Novel variants in ACTL7A and PLCZ1 are associated with male infertility and total fertilization failure.

Total fertilization failure (TFF), which refers to fertilization failure in all mature oocytes, accounting for 5%-10% of in vitro fertilization (IVF) cycles and 1%-3% of intracytoplasmic sperm injection (ICSI) cycles in human. In this study, we recruited three unrelated primary infertile men with repeated cycles of TFF and performed whole-exome sequencing to identify the potential pathogenic variants. We identified homozygous or compound-heterozygous variants of paternal-effect genes ACTL7A and PLCZ1 that followed a Mendelian recessive inheritance pattern. Novel homozygous nonsense variant in ACTL7A [c.C146G: p.S49*] was identified in case 1, who came from a consanguineous family. Ultrastructural observation of ACTL7A-mutated spermatozoa by transmission electron microscopy (TEM) indicated that apparent increased thickness of perinuclear matrix and the acrosome was detached from the nuclear envelop. Besides, two novel compound-heterozygous variants in PLCZ1 were identified in case 2 [c.1174+3A>C:p.?; c.A1274G:p.N425S] and case 3 [c.136-1G>C:p.?; c.G1358A:p.G453D]. Mutated spermatozoa from case 2 with reduced expression of PLCZ1 showed apparent acrosome detachment by TEM analysis. And ICSI with assisted oocyte activation (ICSI-AOA) treatment can partly rescue the TFF. Taken together, our findings revealed that novel biallelic variants in the paternal-effect genes ACTL7A and PLCZ1 were associated with human TFF, which expanding the spectrum of genetic causes and facilitating the genetic diagnosis of male infertility with TFF.

ACROSIN deficiency causes total fertilization failure in humans by preventing the sperm from penetrating the zona pellucida.

STUDY QUESTION: Does a homozygous nonsense mutation in ACR lead to total fertilization failure (TFF) resulting in male infertility in humans? SUMMARY ANSWER: A novel homozygous nonsense mutation of ACR (c.167G>A, p.Trp56X) was identified in two infertile brothers and shown to cause human TFF. WHAT IS KNOWN ALREADY: ACROSIN, encoded by ACR, is a major acrosomal enzyme expressed only in the acrosome of the sperm head. Inhibition of acrosin prevents sperm penetration of the zona pellucida (ZP) in several species, including humans. Acr-knockout in hamsters causes male infertility with completely blocked fertilization. Of note, there are no reports of ACR mutations associated with TFF in humans. STUDY DESIGN, SIZE, DURATION: Whole-exome sequencing (WES) was used for the identification of pathogenic genes for male factor TFF in eight involved couples. PARTICIPANTS/MATERIALS, SETTING, METHODS: Data from eight infertile couples who had experienced TFF during their IVF or ICSI attempts were collected. Functional assays were used to verify the pathogenicity of the potential genetic factors identified by WES. Subzonal insemination (SUZI) and IVF assays were performed to determine the exact pathogenesis of TFF caused by deficiencies in ACROSIN. MAIN RESULTS AND THE ROLE OF CHANCE: A novel homozygous nonsense mutation in ACR, c.167G>A, p.Trp56X, was identified in two additional primary infertile brothers whose parents were first cousins. This rare mutation caused ACROSIN deficiency and acrosomal ultrastructural defects in the affected sperm. Spermatozoa lacking ACROSIN were unable to penetrate the ZP, rather than hampering sperm binding, disrupting gamete fusion, or preventing oocyte activation. These findings were supported by the fertilization success of SUZI and ICSI attempts, as well as the normal expression of ACTL7A and PLCζ in the mutant sperm, suggesting that ICSI without remedial assisted oocyte activation is an optimal treatment for ARCOSIN-deficient TFF. LIMITATIONS, REASONS FOR CAUTION: The absence of another independent pedigree to support our argument is a limitation of this study. WIDER IMPLICATIONS OF THE FINDINGS: The findings expand our understanding of the genes involved in human TFF, providing information for appropriate genetic counseling and fertility guidance for these patients. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Natural Science Foundation of China (grant no. 82201803, 81901541, 82271639, and 32000584), University Synergy Innovation Program of Anhui Province (GXXT-2019-044), and the Nonprofit Central Research Institute Fund of the Chinese Academy of Medical Sciences (grant no. 2019PT310002). The authors declare no conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.

Day after rescue ICSI: eliminating total fertilization failure after conventional IVF with high live birth rates following cryopreserved blastocyst transfer.

