Sperm Aneuploidy Rate Research Articles

Genetic profiling of azoospermic men to identify the etiology and predict reproductive potential

PurposeTo identify germline mutations related to azoospermia etiology and reproductive potential of surgically retrieved spermatozoa, and to investigate the feasibility of predicting seminiferous tubule function of nonobstructive azoospermic men by transcriptomic profiling of ejaculates.Materials and methodsSperm specimens were obtained from 30 men (38.4 ± 6 years) undergoing epididymal sperm aspiration for obstructive azoospermia (OA, n = 19) acquired by vasectomy, or testicular biopsy for nonobstructive azoospermia (NOA, n = 11). To evaluate for a correlation with azoospermia etiology, DNAseq was performed on surgically retrieved spermatozoa, and cell-free RNAseq on seminal fluid (n = 23) was performed to predict spermatogenesis in the seminiferous tubule.ResultsOverall, surgically retrieved sperm aneuploidy rates were 1.7% and 1.8% among OA and NOA cohorts, respectively. OA men carried housekeeping-related gene mutations, while NOA men displayed mutations on genes involved in crucial spermiogenic functions (AP1S2, AP1G2, APOE). We categorized couples within each cohort according to ICSI clinical outcomes to investigate genetic causes that may affect reproductive potential. All OA-fertile men (n = 9) carried mutations in ZNF749 (sperm production), whereas OA-infertile men (n = 10) harbored mutations in PRB1, which is essential for DNA replication. NOA-fertile men (n = 8) carried mutations in MPIG6B (stem cell lineage differentiation), whereas NOA-infertile individuals (n = 3) harbored mutations in genes involved in spermato/spermio-genesis (ADAM29, SPATA31E1, MAK, POLG, IFT43, ATG9B) and early embryonic development (MBD5, CCAR1, PMEPA1, POLK, REC8, REPIN1, MAPRE3, ARL4C). Transcriptomic assessment of cell-free RNAs in seminal fluid from NOA men allowed the prediction of residual spermatogenic foci.ConclusionsSperm genome profiling provides invaluable information on azoospermia etiology and identifies gene-related mechanistic links to reproductive performance. Moreover, RNAseq assessment of seminal fluid from NOA men can help predict sperm retrieval during testicular biopsies.

O-183 Frequent spontaneous abortion in pregnancies followed by ICSI using frozen sperm from patients with testicular germ cell tumor (TGCT)

Abstract Study question Despite the high rate of sperm chromosome abnormalities in testicular germ cell tumor (TGCT), why is newborn aneuploidy rate in pregnancies with TGCT patient normal? Summary answer Miscarriage rate is higher in the pregnancy of TGCT patients than of non-TGCT cancer patients, although the pregnancy rate in ICSI was not significantly different. What is known already Previous reports showed higher sperm aneuploidy in TGCT patients than control males. The sperm aneuploidy rate is high before treatment, and after radical treatments peaked at 6 months after treatment and remained high until 24 months after treatment. However, in the Swedish government base study, the rate of newborn malformations including aneuploidy in all pregnancy with TGCT patients was higher than without TGCT, but not statistically significant. In only natural pregnancies, the rate was not significantly different between the two groups. The cause for discrepancy within sperm aneuploidy and newborn aneuploidy was not well clarified. Study design, size, duration To clarify the mechanism for the purifying selection of aneuploid sperm in conception, we examined ICSI cases with TGCT patients that did not include natural sperm competition and compared to controls with non-TGCT cancer patient, who have normal rate of aneuploid sperm. By investigating ICSI outcomes, we aimed to determine which stage of embryonic development are affected by the sperm of TGCT patients. Participants/materials, setting, methods Under ethical review of Yokohama city university, the 10 TGCT patients (123 oocytes) and 16 non-TGCT cancer patients (251 oocytes) who underwent ICSI with their frozen sperm from 2012 to 2021 were enrolled. Fertilization, embryo viability and embryo transfer outcomes (pregnancy and miscarriage rate) were examined between two groups underwent ICSI with cryopreserved sperm for fertility preservation. Clinical information was retrospectively collected from medical records. Main results and the role of chance The patients' age of sperm cryopreservation was 21-56 years old. 10 cases of TGCT were diagnosed as seminoma (3), non seminoma (6), and unknown (2). 15 cases of non-TGCT were diagnosed as malignant blood diseases (9), prostate cancer (2), bladder cancer (1), and others (3). In both groups, all patients were treated by ICSI with cryopreserved sperm obtained prior to chemotherapy. The mean age of female partners in TGCT was 33.2±3.5 years and not different with 35.8±3.4 years in non-TGCT meaning no different age factor. The fertilization rate, viable embryo rate, pregnancy rate and chemical abortion rate of TGCT vs non-TGCT group were 76.4% vs 67.7%, 64.3% vs 61.7%, 37.1% vs 21.7%, 7.1% vs 23.1%, respectively. Those developmental evaluations were not significantly different between two groups. However, spontaneous abortion rate was significantly higher 46.2% in TGCT group than 10% in non-TGCT group (p < 0.05). Further, in our follow-up, no congenital malformations in the babies born in either group (5 babies in TGCT vs 7 babies in non-TGCT group). The outcome of ICSI using sperm in TGCT patients show normal pregnancy rate but include higher spontaneous abortions rate, suggesting aneuploid embryo were negatively purifying selected under post-implantation stage. Limitations, reasons for caution Because ICSI with fertility-preserving frozen sperm in TGCT patient is rare even in reproduction center in general university hospital, single center analysis is still small and limited. More case reports and studies for TGCT fertility preservation are needed for more accurate evaluation. Wider implications of the findings In TGCT patients, chromosome aberrations and DNA fragmentation of sperm may not be apparent in natural pregnancies with normal sperm competition, but may become apparent as spontaneous abortions when ICSI were performed, suggesting PGT-A can predict and avoid the hidden risk of repeated pregnancy loss in ICSI to TGCT patients. Trial registration number not applicable

