0.05). These findings suggest that the breeders with lower egg laying rate also characterized by deteriorate ovary function indicated by lower antioxidant capacity and higher cell apoptosis rate. Dietary supplementation with tributyrin increased egg albumen quality, decreased ovarian pro-apoptosis related gene expression to improve reproductive tract function; and the positive effect on egg albumen quality is more pronounced in average reproductive breeders.

Protecting and Extending Fertility for Females of Wild and Endangered Mammals

Chapter

Apr 2019

Sustaining viable populations of any wildlife species requires a combination of adequate habitat protection and a good understanding of environmental and biological factors (including reproductive mechanisms) that ensure species survival. Thousands of species are under threat of extinction due to habitat loss/degradation, overexploitation, pollution, disease, alien species invasions, and urban sprawl. This has served as incentive for intensive management of animal populations, both ex situ (in captivity) and in situ (living in nature). Assisted reproductive technologies developed for addressing human infertility and enhancing livestock production have shown encouraging promise in a few wildlife species. However, species-specific physiological variations and a lack of fundamental knowledge have limited how these tools can be used to help rapidly re-build endangered species numbers. Despite limitations, there is enormous potential in applying human-related fertility preservation strategies to wild animals, especially approaches that could assist in managing or “rescuing” gametes and gonadal tissues of genetically valuable individuals. Indeed, one of the highest priorities in wildlife ex situ management is sustaining all existing genetic diversity to (1) preserve heterozygosity to avoid inbreeding depression and (2) ensure species integrity and the persistence of genomic adaptability to environmental changes. There are components of the rapidly emerging field of oncofertility in women that are highly compatible with preserving valuable germplasms of individuals or populations of threatened wildlife. Strategies associated with oocyte and ovarian tissue cryopreservation or follicle in vitro culture are especially attractive for protecting and extending fertility for wild females. Given adequate attention and more basic studies, we predict that these approaches could assist in the intensive and practical management of gene diversity in endangered species.

Art in women of advanced fertility age

Article

May 2018
Minerva Ginecol

Over the past four decades, the postponement of childbearing has continued to increase, significantly impacting the age of first-time mothers. Female age is the most significant factor influencing clinical outcome in IVF. Overcome the limits of the age-related decline of fertility is a challenge for fertility experts. In the decrease of fertility in late reproductive age, a lot of factors related with advancing women's age and general health can be considered, however the dominant regulator of this age- dependent loss of fertility is the ovary. The key factors in the ovarian aging are oxidative stress, abnormalities of the meiotic spindle, decrease of function and number of oocyte mitochondria, alteration of Sirtuins and androgen deficiency. The aim of this review is to assess the main biological factors involved in the female reproductive ageing according to the recent literature, focusing on oocyte-dependent ones, as well as the possible therapeutic strategies in ART.

In Patients with Only One or Two Oocytes, Is IVF-ET or ICSI Better?

Chapter

Jan 2015

One of the aims of assisted reproduction technologies (ART) is the recruitment of multiple follicles ensuring the recovery of good-quality oocytes upon controlled ovarian hyperstimulation (COH). In recent years, the number of patients in whom few oocytes are obtained in response to COH is increasing. This phenomenon mainly is probably related to the postponement of childbearing to the fourth decade of life. In this group of patients, multifollicular response to COH remains a challenge, but the optimisation of laboratory strategies may help to maximise their chances of pregnancy. Ovarian response to COH varies widely among patients and is strictly dependent on the size of the ovarian pool of resting follicles, the so-called ovarian reserve [1]. In women with a reduced ovarian reserve, a poor ovarian response results in a low number of retrieved oocytes despite the high dose of gonadotropins administered. Hence, although tests for predicting ovarian reserve are available [2], the parameter that best categorises a woman as a ‘poor responder’ remains the ovarian response itself. The incidence of poor ovarian response (POR) is estimated between 9 and 24 % [3–5]. This value increases with age [3, 5] reaching about 50 % in women over 40 years [6]. Women who respond poorly to COH have pregnancy rates that vary from 7.6 to 17.5 %, while in normal responders, they vary from 25.9 to 36.7 %. Female age plays a distinct role in predicting poor response to COH; in fact, older poor responders have lower pregnancy rates (ranging between 1.5 and 12.7 %) compared with younger poor responders (ranging between 13.0 and 35 %) [2]. A second predicting factor of pregnancy outcome in poor responders is the degree of poor response. A lower number of retrieved oocytes results in fewer embryos to transfer and a lower chance of pregnancy, in addition to the expected negative effect of poor ovarian function on oocyte quality.

Activin Decoy Receptor ActRIIB:Fc Lowers FSH and Therapeutically Restores Oocyte Yield, Prevents Oocyte Chromosome Misalignments and Spindle Aberrations, and Increases Fertility in Midlife Female SAMP8 Mice

Article

Full-text available

Dec 2015
ENDOCRINOLOGY

Women of advanced maternal age (AMA, age ≥35) have increased rates of infertility, miscarriages, and trisomic pregnancies. Collectively these conditions are called "egg infertility." A root cause of egg infertility is increased rates of oocyte aneuploidy with age. AMA women often have elevated endogenous FSH. Female senescence-accelerated mouse-prone-8 (SAMP8) has increased rates of oocyte spindle aberrations, diminished fertility, and rising endogenous FSH with age. We hypothesize that elevated FSH during the oocyte's FSH-responsive growth period is a cause of abnormalities in the meiotic spindle. We report that eggs from SAMP8 mice treated with eCG for the period of oocyte growth have increased chromosome and spindle misalignments. Activin is a molecule that raises FSH, and ActRIIB:Fc is an activin decoy receptor that binds and sequesters activin. We report that ActRIIB:Fc treatment of midlife SAMP8 mice for the duration of oocyte growth lowers FSH, prevents egg chromosome and spindle misalignments, and increases litter sizes. AMA patients can also have poor responsiveness to FSH stimulation. We report that whereas eCG lowers yields of viable oocytes, ActRIIB:Fc increases yields of viable oocytes. ActRIIB:Fc and eCG co-treatment markedly reduces yields of viable oocytes. These data are consistent with the hypothesis that elevated FSH contributes to egg aneuploidy, declining fertility, and poor ovarian response, and that ActRIIB:Fc can prevent egg aneuploidy, increase fertility, and improve ovarian response. Future studies will continue to examine whether ActRIIB:Fc works via FSH and/or other pathways, and whether ActRIIB:Fc can prevent aneuploidy, increase fertility, and improve stimulation responsiveness in AMA women.

Fertilität bei Männern

Article

Dec 2015

Michael Zitzmann

Eine Elternschaft im späteren Lebensalter wird von vielen Menschen als vorteilhaft wahrgenommen, da die Lebensbedingungen in vielerlei Hinsicht häufig stabiler sind. Auch gibt es Männer, die eine zweite Familie gründen möchten. Damit sich die ältere Vaterschaft tatsächlich erfüllt, sind deren biologische Besonderheiten zu beachten.

Effects of maternal ageing on ICSI outcomes and embryo development in relation to oocytes morphological characteristics of birefringent structures

Article

May 2014

The aim of this study was to determine the morphological characteristics of the older reproductive aged women's oocytes and to reveal the influence of these characteristics on intra-cytoplasmic sperm injection (ICSI) outcomes. The oocytes of women older than 35 years of age were evaluated retrospectively. Non-invasive polarization microscopy (PolScope) examinations of mature oocytes were performed by measurement of meiotic spindles’ length, area and retardance and zona pellucida thickness and retardance. Fertilization and conception competence and the correlation with the birefringent structures were assessed. Two hundred and thirteen mature oocytes from 54 women were evaluated with a PolScope. Length of the meiotic spindle was shown to be related to fertilization success of women with advanced maternal age. In conclusion, the PolScope is a useful device used to identify the oocyte quality. Quantitative measurements of meiotic spindle parameters may be valuable for the selection of high-quality oocytes that have the potential for embryo development in the in vitro fertilization (IVF) laboratory of women older than 35 years of age who are mostly poor responders.

Fertilität bei Männern über 40 Jahren

Article

Feb 2014

Michael Zitzmann

Hintergrund Eine Elternschaft im späteren Lebensalter wird von vielen Menschen als vorteilhaft wahrgenommen, da oft in vielerlei Hinsicht stabilere Lebensbedingungen gegeben sind. Auch gibt es Männer, für die eine zweite Familie eine Lebensoption darstellt. Daher rücken auch biologische Aspekte der älteren Vaterschaft in den wissenschaftlichen und klinischen Fokus. Altersabhängige Faktoren mit Einfluss auf die männliche Fertilität Das Alter beeinflusst die männliche Fertilität durch eine Reihe von Faktoren, die in ihrer Gänze nicht komplett verstanden sind. Die Spermienproduktion und -motilität nimmt aufgrund der verfallenden testikulären Feinarchitektur mit zunehmendem Alter ab. Auch nimmt mit dem Alter des Mannes die Fekundität durch weitere Faktoren ab: Ein gestörter Schwangerschaftsverlauf wird oft beobachtet. Einige sehr seltene autosomal-dominante Erkrankungen sind deutlich mit dem väterlichen Alter assoziiert. Epigenetische Effekte Hinzu kommen epigenetische Effekte, die mit neurokognitiven Störungen und möglicherweise sogar metabolischen Dysbalancen vergesellschaftet sind. Solche Effekte können sich, einmal ausgelöst, offensichtlich über mehrere Generationen erstrecken. Dabei wird ein Alter des Mannes über 40 Jahre bereits als biologischer Einflussfaktor angesehen, der möglicherweise jedoch durch ein jüngeres Alter der Partnerin ausgeglichen werden kann – zumindest in Teilaspekten. Eine entsprechende Beratung sollte auf jeden Fall patientenorientiert sein. Statistische Wahrscheinlichkeiten sind gegen individuelle Wünsche abzuwägen.