STUDY QUESTION: What is the impact of day after rescue ICSI (r-ICSI) on success of fresh and frozen embryo transfers? SUMMARY ANSWER: The use of r-ICSI can virtually allay fears of total fertilization failure (TFF) after conventional IVF (C-IVF) and achieve high live birth rates after frozen blastocyst transfer. WHAT IS KNOWN ALREADY: More infertility clinics have resorted to the use of ICSI in place of C-IVF in IVF treatment owing to fear of TFF or a low fertilization rate. r-ICSI has been attempted either on the day of IVF or the day after. Day after r-ICSI has proved unsuccessful in the past. STUDY DESIGN, SIZE, DURATION: A retrospective data analysis was performed of 16 608 qualifying cases between April 2010 and July 2021 conducted at a single private academically affiliated fertility clinic. PARTICIPANTS/MATERIALS, SETTING, METHODS: r-ICSI was performed principally on patients with >4 metaphase II oocytes, showing no signs of fertilization 18 h after C-IVF. C-IVF was performed on patients who had >4 million total motile sperm after preparation. r-ICSI was then performed 18-24 h after insemination, using the sperm sample from the previous day. r-ICSI fertilization rates, cryopreservation of cleavage and blastocysts embryos, and pregnancy rates after fresh or frozen transfer were then assessed. MAIN RESULTS AND THE ROLE OF CHANCE: r-ICSI was performed on 377 patients (2.3% of eligible retrieval cycles) who had a mean (±SD) female and male age of 35.9 ± 4.5 and 38.1 ± 9.1 years, respectively. A total of 5459 oocytes were initially retrieved. Of the oocytes undergoing r-ICSI, 2389 (49.5%) fertilized normally, and 205 (54.4%) patients underwent a fresh embryo transfer. The live birth rates were 23/186 (12.3%) for fresh cleavage and 5/19 (26.3%) for fresh blastocyst stage transfers. In 145 cycles a blastocyst was frozen, and 137 transfers were performed with a 64/137 (46.7%) live birth rate. Of the 377 cycles receiving r-ICSI only, 25 of the qualifying cases failed to have any fertilization, reducing TFF to 25/16 608 (0.15%). LIMITATIONS, REASONS FOR CAUTION: This was a single-center retrospective study on a specific subset of patients, which may limit its generalizability to other clinics. WIDER IMPLICATIONS OF THE FINDINGS: r-ICSI allows a second opportunity to fertilize oocytes despite poor initial outcomes. Patients who had a frozen blastocyst transfer achieved high live birth rates, indicating that a resynchronization of the embryo with the endometrium can optimize r-ICSI cases. r-ICSI allays fears of TFF when using C-IVF, providing evidence that the overuse of ICSI in patients without male factor may not be warranted. STUDY FUNDING/COMPETING INTEREST(S): The study was internally funded by Boston IVF. The authors declare that they have no conflict of interest in relation to the data published in the article. TRIAL REGISTRATION NUMBER: N/A.

Development and validation of a conventional in vitro total fertilization failure prediction model.

BACKGROUND: Conventional total fertilization failure (TFF) is a challenging problem for clinicians. The predictive model developed in this study aims to predict the individual probability of conventional in vitro total fertilization failure. METHODS: The prediction model was developed based on 1635 patients who underwent first-attempt in vitro fertilization (IVF) cycles from January 2018 to January 2020. Total fertilization failure and normal fertilization occurred in 218 and 1417 cycles, respectively. Multivariate logistic regression analyses were used to develop the prediction model. Performance of our model was evaluated using calibration (Hosmer-Lemeshow test) and discrimination (area under the receiver operating characteristic curve [AUC]). RESULTS: Thirteen risk factors for TFF were included in the prediction model, as follows: female age; female body mass index; infertility duration; number of oocytes retrieved; stimulation protocol; infertility etiology; infertility diagnosis; male age; sperm concentration; total sperm motility; normal sperm morphology percentage; swim-up sperm motility; and swim-up sperm concentration. The AUC of our model was 0.815 (95% CI: 0.783-0.846), indicating satisfactory discrimination performance. CONCLUSION: Considering female and male factors (especially sperm parameters), we established a model that predicts the probability of TFF in conventional IVF procedures that will be helpful in the laboratory supporting IVF to facilitate physicians in determining optimal treatment.

Fertilization and neonatal outcomes after early rescue intracytoplasmic sperm injection: a retrospective analysis of 16,769 patients.