Globozoospermia: A Case Report and Systematic Review of Literature.

Globozoospermia is a genetic syndrome characterized by the presence of round-headed spermatozoa and infertility due to the inability of these spermatozoa to fertilize the oocyte. In this article, we present the clinical case of a young globozoospermic patient with a new, not yet described mutation of the DPY19L2 gene. We also performed a systematic review of the literature on gene mutations, the outcome of assisted reproductive techniques, and the risk of transmission of abnormalities to the offspring in patients with globozoospermia and made recommendations to offer a more appropriate clinical management of these patients. We performed a systematic search in the PubMed, Google Scholar, and Scopus databases from their inception to December 2021. The search strategy included the combination of the following Medical Subjects Headings (MeSH) terms and keywords: "globozoospermia", "round-headed spermatozoa", "round head spermatozoa", "intracytoplasmic sperm injection", "ICSI", "offspring", "child health", "assisted reproductive technique outcome". All the eligible studies were selected following the PECOS (Population, Exposure, Comparison/Comparator, Outcomes, Study design) model. The quality of included studies was assessed by applying the "Cambridge Quality Checklists". The main genes involved in the pathogenesis of globozoospermia are DPY19L2, SPATA16, PICK1, GGN, SPACA1, ZPBP, CCDC62, and CCNB3 genes. Other genes could also play a role. These include C2CD6, C7orf61, CCIN, DNH17, DNH6, PIWIL4, and CHPT1. Globozoospermic patients should undergo ART to achieve fertility. In particular, intracytoplasmic sperm injection with assisted oocyte activation or intracytoplasmic morphologically-selected sperm injection appears to be associated with a higher success rate. Patients with globozoospermia should also be evaluated for the high rate of sperm aneuploidy which appears to influence the success rate of ART but does not appear to be associated with an increased risk of transmission of genetic abnormalities to offspring. This systematic review summarizes the evidence on the gene panel to be evaluated, ICSI outcomes, and the health of the offspring in patients with globozoospermia. Evidence-based recommendations on the management of patients with globozoospermia are provided.

THE EFFECT OF GERMLINE MUTATIONS ON THE ABILITY OF SURGICALLY RETRIEVED SPERMATOZOA FROM AZOOSPERMIC MEN TO SUPPORT EMBRYONIC DEVELOPMENT