Effects of maternal aging on localizations and expression levels of DNA methyltransferases and chromosome architecture in in-vivo matured mouse oocytes

Article

Full-text available

Sep 2013

This paper investigated the age-related changes in the expression patterns of maintenance methyltransferase (DNMT1) and de novo methyltransferases (DNMT3a, 3b, 3L) and the chromosome architecture in in-vivo matured mouse oocytes using two-photon laser-scanning microscope. Our results showed that (1) DNMT1 and DNMT3a, 3b, 3L in the oocytes of pubertal mice were located in the cortical region of oocyte cytoplasm. In aging groups, DNMT1 was also located in the cortical region. However, DNMT3a, 3b, 3L had a relatively wider distribution in the oocyte cytoplasm and appeared near the chromosomes. These differences between pubertal and aging groups suggested that aging might affect DNA methylation; (2) the expression of DNMT1, and DNMT3a, 3b in aging groups increased significantly compared to pubertal groups, while, the expression of DNMT3L decreased. These results might be explained by the compensation mechanism among DNMTs, which might be impervious to aging; (3) aging caused increased errors in the distribution and three-dimensional morphology of chromosomes, including the increased total volume and surface area, the high ratio of height to diameter of a circular cylinder enclosing the chromosomes (H/D). Our work provided morphological information for the studies of age-related decline in oocyte qualities.

Effects of age on male fertility

Article

Aug 2013

Michael Zitzmann

Later parenting is considered by many to have advantages, parents-to-be may feel themselves more stable to rear children. In addition, many men start a second family later in life. Thus, paternal age becomes an emerging issue. Aging affects male fertility by a scope of factors, which are not fully understood to date. Generally, the amount of produced sperm cells as well as their motility decreases with age, as testicular histological architecture deteriorates. Decreased fecundity and an increased risk for disturbed pregnancies occur with advancing paternal age. Some rare autosomal dominant pathologies are clearly related to paternal age. Altered patterns of epigenetics/gene expression in aging sperm seem to affect a range of neurocognitive disorders and also metabolic dyshomeostasis across generations. Such effects refer to men older than 40 years and may have impact on socio-economic issues. Nevertheless, councelling of older men seeking paternity should be patient-oriented and weigh statistical probabilities against the right for individual life-planning.

Ceria nanoparticles boost activity of aged murine oocytes

Article

Full-text available

Nov 2012

Oocyte meiotic maturation and viability of follicular granulosa cells in young and old experimental Balb/C and CBA mice in the presence of ceria nanoparticles were studied. Treatment of old Balb/c mice with ceria nanoparticles for three days once a day (at a dose of 45 mg/kg) leads to a positive effect on reproductive system. The number of oocytes in follicles increases and this effect is accompanied by an increase in the number of oocytes at metaphase I and metaphase II. The number of living granulosa cells increases, while percentage of the necrotic and apoptotic ones decreases relative to control group. Data obtained on CBA mice provided additional evidence for positive effect of ceria nanoparticles. Depending on the initial state of the reproductive system and dose of ceria nanoparticles the effectiveness of the treatment will vary. In case of old mice ceria nanoparticles protect ovarian cells against oxidative damage, working as anti-aging agent. The litter size in old mice treated with the CNs increases too.

Is it best to cryopreserve human cumulus-free immature oocytes before or after in vitro maturation?

Article

Jun 2012
CRYOBIOLOGY

Freezing unfertilized oocytes is an option for females without a partner, either to preserve their fertility prior to sterilizing cancer treatment or for social reasons. Our study considered whether it is best to freeze immature human oocytes at the germinal vesicle (GV) stage, prior to in vitro maturation (IVM) or at metaphase-II (M-II), after IVM. Sibling GV-stage oocytes from stimulated ICSI cycles were allocated to freezing either prior to (n=109) or after (n=107) IVM. Cumulus-free oocytes were cryopreserved using a choline-substituted slow-freezing protocol and matured in a defined medium, with analysis of chromatin, microtubules, and microfilaments by three-dimensional imaging. Cryopreserved oocytes were compared with oocytes matured in vitro but never frozen (n=114). Survival was similar between oocytes frozen before or after IVM (69.7% vs. 70.5%). Polar body extrusion after IVM was lower in oocytes frozen at the GV stage versus those matured and then frozen (51.3% vs. 75.7%) or not frozen (75.4%). Stratification by patient age (<36 and ⩾36year) showed no difference in oocyte survival or maturation. Oocytes frozen as GVs showed elevated proportions of spontaneous activation (with or without polar body), an effect augmented by patient age. Spindle and chromosome configurations were disrupted to similar extents in both groups of frozen oocytes, with no further detrimental effect of patient age. The length, width, and volume of bipolar M-II spindles were comparable in all three groups. When frozen as GVs, oocytes exhibited decreased maturation and increased spontaneous activation, suggesting that it is best to freeze oocytes at M-II.

Advanced paternal age and reproductive outcome

Article

Full-text available

Dec 2011
ASIAN J ANDROL

Women have been increasingly delaying the start of motherhood in recent decades. The same trend is seen also for men. The influence of maternal age on fertility, chromosomal anomalies, pregnancy complications, and impaired perinatal and post-natal outcome of offspring, has been thoroughly investigated, and these aspects are clinically applied during fertility and pregestational counseling. Male aging and reproductive outcome has gained relatively less attention. The purpose of this review is to evaluate updated and relevant literature on the effect of paternal age on reproductive outcome.

What should be the first-line treatment for unexplained infertility in women over 40 years of age - ovulation induction and IUI, or IVF?

Article

Dec 2009

The tendency to postpone childbearing in developed countries and the relatively high rate of infertility in older women contribute to an increase in the portion of women aged 40 years and older opting for infertility treatments. The main factor for infertility in this group is oocyte senescence, but since this process does not have a specific diagnosis many of those will be classified as having 'unexplained infertility'. The efficacy of the traditional clinical approach for 'unexplained infertility' in older women is questionable. Reviewing the current literature, clomiphene citrate seems to be inefficient in this group of patients, while delivery rates of gonadotrophins and intrauterine insemination cycles are less than 5%. Although low in absolute terms, IVF is more efficient. The chance for delivery, however, diminishes with each year of age above 40. Therefore, after a short trial of gonadotrophins and intrauterine insemination, women aged 40-41 years should be quickly referred to IVF. At an older age, IVF is the primary treatment option.

Ovary and ovulation: DNA fragmentation of oocytes in aged mice

Article

Full-text available

Jul 1996

To investigate whether female fertility decreases with age due to poor oocyte quality, we examined the presence of DNA fragmentation in ovulated oocytes from young, mature and aged mice. Oocytes from three age groups of female mice (7–8, 20–24 and 40–48 weeks) were retrieved from the ovlducts 15 h after human chorionic gonadotrophin (HCG) injection.. Oocytes from each mouse were incubated in a CO2 Incubator for 0–60 h in human tubal fluid (HTF). After incubation, each oocyte was stained with the terminal deoxynucleotidyl transferase-mediated dUDP nick-end labelling (TUNEL) method. The rate of DNA fragmentation (interpreted as apoptotic changes) was significantly higher for oocytes from aged mice, and the fertilization rate was significantly lower, compared with oocytes from young and mature mice. Our results suggest that DNA fragmentation of oocytes might be one of the reasons for poor oocyte quality and lower fertility in the aged group.

Effect of Treating Induced Mitochondrial Damage on Embryonic Development and Epigenesis

Data

Full-text available

Mar 2005

Germinal vesicle transplantation (GVT) has been proposed as a possible treatment to correct age-related oocyte aneuploidy caused by dysfunctional ooplasm. How healthy ooplasm regu-lates normal meiosis and subsequent development has yet to be elucidated, but impaired mitochondrial metabolism may be at-tributable to incomplete segregation of the oocyte chromo-somes. In the present study, after ooplasmic mitochondrial dam-age by photoirradiating chloromethyl-X-rosamine, examination of the oocyte nuclei's ability to survive after transfer into healthy ooplasts was performed. To assess their fertilizability and poten-tial for development, GVT oocytes were fertilized by intracyto-plasmic sperm injection (ICSI) and transferred to foster mice. Condition of the offspring at birth was assessed, and epigenetic analysis was performed. Photosensitization consistently inhibit-ed oocyte maturation. However, after GVT of photosensitized nuclei into healthy ooplasts, 67.2% were reconstituted, and 76.2% of these matured normally, with an overall rate of 51.2%, much higher than that (6.0%) in the mitochondrially injured oocytes. After ICSI, 65.8% (52/79) of GVT oocytes were fertilized normally, and 21.1% (11/52) eventually reached the blastocyst stage. The transfer of 132 two-cell GVT embryos into the oviducts of pseudopregnant females resulted in 17 appar-ently healthy live offspring. For some key developmental genes, a high level of expression was identified in the GVT and ''res-cue''-derived fetal adnexa. Thus, one can induce in oocyte mi-tochondria a photosensitization-based type of damage, which consistently inhibits GV breakdown, meiotic spindle formation, chromosomal segregation, and polar body extrusion. Germinal vesicle transplanted and rescued oocytes were able to undergo maturation, fertilization, and embryonic cleavage and, ultimate-ly, to develop to term. This approach may provide a model with which to study the age-related ooplasmic dysfunction seen in human oocytes. fertilization, gamete biology, gene regulation, genomic imprint-ing, germinal vesicle transplantation, meiosis, mitochondrial pho-tosensitization, oocyte development, ooplasmic dysfunction

Accelerated disappearance of ovarian follicles in mid-life: Implications for forecasting menopause