PURPOSE: To evaluate the efficacy and safety of short-term insemination and early-rescue intracytoplasmic sperm injection (ICSI), an approach that rescued oocytes with unclear second polar body 6 h after initial insemination by ICSI (early R-ICSI) to avoid total or near-total fertilization failure in conventional in vitro fertilization (IVF). METHODS: We performed a retrospective study in 16,769 patients (short-term IVF, n = 12,094; ICSI, n = 3452; early R-ICSI, n = 1223) who received IVF/ICSI treatment in our hospital from January 2009 to October 2018. Fertilization and clinical outcomes were compared among those three groups. RESULTS: When considering the R-ICSI embryos in the early R-ICSI group independently, the rates of fertilization and day-3 cleaved embryos in 2PN oocytes were comparable, the rates of fertilization (2PN) and high-quality embryos were lower, whereas the multi-PN fertilization rate (3.27%) was significantly higher than the ICSI group (1.26%). The difference of clinical pregnancy rate between the part of transferred R-ICSI embryos (40.81%) and the ICSI group (44.73%) remained nonsignificant. Furthermore, the rate of congenital birth defects in the early R-ICSI group (0.99%) was not significantly different from those in the short-term IVF (0.76%) and ICSI groups (1.07%). CONCLUSION: Despite the multi-PN fertilization rate, our study highlights early R-ICSI as a safe and effective alternative in assisted reproduction to decrease complete IVF fertilization failure and reduce ICSI utilization. Additional large amount and long-term follow-up studies are needed to further validate the use of early R-ICSI.

Influence of the sperm DNA fragmentation index on the outcome of rescue ICSI and the clinical value of rescue ICSI. / 精子DNA断裂指数对补救ICSI结局的影响及补救ICSI的临床价值.

OBJECTIVES: As a remedy for the failure of in vitro fertilization (IVF), rescue intracytoplasmic sperm injection (R-ICSI) has been widely carried out, but it has failed to significantly improve the fertilization rate and clinical pregnancy rate. Sperm DNA fragmentation index (DFI) was highly correlated with pregnancy outcome of artificial assisted reproduction. This study aims to investigate the effect of the sperm DFI on the outcome of R-ICSI and the clinical value of R-ICSI. METHODS: This retrospective analysis was conducted among 140 infertile couples receiving R-ICSI in from January 2014 to December 2019. The subjects were assigned into a total fertilization failure (TFF)+low DFI group (R-ICSI after TFF and DFI<30%) (n=63), a TFF+high DFI group (R-ICSI after TFF and DFI≥30%) (n=16), a partial fertilization failure (PFF)+low DFI group (R-ICSI after PFF and DFI<30%) (n=52), a PFF+high DFI group (R-ICSI after PFF and DFI≥30%) (n=9). All transferred embryos were come from R-ICSI. The general clinical data [infertility duration, male age, female age, basal serum level of follicle stimulating hormone (FSH), basal serum level of luteinizing hormone (LH), antral follicle count, endometrial thickness of human chorionic gonadotropin (HCG) day, and eggs] and R-ICSI cycle outcomes (fertilization rate, normal fertilization rate, cleavage rate, good embryo rate, implantation rate, clinical pregnancy rate and live birth rate) were analyzed. In addition, the effect of R-ICSI on the fertilization outcome of conventional IVF total fertilization failure and partial fertilization failure was explored. RESULTS: There was no significant difference in the general clinical data and R-ICSI cycle outcome between the TFF+low DFI group and the TFF+high DFI group (all P>0.05). There was no significant difference in the general clinical data between the PFF+low DFI group and the PFF+high DFI group (all P>0.05). The fertilization rate and normal fertilization rate in the PFF+low DFI group were significantly higher than those in the PFF+high DFI group (85.40% vs 72.41%, 71.90% vs 58.62%, respectively; both P<0.05). However, there was no significant difference in cleavage rate, good embryo rate, implantation rate, clinical pregnancy rate, and live birth rate between the 2 groups (all P>0.05). The R-ICSI cycle of TFF: A total of 79 fresh cycles, 57 fresh transplant cycles, a total of 761 unfertilized oocytes, and 584 M II oocytes were treated with R-ICSI, the fertilization rate was 83.22%, the normal fertilization rate was 75.51%, the cleavage rate was 98.15%, the good embryo rate was 40.74%, the implantation rate was 30.56%, and the clinical pregnancy rate was 43.86%; 29 live births were obtained. The R-ICSI cycle of PFF: A total of 61 fresh cycles, 31 fresh transplant cycles, a total of 721 unfertilized oocytes, and 546 M II oocytes were treated with R-ICSI; the fertilization rate was 83.33%, the normal fertilization rate was 69.78%, the cleavage rate was 97.36%, the good embryo rate was 44.39%, the implantation rate was 25.42%, and the clinical pregnancy rate was 45.16%; 12 live births were obtained. CONCLUSIONS: In the case of partial fertilization failure of IVF, the sperm DFI affects the fertilization rate and normal fertilization rate of R-ICSI; whether it is a TFF of IVF or PFF of IVF, ICSI can be used as an effective remedy way.

Novel bi-allelic variants in ACTL7A are associated with male infertility and total fertilization failure.

