Number Of High Quality Oocytes Research Articles

Evaluation of Seasonal Heat Stress on Transcriptomic Profiles and Global DNA Methylation of Bovine Oocytes.

Heat stress affects oocyte developmental competence and is a major cause of reduced fertility in heat stressed cattle. Negative effects of heat stress on the oocyte have been observed at morphological, biochemical and developmental levels. However, the mechanisms by which heat stress affects the oocyte at the transcriptional and epigenetic levels remain to be further elucidated. Here we aimed to investigate the effect of heat stress on oocyte quality, transcriptomic profiles and DNA methylation of oocytes collected through the transition from spring to summer under Louisiana conditions. Summer season resulted in a lower number of high quality oocytes obtained compared to the spring season. There was no difference in in vitro maturation rates of oocytes collected during spring as compared to summer. RNA sequencing analysis showed that a total of 211 and 92 genes were differentially expressed as a result of heat stress in GV and MII oocytes, respectively. Five common genes (E2F8, GATAD2B, BHLHE41, FBXO44, and RAB39B) were significantly affected by heat in both GV and MII oocytes. A number of pathways were also influenced by heat stress including glucocorticoid biosynthesis, apoptosis signaling, and HIPPO signaling in GV oocytes, and Oct4 pluripotency, Wnt/beta-catenin signaling, and melatonin degradation I in MII oocytes. In addition, fluorescent immunocytochemistry analysis showed no difference in global levels of DNA methylation and DNA hydroxymethylation at either the GV or MII stage between spring and summer oocytes. The results of this study contribute to a better understanding of the effect of heat stress on the molecular mechanisms altered in bovine oocytes.

A modern approach to the management of candidates for assisted reproductive technology procedures.

The growing use of assisted reproductive technologies (ART) and the incorrect information from mass media, determined in the public the wrong idea that the right moment of life for programming a pregnancy can be delayed well beyond the physiological fertile age. Spare and insufficient health authorities' interventions are driven to explain to the general population the reduction of the fertility potential of couples and particularly of women with advancing age. This situation, characterized by more and more women seeking for pregnancy after age 38-40 imposes to specialists in Reproductive Medicine an honest and transparent counselling. Today, more than ever, it is pertinent to talk about the need of an "ethic approach" to ART, by which the specialist takes care of all the aspects inherent to infertility, such as the strong motivations of the couples in searching a child, the wrong perception of ART infallibility, the incorrect advertising in the mass media about the pregnancy of elderly actresses and show-girls, and finally, the enormous amount of commercial interests revolving around the "business" of in-vitro fertilization. In this context, the ideal policy that an ART center should adopt entails the correct and rapid identification of the characteristics of the couple, the exploitation of women ovarian reserve to obtain the right number of high quality oocytes, the protection of patients' health, the identification of the embryos with the highest chances of implantation and the reduction of the time to pregnancy. Here we analyse how to obtain each of these goals, through a literature review and expert clinical opinion.

A modified protocol for <i>in vitro</i> maturation of mouse oocytes from secondary preantral follicles

A 2-step culture system was designed and tested for the in vitro maturation efficiency of oocytes from pre-puberty preantral follicles of FVB/N inbred mice. The following modifications were made: 1) The concentration of ITS was reduced by half in the basal MIF medium to minimize uncoordinated growth between oocyte and GC cells; 2) Heterogeneous preantral follicles were cultured in groups of 3 - 5 follicles in hanging drops of medium with reduced concentration of ITS for six days to induction follicular aggregation. This hanging drop method mimics a 3-D IVM culture system at the early stage of cultivation in which the sphere structure of each follicle is well maintained. It also enables follicles in each aggregate to communicate with each other, synchronize their growth, and thus prevent immature follicular rupture. 3) Medium was further supplemented with retinoic acid to enhance developmental capacity of meiotically arrested oocytes. After a 14-day culture in vitro, ~37% of the collected inbred preantral follicles completed nuclear maturation. Approximately 94% of the mature oocytes tested were able to be fertilized; and 77% of them developed into healthy embryos. These results demonstrate that our IVM system is reliable to produce a satisfactory number of high quality oocytes. In addition, multiple cytoplasmic parameters, including gene expression of key regulators, chromosome/spindle organization, mitochondrial proliferation and distribution, and total ATP content were explored to characterize the supportive and limiting components of our IVM system so that the culture system can be further optimized.