Article

Full-text available

Dec 1992

Menopause is triggered by the number of ovarian follicles falling below a threshold number and is irreversible because oogonial stem cells disappear after birth. Since it is the result of programmed disappearance of a limited store of follicles, menopause can be predicted using mathematical models based on total follicle counts at different ages. Our model shows follicle numbers decline bi-exponentially rather than as a simple exponential function of age, as had been assumed, with a first exponential rate parameter of -0.097 and a second of -0.237. The change occurred when numbers had fallen to the critical figure of 25,000 at age 37.5 years. The unexpectedly faster rate of ovarian ageing afterwards lowers the follicle population to 1000 at approximately 51 years, and was adopted as the menopausal threshold because it corresponds to the median age of menopause in the general population. Had the earlier rate persisted menopause would not be expected until 71 years. The impact of step reductions of follicle numbers on the prospective span of menstrual life was predicted by the model. A reduction by 50% before age 30 years resulted in the threshold being reached at 44 years and 0.6 year later for every subsequent year until age 37.5 years after which it is reached at 48 years. A reduction of 90% in childhood before age 14 years could result in menopause as early as 27 years, with increments of 0.6 year per year afterwards until after 37.5 years when it is expected at age 41 years.(ABSTRACT TRUNCATED AT 250 WORDS)

DNA fragmentation of oocytes in aged mice

Article

Full-text available

Aug 1996

To investigate whether female fertility decreases with age due to poor oocyte quality, we examined the presence of DNA fragmentation in ovulated oocytes from young, mature and aged mice. Oocytes from three age groups of female mice (7-8, 20-24 and 40-48 weeks) were retrieved from the oviducts 15 h after human chorionic gonadotrophin (HCG) injection. Oocytes from each mouse were incubated in a CO2 incubator for 0-60 h in human tubal fluid (HTF). After incubation, each oocyte was stained with the terminal deoxynucleotidyl transferase-mediated dUDP nick-end labelling (TUNEL) method. The rate of DNA fragmentation (interpreted as apoptotic changes) was significantly higher for oocytes from aged mice, and the fertilization rate was significantly lower, compared with oocytes from young and mature mice. Our results suggest that DNA fragmentation of oocytes might be one of the reasons for poor oocyte quality and lower fertility in the aged group.

The oxidizing agent tertiary butyl hydroperoxide induces disturbances in spindle organization, c-meiosis, and aneuploidy in mouse oocytes

Article

Full-text available

Jan 1997

It has been recently proposed that a concomitant generation of oxidative stress of oocytes with increasing maternal age may be a major factor responsible for the age-related increase in aneuploid conceptions. As a preliminary step in the testing of this hypothesis, we need to confirm that oxidative stress in itself can induce errors in chromosome segregation. In order to achieve this goal, germinal vesicle (GV)-stage mouse oocytes from unstimulated ICR and (C57BL x CBA) F1 hybrid female mice were matured in vitro for 9 h for metaphase I (MI) oocytes or 16 h for metaphase II (MII) oocytes in the presence of varying concentrations of the oxidizing agent tertiary-butyl hydroperoxide (tBH). MII oocytes from (C57BL x CBA) F1 hybrid mice were fixed and C-banded for karyotyping analysis. MI and MII oocytes from ICR mice were fixed and stained with the DNA-fluorescent probe 4',6-diamidino-2-phenylindole (DAPI) to detect abnormalities in chromosomal distribution. Meiosis I and meiosis II spindles from ICR mice were visualized by confocal immunofluorescence microscopy. Data from these experiments demonstrate that in-vitro exposure of mouse oocytes to tBH during meiosis I reduces the length (pole-to-pole distance) and width (diameter at the equator of the spindle) of meiosis I and meiosis II spindles. This reduction is associated with an increase in the percentage of oocytes showing chromosome scattering and clumping on the MII plate, and of aneuploidy (hyperhaploidy) in MII oocytes. However, tBH at the concentrations used in the present study has only a minimal negative effect on the frequency of meiotic maturation. These results suggest that oxidative stress during meiotic maturation in vitro may induce chromosomal errors that are undetectable in the living oocyte and whose developmental consequences may become manifest after fertilization.

Oxidative stress and protection against reactive oxygen species in the pre-implantation embryo and its surrounding

Article

Full-text available

Mar 2001

Oxidative stress is involved in the aetiology of defective embryo development. Reactive oxygen species (ROS) may originate from embryo metabolism and/or embryo surroundings. Embryo metabolism generates ROS via several enzymatic mechanisms. The relative contribution of each source seems different depending on the species, the stage of development, and the culture conditions. Several exogenous factors and culture conditions can enhance the production of ROS by embryos. ROS can alter most types of cellular molecules, and also induce development block and retardation. Multiple mechanisms of embryo protection against ROS exist, and these have complementary actions. External protection, present in follicular and tubal fluids, mainly comprises non-enzymatic antioxidants such as hypotaurine, taurine and ascorbic acid. Internal protection mainly comprises antioxidant enzymes: superoxide dismutase, glutathione peroxidase and gamma-glutamylcysteine synthetase. Transcripts encoding for these enzymes are present in the oocyte, embryo and oviduct. It may be important that these transcripts are stored during oocyte maturation in order to allow the embryo to acquire the aptitude to develop. It is now common to add antioxidant compounds to culture media. Nevertheless, maintaining the pro-oxidant-antioxidant equilibrium in embryos through such supplementation is a complex problem. Further studies are necessary to limit oxidative stress during embryo culture.

Mitochondrial aggregation patterns and activity in human oocytes and preimplantation embryos

Article

Full-text available

Jun 2001

Mitochondria play a vital role in the metabolism of energy-containing compounds in the oocyte cytoplasm to provide adenosine trisphosphate for fertilization and preimplantation embryo development. In this study, ratiometric confocal microscopy with the mitochondrion-specific membrane potential-sensitive fluorescence dye JC-1 (5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolyl-carbocyanine iodide) was used to measure the activity of mitochondria in human oocytes and developing preimplantation embryos. Mitochondria in oocytes and embryos were characterized by distinct localized aggregation patterns. These patterns however did not determine localized regions of heterogeneity in mitochondrial activity. Mitochondrial activity was analysed during oocyte maturation and after fertilization. The activity of mitochondria in fresh metaphase II oocytes was negatively correlated with maternal age. This trend continued when the activity of developing embryos was analysed. Mitochondrial activity was strongly correlated with the rate of embryo development on day 3 after fertilization, but not on day 2. Partial regression analysis showed that the rate of cleavage of preimplantation embryos was more highly correlated with embryo mitochondrial activity than maternal age. These data suggest that the efficiency of mitochondrial respiration in oocytes and preimplantation embryos is closely correlated with the programmed rate of embryo development, and suggest that maternal age further influences this factor. The loss of mitochondrial activity in oocytes obtained from ageing couples may therefore contribute to lower embryo development and pregnancy rates observed during cycles of IVF.

The variability of female reproductive aging

Article

Full-text available

Nov 2001

The delay in childbearing is an important societal change contributing to an increasing incidence of subfertility. The prevailing concept of female reproductive ageing assumes that the decline of both quantity and quality of the oocyte/follicle pool determines an age-dependent loss of female fertility. There is an apparent discrepancy between the ability to maintain a regular ovulatory cycle pattern and the several years earlier cessation of female fertility. This latter is largely explained by an age-related increase of meiotic non-disjunction leading to chromosomal aneuploidy and early pregnancy loss, such that most embryos from women > or =40 years old are chromosomally abnormal and rarely develop further. The final stage of reproductive ageing-the occurrence of menopause-shows a huge variation between women. Age at last birth in natural fertility populations, which marks the end of female fertility, shows an identically wide variation as age at menopause, but occurs on average 10 years earlier. Given the high heritability for age at menopause, the variation in both age of menopause and last birth are probably under genetic control by the same set of genes. Some of those genes must carry heritable variants which modulate the rate of ovarian ageing and give rise to the wide age variations for the various phases of reproductive ageing.

Antioxidant enzymatic defences in human follicular fluid: Characterization and age-dependent changes

Article

Full-text available

Dec 2003

The aim of this work was to study the antioxidant enzymatic defences in human follicular fluid and investigate their possible changes during reproductive ageing. To this end, we tested the specific activities and protein expression of enzymes involved in reactive oxygen species (ROS) scavenging and in detoxification of ROS byproducts in follicular fluid from young (range 27-32 years, n = 12) and older (range 39-45 years, n = 12) women participating in an IVF programme. Results show that all the tested enzymes [superoxide dismutase (SOD), catalase, glutathione peroxidase, glutathione transferase, glutathione reductase] were significantly expressed in human follicular fluid. However, when the two age groups were compared, we found that follicular fluid from older women exhibited a reduced level of glutathione transferase and catalase activities and a higher level of SOD activity. Immunoblot analysis revealed that ageing was associated with decreased protein expression of GST Pi isoform and did not affect SOD and catalase protein expression. Taken together, these findings indicate that reproductive ageing is accompanied by a change in the antioxidant enzymatic pattern that could impair ROS scavenging efficiency in the follicular environment.

Article

Full-text available

Feb 2004

Age-related decline of fertility in women is the result of the decline in both quantity and quality of the resting ovarian follicle pool. The aim of the present study was to determine whether the decline of follicle quality with age is reflected by ultrastructural changes in the resting follicle pool. Ovarian biopsy specimens were obtained by laparoscopy from seven healthy women aged 25-32 yr (young group) and from 11 healthy women aged 38-45 yr (advanced-age group). A total of 182 resting follicles from the young group were compared with 81 resting follicles from the advanced-age group for signs of age-related changes by transmission-electron microscopy. The ooplasmic fraction of vacuoles was increased (P = 0.02), and the fraction of mitochondria decreased (P = 0.005), in the advanced-age group. Also, the density of the mitochondrial matrix (P < 0.001) and the frequency of dilated smooth endoplasmic reticulum (SER; P = 0.001) and Golgi complex (P = 0.02) were increased with age. The frequencies of ruptured mitochondrial membranes (P = 0.001) and dilated SER (P = 0.003) were increased with age in the granulosa cells. Overall follicle-quality scores, which should reflect atretic changes, were not different for the young and advanced-age groups. In conclusion, in resting follicles, the morphological changes with age are different from the changes seen in quality decline by atresia. The morphological changes with age specifically involved the mitochondria, the SER, and the Golgi complex, and they may be the cause of atresia on initiation of follicular growth because of the substantial increase in metabolic requirements.

