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PubMed2005

Oocyte control of metabolic cooperativity between oocytes and companion granulosa cells: energy metabolism.

Intercellular communication between oocytes and granulosa cells is essential for normal follicular differentiation and oocyte development. Subtraction hybridization was used to identify genes more highly expressed in cumulus cells than in mural granulosa cells of mouse antral follicles. This screen identified six genes involved in glycolysis: Eno1, Pkm2, Tpi, Aldoa, Ldh1, and Pfkp. When oocytes were microsurgically removed from cumulus cell-oocyte complexes, the isolated cumulus cells exhibited decreased expression levels of genes encoding glycolytic enzymes, glycolysis and activity of the tricarboxylic acid (TCA) cycle. These decreases were prevented by culturing the cumulus cells with paracrine factors secreted by fully grown oocytes. Paracrine factors from fully grown oocytes exhibited greater ability than those from growing oocytes to promote expression of genes encoding glycolytic enzymes and glycolysis in the granulosa cells of preantral follicles. However, neither fully grown nor growing oocytes secreted paracrine factors affecting activity of the TCA cycle. These results indicate that oocytes regulate glycolysis and the TCA cycle in granulosa cells in a manner specific to the population of granulosa cells and to the stage of growth and development of the oocyte. Oocytes control glycolysis in granulosa cells by regulating expression levels of genes encoding glycolytic enzymes. Therefore, mouse oocytes control the intercellular metabolic cooperativity between cumulus cells and oocytes needed for energy production by granulosa cells and required for oocyte and follicular development.

Animals↗
PubMed2006

Comparison of in-vitro outcomes from cryopreserved oocytes and sibling fresh oocytes.

In Italy, the restrictive IVF law generalizes the indication for oocyte freezing for surplus oocytes in 78.5% of in-vitro assisted reproductive cycles. With a view to understanding better what the prospects for intracytoplasmic sperm injection (ICSI) on frozen-thawed oocytes might be, the consequences of freeze-thaw procedures on fertilization, cleavage rates and embryo quality obtained from frozen-thawed oocytes were studied and compared with the results obtained from sibling fresh oocytes. Eleven IVF and 29 ICSI on 76 and 169 fresh oocytes were performed and the corresponding 40 ICSI on 221 sibling frozen-thawed oocytes. There was no difference in terms of fertilization rate between fresh and sibling frozen-thawed oocytes. The cleavage rate (98.0 and 94.4% with fresh oocytes in IVF and ICSI; 77.3% with frozen-thawed oocytes in ICSI; P < 0.001) and embryo quality (grade I embryos over total embryos: 36.7 and 22.2% with fresh oocytes in IVF and ICSI; 12.1% with frozen-thawed oocytes in ICSI; respectively P < 0.001 and P < 0.05) were statistically lower after oocyte cryopreservation. The significant decrease in meiotic spindle retrieval rate before freezing (62.4%) and after thawing procedures (43.4%; P < 0.001) suggests that cryoconservation induces irreversible damage to microtubule repolymerization. The consequences of oocyte cryopreservation procedures on embryo development are reviewed.

Adult↗
PubMed2001

Influence of different stages of the oestrous cycle on cumulus-oocyte communications in canine oocytes: effects on the efficiency of in vitro maturation.