The effect of withholding gonadotropin (a coasting period) on the outcome of in vitro fertilization cycles

Objective The purpose of this study was to evaluate the effects of coasting (withdrawing gonadotropin administration) on the cycle outcomes, including total number and quality of oocytes and embryos and pregnancy rate in patients undergoing in vitro fertilization. Study design Sixty-seven patients undergoing coasting were evaluated in a retrospective study at the Royan Institute between May 2002 and June 2003. All patients underwent standard long protocol with gonadotropin-releasing hormone (GnRH) analogue and human menopausal gonadotropin (HMG) and then in vitro fertilization and embryo transfer (IVF-ET). Coasting was considered when the estradiol level was more than 3000 pg/ml and the number of follicles >10 in each ovary. HCG was administered whenever estradiol reached ≤3000 pg/ml and then the (IVF-ET) program was carried out. According to coasting periods, patients were divided into two groups: coasting period ≤3 days and coasting period >3 days. Statistical comparisons were performed using Student's t test and Fisher's exact test. Results There were no significant differences between the two groups with regard to mean age, body mass index (BMI), number of polycystic ovary syndrome (PCOS) patients, number of HMG ampoules and stimulation duration. The total number of retrieved oocytes decreased significantly in patients with more than 3 days of coasting ( P = 0.04). The number of high quality oocytes also decreased in this group; however, this did not reach significant levels. There were no significant differences between the two groups with regard to fertilization and pregnancy rate. None of the patients developed severe ovarian hyperstimulation syndrome (OHSS). Conclusion This study reveals that a long coasting period (>3 days) has a negative effect on the number of oocytes, although the fertilization and pregnancy rates are not affected. Prospective randomized studies with larger sample sizes are needed to compare coasting with other procedures.

The Evaluation of Withholding Gonadotropins (Coasting) Effects on the Outcome of In-Vitro Fertilization Cycles

Ovarian hyper stimulation syndrome (OHSS) is a rare life threatening complication of controlled ovarian stimulation for in vitro fertilization (IVF). To reduce this risk several approaches have been used. Withholding gonadotropin administration or coasting is one of the most effective and popular methods used to prevent OHSS. However the effect of coasting on the number and quality of oocyte and embryo is still controversial. In addition, the duration of coasting seems to be a prognostic factor for the outcome of IVF. The purpose of this study was to evaluate the coasting effects on the cycle outcome in patients undergoing in vitro fertilization Retrospective study Sixty-seven patients undergoing coasting were evaluated in Royan research center between May 2002 and June 2003.The study was approved by our Institutional Review Board and University of Jahad Ethics committee. After signing informed consent forms, all patients underwent standard long protocol with GnRH analogue and HMG and then in vitro fertilization and embryo transfer (IVF-ET). Coasting was considered when estradiol level was more than 3000 pg/ml and the number of follicle >10 in each ovary. HCG was administered whenever estradiol reached ≤ 3000 pg/ml and then IVF-ET program was carried out. According to coasting period, patients were divided in two groups: (coasting period ≤ 3 days and coasting period > 3 days). Statistical comparisons were performed using student t test and Fisher exact test. P value <0.05 was considered as significant level. There were no significant differences between two groups in mean age, BMI, percent of PCOS (poly cystic ovarian syndrome) patients, number of HMG ampoules and stimulation duration. A total number of retrieved oocytes decreased significantly in patients with more than 3 days of coasting(6.2 ± 4.2 vs. 9.5±5.3)(P value=0.04).Also the number of high quality oocytes decreased in this group; however they did not reach to significant levels (5.1 ±.2.9 vs. 7.6 ± 4.7) (P value = 0.06) There were no significant differences between two groups in fertilization and pregnancy rate. None of patients developed severe OHSS. This study reveals that coasting effectively prevents OHSS however long coasting period (>3 days) has negative effect on the number of oocytes, although the fertilization and pregnancy rates do not be affected. Prospective randomized study comparing coasting with other procedures is recommended.

Effect of polyunsaturated fatty acid supplementation on biophysical parameters and chilling sensitivity of ewe oocytes.

