Chromosome abnormalities in spermatozoa of patients with azoospermia and normal somatic karyotype

Since epididymal and testicular spermatozoa of azoospermic patients are frequently used for intracytoplasmic sperm injection (ICSI), many studies have been carried out to evaluate their karyotype. This article will review all published data on this topic. In most of the studies, spermatozoa have been retrieved from the testis or the epididymis of patients with nonobstructive (NOA) or obstructive (OA) azoospermia, respectively. Sperm aneuploidy has been evaluated by fluorescence in situ hybridization using probes for sex chromosomes and an array of autosomes. A significantly higher sperm aneuploidy rate has been reported in patients with NOA and OA compared to ejaculated spermatozoa, mainly for sex chromosomes. The magnitude of the increase varies between studies, probably because of the heterogeneity of case selection as well as of the methodology employed. The majority of the studies reported that patients with NOA have a greater sperm aneuploidy rate compared to OA. The greater frequency of sperm aneuploidy in azoospermic patients increases the risk of transmitting a karyotype abnormality to the offspring generated by ICSI.     

Syndrome de klinefelter et conseil génétique

Klinefelter’s syndrome is a common sex chromosomal aberration generally characterized by hypergonadotrophic hypogonadism and azoospermia. However, spermatogenesis impairment is variable and severe oligozoospermia can be found in some men, particularly those exhibiting a mosaic karyotype 47,XXY/ 46,XY. New reproductive technologies, such as intracytoplasmic sperm injection (ICSI), allow Klinefelter patients to have a progeny, even those who are azoospermic after testicular sperm recovery. The question therefore arises of whether or not there is a genetic risk for pregnancies from affected fathers. Sperm karyotyping, by in vitro penetration of zona-free hamster eggs or by fluorescence in-situ hybridization (FISH), is a method of choice for measuring aneuploidy rate in spermatozoa of patients carrying gonosomal abnormalities. A theoretical model would predict a high level of 24,XX and/or 24,XY disomic sperm cells in Klinefelter patients if 47,XXY spermatogonia were able to complete meiosis and achieve spermatogenesis. Interestingly, current observations show that the rate of abnormal spermatozoa in these patients is low, around 1–2%, which indicates that only 46,XY spermatogonia can produce mature sperm cells and that oligozoospermic Klinefelter patients probably carry a 47,XXY / 46,XY mosaicism, at least at the testicular level. However, this low but statistically significant level of disomic spermatozoa emphasizes the fact that their spermatogenesis occurs in a compromised environment which could increase the risk of meiotic errors. Therefore, the possible occurrence of autosomal aneuploidies in children born from Klinefelter fathers leads to the following recommendations: a) individual analysis by FISH of the sperm aneuploidy rate in each Klinefelter patient candidate for ICSI; b) proposal of fetal karyotyping after amniocentesis in pregnancies obtained by this technique.     

Is there a relationship between sperm chromosome abnormalities and sperm morphology?

This review explores the relationship between sperm chromosomal constitution and morphology. With the advent of techniques for obtaining information on the chromosome complements of spermatozoa, this relationship has been studied in fertile men and in men with a high frequency of chromosomal abnormalities. Using human sperm karyotype analysis, no relationship between sperm chromosome abnormalities and morphology was found in fertile men, translocation carriers or post-radiotherapy cancer patients. Fluorescence in situ hybridization (FISH) analysis has not generally revealed a specific association between morphologically abnormal sperm and sperm chromosome abnormalities, but has indicated that teratozoospermia, like other forms of abnormal semen profiles (aesthenozoospermia, oligozoospermia) is associated with a modest increase in the frequency of sperm chromosome abnormalities. However, FISH studies on some infertile men and mouse strains have suggested that certain types of morphologically abnormal spermatozoa, such as macrocephalic multitailed spermatozoa, are associated with a very significantly increased frequency of aneuploidy. Thus, there may be an association between sperm morphology and aneuploidy in infertile men with specific abnormalities.     