The aim of this study was to investigate the effects of different stages of the oestrous cycle on the communications between dog oocytes and cumulus cells, as well as their possible relationships with the meiotic competence of oocytes matured in vitro. Ovaries were harvested from bitches undergoing anoestrus or late pro-oestrus. Maturation of cumulus-oocyte complexes (COCs) was performed in TCM-199 with 0.3% (w/v) BSA, 0.5 iu FSH ml-1 and 0.5 iu LH ml-1 for 72 h. At the time of collection (0 h), and after 24, 48 or 72 h of culture, some oocytes were stained with Lacmoid to evaluate the nuclear stage of maturation. At 0, 24 or 48 h of in vitro maturation (IVM), oocytes were microinjected (n = 84) with a 3% (w/v) Lucifer yellow solution to evaluate the presence of cumulus-oocyte communications through gap junctions. At t = 0, communications were open in 89% of COCs collected during late pro-oestrus, whereas such communications were not evident in any of the oocytes collected during anoestrus. Communications remained permeable for the first 24 h of maturation in many late pro-oestrous oocytes (67%), whereas no fluorescence diffusion was observed at 48 h of maturation. The presence of cumulus-oocyte communications in late pro-oestrous oocytes was accompanied by their ability to resume meiosis and reach telophase I or metaphase II at higher rates than in oocytes collected during anoestrus (11.1% versus 0%, respectively; P < 0.01). These results indicate that, during anoestrus, there are no permeable gap junctions between oocytes and surrounding cumulus cells at the time of isolation from the ovary. This finding indicates that such oocytes are unsuitable for IVM, as confirmed by their inability to reach telophase I or metaphase II in culture. In contrast, a positive correlation was observed between the presence of permeable gap junctions and meiotic competence in COCs isolated at late pro-oestrus. However, the small proportion of oocytes reaching telophase I and metaphase II indicates that further efforts to define better culture conditions are still required.

Anestrus↗
PubMed1996

Aspects of follicle and oocyte stage that affect in vitro maturation and development of bovine oocytes.

Success of in vitro maturation (IVM) and production of bovine embryos as related to aspects of follicle source and oocyte size were evaluated. First, it was determined that bovine oocytes continue growing in all follicular sizes studied, including >1- to 15-mm follicles. Populations of oocytes were collected from surface visible (peripheral) and cortical follicles from the same ovaries. When the number of oocytes from both peripheral and cortical follicles was combined, the yield of oocytes was approximately double that collected from 1 ovarian site alone. Oocytes from cortical follicles were smaller than those from the surface population, and the smaller cortical oocytes had a lower potential for both meiotic maturation and embryo development Only cortical oocytes with the largest diameters underwent IVM and subsequently developed to blastocysts at rates comparable to oocytes from peripheral follicles. As the diameter of the oocytes recovered from peripheral follicles increased, so did their developmental potential. When the stage of the estrous cycle was observed, it was found to have no effect on developmental potential. Finally, oocytes which extruded polar bodies at an earlier time during maturation were, on average, larger than those which extruded polar bodies later. The results serve a practical purpose in assisting selection of oocytes capable of developing into blastocysts and they give useful correlates of oocyte competencies based on knowledge of follicle source and oocyte stage.

Journal Article↗
PubMed2002

Oocyte and embryo quality after coasting: the experience from oocyte donation.

BACKGROUND: Oocyte donation provides us with an opportunity to study the clinical outcome of oocytes, retrieved from women undergoing coasting, in recipients in whom endometrial receptivity is unaltered by the coasting procedure. Thus, our aim was to describe oocyte donation outcome in donors undergoing coasting, the oocyte and embryo quality obtained from these cycles, and to determine the influence of coasting duration in the cycle outcome. METHODS: Matched-paired analysis included 15 oocyte donors with high response to ovarian stimulation and submitted to coasting and 15 oocyte donors with normal response to ovarian stimulation and not undergoing coasting. There were 38 oocyte recipients who shared oocytes from the donors under coasting and 37 from donors not undergoing coasting. RESULTS: Both groups of donors were comparable in terms of days and dose of ovarian stimulation, oocytes retrieved, metaphase II oocytes obtained, and in the appearance of ovarian hyperstimulation syndrome. Both groups of oocyte recipients were comparable in male-associated factor, pregnancy and implantation rates, as well as in embryo quality. Recipients from donors with coasting for >4 days had significantly lower implantation and pregnancy rates. CONCLUSIONS: the outcome of oocyte donation from donors undergoing coasting is not impaired, as good implantation and pregnancy rates are achieved. Embryo quality, according to our current standards, does not seem to be compromised by coasting itself. However, if coasting in oocyte donors is prolonged for >4 days there is a significant decrease in both implantation and pregnancy rates.

