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Study Question: Do women ≥40 years old without a male partner who utilize donor sperm have the same reproductive outcomes as those who utilize their partner's sperm?Summary Answer: After controlling for relevant confounders, women ≥40 years old using donor sperm for IVF have significantly higher odds of having a live birth compared to those utilizing their partner's sperm.What Is Known Already: Women who are unpartnered or in same-sex relationships are by definition not infertile, but may choose to conceive using donor sperm. It is not known how IVF outcomes are affected with the use of donor sperm compared to women utilizing their partner's sperm, particularly at very advanced maternal ages.Study Design, Size, Duration: This is a retrospective cohort study conducted at a university-affiliated center of women undergoing IVF with fresh embryo transfer between 2008 and 2018.Participants/materials, Setting, Methods: Patients were divided into two groups based on the ejaculated sperm source utilized: donor or partner sperm. Live birth rate was the primary outcome. Pregnancy rate was the secondary outcome. Multivariable logistic regression was performed and adjusted for age, the developmental stage of the embryo, and the number of embryos transferred. Unadjusted odds ratio (OR) and adjusted OR (aOR) with 95% CI for pregnancy and live birth were estimated. Statistical significance was denoted by P < 0.05.Main Results and the Role Of Chance: A total of 3910 cycles in women ≥40 years old were analyzed, of which 307 utilized donor sperm and 3603 utilized their partner's sperm to conceive. In the univariate analysis, patients utilizing donor sperm were found to have similar pregnancy rates as those utilizing partner sperm (41.0 vs 39.8%, OR: 0.95, 95% CI: 0.75-1.20). After adjusting for age, the number of embryos transferred and the developmental stage of the embryos, the model estimates did not vary (aOR: 1.22, 95% CI: 0.95-1.56). Similarly, the univariate analysis for live birth did not demonstrate a difference between groups (19.2 vs 17.8%, OR: 0.91, 95% CI: 0.67-1.22). However, after a similar adjustment was made for confounders, the use of donor sperm was associated with statistically significant increased odds of live birth (aOR: 1.38, 95% CI: 1.01-1.88).Limitations, Reasons For Caution: As with any retrospective study, the potential for residual confounding exists, despite attempts to control for this with regression modeling.Wider Implications Of the Findings: Women ≥40 years old who are unpartnered or in same-sex relationships can be counseled that their odds of a live birth are slightly better than women in heterosexual relationships utilizing their partner's sperm. These findings serve to further refine and individualize counseling on the expected IVF outcomes for women in this population.Study Funding/competing Interest(s): No funding was sought for this study. The authors declare no competing interests.Trial Registration Number: N/A. [ABSTRACT FROM AUTHOR]

Study Question: Does ovarian stimulation affect embryo euploidy rates or live birth rates (LBRs) after transfer of euploid embryos?Summary Answer: Euploidy rates and LBRs after transfer of euploid embryos are not significantly influenced by gonadotropin dosage, duration of ovarian stimulation, estradiol level, follicle size at ovulation trigger or number of oocytes retrieved, regardless of a woman's age.What Is Known Already: Aneuploidy rates increase steadily with age, reaching >80% in women >42 years old. The goal of ovarian stimulation is to overcome this high aneuploidy rate through the recruitment of several follicles, which increases the likelihood of obtaining a euploid embryo that results in a healthy conceptus. However, several studies have suggested that a high response to stimulation might be embryotoxic and/or increase aneuploidy rates by enhancing abnormal segregation of chromosomes during meiosis. Furthermore, a recent study demonstrated a remarkable difference in euploidy rates, ranging from 39.5 to 82.5%, among young oocyte donors in 42 fertility centres, potentially suggesting an iatrogenic etiology resulting from different stimulation methods.Study Design, Size, Duration: This is a retrospective cohort study that included 2230 in vitro fertilisation (IVF) with preimplantation genetic testing for aneuploidy (PGT-A) cycles and 930 frozen-thawed single euploid embryo transfer (FET) cycles, performed in our centre between 2013 and 2017.Participants/materials, Setting, Methods: A total of 12 298 embryos were analysed for ploidy status. Women were divided into five age groups (<35, 35-37, 38-40, 41-42 and >42 years old). Outcomes were compared between different durations of stimulation (<10, 10-12 and ≥13 days), total gonadotropin dosages (<4000, 4000-6000 and >6000 IU), numbers of oocytes retrieved (<10, 10-19 and ≥20 oocytes), peak estradiol levels (<2000, 2000-3000 and >3000 pg/mL), and sizes of the largest follicle on the day of trigger (<20 and ≥20 mm).Main Results and the Role Of Chance: Within the same age group, both euploidy rates and LBRs were comparable between cycles regardless of their differences in total gonadotropin dosage, duration of stimulation, number of oocytes harvested, size of the largest follicles or peak estradiol levels. In the youngest group, (<35 years, n = 3469 embryos), euploidy rates were comparable between cycles with various total gonadotropin dosages (55.6% for <4000 IU, 52.9% for 4000-6000 IU and 62.3% for >6000 IU; P = 0.3), durations of stimulation (54.4% for <10 days, 55.2% for 10-12 days and 60.9% for >12 days; P = 0.2), number of oocytes harvested (59.4% for <10 oocytes, 55.2% for 10-19 oocytes and 53.4% for ≥20 oocytes; P = 0.2), peak estradiol levels (55.7% for E2 < 2000 pg/mL, 55.4% for E2 2000-3000 pg/mL and 54.8% for E2 > 3000 pg/mL; P = 0.9) and sizes of the largest follicle (55.6% for follicles <20 mm and 55.1% for follicles ≥20 mm; P = 0.8). Similarly, in the oldest group (>42 years, n = 1157 embryos), euploidy rates ranged from 8.7% for gonadotropins <4000 IU to 5.1% for gonadotropins >6000 IU (P = 0.3), from 10.8% for <10 days of stimulation to 8.5% for >12 days of stimulation (P = 0.3), from 7.3% for <10 oocytes to 7.4% for ≥20 oocytes (P = 0.4), from 8.8% for E2 < 2000 pg/mL to 7.5% for E2 > 3000 pg/mL (P = 0.8) and from 8.2% for the largest follicle <20 mm to 8.9% for ≥20 mm (P = 0.7). LBRs after single FET were also comparable between these groups.Limitations, Reasons For Caution: Although this large study (2230 IVF/PGT-A cycles, 12 298 embryos and 930 single FET cycles) demonstrates the safety of ovarian stimulation in terms of aneuploidy and implantation potential of euploid embryos, a multi-centre study may help to prove the generalisability of our single-centre data.Wider Implications Of the Findings: These findings reassure providers and patients that gonadotropin dosage, duration of ovarian stimulation, estradiol level, follicle size at ovulation trigger and number of oocytes retrieved, within certain ranges, do not appear to significantly influence euploidy rates or LBRs, regardless of the woman's age.Study Funding/competing Interest(s): No external funding was received and there are no competing interests to declare.Trial Registration Number: N/A. [ABSTRACT FROM AUTHOR]

