Subsequent Intracytoplasmic Sperm Injection Cycles Research Articles

P-066 A multi-bioassay approach to identify male gamete fertilization dysfunction

Abstract Study question Can a multi-bioassay approach identify a specific male gamete dysfunction often overlooked by a standard semen analysis? Summary answer In cases of persistent fertilization failure, additional testing may assist in pinpointing underlying factors contributing to compromised sperm embryo developmental competence and tailor treatment. What is known already Conventional semen analysis, while informative, ineffectively captures the complete spectrum of male gamete dysfunction. Despite adequate sperm concentration and motility, some couples may still experience fertilization failure with Intracytoplasmic Sperm Injection (ICSI). This highlights the complexity of male infertility, that extends beyond standard parameters. In addressing sperm-related fertilization challenges, various assisted oocyte activation (AOA) methods have been proposed to trigger intracellular Ca2+ release, leading to calcium spikes and meiosis initiation. Study design, size, duration In the past 10 months, we identified couples (n = 50) with a history of poor ICSI fertilization, exhibiting suboptimal head morphology. Screening for acrosomal dysfunction involved assessing the presence of PLCxand related gene mutations. We measured the degree of chromatin compaction by histone-to-protamine ratio and evaluated sperm chromatin fragmentation (SCF). To correct for eventual acrosomal dysfunction, couples were counseled to undergo subsequent ICSI cycles with AOA. Clinical outcome was compared between historical and treatment cycles. Participants/materials, setting, methods Fifty men with a history of poor ICSI fertilization were included. A standard semen analysis was conducted according to WHO guidelines. The presence of PLCx was measured through immunofluorescent staining (normal threshold, ≥ 30%), corroborated by gene mutation detection though DNA-seq. Sperm histone-to-protamine ratio was evaluated by Aniline Blue (normal threshold, <15% histone retention). SCF was assessed utilizing the TUNEL assay (normal threshold, ≤15%). AOA was performed using ionomycin or recombinant PLCx protein. Main results and the role of chance Fifty men experienced a poor clinical outcome following ICSI with a fertilization of 7.4% (48/647), embryo implantation of 6.6% (1/15), clinical pregnancy of 6.6% (1/15), and pregnancy loss rate of 100% (1/1). Their semen analysis, interestingly, revealed the following semen parameters: 3.0±1 mL volume, 50.0±20 x106/mL concentration, 41.4±21% motility, and 1.9±1% normal morphology. Nonetheless, PLCx content was diminished, with values ranging from 2-27%. Genomic assessment confirmed mutations in PLCx, DPY19L2, SPATA16 and PICK1 genes, all implicated in sperm nuclear remodeling and acrosome formation. An elevated histone content at 29.7% was positively correlated with an increased SCF at 18.0% (R2=0.6, P<0.001). In subsequent cycles employing AOA, the normal fertilization increased to 42.3% (305/721), the embryo implantation reached 13.0% (9/69), the clinical pregnancy rose to 21.1% (8/38), and the delivery rate was 21.1% (8/38), without any pregnancy loss. (P<0.001). Limitations, reasons for caution While this study affirms the effectiveness of a multifaceted assessment in pinpointing specific sperm dysfunctions and customizing treatment, it is still a retrospective observation requiring independent validation in a larger population. Wider implications of the findings Semen analysis alone fails to address specific aspects of male gamete dysfunction. Incorporating additional bioassays to assess chromatin structure, integrity, and acrosomal function offers valuable insights for fertility management in tailoring personalized treatments. This comprehensive approach enhances the understanding of male reproductive health, maximizing the potential for successful clinical outcome. Trial registration number not applicable

(264) Outcome of Intra-Cytoplasmic Sperm Injection ICSI with Testicular Sperm Aspiration (TESA) In Patients with High Sperm DNA fragmentation, Hospital Experience