Age-associated alteration of gene expression patterns in mouse oocytes

Article

Full-text available

Nov 2004

Decreasing oocyte competence with maternal aging is a major factor in human infertility. To investigate the age-dependent molecular changes in a mouse model, we compared the expression profiles of metaphase II oocytes collected from 5- to 6-week-old mice with those collected from 42- to 45-week-old mice using the NIA 22K 60-mer oligo microarray. Among approximately 11,000 genes whose transcripts were detected in oocytes, about 5% (530) showed statistically significant expression changes, excluding the possibility of global decline in transcript abundance. Consistent with the generally accepted view of aging, the differentially expressed genes included ones involved in mitochondrial function and oxidative stress. However, the expression of other genes involved in chromatin structure, DNA methylation, genome stability and RNA helicases was also altered, suggesting the existence of additional mechanisms for aging. Among the transcripts decreased with aging, we identified and characterized a group of new oocyte-specific genes, members of the human NACHT, leucine-rich repeat and PYD-containing (NALP) gene family. These results have implications for aging research as well as for clinical ooplasmic donation to rejuvenate aging oocytes.

Effect of Treating Induced Mitochondrial Damage on Embryonic Development and Epigenesis

Article

Full-text available

Apr 2005

Germinal vesicle transplantation (GVT) has been proposed as a possible treatment to correct age-related oocyte aneuploidy caused by dysfunctional ooplasm. How healthy ooplasm regulates normal meiosis and subsequent development has yet to be elucidated, but impaired mitochondrial metabolism may be attributable to incomplete segregation of the oocyte chromosomes. In the present study, after ooplasmic mitochondrial damage by photoirradiating chloromethyl-X-rosamine, examination of the oocyte nuclei's ability to survive after transfer into healthy ooplasts was performed. To assess their fertilizability and potential for development, GVT oocytes were fertilized by intracytoplasmic sperm injection (ICSI) and transferred to foster mice. Condition of the offspring at birth was assessed, and epigenetic analysis was performed. Photosensitization consistently inhibited oocyte maturation. However, after GVT of photosensitized nuclei into healthy ooplasts, 67.2% were reconstituted, and 76.2% of these matured normally, with an overall rate of 51.2%, much higher than that (6.0%) in the mitochondrially injured oocytes. After ICSI, 65.8% (52/79) of GVT oocytes were fertilized normally, and 21.1% (11/52) eventually reached the blastocyst stage. The transfer of 132 two-cell GVT embryos into the oviducts of pseudopregnant females resulted in 17 apparently healthy live offspring. For some key developmental genes, a high level of expression was identified in the GVT and "rescue"-derived fetal adnexa. Thus, one can induce in oocyte mitochondria a photosensitization-based type of damage, which consistently inhibits GV breakdown, meiotic spindle formation, chromosomal segregation, and polar body extrusion. Germinal vesicle transplanted and rescued oocytes were able to undergo maturation, fertilization, and embryonic cleavage and, ultimately, to develop to term. This approach may provide a model with which to study the age-related ooplasmic dysfunction seen in human oocytes.

Fertility and ageing

Article

Full-text available

May 2005

The late 20th century trend to delay birth of the first child until the age at which female fecundity or reproductive capacity is lower has increased the incidence of age-related infertility. The trend and its consequences have also stimulated interest in the possible factors in the female and the male that may contribute to the decline in fecundity with age; in the means that exist to predict fecundity; and in the consequences for pregnancy and childbirth. In the female, the number of oocytes decreases with age until the menopause. Oocyte quality also diminishes, due in part to increased aneuploidy because of factors such as changes in spindle integrity. Although older male age affects the likelihood of conception, abnormalities in sperm chromosomes and in some components of the semen analysis are less important than the frequency of intercourse. Age is as accurate as any other predictor of conception with assisted reproductive technology. The decline in fecundity becomes clinically relevant when women reach their mid-30s, when even assisted reproduction treatment cannot compensate for the decline in fecundity associated with delaying attempts at conceiving. Pregnancies among women aged >40 years are associated with more non-severe complications, more premature births, more congenital malformations and more interventions at birth.

Role of oxidative stress in female reproduction. Reprod Biol Endocrinol 3:28

Article

Full-text available

Feb 2005
REPROD BIOL ENDOCRIN

In a healthy body, ROS (reactive oxygen species) and antioxidants remain in balance. When the balance is disrupted towards an overabundance of ROS, oxidative stress (OS) occurs. OS influences the entire reproductive lifespan of a woman and even thereafter (i.e. menopause). OS results from an imbalance between prooxidants (free radical species) and the body's scavenging ability (antioxidants). ROS are a double-edged sword - they serve as key signal molecules in physiological processes but also have a role in pathological processes involving the female reproductive tract. ROS affect multiple physiological processes from oocyte maturation to fertilization, embryo development and pregnancy. It has been suggested that OS modulates the age-related decline in fertility. It plays a role during pregnancy and normal parturition and in initiation of preterm labor. Most ovarian cancers appear in the surface epithelium, and repetitive ovulation has been thought to be a causative factor. Ovulation-induced oxidative base damage and damage to DNA of the ovarian epithelium can be prevented by antioxidants. There is growing literature on the effects of OS in female reproduction with involvement in the pathophysiology of preeclampsia, hydatidiform mole, free radical-induced birth defects and other situations such as abortions. Numerous studies have shown that OS plays a role in the pathophysiology of infertility and assisted fertility. There is some evidence of its role in endometriosis, tubal and peritoneal factor infertility and unexplained infertility. This article reviews the role OS plays in normal cycling ovaries, follicular development and cyclical endometrial changes. It also discusses OS-related female infertility and how it influences the outcomes of assisted reproductive techniques. The review comprehensively explores the literature for evidence of the role of oxidative stress in conditions such as abortions, preeclampsia, hydatidiform mole, fetal embryopathies, preterm labour and preeclampsia and gestational diabetes. The review also addresses the growing literature on the role of nitric oxide species in female reproduction. The involvement of nitric oxide species in regulation of endometrial and ovarian function, etiopathogenesis of endometriosis, and maintenance of uterine quiescence, initiation of labour and ripening of cervix at parturition is discussed. Complex interplay between cytokines and oxidative stress in the etiology of female reproductive disorders is discussed. Oxidant status of the cell modulates angiogenesis, which is critical for follicular growth, corpus luteum formation endometrial differentiation and embryonic growth is also highlighted in the review. Strategies to overcome oxidative stress and enhance fertility, both natural and assisted are delineated. Early interventions being investigated for prevention of preeclampsia are enumerated. Trials investigating combination intervention strategy of vitamin E and vitamin C supplementation in preventing preeclampsia are highlighted. Antioxidants are powerful and there are few trials investigating antioxidant supplementation in female reproduction. However, before clinicians recommend antioxidants, randomized controlled trials with sufficient power are necessary to prove the efficacy of antioxidant supplementation in disorders of female reproduction. Serial measurement of oxidative stress biomarkers in longitudinal studies may help delineate the etiology of some of the diosorders in female reproduction such as preeclampsia.

Age-dependent changes in the expression of superoxide dismutases and catalase are associated with ultrastructural modifications in human granulosa cells

Article

Full-text available

Nov 2006

Limited knowledge exists about changes in follicle quality associated with age. The aim of this work was to investigate whether ageing may cause oxidative stress-mediated alterations in human granulosa cells (GCs) from periovulatory follicles. GCs employed in this study were obtained from follicular aspirates of 20 younger women (range 27-32 years) and 20 older women (range 38-41 years) undergoing an IVF treatment. Results obtained from comparative RT-PCR analysis revealed that the mean relative levels of mRNAs coding for superoxide dismutases, Cu, ZnSOD (SOD1), MnSOD (SOD2) and catalase were significantly decreased in women > or =38 years (P < 0.05, Student's t-test). These changes were associated with a reduced expression of SOD1, SOD2 and catalase at the protein level. When examined at an ultrastructural level, most of the GCs from this group showed defective mitochondria and fewer lipid droplets than those observed in the younger group. These results indicate that GCs from older patients suffer from age-dependent oxidative stress injury and are taken as an evidence for reduced defence against reactive oxygen species (ROS) in GCs during reproductive ageing.

The Role of Mitochondrial Function in the Oocyte and Embryo

Article

Full-text available

Feb 2007
Curr Top Dev Biol

Mitochondria have long been known to be the powerhouses of the cell but they also contribute to redox and Ca2+ homeostasis, provide intermediary metabolites and store proapoptotic factors. Mitochondria have a unique behavior during development. They are maternally transmitted with little (if any) paternal contribution, and they originate from a restricted founder population, which is amplified during oogenesis. Then, having established the full complement of mitochondria in the fully grown oocyte, there is no further increase of the mitochondrial population during early development. The localization of mitochondria in the egg during maturation and their segregation to blastomeres in the cleaving embryo are strictly regulated. Gradients in the distribution of mitochondria present in the egg have the potential to give rise to blastomeres receiving different numbers of mitochondria. Such maternally inherited differences in mitochondrial distribution are thought to play roles in defining the long-term viability of the blastomere in some cases and embryonic axes and patterning in others. Mitochondria may also regulate development by a number of other means, including modulating Ca2+ signaling, and the production of ATP, reactive oxygen species, and intermediary metabolites. If the participation of mitochondria in the regulation of sperm-triggered Ca2+ oscillations is now well established, the role of other properties of mitochondrial function during development remain largely unexplored probably due to the difficulty of accessing the mitochondrial compartment in an embryo. Maintaining a functional complement of maternally derived mitochondria is vital for the early embryo. Mitochondrial dysfunction may not only compromise developmental processes but also trigger apoptosis in the embryo. This dual role for mitochondria (to maintain life or to commit to cell death) may well represent a quality control system in the early embryo that will determine whether the embryo proceeds further into development or is quickly eliminated.