Purpose: To demonstrate whether the standard morphokinetic markers used for embryo selection have a similar relationship to blastocyst formation and implantation in two large clinical data sets. Methods: This is a retrospective cohort analysis striving to answer two distinct questions utilizing data sets from two large IVF clinics. Blastocysts (BL) and implanted blastocysts (I) in both clinics, IVI-Valencia (BL = 11,414, I = 479) and WMC (BL = 15,902; I = 337), were cultured in a time-lapse system (EmbryoScope, Vitrolife, Sweden). The study was designed to assess the relationship between early morphokinetic hallmarks and BL development, with a secondary analysis of implantation rates following single-embryo day 3 and day 5 transfers. Results: We performed a detailed graphical analysis for t3, t5, duration of the second cell cycle (cc2) (t3-t2), and the ratio (t5-t3)/(t5-t2). The t5 timing was not affected between the clinics. However, Weill Cornell Medicine's (WCM) proportions were significantly affected by having BL vs. not. A significant decrease of blastocysts with longer t5 in WCM data, while t5 was more informative in the IVI data set for the implantation rate. Conclusions: Morphokinetic intervals for early cleavages were distributed differently between the clinics. Incorporation of embryo-selection algorithms depends on the individual clinic's selected developmental hallmarks, all of which must be validated before incorporation into clinical practice. [ABSTRACT FROM AUTHOR]

Objective: To design a reproductive treatment algorithm based on the sperm DNA fragmentation (SDF) for couples with unexplained infertility following a poor intrauterine insemination (IUI) outcome.Design: Couples that failed IUI with no apparent reproductive issue in both partners were allocated to diverse reproductive treatments on the basis of SDF.Setting: Reproductive medical center in an academic setting.Patient(s): Over 4 years, couples with an unexpected poor IUI outcome and no apparent female or male partner reproductive issues were recruited.Intervention(s): IUI, IVF, and ICSI were performed in the standard fashion following sperm SDF assays.Main outcomes measure(s): Fertilization rate, implantation rate, pregnancy characteristics, and delivery rates.Result(s): A total of 354 couples with unexplained infertility and normal semen parameters underwent 1133 IUI cycles. Clinical pregnancy rate (CPR) with IUI at our center in an age-matched cohort is 23.9% while the study cohort had 1.8%. Following SDF assessment, couples with failed IUI attempts but normal SDF (SCSA 9.8 ± 4.6%; TUNEL 11.8 ± 6.2%) underwent IVF with a CPR of 12.7%; those with abnormal SDF underwent ICSI with ejaculated spermatozoa, resulting in a CPR of 18.7%. This group included couples with normal SDF that had failed IVF. Couples with abnormal SDF that failed ICSI with ejaculated spermatozoa achieved a CPR of 31.0% with surgically retrieved spermatozoa.Conclusion(s): Couples with unexplained infertility that present with unexpectedly poor IUI outcomes can be funneled into a treatment algorithm guided by the integrity of the sperm genome for higher chances of pregnancy using an alternate method of insemination. [ABSTRACT FROM AUTHOR]

The first conception outside of the human body that led to the birth of Louise Brown was a tremendous accomplishment, which opened the door to the utilization of assisted reproductive techniques globally. This brought the understanding that accomplishing life in a dish required several steps, the most obvious being the timing and characteristics of fertilization. It soon became obvious in the 1980s that the most disappointing phenomenon was unexpected and complete fertilization failure. Among the approaches that were attempted to treat male factor infertility, ICSI surfaced as the technique that brought the ratio of the gametes to 1:1 and was also able to grant consistent fertilization and a higher pregnancy rate. ICSI has now been implemented for a quarter of a century, proving itself as the ultimate technique utilizing ejaculated spermatozoa independent of the semen parameters and is the sole insemination method to be used with surgically retrieved spermatozoa. There are currently various indications for ICSI that are widely adopted, rendering it the most popular insemination method worldwide. The reliability of ICSI ensures its employment in upcoming techniques involving in vitro spermatogenesis and neogametogenesis. [ABSTRACT FROM AUTHOR]

Study Question: Is there a benefit to assessing ploidy in delayed embryos reaching the morula stage on Day 6 of development?Summary Answer: Day-6 morulae should be considered for biopsy in women <40 years old undergoing preimplantation genetic testing for aneuploidy (PGT-A) because they are associated with acceptable, albeit reduced, euploidy and implantation rates (IRs).What Is Known Already: Embryo development and morphology have been shown to correlate with aneuploidy and pregnancy rates. During PGT-A cycles, embryos are biopsied if they reach the blastocyst stage by Day 5 or 6, whereas slow-developing embryos are typically deselected and discarded. Determining the viability of slow-developing embryos is particularly relevant for women undergoing PGT-A who have diminished ovarian reserve and a relatively low blastocyst yield.Study Design, Size, Duration: This is a retrospective cohort study that was performed at an academic medical center. Patients who underwent IVF with PGT-A were reviewed for inclusion.Participants/materials, Setting, Methods: A total of 1615 cycles were reviewed. All cycles which involved a biopsy of a cavitating or compacted morula on Day 6 were included (n = 763). PGT-A was performed using array comparative genomic hybridization. The aneuploidy and implantation of morulae were compared to those of blastocysts originating from the same couples.Main Results and the Role Of Chance: The study included 763 cycles in which 1260 morulae and 3014 blastocysts were biopsied. Women were divided into four age groups (<35, 35-37, 38-39 and ≥40 years): the prevalence of aneuploidy was consistently lower among blastocysts (40.3, 50.8, 56 and 78.3%, respectively) than among compacted morulae (68.7, 75.5, 88.9 and 98.1%, respectively) and cavitating morulae (57, 66.4, 81 and 91.6%, respectively) throughout the different age groups (P < 0.001). Of note, the majority of compacted morulae (98.1%) and cavitating morulae (91.6%) were aneuploid in women aged ≥40 years. Compacted and cavitating morulae had significantly higher rates of complex aneuploidy, which involves ≥3 chromosomes, compared with blastocysts (P < 0.001). Furthermore, euploid morulae were associated with a significantly lower IR (28.2 versus 54.6%; P = 0.002) and live birth rate (23.1 versus 55.0%; P = 0.001) compared to euploid blastocysts.Limitations Reasons For Caution: This study confirms that Day-6 morulae should not be discarded in young women undergoing PGT-A. However, a potential drawback of biopsying embryos at the morula stage is the inability to distinguish between inner cell mass and trophectoderm cell origin. The sample size of euploid morula transfer cycles in this study was limited. Thus, a larger cohort would be beneficial to validate the reassuring live birth and spontaneous abortion rates reported here. Furthermore, the reproducibility of our findings should be determined at different centers.Wider Implications Of the Findings: Although Day-6 morulae are associated with higher aneuploidy rates and lower IRs compared to blastocysts, they still yielded successful pregnancies. Therefore, testing Day-6 morulae should be considered, especially for women <40 years old who are undergoing PGT-A with a small cohort of available blastocysts for biopsy.Study Funding/competing Interest(s): The authors have nothing to disclose. They received no specific funding for this work.Trial Registration Number: N/A. [ABSTRACT FROM AUTHOR]

Among infertile couples, 25% involve both male and female factors, while male factor alone accounts for another 25% due to oligo-, astheno-, teratozoospermia, a combination of the three, or even a complete absence of sperm cells in the ejaculate and can lead to a poor prognosis even with the help of assisted reproductive technology (ART). Intracytoplasmic sperm injection (ICSI) has been with us now for a quarter of a century and in spite of the controversy generated since its inception, it remains in the forefront of the techniques utilized in ART. The development of ICSI in 1992 has drastically decreased the impact of male factor, resulting in millions of pregnancies worldwide for couples who, without ICSI, would have had little chance of having their own biological child. This review focuses on the state of the art of ICSI regarding utility of bioassays that evaluate male factor infertility beyond the standard semen analysis and describes the current application and advances in regard to ICSI, particularly the genetic and epigenetic characteristics of spermatozoa and their impact on reproductive outcome. [ABSTRACT FROM AUTHOR]