Experimental mild increase in testicular temperature has drastic,but reversible,effect on sperm aneuploidy in men: A pilot study

In mammals testicular and epididymal temperature increase impairs spermatogenesis. This experimental study investigates the effects of a mild testis temperature increase (i.e. testis temperature remains below core body temperature) on sperm aneuploidy in men. In 5 fertile volunteers a testicular temperature increase was induced by maintaining the testes at suprascrotal position using specially designed underwear for 15 ± 1 h daily for 120 consecutive days. After heating men were followed for next 180 days. A control group (27 men) was recruited. Semen samples were collected before, during and after heating period and analyzed for chromosomes X, Y and 18 for aneuploidy using FISH. A total of 234,038 spermatozoa were studied by FISH. At day 34 of heating, mean sperm aneuploidy values were not modified. From day 34 of heating until day 45 post heating, FISH evaluation was not possible due to the drastic fall of sperm count. At day 45 post-heating total sperm aneuploidy percentage was twice higher than before heating whereas. Sex disomy (sperm XY18), sex chromosome nullisomy (sperm 18) were significantly higher than controls. These effects were completely reversed at 180 days post heat exposure. Conclusion: A mild rise in testicular temperature significantly increases sperm aneuploidies, reflecting an effect on the meiosis stage of spermatogenesis. The effect of heating was reversible and suggests that recovery of aneuploidy to normal values requires at least two cycles of spermatogenesis. Nonetheless, the low number of volunteers was a limitation of this pilot study and warrants further research on larger population.     

Chromosome abnormalities in sperm from infertile men with normal somatic karyotypes: teratozoospermia

Teratozoospermia is characterized by the presence of spermatozoa with abnormal morphology in sperm. This condition is frequently associated with infertility and intracytoplasmic sperm injection (ICSI) is frequently used as the treatment of choice. However, the use of ICSI has created consequential debate concerning the genetic risk for the offspring. Fluorescence in situ hybridization technique (FISH), allowing the specific identification of human chromosomes in sperm nuclei, has been used to study chromosome abnormalities in sperm from men with teratozoospermia and a normal karyotype. In this review, we present studies that have tried to determine if men with a normal blood karyotype but suffering from teratozoospermia present a higher aneuploidy frequency. The literature is limited to three forms of teratozoospermia. The first group consists of "polymorphic teratozoospermia", where a majority of spermatozoa display more than one type of abnormality. In this case, only a slight increase in aneuploidy frequency is observed, which cannot be differentiated from the results observed in oligo-astheno-teratozoospermia (OAT). The second group, named "globozoospermia", is characterized by round spermatic heads, absence of acrosome and disorganization of mid-piece and tail. In this case, some studies have shown a significant, but moderate, increase in the aneuploidy frequency for acrocentrics and sex chromosomes. The aneuploidy frequency remains low, also ICSI can be proposed to these patients, but few successes occur. The third group consists of "enlarged head teratozoospermia", where almost all spermatozoa have an enlarged head, multiple tail and abnormal acrosome. In this case a very high level of missegregation is observed, leading to nearly 100% aneuploidy. In this particular group, ICSI must be refuted, and patients have to be redirected to other possibilities, like sperm donation.     

Germ-cell nondisjunction in testes biopsies of men with idiopathic infertility.

Intracytoplasmic sperm injection (ICSI) has been used in combination with testicular sperm extraction to achieve pregnancies in couples with severe male-factor infertility, yet many of the underlying genetic mechanisms remain largely unknown. To investigate nondisjunction in mitotic and meiotic germ cells, we performed three-color FISH to detect numeric chromosome aberrations in testicular tissue samples from infertile men confirmed to have impaired spermatogenesis of unknown cause. FISH was employed to determine the rate of sex-chromosome aneuploidy in germ cells. Nuclei were distinguished as haploid or diploid, respectively. The overall incidence of sex-chromosome aneuploidy in germ cells was found to be significantly higher (P<.00001) in all three abnormal histopathologic patterns (range 39.0%-43.5%) as compared with normal controls (29.1%). The relative ratio of normal to aneuploid nuclei in the diploid cells of patients with impaired spermatogenesis was approximately 1.0, a >300% decrease when compared with the 4.42 ratio detected in patients with normal spermatogenesis. These results provide direct evidence of an increased incidence of sex-chromosome aneuploidy observed in germ cells of men with severely impaired spermatogenesis who might be candidates for ICSI with sperm obtained directly from the testis. The incidence of aneuploidy was significantly greater among the diploid nuclei, which suggests that chromosome instability is a result of altered genetic control during mitotic cell division and proliferation during spermatogenesis.     

