Healthy Oocytes Research Articles

An FSH-lowering activin disrupting therapy increases yield of healthy oocytes in a mouse model of female midlife reproductive aging

An FSH-lowering activin disrupting therapy increases yield of healthy oocytes in a mouse model of female midlife reproductive aging

Regulators of ovarian preantral follicle development.

Preantral follicle development has become an increasingly recognized area of study in the last two decades. Factors that regulate the growth survival and differentiation of these small, yet complex structures have been identified. The field of fertility preservation and a need for increased numbers of mature oocytes for stem cell research revealed how little we knew of how follicles got from the primordial stage to the antral stage with a healthy and competent oocyte inside. This work discusses the role of gonadotropins in regulating preantral follicles and also the role of the TGF-β family members and their associated Smad signaling molecules in preantral follicle development. Preantral follicle development is a necessary step to fertility in females and further understanding of this process is essential for progress in both infertility care and the enlarging field of in vitro folliculogenesis.

MicroRNA in Ovarian Biology and Disease.

MicroRNAs (miRNAs) are posttranscriptional gene regulatory molecules that show regulated expression within ovarian tissue. Most research investigating miRNAs in the ovary has relied exclusively on in vitro analyses. In this review, we highlight those few studies in which investigators have illustrated an in vivo effect of miRNAs on ovarian function. We also provide a synopsis of how these small noncoding RNAs can impact ovarian disease. miRNAs have great potential as novel diagnostic biomarkers for the detection of ovarian disease and in the assisted reproductive technologies (ART) for selection of healthy viable oocytes and embryos.

Direct actions of androgens on the survival, growth and secretion of steroids and anti-Müllerian hormone by individual macaque follicles during three-dimensional culture.

What are the direct effects of androgens on primate follicular development and function at specific stages of folliculogenesis? Androgen addition altered primate follicle survival, growth, steroid and anti-Müllerian hormone (AMH) production, and oocyte quality in vitro, in a dose- and stage-dependent manner. Androgens have local actions in the ovary, particularly in the developing follicles. It is hypothesized that androgen promotes early follicular growth, but becomes detrimental to the antral follicles in primates. In vitro follicle maturation was performed using rhesus macaques. Secondary (125-225 µm) follicles were mechanically isolated from 14 pairs of ovaries, encapsulated into alginate (0.25% w/v), and cultured for 40 days. Individual follicles were cultured in a 5% O2 environment, in alpha minimum essential medium supplemented with recombinant human FSH. Follicles were randomly assigned to experiments of steroid ablation by trilostane (TRL), testosterone (T) replacement and dihydrotestosterone (DHT) replacement. Follicle survival and growth were assessed. Follicles with diameters ≥500 μm at Week 5 were categorized as fast-grow follicles. Pregnenolone (P5), progesterone (P4), estradiol (E2) and AMH concentrations in media were measured. Meiotic maturation and fertilization of oocytes from recombinant human chorionic gonadotrophin-treated follicles were assessed at the end of culture. Compared with controls, TRL exposure reduced (P < 0.05) follicle survival, antrum formation rate and follicle diameters at Week 5. While P5 concentrations increased (P < 0.05) following TRL treatment, P4 levels decreased (P < 0.05) in fast-grow follicles at Week 5. Few healthy oocytes were retrieved from antral follicles developed in the presence of TRL. T replacement with TRL increased (P < 0.05) follicle survival and antrum formation at Week 5, compared with TRL alone, to levels comparable to controls. However, high-dose T with TRL decreased (P < 0.05) diameters of fast-grow follicles. Although P4 concentrations produced by fast-grow follicles were not altered by T in the presence of TRL, there was a dose-dependent increase (P < 0.05) in E2 levels at Week 5. High-dose T with TRL decreased (P < 0.05) AMH production by fast-grow follicles at Week 3. More healthy oocytes were retrieved from antral follicles developed in TRL+T compared with TRL alone. DHT had the similar effects to those of high-dose T, except that DHT replacement decreased (P < 0.05) E2 concentrations produced by fast-grow follicles at Week 5 regardless of TRL treatment. This study reports T and DHT actions on in vitro-developed individual primate (macaque) follicles, which are limited to the interval from the secondary to small antral stage. The above findings provide novel information on the role(s) of androgens in primate follicular development and oocyte maturation. We hypothesize that androgens promote pre-antral follicle development, but inhibit antral follicle growth and function in primates. While androgens can act positively, excess levels of androgens may have negative impacts on primate folliculogenesis. NIH U54 RR024347/RL1HD058294/PL1EB008542 (Oncofertility Consortium), NIH U54 HD071836 (SCCPIR), NIH ORWH/NICHD 2K12HD043488 (BIRCWH), NIH FIC TW/HD-00668, ONPRC 8P51OD011092. There are no conflicts of interest.