Study Question: Is supraphysiologic estradiol (E2) an independent predictor of low birth weight (LBW) in singletons born after fresh IVF-embryo transfer (ET) cycles?Summary Answer: Our results suggest that E2 > 2500 pg/ml is an independent predictor for LBW in full-term singletons born to normal responder patients undergoing fresh IVF-ET cycles.What Is Known Already: The pathogenesis of LBW in IVF singletons remains unknown. However, recent studies have suggested that the hyperestrogenic milieu generated during ovarian stimulation may create a sub-optimal peri-implantation environment, leading to placental dysfunction, and therefore, LBW.Study Design, Size, Duration: Retrospective cohort study of normal responder patients, <40 years old, undergoing fresh IVF-ET cycles resulting in live singleton births between January 2005 and June 2014.Participants/materials, Setting, Methods: A total of 6419 patients had live births after fresh IVF-ET during the study period, of which 2348 (36.6%) patients were excluded due to multiple gestation, vanishing twins or incomplete records. Perinatal outcomes recorded for all patients included birth weight, gestational age (GA) at delivery, mode of delivery and gender. Term birth, preterm birth (PTB) and LBW incidence proportions were plotted against E2 level on the day of trigger. The term LBW incidence proportion (i.e. singletons born at GA ≥ 37 weeks with birth weight <2500 g) was considered the primary outcome of interest.Main Results and the Role Of Chance: A total of 4071 patients with live singleton births were included. The median age, BMI, E2 level and birth weight for the study cohort was 36 (33-39) years, 22.3 (20.4-25.0) kg/m2, 1554 (1112.7-2179) pg/ml and 3289 (2920-3628) g, respectively. The incidence proportion of LBW rose from 6.4% (E2 2001-2500 pg/ml) to 20.7% (E2 3501-4000 pg/ml), without a corresponding rise in the incidence proportion of PTB. The odds of term LBW with E2 > 2500 pg/ml were 6.1-7.9 times higher compared to the referent E2 group. Multivariable logistic regression analysis revealed that E2 was an independent predictor for term LBW, even after adjusting for age, BMI, race, parity, infertility diagnosis, duration of ovarian stimulation, gonadotropin dosage and method of insemination (adjusted odds ratio 10.8, 95% CI 9.2-12.5). Receiver operating characteristic analysis generated an AUC estimate of 0.85 for E2 level as a predictor of LBW.Limitations Reasons For Caution: The current study did not include analyses of hypertensive disorders of pregnancy or placental abnormalities. Furthermore, all patients were normal responders and of normal BMI, possibly limiting the overall generalizability of the study. Finally, as with any retrospective study, prospective data are required to validate the role of E2 in predicting LBW.Wider Implications Of the Findings: Our results emphasize the importance of minimizing the supraphysiologic elevations of E2 levels during ovarian stimulation in fresh IVF-ET cycles. This, in turn, can optimize the early peri-implantation environment and mitigate adverse perinatal outcomes such as LBW.Study Funding/competing Interest(s): Dr Paul J. Christos was partially supported by the following grant: Clinical and Translational Science Center at Weill Cornell Medical College (UL1-TR000457-06).Trial Registration Number: N/A. [ABSTRACT FROM AUTHOR]

Thiazolidinediones (TZDs) are insulin sensitizers used for treatment of diabetes. We have previously reported that TZDs reduce estrogen synthesis by inhibiting aromatase activity in human granulosa cells (HGC). Multiple clinical trials demonstrated that TZDs increase the risk of fractures in postmenopausal women with type 2 diabetes. We studied mouse osteoblasts alone or in a co-culture with HGC to determine whether TZD inhibition of aromatase plays a role in their effects on bone metabolism. Mouse osteoblasts were cultured with and without HGC, and incubated in a medium with or without testosterone, pioglitazone or rosiglitazone. Cell growth, oleic acid uptake, alkaline phosphatase activity, and osteocalcin production were measured. TZDs inhibited estradiol production by up to 84 % in HGC/mouse osteoblast co-cultures. TZDs induced mouse osteoblast death and increased oleic acid uptake. TZDs also inhibited alkaline phosphatase activity (58-75 %, p < 0.046) and osteocalcin production (52-75 %, p < 0.031). For all the parameters, there were no signifi cant differences between the osteoblast cultures alone and the HCG/osteoblast co-cultures. TZD effects on osteoblast viability, oleic acid uptake, alkaline phosphatase and osteocalcin production are independent of their effects on aromatase. [ABSTRACT FROM AUTHOR]

The eff ects of rosiglitazone or pioglitazone (thiazolidinediones, TZDs) on estrogen production and aromatase activity in human ovarian cells were examined. Human granulosa cells were incubated in the tissue culture medium supplemented with androstenedione or testosterone, with or without insulin, TZDs, or type 1 17 β -hydroxysteroid- dehydrogenase (17 β -HSD) inhibitor. Estrogen concentrations in the conditioned medium, aromatase mRNA and protein expression in the cells and androgen substrate binding to aromatase were measured. With androstenedione as substrate, rosiglitazone or pioglitazone inhibited estrone production by up to 22 % (p < 0.012) while type 1 17 β -HSD inhibitor enhanced this eff ect of rosiglitazone or pioglitazone by 37 % (p < 0.001) and by 67 % (p < 0.001), respectively. With testosterone as substrate, rosiglitazone or pioglitazone inhibited estradiol production by 32 % (p < 0.001). With 3 H-testosterone as substrate, rosiglitazone or pioglitazone inhibited the 3 H-tritiated water release by the cultured cells by 45 % and 35 % , respectively, thus directly demonstrating inhibition of aromatase. Rosiglitazone or pioglitazone, however, had no signifi cant eff ect on aromatase mRNA or protein expression. Rosiglitazone or pioglitazone inhibited 125 I-androstenedione and 125 I-testosterone binding to aromatase by 38 % (p < 0.001). It was concluded that rosiglitazone or pioglitazone inhibit estrogen synthesis in human granulosa cells by interfering with androgen binding to aromatase. [ABSTRACT FROM AUTHOR]

The article focuses on germ cell transplantation and stem-cell based therapy for spermatogenesis. The transplantation and the therapy is used for the treatment of impaired spermatogenesis. It examines different methods for manufacturing gametes. It describes different nuclear transplantation methods designed to induce haploidization of the somatic nucleus in the ootid to yield gametes. It also examines the recently acquired knowledge in the proliferation of in vivo-derived gonocvtes and their differentiation from ESC-derived primordial germ cells (PGCs).

Purpose: To analyze the success of autologous endometrial coculture (AECC) in improving embryo quality and pregnancy outcome based on the histologic characteristic of the biopsy. Methods: Prospective study of 86 consecutive patients undergoing IVF utilizing AECC. Results: The patients were on average 37.4 ± 4.0 years with a history of 2.6 ± 1.8 failed previous attempts. An overall clinical pregnancy rate of 45.3% per ET was found. The embryos grown in AECC were of an improved quality in comparison to those grown in conventional media. 33.7% (29/86) of the biopsies were out of phase (>3 days). In-phase (IP) and OOP (out of phase) specimens both demonstrated an improvement in embryo quality. However, OOP endometrial biopsies that displayed significant retarded endometrial development (< cycle day 19) did not demonstrate an improvement in embryos grown on AECC as compared to IP endometrial biopsies or OOP endometrial biopsies that demonstrated at least an endometrial development of cycle day 19. Conclusions: We have demonstrated a significant improvement in embryo quality with AECC. We have also demonstrated that histologic dating of the endometrium is predictive of IVF outcome when utilizing AECC. [ABSTRACT FROM AUTHOR]