Fertilité et aneuploïdies spermatiques après traitement par radiothérapie et/ou chimiothérapie pour cancer du testicule ou lymphome

Improvements in cancer therapy have considerably modified patient survival rates over recent years. However, the side effects of these treatments especially the effects on fertility, must be taken into account. Anticancer therapy can transiently inhibit spermatogenesis. Factors such as pretreatment semen parameters and the type of chemotherapy or radiotherapy may influence recovery of spermatogenesis, but it is still impossible to predict the probability of and time to recovery for each patient. Sperm banking remains the only way to prevent the effects of cancer treatment on male fertility. Another possible effect of chemotherapy or radiotherapy is genetic damage to germ cells. For instance, chromosomal abnormalities in viable sperm produced by these patients after recovery of spermatogenesis may result in fetal death or congenital abnormalities in their offspring. It has been fairly well documented that, during the first three months after treatment, DNA breaks and abnormal chromosomal segregation induced by chemotherapy/radiotherapy lead to structural and numerical chromosomal abnormalities in spermatozoa, respectively. However, the long-term effects on genetic sperm content have not been clearly established. The results of published studies are contradictory and are based on limited numbers of patients (maximum of 6). We present the preliminary results of a retrospective study concerning patients treated for testicular cancer or lymphoma between 1995 and 2000. Fluorescence in situ hybridization (FISH) analysis of chromosomes X, Y and 18 was performed on sperm collected one to five years after treatment and compared to the data obtained for non-affected fertile men. For four out of 13 patients, we found a significantly increased frequency of aneuploidy rates (mainly XY disomy and diploidy), and these results did not appear to be correlated with sperm count, sperm morphology or post-treatment duration. In conclusion, increased sperm aneuploidy rates appear to only concern a small number of patients, to varying degrees and without any predictive factors. According to published data and our preliminary results, we recommend waiting at least two years before starting ART (Assisted Reproduction Therapy) for patients treated for testicular cancer or lymphoma. Moreover, FISH analysis could be helpful to choose between ART with post-treatment sperm or cryopreserved sperm.     

Increased Frequency of Aneuploidy in Long-Lived Spermatozoa

Aneuploidy commonly causes spontaneous abortions, stillbirths, and aneuploid births in humans. Notably, the majority of sex chromosome aneuploidies in live births have a paternal origin. An increased frequency of aneuploidy is also associated with male infertility. However, the dynamics and behavior of aneuploid spermatozoa during fertilization in humans have not been studied in detail. Therefore, we compared the frequency of aneuploidy and euploidy in live spermatozoa from normozoospermic men over a 3-day period. To assess the dynamics and behavior of aneuploid spermatozoa, we simultaneously evaluated sperm viability using the hypo-osmotic swelling test and sperm aneuploidy using fluorescence in situ hybridization. Whereas the frequency of viable euploid spermatozoa significantly decreased over 3 days, the frequency of viable spermatozoa with aneuploidy interestingly showed a time-dependent increase. In addition, spermatozoa with abnormal sex chromosomes survived longer. To compared with spermatozoa with other swelling patterns, those with tail-tip swelling patterns had a lower frequency of aneuploidy at all time points. This study revealed the novel finding that the frequency of aneuploid spermatozoa with fertilization capability significantly increased compared to that of euploid spermatozoa over 3 days, suggesting that aneuploid spermatozoa can survive longer than euploid spermatozoa and have a greater chance of fertilizing oocytes.     