Abstract Introduction Sperm DNA fragmentation (SDF) is considered as an important biomarker in the assessment of male fertility potential with contradictory results regarding its effect on ICSI. In our study we focus on the outcomes of Intra-Cytoplasmic Sperm Injection (ICSI) in male patients with high SDF using testicular spermatozoa (TESA). Objective To evaluate intra- cytoplasmic sperm injection (ICSI) outcomes in male patients with high SDF using testicular spermatozoa and analysis of the cofactors (Age, Smoking, Chemical Exposure and usage of Anti-oxidants agents) Methods Retrospective study, institutional experience. Results In our study, we evaluated 41 men with high-SDF levels who had a previous ICSI cycle from their ejaculates. A subsequent ICSI cycle was performed using spermatozoa retrieved through testicular sperm aspiration. The mean (SD) SDF level is 33.7%. live birth rate was 29.2% (12 out of 41) In addition of that, we analyzed the cofactors of men such as Age, Smoking, Chemical Exposure and usage of Anti-oxidants agents. The mean age is 33 %. Men received anti oxidants agent prior to TESA were 9 out of 12 (75%) Conclusions Intra-cytoplasmic sperm injection (ICSI) outcomes in male patients with high SDF using testicular spermatozoa has higher clinical pregnancy and higher chance of live-birth. The rate is higher with those who received anti-oxidant supplements for at least 6 moths. Disclosure No.

P-080 Enzymatic tissue processing after testicular biopsy in non-obstructive azoospermia enhances sperm retrieval and cumulative live birth rates

Abstract Study question What is the added value of enzymatic processing of testicular biopsies on testicular sperm retrieval rates in patients with non-obstructive azoospermia (NOA)? Summary answer In addition to mechanical mincing, enzymatic digestion increased sperm retrieval rates in testicular biopsies of NOA patients with 48.9%. What is known already Many studies focus on the surgical approach to optimize recovery of testicular sperm in NOA, and in spite of that, there is still controversy whether the type of surgery makes any difference as long as multiple biopsies are taken. Few studies, however, focus on the role of the IVF laboratory and the benefit of additional lab procedures, like enzymatic digestion, in order to optimize sperm retrieval rates and CLB rate per TESE. Study design, size, duration A retrospective single-center cohort study including all patients who underwent their first TESE by open multiple-biopsy method from January 2004 till July 2022. Only patients with a normal karyotype, absence of Y-q deletions and a strict diagnosis of NOA based on histology were included. Primary outcome was sperm retrieval after mincing or enzymes for intracytoplasmic sperm injection (ICSI). Secondary outcome was CLB after ICSI with fresh TESE, and subsequent ICSI cycles with frozen TESE. Participants/materials, setting, methods Multiple biopsies were obtained from the testis, unilateral or bilateral, on the day of oocyte retrieval. Upon mechanical mincing, dishes were searched for 30 min; when no or insufficient numbers of spermatozoa were observed, enzymatic treatment was performed using collagenase type IV. Multivariable regression analysis was performed to predict CLB by adjusting for the following confounding factors: male age, male FSH level, cryptorchidism, enzymatic digestion, number of oocytes and female age. Main results and the role of chance Hundred-eighteen patients were included of which 61.0% had successful sperm retrieval. Spermatozoa were retrieved after mechanical mincing (23.7%; 28/118) or after additional enzymatic digestion of the remaining 90 patients (48.9%; 44/90). Mean male characteristics were not different between patients with sperm retrieval after mincing or enzymes: age (34.5 vs 34.5 y), testis volume (10.2 vs 10.6 ml), FSH (17.8 vs 16.9 IU/l), cryptorchidism (21.4 vs 34.1%) and histological diagnosis (Sertoli Cell Only 53.6 vs 47.7%, maturation arrest 21.4 vs 38.6%, sclerosis/atrophy 25.0 vs 13.6%), respectively. Of the 72 patients with sperm available for ICSI, 23/72 (31.9%) obtained a LB after the injection with fresh testicular sperm (fresh and frozen embryo transfers). Forty-nine patients remained without LB, of which 34 had supernumerary testicular sperm frozen. Of these, 9/47 (47.4%) had a LB after ICSI with frozen testicular sperm, giving rise to a total CLB per TESE of 32/118 (27.1%) or 32/72 (44.4%) CLB per TESE with sperm. Of the female characteristics (couples with sperm available), only female age (30.3 vs 32.7 y – p = 0,042) was significantly lower in the group with a live birth. Multivariable logistic regression analysis showed that enzymatic digestion was associated with significant decrease of CLB per TESE. Limitations, reasons for caution Limitations of the study are related to the retrospective design. The selection of only NOA patients with specific characteristics (normal karyotype and absence Y-q deletion) and having their first TESE ever strengthens our findings. Whether enzymatic digestion after a failed TESE without digestion may improve retrieval rate remains undecided. Wider implications of the findings Enzymatic processing increases the sperm retrieval rate from testicular biopsies of NOA patients compared to mechanical mincing, demonstrating the importance of an appropriate laboratory protocol. NOA patients should be counselled that if sperm has been found after enzymatic digestion, their chances to father a genetically own child will be lower. Trial registration number not applicable

