Estella Jones

Does storage time influence postthaw survival and pregnancy outcome? An analysis of 11,768 cryopreserved human embryos

OBJECTIVE To evaluate the impact of cryopreservation storage duration on embryo survival, implantation competence, and pregnancy outcome. DESIGN Retrospective study. SETTING Academic tertiary-referral infertility center. PATIENT(S) In vitro fertilization patients and recipients of oocyte donation cycles who had cryopreserved embryos and underwent at least one thaw cycle from 1986 to 2007. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Postthaw survival proportion and implantation, clinical pregnancy, miscarriage, and live birth rates. RESULT(S) Length of storage time did not have a significant effect on postthaw survival for IVF or oocyte donation cycles, or for embryos frozen at the pronuclear or cleavage stages. There was no significant impact of the duration of storage on clinical pregnancy, miscarriage, implantation, or live birth rate, whether from IVF or oocyte donation cycles. Logistic regression analysis demonstrated that the length of storage time or developmental stage at freezing were not predictive of embryo survival or pregnancy outcome. Only oocyte age, survival proportion, and number of transferred embryos were positive predictors of pregnancy outcome. CONCLUSION(S) Cryostorage duration did not adversely affect postthaw survival or pregnancy outcome in IVF or oocyte donation patients.

Pregnancy-specific β-1-glycoprotein 1 and human leukocyte antigen-E mRNA in human sperm: differential expression in fertile and infertile men and evidence of a possible functional role during early development

BACKGROUND Mature spermatozoa contain thousands of mRNA transcripts. It has been recently shown that human sperm can deliver RNA into the oocyte, suggesting that mRNAs might have a role before or after fertilization. Human embryos express PSG1 (pregnancy-specific beta-1-glycoprotein 1) and HLA-E (human leukocyte antigen-E), molecules playing a role in implantation and early development. We compared PSG1 and HLA-E sperm mRNA levels in fertile and infertile men and we tested the hypothesis that these transcripts are selectively retained in the newly formed zygote. METHODS Real-time RT-PCR was used to analyze sperm mRNA levels (n = 11 fertile, n = 31 infertile patients) of PSG1, HLA-E and PRM2 (protamine 2). The presence of PSG1 and HLA-E proteins was evaluated by western blot in sperm protein extracts (n = 3). Using ICSI of human sperm into hamster oocytes we evaluated the permanence of these mRNAs at different time points (n = 10 for each time) after fertilization. RESULTS PSG1, HLA-E and PRM2 transcripts were demonstrated in ejaculated sperm. The fertile group showed significantly higher levels of PSG1 and HLA-E mRNA (both P < 0.05) than the infertile group, whereas PRM2 levels were not significantly different. However, PSG1 and HLA-E proteins were not found in ejaculated sperm. Following ICSI, PRM2 was undetectable after fertilization; conversely, PSG1 and HLA-E transcripts remained detectable for at least 24 h of zygotic development. CONCLUSIONS We provide new evidence that indicates that human sperm deliver transcripts that may have a role in early embryo development and decreased levels of these transcripts may be associated with infertility.

Impact of transabdominal ultrasound guidance on performance and outcome of transcervical uterine embryo transfer.

AbstractPurpose: To determine the impact of transabdominal ultrasound guidance on embryo transfer during IVF therapy. Methods: Retrospective analysis of 823 consecutive embryo transfers. Three hundred and sixty-seven procedures performed with transabdominal ultrasound guidance were compared to 456 cases performed with the “clinical touch” method. Results: Ultrasound-guided embryo transfer yielded higher, but not statistically significant, clinical pregnancy (48% vs. 44%) and implantation rates (22% vs. 20%). The incidence of multiple pregnancies, ectopic and multiple pregnancy rates were similar. The frequency of negative factors typically associated with difficult transfers, such as requirement of use of tenaculum, and presence of blood or mucus in the catheter tip, were significantly lower in the ultrasound-guided group in comparison with the clinical touch group. Ultrasound-guided embryo transfer was associated with a significantly increased easiness of transfer performance; 95% of the transfers were rated as very easy in the ultrasound-guidance group compared to 87% in the clinical touch group. The use of a soft pass catheter was the only variable independently and significantly associated with pregnancy success (odds ratio = 2.74). Conclusion(s): Ultrasound-guidance facilitates embryo transfer and in combination with the use of a soft catheter should be implemented to optimize embryo transfer results.