To identify eventual germline mutations involved in spermatogenesis and the ability of spermatozoa retrieved from azoospermic men to support embryonic development. For over 20 months, we recruited infertile men undergoing epididymal aspiration for obstructive (OA; n=17) and testicular retrieval for nonobstructive (NOA; n=10) origins. Three men functioned as fertile controls. Gene mutations were compared according to the etiology of the azoospermia and in relation to whether they were able to generate a pregnancy (fertile) or not (infertile). Specimens were provided by consenting men, who were being treated at our center for infertility. DNA was extracted and amplified from at least 500 spermatozoa (DNA concentration, 705±562 ng/ul; quality, 1.7±0.1 nm). Following NGS, gene mutations, duplications, and deletions were detected using the CLC Genomic Server 9.0. Genes were considered duplicated or deleted when the read depth was >1.5 or <0.5 times the median read depth in the control. Common gene mutations in the OA and NOA cohorts were assessed according to the couples’ clinical outcomes. Of the 27 couples (paternal age, 41.3±5yrs) included, 17 OA men underwent epididymal sperm retrieval with an average concentration of 1.3±3x106/ml and 7±14% motility, while 10 NOA men yielded twitching spermatozoa with a concentration of 0.03±0.2x106/ml and 0.5±1% motility. Whole genome karyotype assessment showed an overall sperm aneuploidy rate below only 2%, specifically 1.7% in the OA and 1.9% in the NOA cohort. In the OA group overall, only 3 housekeeping genes were mutated (ATP4A, SLC17A7, and OR1D4). However, in the NOA cohort, 5 genes involved in RNA transcription (POLR2L), apoptosis (AP5M1), and basic spermiogenic function (AP1S2, AP1G2, and APOE) were deleted. The OA men and their partners (maternal age, 34.8±3yrs) underwent 17 ICSI cycles, resulting in a pregnancy and delivery rate of 47.1% (8/17). For those able to reproduce (n=8), all shared a mutation in ZNF749, which is mainly responsible for spermatogenesis. Those who remained infertile had a common mutation in PRB1, a gene involved in DNA replication. When NOA men were treated in 10 ICSI cycles with their partners (maternal age, 38.2±2yrs), the pregnancy rate was 70% (7/10). Of those 7 men, 1 mutation (MPIG6B) related to stem cell lineage differentiation was in common. The remaining 6 who remained childless had mutations in genes associated with spermatogenesis (6 genes), apoptosis (4 genes), acrosomal function (2 genes), and, most importantly, in early embryonic development (MBD5, CCAR1, PMEPA1, POLK, REC9, REPIN1, MAPRE3, and ARL4C). Screening infertile men for germline mutations can help characterize spermatogenic function in relation to the etiology of their azoospermia and provide valuable information on their reproductive potential. In men capable of reproducing, gene mutations were limited to those associated with spermatogenic function. For men unable to reproduce, particularly the NOA cohort, the mutated genes were mostly related to multiorgan functions and embryo development.

Aneuploidy Rate, DNA Fragmentation, Tubulin and Centrin Content in Different Groups of Male Infertility

In this study, we aimed to compare DNA fragmentation ratios, centrosomal protein concentrations (centrin and tubulin) and numerical chromosomal abnormalities between four groups including asthenozoospermia, teratozoospermia, oligoasthenozoospermia and normozoospermia patients, and to emphasize the necessity of investigating genetic and protein content of the spermatozoa also in patients with a semen analysis within normal reference range. Evaluation of semen analysis of the patients was performed based on WHO 2010 criteria. As a result of the statistical analysis, it was observed that the sperm DNA fragmentation rate in the normozoospermia group (6.50 ± 3.46%) was significantly lower than those in the teratozoospermia (19.69 ± 9.86%) and oligoasthenoteratozoospermia group (32.47± 14.13%) (p<0.001). For centrin and tubulin proteins, highest concentrations were observed in normozospermia group (p<0.05). When incidence of numerical abnormalities were compared between four groups, it was found that there were statistically significant differences between groups in terms of nullisomies X, Y and 13, and 21 (p<0.05). In conclusion, we suggest that abnormalities in semen parameters seem to be associated with chromosomal abnormalities; and sperm aneuploidy rate is also at a considerable level even in normozoospermic patients. Keywords: DNA fragmentation, teratozoospermia, centrin, tubulin DOI: 10.7176/JMPB/68-04 Publication date: September 30 th 2020

Sperm aneuploidy and recurrent pregnancy loss: A systematic review and meta-analysis

Background and Objective:Studies have reported that the spouses of patients with recurrent miscarriage have a high rate of sperm aneuploidy,The types and the methods of determination of sperm aneup...

Cannabis consumption might exert deleterious effects on sperm nuclear quality in infertile men.