Assessing male gamete genome integrity to ameliorate poor assisted reproductive technology clinical outcome

To assess the role of evaluating sperm chromatin fragmentation (SCF) as a tool to guide treatment in couples who achieved unexpectedly poor clinical outcomes after intracytoplasmic sperm injection (ICSI). We identified couples with an unexpectedly suboptimal clinical outcome after ICSI who were then screened for SCF. Consequently, the same couples were counseled to undergo a subsequent ICSI cycle using either ejaculates processed by microfluidic sperm selection (MFSS) or spermatozoa retrieved from the testis, and clinical outcomes were compared between history and treatment cycles. To confirm the sole effect of a compromised male gamete, we compared the ICSI outcome in cycles where male gametes with abnormal SCF were used to inseminate autologous and donor oocytes. Finally, to eliminate an eventual confounding female factor component, we compared the clinical outcome of ICSI cycles using sibling donor oocytes injected with spermatozoa with normal or abnormal SCF. Academic reproductive medicine center point of care. The patient population consisted of 76 couples with reproductively healthy and relatively young female partners and male partners with compromised semen parameters, but suitable for ICSI. In a subanalysis, we identified 67 couples with abnormal SCF who underwent ICSI cycle(s) with donor oocytes. Furthermore, we identified 29 couples, 12 with normal SCF and 17 with abnormal, uncorrected SCF, and 7 couples with abnormal, corrected SCF vs. a control, who used sibling donor oocytes for their ICSI cycle(s). For couples who resulted in surprisingly low clinical outcomes after ICSI, despite semen parameters adequate for ICSI and a normal female infertility evaluation, a SCF assessment was performed on the semen specimen using the terminal deoxynucleotidyl transferase-mediated fluorescein-deoxyuridine triphosphate nick-end labeling (TUNEL) assay. The couples then underwent a subsequent ICSI cycle with spermatozoa processed by MFSS or surgically retrieved. Moreover, cycles with donor oocytes were used to confirm the sole contribution of the male gamete. Clinical outcomes, such as fertilization, embryo implantation, clinical pregnancy, delivery, and pregnancy loss rates were compared between history and treatment cycle(s) using ejaculated spermatozoa selected by MFSS or from a testicular biopsy, taking into consideration the level of SCF. In a subanalysis, we reported the clinical outcomes of 67 patients who used donor oocytes and compared them with cycles where their own oocytes were used. Furthermore, we compared the ICSI clinical outcomes between cycles using sibling donor oocytes injected with low or high SCF with or without sperm intervention aimed at correcting, or alleviating the degree of SCF. In a total of 168 cycles, 76 couples had in a prior cycle a 67.1% fertilization rate, and clinical pregnancy and pregnancy loss rates of 16.6% and 52.3%, respectively. After testing for SCF, the DNA fragmentation rate was 21.6%. This led to a subsequent ICSI cycle with MFSS or testicular sperm extraction, resulting in clinical pregnancy and delivery rates of 39.2%, and 37.3%, respectively. The embryo implantation rate increased to 23.5%, whereas the pregnancy loss rate decreased to 5% in the treatment cycle. This was particularly significant in the moderate SCF group, reaching embryo implantation, clinical pregnancy, and delivery rates of 24.3%, 40.4%, and 36.2%, respectively, and reducing the pregnancy loss rate to 10.5% in post-sperm treatment cycles. In 67 patients with high SCF who used donor oocytes, a significantly higher fertilization rate of 78.1% and embryo implantation rate of 29.1% were reported, compared with those in couples also with an elevated SCF who used their own. Interestingly, the clinical pregnancy and delivery rates only increased slightly from 28.0%-36.1% and from 23.7%-29.2%, respectively. To further control for a female factor, we observed couples who shared sibling donor oocytes, 17 with normal SCF and 12 with abnormal (uncorrected) SCF. Interestingly, the abnormal SCF group had impaired fertilization (69.3%), embryo implantation (15.0%), and delivery (15.4%) rates. For an additional 15 couples who split their donor oocytes, 8 had normal SCF, and although 7 couples originally had abnormal SCF, 4 used microfluidic processing, 2 used testicular spermatozoa, and 1 used donor spermatozoa to alleviate the degree of SCF, resulting in comparable clinical outcomes with the normal SCF group. A superimposed male factor component may explain the disappointing ICSI outcome in some couples despite reproductively healthy female partners. Therefore, it may be useful to screen couples for SCF to guide treatment options and maximize chances of a successful pregnancy. The improved, but suboptimal pregnancy and delivery outcomes observed in couples using donor oocytes confirmed the exclusive detrimental role that the male gamete exerted on embryo development despite the presence of putative oocyte repair mechanisms.