STUDY QUESTION: What are the genetic causes of total fertilization failure (TFF) in a proband suffering from male infertility? SUMMARY ANSWER: Novel compound heterozygous variants (c.[463C>T];[1084G>A], p.[(Arg155Ter)];[(Gly362Arg)]) in actin-like protein 7A (ACTL7A) were identified as a causative genetic factor for human TFF. WHAT IS KNOWN ALREADY: ACTL7A, an actin-related protein, is essential for spermatogenesis. ACTL7A variants have been reported to cause early embryonic arrest in humans but have not been studied in human TFF. STUDY DESIGN, SIZE, DURATION: We recruited a non-consanguineous family whose son was affected by infertility characterized by TFF after ICSI. Whole-exome sequencing was used to identify the potential pathogenic variants. Artificial oocyte activation (AOA) after ICSI was performed to overcome TFF and any resulting pregnancy was followed up. PARTICIPANTS/MATERIALS, SETTING, METHODS: Sanger sequencing was performed to validate the variants. Pathogenicity of the identified variants was predicted by in silico tools. The ultrastructure of spermatozoa was studied by transmission electron microscopy (TEM). Immunofluorescence staining and western blotting were used to investigate the mechanism of the variants on the affected spermatozoa. MAIN RESULTS AND THE ROLE OF CHANCE: Novel compound heterozygous variants in ACTL7A (c.[463C>T];[1084G>A], p.[(Arg155Ter)];[(Gly362Arg)]) were identified in a family with TFF after ICSI. In silico analysis predicted that the variants lead to a disease-causing protein. TEM showed that the ACTL7A variants caused ultrastructural defects in the acrosome and perinuclear theca. Protein expression of ACTL7A and phospholipase C zeta, a key sperm-borne oocyte activation factor, was significantly reduced in the affected sperm compared to healthy controls, suggesting that the ACLT7A variants lead to an oocyte activation deficiency and TFF. AOA by calcium ionophore (A23187) after ICSI successfully rescued the TFF and achieved a live birth for the patient with ACTL7A variants. LIMITATIONS, REASONS FOR CAUTION: Given the rarity of sperm-associated TFF, only one family with an only child carrying the ACTL7A variants was found. In addition, the TFF phenotype was not assessed in two or more ICSI cycles, due to the intervention in ICSI with AOA after one failed ICSI cycle. Further studies should validate the ACTL7A variants and its effect on male infertility in larger independent cohorts. WIDER IMPLICATIONS OF THE FINDINGS: : Our findings revealed a critical role of ACTL7A in male fertility and identified bi-allelic variants in ACTL7A associated with human TFF, which expands the genetic spectrum of TFF and supports the genetic diagnosis of TFF patients. We also rescued TFF by AOA and obtained a healthy live birth, which provides a potentially effective intervention for patients with ACTL7A pathogenic variants. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Natural Science Foundation of China (81971374 and 81401267). No conflicts of interest were declared. TRIAL REGISTRATION NUMBER: N/A.

Should rescue ICSI be re-evaluated considering the deferred transfer of cryopreserved embryos in in-vitro fertilization cycles? A systematic review and meta-analysis.

BACKGROUND: Total fertilization failure represents a particularly frustrating condition for couples undergoing in vitro fertilization. With the aim of reducing the occurrence of total fertilization failure, intracytoplasmic sperm injection (ICSI) has become the first choice over conventional in vitro fertilization (IVF) procedures although evidence of improved results is still debated and its use in couples without male factor infertility is not recommended. Among the strategies potentially useful to promote the use of conventional IVF, we herein call attention to the late rescue ICSI, which consists in performing ICSI after 18-24 h from conventional insemination on oocytes that show no signs of fertilization. This treatment has however been reported to be associated with a low success rate until recent observations that embryos derived from late rescue ICSI may be transferred after cryopreservation in a frozen-thawed cycle with improved results. The aim of the present study was to assess whether frozen embryos deriving from rescue ICSI performed about 24 h after conventional IVF may represent a valuable option for couples experiencing fertilization failure. METHODS: A systematic review on the efficacy of late rescue ICSI was performed consulting PUBMED and EMBASE. RESULTS: Including twenty-two original studies, we showed that clinical pregnancy rate per embryo transfer and implantation rate obtainable with fresh embryo transfers after rescue ICSI are not satisfactory being equal to 10 and 5%, respectively. The transfer of cryopreserved rescue ICSI embryos seems to offer a substantial improvement of success rates, with pregnancy rate per embryo transfer and implantation rate equal to 36 and 18%, respectively. Coupling rescue ICSI with frozen embryo transfer may ameliorate the clinical pregnancy rate for embryo transfer with an Odds Ratio = 4.7 (95% CI:2.6-8.6). CONCLUSION: Results of the present review support the idea that r-ICSI coupled with frozen embryo transfer may overcome most of the technical and biological issues associated with fresh transfer after late r-ICSI, thus possibly representing an efficient procedure for couples experiencing fertilization failure following conventional IVF cycles. TRIAL REGISTRATION: Prospero registration ID: CRD42021239026 .