Can cannabis consumption alter sperm nuclear integrity in infertile men? A retrospective cross-sectional study conducted between July 2003 and December 2013, which included 54 men who consulted for male-factor infertility. Twenty-seven infertile men who were regular cannabis users were matched to 27 infertile men who were cannabis non-users. To complement the conventional semen parameter and plasma hormone level assessments, sperm nuclear alterations were explored using fluorescence in-situ hybridization to assess numerical chromosomal abnormalities, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling to investigate DNA fragmentation, aniline blue staining to examine chromatin condensation and a motile sperm organelle morphology examination to detect vacuoles in sperm heads. The rates of sperm aneuploidy (P=0.0044), diploidy (P=0.037), total chromosome abnormalities (P = 0.0027) and DNA fragmentation (P = 0.027) were significantly higher in cannabis users than in non-cannabis users. Cannabis consumption might have deleterious effects on sperm nuclear quality in infertile men by increasing numerical chromosome abnormalities and DNA fragmentation. Cannabis consumption induces these detrimental effects on the progression of spermatogenesis from meiotic stages to spermiogenesis and potentially on post-testicular sperm maturation in infertile men. Any potential findings, however, need to be validated with larger sample size, and our data are only exploratory findings.

Paternal age: Negative impact on sperm genome decays and IVF outcomes after 40 years.

This study assessed sperm quality declining on relation to paternal age and its impact on in vitro fertilization (IVF) outcomes in order to estimate the APA (Advanced Paternal Age) cutoff. For this, 83 couples undergoing IVF treatment for male factor infertility were enrolled. The women age was ≤39 years, whereas the men were divided in two groups: APA (n = 41; age ≥ 40 years) and young (Y) (n = 42; age < 40 years). Conventional semen parameters (volume, concentration, motility, vitality, and morphology) were analyzed in the collected sperm samples. Furthermore, sperm genome decays (SGD) was assessed by TUNEL assay (DNA fragmentation), aniline blue staining (chromatin decondensation), and fluorescent in situ hybridization (aneuploidy). No significant difference was found concerning the conventional semen parameters between APA and Y groups. Conversely, SGD analysis showed increased DNA fragmentation; chromatin decondensation and sperm aneuploidy rates in the APA group (respectively, 41%, 43%, and 14% vs. 25%, 23%, and 4% in Y group). IVF outcomes also were affected by paternal age as indicated by the rates of cancelled embryo transfers, clinical pregnancy and miscarriage in the two groups APA and Y (29%, 17%, and 60% vs. 10%, 32%, and 42%). Finally, statistical analysis of the results suggests that the age of 40 should be considered as the APA cutoff during ART attempts.

Sperm aneuploidy after testicular cancer treatment: data from a prospective multicenter study performed within the French Centre d’Étude et de Conservation des Oeufs et du Sperme network

To study sperm aneuploidy in a population of testicular cancer (TC) patients treated with the use of either bleomycin-etoposide-cisplatin (BEP) chemotherapy or radiotherapy. Multicenter prospective longitudinal study of TC patients analyzed before treatment and after 3, 6, 12, and 24months (T3-T24). Fifty-four TC patients and a control group of 10 fertile sperm donors. University hospital laboratories. Routine semen analyses; sperm aneuploidy and diploidy. Comparison of sperm characteristics and sperm chromosome abnormalities during TC patient follow-up. Semen characteristics recovered pretreatment values 12months after radiotherapy and 24months after more than two BEP cycles. A significant increase in sperm disomy YY and XX was observed in the TC group before treatment compared with the control group. After more than two BEP cycles, the mean sperm aneuploidy rate increased significantly at T12 and reached the pretreatment value at T24. After radiotherapy, the mean sperm aneuploidy returned to the pretreatment value at T12. At T24, nearly 40% of TC patients did not recover their pretreatment sperm aneuploidy rate. Genetic counseling of TC patients should include information on the potential elevated risk of aneuploid conceptus from sperm recovered after treatment and the necessity to postpone conception up to ≥12months after radiotherapy and ≥24months after more than two BEP chemotherapy cycles. However, few men receiving one or two BEP cycles and some dropouts are the main limitations of this study.

Impact of Hodgkin or non-Hodgkin lymphoma and their treatments on sperm aneuploidy: a prospective study by the French CECOS network

