Predict Embryo Quality Research Articles

Zona pellucida birefringence in in vivo and in vitro matured oocytes

To evaluate zona pellucida birefringence (ZPB) in immature and mature oocytes collected after controlled ovarian stimulation and to assess the influence of ZPB on oocyte development. Prospective study. Private assisted reproduction centre. Thirty patients undergoing intracytoplasmic sperm injection. The ZPB of mature and immature oocytes was evaluated using a polarization imaging software module, and the oocytes were classified as high birefringence (HB) or low birefringence. The ZPB of in vivo and in vitro matured oocytes and its influence on spontaneous nuclear maturation in vitro, fertilization, and embryo quality. The percentage of HB oocytes was higher in immature than in mature oocytes (40.1 vs. 23.6%). Among immature oocytes, an increased percentage of HB in prophase-I stage oocytes compared to metaphase I stage oocytes was also observed (50.7 vs. 25.0%). However, the percentage of HB oocytes did not change when comparing oocytes before and after in vitro maturation for both prophase I and metaphase I oocytes. No influence of ZPB was observed on the spontaneous in vitro maturation potential. Exclusively for metaphase II retrieved oocytes, a positive influence of ZPB on fertilization (odds ratio [OR], 1.78; 95% confidence interval [CI], 1.27-2.49) and embryo quality (OR, 2.28; 95% CI, 1.04-4.99) was noted. ZPB may be a useful tool to predict embryo quality for metaphase-II oocytes. Moreover, the completion of nuclear changes in the production of metaphase-II oocytes in vitro may not reflect their molecular maturity.

Correlationship of antimullerian hormone (AMH) in follicular fluid with oocyte, embryo quality and pregnancy outcome in ART cycles

OBJECTIVE: To verify the correlationship of AMH concentration in collective follicular fluid with the quality of oocytes, embryos and pregnancy outcome in down regulated IVF cycles. DESIGN: Prospective study. MATERIALS AND METHODS: Controlled ovarian hyperstimulation with gonadotropins (225 IU daily) was done in 206 women undergoing IVF cycles. Egg collection was done after 34 hours of injection hCG. Follicular fluid (FF) from lead follicles (measuring > 15 mm in diameter) was collected and equal volume from each follicle was pooled. Estradiol, progesterone along with AMH concentration and protein content of the pooled FF were measured. Retrieved oocytes and resultant embryos were graded as per gradual embryo scoring method. D3 or D5 embryo transfer was done in 196 cycles. Day 14 serum β hCG level > 25 mIU /ml was considered as positive indication of pregnancy. Two weeks later, fetal sac with cardiac activity on transvaginal sonography was considered as clinical pregnancy. Statistical analysis of the data for three groups viz. low, medium and high as per < 25, 25 -75 and > 75 percentile of FF AMH was done. Student t test to differentiate pregnant and nonpregnant groups, P value, one way ANOVA to compare variance in three groups of FF AMH concentration, correlation coefficient, receiver operator characteristic (ROC) was done to establish cutoff values of FF AMH to predict embryo quality and pregnancy outcome. RESULTS: Out of 72 clinical pregnancies obtained in196 embryo transfers, 70 pregnancies occurred in low and medium FF AMH groups irrespective of age and poor or good responders. The Pearson r for pregnancy outcome in low and medium FF AMH groups was 0.7812 and 0.5923 as compared to -0.7354 in high FF AMH group (P value < 0.0001). The FF E2 showed inverse correlationship with FF AMH concentration (r = - 0.5624). A better oocyte and embryo quality was found in lower levels of AMH in follicular fluid (34.9 ± 13.2 ng / g protein) as compared to higher levels of AMH levels (58.2 ± 9.8 ng/g protein). ROC AUC of FF AMH for pregnancy outcome was observed as 87 % (sensitivity of 90.9 % and specificity of 71.4 %) with cutoff value < 27 ng/g protein where likelihood ratio was 1.0. CONCLUSIONS: Lower values of FF AMH favored higher FF E2 concentrations with a better oocyte and embryo quality resulting in positive pregnancy outcome in ART cycles.

Soluble human leukocyte antigen-G and pregnancy success

Non-invasive methods of assessing embryo quality are critical for pregnancy success following IVF or intracytoplasmic sperm injection (ICSI). The addition of new non-invasive morphological and biochemical analyses may further improve pregnancy success, allowing the transfer of a single embryo, thereby reducing the risks involved in multiple births following IVF/ICSI. The presence of a protein, soluble human leukocyte antigen-G (sHLA-G), in embryo cultures has been suggested as a way to non-invasively predict embryo quality and pregnancy success, especially when used in conjunction with current embryo quality assessment methods. Detection of sHLA-G in embryo culture medium has been correlated with pregnancy success in 12 studies, but three studies were not able to detect sHLA-G. This is a review of the literature on sHLA-G detection in IVF/ICSI, and reasons are proposed for the reported discrepancies, as well as guidelines for reporting of data in future studies. Furthermore, it is suggested that the use of an HLA-G transgenic mouse model would advance understanding of the mechanism of action of sHLA-G in preimplantation embryos and its correlation to embryo health and viability. Research on a mouse model, combined with clinical studies, should enable the development of a fast and reliable method for utilizing sHLA-G detection to improve pregnancy success after IVF/ICSI.