A new NLRP5 mutation causes female infertility and total fertilization failure.

OBJECTIVE: The genetic reasons of total fertilization failure (TFF), especially regarding suspected oocyte-related factors, are largely unknown. Only PLCZ1, TLE6 and WEE2 mutations have been shown to result in human fertilization failure. The aim of this study is to report a new NLRP5 mutation which causes female infertility and total fertilization failure. METHODS: A patient presenting TFF after two failed cycles was included. In the first cycle, 13 oocytes were obtained, all of which were inseminated by IVF, but there was no sign of fertilization. In the second cycle, 11 oocytes were obtained, 8 of which were matured; however, none were fertilized upon ICSI and ICSI-AOA. Peripheral blood samples were taken from the patient and her family members. The whole-exome sequencing was performed on them. RESULTS: We identified two heterozygous missense variants NLRP5 (NM_153447.4: c.1598G > C and 1919 T > G; p.Arg533Pro and Leu640Arg) which could cause the incidence of TFF. The first mutation of c.1598G > C (p.Arg533Pro) was inherited from her father and the second mutation of c.1919T > G (Leu640Arg) was inherited from her mother. The sister with normal fertility did not carry these variants. CONCLUSIONS: A new NLRP5 mutation causes female infertility and the incidence of TFF.

Effect of early cumulus cells removal and early rescue ICSI on pregnancy outcomes in high-risk patients of fertilization failure.

OBJECTIVE: The objective of this retrospective study was to evaluate clinical outcomes of early cumulus cells removal and early rescue intracytoplasmic sperm injection (ICSI) in high-risk patients of fertilization failure during human in vitro fertilization (IVF). METHODS: A total of 5,518 patients were enrolled between January 2014 to December 2016. Of which 505 couples performed short insemination with >30% fertilization rate were included in short-term IVF group, 102 patients detected total fertilization failure (TFF) were treated with early rescue ICSI (R-ICSI group), and 4911 couples underwent conventional IVF with overnight co-incubation of gametes (traditional IVF group). The clinical outcomes were analyzed among the three groups. RESULTS: The embryo implantation rates (40.34%, 39.78% and 42.42% for traditional, short-term IVF and R-ICSI groups, respectively) were comparable in the three groups. The clinical pregnancy rates among traditional IVF group, short-term IVF group and R-ICSI group were 57.95%, 57.03% and 60.78%, respectively, and the difference among three groups didn't reach significance. CONCLUSION: The present study indicated that short insemination had no detrimental effects on clinical outcomes in human IVF and could prevent the occurrence of TFF combined with early rescue ICSI for high-risk patients of fertilization failure, which attained acceptable pregnancy outcomes.

Human sperm devoid of germinal angiotensin-converting enzyme is responsible for total fertilization failure and lower fertilization rates by conventional in vitro fertilization.

In conventional in vitro fertilization (IVF), complete failure of fertilization occurs in 5% to 15% of treatments. Although the causes may be unclear, sperm defects appear to be the major contributor. However, a convincing test is not yet available that can predict the risk of fertilization failure. In this study, we found that germinal angiotensin-converting enzyme (gACE) (also called testicular ACE) was undetectable in sperm from patients who had total fertilization failure (TFF) and lower fertilization rates (LFRs) by IVF based on Western blot and indirect immunofluorescence analyses. Additionally, almost all of the patients without gACE on sperm (23 of 25) manifested a TT genotype of the rs4316 single-nucleotide polymorphism of ACE. Overall, our results indicate that the absence of gACE expression is responsible for TFF and LFRs by IVF. The rs4316 polymorphism of ACE might be associated with infertility in those patients. We conclude that sperm lacking gACE may be recognized before commencing IVF and that the patients may be directed instead to consider intracytoplasmic sperm injection.

Total fertilization failure in intra-cytoplasmic sperm injection cycles--classification and management.

In this retrospective cohort study we intended to propose a classification and preliminary management strategy for couples exhibiting total fertilization failure (TFF) in intra-cytoplasmic sperm injection (ICSI) cycles. Sixteen couples with a total of 27 cycles exhibiting TFF, age <40 and/or more than four M2 oocytes aspirated were enrolled. While TFF occurred in 4.3% of all 3723 ICSI cycles, in women younger than 40 with at least 5 M2 oocytes the TFF rate was 0.7%. Indications for ICSI were severe male factor and unexplained infertility. Of the 16 couples with TFF, 4 demonstrated a single episode of TFF, with either subsequent or former normal fertilizations, thus implying possible sporadic faulty laboratory conditions. Ten couples demonstrated repeated total or very low fertilization rates, hinting at a gamete defect not overcome by ICSI. Two couples experienced TFF in the first and only cycle performed at our unit. We conclude that initial and repeated TFF hints at severe gamete defects for which only donor gametes may prove successful while sporadic TFF events could simply imply a technical modifiable condition.

