Standard Intracytoplasmic Sperm Injection Research Articles

Low birth defects by deselecting abnormal spermatozoa before ICSI

Consistent evidence from meta-analysis has linked assisted conception by IVF, and particularly intracytoplasmic sperm injection (ICSI), with an increased risk of major birth defects. To compare the risk of major malformations of children born after standard ICSI and after intracytoplasmic injection of morphologically selected spermatozoa (IMSI), a prospective population-based study was conducted from 2005 to 2010. ICSI and IMSI were performed in only one assisted reproduction unit according to its classification of spermatozoa and using fresh semen. Medical data and follow up during 2 years of 1028 infants were collected. Major malformations were identified and classified by an external independent physician. The two groups were similar concerning the parents’ age, treatment, number of oocytes recovered, days of transfer, gestational age and birthweight. However, major malformations were significantly lower with IMSI (6/450, 1.33%) versus ICSI (22/578, 3.80%; adjusted odds ratio 0.35, 95% confidence interval 0.14–0.87, P=0.014), mainly affecting boys (adjusted odds ratio 2.84, 95% confidence interval 1.24–6.53, P=0.009). In conclusion, the significantly decreased risk of major birth defects associated with IMSI remained decreased after multivariate adjustment and highlights the beneficial effect of sperm selection before ICSI.The aim of this study was to compare the birth defects in children born after intracytoplasmic sperm injection (ICSI) and intracytoplasmic morphologically selected sperm injection (IMSI). We conducted a prospective population-based study from 2005 to 2010. Medical data and follow up at 2 years of age for 1028 infants were collected. We show that the major malformations were significantly lower in the IMSI group versus the ICSI group, and they mainly affected the boys. We found less abnormality when abnormal spermatozoa, according to the Cassuto Barak classification, were deselected at high magnification and not injected. In conclusion, the significantly decreased risk of major birth defects associated with IMSI remained decreased after multivariate adjustment and highlights the beneficial effect of spermatozoon selection before ICSI.

Perinatal outcomes in 375 children born after oocyte donation: a Danish national cohort study

To describe perinatal outcomes in children born after oocyte donation (OD) compared with in vitro fertilization (IVF), intracytoplasmic sperm injection (ICSI), and spontaneous conception (SC). National cohort study. Fertility clinics. Three hundred seventy-five children born after OD during the period 1995-2010. None. Mean birth weight, mean gestational age, risks of low birth weight (LBW), preterm birth (PTB), congenital malformations, cesarean delivery, preeclampsia, and admittance to neonatal intensive care unit. We found an increased risk of PTB in OD pregnancies. The adjusted odds ratio (AOR) of PTB in OD singletons was 1.8 (95% CI, 1.2-2.69), 2.5 (95% CI, 1.7-3.6), and 3.4 (95% CI, 2.3-4.9) compared with IVF, ICSI, and SC, respectively. The risk of LBW was also increased. The AOR of LBW was 1.4 (95% CI, 0.9-2.2), 1.8 (95% CI, 1.2-2.8), and 2.6 (95% CI, 1.7-4.0) compared with IVF, ICSI, and SC. The risk of preeclampsia was increased in OD pregnancies with an AOR of 2.9 (95% CI, 1.8-4.6), 2.8 (95% CI, 1.7-4.5), and 3.1 (95% CI, 1.9-4.9) compared with IVF, ICSI, and SC. After additional adjustment for preeclampsia, perinatal outcome improved. Among the twins, the difference between the groups was less pronounced. Pregnancies after OD have a poorer perinatal outcome than those after standard IVF and ICSI mainly because of the high prevalence of preeclampsia.

Frozen-thawed spermatozoa from oligozoospermic ejaculates are susceptible to in situ DNA fragmentation in polyvinylpyrrolidone-based sperm-immobilization medium

To elucidate the effect of sperm immobilization media that are and are not based on polyvinylpyrrolidone (PVP) on the DNA integrity of fresh and frozen-thawed spermatozoa during standard intracytoplasmic sperm injection (ICSI) conditions. Experimental prospective study. Embryology research laboratory. Forty-six ejaculates from normozoospermic and oligozoospermic men. Assessment of sperm DNA fragmentation by single-cell gel electrophoresis assay. DNA integrity of fresh and frozen-thawed spermatozoa from normozoospermic and oligozoospermic ejaculates exposed to PVP-based and non-PVP-based media. Exposure of fresh and frozen thawed spermatozoa from normozoospermic and oligozoospermic ejaculates to PVP-based medium in an ICSI dish for 30 minutes statistically significantly increased the DNA fragmentation. In contrast, the extent of DNA fragmentation in non-PVP-based medium did not statistically significantly differ from control. A PVP-based medium can induce a statistically significant amount of sperm DNA fragmentation in an ICSI dish, and frozen-thawed sperm from oligozoospermic ejaculates are more susceptible to in situ DNA fragmentation.