Adult↗
PubMed1985

Inhibition of nuclear maturation in fully grown porcine and mouse oocytes after their fusion with growing porcine oocytes.

Porcine ovarian oocytes, isolated from follicles of 5 mm in diameter (large oocytes), were fused either together or with oocytes isolated from follicles of 0.5 mm in diameter (small oocytes). In giant cells composed of two large oocytes (control) germinal vesicle breakdown (GVBD) occurred and two metaphase I chromosome sets (M I) were observed 24 to 30 h after fusion. By contrast, in giant cells composed of one large and one small porcine oocyte, both germinal vesicles (GVs) remained well conserved after 24-30 h of culture. An identical situation was observed after fusion and cultivation of small porcine and large mouse oocytes isolated from preovulatory follicles. The results demonstrate the presence of inhibiting activity in the ooplasm of small porcine oocytes that prevents nuclear maturation of large porcine and mouse oocytes fused to them. This maturation inhibiting activity can be overcome by preincubating large porcine oocytes for more than 14 h before fusion with small oocytes. During preincubation the ooplasm produces sufficient amount of maturation promoting factor (MPF) to overcome the inhibiting activity present in small porcine oocytes thus inducing GVBD and chromatin condensation both in small and large oocytes.

Animals↗
PubMed2000

Metaphase I arrest and spontaneous parthenogenetic activation of strain LTXBO oocytes: chimeric reaggregated ovaries establish primary lesion in oocytes.

Oocytes of strain LT mice, and related strains such as LTXBO, exhibit a high incidence of arrest in the progression of meiosis at metaphase I (MI) and in spontaneous parthenogenetic activation. Activation of these oocytes within the ovary leads to the formation of ovarian teratomas. In this study, the role of the oocyte's companion granulosa cells, the cumulus cells, was investigated using fully grown oocytes matured in vitro after isolation from LTXBO mice. Results showed that the role of cumulus cells in MI arrest is dichotomous. Cumulus cells temporarily helped to sustain MI arrest, but they also promoted a delayed progression to metaphase II. Cumulus cells also promoted parthenogenetic activation that occurred in association with the delayed progression to metaphase II. Next, the question of whether the lesion(s) promoting MI arrest and spontaneous activation is due to defects in the somatic cells or is intrinsic to the oocyte was addressed using chimeric reaggregated ovaries. An improved method for completely exchanging the germ cell and the somatic cell compartments of ovaries from newborn mice is described. These chimeric reaggregated ovaries, grafted beneath the renal capsule of SCID mice, allowed the complete development of LTXBO oocytes to occur in association with somatic cells from control (B6SJLF(1)) ovaries and development of control oocytes in association with LTXBO somatic cells. Oocyte growth and follicular development appeared generally normal in reaggregated ovaries. High incidences of MI arrest and spontaneous activation of LTXBO oocytes occurred regardless of the genotype of the somatic cells. Moreover, there was a low incidence of MI arrest and spontaneous activation of control oocytes, even though they underwent complete development and maturation associated with LTXBO somatic cells. It is concluded that the phenotypes of MI arrest and parthenogenetic activation in LTXBO oocytes are defects caused by lesions intrinsic to the oocyte. Nevertheless, the oocyte's companion somatic cells play crucial roles in the expression of these lesions.

Animals↗
PubMed2006

Effect of gonadotropins during in vitro maturation of feline oocytes on oocyte-cumulus cells functional coupling and intracellular concentration of glutathione.