Purpose: To examine the roles of Interleukin-1 (IL-l) and IL-1 receptor antagonist (IL-1ra), in in vitro embryo development and subsequent pregnancy outcome. Methods: Maternal serum utilized to supplement embryo growth in IVF cycles was analyzed for the presence of IL-1 cytokines. Results: The maternal serum that was utilized to supplement the embryo media was found to have measurable amounts of IL-1β and IL-1ra. Conclusions: Relative antagonism of the IL-1 system was positively associated with embryo development and pregnancy outcome. [ABSTRACT FROM AUTHOR]

Background: Breast cancer chemotherapy commonly causes premature ovarian failure and infertility. Because increased estrogen levels are thought to be potentially risky in breast cancer patients, natural cycle IVF (NCIVF) has been used to preserve fertility and treat infertility in these women.Methods: Twelve women with breast cancer received 40-60 mg tamoxifen for 6.9 +/- 0.6 days beginning on days 2-3 of their menstrual cycle (15 cycles), and had IVF (TamIVF) with either fresh embryo transfer (six cycles) or cryopreservation (nine cycles). They were compared to a retrospective control group (n = 5) who had natural cycle IVF (NCIVF, nine cycles).Results: Cycle cancellation was significantly less frequent in TamIVF, compared with NCIVF (1/15 versus 4/9, P < 0.05). Compared with NCIVF, TamIVF patients had a greater number of mature oocytes (1.6 +/- 0.3 versus 0.7 +/- 0.2, P = 0.03) and embryos (1.6 +/- 0.3 versus 0.6 +/- 0.2, P = 0.02) per initiated cycle. TamIVF resulted in the generation of embryo(s) in every patient (12/12) while only three out of five patients had an embryo following NCIVF. Two out of six patients in TamIVF, and 2/5 in NCIVF conceived. One patient in the TamIVF group delivered a set of twins. After a mean follow up of 15 +/- 3.6 months (range 3-54), none of the patients had a recurrence of cancer.Conclusions: Tamoxifen stimulation appears to result in a higher number of embryos and may provide a safe method of IVF and fertility preservation in breast cancer patients. [ABSTRACT FROM AUTHOR]

Context: In reproductive-age women, one of the common adverse effects of chemotherapy and radiotherapy is premature ovarian failure. In addition, a significant number of women experience early menopause due to oophorectomy performed for benign indications.Objective: To develop an ovarian transplantation technique to preserve endocrine function in women undergoing sterilizing radiotherapy and/or chemotherapy, or oophorectomy.Design and Setting: Case study of 2 patients in New York who received autologous ovarian transplantation (patient A, November 1999; patient B, April 2000) to the forearm prior to pelvic radiotherapy or after oophorectomy.Participants: Patient A is a 35-year-old woman with stage IIIB squamous cell cervical carcinoma and patient B is a 37-year-old woman with recurrent benign ovarian serous cysts.Main Outcome Measures: Follicular development evident by ultrasound examination; cyclical production of estradiol and progesterone; restoration of serum follicle-stimulating hormone, luteinizing hormone, and testosterone levels to nonmenopausal range; and disappearance of menopausal symptoms.Results: Menopause was confirmed immediately after the transplantation in both patients by serum follicle-stimulating hormone measurements (patient A, 47 mIU/mL; patient B, 50.7 mIU/mL). In patient A, follicle development was noted by physical and ultrasound examinations approximately 10 weeks after the transplantation. The mean (SE) follicle-stimulating hormone and luteinizing hormone levels decreased to 8.6 (0.4) mIU/mL and 12.8 (0.8) mIU/mL, respectively. The peripheral estradiol levels showed cyclical variation (mean [SE], 115 [9.2] pg/mL [422 (33.8) pmol/L), and during the 18-month follow-up, a dominant follicle developed each month. The estradiol levels from the right cubital vein were consistent with ovarian vein measurements (mean [SE], 1069 [269] pg/mL [3924 (987.5) pmol/L]). Percutaneous oocyte aspirations yielded a mature oocyte. In patient B, ovarian function was demonstrated by ultrasound visualization of a 9-mm follicle by 6 months after transplantation. Thereafter, the patient had spontaneous menstruation every 25 to 28 days. Ovulation was further confirmed by midluteal progesterone measurements (range, 7-10.1 ng/mL; mean [SE], 8.5 [0.9] ng/mL). Patient B's ovarian graft was still functional 10 months after the transplantation.Conclusions: Subcutaneous ovarian transplantation appears to be a relatively simple, novel technique to preserve endocrine function in women undergoing sterilizing cancer therapy or surgery. [ABSTRACT FROM AUTHOR]

Background: Ethical constraints limit the ability to study peri-implantation phase human endometrium. In this study, the donor oocyte model was used to study candidate endometrial markers of uterine receptivity.Methods: Archived, paraffin-embedded tissue obtained by endometrial biopsy during cycle days 21-23 of patients undergoing 'mock' hormonal treatment cycles were evaluated by standard histological criteria and immunohistochemical staining for alpha v beta 3 integrin and glycodelin. All of these patients (n = 101) had undergone a donor oocyte embryo transfer cycle utilizing the exact same hormonal protocol.Results: Histological evaluation revealed 62 (61.3%) in-phase, 34 (33.7%) dyssynchronous, 2 (2.0%) immature and 3 (3.0%) advanced endometria. The clinical outcomes of patients with either in-phase or dyssynchronous endometria were similar. Very strong correlations were noted between endometrial glandular dating and either alpha v beta 3 integrin or glycodelin immunostaining intensity (P < 0.001 for both). Glycodelin and alpha v beta 3 integrin immunostaining intensities were also highly correlated with each other (P < 0.001).Conclusions: Throughout the time period corresponding to the putative window of maximal endometrial receptivity (cycle days 21-23) a dynamic process was observed in exogenous hormonal replacement cycles characterized by a rapid histological advancement of endometrial glandular elements as well as progressive alpha v beta 3 integrin and glycodelin expression. [ABSTRACT FROM AUTHOR]

Transplanting a germinal vesicle (GV) from an aged woman's oocyte into a younger ooplasm has been proposed as a possible way to reduce the incidence of oocyte aneuploidy which is considered to be responsible for age-related infertility. In this study, we have assessed the efficiency of each step involved in nuclear transplantation—specifically cell survival, nuclear-cytoplasmic reconstitution, and the capacity of the reconstituted oocytes for in-vitro maturation. In addition, we have evaluated the fertilizability and karyotypic status of the manipulated oocytes by intracytoplasmic sperm injection (ICSI) and fluorescent in-situ hybridization technique respectively. Nuclear transplantation was accomplished with an overall efficiency of 73%. Due to the limited availability of materials, most nuclear transplantation procedures were performed between sibling oocytes. The maturation rate of 62% following reconstitution was comparable with that of control oocytes, as was the incidence of aneuploidy among the reconstituted oocytes. The ICSI results of the reconstituted oocytes yielded a survival rate of 77%, a fertilization rate of 52%, and a satisfactory early embryonic cleavage. Furthermore, in a limited number of observations where the nucleus of an aged oocyte was transferred into a younger ooplasm, there was an appropriate chromosomal segregation. These findings demonstrate that human oocytes reconstituted with GV nuclei are able to undergo maturation, fertilization, and early embryo cleavage, and maintain a normal ploidy. Although in-vitro maturation seems to be a limiting step, this technique would allow us to investigate further the nuclear-ooplasmic relationship during meiotic maturation. [ABSTRACT FROM PUBLISHER]