Outcome of intracytoplasmic sperm injection in azoospermic patients : Stressing the liaison between the urologist and reproductive medicine specialist

OBJECTIVES To analyze the outcome of intracytoplasmic sperm injection (ICSI) cycles in infertile couples in whom the main diagnosis of infertility was azoospermia of obstructive and nonobstructive origin. METHODS Eighty-three consecutive ICSI cycles were carried out with retrieved testicular or epididymal spermatozoa, 60 cycles in 32 patients with obstructive azoospermia and 23 cycles in 12 patients with nonobstructive azoospermia. Fifty-four testicular biopsies (testicular sperm extraction) and 18 epididymal aspirations (microepididymal sperm aspiration) were performed.Results. Motile spermatozoa were recovered in 65 cycles (90.3%). In another 3 (4.2%), nonmotile spermatozoa were retrieved. In 4 patients (5.5%), sperm could not be recovered. In 11 cycles, frozen sperm from a previous procedure were used. A significantly lower fertilization rate (64% versus 73%, P = 0.02), clinical pregnancy rate (13% versus 47%, P <0.001), and good embryo quality rates (35% versus 56%, P = 0.009) were observed in patients with nonobstructive azoospermia. In patients with obstructive azoospermia, no significant differences were observed when the outcome was analyzed on the basis of the sperm origin (ie, from testicular sperm extraction or microepididymal sperm aspiration). CONCLUSIONS When combining testicular sperm extraction or microepididymal sperm aspiration with ICSI in patients with obstructive azoospermia, the results in terms of fertilization, implantation, and pregnancy rates were similar to those found in patients with nonazoospermic obstruction who underwent ICSI with ejaculated sperm. Patients with nonobstructive azoospermia had lower fertilization, embryo quality, and pregnancy rates than did those with obstructive azoospermia, probably because of severe defects in spermatogenesis, leading to poor gamete quality. The urologist and reproductive endocrinologist now have an excellent therapeutic option to offer men with previously intractable infertility.

Positioning of Chromosomes in Human Spermatozoa Is Determined by Ordered Centromere Arrangement

The intranuclear positioning of chromosomes (CHRs) is a well-documented fact; however, mechanisms directing such ordering remain unclear. Unlike somatic cells, human spermatozoa contain distinct spatial markers and have asymmetric nuclei which make them a unique model for localizing CHR territories and matching peri-centromere domains. In this study, we established statistically preferential longitudinal and lateral positioning for eight CHRs. Both parameters demonstrated a correlation with the CHR gene densities but not with their sizes. Intranuclear non-random positioning of the CHRs was found to be driven by a specific linear order of centromeres physically interconnected in continuous arrays. In diploid spermatozoa, linear order of peri-centromeres was identical in two genome sets and essentially matched the arrangement established for haploid cells. We propose that the non-random longitudinal order of CHRs in human spermatozoa is generated during meiotic stages of spermatogenesis. The specific arrangement of sperm CHRs may serve as an epigenetic basis for differential transcription/replication and direct spatial CHR organization during early embryogenesis.

Successful extracorporeal mature oocyte harvesting after laparoscopic oophorectomy following controlled ovarian hyperstimulation for the purpose of fertility preservation in a patient with borderline ovarian tumor

PurposeTo report the successful extracorporeal recovery of mature oocytes after laparoscopic oophorectomy following ovarian hyperstimulation for the purpose of fertility preservation in a patient with recurrent serous borderline ovarian tumor.MethodsA 25-year-old nulligravida woman presented with recurrence of a borderline serous adenocarcinoma in the right ovary after been treated conservatively with left oophorectomy for the same.Result(s)The patient underwent ovarian stimulation followed by a laparoscopic oophorectomy and ex-vivo retrieval of oocytes. Twenty two oocytes were recovered: fourteen metaphases II, two metaphases I, five prophases I and one degenerate.Conclusion(s)Mature oocytes were successfully retrieved ex-vivo from the hyperstimulated ovary recovered via laparoscopy. The procedure can be performed in a quick manner, with standard equipment, without damaging the ovary, the follicles or the oocytes, and without the risk of cancer cell spillage associated with the standard transvaginal oocyte retrieval if there is concern of ovarian surface/peritoneal metastatic disease.