The benefit of artificial oocyte activation is dependent on the fertilization rate in a previous treatment cycle

Following intracytoplasmic sperm injection (ICSI), some patients present low or zero fertilization rates. Artificial oocyte activation has been proposed as a suitable means to overcome this problem. This study applied artificial oocyte activation in patient cohorts with a history of no fertilization (0%, group 1), fertilization between 1 and 29% (group 2) or fertilization between 30 and 50% (group 3) in initial ICSI cycles. In the following treatment cycles, oocytes were activated after ICSI using calcium ionophore. Fertilization, pregnancy and take-home baby rates were compared with the previous cycle without activation. In group 1, fertilization rate was 41.6%, embryos for transfer were available in 82.1% of cycles, giving a clinical pregnancy rate of 18.8% and take-home baby rate of 12.8%. In group 2, despite a lower transfer rate (87.9% versus 100%, P<0.05), there were higher fertilization and clinical pregnancy rates (44.4% versus 19.3% and 31.4% versus 12.8%, respectively, P<0.05) and take-home baby rate was 24.1% versus 12.8%. In group 3, fertilization rates differed (56.1% versus 36.8%; P<0.001) but all other parameters were similar. Artificial oocyte activation has great potential especially in patients showing compromised fertilization rates below 30% after standard ICSI.Following intracytoplasmic sperm injection (ICSI), some patients present very low or even zero fertilization rates after ICSI. Artificial oocyte activation has been proposed as a suitable means to overcome this problem. We applied artificial oocyte activation in patients which presented a history either no fertilization, fertilization between 0 and 30% or fertilization between 30 and 50% in initial ICSI cycles. In the following treatment cycles, oocytes were activated after ICSI using a calcium ionophore. Fertilization, pregnancy and take-home baby rates were compared to the previous cycle without activation. For the groups with previously 0% or 1–29% fertilization, we noted higher fertilization rates and clinical pregnancy rates per embryo transfer. For the group with moderate fertilization, only fertilization rates differed but all other parameters were not significantly different. From these data we conclude that artificial oocyte activation has a great potential especially in patients which show a compromised fertilization rate below 30% in a standard ICSI cycle.

188 HUMAN FOLLICULAR FLUID IN EQUINE IN VITRO MATURATION MEDIUM SUPPORTS IN VITRO MATURATION AND VIABLE INTRACYTOPLASMIC SPERM INJECTION-BLASTOCYST PREGNANCIES

The aim of this study was 2-fold: (1) To determine if altrenogest-treated mares will yield higher numbers of quality in vitro-matured (IVM) oocytes than early pregnant mares and cycling/control mares and (2) if the addition of human follicular fluid (HFF) to IVM medium can support IVM and viable pregnancies from in vitro-produced blastocysts. In this study, 18 mares were assigned to 3 equally sized treatment groups and each mare was subjected to follicle aspiration every 10 to 11 days without monitoring follicular growth. The 3 treatment groups were altrenogest-treated mares (0.044 mg kg–1 of PO daily), early pregnant mares (30–110 days) and control/cycling mares. Using transvaginal ultrasound guidance, all visible follicles were aspirated. Altrenogest-treated mares each yielded more follicles (8.75) per aspiration session when compared with the control mare group (5.75) and the pregnant mare group (3.72), but there was no difference in oocyte recovery rates among the groups (Table 1). A limited number of these oocytes were subjected to in vitro maturation. After heated (38.5°C) transport of oocytes to an off-site laboratory, the oocytes were placed in maturation medium containing 10% HFF obtained from preovulatory follicles after ovulation induction, 20% serum substitute supplement and no hormones for 36 h. This approach yielded a maturation rate of 61.8, 68.8 and 82.0% for the altrenogest, pregnant and control treatment groups, respectively (not significant). Mature oocytes (n = 65) were injected with frozen-thawed sperm using a standard intracytoplasmic sperm injection (ICSI) technique. Four expanding blastocysts (Table 1) were selectively transported back to the embryo transfer facility and transcervically transferred into recipient mares on Day 6 post-ICSI. These 4 transfers resulted in 2 viable, normally progressing pregnancies, ongoing beyond 60 days of gestation. Both pregnancies resulted from the altrenogest-treated aspiration group. In this study we concluded that (1) altrenogest-treated mares provide more follicles and may be a better source of viable immature oocytes for the production of ICSI embryos and foals, but their overall advantage is unclear; (2) addition of HFF to IVM media, in the absence of added gonadotropins, can support oocyte maturation, blastocyst production and viable pregnancies; (3) an aspiration schedule of every 10 to 11 days without ultrasonic monitoring can yield viable immature oocytes, capable of producing ICSI blastocysts, resulting in viable pregnancies. Table 1.Altrenogest-treated mares compared to early pregnant mares and control mares