To assess sperm production and aneuploidy in Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) before and after treatments. Multicenter, prospective, longitudinal study of lymphoma patients analyzed before treatment and after 3, 6, 12, and 24months. University hospitals. Forty-five HL and 13 NHL patients were investigated before and after treatment. Treatment regimens were classified in two groups: ABVD (doxorubicin, bleomycin, vinblastine, dacarbazine) with or without (±) radiotherapy, and CHOP (doxorubicin, cyclophosphamide, vincristine, prednisone)/MOPP-ABV (mechlorethamine, oncovin, procarbazine, prednisone-doxorubicin, bleomycin, vinblastine). A control group of 29 healthy men was also studied. Semen analyses and aneuploidy study by FISH were performed at each time point. Comparison of mean sperm characteristics and percentage of sperm aneuploidy rates before and after treatment. Before treatment, HL and NHL men had altered semen characteristics and higher sperm aneuploidy rates (median 0.76 [interquartile range 0.56-0.64]) than the control group (0.54 [0.46-0.74]). After treatment, sperm production was significantly lowered 3 and 6months after ABVD ± radiotherapy or CHOP/MOPP-ABV. After ABVD ± radiotherapy, the aneuploidy rate increased significantly only at 3months, and values obtained 1 or 2years later were lower than pretreatment values. In contrast, in the CHOP/MOPP-ABV treatment group, semen characteristics and aneuploidy rate did not return to normal levels until 2years after treatment. Lymphoma itself has consequences on sperm aneuploidy frequency before treatment. Moreover, lymphoma treatments have deleterious effects on sperm chromosomes related to treatment type and time since treatment. Patient counseling is essential concerning the transient but significant sperm aneuploidy induced by lymphoma and its treatments.

Effect of environmental endocrine disruptor's exposure on sperm quality and aneuploidy rates in fertile sperm donors

Effect of environmental endocrine disruptor's exposure on sperm quality and aneuploidy rates in fertile sperm donors

Chromosomal polymorphic variants increase aneuploidies in male gametes and embryos

ABSTRACTChromosomal polymorphisms involve heterochromatic regions and occur in the general population. However, previous studies have reported a higher incidence of these variants in infertile patients. The aim of this study was to examine the relationship between polymorphic variants and infertility and their association with aneuploidies in male gametes and embryos. We retrospectively considered 1,551 cytogenetic studies involving infertile patients (study group; n=866) and oocyte/sperm donors as the control group (n=685). We had detected 168 polymorphisms in the study group and 92 in the control group. An increase in the frequency of polymorphic variants was observed among infertile patients (19.4% study group vs. 13.4% control group; P < 0.01). Sperm aneuploidies among 145 infertile men were evaluated by fluorescent in situ hybridization (FISH). The frequency of infertile men with increased rates of sperm aneuploidy was higher among polymorphism carriers. Twenty men showed an abnormal rate of sperm aneuploidy in the carrier group (n=53) vs. 15 in the non-carrier group (n=92) (37.7% vs. 16.3%, respectively; P < 0.01). Finally, aneuploidies in blastocysts (n=301) resulting from donated oocytes were also examined by array comparative genomic hybridization (array-CGH). Significant differences were reported in the embryo aneuploidy rate between female carriers and non-carriers in oocyte donation cycles (50.0% vs. 27.6%; P < 0.001). This study suggests that polymorphic variants have an impact on fertility. Moreover, our results show a relationship between polymorphisms and aneuploidy in spermatozoa and embryos.Abbreviations: FISH: fluorescent in situ hybridization; CGH: comparative genomic hybridization; ESHRE: European Society of Human Reproduction and Embryology; ASRM: American Society for Reproductive Medicine; RPL: recurrent pregnancy loss; WHO: World Health Organization; ISCN: International System for Human Cytogenetic Nomenclature guidelines; WGA: whole genome amplification; SPSS: Statistical Package for Social Sciences

Shedding Light on the Nature of Seminal Round Cells.