Presence of a cytoplasmic halo and complete nucleolar alignment is predictive of implantation rates and pregnancy rates for patients 35–39 years old but not for patients less than 35

To determine the relationship between age, the presence of a cytoplasmic halo, nuclear precursor body alignment (NPB), and pronuclear (PN) morphology grade in human embryos with respect to pregnancy outcomes. Retrospective study in a university based IVF center. All patients undergoing ART from 2001 – 2006 who gave IRB consent were considered for analysis. Zygotes with a PN grade >1.0 were considered morphologically abnormal and excluded. The remaining 1579 embryos from 714 patients were considered normal and were included in our study. Variables analyzed were cytoplasmic halo (complete or partial) and complete NPB alignment. Clinical pregnancy rate (CPR) and implantation rate (IR) for transfers where a NPB pattern or halo was homologous within the transfer cohort were statistically evaluated by Fisher's Exact Test and Wilcoxon Rank Sum test. For transfers with PN grade ≤1.0, the CPR was 51% with an IR of 29.5%. CPR and IR were analyzed based on the presence or absence of a cytoplasmic halo, for the presence or absence of complete NPB alignment, and for the presence of both features for patients <35 years (Table 1) and patients 35–39 (Table 2). Table 1Patients <35 years of ageCytoplasmic HaloComplete NPB AlignmentHalo and Complete AlignmentPresentAbsentPresentAbsentPresentAbsentTransfer2472911713871182CPR51.1%48.3%52.1%52.2%50.7%52.8%IR33.1%30.5%31.4%32.2%30.8%33.2% Open table in a new tab Table 2Patients 35–39 years of ageCytoplasmic HaloComplete NPB AlignmentHalo and complete AlignmentPresentAbsentPresentAbsentPresentAbsentTransfers911321651487CPR58.2%130.8%66.7%49.2%85.7%346.0%IR30.1%315.4%36.5%25.3%50.6%323.1%1 P=0.08, 2 P=0.10, 3 P<0.01. Open table in a new tab 1 P=0.08, 2 P=0.10, 3 P<0.01. With the current debate of PN morphology and its ability to predict embryo quality, our data supports using cytoplasmic halo and nuclear precursor body alignment as predictors for successful transfers. Several studies have proposed that pregnancy and live birth rates do not correlate with PN scoring. While differences in zygote morphology examined in this study do not affect outcome in young patients, the presence of a halo and complete NPB alignment is associated with statistically higher IR and CPR in patients 35–39. This suggests that the combined assessment of halo and NPB alignment is useful in embryo selection for patients with advanced maternal age.

O-173: Serum anti-mullerian hormone (AMH) and antral follicle count (AFC) are better predictors of embryo quality and blastocyst formation than patient age and follicle stimulating hormone (FSH)

Both age & FSH have been extensively evaluated as a predictor of ovarian stimulation and ART success; however, both have limitations. There is, therefore, the need for additional & more accurate pre-ART cycle prognostic testing. While AMH has been found to be associated with poor ovarian reserve & ART outcomes, there is inadequate information regarding the ability of AMH to predict embryo characteristics. The present objective is to evaluate age, AFC, FSH, & AMH and their ability to predict overall embryo quality & blastocyst transfer.

187 INCREASED NUMBER OF EXPANDED BOVINE BLASTOCYSTS IN SOF AND CR1 RELATIVE TO KSOM

Assessment of morphological stage grade is a subjective procedure. Stage grade is of vital importance to, among other things, recipient synchrony for the purpose of establishing successful pregnancies. Asynchronous embryo transfer has led to decreases in pregnancy rates (Farin et al. 1995 Biol. Reprod. 52, 676–682) and has been implicated in contributing to large offspring syndrome (Young et al. 1996 Theriogenology 45, 231). Differences in embryo kinetics based on culture conditions have been well documented (Mello et al. 2005 Reprod. Fert. Dev. 17, 221 abst). Whether such differences are the result of species, breed, metabolic stress, sire effects, or separation from an in vivo environment has yet to be determined. The correlation between oxygen respiration rates and embryo morphology as well as embryo diameter in bovine embryos produced in vitro has shown promise in the development of a more objective predictor of embryo quality and perhaps pregnancy initiation (Lopes et al. 2005 Reprod. Fert. Dev. 17, 151 abst). As well, recent examination of gene expression patterns of in vitro-derived bovine embryos seems to indicate that longer periods of in vitro culture are associated with lower rates of embryo survival (Lonergan et al. 2006 Theriogenology 65, 137–152). We hypothesize that differences do exist in the number, rate, and morphological appearance of blastocysts and that these parameters are in large part based on culture conditions in vitro. The objective of this experiment was to determine the timing and distribution of blastocyst formation of in vitro-produced bovine embryos cultured in SOF8, CR18AA, and KSOM8, under a standard incubation environment. Bovine ovaries from a local abattoir were aspirated and matured for 18-22. Oocytes were fertilized with frozen-thawed Percoll-separated semen from a Holstein bull. Presumptive zygotes were vortexed to remove cumulus cells and placed into 3 different culture media in a highly humidified atmosphere containing 20% oxygen, 5% carbon dioxide, and compressed air at 38.5�C. Embryos were evaluated specifically at 168 h post-insemination (Day 7) and assigned a morphological stage grade (IETS) to determine fixed time point differences. A total of 6 complete replicates were performed. Only embryos exhibiting the presence of a blastocoel at this time were documented (early blast, mid-blast, expanded blast). At 168 h post-insemination, there were no significant differences in the total number of embryos reaching early or mid-blast stage in any of the media. However, chi-square analysis revealed an increase in the number of expanded blastocysts in SOF (n = 813) and CR1 (n = 838) treatments compared to KSOM (n = 824; P &amp;lt; 0.0001). Expanded blastocysts in SOF were also greater in number than in CR1 (P &amp;lt; 0.05). Embryo selection based on development to the expanded blastocyst stage on Day 7 may prove useful in increasing pregnancy rates, and may validate qualitative correlations based on oxygen consumption and gene expression profiles for embryos produced in vitro.