Management of endometrial polyps diagnosed before or during ICSI cycles

This retrospective study aimed to shed light on the management options of endometrial polyps diagnosed before or during intracytoplasmic sperm injection (ICSI) treatment. The study included all fresh ICSI cycles performed in the Anatolia IVF Center between July 2005 and January 2009. Group 1 consisted of 47 patients who were diagnosed with an endometrial polyp before their ICSI cycle. All patients diagnosed with an endometrial polyp by transvaginal ultrasonography before the ICSI cycle underwent hysteroscopic polyp resection. Group 1 was compared with 47 matched control patients without endometrial polyps who underwent standard ICSI cycles (group 2). Group 3 included 128 patients diagnosed with an endometrial polyp during stimulation in their ICSI cycles. Group 3 was compared with 128 matched control patients without endometrial polyps who underwent standard ICSI cycles (group 4). Patients diagnosed with an endometrial polyp before ICSI cycles were similar to their controls with regard to clinical pregnancy (29.8% versus 38.3%) and live-birth (25.5% versus 31.9%) rates per transfer, as were patients diagnosed with an endometrial polyp during ovarian stimulation (clinical pregnancy rates 45.3% versus 46.9%; live-birth rates 40.6% versus 39.8%). In conclusion, further studies are required to identify the most appropriate management of endometrial polyps. In our retrospective study, we aimed to shed light on management options of endometrial polyps diagnosed before or during intracytoplasmic sperm injection (ICSI) treatment. This study included all fresh ICSI cycles performed at the Anatolia IVF Center between July 2005 and January 2009. Group 1 consisted of 47 patients who were diagnosed with an endometrial polyp before their ICSI cycle. All patients diagnosed with an endometrial polyp by transvaginal ultrasonography before the ICSI cycle underwent hysteroscopic polyp resection. Group 1 was compared was compared to 47 matched control patients without endometrial polyps who underwent standard ICSI cycles (group 2). Group 3 included 128 patients diagnosed with an endometrial polyp during stimulation in their ICSI cycles. Group 3 was compared to 128 matched control patients without endometrial polyps who underwent standard ICSI cycles (group 4). Patients diagnosed with an endometrial polyp before ICSI cycles were similar to their controls with regard to pregnancy rates and outcomes. Patients diagnosed with an endometrial polyp during ovarian stimulation were similar to their controls with regard to pregnancy rates and outcomes. In conclusion, endometrial polyps <1.5 cm occurring during ovarian stimulation do not require any intervention. Resection of an endometrial polyp <1.5 cm detected before an ICSI cycle is not required and might be even harmful.