Conflicts of Interest and Effective Oversight of Assisted Reproduction Using Donated Oocytes.

Oocyte donation raises conflicts of interest and commitment for physicians but little attention has been paid to how to reduce these conflicts in practice. Yet the growing popularity of assisted reproduction has increased the stakes of maintaining an adequate oocyte supply and (where appropriate) minimizing conflicts. A growing body of professional guidelines, legal challenges to professional self-regulation, and empirical research on the practice of oocyte donation all call for renewed attention to the issue. As empirical findings better inform existing conflicts and their potential harms, we can better attempt to reduce these conflicts. To that end, the article first describes the nature of conflicts in oocyte donation and relevant regulations and professional guidelines. We then describe studies on conflicts at four phases of oocyte donation: recruitment, screening, stimulation, and post-stimulation monitoring. Next we consider three models for conflict reduction in medicine generally: improved professional self-regulation, outright restriction like Stark anti-referral laws, or the use of conflict mediators, like in living organ donation. We ultimately conclude that improved professional self-regulation is a reasonable starting place for oocyte donation.

Oocyte glycoproteins regulate the form and function of the follicle basal lamina and theca cells

Maintaining follicle integrity during development, whereby each follicle is a functional unit containing a single oocyte, is essential for the generation of healthy oocytes. However, the mechanisms that regulate this critical function have not been determined. In this paper we investigate the role of the oocyte in maintaining follicle development. To investigate this role, we use a mouse model with oocyte-specific deletion of C1galt1 which is required for the generation of core 1-derived O-glycans. The loss of oocyte-generated O-glycans results in the joining of follicles and the generation of Multiple-Oocyte Follicles (MOFs). The aim was to determine how Mutant follicle development is modified thus enabling follicles to join. Extracellular matrix and follicle permeability were studied using histology, immunohistochemistry and electron microscopy (EM). In ovaries containing Mutant Oocytes, the Follicle basal lamina (FBL) is altered both functionally and structurally from the primary stage onwards with Mutant follicles possessing unexpectedly thicker FBL. In Mutant ovaries, the theca cell layer is also modified with intermingling of theca between adjacent follicles. MOF function was analysed but despite increased numbers of preantral MOFs in Mutants, these do not reach the preovulatory stage after gonadotrophin stimulation. We propose a model describing how oocyte initiated changes in FBL and theca cells result in follicles joining. These data reveal new and important roles for the oocyte in follicle development and follicle integrity.

Inhibition of zygotic DNA repair: transcriptome analysis of the offspring in trout (Oncorhynchus mykiss).

Zygotic repair of the paternal genome is a key event after fertilization. Spermatozoa accumulate DNA strand breaks during spermatogenesis and can suffer additional damage by different factors, including cryopreservation. Fertilization with DNA-damaged spermatozoa (DDS) is considered to promote implantation failures and abortions, but also long-term effects on the progeny that could be related with a defective repair. Base excision repair (BER) pathway is considered the most active in zygotic DNA repair, but healthy oocytes contain enzymes for all repairing pathways. In this study, the effects of the inhibition of the BER pathway in the zygote were analyzed on the progeny obtained after fertilization with differentially DDS. Massive gene expression (GE; 61 657 unique probes) was analyzed after hatching using microarrays. Trout oocytes are easily fertilized with DDS and the high prolificacy allows live progeny to be obtained even with a high rate of abortions. Nevertheless, the zygotic inhibition of Poly (ADP-ribose) polymerase, upstream of BER pathway, resulted in 810 differentially expressed genes (DEGs) after hatching. DEGs are related with DNA repair, apoptosis, telomere maintenance, or growth and development, revealing a scenario of impaired DNA damage signalization and repair. Downregulation of the apoptotic cascade was noticed, suggesting a selection of embryos tolerant to residual DNA damage during embryo development. Our results reveal changes in the progeny from defective repairing zygotes including higher malformations rate, weight gain, longer telomeres, and lower caspase 3/7 activity, whose long-term consequences should be analyzed in depth.