Chromosome abnormalities in sperm from infertile men with normal somatic karyotypes: oligozoospermia

Recently, intracytoplasmic sperm injection (ICSI) has been extremely successful for the treatment of male infertility. However, transmission of cytogenetic defects to offspring is a great concern. There are two types of cytogenetic problems in patients seeking ICSI; one is the transmission of genetic defects from patients with constitutional chromosomal abnormalities and the second is the generation of de novo defects in infertile men. Generally about 5.1% of infertile men have chromosomal abnormalities. Among such infertile men, men with severe spermatogenesis defects, including oligozoospermia and azoospermia, are subjects for ICSI. Therefore it is very important to obtain cytogenetic information in these infertile patients. Furthermore, oligozoospermic men with a normal somatic karyotype also have increased frequencies of sperm chromosome abnormalities. Oligozoospermia is usually associated with other sperm alterations, for example oligoasthenozoospermia, oligoteratozoospemia and oligoasthenoteratozoospermia. In this review, the relationship between sperm concentration and sperm aneuploidy frequencies has been analyzed. The inverse correlation between the frequency of sperm aneuploidy and concentration has been reported in extensive studies. Especially in severe oligozoospermia, a significantly higher frequency of sex chromosome aneuploidy has been observed and this has been corroborated in recent clinical outcome data of ICSI.     

Génotoxicité des chimiothérapies et radiothérapies: Quelles sont les conséquences pour le spermatozoïde humain?

The prognosis of cancer in young men of childbearing potential has been considerably improved over recent decades as a result of therapeutic progress. Chemotherapy and radiotherapy have well known effects on spermatogenesis. Apart from quantitative and qualitative impairment of spermatogenesis, animal studies have also demonstrated nuclear lesions (aneuploidy, presence of adducts, DNA fragmentation, etc.) and sometimes lesions affecting the F1 and F2 generations. Chromosomal studies of human spermatozoa after radiotherapy have demonstrated an increased frequency of chromosomal anomalies. The first studies concerning the effects of chemotherapy used the heterospecific fertilization technique to demonstrate spermatozoal chromosomal anomalies. More recently, thefluorescence in situ hybridization (FISH) technique has been used to study several chromosomes on a large number of spermatozoa. The results of various studies based on small sample sizes vary as a function of the therapeutic protocol administered and the time of sperm collection in relation to the end of treatment. We studied 5 patients who provided a semen sample 6 to 17 months after completing the BOE chemotherapy protocol (Bleomycin, Etoposide, Cisplatin). We demonstrated an increased rate of aneuploid and diploid spermatozoa. The results of our study and those reported by R. Martin et al. [45, 47] suggest the possibility of a transient effect of chemotherapy on gamete chromosomes. Other studies, conducted in the context of Hodgkin’s disease, have demonstrated the transient nature of the aneuploidy effect. Apart from the harmful action on chromosomes, treatments could also damage spermatozoal DNA. Studies conducted on larger sample sizes and using other methods of analysis therefore appear to be essential. In the meantime, it appears preferable to systematically propose semen cryopreservation before treatment and to provide very cautious advice to patients desiring a pregnancy soon after completion of treatment.     

Relationship between clinical phenotype, semen parameters and aneuploidy frequency in sperm nuclei of 50 infertile males

The purpose of this study was to analyse the frequency of disomy for chromosomes 1, 13, 14, 18, 21, 22, X and Y in sperm nuclei of 50 infertile men and 10 healthy probands of proven fertility. Semen parameters (sperm count, global motility and morphology), urological clinical examination, genital ultrasound and lymphocyte karyotyping were performed for each patient. Disomy frequency was established by fluorescence in situ hybridization by using whole chromosome paint probes. The mean rate of disomy for the various autosomes studied was higher in infertile males than in subjects of proven fertility. Interchromosomal and interindividual differences in the disomy frequency were observed between the 50 patients. The mean frequency of homodisomy YY and heterodisomy XY was increased in spermatozoa of patients with low semen quality parameters (0.24% and 0.54%, respectively). The disomy frequency in infertile males was directly correlated with the severity of oligospermia. However, no relationship was established between aneuploidy rate, sperm motility, morphology or clinical phenotype. These results support the hypothesis that, during spermatogenesis of males with sperm parameter alterations, a decreased frequency of meiotic chromosome pairing and crossing over may lead to spermatogenesis arrest at the meiosis stage and/or to an increase of meiotic nondisjunctions. Meiotic arrest in some germ cells may be responsible for oligospermia and nondisjunctions in other cells for aneuploidy in mature male gametes.     