Reorganisation of human sperm nuclear architecture during formation of pronuclei in a model system

By fertilisation, two terminally differentiated cells, namely the egg and spermatozoon, are combined to create a totipotent zygote. During this process, the inactive sperm nucleus is transformed into a functional male pronucleus. Recent studies demonstrate that human sperm chromatin has an elaborate multilevel organisation, but almost nothing is known about how sperm chromosomes are transformed during fertilisation. Because of ethical reasons and technical complications, experimentation with human embryos is generally unworkable and adequate model systems are necessary to study the formation of male pronuclei. Here, we analyse remodelling of human sperm chromatin and chromosome architecture in Xenopus egg extracts using immunofluorescent localisation of protamines and centromere protein A, as well as fluorescence in situ hybridisation localisation of major alpha-satellite DNA and whole chromosome territory (CT). We demonstrate noticeable relocalisation of centromeres and remodelling of CT during the decondensation-recondensation cycle, mimicking cellular events that occur in the paternal genome in vivo during fertilisation.

Total quality improvement in the IVF laboratory: choosing indicators of quality

The purpose of this paper is to describe a programme of total quality improvement (TQI) within the IVF laboratory and to provide specific examples of indicators that could be used in such a TQI programme. Although TQI is sometimes confused with quality control (QC) and quality assurance (QA), there are major differences between the three quality plans: (i) QC is an activity designed to ensure that a specific element within the laboratory is functioning correctly; (ii) QA is a comprehensive programme designed to looks at a laboratory as a whole and to identify problems or errors that exist in an attempt to improve the entire process; (iii) TQI is also a comprehensive monitoring process designed not only to detect and eliminate problems, but also to enhance a laboratorys performance by exploring innovation and developing flexibility and effectiveness in all processes. Indicators used in a TQI plan should be objective, relevant to the laboratory, and measure a broad range of specific events or aspects of treatment that reflect the quality of care. Threshold values for each of the indicators should be based on how the specific protocols used in the laboratory impact the outcomes and the nature of the indicators on quality of care.

Elective transfer of two embryos: Reduction of multiple gestations while maintaining high pregnancy rates

Purpose: To determine if the elective transfer of two embryos reduced the incidence of multiple gestations while maintaining high pregnancy rates. Methods: IVF patients and recipients of oocyte donation with an elective day-3 transfer of 2 or 3 embryos were studied. Result(s): In IVF, the elective transfer of 2 embryos resulted in similar pregnancy rate but significantly reduced the overall incidence of multiple gestations (20% versus 39%) when compared to the elective transfer of 3 embryos. Twin gestations decreased from 28% to 19%, and triplets significantly decreased from 9% to 1%. In oocyte donation, the elective transfer of 2 embryos resulted in similar pregnancy rate but also significantly reduced the overall incidence of multiple gestations (26% versus 48%), with twins decreasing from 34% to 24%, and with a significant reduction of triplets (13% versus 2%). Conclusions: In IVF and oocyte donation, the elective transfer of 2 embryos resulted in similar pregnancy rates and significantly reduced multiple gestation rates when compared to the elective transfer of 3 embryos.

Kinetics of human male pronuclear development in a heterologous ICSI model

PurposeTo evaluate human sperm nuclear chromatin decondensation in a heterologous ICSI system using hamster ova injected with human sperm.Materials and methodsFrozen hamster oocytes were injected with Triton X-100 treated sperm and fixed at different time points post ICSI. Oocytes injected with non-treated sperm served as controls. Male pronuclear decondensation was evaluated after staining with DAPI.ResultsSperm cells with partially destroyed membranes and depletion of the acrosome decondense more rapidly and to a greater extent than membrane/acrosome intact cells. Marked variability in pronuclear size was observed for any time point post ICSI, which most probably reflects the heterogeneity in the mature human sperm population.ConclusionRemodeling of male gamete nuclei in this heterologous ICSI mimics events that occur during natural fertilization in humans and therefore this approach may be used for studies of human sperm chromosomes transformations.