Timing the window of implantation by nucleolar channel system prevalence matches the accuracy of the endometrial receptivity array

To test if nucleolar channel system (NCS) prevalence matches the accuracy of the endometrial receptivity array (ERA) for identification of the window of endometrial receptivity. Comparative retrospective study, May 2008-May 2012. University-affiliated infertility clinic. Forty-nine healthy oocyte donors, regularly cycling, aged 20-34 years with a body mass index of 19-25 kg/m2. Endometrial biopsies were collected throughout the menstrual cycle. All samples underwent transcriptomic signature identification by ERA testing (performed in a prior study) and quantification of NCS prevalence by using indirect immunofluorescence (performed in the present study). Concordance of ERA results determining the window of implantation with NCS prevalence was statistically analyzed using the kappa index. Based on dating according to the luteinizing hormone surge, specimens were dichotomized into receptive (n=24) and nonreceptive (n=25). The NCS prevalence was expressed as percentage of NCSs per endometrial epithelial cells in each endometrial biopsy. Concordance of ERA and NCS dating vs. luteinizing hormone yielded comparable kappa indices of 0.878 and 0.836, respectively. Direct comparison of ERA and NCS dating resulted in a kappa index of 0.796. Prevalence of NCS identifies the window of endometrial receptivity previously identified by their transcriptomic signature using the ERA.

Heritability and impact of environmental effects during pregnancy on antral follicle count in cattle

Previous studies have documented that ovarian antral follicle count (AFC) is positively correlated with number of healthy follicles and oocytes in ovaries (ovarian reserve), as well as ovarian function and fertility in cattle. However, environmental factors (e.g., nutrition, steroids) during pregnancy in cattle and sheep can reduce AFC in offspring. The role that genetic and environmental factors play in influencing the variability in AFC and, correspondingly, the size of the ovarian reserve, ovarian function, and fertility, are, however, poorly understood. The present study tests the hypothesis that variability in AFC in offspring is influenced not only by genetic merit but also by the dam age and lactation status (lactating cows vs. nonlactating heifers) and milk production during pregnancy. Antral follicle count was assessed by ultrasonography in 445 Irish Holstein-Friesian dairy cows and 522 US Holstein-Friesian dairy heifers. Heritability estimates for AFC (± standard error) were 0.31±0.14 and 0.25±0.13 in dairy cows and heifers, respectively. Association analysis between both genotypic sire data and phenotypic dam data with AFC in their daughters was performed using regression and generalized linear models. Antral follicle count was negatively associated with genetic merit for milk fat concentration. Also, AFC was greater in offspring of dams that were lactating (n=255) compared with nonlactating dams (n=89) during pregnancy and was positively associated with dam milk fat concentration and milk fat-to-protein ratio. In conclusion, AFC in dairy cattle is a moderately heritable genetic trait affected by age or lactation status and milk quality but not by level of dam’s milk production during pregnancy.

Mitochondrial Patterns in Bovine Oocytes with Different Meiotic Competence Related to Their in vitro Maturation

This study was designed to specify chromatin and mitochondrial patterns in bovine oocytes with different meiotic competence in relation to maturation progress, resumption of meiosis, MII onset and completion of maturation. Oocytes with greater or lesser meiotic competence, recovered separately from medium (MF) and small follicles (SF), were categorized according to morphology. Four oocyte categories, healthy and light-atretic MF and healthy and light-atretic SF oocytes were matured and collected at 0, 3, 7, 16 and 24h of maturation. Specific differences in terms of chromatin and mitochondrial patterns were found among the maturing oocyte categories. Resumption of meiosis was accelerated in light-atretic oocytes, as compared with healthy oocytes, regardless of their meiotic competence. More competent oocytes activated mitochondria twice during maturation, before resumption of meiosis and before completion of maturation, while less competent oocytes did it only once, before completion of maturation. Changes in mitochondrial activity differed in light-atretic compared with healthy in both more and less competent oocytes. Healthy meiotically more competent oocytes formed clusters and produced ATP for the whole time of maturation until its completion, while light-atretic more competent oocytes and healthy less competent oocytes reduced these activities earlier, at MII onset. Contrary to these oocyte categories, light-atretic less competent oocytes increased cluster formation significantly before resumption of meiosis. It can be concluded that bovine oocytes with different meiotic competence and health differed in the kinetics of mitochondrial patterns during maturation.