Health of IVM children

In vitro maturation (IVM) is an emerging type of assisted reproductive technology (ART), in which immature as opposed to mature oocytes are collected from the ovaries and allowed to mature in vitro prior to fertilisation by either standard in vitro fertilisation (IVF) on intracytoplasmic sperm injection (ICSI). Since the first report of a human birth resulting from IVM in 1991 [1], it is estimated that over 1,300 babies have been born following the technique [2] (in contrast to an estimated 3 million babies conceived by standard IVF and ICSI over the last 30 years). IVM offers a number of potential benefits when compared to standard IVF, principally related to the avoidance of high doses of hormonal stimulation with gonadotrophins and GnRH analogues. In patients with polycystic ovary syndrome (PCOS) this avoids the risk of ovarian hyperstimulation syndrome (OHSS). The high costs of hormonal therapy also make IVM cheaper than conventional alternatives. Immature oocyte retrieval, without the need to wait for hormonal stimulation and in vivo maturation, offers the possibility of expedient oocyte cryopreservation for patients diagnosed with cancers without delaying cancer treatment. However, despite recent improvements, pregnancy rates with IVM remain lower than with conventional IVF, and higher miscarriage rates have been reported with IVM [2]. For these reasons IVM has yet to become widely used in the field of ART. Given that IVM is not yet a mainstream ART technique, it is not surprising that there is very limited follow-up data regarding the health of children conceived following its use. This is in contrast to a significant volume of high quality studies that have followed-up children conceived following standard IVF and ICSI. In this article we discuss hypothetical concerns regarding the health of IVM children, and review the few studies published in this area. Health of IVF & ICSI children High multiple pregnancy rates continue to be the single greatest factor adversely impacting upon the outcome of ART pregnancies [3, 4], with resulting increased risk of preterm delivery, low birth weight, and perinatal mortality. However, evidence regarding the health of ART singletons when compared to spontaneously conceived singletons is generally reassuring. The majority of studies investigating the risk of congenital malformations, neurodevelopmental outcome, physical health, growth, and psychosocial well-being in ART children are reassuring [5, 6]. Concerns have been raised that ART children may exhibit higher rates of cancer [7] and rare imprinting disorders [8] (such as Beckwith-Wiedemann Syndrome and Angelman Syndrome), however larger studies are required to confirm these associations. Risks of infertility and cardiovascular disease in later life remain unexplored areas of concern and require follow-up of cohorts of ART children to continue into adulthood.

A sperm viability test using SYBR-14/propidium iodide flow cytometry as a tool for rapid screening of primary ciliary dyskinesia patients and for choosing sperm sources for intracytoplasmic sperm injection

Spermatozoa viability tests based on dual-color flow cytometry after staining with Sybr-14/propidium iodide were performed on 44 men with complete asthenospermia for primary ciliary dyskinesia (PCD) screening, and seven were identified with PCD by electron microscopy of ultrastructural ciliary defects. Six PCD patients underwent eight intracytoplasmic sperm injection therapy cycles using ejaculated sperm or testicular sperm, obtaining a mean fertilization rate of 46.6%, with three healthy babies born and one in utero at the time of writing.

Embryo yield after in vitro fertilization in women undergoing embryo banking for fertility preservation before chemotherapy

To evaluate embryo yield after IVF in patients undergoing embryo banking before chemotherapy. A retrospective cohort study. Hospital-based academic medical center. Thirty-eight women diagnosed with cancer or autoimmune disease presenting for IVF cycles, with or without intracytoplasmic sperm injection (ICSI), for embryo cryopreservation before any therapy were compared with 921 presumably fertile women undergoing IVF for male factor infertility from January 2001 through October 2007. Standard IVF or ICSI protocol, embryo freezing, and ET. The number of 2 pronuclear (2PN) embryos created and suitable for cryopreservation or transfer. No statistically significant differences were observed between preservation and male factor groups for number of embryos, number of oocytes, or amount of gonadotropin needed to stimulate follicular development. Peak serum E(2) levels were significantly lower for women with disease-seeking fertility preservation. Women facing chemotherapy as treatment for cancer or systemic autoimmune disease infrequently undergo fertility preservation. If offered this potentially fertility-preserving option, these data suggest equivalent embryo yield compared with women with infertile male partners. Our data report no significant complications in subsequent births in those who sought fertility preservation, which is informative and encouraging for these women and their providers.

Congenital malformations after assisted reproduction: risks and implications for prenatal diagnosis and fetal medicine

Congenital malformations after assisted reproduction: risks and implications for prenatal diagnosis and fetal medicine

The effectiveness of intracytoplasmic sperm injection combined with piezoelectric stimulation in infertile couples with total fertilization failure