Assisted reproductive technology surveillance--United States, 2005

Article

Full-text available

Jul 2008

Assisted reproductive technology (ART) includes fertility treatments in which both eggs and sperm are handled in the laboratory (i.e., in vitro fertilization and related procedures). Patients who undergo ART procedures are more likely to deliver multiple-birth infants than women who conceive naturally. Multiple births are associated with increased risk for mothers and infants (e.g., pregnancy complications, premature delivery, low-birthweight infants, and long-term disability among infants). This report presents the most recent national data and state-specific results. 2005. In 1996, CDC initiated data collection regarding ART procedures performed in the United States, as mandated by the Fertility Clinic Success Rate and Certification Act of 1992 (FCSRCA) (Public Law 102-493 [October 24, 1992]). Beginning with 2004, CDC has contracted with a statistical survey research organization, Westat, Inc., to obtain data from ART medical centers in the United States. Westat, Inc., maintains CDC's web-based data collection system called the National ART Surveillance System (NASS). In 2005, a total of 134,260 ART procedures were reported to CDC. These procedures resulted in 38,910 live-birth deliveries and 52,041 infants. Nationwide, 73% of ART procedures used freshly fertilized embryos from the patient's eggs, 15% used thawed embryos from the patient's eggs, 8% used freshly fertilized embryos from donor eggs, and 4% used thawed embryos from donor eggs. Overall, 42% of ART transfer procedures resulted in a pregnancy, and 35% resulted in a live-birth delivery (delivery of one or more live-born infants). The highest live-birth rates were observed among ART procedures that used freshly fertilized embryos from donor eggs (52%). The highest numbers of ART procedures were performed among residents of California (18,655), New York (12,032), Illinois (9,449), New Jersey (9,325), and Massachusetts (8,571). These five states also reported the highest number of live-birth deliveries. Of 52,041 infants born through ART, 49% were born in multiple-birth deliveries. The multiple-birth risk was highest for women who underwent ART transfer procedures that used freshly fertilized embryos from either donor eggs (41%) or their own eggs (32%). Approximately 1% of U.S. infants born in 2005 were conceived through ART. Those infants accounted for 17% of multiple births nationwide. Approximately 9% of ART singletons, 57% of ART twins, and 95% of ART triplets or higher-order multiples were low birthweight. Similarly, 15% of ART singletons, 66% of ART twins, and 97% of ART triplets or higher-order multiples were born preterm. Whether an ART procedure resulted in a pregnancy and live-birth delivery varied according to different patient and treatment factors. ART poses a major risk for multiple births that are associated with adverse maternal and infant outcomes (e.g., preterm delivery, low birthweight, and infant mortality). This risk varied according to the patient's age, the type of ART procedure performed, the number of embryos available for transfer to the uterus, the number actually transferred, and the day of transfer (day 3 or day 5). ART-related multiple births represent a sizable proportion of all multiple births nationwide and in selected states. To minimize the adverse maternal and child health effects that are associated with multiple pregnancies, ongoing efforts to limit the number of embryos transferred in each ART procedure should be continued and strengthened. Adverse maternal and infant outcomes (e.g., low birthweight and preterm delivery) associated with ART treatment choices should be explained fully when counseling patients who are considering ART.

A Preliminary Report on Oocyte Donation Extending Reproductive Potential to Women over 40

Article

Mar 1991

Role of oxidative stress in female reproduction

Article

Jan 2005

Expression of genes encoding antioxidant enzymes in human and mouse oocytes during the final stages of maturation

Article

Aug 1999
MOL HUM REPROD

Saoussane El Mouatassim

Association between spindle assembly checkpoint expression and maternal age in human oocytes

Article

Jan 2001
MOL HUM REPROD

Nury M Steuerwald

The spindle assembly checkpoint modulates the timing of anaphase initiation in response to the improper alignment of chromosomes at the metaphase plate. If defects are detected, a signal is transduced to halt further progression of the cell cycle until correct bipolar attachment to the spindle is achieved. The mitotic arrest deficient (MAD2) and budding uninhibited by benomyl (BUB1) genes encode conserved kinetochore-associated proteins believed to be components of the checkpoint regulatory pathway. A failure in this surveillance system could lead to genomic instability that may underlie the increased incidence of aneuploidy in the gametes of older women. To explore this possibility, the concentrations of these transcripts in human oocytes at various stages of maturation were determined by real-time rapid cycle fluorescent reverse transcription–polymerase chain reaction (RT–PCR). The results obtained following quantitative analysis suggest that these messages degrade as oocytes age. Potentially, this may impair checkpoint function in older oocytes and may be a contributing factor in age-related aneuploidy.

Antioxidant enzymatic defences in human follicular fluid: characterization and age-dependent changes

Article

Nov 2003
MOL HUM REPROD

M.C. Carbone

The aim of this work was to study the antioxidant enzymatic defences in human follicular fluid and investigate their possible changes during reproductive ageing. To this end, we tested the specific activities and protein expression of enzymes involved in reactive oxygen species (ROS) scavenging and in detoxification of ROS byproducts in follicular fluid from young (range 27‐32 years, n = 12) and older (range 39‐45 years, n = 12) women participating in an IVF programme. Results show that all the tested enzymes [superoxide dismutase (SOD), catalase, glutathione peroxidase, glutathione transferase, glutathione reductase] were significantly expressed in human follicular fluid. However, when the two age groups were compared, we found that follicular fluid from older women exhibited a reduced level of glutathione transferase and catalase activities and a higher level of SOD activity. Immunoblot analysis revealed that ageing was associated with decreased protein expression of GST Pi isoform and did not affect SOD and catalase protein expression. Taken together, these findings indicate that reproductive ageing is accompanied by a change in the antioxidant enzymatic pattern that could impair ROS scavenging efficiency in the follicular environment.

Fertilization and early embryology: Influence of maternal age on meiotic spindle assembly oocytes from naturally cycling women

Article

Oct 1996

To examine the effects of maternal ageing on the meiotic apparatus, we obtained oocytes from naturally cyding women in two age groups, including younger (aged 20–25 years) and older (aged 40–45 years) women. Using high- resolution confocal microscopy we obtained a detailed picture of the meiotic spindle and chromosome placement during various phases of meiosls. Our data revealed that the meiotic spindle in older women is frequently abnormal, both with regard to chromosome alignment and the micro- tubule matrix that comprise the meiotic spindle. The spindle in 79% of the oocytes from the older group exhibited abnormal tubulin placement and one or more chromosomes were displaced from the metaphase plate during the second meiotic division. In contrast, only 17% of the oocytes from the younger age group exhibited aneuploid conditions. The majority of eggs from this group possessed a well ordered, meiotlc spindle containing chromosomes that were fully aligned within a distinct metaphase plate in the spindle. Chromosome management during meiosis is directed by microtubule assembly within the spindle. These data suggest that the regulatory mechanisms responsible for assembly of the meiotic spindle are significantly altered in older women, leading to the high prevalence of aneuploidy.

Genetics of oocyte ageing

Article

Jan 1998
MATURITAS

Aging of Oocyte, Ovary, and Human Reproduction

Article

Jan 2006
ANN NY ACAD SCI

We review age-related changes in the ovary and their effect on female fertility, with particular emphasis on follicle formation, follicle dynamics, and oocyte quality. The evidence indicates that the developmental processes leading to follicle formation set the rules determining follicle quiescence and growth. This regulatory system is maintained until menopause and is directly affected in at least some models of premature ovarian failure (POF), most strikingly in the Foxl2 mouse knockout, a model of human POF with monogenic etiology (blepharophimosis/ptosis/epicanthus inversus syndrome). Several lines of evidence indicate that if the ovarian germ cell lineage maintains regenerative potential, as recently suggested in the mouse, a role in follicle dynamics for germ stem cells, if any, is likely indirect or secondary. In addition, age-related variations in oocyte quality in animal models suggest that reproductive competence is acquired progressively and might depend on parallel growth and differentiation of follicle cells and stroma. Genomewide analyses of the mouse oocyte transcriptome have begun to be used to systematically investigate the mechanisms of reproductive competence that are altered with aging. Investigative and therapeutic strategies can benefit from considering the role of continuous interactions between follicle cells and oocytes from the beginning of histogenesis to full maturation.

Germline stem cells and neo-oogenesis in the adult human ovary

Article

Jun 2007

It remains unclear whether neo-oogenesis occurs in postnatal ovaries of mammals, based on studies in mice. We thought to test whether adult human ovaries contain germline stem cells (GSCs) and undergo neo-oogenesis. Rather than using genetic manipulation which is unethical in humans, we took the approach of analyzing the expression of meiotic marker genes and genes for germ cell proliferation, which are required for neo-oogenesis, in adult human ovaries covering an age range from 28 to 53 years old, compared to testis and fetal ovaries served as positive controls. We show that active meiosis, neo-oogenesis and GSCs are unlikely to exist in normal, adult, human ovaries. No early meiotic-specific or oogenesis-associated mRNAs for SPO11, PRDM9, SCP1, TERT and NOBOX were detectable in adult human ovaries using RT-PCR, compared to fetal ovary and adult testis controls. These findings are further corroborated by the absence of early meiocytes and proliferating germ cells in adult human ovarian cortex probed with markers for meiosis (SCP3), oogonium (OCT3/4, c-KIT), and cell cycle progression (Ki-67, PCNA), in contrast to fetal ovary controls. If postnatal oogenesis is confirmed in mice, then this species would represent an exception to the rule that neo-oogenesis does not occur in adults.