The impact of using the combined oral contraceptive pill for cycle scheduling on gene expression related to endometrial receptivity

Does the combined oral contraceptive pill (COCP) change endometrial gene expression when used for cycle programming? COCP used for scheduling purposes does not have a significant impact on endometrial gene expression related to endometrial receptivity. Controversy exists around COCP pretreatment for IVF cycle programming, as some authors claim that it might be detrimental to the live birth rate. Microarray technology applied to the study of tissue gene expression has previously revealed the behavior of genes related to endometrial receptivity under different conditions. Proof-of-concept study of 10 young healthy oocyte donors undergoing controlled ovarian stimulation (COS) recruited between June 2012 and February 2013. Microarray data were obtained from endometrial biopsies from 10 young healthy oocyte donors undergoing COS with GnRH antagonists and recombinant FSH. In group A (n = 5), COCP pretreatment was used for 12-16 days, and stimulation began after a 5-day pill-free interval. Stimulation in group B (n = 5) was initiated on cycle day 3 after a spontaneous menses. Endometrial biopsies were collected 7 days after triggering with hCG. No individual genes exhibited increased or decreased expression (fold change (FC) >2) in patients with prior COCP treatment (group A) compared with controls (group B). However, the results of the functional analysis showed a total of 11 biological processes that were significantly enriched in group A compared with group B (non-COCP). The Endometrial Receptivity Array (ERA) has only been validated on endometrial samples obtained in natural cycles and after hormonal replacement treatment (HRT). Therefore, it was not possible in this study to classify the endometrial samples as receptive or non-receptive. We used the ERA to focus on 238 genes that are intimately related to endometrial receptivity, thus simplifying the analysis and understanding of the data. Cycle scheduling is common in IVF units and is used to avoid weekend retrievals and/or to distribute evenly the workload for better efficiency. Our failure to detect any relevant changes in the genes related to the window of implantation when cycles were programmed with COCP pretreatment suggests that, despite controversial clinical results in previous studies, the use of COCPs in this way does not affect uterine receptivity adversely. Funding for this study was provided by an unrestricted grant from Merck Sharp & Dohme. C.S. and A.P. are co-inventors (with Patricia Diaz-Gimeno) of the Endometrial Receptivity Array and hold the patent. The other authors have no conflicts of interest to declare. EudraCT registration number is 2011-003250-34.

Plasma Antimullerian Hormone as a Predictor of Ovarian Antral Follicular Population in Bos indicus (Nelore) and Bos taurus (Holstein) Heifers

In Bos taurus cattle, antimullerian hormone (AMH) has been demonstrated to have a high degree of correlation with ovarian antral follicle count and the number of healthy follicles and oocytes. To document the correlation between the plasma concentration of AMH and follicular number in Bos indicus and Bos taurus heifers, Nelore (Bos indicus, n=16) and Holstein heifers (Bos taurus, n=16) had their ovarian follicular waves synchronized. After synchronization, ovarian antral follicular population (AFP) was evaluated three times at 60-day (d) intervals (T-120 d, 120days before plasma AMH determination; T-60 d, 60days before; and T0, at the time of plasma AMH determination). The plasma AMH concentration was positively correlated with the number of ovarian follicles on the day of the follicular wave emergence in Bos indicus (Nelore) and Bos taurus (Holstein) heifers at each evaluation time (p<0.05). The AFP was higher in Bos indicus (Nelore) than in Bos taurus (Holstein) heifers (p<0.05). Similarly, the AMH concentration was higher in Bos indicus (Nelore) than in Bos taurus (Holstein) heifers (p<0.0001). When heifers were classified as to present high or low AFP according to the mean of the AFP within each genetic group, high-AFP heifers presented a greater (p<0.0001) AMH concentration than low-AFP heifers, regardless of the genetic group. In conclusion, the AFP is positively correlated with plasma AMH concentration in both Bos indicus (Nelore) and Bos taurus (Holstein) heifers. Furthermore, Bos indicus (Nelore) heifers presented both greater plasma AMH concentrations and AFP than Bos taurus (Holstein) heifers.