Historically, follicular stimulation protocols have included both FSH and LH in an attempt to mimic the physiology of normal human folliculogenesis. However, many recent gonadotrophin administration regimens have completely eliminated LH bioactivity. The importance and the amount of LH necessary for optimal follicular stimulation has been a topic of debate. Several recent studies have added to our understanding of the actions of androgens, oestrogens, gonadotrophins, and insulin on the follicle-oocyte unit, allowing a less speculative approach. Moreover, the availability of human gonadotrophins synthesized by recombinant DNA technology and gonadotrophin-releasing hormone (GnRH) antagonists, should soon permit a precise in-vivo assessment and re-evaluation of the historical 2-cell, two-gonadotrophin hypothesis. These pharmacological tools may also provide essential insights into the physiological roles of FSH and LH in human follicular development and oocyte maturation. The recombinant gonadotrophins give clinicians the unique opportunity to tailor ovarian stimulation regimens according to the patient's medical history, in an effort both to maximize oocyte yield and to improve oocyte quality. [ABSTRACT FROM AUTHOR]

Although great progress has been made in both the investigation and treatment of infertility, a considerable number of patients still fail to conceive. Spermatogenic failure and/or oocyte ageing appear to be responsible for a large proportion of cases. The use of donor gametes may bring legal, ethical and even social problems of acceptance that can discourage infertile couples from the donor route. Fortunately, emerging reproductive technologies and preliminary results from animal experiments provide some hope for alternative sources of gametes through which these infertile patients can finally conceive their own genetic child. In conjunction with intracytoplasmic sperm injection (ICSI), fertilization of human oocytes with immature sperm precursors, e.g. spermatids and even secondary spermatocytes, has resulted in healthy babies. Pregnancies have also resulted from the use of spermatids derived from in-vitro spermatogenesis. In the mouse, even primary spermatocytes appear able to participate in normal embryogenesis. In view of the possibility for transplantation and even xenotransplantation of spermatogonia to a host testis in animals, a similar use of human male stem cells might provide an attractive source for the treatment of males with arrested spermatogenesis, as well as male cancer patients. Transplantation of somatic cell nuclei and their haploidization within oocytes may prove to be a practical way of eradicating age-related aneuploidy and so constitute an innovative source of healthy oocytes. Most importantly, however, the safety of the procedures described here needs to be proven before their application to the human arena. Finally, we discuss the implications of cytoplasmic quality and of genetic imprinting in the context of these manipulations. [ABSTRACT FROM AUTHOR]

To assess the association of zona pellucida micromanipulation and subsequent development of monozygotic twins, cases of assisted embryo hatching (AH) and intracytoplasmic sperm injection (ICSI) were identified and related to treatment type, implantation and zygosity data. Embryology records from all patients undergoing in-vitro fertilization (IVF) at this centre from January 1995 to March 1998 were reviewed. In this study, 3546 transfer cycles were completed, with clinical pregnancy established in 1911 (54% per transfer) patients undergoing a single IVF cycle. These pregnancies occurred in 1674 (88%) IVF cycles, 120 (6%) donor oocyte cycles (DER), and 117 (6%) frozen embryo transfer (FET) cycles. During the study period, 23 cases of monozygotic (MZ) twins were identified, representing an overall frequency of 1.2%. Chorionicity was determined by transvaginal ultrasound at 7 weeks when the number of embryos transferred was less than the number of fetal heart-beats, or when >1 fetal heartbeat per gestational sac was seen. Zygosity was confirmed by placental evaluation at delivery, and corroborated the antenatal diagnosis in all cases. Among IVF study patients the frequency of MZ twinning was not statistically different between zona manipulated and zona intact subgroups. While this investigation is the largest to date describing the relationship between MZ twins and zona procedures, studies with even greater statistical power are needed to clarify it more precisely, particularly in DER and FET settings. A greater overall frequency of MZ twinning for IVF patients may be a function of the higher number of embryos transferred in IVF, rather than discrete zona manipulations. [ABSTRACT FROM AUTHOR]

Heat shock proteins (HSP) were first identified in cells after exposure to elevated temperature. Subsequently HSP have been identified as a critical component of a very complex and highly conserved cellular defence mechanism to preserve cell survival under adverse environmental conditions. HSP are preferentially expressed in response to an array of insults, including hyperthermia, free oxygen radicals, heavy metals, ethanol, amino acid analogues, inflammation and infection. HSP interact with intracellular polypeptides and prevent their denaturation or incorrect assembly. In addition HSP are also involved in several processes essential for cellular function under physiological conditions. HSP production is enhanced during in-vitro embryo culture and they are among the first proteins produced during mammalian embryo growth. The spontaneous expression of HSP as an essential part of embryo development is well documented and the presence or absence of HSP influences various aspects of reproduction in many species. Finally, HSP are immunodominant antigens of numerous microbial pathogens, e.g. Chlamydia trachomatis, which have been recognized as the main cause of tubal infertility. Many couples with fertility problems have had a previous genital tract infection, have become sensitized to microbial HSP, and a prolonged and asymptomatic infection may trigger immunity to microbial HSP epitopes that are also expressed in man. Antibodies to both bacterial and human HSP are present at high titres in sera and hydrosalpinx fluid of many patients undergoing in-vitro fertilization (IVF). In a mouse in-vitro embryo culture model, these antibodies impaired the mouse embryo development at unique developmental stages. Recent studies indicate an association between a previous infection, immunity to HSP and reproductive failure. [ABSTRACT FROM PUBLISHER]

Context: Sickle cell anemia is a common autosomal recessive disorder. However, preimplantation genetic diagnosis (PGD) for this severe genetic disorder previously has not been successful.Objective: To achieve pregnancy with an unaffected embryo using in vitro fertilization (IVF) and PGD.Design: Laboratory analysis of DNA from single cells obtained by biopsy from embryos in 2 IVF attempts, 1 in 1996 and 1 in 1997, to determine the genetic status of each embryo before intrauterine transfer.Setting: University hospital in a large metropolitan area.Patients: A couple, both carriers of the recessive mutation for sickle cell disease.Interventions: Standard IVF treatment, intracytoplasmic sperm injection, embryo biopsy, single-cell polymerase chain reaction and DNA analyses, embryo transfer to uterus, pregnancy confirmation, and prenatal diagnosis by amniocentesis at 16.5 weeks' gestation.Main Outcome Measure: DNA analysis of single blastomeres indicating whether embryos carried the sickle cell mutation, allowing only unaffected or carrier embryos to be transferred.Results: The first IVF attempt failed to produce a pregnancy. Of the 7 embryos analyzed in the second attempt, PGD indicated that 4 were normal and 2 were carriers; diagnosis was not possible in 1. Three embryos were transferred to the uterus on the fourth day after oocyte retrieval. A twin pregnancy was confirmed by ultrasonography, and subsequent amniocentesis revealed that both fetuses were unaffected and were not carriers of the sickle cell mutation. The patient delivered healthy twins at 39 weeks' gestation.Conclusion: This first unaffected pregnancy resulting from PGD for sickle cell anemia demonstrates that the technique can be a powerful diagnostic tool for carrier couples who desire a healthy child but wish to avoid the difficult decision of whether to abort an affected fetus. [ABSTRACT FROM AUTHOR]