To assess the effectiveness of intracytoplasmic sperm injection (ICSI) combined with piezoelectric stimulation in infertile couples with a history of total fertilization failure (TFF). Prospective controlled trial. Clinical IVF laboratory. Seventy-one couples undergoing ICSI on sibling oocytes having at least one previous ICSI attempt with TFF. ICSI or ICSI with piezoelectric activation. Fertilization rate. The patients were allocated to two groups: group I included 21 patients with only one previous TFF and group II included 50 patients with more than one previous TFF. Collectively, a total of 823 metaphase II (MII) oocytes were retrieved in 78 oocyte retrievals. In Group I, combined ICSI with piezoelectric stimulation was applied to 123/211 (58.2%) of MII oocytes (group IA), whereas standard ICSI procedure was applied to 88/211 (41.8%) of MII oocytes (group IB). The fertilization rate was 62% and 12% in group IA and group IB respectively. In group II, piezoelectric activation was applied in all 612 MII oocytes, of which 296 (48.3%) were fertilized. The rates for implantation and pregnancy/embryo transfer were obtained as 30.6% and 44.1%, respectively. Piezoelectric activation seems to improve IVF outcome in patients with previous TFF history.

Developmental and Genetic Outcomes in Children Conceived Through Assisted Reproductive Technologies

After completing this article, readers should be able to: 1. Describe the outcomes of assisted reproductive technologies (ART) for singleton, twin, and other multiple births. 2. Describe the role of fertility in adverse outcomes seen with ART births. 3. Review the association of birth defects with ART. 4. Delineate the association of disease of genomic imprinting with ART. 5. Describe the relationship between ART and the subsequent incidence of neurodevelopmental sequelae. In the 1977 ruling “Carey v. Population Services International,” the United States Supreme Court ruled that the decision to bear children is constitutionally protected. (1) Significant interest already had been shown in the development and improvements of in vitro fertilization (IVF) for infertile couples. The first human pregnancy and human birth using IVF were reported by Steptoe and Edwards in the United Kingdom. (2) Their work resulted in the first baby born via reproductive technologies, Louise Brown, born on July 25, 1978, at Oldham General Hospital in Oldham, United Kingdom. (3) She was born via a planned cesarean section, and her birthweight was 2.61 kg. The first successful viable IVF in the United States was performed by Jones and Seager-Jones in 1981 in Norfolk, Virginia. (4) Assisted reproductive technologies (ART) have seen a recent surge in popularity. The Centers for Disease Control and Prevention (CDC) reported that 122,872 cycles of ART were initiated in 2003, resulting in the delivery of 48,756 neonates, (5) accounting for approximately 1% of all neonates delivered in the United States. The percentage is higher in many countries, including Denmark, where it is estimated that 5% of all deliveries are with the assistance of ART. (6) Couples pursue ART for myriad reasons, including tubal transport factors, ovulatory dysfunction, uterine factors, endometriosis, male- and female-specific factors, and when a cause of infertility is unknown. (5) It would be very …

P-996: Relationship among HBA, sperm motility, Kruger morphology with fertilization rate and pregnancy outcome in patients treated with IVF or ICSI

Semen analysis with Kruger morphology is widely used to assess male fertility potential. More recently hyaluronan-binding protein has been established as a biomarker for mature, functional sperm. Consequently the HBA assay (Sperm-Hyaluronan Binding assay) has been developed to differentiate functional integrity and fertilization potential of sperm cells. We investigated the relationships among HBA score, sperm motility, and Kruger morphology with fertilization rate in patients undergoing in vitro fertilization (IVF) with standard insemination or intracytoplasmic sperm injection (ICSI). Prospective comparative data analysis. The Andrology and Embryology laboratories at the Kaiser Permanente Center for Reproductive Health performed HBA assay in conjunction with a standard semen analysis with Kruger morphology for patients undergoing IVF. The decision to perform standard IVF or ICSI was made by the primary physicians and the patients. Sperm HBA score, sperm motility and Kruger morphology before and after density gradient preparation were compared and correlated with fertilization rate of oocytes inseminated with standard insemination or ICSI. Linear regression analysis was performed. We studied prospectively 31 cases in which 20 cases were treated with ICSI, 7 cases were treated with IVF, and 4 cases were performed split IVF/ICSI. In the ICSI treatment group, the fertilization rate had no correlation with sperm motility, Kruger morphology, or HBA score (r = -0.15, -0.14, and 0.07 respectively). We achieved high fertilization rates (50 - 100%) in the ICSI treatment group even when the sperm HBA score was less than 10 either before or after density gradient preparation. In general, density gradient preparation of sperm improved its motility and morphology. However, it did not necessarily improve the HBA score. In fact, some sperm samples after density gradient preparation dropped in HBA score dramatically. In the IVF treatment group, there was a positive trend between fertilization rate and initial sperm motility and HBA score. The fertilization rate, as previously suggested by Kruger et al, showed little correlation (r = -0.15) with Kruger morphology when the later score was greater than or equal to 4. HBA assay, sperm motility and Kruger morphology have all been suggested as indicators for differentiating male fertility potential. Based on our data, we can not recommend any one of them individually as a discriminating factor for IVF or ICSI decision making except when Kruger morphology score is ≤ 4. Our results demonstrate a reduction of HBA score after density gradient sperm preparation of some sperm populations. This observation raises a concern about potential damage of membrane integrity and/or unintended acrosome reaction during sperm preparation, both of which may lead to compromised fertilization potential. Future studies are needed to explore the utility of HBA assay in assessing sperm functional integrity after processing.