Cloned Microglias with Novel Delivery System in Multiple Sclerosis

Multiple Sclerosis (MS) is a chronic inflammatory neurological disease of the Central Nervous System (CNS), characterized by demyelination and activation of microglia. Mitochondrial mutations and dysfunctions in microglial cells are thought to contribute to the detrimental effects of neuroinflammation seen in MS. The Somatic Nuclear Transfer (SCNT) technology offers a more practical mode of therapy in MS, this method would attempt to dilute and/or progressively replace the mutated and activated microglia with cloned Olfactory Ensheathing Cells (OEC) with remyelinating and scavenging properties which would attempt to limit the progression of MS. Applying SCNTderived Embryonic Stem (ES) cells based therapy by cloning Olfactory Ensheathing Cells (OEC), engineered with an autologous nuclear component of the recipient OEC with a healthy donor oocyte. The inner cell mass of the subsequently developed blastocyst would be the source to generate the radial microglia to be used for cell therapy in MS. The novel proposed transcribrial route device offers a painless mode of cell transplantation to the brain. This mode of generating cloned glia and its transplantation to the brain is expected to replace the mutated and activated microglia of the patients with MS and use the regenerative and remyelinating and scavenging properties of the OEC’s, as has been seen in clinical trials in patients with spinal cord injuries. The use of SCNT to develop isogenic ES cellbased therapies for the prevention and treatment of MS associated with mtDNA mutations may open a new avenue of designer’s targeted cell therapy unique for the patients with MS. The proposed “transcribrial device” to access the brain can be an advantageous route of delivery of cloned cells to the brain.

Mitochondrial DNA Mutation in Microglia Can Be Treated by SCNT Cloning and Not by Reprogramming of Olfactory Ensheathing Cells in the Multiple Sclerosis Treatment

Background: Multiple sclerosis (MS) is a demyelinating chronic inflammatory neurological disease of the central nervous system (CNS), characterized by activation of microglia. Mitochondrial mutations (mtDNA.mutat) and dysfunctions in microglial cells are thought to centrally contribute to the damaging effects of neuroinflammation seen in MS. Objective: The somatic nuclear transfer (SCNT) technology offers a more practical mode of therapy in MS, This method would attempt to dilute and/or progressively replace the (mtDNA.mutat) and activated microglia with cloned olfactory ensheathing cells (OEC) with remyelinating and scavenging properties which would attempt to limit the progression of MS. Proposed Methods: Applying SCNT-derived embryonic stem (ES) cells based therapy by cloning olfactory ensheathing cells (OEC), engineered with an autologous nuclear component of the recipient OEC with a healthy donor oocyte. The inner cell mass of the subsequently developed blastocyst would be the source to generate the microglia. The novel proposed transcribrial route device offers a painless mode of cell transplantation to the brain. Expected Results: This mode of generating SCNT cloned glia and its transplantation to the brain is expected to replace the (mtDNA.mutat) and activated microglia of the patients with MS and use the regenerative and remyelinating and properties of the OEC’s, as has been seen in recent clinical trials in paraplegic patients with spinal cord injuries. Conclusion: The use of SCNT in contrast to the reprogramming adult somatic cell is expected to develop isogenic ES cell-based therapies for the prevention and treatment of MS associated with mtDNA mutations that may open a new avenue of designer’s targeted cell therapy unique for the patients with MS. The proposed “transcribrial device” to access the brain can be an advantageous route of delivery of cloned cells to the brain.