Information about the mechanisms of meiotic arrest and resumption of meiosis in feline oocytes is still limited. The aim of this study was to investigate the effect of the presence of gonadotropins during IVM, on meiotic progression in relation to the status of gap junction mediated communications between oocyte and cumulus cells, to the cAMP intracellular content, and to the intra-oocyte concentration of glutathione (GSH) in feline oocytes. Our results indicated that about 50% of cumulus-oocyte complexes (COCs) showed functionally open communications at the time of collection, while the remainder were partially or totally closed. After 3h of culture, the percentage of COCs with functional gap junctions was significantly greater in the group matured in the presence of gonadotropins than in those matured without them, where an interruption of communications was observed. Moreover, this precocious uncoupling was associated with a moderate increase of cAMP concentration in the oocyte, lower than in the group exposed to gonadotropins. Intra-oocyte glutathione levels decreased significantly after 24h of IVM, whether gonadotropins were present or absent during the culturing process. The presence of thiol compounds in the IVM medium induced an intra-oocyte GSH concentration significantly higher than that found in oocytes cultured without these compounds, and similar to the GSH content of immature oocytes. Moreover, the intracellular GSH concentration increased as meiosis progressed. The present study suggests that in feline oocytes, gonadotropins affect the dynamic changes in communications between oocyte and cumulus cells during IVM. However, the intracellular concentration of GSH is not influenced by the gonadotropin stimulation. Moreover, the presence of gonadotropins and thiol compounds results in an increase of GSH levels along with meiotic progression of the oocytes.

Animals↗
PubMed2006

In vivo maturation of oocytes by extending the interval between human chorionic gonadotropin administration and oocyte retrieval.

OBJECTIVE: To find whether the percentage of mature oocytes can be increased in the following ICSI cycle, in a selected group of patients with >or=47% immature oocytes, with the prolongation of hCG-to-oocyte retrieval interval. DESIGN: Randomized study. SETTING: In vitro fertilization unit in a medical center in Israel. PATIENT(S): From January 2003 to June 2005, in 72 of 2,650 intracytoplasmic sperm injection cycles (2.8%), >or=47% of the total aspirated oocytes were immature (group A). In the following treatment cycle, with similar controlled ovarian hyperstimulation, hCG was planned to be injected in the same interval (group B) as well as 3-4 hours earlier (group C). In fact, the hCG-to-oocyte retrieval interval in group C was statistically significantly longer (38.6 +/- 1.2 hours) compared with that in group B (35.3 +/- 0.7 hours). INTERVENTION(S): The clinical characteristics of the patients, ovarian stimulation, hormonal profile, and the intracytoplasmic sperm injection outcome of the various groups were assessed. MAIN OUTCOME MEASURE(S): The number and maturity of the aspirated oocytes. RESULT(S): The mean number of ampules needed for stimulation, treatment duration, serum E(2), and P level on the hCG day were similar in the various groups. Escaped ovulation did not occur in any of the patients. A larger number of aspirated oocytes, a statistically significantly larger percentage of mature oocytes (72%), and a statistically significantly lower percentage of metaphase I (12%), germinal vesicle (18%), and degenerated oocytes (8%) were found in group C compared with group A (50%; 16%, 21%, and 13%, respectively). Similar morphological distribution of the aspirated oocytes was observed among groups B and C. Clinical implantation and pregnancy rates were higher in group C compared with group B. CONCLUSION(S): In a selected group of patients with >or=47% immature oocytes per retrieval, a larger cohort of mature oocytes can be recruited by extending the hCG- to-oocyte retrieval interval in a subsequent cycle. This improved outcome directly affects the quality of embryos available for transfer and thus increases pregnancy rate.

Adult↗
PubMed2003

Development of bovine oocytes reconstructed with a nucleus from growing stage oocytes after fertilization in vitro.