P-249: Successful clinical pregnancy following the transfer of embryos frozen and thawed twice: A case report

To report the ongoing clinical pregnancy of an IVF patient following the transfer of embryos frozen/thawed at the two pronucleus stage (2PN) then cultured to day 5 and frozen/thawed again at the blastocyst stage. Case Report. An IVF cycle was performed using a normal ovarian stimulation protocol with GnRH agonist down regulation, recombinant FSH stimulation and recombinant βhCG ovulation-induction. Half of oocytes retrieved underwent standard insemination and the remaining half was fertilized by intracytoplasmic sperm injection (ICSI). Embryo culture was performed using Quinn’s Advantage Cleavage Media (QAC) supplemented with 10% v/v Serum Substitute Supplement (SSS) in a humidified atmosphere of 5% CO2, 5% O2 and 90% N2 at 37° C. All resulting 2PN’s were cryopreserved using a standard propanediol and sucrose slow freezing protocol in a controlled-rate liquid nitrogen (LN2) freezer. 2PN’s were thawed with a multi-step thaw protocol in sucrose solutions at room temperature. The zygotes were then cultured again in QAC Medium for 48 hrs under similar culture conditions. Blastocyst culture was performed using Quinn’s Blastocyst Medium also in a humidified low oxygen environment at 37°C to Day 5 and Day 6. Blastocyst cryopreservation was performed on Day 6 on all remaining blastocysts of grade 3BB or better using a two-step slow-freezing protocol (Menezo) with glycerol and sucrose in a controlled rate LN2 freezer. Blastocysts were thawed with a two-step thaw protocol in sucrose solutions (Menezo). Recovery and re-expansion was conducted in Quinn’s Blastocyst Medium supplemented with 20% SSS in a humidified low oxygen environment at 37°C for two hours before embryo transfer. Laser assisted hatching was performed on all frozen embryos transferred. A 34-year-old female (gravida 0) with a diagnosis of polycystic ovarian disease and BMI of 37.6 underwent controlled ovarian hyperstimulation for IVF. The cycle resulted in 29 oocytes which were split between standard insemination and ICSI. 17- 2PN’s, 8 from standard insemination and 9 from ICSI, were observed on Day 1 all of which were cryopreserved due to risks associated with ovarian hyperstimulation syndrome. Since then, three FET cycles followed. First 5 zygotes from the IVF group were thawed yielding the transfer of 2 cleavage stage embryos and 3 embryos cultured to the blastocyst stage. One blastocyst (5AB) was re-frozen. After a negative pregnancy test the patient underwent a second FET cycle. 5 zygotes from the ICSI group were thawed yielding the transfer of 3 cleavage stage embryos and 2 embryos cultured to the blastocyst stage. 2 blastocysts were frozen (4AA, 3BA). Again a negative pregnancy test ensued. The third FET cycle was composed of a blastocyst thaw using the one blastocyst (5AB) from the first FET cycle and both of the blastocysts (4AA,3BA) from the second FET cycle. All 3 blastocysts survived (90%, 70%, 70%) the thaw to be transferred following laser assisted hatching. A positive pregnancy test resulted and subsequent 6 and 8 week ultrasound scans revealed a viable singleton intrauterine pregnancy. Human embryos are capable of withstanding more than one freeze/thaw cycle without impairing their ability to develop and implant. Furthermore, additional data relating to the favorable outcome of fetuses born from twice frozen/thawed embryos is needed before such practices are recommended routinely.