The commercial availability of highly purified, s.c. administered urinary follicle stimulating hormone (FSH) preparations for ovarian stimulation marked the beginning of a new era in the treatment of infertility. As these new formulations contain essentially no luteinizing hormone (LH), supplemental LH may be needed for optimal folliculogenesis. It was the aim of this pilot study to compare fertilization rates, embryo morphology, implantation rates and pregnancy outcomes prospectively in two age-matched patient groups: women who received highly purified FSH (FSH-HP) (n = 17), and women who received FSH-HP plus recombinant human LH (rhLH, n = 14) throughout ovarian stimulation. All patients received mid-luteal pituitary down-regulation with s.c. gonadotrophin-releasing hormone agonist (GnRHa) (leuprolide). Mean implantation rates were 26.9 and 11.9% in the FSH-HP only and FSH-HP + rhLH groups respectively. The mean clinical pregnancy/initiated cycle rate was 64.7 and 35.7% for the FSH-HP only and FSH-HP + rhLH patients respectively. FSH-HP patients and FSH-HP + rhLH patients achieved clinical pregnancy/transfer rates of 68.8 and 45.5% respectively. One patient in the FSH-HP + rhLH group had a spontaneous abortion; no pregnancy losses occurred in the FSH-HP only group. There were more cancellations for poor ovarian response among FSH-HP + rhLH patients (n = 3) than among FSH-HP patients (n = 1). The trend toward better pregnancy outcomes among patients who received FSH-HP without supplemental rhLH did not reach statistical significance. It is postulated that appropriate endogenous LH concentrations exist despite luteal GnRHa pituitary suppression, thereby obviating the need for supplemental LH administration. [ABSTRACT FROM AUTHOR]

Transplanting a germinal vesicle (GV) to another enucleated oocyte provides a possible way to avoid age-related aneuploidy in metaphase II (MII) oocytes from older women. This study was conducted to examine the efficiency of each step of nuclear transplantation as reflected in the survival and maturation capacity of immature mouse oocytes subjected to this procedure. GV stage oocytes were retrieved from unstimulated ovaries. A GV removed with a small amount of cytoplasm (karyoplast) was transferred subzonally into a previously enucleated oocyte, which was then exposed to direct current to promote fusion. Such reconstituted oocytes were placed in culture to allow maturation, and some that had extruded a first polar body were fixed and processed for chromosome analysis. Each step of nuclear transplantation - survival, enucleation, grafting, and reconstitution - was successful in >90%, with the overall efficiency of reconstitution being 80%. The observation of normal karyotypes confirmed that the procedure did not increase chromosomal aneuploidy. An electrolytic medium, revealed to be superior for the reconstitution procedure, also allowed haploidization of the transplanted nucleus. These findings suggest that this technique can be applied to study the effects of a 'younger' woman's ooplasm on the disjunction of an 'older' woman's chromosomes during meiosis I. [ABSTRACT FROM AUTHOR]

The evident ability of the intracytoplasmic sperm injection (ICSI) procedure to achieve high fertilization and pregnancy rates regardless of semen characteristics has induced its application with spermatozoa surgically retrieved from azoospermic men. Here, ICSI outcome was analysed in 308 cases according to the cause of azoospermia; four additional cycles were with cases of necrozoospermia. All couples were genetically counselled and appropriately screened. Spermatozoa were retrieved by microsurgical epididymal aspiration or from testicular biopsies. Epididymal obstructions were considered congenital (n = 138) or acquired (n = 103), based on the aetiology. Testicular sperm cases were assessed according to the presence (n = 14) or absence (n = 53) of reproductive tract obstruction. The fertilization rate using fresh or cryopreserved epididymal spermatozoa was 72.4% of 911 eggs for acquired obstructions, and 73.1% of 1524 eggs for congenital cases; with clinical pregnancy rates of 48.5% (50/103) and 61.6% (85/138) respectively. Spermatozoa from testicular biopsies fertilized 57.0% of 533 eggs in non-obstructive cases compared to 80.5% of 118 eggs (P = 0.0001) in obstructive azoospermia. The clinical pregnancy rate was 49.1% (26/53) for non-obstructive cases and 57.1% (8/14) for testicular spermatozoa obtained in obstructive azoospermia, including three established with frozen-thawed testicular spermatozoa. In cases of obstructive azoospermia, fertilization and pregnancy rates with epididymal spermatozoa were higher than those achieved using spermatozoa obtained from the testes of men with non-obstructive azoospermia. [ABSTRACT FROM PUBLISHER]

The relationship between a previously undetected Chlamydia trachomatis infection, tubal infertility, immunity to heat shock proteins and subsequent in-vitro fertilization (IVF) outcome was evaluated. Women with tubal occlusion, with or without hydrosalpinges, and no history of C. trachomatis infection were tested for circulating antibodies to the human 60-kDa heat shock protein (Hhsp60), the C. trachomatis 10-kDa heat shock protein (Chsp10) and C. trachomatis surface antigens prior to their initial IVF cycle. Sera were obtained from 50 women whose male partners were infertile, 58 women with tubal occlusion but no hydrosalpinx and 39 women with tubal occlusions plus hydrosalpinx. Clinical pregnancies were documented in 68% of the women with male factor infertility. This was higher than the 43.1% rate in women with tubal occlusions (P = 0.04) and the 41% rate in women with hydrosalpinx (P = 0.02). C. trachomatis antibodies were present in one (2%) women with male factor infertility as opposed to 15 (25.9%) women with tubal occlusion (P = 0.003) and 13 (33%) with hydrosalpinx (P < 0.0001). Antibodies to Chsp10 were more prevalent in women with hydrosalpinx (46.8%) than in women with male factor infertility (P < 0.0001, 6%) or tubal occlusion (P = 0.0009, 15.5%). Hhsp60 antibodies were equally more prevalent in women with tubal occlusion plus (46.8%) or minus hydrosalpinx (41.4%) than in women with male factor infertility (P < 0.0002). Hhsp60 was more prevalent in those women positive for Chsp10 (P = 0.02) or C. trachomatis (P = 0.04) antibodies than in women lacking these antibodies. There was no relationship between any of the antibodies measured in sera and IVF outcome. [ABSTRACT FROM AUTHOR]

The aim of this study was to analyse the various reactions displayed by the oolemma to the penetrating pipette during intracytoplasmic sperm injection (ICSI) and correlate them with clinical factors, oocyte survival and fertilization patterns. Three types of oolemma responses were observed: normal breakage, when the injection needle created an invagination that ruptured at the approximate centre of the egg; sudden breakage, when the membrane broke without creating a funnel; and difficult breakage, when the membrane did not break or broke after several penetration attempts. A total of 2928 oocytes were analysed with the following observations: 73.9% (n = 2164) experienced normal breakage, 11.8% (n = 345) sudden breakage, and 14. 3% (n = 419) difficult breakage. The survival rate and number of normally fertilized oocytes were significantly lower and the incidence of digynic oocytes was significantly higher in the sudden breakage group; furthermore, in this group a significantly shorter length of stimulation was observed along with lower serum oestradiol concentrations when compared to oocytes experiencing normal and difficult breakage patterns. These recorded patterns were predictive of the survival and fertilization ability of the injected oocytes, as well as the incidence of digyny. The link between membrane behaviour and various clinical parameters appears to indicate a correlation between the modality of stimulation and oolemma characteristics. [ABSTRACT FROM AUTHOR]

To evaluate the role of endometrial thickness and pattern in in-vitro fertilization (IVF), these parameters were prospectively measured in 516 cycles of IVF with embryo transfer at our clinic. Pregnancy and embryo implantation rates were assessed for each mm of endometrial thickness and for each of three endometrial patterns. Embryo implantation, clinical and ongoing pregnancy rates were significantly higher in the patients with an endometrial thickness >9 mm (24.4, 48.6 and 42.2% respectively) compared with those of <9 mm (14.3, 16.0 and 11.7% respectively; < 0.005). Endometrial thickness was negatively influenced by age and positively influenced by oestradiol concentration. The majority of patients (69.8%) exhibited a ‘ring’ endometrial pattern. Embryo implantation and clinical pregnancy (statistically significant), as well as ongoing pregnancy rates (not statistically significant), were lower in patients exhibiting the ‘solid’ pattern. Endometrial thickness is independent of pattern in its effect on pregnancy outcome. In conclusion, endometrial thickness >9 mm as well as ring and intermediate endometrial patterns denoted a more favourable prognosis for pregnancy in IVF but thinner endometrium and those exhibiting a solid configuration had an acceptable pregnancy outcome. [ABSTRACT FROM PUBLISHER]