Identification and treatment of men with phospholipase Cζ–defective spermatozoa

To identify and treat the gamete responsible for complete fertilization failure with intracytoplasmic sperm injection (ICSI) using a newly proposed assisted gamete treatment (AGT). Prospective cohort study. Center for reproductive medicine. One-hundred and fourteen couples with an adequate number of spermatozoa for ICSI and a fertilization rate of ≤10%, after controlling for maternal age. Couples with an oocyte-related oocyte activation deficiency (OAD) underwent a subsequent cycle with a modified superovulation protocol; couples with sperm-related OAD had an additional genetic and epigenetic assessment to identify mutations and expression levels of the corresponding genes. Treatment cycle outcome for couples undergoing ICSI with either a modified superovulation protocol or AGT compared with their historical cycle. A total of 114 couples matched the inclusion criteria, representing approximately 1.3% of the total ICSI cycles performed at our center, with age-matched controls. Fifty-two couples were confirmed negative for sperm-related OAD by the phospholipase Cζ (PLCζ) assay, indicating oocyte-related factors in their failed fertilization cycles. Couples were treated by one of two AGT protocols, AGT-initial or AGT-revised, in a subsequent attempt that was compared with their historical cycle. Subsequent ICSI cycles with a tailored superovulation protocol yielded significantly higher fertilization (59.0% vs. 2.1%) and clinical pregnancy (28.6% vs. 0) rates. In 24 couples (mean ± standard deviation: maternal age, 35.6 ± 5 years; paternal age, 39.8 ± 6 years) sperm-related OAD was confirmed; in four men, a deletion on the PLCZ1 gene was identified. Additional mutations were also identified of genes supporting spermiogenesis and embryo development (PIWIL1, BSX, NLRP5) and gene deletions confirming a complete absence of the subacrosomal perinuclear theca (PICK1, SPATA16, DPY19L). Subsequent AGT treatment provided higher fertilization (42.1%) and clinical pregnancy (36% vs. 0%) rates for couples with a history of impaired (9.1%) fertilization. A comparison of the two AGT protocols, AGT-initial or AGT-revised, revealed that the latter yielded even more favorable fertilization (37.6% vs. 45.9%) and clinical pregnancy (21.1% vs. 83.3%) rates. In couples with an oocyte-related OAD, tailoring the superovulation protocol resulted in successful fertilization, term pregnancies, and deliveries. In couples with a sperm-related OAD as determined by PLCζ assay, mouse oocyte activation test, and the assessment of gene mutations and function, AGT was successful. The AGT-revised protocol yielded an even higher fertilization rate than the AGT-initial protocol, resulting in the birth of healthy offspring in all couples who achieved a clinical pregnancy.

ICSI outcome in patients with high DNA fragmentation: Testicular versus ejaculated spermatozoa.

Sperm DNA fragmentation (SDF) has emerged as an important biomarker in the assessment of male fertility potential with contradictory results regarding its effect on ICSI. The aim of this study was to evaluate intracytoplasmic sperm injection (ICSI) outcomes in male patients with high SDF using testicular versus ejaculated spermatozoa. This is a prospective study on 36 men with high-SDF levels who had a previous ICSI cycle from their ejaculates. A subsequent ICSI cycle was performed using spermatozoa retrieved through testicular sperm aspiration. Results of the prior ejaculate ICSI were compared with those of the TESA-ICSI. The mean (SD) SDF level was 56.36% (15.3%). Overall, there was no difference in the fertilization rate and embryo grading using ejaculate and testicular spermatozoa (46.4% vs. 47.8%, 50.2% vs. 53.4% respectively). However, clinical pregnancy was significantly higher in TESA group compared to ejaculated group (38.89% [14 of 36] vs. 13.8% [five of 36]). Moreover, 17 live births were documented in TESA group, and only three live births were documented in ejaculate group (p<.0001). We concluded that the use of testicular spermatozoa for ICSI significantly increases clinical pregnancy rate as well as live-birth rate in patients with high SDF.