Studies on the influence of aneuploidy of spermatozoa obtained from patients with oligozoospermia on their structure

The presence of aneuploidy in spermatozoa influences their biological characteristics, especially their ability to fertilise the ovum. The aim of the present study was to investigate if aneuploidy is accompanied by any changes in the morphology of spermatozoa in oligozoospermic patients. For this purpose, the percentage of aneuploid cells in sperm and the correlation between the specific morphological forms of spermatozoa and aneuploidy were evaluated. The study proved a negative correlation between DNA content of aneuploid and normal spermatozoa. A weak positive correlation was demonstrated between the presence of aneuploid spermatozoa and DNA content of spermatozoa with large heads. No such correlations could be detected for DNA content of the remaining morphological forms of spermatozoa. Thus, men with a lowered number of spermatozoa and/or with abnormal spermatozoal morphology should have their spermatozoal DNA content tested in order to evaluate the degree of aneuploidy, especially in cases where in vitro fertilisation is intended.     

Aneuploidy in human spermatozoa

We reviewed the frequency and distribution of disomy in spermatozoa obtained by multicolor-FISH analysis on decondensed sperm nuclei in (a) healthy men, (b) fathers of aneuploid offspring of paternal origin and (c) individuals with Klinefelter syndrome and XYY males. In series of healthy men, disomy per autosome is approximately 0.1% but may range from 0.03 (chromosome 8) to 0.47 (chromosome 22). The great majority of authors find that chromosome 21 (0.18%) and the sex chromosomes (0.27%) have significantly elevated frequencies of disomy although these findings are not universal. The total disomy in FISH studies is 2.26% and the estimated aneuploidy (2× disomy) is 4.5%, more than double that seen in sperm karyotypes (1.8%). Increased disomy levels of low orders of magnitude have been reported in spermatozoa of some normal men (stable variants) and in men who have fathered children with Down, Turner and Klinefelter syndromes. These findings suggest that men with a moderately elevated aneuploidy rate may be at a higher risk of fathering paternally derived aneuploid pregnancies. Among lifestyle factors, smoking, alcohol and caffeine have been studied extensively but the compounding effects of the 3 are difficult to separate because they are common lifestyle behaviors. Increases in sex chromosome abnormalities, some autosomal disomies, and in the number of diploid spermatozoa are general features in 47,XXY and 47,XYY males. Aneuploidy of the sex chromosomes is more frequent than aneuploidy of any of the autosomes not only in normal control individuals, but also in patients with sex chromosome abnormalities and fathers of paternally derived Klinefelter, Turner and Down syndromes.     

Abnormal Sperm Parameters in Humans Are Indicative of an Abortive Apoptotic Mechanism Linked to the Fas-Mediated Pathway

The life cycle of many cell types can hinge on the presence of death factors that can control programmed cell death. The Fas-mediated apoptotic pathway has been implicated in controlling apoptosis during spermatogenesis in a number of mammalian species. In the human, the presence of nuclear DNA damage in ejaculated spermatozoa has pointed to a possible role for apoptosis during spermatogenesis. The presence of other molecular markers of apoptosis has, however, not been shown. More importantly, differences in these markers have not been investigated in men with normal and abnormal sperm parameters. In this study we examine for the presence of the cell surface protein Fas in ejaculated human spermatozoa. Ejaculated spermatozoa (55 samples) were labeled with anti-human Fas antibody and the number of spermatozoa displaying Fas were counted using a fluorescence-activated cell sorter (FACS). In 30/31 (96.8%) normal males (>20 million sperm per milliliter), less than 10% of the spermatozoa were Fas positive. In contrast, 14/24 (58.3%) oligozoospermic samples (<20 million sperm per milliliter) contained more than 10% Fas-positive spermatozoa. Similar differences were observed in men whose spermatozoa had poor motility and morphology. These results indicate that apoptosis is a major mechanism in regulating spermatogenesis in the human and that there are clear differences in molecular markers of apoptosis between males with normal and abnormal sperm parameters. We propose that the presence of Fas-labeled spermatozoa in the ejaculate of these men is indicative of an "abortive apoptosis" having taken place, whereby the normal apoptotic mechanisms have misfunctioned, have been overridden, or have not been completed.     