A Preliminary Report on Oocyte Donation Extending Reproductive Potential to Women over 40

Article

Nov 1990

Fertility in women 40 years of age or older is decreased, and in those with ovarian failure it is thought to be irrevocably lost. The donation of oocytes to young (less than 35 years old) women with ovarian failure has allowed many considered infertile a chance to become pregnant. In these women gonadal hormone replacement results in an endometrium receptive to implantation. It is not known whether the endometrial response to such replacement is decreased in women over the age of 40. To test the efficacy of oocyte donation to older women, we enrolled seven women 40 to 44 years old with ovarian failure in a trial of hormone replacement and embryo transfer, using oocytes obtained from women undergoing ovarian hyperstimulation solely for gamete donation. Seven stimulated cycles in the donors that were synchronized with nine cycles in the recipients resulted in eight embryo transfers. Five viable pregnancies were established, one with twins. A sixth pregnancy ended in miscarriage. Five normal infants were delivered by cesarean section, and one stillborn infant was delivered vaginally. The outcomes were compared with those in women under the age of 40 with ovarian failure who were also participating in our donor-oocyte program and in infertile ovulating women 40 or older who were undergoing standard in vitro fertilization. No significant differences in rates of implantation or ongoing pregnancy were noted in older women as compared with younger women receiving donated embryos. These rates, however, were higher than the rates in the infertile ovulating women of similar age who were undergoing standard in vitro fertilization. These preliminary results suggest that the endometrium retains its ability to respond to gonadal steroids and provides a receptive environment for embryo implantation and gestation even in older women.

Gene Expression During Oogenesis and Oocyte Development in Mammals

Article

Feb 1985

Rosemary F. Bachvarova

The ultimate goal in the study of gene expression in oocytes is to understand the special characteristics of the genetic program that confer totipotency on the egg. Totipotency includes both the ability to generate differentiated cells of every description among its progeny and the ability to develop as an organized whole organism. What switches and circuits among genes and gene products maintain or renew such potential at each generation? What special cytoplasmic organization underlies the pattern appearing in the early embryo? Obviously, we are a long way from answering this kind of question.

Aetiology of age-associated aneuploidy: A mechanism based on the 'free radical theory of ageing'

Article

Jul 1995

J J Tarín

A general model is put forward to explain the mechanism by which age-associated aneuploidies are produced. This is based on the free radical theory of ageing, which assumes a rise in oxidative stress with age. It is proposed that determination of indicators of oxidative stress in oocytes from various sources could be a first step in the testing of this hypothesis.

Mitochondrial deoxyribonucleic acid deletions in oocytes and reproductive aging in women

Article

Oct 1995
FERTIL STERIL

To determine whether oocytes from women harbor deletions in mitochondrial DNA (mtDNA) and whether deleted mtDNA is more common in oocytes from older women than oocytes from younger women. A polymerase chain reaction (PCR)-based strategy, which depends on deletions approximating otherwise widely separated primers to demonstrate mtDNA deletions in individual oocytes, was used. Yale In Vitro Fertilization Clinic and Laboratory at Yale University School of Medicine. Primers flanked a region of the mitochondrial genome in which long direct repeated sequence predispose to deletions. The primers identified the 0.5-kb "common" deletion. Deleted mtDNA was represented by a 0.5-kb band when primers separated by 5 kb were used. Control reactions used primers that amplify mtDNA outside the deletion hotspot. Positive controls included brain and/or muscle from aged individuals, and negative controls included fetal tissue and DNA-free blanks. Nested primers confirmed the specificity of the deleted product. Unfertilized oocytes, muscle, and brain tissue contained PCR products consistent with deleted mtDNA. Fetal tissue lacked the mtDNA deletion product. Deleted mtDNA was detected in single oocytes. Oocytes from older women were more likely to contain deleted mtDNA than oocytes from younger women. Deleted mtDNA in unfertilized oocytes may serve as a marker of oocyte senescence.

Influence of maternal age on meiotic spindle assembly in oocytes from naturally cycling women

Article

Nov 1996

To examine the effects of maternal ageing on the meiotic apparatus, we obtained oocytes from naturally cycling women in two age groups, including younger (aged 20-25 years) and older (aged 40-45 years) women. Using high-resolution confocal microscopy we obtained a detailed picture of the meiotic spindle and chromosome placement during various phases of meiosis. Our data revealed that the meiotic spindle in older women is frequently abnormal, both with regard to chromosome alignment and the microtubule matrix that comprise the meiotic spindle. The spindle in 79% of the oocytes from the older group exhibited abnormal tubulin placement and one or more chromosomes were displaced from the metaphase plate during the second meiotic division. In contrast, only 17% of the oocytes from the younger age group exhibited aneuploid conditions. The majority of eggs from this group possessed a well ordered, meiotic spindle containing chromosomes that were fully aligned within a distinct metaphase plate in the spindle. Chromosome management during meiosis is directed by microtubule assembly within the spindle. These data suggest that the regulatory mechanisms responsible for assembly of the meiotic spindle are significantly altered in older women, leading to the high prevalence of aneuploidy.

Genetics of oocyte aging

Article

Nov 1998
MATURITAS

Ursula Eichenlaub-Ritter

Correlations between parental age, aneuploidy in germ cells and recent findings on aetiological factors in mammalian trisomy formation are reviewed. Data from observations in human oocytes, molecular studies on the origin of extra chromosomes in trisomies, experiments in a mouse model system, and transgenic approaches are shown. Errors in chromosome segregation are most frequent in meiosis I of oogenesis in mammals and predominantly predispose specific chromosomes and susceptible chiasmate configurations to maternal age-related nondisjunction. Studies on spindle structure, cell cycle and chromosome behaviour in oocytes of the CBA/Ca mouse used as a model for the maternal age-effect suggest that hormonal homeostasis and size of the follicle pool influence the quality, maturation competence and spindle size of the mammalian oocyte. Predisposition to errors in chromosome segregation are critically dependent on altered cell cycles. Compromised protein synthesis and mitochondrial function affect maturation kinetics and spindle formation, and cause untimely segregation of chromosomes (predivision), mimicking an aged phenotype. Altered cell cycles and untimely resolution of chiasmata but also nondisjunction of late segregating homologues caused by asynchrony in cytoplasmic and nuclear maturation appear to be causal to errors in chromosome segregation with advanced maternal age. Oocytes appear to lack checkpoints guarding against untimely chromosome segregation. Genes and exposures affecting pool size, hormonal homeostasis and interactions between oocytes and their somatic compartment and thus quality of follicles and oocytes have the potential to critically influence chromosome distribution in female meiosis and affect fertility in humans and other mammals.

Intrafollicular influences on human oocyte developmental competence: Perifollicular vascularity, oocyte metabolism and mitochondrial function

Article

Aug 2000

Jonathan van Blerkom

While genetic and epigenetic factors have been associated with the developmental competence of human oocytes and embryos produced by IVF, how such factors develop or influence the oocyte remain to be explained. This paper reviews evidence which suggests that the degree of perifollicular vascular expansion associated with increased rates of blood flow are developmentally important for the generation of a normal follicle and competent oocyte. The degree of vascular development is follicle specific and differences between follicles might reflect their unique abilities to regulate angiogenic growth factor(s) production by the follicle cells in response to hypoxia. The notion that mitochondrial function in oocytes and early embryos could be influenced by intrafollicular conditions, and that differential patterns of mitochondrial segregation occur in blastomeres during early cleavage, is discussed with respect to the role of these organelles as critical determinants of developmental competence.

Association Between Spindle Assembly Checkpoint Gene Expression and Maternal Age in Human Oocytes

Article

Feb 2001

The spindle assembly checkpoint modulates the timing of anaphase initiation in response to the improper alignment of chromosomes at the metaphase plate. If defects are detected, a signal is transduced to halt further progression of the cell cycle until correct bipolar attachment to the spindle is achieved. The mitotic arrest deficient (MAD2) and budding uninhibited by benomyl (BUB1) genes encode conserved kinetochore-associated proteins believed to be components of the checkpoint regulatory pathway. A failure in this surveillance system could lead to genomic instability that may underlie the increased incidence of aneuploidy in the gametes of older women. To explore this possibility, the concentrations of these transcripts in human oocytes at various stages of maturation were determined by real-time rapid cycle fluorescent reverse transcription-polymerase chain reaction (RT-PCR). The results obtained following quantitative analysis suggest that these messages degrade as oocytes age. Potentially, this may impair checkpoint function in older oocytes and may be a contributing factor in age-related aneuploidy.

Cellular and Morphological Traits of Oocytes Retrieved from Aging Mice after Exogenous Ovarian Stimulation

Article

Aug 2001

The present study aims to shed light on the origin of abnormal oocytes ovulated by aged females. In order to reach this goal, cellular and morphological traits of ovulated oocytes from hybrid (C57Bl/6JIco female x CBA/JIco male) female mice retrieved after exogenous ovarian stimulation at the age of 12, 40-42, 50-52, or 57-62 wk were analyzed. Aging of female mice was associated with 1) decreased number of ovulated oocytes; 2) increased percentage of cumulus-free oocytes; 3) raised percentage of oocytes with intracellular mitochondrial aggregates; 4) reduced percentage of oocytes displaying a normal distribution of chromosomes in the metaphase-II plate; 5) increased percentage of normal oocytes exhibiting a DNA-containing polar body (PB); 6) higher percentage of oocytes with chromosome scattering; 7) increased percentage of chromosome-scattered oocytes without a DNA-containing PB and with intracytoplasmic mitochondrial aggregates; 8) raised percentage of oocytes exhibiting chromosome decondensation; 9) lower percentage of chromosome-decondensed oocytes lacking both a DNA-containing PB and intracytoplasmic mitochondrial aggregates; 10) increased percentage of abnormal/degenerated oocytes; 11) reduced percentage of abnormal/degenerated oocytes displaying cellular fragmentation; and 12) higher percentage of abnormal/degenerated oocytes with mitochondrial aggregates exhibiting no nuclear/chromosomal DNA fluorescence, cellular fragmentation, milky or dark cytoplasm, or cellular remains enclosed by the zona pellucida. Although several studies suggest aging females may ovulate aged or overripened oocytes, these data support the hypothesis that old females ovulate an increased percentage of atretic/apoptotic oocytes coming from rescued follicles that would have become atretic earlier in life.