Uncommon but devastating event: total fertilisation failure following intracytoplasmic sperm injection.

Fertilisation with intracytoplasmic sperm injection (ICSI) is a consequence of complex molecular interactions between spermatozoon and oocyte. Disruption of the process obviously prompts a frustrating event called total fertilisation failure (TFF). Up to 3% of ICSI cycles may result in TFF, and brief counselling for subsequent cycle management is indispensable. Within this perspective, ICSI cycles of a centre over a 10-year period were analysed to document TFF cases. Initial TFF after ICSI and subsequent ICSI cycle of the same cases were documented to clarify predictive factors of successful outcomes after initial TFF. In subsequent cycles, assisted oocyte activation (AOA) with calcium ionophore and Hypo-osmotic swelling test (HOST)/pentoxifilline for sperm selection was used. In the current analysis, successful fertilisation was achieved in 85% of the cases with previous TFF. The significant contributing factors for successful fertilisation in the latter cycle were: improved oocyte quantity and better sperm morphology. In conclusion, sporadic TFF event in the first and only cycle is usually a technically modifiable condition, but repeated TFF could indicate possible gamete defects, which might not be overcomed in the next modified ICSI cycle.

Sperm involved in recurrent partial hydatidiform moles cannot induce the normal pattern of calcium oscillations

To assess the Ca2+-releasing ability of sperm involved in partial hydatidiform moles. Analysis of the activating and Ca2+-releasing ability of human sperm. University hospital research laboratory. Patients undergoing intracytoplasmic sperm injection (ICSI) treatment. Microinjection of mouse and human oocytes with sperm. Measurement of the fertilizing and Ca2+-releasing ability of human sperm. The mouse oocyte Ca2+ analysis showed that only 19.0% (4/21) of the mouse oocytes injected with sperm involved in molar pregnancies exhibited a normal pattern of Ca2+ oscillations versus 63.2% (36/57) of those injected with control sperm. Further, 83.3% (15/18) of donated in vitro-matured human oocytes injected with deficient sperm did not exhibit any Ca2+ release, while 76.9% (10/13) failed to show normal pronuclear development. Yet the sperm oocyte activation factor phospholipase C zeta (PLCζ) was present in the majority (96.6%, n=113) of the analyzed sperm at a normal expression level. Eventually, fertilization failure was overcome with assisted oocyte activation in subsequent therapeutic ICSI cycles, which led to normal deliveries. Sperm that previously provoked recurrent partial hydatidiform mole pregnancies due to dispermic fertilization is not able to activate human oocytes or trigger the normal pattern of Ca2+ oscillations in mouse and human oocytes in vitro.

Pregnancy outcome and neonatal data of children born after intracytoplasmic sperm injection with a different duration of cryopreservation of spermatozoa obtained through testicular sperm extraction

ObjectiveTo evaluate the impact of the duration of cryopreservation of testicular sperm on clinical and neonatal outcomes after intracytoplasmic sperm injection (ICSI) for patients experiencing azoospermia. Materials and methodsThis study included 132 azoospermic men who participated in 212 ICSI cycles. The participating men underwent testicular biopsy for the cryopreservation of tissue to be used for subsequent ICSI cycles. The duration of the storage of testicular sperm was determined by the time of oocyte retrieval. Fertilization, embryo development in vitro, pregnancy rates, and neonatal outcomes were assessed. ResultsAlthough the mean percentage of viability decreased from 73.2% prior to freezing to 50.2% after thawing, viable spermatozoa were visualized subsequent to thawing of the tissue samples for all patients. The potential for fertilization and subsequent embryonic development was not influenced by the duration of sperm cryopreservation up to 2 years longer. The pregnancy outcomes also varied independently of the duration of sperm cryopreservation. The duration of storage did not appear to affect the neonatal outcomes adversely, including the Apgar score and intensive care unit admission rates, although neonatal outcomes were influenced by advanced maternal age. It also has no obvious impact on the major and minor congenital malformation rate of the newborns. ConclusionICSI outcomes, pregnancy outcome, neonatal outcome, and congenital malformation rate appear not to be affected by the duration of the period of cryostorage. An earlier start of the ICSI cycle following the testicular sperm cryopreservation is preferable because longer preservation is associated with more advanced maternal age.