Six cases of identical twin pregnancies which occurred in 2163 cycles of in-vitro fertilization during a 3 year period are reported. Monozygosity was confirmed when the number of fetuses exceeded the number of embryos replaced (n = 4) or when two concepti were observed in a single amniotic sac (n = 2). Each of the reported pregnancies resulted from replacement of embryos either with naturally thin zonae pellucidae or embryos whose zonae had been breached during micromanipulation for assisted fertilization (subzonal sperm insertion) or assisted hatching. That such cases exclusively gave rise to monozygosity suggests a link between the physical state of the zona pellucida, hatching, and generation of identical twins. [ABSTRACT FROM AUTHOR]

Preimplantation genetic diagnosis was performed in 122 embryos obtained by IVF from 11 patients carriers of haemophilia, Duchenne's muscular dystrophy, Barth's syndrome, cystic fibrosis, Pelizaeus-Merzbacher syndrome or Rett's syndrome. After multiplex polymerase chain reaction (PCR) or fluorescence in situ hybridization (FISH) analysis with multiple probes, 28 embryos diagnosed as not affected were replaced. Of these, eight implanted (28%) and produced three ongoing pregnancies, three deliveries of four babies and a biochemical pregnancy. However, one case screened for cystic fibrosis was misdiagnosed and the pregnancy was terminated. In order to evaluate the efficiency of multiplex PCR, 55 non-replaced embryos were reassessed by PCR or by FISH. Identical results were obtained in all cases. However, one embryo which had only X-chromosome specific amplification by PCR was found to be XO in all its cells by FISH. Although multiplex PCR is demonstrated to be reliable for sexing of human embryos, FISH has the additional advantages of supplying ploidy assessment while not being affected by contamination. [ABSTRACT FROM AUTHOR]

A new technology called primer extension preamplification (PEP), which has been applied to single spermatozoa, increases the amount of polymerase chain reaction (PCR) templates by amplifying DNA of the whole genome. The current investigation was aimed at applying PEP to single human blastomeres. Two blastomeres with nuclei from arrested embryos were selected for this study. Using three different PEP protocols (experiments I, II and III), DNA from single blastomeres was amplified using 15-base oligonucleotide random primers. The efficiency of the procedure was determined by further amplifications of aliquots of the PEP products with two specific sequences. Three aliquots from each PEP product were used as PCR templates for the human X chromosome (X) or the exon 10 of the cystic fibrosis gene (CF). PCR amplified products were analysed by gel electrophoresis. In experiment I, when X primers were used, positive signals were detected in all 10 embryos (100%), 90.0% (18/20) of the blastomeres, and in 80.0% (96/120) of the replicates. When CF primers were amplified, all embryos (100%, 10/10), 90.9% (18/20) of the blastomeres and 78.3% (47/60) of the replicates were positive. In experiment II, efficiency was significantly reduced when total time for the procedure was minimized from 8 h to 5 h and 45 min. Although the time was further reduced to 4 h and 40 min in experiment III, the efficiency remained the same as in experiment I when the volume of PEP was reduced from 60 microliters (experiments I and II) to 40 microliters. One out of 132 control replicates (0.8%) was contaminated.(ABSTRACT TRUNCATED AT 250 WORDS) [ABSTRACT FROM AUTHOR]

Assisted hatching by zona drilling using acidic Tyrode's solution was performed during three randomized trials in 330 in-vitro fertilization patients. The trials were designed in order to study the overall effect of the procedure and whether characteristic patient [i.e. maternal age and basal levels of follicle stimulating hormone (FSH)] and embryonic features (i.e. zona pellucida thickness) are important for the decision to perform assisted hatching routinely. Couples ( = 137; Trial 1) in whom the female partner had normal basal FSH levels were randomized in a control group (without micro-manipulation) and a zona drilling group (all embryos micro-manipulated). The incidence of implantation (67/239; 28%) of zona-drilled embryos compared favourably with that of control embryos (49/229; 21%), but the difference was not significant. Retrospective analysis revealed that those embryos whose zonae were thicker than 15 μon were rescued. In order to test the validity of this finding, selective assisted hatching was performed on embryos with a poor prognosis in 163 other patients (Trial 2). The couples were randomized into a control group and a group in which embryos were selectively zona-drilled, based on zona thickness and other embryonic features. The rate of embryonic implantation in the selectively zona-drilled group was 25% (70/278), significantly ( < 0.05) higher than that of control embryos (51/285; 18%). Although it was demonstrated retrospectively and prospectively that assisted hatching by zona drilling is effective in embryos with thick zonae (≧$15 μm), patients whose embryos have thin (<13 μm) zonae may be jeopardized by the procedure. Selective assisted hatching appeared most effective in women over 38 and in those with elevated basal FSH levels. [ABSTRACT FROM PUBLISHER]

Our objective was to evaluate and compare the efficacy and mechanisms of co-culturing mouse embryos with Vero cells in both Dulbecco's modified Eagle's medium/Ham's F12 (DMEM/F12) and human tubal fluid (HTF) culture medium. Two-cell CB6F1 mouse embryos were cultured either in the presence of Vero cells (group A) or in culture medium alone (group B). In DMEM/F12 significantly more morulae developed in group A than in group B on day 3 (91 versus 23%; < 0.01). In contrast, the mouse embryos grew rapidly in HTF and significant differences were noted only in later embryonic stages (on day 5; 86% and 50%, < 0.01 of group A and B respectively, hatching or hatched). Similar experiments using DF1 and ICR mouse strains also revealed enhanced embryo development in the presence of Vero cells. To determine whether the embryo-enhancing effects of Vero cells were due to the removal of toxins or to the secretion of embryotrophic factors, ICR mouse embryos were cultured in fresh media with cells (group A), without cells (group B) and in cell-free conditions using cell-conditioned media which were obtained in the presence (group C) or absence (group D) of embryos. These results demonstrated that completion of hatching was highest (52%; < 0.01) in group A after 6 days in culture. There were no significant differences between groups B, C and D (rates of total hatching 18, 17 and 17%, respectively). It is concluded that Vero cells improve the development of mouse embryos and this is likely to be due to removal of substances inhibitory to development. The beneficial effects of Vero cells in HTF in later developmental stages may be due to the removal of toxins resulting from media degradation. [ABSTRACT FROM PUBLISHER]

The impact of intracytoplasmic sperm injection (ICSI) on cryopreserved zygotes and embryos was evaluated by comparing embryo survival and implantation between embryos derived from ICSI and those derived from standard insemination procedures. The study included patients whose excess zygotes and embryos were cryopreserved between September 1993 and December 1994 and who subsequently underwent a frozen embryo transfer. Embryo survival, clinical pregnancy rates per transfer and pregnancy outcome were compared. Three hundred and thirty eight cryopreservation cycles, during which 1471 embryos were cryopreserved, were included in this study. Of those, 961 were derived from oocytes fertilized by insemination in vitro and 510 were derived from oocytes fertilized by ICSI. A total of 690 of the embryos (451 in the insemination group and 239 in the ICSI group) have since undergone a thaw cycle. The embryo survival rates were similar between the two groups (70.5 and 73.2%, insemination and ICSI respectively) and were not significantly affected by the stage at cryopreservation. There was no significant difference in pregnancy rates per transfer (31.8 and 32.3%), the preclinical pregnancy loss rate (16.7 and 23.8%), or the clinical miscarriage rate (16.7 and 23.8%) between the insemination and the ICSI groups respectively. It is concluded that ICSI does not have an adverse impact on the survival and successful implantation of cryopreserved and thawed embryos. [ABSTRACT FROM PUBLISHER]