IntroductionIn this investigation we assess the incidence of round cells (RCs) in semen samples in our infertile patient population and their significance on intracytoplasmic sperm injection (ICSI) cycle outcomes. We also evaluate the usefulness of RCs as indicators of bacterial infection and highlight the origin of this cell-type, as well as its role in the human ejaculate.Patients and MethodsIn a prospective fashion, a total of 4,810 ejaculated samples were included in the study during a period of 24 months. RCs were characterized for white blood cell (WBC) components versus exfoliated germ cells by testing for multiple markers of ploidy as well as protamine assays. Cases displaying ≥ 2 x 106/ml RCs were screened for bacteria. Raw specimens containing RC were processed by peroxidase and other leukocyte assays, specific stains for protamines were used to identify spermiogenic stage, aneuploidy (FISH) assessment was carried out, and the presence of various Sertoli-cell cytoplasmic remnants was analyzed to identify and characterize immature germ cells. The effect of RC on clinical outcome was assessed in specimens used for ICSI.ResultsThe average age of the men involved was 39.2 ± 7 years. Semen samples had a mean concentration of 40.7 ± 31 x 106/ml, motility of 42.6 ± 35%, and morphology of 2.3 ± 2%. RCs were identified in 261 specimens, representing a proportion of 5.4%. Men with RCs had comparable age but lower sperm concentration and morphology than the control group (P<0.001). The aneuploidy rate of 4.3% in RCs group was remarkably higher than the control group (2.3%; P<0.001). Sperm aneuploidy rate positively correlated with the number of RCs (P<0.001). Of 44 men, 17 of them in 18 cycles had up to 1.9 x 106/ml RCs without affecting fertilization and clinical pregnancy rates when compared to controls (n = 365 cycles). In 27 men undergoing 33 ICSI cycles with ≥ 2 x 106/ml RCs, the fertilization rate trended lower and the miscarriage rate was significantly increased (P = 0.05). There was lack of correlation between RC and bacteriological growth. Specific markers indicated that seminal RCs are mostly immature germ cells encased in the remnants of Sertoli cell cytoplasm. Moreover, their modest protamine content and their haploid status confirm that they are post-meiotic. Sequential observation in the same man showed that RC episodes were followed by an amelioration of semen parameters, and interestingly, the episodic occurrence of RCs often coincides with flu season peaks.ConclusionsSeminal RCs are not a marker of infectiousness but rather a transient indicator of spermatogenic insult that possibly occurs in most men following a mild and transient ailment such as the flu.

Relationship between sperm aneuploidy, sperm DNA integrity, chromatin packaging, traditional semen parameters, and recurrent pregnancy loss

To study the possible relationship between sperm aneuploidy, sperm DNA integrity, chromatin packaging, traditional semen parameters, and recurrent pregnancy loss (RPL). Descriptive study. University-affiliated tertiary teaching. A total of 22 couples with history of RPL and 20 fertile men. Semen samples from case and control men were examined for differences in semen parameters, DNA fragmentation, chromatin condensation, and sperm aneuploidy. Sperm DNA and chromatin integrity and sperm aneuploidy. Sperm progressive motility (30.2% vs. 51.5%) was significantly lower and abnormal morphology (74.8% vs. 54.2%) was significantly higher in the RPL group versus the control group, respectively. The percentage of fragmented DNA was significantly increased in the RPL group (17.1% vs. 10.2%) as well as the rate of spermatozoa with nuclear chromatin decondensation (23.6% vs. 11.8%). There was a significantly higher sperm aneuploidy rate among the RPL group as well. The increase in abnormal sperm parameters, sperm DNA fragmentation, nuclear chromatin decondensation, and sperm aneuploidy suggest possible causes of unexplained RPL.

Sperm Selection Method in ICSI

Abstract Since the establishment of intracytoplasmic sperm injection (ICSI) in 1992, sperms have been observed, evaluated, and selected under an inverted microscope at 400 × magnification for ICSI. Intracytoplasmic morphologically selected sperm injection (IMSI) is a technique in which sperms are selected for micro insemination by more precisely observing their morphology, at increased magnification under an inverted microscope used for micro-insemination and enhancing the image resolution. Approximately 20 years have passed since IMSI was developed, and its efficacy has only been confirmed in cases with a high rate of sperm aneuploidy, a high rate of sperm DNA fragmentation, and repeated ICSI failures. It is well-known that the quality of sperms has a profound effect on the quality of embryos which may eventually become children. Therefore, when micro-insemination is performed, sperms should be selected by not only paying attention to their motility, but also by closely observing their morphology. Those ...

Fluorescence in situ hybridization detects increased sperm aneuploidy in men with recurrent pregnancy loss

To investigate, in men presenting with recurrent pregnancy loss (RPL), the prevalence of sperm autosome and sex chromosome aneuploidy. Retrospective study. Male infertility clinic. A total of 140 men with RPL provided semen samples, and five normozoospermic controls provided 140 semen samples for comparison. Recurrent pregnancy loss, documented in the female partners, was defined as a prior miscarriage and/or recurrent IVF/intracytoplasmic sperm injection failure. Fluorescence in situ hybridization (FISH) was used to detect numerical abnormalities in sex chromosomes (X, Y) and autosomes (13, 18, 21) in ejaculated sperm. Sperm aneuploidy in men with RPL and normozoospermic controls. Men with RPL had a greater percentage of sperm aneuploidy within the sex chromosomes and chromosomes 18 and 13/21 (1.04% vs. 0.38%; 0.18% vs. 0.03%; 0.26% vs. 0.08%). In total, 40% of men with normal sperm density and motility had abnormal sperm aneuploidy in all the chromosomes analyzed. Men with abnormal sperm density and motility had a higher proportion of sperm sex chromosome aneuploidy than men with normal density/motility (62% vs. 45%). Men with normal strict morphology (>4%) had lower rates of sex chromosome and sperm aneuploidy than men with abnormal strict morphology (28% vs. 57%). There was no association between sperm DNA fragmentation and sperm aneuploidy. Men with RPL have increased sperm aneuploidy compared with controls. A total of 40% of men with RPL and normal sperm density/motility had abnormal sperm aneuploidy. Men with oligoasthenozoospermia and abnormal strict morphology had a greater percentage of sperm aneuploidy compared with men with normal semen parameters.