Endometrial expression of selected genes in patients achieving pregnancy spontaneously or after ICSI and patients failing at least two ICSI cycles

The objective of this study was to identify the endometrial gene expression profile in receptive phase, which could represent a useful prognostic tool for selecting IVF patients. Endometrial expression of 47 selected genes biopsied during the window of implantation in natural cycles was compared between patients who achieved a successful pregnancy spontaneously or after subsequent intracytoplasmic sperm injection (ICSI) cycles and patients who did not achieve a pregnancy after at least two failed ICSI cycles. The comparative analysis showed significantly different levels of expression in 19 genes, five implicated in apoptosis (CASP8, FADD, CASP10, APAF1, ANXA4), three in immunity (LIF, SPP1, C4BPA), five in transcriptional activity (MSX1, HOXA10, MSX2, HOXA11, GATA2), two in lipid metabolism (LEPR, APOD) and four in oxidative metabolism (AOX1, ALDH1A3, GPX3, NNMT). The evidence for these genes being differently expressed could represent the starting point of identifying the ideal receptive endometrial gene expression profile, which could be used in the future as a prognostic tool for IVF patients.Gene expression analysis technology has opened new important perspectives on the study of the physiological processes of different tissues and organs. Specifically for the endometrium, it would be really interesting to find out an endometrial gene expression profile of receptive phase, which could be used in future as a useful prognostic tool for selecting IVF patients. To achieve this aim, the objective of the present paper was the comparison of endometrial expression in natural cycles of 47 selected genes between the biopsies of patients who achieved a successful pregnancy, either spontaneously or after subsequent ICSI cycles, and those of patients who did not achieve a pregnancy after at least two failed ICSI cycles. The comparative analysis showed a significant different expression in 19 genes: five implicated in programmed cell death, known as apoptosis (CASP8, FADD, CASP10, APAF1, ANXA4), three in immunity (LIF, SPP1, C4BPA), five in transcriptional activity (MSX1, HOXA10, MSX2, HOXA11, GATA2), two in lipid metabolism (LEPR, APOD) and four in oxidative metabolism (AOX1, ALDH1A3, GPX3, NNMT). The evidence of these genes being differently expressed could represent the starting point of identifying the ideal receptive endometrial gene expression profile which could be used in the future as a prognostic tool for IVF patients.

The Well-of-the-Well system: an efficient approach to improve embryo development

Transfer of human embryos at the blastocyst stage may offer considerable benefits including an increased implantation rate and a decreased risk of multiple pregnancies; however, blastocyst culture requires an efficient and reliable in-vitro embryo culture system. In this study, the effect of the Well-of-the-Well (WOW) system consisting of microwells formed on the bottom of the culture dish was tested in three mammalian species, including humans. The WOW system resulted in significant improvement when comparing the drops for culture of in-vitro-matured and parthenogenetically activated porcine oocytes, and in-vivo-derived mouse zygotes. In human embryos, using a sibling oocyte design, embryos cultured in WOW developed to the blastocyst stage in a significantly higher proportion than did embryos cultured traditionally (55% in WOW and 37% in conventional culture; P < 0.05). In a separate study, also in human, a total of 48 patients with a cumulative 214 unsuccessful previous IVF cycles were selected for the trials. In subsequent intracytoplasmic sperm injection cycles, oocytes/embryos were cultured individually in the WOW system or in microdrops. Transferable quality blastocyst development (48.9% of cultured zygotes) was observed in the WOW system. Ninety-four blastocysts transferred to 45 patients resulted in clinical pregnancy rates of 48.9%, including nine twin pregnancies, seven single pregnancies, five miscarriages and one ectopic pregnancy. The results indicate that the WOW system provides a promising alternative for microdrop culture of mammalian embryos, including human embryos.

Normal birth after single-embryo transfer in a patient with excessive polypronuclear zygote formation following in vitro fertilization and intracytoplasmic sperm injection