The developmental capacity of reconstructed bovine oocytes that contained nuclei from growing stage oocytes, 70-119 microm in diameter, was assessed after fertilization in vitro. Nuclei from growing stage oocytes of adult ovaries were transferred to enucleated, fully grown germinal vesicle (GV) stage oocytes. After culture in vitro, the reconstructed oocytes matured, forming the first polar body and MII plate. To supply the ability to form pronuclei, the resultant MII plate was transferred to enucleated MII oocytes, which were obtained by in vitro culture of cumulus-oocyte complexes. After fertilization in vitro, 11-15% of the reconstructed oocytes developed to morulae and blastocysts. To assess the ability to develop to term, a total of 27 late morulae and blastocysts were transferred to 19 recipient cows. Of the three cows that subsequently became pregnant, one recipient, who received two embryos derived from reconstructed oocytes with a nucleus from oocytes 100 to 109 microm in diameter, continued the pregnancy to Day 278 of gestation. This pregnancy, however, was unexpectedly a triplet pregnancy that included a set of identical twins and resulted in the premature birth of the calves, followed by death from lack of post-parturient treatment. These results show that bovine oocyte genomes are capable of supporting term development before the oocytes grow to their full size, which suggests that growing stage oocytes can be directly used as a source of maternal genomes.

Animals↗
PubMed1996

Bovine oocyte plasma membrane binding sites for sperm plasma membrane during in vitro oocyte maturation and fertilisation.

The experimental objective was to determine whether the capability of bovine oocyte plasma membrane to bind sperm changes during in vitro oocyte maturation and fertilisation. Binding was quantified by the intensity of tetramethylrhodamine isothiocyanate (TRITC) fluorescence at the periphery of oocytes following incubation with biotinylated sperm plasma membrane proteins and subsequent incubation with TRITC-avidin. Bovine oocytes were matured in vitro. Sample groups were removed after 0,6 and 22 h, or inseminated and further cultured for 24 or 48 h. Oocytes were denuded of cumulus cells and zona pellucida and co-incubated with 56 micrograms biotinylated bovine sperm plasma membrane protein for 45 min in 150 microliters drops of saline-BSA. Controls were incubated for the same time period in the absence of sperm plasma membrane proteins. All oocytes were rinsed, incubated with TRITC-avidin and subsequently fixed and transferred to mounting medium. Oocytes were scanned with a confocal microscope and analysed using ImageQuant software. The binding of sperm plasma membrane was quantified by integrated fluorescent intensity in standardised ellipses spaced around the plasma membrane of the oocyte. Values are expressed as mean intensity units per 320 pixel ellipse. Binding of sperm plasma membrane continued to increase throughout in vitro oocyte maturation and fertilisation (9051, 24318 and 49953 for 0 and 22 h in vitro matured oocytes and fertilised oocytes, respectively; p = 0.0001). A dramatic decrease in sperm plasma membrane binding to the oocyte plasma membrane was observed in 2-cell embryos (mean intensity = 24477, p = 0.0001). The observed binding was primarily due to the binding of sperm plasma membrane proteins, as control oocytes incubated with TRITC-avidin only were barely visible (integrated fluorescence intensity values ranged from 8 to 3757.

Animals↗
PubMed1983

Decline of follicular oocyte maturation inhibitor coincident with maturation and achievement of fertilizability of oocytes recovered at midcycle of gonadotropin-treated women.