Early pronuclear breakdown is a good indicator of embryo quality and viability

To examine whether the timing of pronuclear breakdown can be a predictor of embryo quality and viability. Retrospective comparison of the development and quality of early and late developing zygotes. Infertility and endocrinology unit in a university hospital. One thousand seven hundred eighty-two zygotes obtained in 383 consecutive IVF cycles. Culture of all fertilized embryos. Number of fertilized zygotes showing early pronuclear breakdown at 22-25 hours postinsemination, embryo quality, pregnancy rates (PR), implantation rates. Early pronuclear breakdown embryos had a significantly higher cell number (4.4 +/- 1.2) compared with the late pronuclear breakdown embryos (3.6 +/- 1.4). When comparing the frequency of the early pronuclear breakdown embryos according to the method of fertilization, we failed to find any significant difference between the IVF (37.1%) and the intracytoplasmic sperm injection (ICSI) (41.1%) groups. The transfer of early pronuclear breakdown embryos resulted in a significantly higher clinical pregnancy rate than those with late pronuclear breakdown (48.3% vs. 27.3%). The implantation rate was higher in the early pronuclear breakdown group than in the late pronuclear breakdown group (26.5% vs. 15.1%). Early pronuclear breakdown is a strong indicator of embryo viability, and may be used as an additional criterion in the selection of embryos for transfer.

Embryo quality and IVF treatment outcomes may correlate with different sperm comet assay parameters.

Standard semen parameters have proven poor at predicting the outcomes of IVF treatment cycles. As recent studies suggest that the male genome may play an important role in early embryogenesis, this study attempts to correlate the level of sperm DNA damage in fresh semen and prepared sperm with the outcomes of conventional IVF treatment cycles. Forty patients embarking on IVF treatment were recruited into this prospective observational study. Both fresh semen and PureSperm-prepared sperm were processed using a modified comet assay 3-6 months prior to the patients' IVF treatment cycles. Comet head DNA (mean and integrated head density) and tail DNA parameters (length and moment) were measured separately. Significant correlations between total sperm concentration and between comet length, moment, mean head density with embryo quality were detected in fresh semen and prepared sperm. Surprisingly, no significant correlations between head and tail parameters were detected. Comet head and tail DNA parameters appear to be potentially useful as predictors of embryo quality and IVF outcomes, especially in couples with unexplained subfertility. The lack of correlation between head and tail parameters may be due to a different mechanism of DNA damage within these two compartments.

The predictive value of pronuclear morphology of zygotes in the assessment of human embryo quality

Recent studies have shown that zygote morphology could be used for the assessment of human embryo quality. Pronuclear (PN) morphology is based on certain distinct features seen in zygotes 16-18 h after fertilization. In the present study PN stage morphology was assessed and combined with a single embryo transfer in order to investigate whether currently used zygote classifications are able to predict embryo quality and implantation rates. Zygotes were analysed according to two different classification systems. In the first, a total of 764 zygotes was analysed according to the degree of polarization of nucleolar precursor bodies (NPB). Zygotes with unpolarized PN (i.e. scattered localization of NPB) showed significantly slower (P < 0.005) cleavage rates (38.9%) than zygotes having at least one pronucleus polarized (57.3% and 54%). However, there was no difference in the pregnancy rate in 105 single embryo transfers between the groups. The appearance of a cytoplasmic halo was related to embryo morphology. Embryos derived from halo-positive zygotes had significantly better (P < 0.05) morphology (60.9%) compared to halo-negative derived embryos (52.2%), but in terms of pregnancy rates no difference was found. A total of 1520 zygotes was analysed according to a second classification system, which was based on the number and distribution of NPB. In the comparative analysis, none of the six different classes produced superior quality embryos or higher pregnancy rates in 144 single embryo transfers. Our results indicate that there are no significant differences in embryo quality or implantation/pregnancy rates between proposed zygote classes.