To report an unusual case of idiopathic zygotic polypronuclei. Case report. Major urban infertility referral center. A 34-year-old woman with unexplained infertility. The patient underwent two cycles of controlled hyperstimulation and oocyte retrieval followed by in vitro insemination and intracytoplasmic sperm injection (ICSI). Pregnancy and delivery of a normal infant following transfer of a single preembryo from an ICSI cycle in which only one zygote showed a normal pronuclear number (2PN) and 12 zygotes appeared polypronucleated (> or =3PN). On the first IVF cycle, 23 oocytes were retrieved and inseminated with 240 x 10(3) motile sperm/mL, after which two zygotes showed a normal pronuclear number and 20 zygotes appeared polyploid with three to seven pronuclei. Transfer of two poor-quality day-3 preembryos following assisted hatching did not achieve pregnancy. On the subsequent ICSI cycle, 33 oocytes were retrieved, and 17 mature oocytes were subjected to ICSI, after which only one zygote showed 2PN and 12 zygotes appeared polyploid with three to eight pronuclei. The normally fertilized zygote developed into a poor-quality, day-3 embryo and was subjected to assisted hatching. Transfer of this preembryo resulted in an uneventful pregnancy and birth of a normal infant. Mechanisms other than polyspermia may result in polypronuclear development in some patients.

Disordered zona pellucida–induced acrosome reaction and failure of in vitro fertilization in patients with unexplained infertility

Objective To determine the relationship between the zona pellucida (ZP)–induced acrosome reaction (AR) and fertilization rate and pregnancy rate in standard IVF and the frequency of disordered ZP-induced AR (DZPIAR) in patients with unexplained infertility. Design Prospective study. Setting Academic research and teaching tertiary hospital. Patient(s) Patients with unexplained infertility with normal semen analysis. Intervention(s) None. Main outcome measure(s) Semen analysis, the ZP-induced AR, and measurements of fertilization rate and pregnancy rate with in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI). Result(s) A statistically significant correlation was found for the ZP-induced AR and fertilization rate with standard IVF (n = 65). Patients with DZPIAR (ZP-induced AR of ≤16%) had an average fertilization rate of 23%, in contrast to patients with ZP-induced AR of >16%, who had an average fertilization rate of 61%. The sensitivity and specificity of DZPIAR for prediction of IVF rates <30% and ≥30% were 80% and 86%, respectively. Of 260 patients screened, the frequency of DZPIAR was 29%. Ten patients with DZPIAR had an average fertilization rate of 15% and no pregnancy with initial IVF cycles, and a fertilization rate of 61% (with three live-birth pregnancies) with subsequent ICSI cycles. Another 33 patients with DZPIAR were treated with ICSI alone, with an average fertilization rate of 71% and a live-birth pregnancy rate of 17% per embryo transfer. Sixteen of the patients had live-birth pregnancies (including one set of twins) after undergoing an average of 3.2 embryo transfers. Conclusion(s) Patients with DZPIAR have a low or zero fertilization rate with standard IVF but high fertilization and pregnancy rates with ICSI. Up to 29% of patients with unexplained infertility with normal semen analysis may have this condition, which should be diagnosed and treated with ICSI rather then standard IVF.

Surgical sperm retrieval after previous vasectomy and failed reversal: clinical implications for in vitro fertilization.

To investigate the effect of the interval between previous vasectomy reversal on retrieval rates of epididymal and testicular spermatozoa using percutaneous epididymal sperm aspiration (PESA), or testicular sperm extraction (TESE), and the subsequent reproductive potential of these gametes in intracytoplasmic sperm injection (ICSI) cycles. Sixty-six consecutive sperm retrievals were considered in patients who were azoospermic after previous vasectomy, of whom 54 had had a previous failed reversal, the remainder deciding against a reversal. PESA and TESE retrieval rates were noted, as were the time since vasectomy and the interval between vasectomy and unsuccessful reversal. The presence of palpable epididymal cysts was noted, with their effect on sperm retrieval rates. Fertilization and pregnancy rates were analysed in subsequent ICSI cycles using freshly retrieved spermatozoa or frozen-thawed cryopreserved spermatozoa. All 66 patients had sperm retrieved successfully; the success rates for PESA were not significantly affected by previous failed reversal when compared with patients who had not had a reversal, at 14 of 54 (26%) vs five of 12 (P=0.3). The interval since vasectomy did not affect PESA retrieval rates but there was a significantly poorer retrieval rate for PESA in the presence of palpable epididymal cysts, at seven of 35 (20%) vs 12 of 23 (52%) (P=0.012). Fertilization rates were significantly lower using cryopreserved spermatozoa retrieved from either the epididymis or testis (50% vs 70%, P=0.007), although subsequent implantation and pregnancy rates were not significantly different. Surgical sperm retrieval is successful in all cases of azoospermia secondary to vasectomy, either by PESA or TESE. There are no clinical markers to indicate which patients will have successful PESA after vasectomy, although the presence of epididymal cysts is associated with significantly lower retrieval rates. The reduction in fertilising ability of cryopreserved spermatozoa does not affect clinical pregnancy rates in ICSI cycles.