To examine whether a decline in follicular oocyte maturation inhibitor (OMI) is associated with attainment of oocyte maturation and fertilizability, OMI was measured in follicular fluid (FF) of 39 follicles of 20 normal women given human menopausal gonadotrophin and human chorionic gonadotrophin to induce follicular growth and maturation. Oocytes were aspirated per laparoscope, the fluid was saved, and the egg was observed, incubated, and inseminated with the husband's sperm. Concepti that developed to the 4- to 8-cell stage were transferred to the uterus and the women were followed for pregnancy. OMI activity in each FF was measured by using cultured cumulus-enclosed porcine oocytes (30-40 oocytes per FF sample). Estrogen, progesterone, oocytes (30-40 oocytes per FF sample). Estrogen, progesterone, and delta 4-androstenedione were measured in FF by radioimmunoassay. The FF of 13 preovulatory follicles yielding oocytes that were mature and fertilizable had significantly less OMI activity (mean +/- SEM) (0.58 +/- 0.10 unit/ml) compared to follicles yielding immature oocytes (2.8 +/- 0.56 units/ml; n = 9), atretic oocytes (5.5 +/- 2.5 units/ml; n = 7), or preovulatory oocytes with fractured zonae (1.9 +/- 0.63 units/ml; n = 7). The estrogen concentration (mean +/- SEM) of preovulatory follicles yielding mature fertilizable eggs or mature eggs with fractured zonae was greater (396 +/- 34 ng/ml; n = 20) compared to follicles yielding immature or atretic eggs (203 +/- 59 ng/ml; n = 9 and 97 +/- 47 ng/ml; n = 7, respectively; P less than 0.05). Progesterone concentration (mean +/- SEM; ng/ml) of FF was generally elevated in all preovulatory follicles (635 +/- 53) compared to immature or atretic follicles (230 +/- 64 and 76 +/- 17, respectively; P less than 0.05). It may be concluded that in normal follicle maturation there is a decline in OMI in the follicle containing an oocyte that becomes mature and fertilizable. There is also an increase in estrogen, progesterone, and follicle size. It is also possible to have an abnormal follicle maturation when there is an increase in size as well as FF, estrogen, and progesterone, but withut a decline in OMI--a situation which can lead to production of a nonfertilizable oocyte.

Adult↗
PubMed1998

Timing of oocyte activation, pronucleus formation and cleavage in humans after intracytoplasmic sperm injection (ICSI) with testicular spermatozoa and after ICSI or in-vitro fertilization on sibling oocytes with ejaculated spermatozoa.

In the first study, we evaluated 101 oocytes [2, 4, 6, 8, 16, 18 and 20 h after intracytoplasmic sperm injection (ICSI)] that had been microinjected with testicular spermatozoa. Of the 70 normally fertilized oocytes (69%) 30 (43%) had two pronuclei by 6 h after ICSI. Fifty-one (73%) by 8 h, 69 (99%) by 16 h and four of them by 20 h cleaved to the 2-cell stage. In the second study, 95 cumulus-corona-oocyte complexes (CCOC) were divided into two groups. Forty-seven CCOC were inseminated by conventional in-vitro fertilization (IVF) and 40 metaphase-II oocytes by ICSI. Oocytes were evaluated at 2, 4, 6 (only after ICSI), 8, 10, 12, 18, 20, 22, 24, 26, 28 and 30 h after both ICSI and IVF. After IVF, 35 oocytes were fertilized normally (75%), four of which (11%) had two pronuclei by 8 h, 11 (31%) by 10 h, 27 (77%) by 12 h and 35 (100%) by 14 h. The first cleavages had occurred by 24 h after insemination (four oocytes, 11%). After ICSI, 34 oocytes were fertilized normally (79%), 13 of which (38%) had two pronuclei by 6 h, 27 (79%) by 8 h and 32 (94%) by 10 h. Three oocytes cleaved by 20 h after microinjection (9%) and 19 by 24 h (56%). Pronuclei developed asynchronously in six oocytes after ICSI (18%) as opposed to 16 oocytes after IVF (46%). The results of this study suggest that the timing of pronuclear formation is no different when a testicular spermatozoon is microinjected into the oocytes from when an ejaculated spermatozoon is injected. Secondly, pronuclear development and first cleavage generally take place 4 h sooner after ICSI than after IVF. On the other hand, a higher proportion of oocytes develop two pronuclei asynchronously after IVF than after ICSI.

Adult↗
PubMed2000

Comparison of oocyte activation and Ca2+ oscillation-inducing abilities of round/elongated spermatids of mouse, hamster, rat, rabbit and human assessed by mouse oocyte activation assay.