Identification of a new mutation in the ACTL9 gene in men with unexplained infertility.

BACKGROUND: Infertility is defined as the failure to achieve pregnancy after one year of unprotected intercourse within a marital relationship. Approximately 10%-15% of couples worldwide experience infertility issues, with nearly half of these cases attributed to male factors. Among men with unexplained infertility, genetic mutations have been identified as a potential cause. Studies have indicated that mutations affecting the function of the protein encoded by the ACTL9 gene may play a role in male infertility. METHODS: The purpose of this research was to identify mutations in the ACTL9 gene associated with male infertility in a sample of 40 infertile men with unknown causes. Genomic DNA extraction and PCR amplification were carried out on samples from each individual. The genetic material was then analyzed using Sanger sequencing, followed by bioinformatics and segregation analysis to determine the potential effects of the observed variations. RESULT: A novel genetic variant, c.376G>A (p.Glu126Lys), was identified in an infertile male individual, representing a previously unreported finding that was validated through segregation analyses. This specific variant induces a change from glutamate to lysine at the amino acid level by replacing the nucleotide G with A in the genomic DNA sequence, consequently impacting the secondary structure and function of the protein. CONCLUSIONS: The conclusive analysis of the procedure indicated that this alteration has the potential to interfere with the process of fertilization, ultimately resulting in the complete failure of fertilization (TFF) and causing male infertility.

Clinical Applications of Assisted Oocyte Activation in Couples with Various Reproductive Problems: A Systematic Review.

ICSI may face fertilization failure, prompting the use of assisted oocyte activation (AOA) techniques. While AOA is implemented in infertility clinics, its target patients and definitive application remain uncertain. This systematic review adheres to Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines to assess reproductive outcomes in ICSI-AOA cycles compared to conventional ICSI and evaluate AOA effectiveness in various infertility disorders. A literature search encompassed PubMed, Web of Science, EMBASE, and Scopus databases until December 2023 for relevant English studies. Included studies compared ICSI-AOA with conventional ICSI in couples with prior fertilization failure, utilizing diverse AOA methods. Control groups consisted of sibling oocytes, previous cycles of the same couples, or couples undergoing conventional ICSI. Evaluated outcomes included fertilization, cleavage, embryo quality, implantation, pregnancy, and live birth rates. Article screening and data extraction were performed by two authors, with risk of bias assessed by another investigator. Out of 3088 initially identified articles, 30 studies were included, focusing on fertilization failure (n = 10), female infertility (n = 3), PLCζ defects (n = 4), poor sperm quality (n = 4), Globozoospermia (n = 4), and surgically retrieved sperm (n = 8). Most studies concluded that AOA could overcome fertilization failure, but success rates varied based on sperm-related or oocyte-related factors in ICSI-AOA cycles. Due to differences in patient inclusion criteria and sample sizes, most studies were not sufficiently similar for pooled analysis, limiting robust conclusions. There is insufficient evidence, particularly from randomized controlled trials (RCTs), to determine the efficacy or safety of ICSI-AOA as a treatment strategy. Registration number is PROSPERO, CRD42024551221.

Total Fertilization Failure: A Single Center Analysis.

Our main objective was to identify the male and female parameters associated with total fertilization failure (TFF) in IVF with nonmasculine indications. The present work, IRB equivalent INS 63209, is a case-control study that evaluated all cases with TFF after conventional IVF at the Center for Human Reproduction from January 2010 to December 2019 (n = 154). As a control group, we analyzed all patients who did not experience fertilization failure after conventional IVF in the same period (n = 475). We evaluated various parameters, both male and female, assessed during infertility treatment, and only cases without masculine etiology (normal seminal parameters) were included. Ages (female and male) were not different between the groups. Moreover, AMH (anti-Müllerian hormone), semen volume, preprocessing concentration and preprocessing motility were not significantly different (P > 0.05). However, the number of collected oocytes (study versus control groups, median [25-75 interquartile]: 2 [1-5] and 5 [3-8]); MII (2 [1-4] and 5 [2-7]); and postprocessing motility (85 [70-90] and 90 [80-95]) were significantly different between both groups (P < 0.05). Furthermore, a logistic regression analysis including all significant data demonstrated that the number of collected oocytes was significantly related to IVF failure. Patients with fewer than 5 oocytes had an OR of - 1.37 (- 0.938 to - 1.827) for TFF after conventional IVF. Our results showed that a lower follicular response to controlled ovarian stimulation, evidenced by a decreased number of collected oocytes, was the most important parameter associated with IVF failure in nonmasculine infertility.

Assisted gamete treatment to pinpoint acquired meiotic maturity and overcome oocyte activation deficiency contributed by both gametes.