Impact of final oocyte maturation using gonadotropin-releasing hormone agonist triggering and different luteal support protocols on endometrial gene expression

To use microarray technology to analyze endometrial gene expression after gonadotropin-releasing hormone agonist (GnRH-a) triggering with four different protocols of luteal support in comparison with results obtained after a human chorionic gonadotropin (hCG) trigger. Prospective, randomized, controlled trial. University-affiliated private assisted-reproduction center. 25 healthy oocyte donors undergoing controlled ovarian stimulation. On day of final oocyte maturation, randomization to [1] GnRH-agonist triggering and luteal support with oral estradiol (2 mg/8 hours) and vaginal progesterone (200 mg/12 hours), [2] GnRH-a and a daily dose of 150 IU of recombinant LH from oocyte pickup, [3] GnRH-a and a single bolus of 60 μg of recombinant hCG on oocyte pickup, [4] GnRH-a and three doses of 20 μg of recombinant hCG separated by 48 hours, or [5] 250 μg of recombinant hCG for trigger and standard luteal support; with endometrial biopsy samples collected 7 days after triggering. Gene expression using the Endometrial Receptivity Array (ERA) and pathway and network analysis of study groups 1-4 compared with controls (group 5). The 56 genes in group 1 (25 up-regulated and 31 down-regulated) exhibited altered expression compared with the 36 genes from group 2 (13 up-regulated and 23 down-regulated), 44 from group 3 (28 up-regulated and 16 down-regulated), and 30 (20 up-regulated and 10 down-regulated) from group4. Differences were seen in endometrial gene expression related to the type of ovulation trigger and luteal support. However, gene expression after the GnRH-a trigger and modified luteal support adding LH/hCG activity more closely resembles the pattern seen in the hCG group. EudraCT 2011-003250-34.

Premature formation of nucleolar channel systems indicates advanced endometrial maturation following controlled ovarian hyperstimulation.

Is there a shift in the timing of nucleolar channel system (NCS) formation following controlled ovarian hyperstimulation (COH)? NCSs appear prematurely following COH compared with natural cycles. During natural cycles, NCSs of endometrial epithelial cell (EEC) nuclei are exclusively present during the window of implantation and are uniformly distributed throughout the upper endometrial cavity. Prospective two-cohort study. Cohorts I and II each consisted of seven volunteers for the duration of three menstrual study cycles that were separated by at least one wash-out or rest cycle, between December 2008 and May 2012. Participants were recruited from a pool of healthy oocyte donors. Consecutive endometrial biopsies were obtained during the same luteal phase on cycle days (CD) 16, 20 and 26 for Cohort I, and on CD14, 22 and 24 for Cohort II, following random assignment to a natural cycle group, a COH cycle group (using a GnRH antagonist), or a COH cycle group receiving luteal phase hormonal supplementation (COH + S). The day of oocyte retrieval was designated CD14 in COH cycles and the day of the LH surge was designated CD13 in natural cycles. Prevalence of NCSs in the nuclei of EECs was quantified using indirect immunofluorescence with an antibody directed against a subset of related nuclear pore complex proteins that are major constituents of NCSs. Progesterone and estradiol levels were measured on the day of each endometrial biopsy. The natural cycle group exhibited peak NCS prevalence on CD20 [53.3%; interquartile range (IQR) 28.5-55.8], which rapidly declined on CD22 (11.8%; IQR 6.3-17.6), CD24 (2.5%; IQR 0.0-9.2) and CD26 (0.3%; IQR 0.0-3.5), and no NCSs on CD14 and 16 defining a short NCS window around CD20. In contrast, in COH and COH + S cycles, NCS prevalence was high already on CD16 (40.4%; IQR 22.6-53.4 and 35.6%; IQR 26.4-44.5, respectively; P = 0.001 compared with CD16 of the natural cycle group, Mann-Whitney), whereas no significant difference in NCS prevalence was detected on any of the other five CDs between the three groups (P > 0.05). The cohort size was small (n = 7) but was offset by the all-or-none presence of NCSs on CD16 in natural versus COH and COH + S cycles and the fact that each subject served as her own control. Premature appearance of NCSs and hence maturation of the endometrium following COH is consistent with previous studies based on histological dating but contradicts studies based on mRNA expression profiling, which reported a lag in endometrial maturation. However, this is the first study of this kind that is based on consecutive endometrial biopsies within the same cycle and that reports such clear-cut differences: no versus robust NCS presence on CD16. Our observation of advanced endometrial maturation following COH may contribute to the reduced implantation rates seen in fresh compared with frozen and donor IVF-embryo transfer cycles. Therefore, the NCS window could serve as a sensitive guide for timing of embryo transfer in frozen and donor cycles. The study was supported by the March of Dimes Birth Defects foundation (1-FY09-363 to U.T.M.); Ferring Pharmaceuticals, Parsippany, NJ; East Coast Fertility, Plainview, NY and the CMBG Training Program (T32 GM007491 to M.J.S.). We report no competing interests.