Oocyte activation and Ca2+ oscillation-inducing abilities of round spermatid (ROS) and elongated spermatid (ELS) of some rodents and human were assessed by their injection into mouse (B6D2F1) oocytes (mouse test). With mice (B6D2F1, ICR) and rat, ROS displayed no oocyte activation or Ca2+ oscillation-inducing abilities. Although ELS could induce activation at 87, 86 and 31% of injected oocytes respectively, most of the intracellular calcium concentration ([Ca2+]i) responses of ELS-injected oocytes did not show oscillation patterns; only several transient [Ca2+]i rises (transient pattern) were seen. Similarly, with hamster, rabbit and human, while ROS could induce oocyte activation efficiently (70, 71 and 52% respectively), most of the [Ca2+]i patterns of injected oocytes were transient patterns, and not oscillation patterns. When ROS nuclei only from these latter species were injected into mouse oocytes, most of the oocytes could not be activated. [Ca2+]i patterns of oocytes injected with immature sperm cells changed from transient pattern to oscillation pattern while the cells were maturing into spermatozoa. With hamster ROS, oocyte-activating factor was found to be distributed mainly in the cytoplasm. It was interesting that there is a dissociation between the timings of appearance of oocyte activation and that of Ca2+ oscillation of oocytes injected with developing immature sperm cells.

Animals↗
PubMed1996

Efficacy of oocytes donated by older women in an oocyte donation programme.

Population and insemination studies indicate that women experience declining fertility with ageing. The question therefore arises whether older women are suitable oocyte donors. This study addresses this issue by examining the relationship between oocyte donor age and clinical outcome in a large oocyte donation programme. We retrospectively reviewed data from 458 consecutive oocyte donation cycles completed by 164 different designated oocyte donors. Data were divided into two groups: group A, cycles with donors aged 21-30 years at the time of follicular aspiration (193 cycles, 88 donors); and group B, cycles with donors aged 31-40 years at the time of follicular aspiration (265 cycles, 86 donors). Five donors, because of ageing during repetitive donations, contributed data to groups A and B. In a given cycle, all oocytes for a recipient came from only one designated donor. Comparing the two donor groups, there was no difference in the amount of gonadotrophin used to achieve optimal stimulation; however, more oocytes were obtained from group A than group B donors (16.8 +/- 6.9 and 15.1 +/- 8.1 respectively, P < 0.05). Similar percentages of oocytes were fertilized in each group, resulting in the transfer of comparable numbers of embryos (4.5 +/- 1.1 and 4.4 +/- 1.3 respectively). Comparable clinical pregnancy rates were achieved (group A, 36%; group B, 37%). The spontaneous abortion rates were also similar (group A, 20%; group B, 12%), resulting in comparable ongoing and delivered pregnancy rates per cycle (group A, 29%; group B, 32%) and per embryo transferred (group A, 6.4%; group B, 7.3%). In conclusion, women of proven fertility should not be excluded from donating oocytes simply because of their age. There exists a cohort of fertile women who resist the decreasing fecundity and increasing spontaneous abortion rates associated with ageing. With careful screening, many women of proven fertility can donate oocytes until the age of 40 years with an efficacy equal to that of younger women. Given the relative shortage of suitable oocyte donors, and increasing requests from recipients with previous donor oocyte babies to obtain oocytes from the same, now older, donor, the findings of this study are of practical clinical importance.

Adult↗
PubMed1994

Time-course of oocyte activation, pronucleus formation and cleavage in human oocytes fertilized by intracytoplasmic sperm injection.