Aneuploidy and DNA fragmentation in sperm of carriers of a constitutional chromosomal abnormality

Among various causes responsible for infertility, it has been admitted for a long time that male infertility can be due to impaired spermatogenesis and/or balanced structural chromosomal abnormalities. Sperm DNA fragmentation is also considered as another cause of infertility. Most of the studies on male infertility have concerned either aneuploidy in the sperm of carriers of constitutional chromosomal abnormalities or sperm DNA fragmentation. This review is aimed at analyzing these 2 parameters in the same patients. Furthermore, we present work on the study of these 2 parameters in the same gametes of 4 carriers of a balanced chromosomal abnormality. Meiotic segregation was analyzed by fluorescent in situ hybridization and DNA fragmentation was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay. It was shown that aneuploidy and DNA fragmentation were increased in the sperm of carriers of a balanced chromosomal abnormality. For all 4 carriers of a balanced structural abnormality, there was a 2-5 times higher proportion of spermatozoa with unbalanced chromosomal content and fragmented DNA than among those with normal/balanced content. Moreover, we found a non-random distribution with more gametes with DNA fragmentation when these arose from a particular segregation mode. The mechanism which would tend to explain our results is abortive apoptosis. In conclusion, both meiotic segregation and DNA fragmentation studies should be integrated in the genetic exploration of male carriers of a chromosomal structural abnormality.     

Detection of sex chromosome aneuploidy in dog spermatozoa by triple color fluorescence in situ hybridization

With the development of a direct visualization of sex chromosome in a single sperm by fluorescence in situ hybridization (FISH) technique, the frequency of aberration (aneuploidy) in spermatozoa in several mammals has been investigated. However, there is no report in the incidence of X-Y aneuploidy in the sperm population of dogs. Therefore, in this study, the aneuploidy in dog spermatozoa was examined by multicolor FISH using specific molecular probes for canine sex chromosomes and autosome. Semen from eight male Labrador retrievers was used as specimen. For decondensation of sperm nuclei, the specimen was treated with 1 M NaOH for 4 minutes at room temperature. Probes for chromosomes X, Y, and 1, labeled with SpectrumGreen, Cy3 and Cy5, respectively, were hybridized with decondensed spermatozoa. Fluorescence in situ hybridization signals in sperm heads were clearly detected in each specimen, regardless of the sperm donor. The FISH signal of at least one of the three probes was detected in all sperm heads examined. There was no significant difference between the theoretical ratio (50:50) and the observed ratio of X and Y chromosomes in spermatozoa of all the eight dogs. Mean percentage of sex chromosome aneuploidy was 0.127% (ranged between 0% and 0.316%). This percentage of canine sex chromosome aneuploidy was lower than the one reported in cattle, horses, river buffalo, and goats sperm, but higher than that observed in mice and sheep.     

Exposure to acrylonitrile induced DNA strand breakage and sex chromosome aneuploidy in human spermatozoa

To explore acrylonitrile (ACN)-induced DNA strand breakage and sex chromosome aneuploidy in human spermatozoa, semen parameters were examined among 30 acrylonitrile-exposed workers according to WHO laboratory manual for the examination of human sperm. DNA strand breakage of sperm cells was investigated among 30 ACN-exposed workers using single cell gel electrophoresis (SCGE). The frequency of sex chromosome aneuploidy in sperm cells was analyzed among nine ACN-exposed workers using fluorescence in situ hybridization (FISH). The geometrical mean of sperm density was 75 x 10(6)ml(-1) in exposure group, significantly lower than 140 x 10(6)ml(-1) in the control. The geometrical mean of sperm number per ejaculum was 205 x 10(6) in exposure group, significantly lower than 280 x 10(6) in the control. The rates of comet sperm nuclei were 28.7% in exposure group, significantly higher than 15.0% in the control. Mean tail length was 9.8 microm in exposure group, longer than 4.3 microm in the control. The frequency of sex chromosome disomy was 0.69% in exposure group, significantly higher than 0.35% in the control. XY-bearing sperm was the most common sex chromosome disomy, with an average rate of 0.37% in exposure group, and 0.20% in the control. XX- and YY-bearing sperm accounted for an additional 0.09 and 0.23% in exposure group, and 0.05 and 0.10% in the control. The results indicate that ACN affect semen quality among ACN-exposed workers. ACN or its metabolites could induce reproductive defects as an in vivo multipotent genotoxic agent by inducing DNA strand breakage and sex chromosome non-disjunction in spermatogenesis.     