Objective: To treat couples with total fertilization failure (TFF) based on a combined oocyte- and sperm-related oocyte activation deficiency by optimizing oocyte response to chemical activation with calcium ionophore. Design: Case report. Setting: Tertiary Hospital. Patients: Two couples with a history of TFF after intracytoplasmic sperm injection intracytoplasmic sperm injection (ICSI). Interventions: To overcome oocyte-related oocyte activation deficiency (OAD), extended in vivo/in vitro oocyte maturation was performed to enhance ooplasmic maturity; to address sperm-related OAD, assisted gamete treatment (AGT) was performed to trigger oocyte activation. Main outcome measures: Treatment cycle outcomes for the 2 couples undergoing ICSI with extended oocyte maturation (EOM) and AGT. Results: We identified 2 couples with TFF after ICSI because of a combined factor of OAD confirmed by phospholipase C zeta expression and genomic assessment. Initial AGT treatment alone failed to enhance fertilization, suggesting superimposed oocyte dysmaturity prohibiting oocytes from responding to chemical stimuli. To address this complex form of OAD, in couple 1, 27 oocytes out of 34 retrieved presented normal metaphase II spindles after EOM; ICSI with AGT yielded a fertilization rate of 63.0% (17/27). All 17 zygotes were cryopreserved initially. Two embryos were thawed and transferred, yielding a monochorionic diamniotic twin pregnancy. Couple 2 underwent 3 ICSI cycles with EOM and AGT; 91.4% (32/35) of oocytes displayed normal metaphase II spindle and achieved an overall fertilization rate of 43.8% (14/32). A total of 12 blastocysts were cryopreserved. A single 46XY blastocyst was thawed and transferred, resulting in a singleton pregnancy. Conclusions: Our study has demonstrated the usefulness of EOM by targeting spindle presence to enhance chemical responses to AGT.

Corrigendum: A novel assisted oocyte activation method improves fertilization in patients with recurrent fertilization failure.

[This corrects the article DOI: 10.3389/fcell.2021.672081.].

Predicting the unexpected total fertilization failure in conventional in vitro fertilization cycles: What is the role of semen quality?

Background: Male and female gametes factors might contribute to the total fertilization failure (TFF). In first in vitro fertilization (IVF) cycles, decision-making of insemination protocol was mainly based on semen quality for the contribution of female clinical characteristics to TFF remained obscure. The purpose of the study was to evaluate the role of semen quality in predicting unexpected TFF. Methods: A single-center retrospective cohort analysis was performed on 19539 cycles between 2013 and 2021. Two algorithms, a Least Absolute Shrinkage and Selection Operator (LASSO) and an Extreme Gradient Boosting (Xgboost) were used to create models with cycle characteristics parameters. By including semen parameters or not, the contribution of semen parameters to the performance of the models was evaluated. The area under the curve (AUC), the calibration, and the net reclassification index (NRI) were used to evaluate the performance of the models. Results: The prevalence of TFF were .07 (95%CI:0.07-0.08), and .08 (95%CI:0.07-0.09) respectively in the development and validation group. Including all characteristics, with the models of LASSO and Xgboost, TFF was predicted with the AUCs of .74 (95%CI:0.72-0.77) and .75 (95%CI:0.72-0.77) in the validation group. The AUCs with models of LASSO and Xgboost without semen parameters were .72 (95%CI:0.69-0.74) and .73 (95%CI:0.7-0.75). The models of LASSO and Xgboost with semen parameters only gave the AUCs of .58 (95%CI:0.55-0.61) and .57 (95%CI:0.55-0.6). For the overall validation cohort, the event NRI values were -5.20 for the LASSO model and -.71 for the Xgboost while the non-event NRI values were 10.40 for LASSO model and 0.64 for Xgboost. In the subgroup of poor responders, the prevalence was .21 (95%CI:0.18-0.24). With refitted models of LASSO and Xgboost, the AUCs were .72 (95%CI:0.67-0.77) and .69 (95%CI:0.65-0.74) respectively. Conclusion: In unselected patients, semen parameters contribute to limited value in predicting TFF. However, oocyte yield is an important predictor for TFF and the prevalence of TFF in poor responders was high. Because reasonable predicting power for TFF could be achieved in poor responders, it may warrant further study to prevent TFF in these patients.

Comparing the pregnancy outcomes of Re­ICSI and ICSI embryos in fresh ET and FET cycles.