With the introduction of intracytoplasmic sperm injection (ICSI), couples with severe male factor infertility have achieved fertilization and clinical pregnancy rates comparable to other in-vitro fertilization (IVF) patients. However, failure of fertilization still occurs in some patients despite the utilization of microsurgical sperm injection techniques. How such fertilization failure after ICSI might impact later ICSI treatment(s) is unknown. In this investigation, couples with complete fertilization failure after ICSI treated from August 1993 to August 1996 were identified (index cycle, n = 21). Additionally, fertilization data from any previous or subsequent infertility treatments were evaluated. Seven patients (33%) had at least one IVF treatment before the index cycle, although no deliveries occurred. Of patients with complete fertilization failure in the index cycle, 48% (n = 10) underwent at least one subsequent ICSI cycle which proceeded to oocyte retrieval. The remainder (n = 11) elected to discontinue treatment. Although six subsequent cycles were cancelled due to poor follicular response (< or = 2 mature oocytes), all patients electing to continue treatment eventually achieved a subsequent embryo transfer. The clinical pregnancy rate per transfer was 45.4% for this group; the delivery and ongoing pregnancy rate per transfer was 36.3%. Review of semen parameters, superovulation characteristics or other clinical parameters during the three study cycles (pre-index, index, and post-index) was not prognostic of fertilization success or reproductive outcomes in later treatments. Fertilization failure with ICSI therefore could not be predicted by prior cycle performance, although total immotility of spermatozoa at time of oocyte retrieval, total teratozoospermia, and low oocyte yield were common characteristics of couples experiencing complete fertilization failure with ICSI. These findings suggest that fertilization failure in one ICSI cycle does not preclude successful fertilization and delivery in a later ICSI treatment. [ABSTRACT FROM AUTHOR]

Electroejaculation has been successfully used for sperm procurement in anejaculatory men desiring fertility. However, electroejaculates typically must have normal sperm numbers but poor motility, morphology, and functional deficiencies. Here we report the pregnancy outcome of a series of couples undergoing combined electroejaculation and in-vitro fertilization (IVF) with intracytoplasmic sperm injection (ICSI). In all, 13 couples underwent a total of 18 cycles. The aetiologies of anejaculation included history of retroperitoneal lymph node dissection for testicular cancers, spinal cord injury and psychogenic causes. ICSI was performed on 192 oocytes, resulting in a fertilization rate of 75.5%. A total of 15 embryo transfers were performed using a total of 51 embryos. Clinical pregnancy rate, as defined by positive fetal heart rate(s) using vaginal sonography, was 55.6% per retrieval; implantation rate was 33.3% per embryo. These rates appear to be similar to those obtained in standard IVF for non-male factor infertility, or ICSI for male factor infertility. The use of ICSI for electroejaculates undoubtedly provides these couples with the highest chance of pregnancy. [ABSTRACT FROM PUBLISHER]

Two-cell mouse embryos (B6D2F1) were cultured in the presence or absence of 100 μg/ml monoclonal antibodies specific for the mammalian 60 kDa (HSP60), 70 kDa (HSP70) and 90 kDa (HSP90) heat shock proteins. Embryo development was evaluated after 3, 5 and 7 days in culture by determining the number of blastocysts, hatched blastocysts and outgrown trophoblasts at the successive time points. At day 3, only 29% (22/75) of the embryos cultured with anti-HSP60 antibody developed to the blastocyst stage (P < 0.0001) as compared to 67% (31/46) of the embryos cultured with anti-HSP70, 72% (43/60) cultured with anti-HSP90, and 79% (49/62) in medium plus mouse IgG1. By day 5, hatched embryos were present in 28% (13/46) of the cultures containing anti-HSP70 (P < 0.0001), as opposed to 57% (34/60) containing anti-HSP90 and 73% (45/62) containing IgG1. At day 7, outgrown trophoblasts were observed in 9% (4/46) of cultures containing anti-HSP70 (P < 0.0001), 45% (27/60) containing anti-HSP90 (P < 0.01) and 66% (41/62) cultured in medium plus IgG1. Antibodies to different heat shock proteins exerted a detrimental effect on mouse embryo development at unique development stages. Immune sensitization to heat shock proteins may be a cause of reproductive failure. [ABSTRACT FROM PUBLISHER]

The purpose of this study was to investigate any influence of maternal and/or paternal age on gamete characteristics and pregnancy outcomes in intracytoplasmic sperm injection (ICSI) cycles. In all, 821 consecutive ICSI cases were analysed retrospectively. While a significant linear decline in semen volume was detected, no significant differences in the concentration, motility or morphology of the spermatozoa were found with paternal ageing. A significant decline in the number of oocytes retrieved and the number of mature oocytes obtained was found with advancing maternal age. An increase in the occurrence of digyny was noted with parental ageing, while no difference in single or bipronuclear fertilization was found. Older women had a decreased incidence of single pronucleus formation and an increase in digyny, but no significant difference in the percentage of oocytes that underwent two-pronuclear fertilization was detected with regard to maternal ageing. Pregnancy outcomes were not influenced by the age of the male partner, while a strong negative correlation was found with maternal ageing. To better analyse male partner ageing as a factor affecting pregnancy outcome, we analysed a subgroup of patients with a female partner aged <35 years who underwent ICSI. No paternal influence on ICSI pregnancy outcome was found in this subgroup of patients. We conclude that the influence of pregnancy outcome after ICSI is related mostly to maternal and not paternal age. [ABSTRACT FROM PUBLISHER]

Certain patients have a tendency for high response to gonadotrophin therapy which is often not ameliorated with prior gonadotrophin-releasing hormone agonist (GnRHa) suppression. As a result, these patients are frequently cancelled and often experience ovarian hyperstimulation syndrome (OHSS) episodes during in-vitro fertilization (IVF)–embryo transfer cycles. Patients with polycystic ovarian syndrome (PCOS) have been noted to be particularly sensitive to exogenous gonadotrophin therapy. We have developed a protocol which is effective in improving IVF outcome in high responder patients, including those with PCOS. Oral contraceptive pills (OCP) are taken for 25 days followed by s.c. leuprolide acetate, 1 mg/day, which is overlapped with the final 5 days of oral contraceptive administration. Low-dose gonadotrophin stimulation is then initiated on the third day of withdrawal bleeding in the form of either human menopausal gonadotrophins or purified urinary folliclestimulating hormone at a dosage of 150 IU/day. Over a 5 year period, we reviewed our experience utilizing this dual method of suppression in 99 cycles obtained in 73 high responder patients. There were only 13 cancellations prior to embryo transfer (13.1%). The clinical and ongoing pregnancy rates per initiated cycle were 46.5 and 40.4% respectively. Only eight patients experienced mild-moderate OHSS following treatment. For those patients who had undergone previous IVF–embryo transfer cycles at our centre, significant improvements were noted in oocyte fertilization rates, embryo implantation rates and clinical/ongoing pregnancy rates with this protocol. Hormonal analyses revealed that the chief mechanism may be through an improved luteinizing hormone/follicle-stimulating hormone ratio following dual suppression. An additional feature of this dual method of suppression is significantly lower serum androgen concentrations, particularly dehydroepiandrosterone sulphate. [ABSTRACT FROM PUBLISHER]