Functional ovarian reserve assessments in young oocyte donors based on FMR1 genotypes and sub-genotypes

Since ovarian aging patterns are associated with genotypes and sub-genotypes of the fragile X mental retardation 1 (FMR1) gene, we investigated if differences in functional ovarian reserve (FOR) are already apparent at young ages in healthy oocyte donors.

Premature formation of nucleolar channel systems indicates advanced endometrial maturation following controlled ovarian hyperstimulation

To evaluate the temporal appearance of nucleolar channel systems (NCSs) in the human secretory endometrium in natural cycles (Nat) and following ovarian hyperstimulation with (OH+S) and without (OH) vaginal progesterone (P4) (Endometrin; Ferring Pharmaceuticals). Is there a shift in NCS formation following controlled ovarian hyperstimulation (COH)? Prospective two-cohort study. Consecutive endometrial biopsies were obtained from healthy, oocyte donors on cycle days (CD) 16, 20 and 26 (cohort I, n=7), or CDs 14, 22 and 24 (cohort II, n=8) following random assignment to a Nat, OH or OH+S group, with at least one ‘wash-out’ cycle between groups. NCS prevalence in the nuclei of endometrial epithelial cells (EECs) was quantified (#NCSs/#EECs x 100, %) using indirect immunofluorescence with an antibody directed against certain nuclear pore complex proteins, that are major constituents of NCSs. Estradiol and P4 levels were measured on the day of each biopsy. Nat group exhibited peak NCS prevalence on CD20 (53%; interquartile range [IQR] 28.5-55.8), rapid decline on CD22, 24 and 26, and no NCSs on CD14 and 16. In contrast, after OH and OH+S, NCS prevalence was high already on CD16 (40.4%; IQR 22.6-53.4 and 35.6%; IQR 26.4-44.5, respectively; p=0.001, Mann-Whitney). No other significant differences were noted between the groups on any of the other five cycle days (p>0.1). In these healthy oocyte donors, NCS prevalence occurs when expected in the mid-luteal phase of natural cycles. In contrast, premature appearance of NCSs and hence maturation of the endometrium is observed following COH. This is the first study of this kind that is based on consecutive endometrial biopsies within the same cycle and that reports all-or-none data. Our observation of advanced endometrial maturation following COH may contribute to the reduced implantation success rates in fresh vs. frozen IVF-ET cycles.

P73 – constitutively open for business

p73 – constitutively open for business

The Role of Ovarian Surface Epithelium in Folliculogenesis during Fetal Development of the Bovine Ovary: A Histological and Immunohistochemical Study

Although many aspects of ovarian differentiation have been established, comparatively little is known about prenatal follicle formation and differentiation of bovine ovaries. The objective of this investigation was to study the role of the surface epithelium during the development of germ cell nests, germ cell cords and follicle formation in the fetal bovine ovary. Associated important proliferation and apoptotic features were further investigated. Additionally, the expression pattern of the S100 protein was detected. A strong increase of mitotic figures was detected in the surface epithelium, germ cell nests and germ cell cords of ovaries with a crown-rump length (CRL) of 13.0-58.0 cm. Oocytes were positively stained with S100 in bovine ovaries from fetuses with a CRL of 21.0 cm. The staining intensity enhanced parallel to increasing oocyte and follicle sizes during the ovary development. In later stages, a strong staining for S100 was observed in healthy oocytes in contradistinction to atretic oocytes where no expression of the S100 protein could be found. In conclusion, increasing mitosis index of surface epithelium cells, as well as oogonia directly beneath the surface epithelium, in combination with open surface connection during stages from a CRL of 11.0-94.0 cm of bovine fetal ovaries could play an important role in the period of time of ongoing folliculogenesis and derivation of granulosa cells. Additionally, S100-positive oocytes in primordial and later follicle stages joined by a high rate of Ki67-positive index in surrounding granulosa cells indicate that in the oocytes the S100 protein can perhaps be a useful marker for intact oocytes in bovine ovaries.