Early rescue intracytoplasmic sperm injection (Re-ICSI) can prevent total fertilization failure (TFF) during conventional in vitro fertilization (IVF). However, the implantation rate of Re-ICSI embryos is lower than that of direct ICSI during fresh embryo transfer (ET). The aim of the present study was to investigate the effect of frozen ET (FET) after Re-ICSI. In the present retrospective study, primary infertility patients that underwent the first Re-ICSI and ICSI treatment, were studied. The clinical pregnancy rate, implantation rate, ectopic pregnancy, abortion rate and live birth rate were analyzed between the Re-ICSI and ICSI groups in fresh ET and FET cycles. The average age of patients between Re-ICSI and ICSI groups in fresh ET and FET cycles was (29.0±3.2 vs. 29.1±3.1, and 29.1±3.3 vs. 28.9±3.0), respectively (P>0.05). Compared with ICSI embryos, the clinical pregnancy, implantation and live birth rates of Re-ICSI embryos were lower in fresh ET cycles. By contrast, there were no significant differences in the pregnancy, implantation and live birth rates between the Re-ICSI and ICSI embryos during the FET cycles. Re-ICSI coupled with FET may overcome the impaired outcomes in fresh ET.

Cumulative live birth rate and neonatal outcomes after early rescue ICSI: a propensity score matching analysis.

STUDY QUESTION: Is early rescue ICSI (E-RICSI) an effective and safe technique compared to conventional ICSI? SUMMARY ANSWER: Despite the higher multi-pronucleus (PN) rate compared to conventional ICSI, E-RICSI did not add extra risks to clinical and neonatal outcomes. WHAT IS KNOWN ALREADY: Based on the finding that the second polar body was released in 80% of fertilized oocytes by 4 h after exposure to spermatozoa and in ∼90% of fertilized oocytes by 6 h, E-RICSI brings forward the timing of rescue ICSI to 6 h after initial insemination, and effectively prevents oocyte aging and embryo-uterus asynchrony. However, some researchers still voice concerns about the efficacy and safety of E-RICSI, and comparative studies are limited. STUDY DESIGN SIZE DURATION: A retrospective cohort study was conducted on patients who underwent conventional ICSI or E-RICSI treatment between January 2015 and December 2020 at a university-affiliated hospital. Using 1:1 propensity score matching, 1496 cases entered each group. PARTICIPANTS/MATERIALS SETTING METHODS: In total, 1496 couples undergoing conventional ICSI oocyte retrieval cycles and 1496 undergoing E-RICSI oocyte retrieval cycles were enrolled in this study, and basic clinical characteristics, embryologic data, clinical outcomes and neonatal data were compared between groups. The embryos in the E-RICSI group were divided into two subgroups: those fertilized by iIVF (IVF subgroup) and those fertilized by E-RICSI (E-RICSI subgroup); the embryologic data, clinical outcomes, and neonatal data for these subgroups were also compared with the conventional ICSI group. Logistic regression was used for statistical analysis with potential confounder adjustment. MAIN RESULTS AND THE ROLE OF CHANCE: The 2PN rate, blastocyst formation rate, and viable blastocyst formation rate of the E-RICSI group were significantly lower compared to the conventional ICSI group (2PN rate: P < 0.001; blastocyst formation rate: P < 0.001; viable blastocyst formation rate: P = 0.004), and the multi-PN rate in the E-RICSI group was significantly higher than the conventional ICSI group (P < 0.001). However, the number of 2PN embryos, normal cleavage embryo rate, Day 3 high-quality cleavage embryo rate, and high-quality blastocyst rate were similar between groups. When considering the IVF embryos and E-RCSI embryos in the E-RICSI group independently, the 2PN rate of the conventional ICSI group was significantly lower than E-RICSI subgroup but higher than the IVF subgroup, whereas the blastocyst formation rate and viable blastocyst formation rate were higher than E-RICSI embryos but comparable to IVF embryos. As for the clinical and neonatal outcomes, the implantation rate of the E-RICSI subgroup was significantly lower than the IVF subgroup but comparable to the conventional ICSI group, while the low birthweight (LBW) rate was significantly lower compared with the conventional ICSI group but similar with the IVF subgroup. No other differences were observed among the three groups for cumulative clinical pregnancy rate, cumulative live birth rate, and the pregnancy outcomes per transfer including clinical pregnancy, ectopic pregnancy, miscarriage, and live birth, either in fresh or frozen embryo transfer cycles. Furthermore, neonatal outcomes, including cesarean section, sex ratio, LBW, preterm birth, and macrosomia, were similar among groups. LIMITATIONS REASONS FOR CAUTION: This study is limited by the retrospective design, limited sample size, and short follow-up period. However, our study underlies the need for large-scale, multi-center randomized controlled trials with long-term follow-up. WIDER IMPLICATIONS OF THE FINDINGS: Short-term insemination (3 h) combined with E-RICSI may be a safe and effective method to prevent the occurrence of total fertilization failure, and patients with normal or borderline sperm could be encouraged to try IVF first. STUDY FUNDING/COMPETING INTERESTS: This study was supported by grants from the National Key & Development Program of China (No. 2021YFC2700603) and the National Natural Science Foundation of China (No. 81801443). The authors declare no conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.