Ageing-associated aberration in meiosis of oocytes from senescence-accelerated mice

Article

Nov 2002

The senescence-accelerated mouse (SAM) has been shown to exhibit ageing-associated mitochondrial dysfunction and oxidative stress, and early decline in fertility. We compared meiotic progression of germinal vesicle oocytes between young (2-3 months) and old (10-14 months) SAM mice using triple immunostaining and fluorescence microscopy as well as Pol-Scope imaging. At 8-9 h of in-vitro maturation (IVM), most young SAM oocytes (86%, 32/37) were at meiosis I (MI) stage, with chromosomes aligned in the mid-region of MI spindles, whereas disrupted MI spindles and/or chromosome misalignments (45%, 18/40) and a few oocytes (20%, 8/40) with abnormal MII spindles were found in old SAM oocytes. At 15-17 h of IVM, old SAM oocytes, despite errors at MI stage, extruded a first polar body at an incidence of 88% (n = 85), which did not differ from that (92%, n = 106) of young SAM oocytes. However, oocytes from old SAM (64%, 32/50) showed aberrant MII, with chromosome misalignment and dispersal, in contrast to normal MII in most young SAM oocytes (87%, 65/75), showing chromosome alignment at the metaphase plate of MII spindles. Moreover, Pol-Scope imaging non-invasively detected disrupted or absent visible spindles and possibly aberrant chromosome alignment. Spindle disruption and/or chromosome misalignments at both MI and MII are associated with maternal ageing in the SAM mouse. Our findings also suggest that meiotic division lacks a competent checkpoint for spindle integrity and chromosome alignment during reproductive ageing-associated oocyte senescence.

Ooplasmic transfer: animal models assist human studies

Article

Jul 2002
REPROD BIOMED ONLINE

Ooplasmic transplantation is based on the premise that ooplasmic components are compromised in some individuals. In theory, the transfer of small amounts of healthy ooplasm can correct such deficits, allowing for improved development and implantation. The technique is based on a well-established background of experimental embryology demonstrating that cytoplasmic manipulation in oocytes and early embryos can be entirely compatible with normal development. Cytoplasm has been manipulated via karyoplast and cytoplast transfer and by cytoplasmic injection. Term development has been obtained following such manipulations in a variety of mammalian species. While some manipulative scenarios have exhibited compromised development, others have exhibited improved development. Developmental problems involving specific epigenetic and mitochondrial incompatibilities have been observed in a very limited subset of animal studies. These studies are based on genetic and physical models that have little relation to the actual substance of ooplasmic transplantation in the human. In fact, the majority of animal studies suggest that ooplasmic transplantation is well-founded and unlikely to result in negative developmental consequences. Furthermore, there are considerable physical, physiological and developmental differences between human and rodent eggs and embryos. These differences suggest that potentially negative issues raised by rodent results may not be relevant in the human.

Aging: A Theory Based on Free Radical and Radiation Chemistry

Article

Aug 1956
J Gerontol

Harman DA

Association of female aging with decreased parthenogenetic activation, raised MPF, and MAPKs activities and reduced levels of glutathione S-transferases activity and thiols in mouse oocytes

Article

Jan 2005

This study aims to determine in the mouse whether oocytes from reproductively old females exhibit a different susceptibility to be parthenogenetically activated when compared to oocytes from young females. At the age of 10-12 (young-female group) or 60-62 (old-female group) weeks, hybrid female mice were superovulated using pregnant mare's serum gonadotropin (PMSG) followed by human chorionic gonadotropin (hCG) 48 hr later. After removing the cumulus cells, oocytes were exposed to any of two different activating protocols: (a) 6-min exposure to 8% ethanol; and (b) treatment with 200 microM thimerosal for 15 min followed by 8 mM dithiothreitol (DTT) for 30 min. Oocytes from old female mice displayed (1) lower total percentage of parthenogenetic activation and extrusion of the second polar body after treatment with either thimerosal + DTT or ethanol; (2) higher M-phase-promoting factor (MPF) and mitogen-activated protein kinases (MAPKs) activities; and (3) lower intracytoplasmic levels of glutathione S-transferases (GSTs) activity and thiols than oocytes from young females. These data show that female aging is associated with higher resistance of oocytes to be parthenogenetically activated, higher MPF and MAPKs activities and lower intracytoplasmic levels of GSTs activity and thiols.

Effect of Female Age on Mouse Oocyte Developmental Competence Following Mitochondrial Injury

Article

Sep 2005

Oocytes from aging ovaries contain mitochondria with morphological and genetic flaws. How these flaws relate to phenotypes of oocyte developmental compromise associated with clinical infertility is not well understood. This study was conducted to investigate the role of mitochondria in the developmental compromises observed with female aging using a mouse model of mitochondrial dysfunction. Oocytes obtained from aging (30-40 wk) (C57BL/6J x CBACaH)F1 (B6CBAF1) hybrid female mice were photosensitized with mitochondrial fluorophore rhodamine-123 for variable durations and compared to similarly treated oocytes derived from pubertal mice (4-6 wk). Blastocyst development of normally fertilized oocytes from both age-groups correlated negatively in mathematically unique profiles with irradiation time, with a more sudden decline in development for oocytes from aging mice. Complete inhibition of blastocyst development occurred following a shorter duration of photosensitization for oocytes from aging compared to pubertal animals (60 vs. 90 sec). Prolonged photosensitization resulted in mitochondrial uncoupling and promoted localized generation of reactive oxygen species, mitochondrial permeabilization, and apoptotic phenotypes. Thus, aging oocytes are more developmentally sensitive to mitochondrial damage than pubertal oocytes but undergo similar metabolic and apoptotic responses. These and future findings may encourage further optimization of laboratory-based strategies to minimize mitochondrial injury to oocytes, particularly those from older women, and improve clinical outcomes for women with age-related etiologies of infertility.

Pellestor F, Anahory T, Hamamah S. Effect of maternal age on the frequency of cytogenetic abnormalities in human oocytes. Cytogenet Genome Res 111: 206-212

Article

Feb 2005
CYTOGENET GENOME RES

The cytogenetic investigation of human oocytes was initiated in the Sixties, and for the last four decades, this field of research has never stopped progressing as new technologies appear. Numerous karyotyping studies and molecular cytogenetic studies have been reported to date, providing a large body of data on the incidence and the distribution of chromosomal abnormalities in human female gametes, but also displaying a great variability in results, which may be essentially attributable to the technical limitations of these in situ methods when applied to human oocytes. Essentially, the most relevant analyses have led to the estimate that 15-20% of human oocytes display chromosome abnormalities, and they have emphasized the implication of both whole chromosome nondisjunction and chromatid separation in the occurrence of aneuploidy in human oocytes. The effect of advanced maternal age on the incidence of aneuploidies has also been investigated in human oocytes. Most previous studies have failed to confirm any relationship between maternal age and aneuploidy frequency in human oocytes, whereas the more recent reports based on large samples of oocytes or polar bodies have provided evidence for a direct correlation between increased aneuploidy frequency and advanced maternal age, and have clarified the contribution of the various types of malsegregation in the maternal age-dependent aneuploidies.

Age-Associated Changes in Mouse Oocytes During Postovulatory In Vitro Culture: Possible Role for Meiotic Kinases and Survival Factor BCL2

Article

Mar 2006

To elucidate molecular mechanisms underlying oocyte senescence, we investigated whether oocytes from female mice of advanced reproductive age exhibit a precocious postovulatory aging that, in turn, may be responsible for the precocious activation of an apoptotic program. During a 9-h in vitro culture, the frequency of oocytes showing MII aberrations, spontaneous activation, and cellular fragmentation increased in old oocytes (P < 0.05), whereas it did not change in the young group. In old oocytes, the activities of MPF (a complex of the cyclin-dependent kinase cdc2 and cyclin B1) and MAPK (mitogen-activated protein kinase) decreased precociously, showing a first drop as early as 3 h after the beginning of in vitro culture (P < 0.05). Immunoblotting and immunocytochemical analysis revealed that, in oocytes of the old group, reduction of BCL2 expression at protein level occurred earlier than in the young group (P < 0.05) and was not associated to the loss of BCL2 transcripts detected by RT-PCR. These changes are followed by an abrupt increase of the rate of TUNEL-positive oocytes after 24 h of culture to a value of 67% +/- 6%. Exposure of young oocytes to 20 microM roscovitine or 20 microM U0126, specific inhibitors of MPF and MAPK, resulted in the decreased percentage of oocytes showing positive immunostaining for BCL2 and in an increased rate of DNA fragmentation. Present results suggest that the developmental competence of oocytes ovulated by aging mice may be negatively influenced by a downregulation of MPF and MAPK activities that in turn induces the activation of a proapoptotic signaling pathway.

SMC1Β-deficient female mice provide evidence that cohesins are a missing link in age-related nondisjunction

Article

Jan 2006

Mitotic chromosome segregation is facilitated by the cohesin complex, which maintains physical connections between sister chromatids until anaphase. Meiotic cell division is considerably more complex, as cohesion must be released sequentially to facilitate orderly segregation of chromosomes at both meiosis I and meiosis II. This necessitates meiosis-specific cohesin components; recent studies in rodents suggest that these influence chromosome behavior during both cell division and meiotic prophase. To elucidate the role of the meiosis-specific cohesin SMC1beta (encoded by Smc1l2) in oogenesis, we carried out meiotic studies of female SMC1beta-deficient mice. Our results provide the first direct evidence that SMC1beta acts as a chiasma binder in mammals, stabilizing sites of exchange until anaphase. Additionally, our observations support the hypothesis that deficient cohesion is an underlying cause of human age-related aneuploidy.

