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

ObjectiveTo test a novel method to select spermatozoa with high chromatin integrity.DesignSpecimens with high sperm chromatin fragmentation (SCF) were selected by density gradient selection (DGS) and microfluidic sperm sorting (MSS).SettingAcademic medical center.Patient(s)Ejaculates from consenting men were processed by DGS/MSS. Couples underwent ICSI cycles with spermatozoa processed by DGS/MSS. Clinical outcomes were evaluated after embryo transfer.Intervention(s)SCF was measured by TUNEL. ICSI with spermatozoa selected by DGS and MSS was performed.Main outcome measure(s)Fertilization, embryo implantation, and pregnancy outcomes were compared between DGS and MSS.Result(s)A total of 23 men had an average SCF of 20.7 ± 10%. After DGS and MSS, the SCF was 12.5 ± 5% and 1.8 ± 1%, respectively. In couples who underwent ICSI, the average SCF was 28.8 ± 9%, which fell to 21.0 ± 9% after DGS and 1.3 ± 0.7% after MSS. Four couples underwent 11 ICSI cycles with DGS and achieved one (25%) pregnancy that resulted in pregnancy loss. In four subsequent ICSI cycles with MSS, an ongoing clinical pregnancy rate of 50% was achieved. Five additional couples underwent 12 cycles of ICSI with DGS. After preimplantation genetic testing for aneuploidy, 30.3% of the embryos were euploid. One pregnancy was achieved, resulting in pregnancy loss. With MSS, 31.5% of the embryos were euploid and 4 couples obtained a pregnancy. Finally, sixteen couples underwent 20 ICSI cycles solely with MSS at our center. Of these couples, 8 had failed 13 ICSI cycles with DGS elsewhere. These couples achieved an overall implantation of 34.5% (10/29) and a pregnancy rate of 58.8% (10/17).Conclusion(s)Microfluidic selection yielded spermatozoa with optimal genomic integrity and improved chances of obtaining a euploid conceptus.

Objective: To identify and treat the gamete responsible for complete fertilization failure with intracytoplasmic sperm injection (ICSI) using a newly proposed assisted gamete treatment (AGT). Design: Prospective cohort study. Setting: Center for reproductive medicine. Patient(s): One-hundred and fourteen couples with an adequate number of spermatozoa for ICSI and a fertilization rate of %10%, after controlling for maternal age. Intervention(s): 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. Main Outcome Measure(s): Treatment cycle outcome for couples undergoing ICSI with either a modified superovulation protocol or AGT compared with their historical cycle. Result(s): 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 Cz (PLCz) 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 AE standard deviation: maternal age, 35.6 AE 5 years; paternal age, 39.8 AE 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. Conclusion(s):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 PLCz 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. (Fertil Steril Ò 2020;114:535-44. Ó2020 by American Society for Reproductive Medicine.) El resumen está disponible en Español al final del artículo.

Evaluation of reproductive quality of spermatozoa by standard semen analysis is often inadequate to predict ART outcome. Men may be prone to meiotic error and have higher proportion of spermatozoa with fragmented chromatin, capable of affecting the conceptus' health. In men with unexplained infertility, supplementary tests may be pivotal to gain insight into the paternal contribution to the zygotic genome. A total of 113 consenting men were included in the study, with an additional 5 donor specimens used as control. Among study participants, 87 were screened for sperm aneuploidy by fluorescent in situ hybridization (FISH) and ranked according to their increasing age. A total of 18 men were assessed by whole exome sequencing and categorized according to their reproductive outcome as either fertile or infertile. Another set of men (n = 13) had their gene expression analyzed by RNA-seq and were profiled according to their reproductive capacity. FISH revealed that the average aneuploidy rate was highest for men over-55 age group (9.6%), while men >55 had the highest average disomy for chromosomes 17(1.2%) and 18(1.3%). ART results for the entire cohort comprised 157 cycles, stratified by paternal age. The youngest age group (25–30 years) had a fertilization rate of 87.7% which decreased to 46.0% in the >55 age group. Clinical pregnancy rate was highest in the 25-30yr group (80.0%) while no pregnancies were attained in the >55 age groups. Pregnancy loss was characterized by a steadily increasing trend, highest in the 51–55 age group (50.0%). NGS was performed on a cohort of patients classified as having recurrent pregnancy loss. This cohort was classified as the infertile group (n = 10) and was compared to a control group (n = 8) consisting of patients successfully treated by ART. Eight couples in 17 ICSI cycles achieved a clinical pregnancy rate of 82.4% while 10 infertile couples treated in 21 cycles achieved a pregnancy rate of 23.8%, all resulting in pregnancy loss. DNA-sequencing on spermatozoa from these patients yielded overall aneuploidy of 4.0% for fertile and 8.6% for the infertile group (P<0.00001). In the infertile cohort, we identified 17 genes with the highest mutation rate, engaged in key roles of gametogenesis, fertilization and embryo development. RNA-seq was performed on patients (n = 13) with normal semen analyses. Five men unable to attain a pregnancy after ART were categorized as the infertile group, while 8 men who successfully sustained a pregnancy were established as the fertile control. Analysis resulted in 86 differentially expressed genes (P<0.001). Of them, 24 genes were overexpressed and 62 were under-expressed in the infertile cohort. DNA repair genes (APLF, CYB5R4, ERCC4 and TNRFSF21) and apoptosis-modulating genes (MORC1, PIWIL1 and ZFAND6) were remarkably under-expressed (P<0.001). Sperm aneuploidy assessment supported by information on gene mutations may indicate subtle dysfunctions of the spermatozoon. Furthermore, by querying noncoding RNA we may gather knowledge on embryo developmen...