Fat supplementation in the diet influences reproductive performance of lactating ruminants. Changes in the fat supply alter fatty acid composition and this can affect physical properties of cell membranes. This study examined the effect of rumen bypass polyunsaturated fatty acid (PUFA) supplementation on oocyte quality, chilling sensitivity, and lipid phase transition in oocytes of the sheep. Ewes were fed a diet supplemented with calcium soaps of fish oil for 13 weeks. More follicles and oocytes were found in the ovaries of ewes supplemented with PUFA than in control ewes. The number of high-quality oocytes was higher in ewes fed PUFA than in control ewes (74.3 and 57.0%, P < 0.05, respectively). Evaluation of phospholipid fatty acid composition indicated that PUFA were present in small proportions in oocytes, and eicosapentaenoic acid and docosahexaenoic acid were absent. Supplementation with PUFA increased the proportion of long chain unsaturated fatty acid in the plasma and cumulus cells phospholipids by 7.4 and 12.7%, respectively (P < 0.05). Integrity of oocyte membranes following chilling (16 degrees C, 15 min) was improved by PUFA supplementation increasing from 62.5 to 90.0% (P < 0.05). Due to changes in the oocyte's fatty acid profile, physical properties of the membrane were changed and the midpoint temperature of lipid transition reduced by 11 degrees C. These results suggest that supplementation of rumen bypass PUFA to ruminant diets can change fatty acid composition of follicle components and influence parameters such as number and quality of oocytes and their chilling resistance.

In vitro development of oocytes from porcine and bovine primary follicles

A limiting factor to realising the full potential of many of the new reproductive techniques is the lack of availability of fertile oocytes. Methods for maturing oocytes in vitro (IVM) have been developed to address this problem but the success rate and quality of embryos produced by IVM is variable. The variation in success may be due to the poor quality of oocytes that are being selected for maturation, since these would be taken from developed antral follicles. To attempt to eliminate this variation and increase the numbers produced, it may be better to use the large source of oocytes from preantral and primordial follicles by developing systems for in vitro growth (IVG). In vitro systems that utilise early growing follicles as a source of oocytes have been developed for laboratory species and these have been successful in producing live young. If successful, IVG in association with IVM would supercede existing technology for assisted reproduction in both humans and animals by making it possible to develop the desired number of high quality oocytes from small amounts of ovarian tissue. However, developing IVG systems for species with follicles that develop over several months presents enormous technical challenges. We have developed systems that permit the growth of individual porcine and bovine preantral follicles for periods of up to 20 days. Porcine follicles grown in micro-wells show a higher rate of survival if grown in the presence of serum than follicles grown under serum free conditions. Oocytes recovered from in vitro grown porcine follicles are capable of reaching metaphase II after in vitro maturation. A similar system has been developed for bovine follicles and survival rate is high under serum free conditions but as yet no oocytes from in vitro grown oocytes have been capable of completing meiotic maturation.

Oocyte recovery and maturation in the American black bear (Ursus americanus): a model for endangered ursids.

A study was conducted to determine if meiotic maturation could be induced in ovarian oocytes of the American black bear (Ursus americanus), a model for gamete "rescue" techniques for endangered ursids. Ovaries obtained from 48 black bears yielded 2,403 oocytes (51.1 +/- 4.9/female), of which 777 (32.3%) were morphologically classified as excellent quality. More total oocytes were recovered from donors that were anestrous compared to luteal/pregnant (P < 0.05) at the time of ovarian excision. Delaying the recovery of oocytes from antral follicles within excised ovaries from 12-24 hr to 25-36 hr had no effect (P > 0.05) on the overall number of high quality oocytes recovered or subsequent maturational ability. The highest incidence of metaphase II was reached between 48 and 60 hr of in vitro incubation. Donor status (anestrous vs. luteal/pregnant) had no influence on the oocyte maturation rate by 24 or 48 hr, but by 60 hr, more (P < 0.05) oocytes recovered from anestrous females (43.9%) had achieved metaphase II compared to luteal/pregnant counterparts (23.1%). In preliminary trials involving endangered ursids, 54 ovarian oocytes were recovered from three aged sun bears (Helarctos malayanus), of which 72.2% were excellent quality and 15.4% matured in vitro to metaphase II. Similarly, 119 antral oocytes were recovered from two aged sloth bears (Melursus ursinus), of which 41.2% were excellent and 17.5% matured in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)