Assisted reproductive technology surveillance—United States, 2003

Article

Jun 2006

In 1996, CDC initiated data collection regarding assisted reproductive technology (ART) procedures performed in the United States, as mandated by the Fertility Clinic Success Rate and Certification Act (FCSRCA) (Public Law 102-493, October 24, 1992). ART includes fertility treatments in which both eggs and sperm are handled in the laboratory (i.e., in vitro fertilization and related procedures). Patients who undergo ART treatments are more likely to deliver multiple-birth infants than women who conceive naturally. Multiple births are associated with increased risk for mothers and infants (e.g., pregnancy complications, premature delivery, low-birthweight infants, and long-term disability among infants). 2003. CDC contracted with the Society for Assisted Reproductive Technology (SART) to obtain data from ART medical centers located in the United States. Since 1997, CDC has compiled data related to ART procedures. In 2003, a total of 122,872 ART procedures were reported to CDC. These procedures resulted in 35,785 live-birth deliveries and 48,756 infants. Nationwide, 74% of ART procedures used freshly fertilized embryos from the patient's eggs; 14% used thawed embryos from the patient's eggs; 8% used freshly fertilized embryos from donor eggs; and 4% used thawed embryos from donor eggs. Overall, 42% of ART transfer procedures resulted in a pregnancy, and 35% resulted in a live-birth delivery (delivery of one or more live-born infants). The highest live-birth rates were observed among ART procedures using freshly fertilized embryos from donor eggs (51%). The highest numbers of ART procedures were performed among residents of California (15,911), New York (15,534), Massachusetts (8,813), Illinois (8,676), and New Jersey (8,299). These five states also reported the highest number of infants conceived through ART. Of 48,756 infants born through ART, 51% were born in multiple-birth deliveries. The multiple-birth risk was highest for women who underwent ART transfer procedures using freshly fertilized embryos from either donor eggs (40%) or their own eggs (34%). Number of embryos transferred, embryo availability (an indicator of embryo quality), and patient's age were also strong predictors of multiple-birth risk. Approximately 1% of U.S. infants born in 2003 were conceived through ART. Those infants accounted for 18% of multiple births nationwide. The percentage of ART infants who were low birthweight ranged from 9% among singletons to 94% among triplets or higher order multiples. The percentage of ART infants born preterm ranged from 15% among singletons to 97% among triplets or higher order multiples. Whether an ART procedure resulted in a pregnancy and live-birth delivery varied according to different patient and treatment factors. ART poses a major risk for multiple births. This risk varied according to the patient's age, the type of ART procedure performed, the number of embryos transferred, and embryo availability (an indicator of embryo quality). ART-related multiple births represent a sizable proportion of all multiple births nationwide and in selected states. Efforts should be made to limit the number of embryos transferred for patients undergoing ART. In addition, adverse infant health outcomes (e.g., low birthweight and preterm delivery) should be considered when assessing the efficacy and safety of ART.

Demographic effects of the introduction of steroid contraception in developed countries

Article

May 2006

Henri Leridon

The use of the contraceptive pill increased very rapidly in the 1970s in many developed countries, and fertility almost simultaneously started to decline. We discuss here the possibility of a causal link between these two major changes. We first provide evidence for a relationship between the spread of oral contraceptive use and the change in fertility in many European countries over the last three or four decades. The situation of specific countries is examined more in depth on the basis of available literature. We then review the various theories attempting to explain these trends and see how the family planning variables are treated in these approaches. At the country level, the conclusion is unambiguous: within individual countries, there is no systematic negative correlation between fertility and contraceptive pill use. The development of hormonal contraception cannot be considered as responsible for either starting or the size of the fertility decline. A more subtle chain of causality must be considered, but there is no agreement on a general theory of fertility changes. Most authors, however, agree that the diffusion of modern contraception has certainly contributed to the reduction in the number of unwanted pregnancies and has also facilitated and favoured the adoption of new (more restrictive) norms for the ideal family size.

Deficit of mitochondria-derived ATP during oxidative stress impairs mouse MII oocyte spindles

Article

Nov 2006
CELL RES

Although the role of oxidative stress in maternal aging and infertility has been suggested, the underlying mechanisms are not fully understood. The present study is designed to determine the relationship between mitochondrial function and spindle stability in metaphase II (MII) oocytes under oxidative stress. MII mouse oocytes were treated with H2O2 in the presence or absence of permeability transition pores (PTPs) blockers cyclosporin A (CsA). In addition, antioxidant N-acetylcysteine (NAC), F0/F1 synthase inhibitor oligomycin A, the mitochondria uncoupler carbonyl cyanide 4-trifluoro-methoxyphenylhydrazone (FCCP) or thapsigargin plus 2.5 mM Ca2+ (Th+2.5 mM Ca2+) were used in mechanistic studies. Morphologic analyses of oocyte spindles and chromosomes were performed and mitochondrial membrane potential (DeltaPsim), cytoplasmic free calcium concentration ([Ca2+]c) and cytoplasmic ATP content within oocytes were also assayed. In a time- and H2O2 dose-dependent manner, disruption of meiotic spindles was found after oocytes were treated with H2O2, which was prevented by pre-treatment with NAC. Administration of H2O2 led to a dissipation of DeltaPsim, an increase in [Ca2+]c and a decrease in cytoplasmic ATP levels. These detrimental responses of oocytes to H2O2 treatment could be blocked by pre-incubation with CsA. Similar to H2O2, both oligomycin A and FCCP dissipated DeltaPsim, decreased cytoplasmic ATP contents and disassembled MII oocyte spindles, while high [Ca2+]c alone had no effects on spindle morphology. In conclusion, the decrease in mitochondria-derived ATP during oxidative stress may cause a disassembly of mouse MII oocyte spindles, presumably due to the opening of the mitochondrial PTPs.

Influence of follicular fluid meiosis-activating sterol on aneuploidy rate and precocious chromatid segregation in aged mouse oocytes

Article

Apr 2007

Follicular fluid meiosis-activating sterol (FF-MAS) protects young oocytes from precocious chromatid separation (predivision). Reduced expression of cohesion and checkpoint proteins and predivision has been hypothesized to occur in age-related aneuploidy in oocytes. To know whether FF-MAS also protects aged oocytes from predivision and from age-related non-disjunction, we analysed chromosome constitution in mouse oocytes matured spontaneously with or without 10 microM FF-MAS and in hypoxanthine (HX)-arrested young and aged oocytes induced to resume maturation by FF-MAS. Messenger RNA for checkpoint protein MAD2 and cohesion protein SMC1beta was compared between oocytes matured with or without FF-MAS. Aged oocytes possessed many bivalents with single distal chiasma at meiosis I. Predivision was especially high in aged oocytes cultured sub-optimally to metaphase II in alpha-minimum essential medium (alpha-MEM). FF-MAS reduced predivision significantly (P < 0.001) but neither reduced non-disjunction nor induced aneuploidy in aged oocytes. Polyploidy was high in FF-MAS-stimulated maturation, in particular in the aged oocytes (P > 0.001). Relative levels of Smc1beta mRNA appeared increased by maturation in FF-MAS, and mitochondrial clustering was restored. Sister chromatids of aged oocytes appear to be highly susceptible to precocious chromatid separation, especially when maturation is under sub-optimal conditions, e.g. in the absence of cumulus and FF-MAS. This may relate to some loss of chromatid cohesion during ageing. FF-MAS protects aged oocytes from predivision during maturation, possibly by supporting Smc1beta expression, thus reducing risks of meiotic errors, but it cannot prevent age-related non-disjunction. Aged oocytes appear prone to loss of co-ordination between nuclear maturation and cytokinesis suggesting age-related relaxed cell cycle control.

Impact of Assisted Reproductive Technologies: A Mitochondrial Perspective of Cytoplasmic Transplantation

Article

Feb 2007
Curr Top Dev Biol

Many of the assisted reproductive techniques associated with maternal aging, disease states, or implantation failure aim to correct poor developmental capacity. These techniques are highly invasive and require the exchange of nuclear or cytoplasmic material from a donor oocyte to compensate for deficiencies inherent in the affected individual. These techniques are based on the assumption that the cytoplasm of the donor oocyte can effectively substitute the necessary component(s) to enable development to proceed. Several studies have attempted to inject cytoplasm from "normal" (young) donors, into aged eggs, again assuming that beneficial components of the cytoplasm are transferred to restore developmental capacity. These invasive assisted reproduction technology (ART) procedures aim to eliminate chromosomal abnormalities, improve the quality of oocytes deficient in some important cytoplasmic factors necessary for maturation and/or subsequent development, and eliminate maternally inherited diseases (particularly mitochondrial myopathies). However, in order to develop such ART, understanding the processes involving mitochondrial DNA replication and transcription is imperative, as asynchrony between mitochondrial and nuclear genomes may cause problems in mitochondrial function, localization, and biogenesis.

Article

Jul 2007
REPROD BIOMED ONLINE

The age-related decline in female fertility has been attributed to a variety of causes including progressive oocyte depletion, meiotic irregularities and mitochondrial dysfunction. However, additional factors could potentially be involved. To explore this possibility, comprehensive analysis of gene expression in human oocytes, discarded following IVF procedures and segregated by age, was undertaken using microarray methods. These findings indicate that the expression of oocyte genes, in a variety of major functional categories including cell cycle regulation, cytoskeletal structure, energy pathways, transcription control, and stress responses, are influenced by maternal age. These results are corroborated by a complementary extensive study using mouse oocytes.

corrigendum: Germline stem cells and follicular renewal in the postnatal mammalian ovary

Article

Apr 2004
NATURE

A basic doctrine of reproductive biology is that most mammalian females lose the capacity for germ-cell renewal during fetal life, such that a fixed reserve of germ cells (oocytes) enclosed within follicles is endowed at birth. Here we show that juvenile and adult mouse ovaries possess mitotically active germ cells that, based on rates of oocyte degeneration (atresia) and clearance, are needed to continuously replenish the follicle pool. Consistent with this, treatment of prepubertal female mice with the mitotic germ-cell toxicant busulphan eliminates the primordial follicle reserve by early adulthood without inducing atresia. Furthermore, we demonstrate cells expressing the meiotic entry marker synaptonemal complex protein 3 in juvenile and adult mouse ovaries. Wild-type ovaries grafted into transgenic female mice with ubiquitous expression of green fluorescent protein (GFP) become infiltrated with GFP-positive germ cells that form follicles. Collectively, these data establish the existence of proliferative germ cells that sustain oocyte and follicle production in the postnatal mammalian ovary.

Oocyte senescence: A firm link to age-related female subfertility

Abstract

No full-text available

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