Chromosomal aneuploidies and DNA fragmentation of human spermatozoa from patients exposed to perfluorinated compounds.

This study investigated chromosomal aneuploidies and DNA damage in spermatozoa from male patients contaminated by perfluorinated compounds (PFCs) in whole blood and seminal plasma. Sperm aneuploidy and diploidy rate for chromosomes 18, X and Y were evaluated by FISH; sperm DNA fragmentation was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling technique coupled to flow cytometry. Our results indicated that PFC contamination was present in 58% of subjects included in the study. A significant increase in alterations of sperm parameters was observed in PFC-positive subjects compared to PFC-negative subjects. As regards the sperm aneuploidy, both disomy and diploidy rates resulted significantly increased in subjects positive for PFC contamination compared to PFC-negative samples. In addition, sperm DNA fragmentation index resulted significantly increased in PFC-contaminated subjects compared to PFC-non-contaminated subjects, with a significant increased level of dimmer DNA fragmentation index. Our results clearly indicate that PFC contamination may detrimentally affect spermatogenesis, disturbing both meiotic segregation and DNA integrity. We could therefore suggest cautions to reduce or eliminate any contact with these compounds because the long-term effects of PFC accumulation in the body are not predictable.

Sperm concentration, hyaluronic acid-binding capacity, aneuploidy and persistent histones in testicular cancer.

Are the quantitative and qualitative characteristics of semen samples of patients with testicular cancer (TC), prior to anticancer therapy, different from infertile oligozoospermic (IO) and normozoospermic (NZ) age-matched men? Sperm concentration in TC patients was significantly decreased with no difference in estimated numerical chromosome aberrations and nuclear decondensation compared with NZ men, while the infertile, oligozoospermic men had significantly poorer sperm qualitative characteristics versus the TC group overall and oligozoospermic patients with TC. Spermatogenesis is altered in TC patients at the time of diagnosis. However, the mechanism responsible for the decreased semen quantity in patients with TC is not well understood. Anticancer treatment may have gonadotoxic side effects and post-treatment fertility cannot be predicted. Before commencing anticancer treatment, cryopreservation may be suggested to preserve fertility but there are no data regarding the risk of genetic aberrations in these sperms. This is a cross-sectional study examining semen from 28 patients with TC, 20 IO and 20 NZ age-matched men attending the Andrology Center and the Sperm Cryopreservation Laboratory of the Medical and Health Science Center, University of Debrecen. Semen samples from patients with TC were collected after orchidectomy, but prior to anticancer treatment. Semen samples from TC patients recruited over a period of 4 years were studied. Based on their sperm concentration, TC patients were subgrouped into an oligozoospermic TC (TCO) and a normozoospermic TC group. For statistical analysis, the normal group (NZ + IO) comprised non-tumorous NZ and IO men. The ejaculates were assessed as per World Health Organization guidelines. Hyaluronic acid (HA)-binding capacity was the functional test. To determine the numerical chromosome aberrations, we used multi-color fluorescence in situ hybridization. Aniline blue (AB) staining was performed as a nuclear decondensation marker test. The results did not reveal any significant difference in disomy of sex chromosomes and chromosome 17, diploidy and estimated numerical chromosome aberrations and AB staining results upon comparing the NZ and TC groups, although the sperm concentration (P < 0.001) and HA-binding capacity (P < 0.001) were lower in the TC group. Estimated numerical chromosome aberrations (P < 0.001), AB staining (P < 0.001) and HA-binding capacity (P = 0.019) were lower in the infertile, oligozoospermic group when compared with the patients with TC. The TCO group had significantly better results in every examined parameter than the infertile, oligozoospermic group. In the non-tumorous control group (NZ + IO), a significant (P < 0.001) correlation (Spearman's rho = r) was found between sperm concentration and aneuploidy rate (r = -0.642), AB staining (r = -0.876) and HA binding (r = 0.842); the HA-binding capacity was related to the aneuploidy rate (r = -0.678) and the AB staining (r = -0.811); and there was significant correlation between aneuploidy and AB staining (r = 0.559). In the TC group, apart from the negative correlation between sperm concentration and estimated chromosomal aberrations (r = -0.642), no other correlations were observed. Data on confounders influencing sperm characteristics, such as smoking, occupational or environmental hazards, alcoholism, co-morbidities and other andrological conditions, were not collected. This is the first study to demonstrate that sperm qualitative characteristics in anticancer therapy naïve oligozoospermic TC patients differ significantly from those in IO men and do not differ from those in NZ men. Our results need to be validated in similar groups of men and in other patient groups with cancer where cryopreservation is advisable. This research was supported by the European Union and the State of Hungary, co-financed by the European Social Fund in the framework of TÁMOP-4.2.4.A/2-11/1-2012-0001 'National Excellence Program'. The authors have no conflict of interest to declare.