Initial experiences of a testicular sperm extraction programme for assisted reproduction in Ireland.

The technique aspirating spermatozoa from the testis is a new development in male infertility treatment. It is appropriate for infertile couples where the male has azoospermia but is still producing live motile spermatozoa in the testes. To describe the initial experiences of a testicular sperm extraction programme (TESE) coupled with intracytoplasmic sperm injection (ICSI) in 18 men during 1998. Spermatozoa were obtained by direct aspiration from the testes using a 16 gauge needle with cannula and negative suction under local anaesthetic. All samples obtained were to be cryopreserved for use in a subsequent ICSI cycle. All five men with congenital bilateral absence of the vas deferens to be carriers of cystic fibrosis gene mutations. No gene deletions were found in their wives. No other cyto or molecular genetic abnormalities were otherwise found. Twenty one procedures were carried out. The mean number of aspirations was 1.72. Eleven samples from 10 men had sperm suitable for ICSI post-freeze. Post-procedure pain was the universal side-effect. Eight couples had a single attempt at ICSI, two couples two each. Fertilisation rate was 71.4%. Two pregnancies were achieved. TESE may give hope in selected cases of azoospermia of fathering a child without the involvement of a third party.

Implications of Complete Fertilization Failure After Intracytoplasmic Sperm Injection for Subsequent Fertilization and Reproductive Outcome

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 J 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 J 10) underwent at least one subsequent ICSI cycle which proceeded to oocyte retrieval. The remainder (n J 11) elected to discontinue treatment. Although six subsequent cycles were cancelled due to poor follicular response (o2 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 postindex) 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.

Implications of complete fertilization failure after intracytoplasmic sperm injection for subsequent fertilization and reproductive outcome.

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.

Fertilization and pregnancy using intentionally cryopreserved testicular tissue as the sperm source for intracytoplasmic sperm injection in 10 men with non-obstructive azoospermia.

Testicular tissue extraction (TESE) to obtain spermatozoa for use with intracytoplasmic sperm injection (ICSI) has recently been employed in patients with non-obstructive azoospermia. Standard protocol is to retrieve a new sample of testis tissue on the day of oocyte recovery. Unfortunately, approximately 30% of men will possess no spermatozoa in their tissue, making ICSI an impossibility. We investigated whether testicular tissue that was intentionally obtained well before any planned ICSI cycle and cryopreserved could then serve as an efficacious sperm source in a subsequent ICSI cycle. This study reports on 10 men with non-obstructive azoospermia who did have spermatozoa found within their testis tissue at the time of TESE and who chose to use their frozen samples as the source of spermatozoa for a later cycle of ICSI. In 19 cycles the overall fertilization rate was 48%. Embryo transfer occurred in 89% of cycles. Two couples have achieved pregnancy (one ongoing, one delivered). All patients except one had multiple vials of frozen tissue remaining following their first cycle. This approach is offered as an alternative to repeated testicular tissue sampling, as the availability of spermatozoa is assured prior to the initiation of ovulation induction. This tissue can be harvested at the same time as diagnostic biopsy, thereby minimizing the number of surgical procedures.

Analysis of 76 total fertilization failure cycles out of 2732 intracytoplasmic sperm injection cycles

From October 1992 to December 1994, 2732 cycles of treatment by intracytoplasmic sperm injection (ICSI) were carried out in couples mainly with severe male-factor infertility. The overall fertilization rate in these 2732 cycles was 71% of intact oocytes. However, in 76 (72 couples) of these cycles, none of the injected oocytes became fertilized, so the total fertilization failure rate was 3% (76/2732 cycles). Details of these 76 cycles were analysed. The results show that total fertilization failure after ICSI may be explained by different factors related to (i) semen characteristics (only immotile or round-headed spermatozoa for ICSI) or (ii) the oocytes (number, abnormal morphology, damage after ICSI). Of 26 couples, 22 achieved fertilization in their subsequent ICSI cycles. In conclusion, total fertilization failure after ICSI for the treatment of severe male-factor infertility was mainly caused by the poor viability of the spermatozoa used for injection; it was also associated with a low number and poor quality of oocytes. Repeated ICSI treatment may be useful or necessary in couples with total fertilization failure.