Effect of time since vasectomy and maternal age on intracytoplasmic sperm injection success in men with obstructive azoospermia after vasectomy

Objective To determine the effects of time since vasectomy and maternal age on intracytoplasmic sperm injection (ICSI) outcome in azoospermic men after vasectomy. Design Retrospective analysis. Setting Assisted conception unit. Patient(s) Thirty-seven azoospermic men (after vasectomy) who were undergoing 56 cycles of ICSI. Intervention(s) Surgical sperm retrieval and standard ICSI protocol. The ICSI cycles were analyzed in four groups, according to years since vasectomy, and were reanalyzed in three groups, according to maternal age. Main outcome measure(s) Fertilization rate, implantation rate, clinical pregnancy rate, and live-birth rate (LBR) per ET. Result(s) No effect of time since vasectomy was seen on any outcome. The highest fertilization rate and LBR were found in the group with the longest time interval. These findings could not be explained by differences in either patient characteristics or stimulation regimes. When reanalyzed by maternal age, there was an improvement in implantation rate and LBR with decreasing maternal age. Live birth rates of 38.5%, 22.7%, and 11.8% were achieved for maternal ages of <32, 32–37, and >38 years, respectively. Logistic regression confirmed a statistically significant effect on outcome of maternal age but not time since vasectomy. Conclusion(s) Our data suggest that maternal age, and not interval since vasectomy, remains the principal determinant of ICSI success in men with obstructive azoospermia after vasectomy.

Effect of contaminated preprocessed semen on fertilization rate and embryo quality in assisted reproductive techniques

We aimed to identify the sources and prevalence of semen contamination from masturbation and determine the effect of bacterospermia on fertilization rate and embryo quality in standard in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). This was a prospective controlled study, in an IVF unit of a university teaching hospital, of 93 consecutive couples undergoing IVF-embryo transfer cycles. We evaluated handwashing; semen collection and processing; and assisted reproductive technology using semen provided by masturbation. The main outcome measures were presence and type of micro-organisms in the semen samples and embryo culture medium; the effect of hand washing on rate of contamination; and the effect of semen contamination on fertilization rate and embryo quality. The first consecutive 52 men of the 93 couples were not instructed to wash their hands before masturbation, and the remainder were so instructed. Forty-nine semen cultures (94.2%) in the first group were contaminated compared to only 16 (39%) in the second (p<0.016); 27 of the 65 positive cultures (41.5%) were contaminated by more than one organism. The most common contaminators were bacteria usually found on the skin. All but four embryo medium cultures were negative. There was no significant difference in fertilization rate and embryo quality by culture findings in either the IVF or the ICSI procedures. We found that a high percentage of manually obtained semen for standard IVF or ICSI procedures was contaminated, but this had no effect on fertilization rate and embryo quality.

Embryo development characteristics in Robertsonian and reciprocal translocations: a comparison of results with non-translocation cases

The effect of translocations on embryo development was evaluated and results were compared in terms of embryo development with those of embryos obtained from standard intracytoplasmic sperm injection (ICSI) cycles. In 23 translocation carriers with 34 cycles, fertilization, pronuclear morphology scoring (PMS), developmental arrest, cleavage and blastocyst formation were evaluated and compared with embryos obtained from non-translocation cases undergoing ICSI ( n = 98 cycles). In 28 cycles, preimplantation genetic diagnosis (PGD) was performed on prezygotes (first and second polar body biopsy for female carriers; n = 3) or on embryos having seven or more blastomeres (blastomere biopsy; n = 25). In six cycles for four couples, probes for translocated chromosomes were not available, so PGD could not be performed. Overall, in translocation cases, a lower fertilization rate, a higher rate of retarded embryo development, and a lower rate of blastocyst formation were observed compared with embryos of non-translocation cases. Fluorescence in-situ hybridization (FISH) analysis showed a 70.9% abnormality rate for reciprocal translocations and 55.0% for Robertsonian translocations respectively. In cases with Robertsonian and reciprocal translocation carriers, the probability of poor embryo development, which may be a result of high segregation abnormalities, may negatively affect the outcome of assisted reproductive techniques. This poor prognosis should also be considered when genetic counselling for translocation is given.