Aneuploidy frequency in spermatozoa of Egyptian men with normal and abnormal semen parameters using fluorescencein situhybridisation

Chromosome anomalies were suggested to be more frequent in infertile males so our case-control study aimed at evaluating the incidence of spermatic aneuploidies in forty males with severe oligoasthenoteratozoospermia (OAT) and comparing it with that in another forty males having normal semen parameters. Semen samples were collected and analysed in the Clinical Pathology Department according to criteria of the World Health Organization (WHO laboratory manual for the examination and processing of human semen, 2010, WHO Press). Fluorescence in situ hybridisation (FISH) was performed on decondensed spermatozoa from fresh semen ejaculates, using dual coloured chromosome-specific DNA probes labelled with fluorochromes to study sperm aneuploidies in chromosomes 13, 21, X and Y. There was no statistical significant difference between cases and controls regarding disomy frequencies for chromosomes 13, 21 or both combined. However, 13, 21 diploidy frequency was significantly higher among OAT cases. Regarding chromosomes X and Y, both cases and controls showed similar results for disomy/diploidy frequency for both chromosomes; however, there was a statistical significant increase in YY disomy/diploidy frequency among OAT patients. X chromosome-bearing spermatozoa were found to be significantly higher among controls. Patients with severe OAT have a higher total sperm aneuploidy rate, regarding chromosomes 13, 21, X and Y but without a statistical significant difference.

FISH and tips: a large scale analysis of automated versus manual scoring for sperm aneuploidy detection.

BackgroundApproximately 1% of the spermatozoa found in ejaculate of healthy men are aneuploid and this rate increases in the population of subfertile and infertile men. Moreover, fertilization with these aneuploid sperm can lead to impaired embryo development. Fluorescent In Situ Hybridization (FISH) is the common cytogenetic tool used for aneuploidy screening on sperm. However, it is a time-consuming technique and cytogenetic or in vitro fertilization laboratories cannot routinely use it and face the increasing demand of such analyses before Assisted Reproductive Techniques (ART). As automation can be a clue for routine practice, this study compares manual and automated scoring of sperm aneuploidy rates using a Metafer Metasystems® device. The results obtained also contribute to global data about FISH on sperm cells.MethodsWe recruited 100 men addressed for sperm cryopreservation. They all signed an informed consent to participate in the study. 29 men were donors or consulted before vasectomy (control group) and 71 were suffering of Hodgkin’s disease or non Hodgkin lymphoma (patient group). One semen sample was collected for each patient, analyzed according to WHO criteria and prepared for a triple-color FISH using centromeric probes for chromosomes 18, X and Y. Automated scoring was performed using a Metafer Metasystems® device.Results507,019 cells were scored. We found a strong concordance between the automated and the manual reading (d < 0.01 in Bland-Altman test). We also did not find a statistically significant difference between the automated and the manual reading using Wilcoxon test for total aneuploidy rate (p = 0.06), sex chromosomes disomy (p = 0.33), chromosome 18 disomy (p = 0.39) and diploidy (p = 0.21). Cumulative rate of total aneuploidy was 0.78% ± 0.212% for patient group and 0.54% ± 0.15 for control group and among this, sex chromosome XY disomy rate was of 0.54% for patient group and 0.27% for control group.ConclusionThis study validates the automated reading for FISH on sperm with a Metafer Metasystems® device and allows its use in a laboratory routine.