Knowledge of the timing of the stages of fertilization in humans is still limited because the time of gamete fusion is not known when pre-ovulatory or in-vitro matured cumulus-enclosed oocytes are inseminated. We therefore studied the morphological nuclear changes in 14 patients' oocytes by means of light microscopic observation at 2, 4, 6, 8, 16, 18 and at 20 h after intracytoplasmic single sperm injection (ICSI). A total of 144 metaphase II oocytes were injected with the spermatozoa of the patients' partners. Out of the 134 oocytes that survived the injection, 93 displayed two pronuclei in the course of the observation period (69%). Out of the 93 normally fertilized oocytes, 21 extruded the second polar body at 2 h after micro-injection (23%) and 63 oocytes at 4 h (68%). Pronuclei appeared as early as 6 h after ICSI in 16 normally fertilized oocytes (17%). At 8 h, 75 (80%) oocytes had two visible pronuclei, at 16 h 92 (99%), at 18 h 76 (82%) and at 20 h 63 (68%). In 24 oocytes (26%) the appearance of pronuclei was asynchronous, while the disappearance of the pronuclei was always synchronous, except in one oocyte. Nine of the 134 successfully injected oocytes showed three equal-sized pronuclei (6.7%). Four of the nine multi-pronucleated oocytes did not extrude the second polar body at all, while the time sequence of appearance of pronuclei was similar to that of the normally fertilized oocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed2004

Effects of follicle size and oocyte maturation conditions on maternal messenger RNA regulation and gene expression in rhesus monkey oocytes and embryos.

The relationship between alterations in gene expression and differences in developmental potential in primate oocytes and embryos was examined. Oocytes from 3 sources were used for these studies: 1) in vivo-matured oocytes from monkeys stimulated with FSH and hCG, 2) in vitro-matured oocytes from large follicles of monkeys primed with FSH, and 3) in vitro-matured oocytes from small follicles from nonstimulated (NS) monkeys. Following in vitro fertilization, embryos from these oocytes displayed high, moderate, and low developmental competence, respectively. Oocytes from NS females displayed aberrant accumulation of a number of maternal mRNAs, followed by precocious loss of many maternal mRNAs by the 2-cell stage. Embryos from NS oocytes displayed alterations in expression of key transcription factors after the 8-cell stage. Oocytes and embryos from FSH-stimulated females also displayed alterations in gene expression relative to hCG-stimulated females, but these alterations were much less severe than those observed for NS oocytes and embryos. Our data are consistent with the hypothesis that continued development and maturation of the oocyte within the ovarian follicle in vivo facilitates the production of oocytes of the highest developmental potential, and that in vitro conditions may not support this process as effectively due to differences in the extracellular milieu. These observations are relevant to understanding the role of the in vivo environment on oocyte maturation, and the potential effects of in vitro maturation on human assisted reproduction methods.

Animals↗
PubMed2005

Mouse oocytes regulate metabolic cooperativity between granulosa cells and oocytes: amino acid transport.

A search for genes expressed more highly in mouse cumulus cells than mural granulosa cells by subtraction hybridization yielded Slc38a3. SLC38A3 is a sodium-coupled neutral amino acid transporter having substrate preference for l-glutamate, l-histidine, and l-alanine. Detectable levels of Slc38a3 mRNA were found by in situ hybridization in granulosa cells of large preantral follicles, but levels were higher in all granulosa cells of small antral follicles; expression became limited to cumulus cells of large antral follicles. Expression of Slc38a3 mRNA in granulosa cells was promoted by fully grown oocytes from antral follicles but not by growing oocytes from preantral follicles. Fully grown oocytes were dependent on cumulus cells for uptake of l-alanine and l-histidine but not l-leucine. Fully grown but not growing oocytes secreted one or more paracrine factors that promoted cumulus cell uptake of all three amino acids but of l-alanine and l-histidine to a much greater extent than l-leucine. Uptake of l-leucine appeared dependent primarily on contact-mediated signals from fully grown oocytes. Fully grown oocytes also promoted elevated levels of Slc38a3 mRNA and l-alanine transport by preantral granulosa cells, but growing oocytes did not. Therefore, fully grown oocytes secrete one or more paracrine factors that promote cumulus cell uptake of amino acids that oocytes themselves transport poorly. These amino acids are likely transferred to oocytes via gap junctions. Thus, oocytes use paracrine signals to promote their own development via metabolic cooperativity with cumulus cells. The ability of oocytes to mediate this cooperativity is developmentally regulated and acquired only in later stages of oocyte development.

Alanine↗