Laser-assisted intracytoplasmic sperm injection

Objective: To improve oocyte survival and fertilization rates after intracytoplasmic sperm injection (ICSI) in patients with inherent oocyte fragility. Design: Pilot feasibility study and case report. Setting: Private hospital. Patient(s): Infertile couple with repeated failures of ICSI caused by oocyte degeneration. Intervention(s): Laser-assisted drilling of the zona pellucida followed by ICSI. Main Outcome Measure(s): Oocyte survival, fertilization, and pregnancy. Results: In a couple with four previous ICSI failures because of poor oocyte survival (33.3%, 0%, 20%, and 18%), a fifth attempt using laser-assisted ICSI resulted in the survival of 8 oocytes out of 13 injected. Normal fertilization occurred in 5 oocytes, and a clinical pregnancy was established. Conclusion(s): Performing ICSI through a laser-drilled hole in the zona pellucida reduces the risk of oocyte damage related to deformation during the initial phase of the microinjection procedure. This modification of ICSI appears to be suitable for patients whose oocyte show inherent fragility and high degeneration rates after the standard ICSI procedure.

Intracytoplasmic sperm injection could minimize the incidence of prematurely condensed human sperm chromosomes

Objective: To compare the fertilization and prematurely condensed human sperm chromosomes (PCCs) rates between two intracytoplasmic sperm injection (ICSI) techniques. Design: A retrospective study. Setting: The data were obtained from the University of British Columbia in vitro fertilization (IVF) laboratory. Patient(s): ICSI cycles (n = 105) were performed for couples suffering from severe male-factor infertility and dysfunction of fertilization. Intervention(s): Two types of ICSI techniques were used for ICSI procedures. Main Outcome Measure(s): Fertilization and pregnancy rates in group B using the improved ICSI technique were compared with those of group A using the standard ICSI technique. Unfertilized oocytes from the two groups were studied with cytogenetic methods. Result(s): Oocyte damage dropped from 14.8% in group A to 5.3% in group B. Normal fertilization for each group was 57.3% and 88.4%, respectively ( P<.05). Pregnancy rate per egg retrieval was 15.6% in group A and 27.4% in group B ( P<.05). PCCs occurred in 19.4% of unfertilized oocytes in group A and did not occur in group B. Conclusion(s): This study indicates that ICSI not only yields high fertilization rates, but also minimizes the incidence of PCCs. It may be directly related to two crucial steps (immobilization of sperm and aspiration of oocyte cytoplasm) used in ICSI procedures. This study also suggests that it is possible to overcome one cause of IVF failure resulting from the formation of PCCs by using the improved ICSI technique.

Autologous cumulus cell coculture improves embryo quality and blastocyst growth of icsi-derived human embryos

Intracytoplasmic sperm injection (ICSI) is commonly used to treat severe male factor infertility and results in a high fertilization rate. As with standard IVF embryos, ICSI-derived embryos vary in quality and implantation potential. Since embryo quality and implantation potential are two rate-limiting factors of IVF success, alternative culture systems, such as blastocyst culture and coculture systems, have been developed and implemented. We have recently reported that a simplified coculture system using the patients’ own cumulus tissue results in improved embryo quality and implantation rates for embryos fertilized using standard microdroplet insemination. In this study, we have evaluated autologous cumulus cell coculture in embryos resulting from ICSI. Embryo quality, selection for embryo transfer, and blastocyst development of nontransferred embryos was evaluated in 303 embryos fertilized by ICSI (303/402 [75.4%]). Following ICSI, oocytes were placed in individual 50 μL drops of HTF medium and randomly supplemented with cumulus tissue dissected from the oocytes prior to treatment of the oocyte/cumulus mass with trypain. Embryo quality was evaluated at 72 hours post–oocyte harvest. Percent blastocyst development of nontransferred embryos was determined at 144 hours post–oocyte harvest. The mean embryo quality score was significantly (P<.05) higher for embryos cultured with cumulus tissue (5.1 ± 0.21) than those cultured without cumulus (4.3 ± 0.23). Additionally, 64.1% (84/131) of the coculture embryos were selected for transfer compared to 43.6% (75/172) for standard ICSI embryos (P<.05). Lastly, nontransferred embryos continued to the blastocyst stage at a higher rate (P<.05) when cocultured (18/47 [38.3%] vs. 19/97 [19.6%]). These data indicate that coculture of ICSI embryos with autologous cumulus tissue results in improved embryo quality and developmental potential. Autologous cumulus culture is an advantageous coculture system due to its ease and safety. These data confirm the benefit of the system in ICSI-derived embryos.