Sperm of Doradidae (Teleostei: Siluriformes)

Spermatic characteristics were studied in 10 species representing several distinct groups within the catfish family Doradidae. Interestingly, different types of spermatogenesis, spermiogenesis and spermatozoa are correlated with intrafamilial groups previously proposed for Doradidae. Semi-cystic spermatogenesis, modified Type III spermiogenesis, and biflagellate sperm appear to be unique within Doradidae to the subfamily Astrodoradinae. Other doradid species have sperm with a single flagellum, cystic spermatogenesis, and spermiogenesis of Type I (Pterodoras granulosus, Rhinodoras dorbignyi), Type I modified (Oxydoras kneri), or Type III (Trachydoras paraguayensis). Doradids have an external mode of fertilization, and share a few spermatic characteristics, such as cystic spermatogenesis, Type I spermiogenesis and uniflagellate sperm, with its sister group Auchenipteridae, a family exhibiting sperm modifications associated with insemination and internal fertilization. Semi-cystic spermatogenesis and biflagellate spermatozoa are also found in Aspredinidae, and corroborate recent proposals that Aspredinidae and Doradoidea (Doradidae + Auchenipteridae) are sister groups and that Astrodoradinae occupies a basal position within Doradidae. The co-occurrence in various catfish families of semi-cystic spermatogenesis and either biflagellate spermatozoa (Aspredinidae, Cetopsidae, Doradidae, Malapturidae, Nematogenyidae) or uniflagellate sperm with two axonemes (Ariidae) reinforces the suggestion that such characteristics are correlated. Semi-cystic spermatogenesis and biflagellate sperm may represent ancestral conditions for Loricarioidei and Siluroidei of Siluriformes as they occur in putatively basal members of each suborder, Nematogenyidae and Cetopsidae, respectively. However, if semi-cystic spermatogenesis and biflagellate sperm are ancestral for Siluriformes, cystic spermatogenesis and uniflagellate sperm have arisen independently in multiple lineages including Diplomystidae, sister group to Siluroidei.     

Estimation du taux d’aneuploïdies des spermatozoïdes épididymaires prélevés en vue d’ICSI chez des hommes à caryotype normal

Over the last ten years, fluorescent in situ hybridization in decondensed sperm nuclei has been used to study the chromosomal constitution of human spermatozoa. Studies have estimated that the disomy rate per chromosomal pairs is between 0.15% and 0.3%. The aim of this study was to evaluate the aneuploidy rate of human epididymal spermatozoa extracted from five men with obstructive azoospermia undergoing IVF. Genetic studies (karyotypes, Y micodeletion syndrome and mutation of the CFTR gene) did not reveal any abnormality. Disomy frequencies were determined by X-Y-8 multicolour fluorescence in situ hybridisation on 18,013 epididymal spermatozoa and 20,000 spermatozoa from healthy donors (control group). No significant difference was found between epididymal and ejaculated samples. However, isolated non-significant differences were observed between one of the patients and the control group. In conclusion, the present findings suggests that there is no increased risk for de novo chromosomal aberrations after IVF therapy with epididymal spermatozoa of men with obstructive azoospermia.     

Sperm aneuploidy and DNA fragmentation in unexplained recurrent pregnancy loss: a multicenter case-control study

Background

Recurrent pregnancy loss (RPL) is defined as the loss of at least three pregnancies in the first trimester. Although the most common cause is embryo aneuploidy, and despite female checkup and couple karyotyping, in about 50% of cases RPL remain unexplained. Male implication has little been investigated and results are discordant. In this context, we conducted a multi-center prospective case-control study to investigate male gamete implication in unexplained RPL.

Methods

A total of 33 cases and 27 controls were included from three university hospitals. We investigated environmental and family factors with a detailed questionnaire and andrological examination, sperm characteristics, sperm DNA/chromatin status using the sperm chromatin structure assay (SCSA) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and sperm aneuploidy using fluorescence in situ hybridization (FISH). The Mann-Whitney test and the Wilcoxon or Fisher exact tests were used. A non-parametric Spearman correlation was performed in order to analyze the relationship between various sperm parameters and FISH and sperm DNA fragmentation results.

Results

We found significant differences between cases and controls in time to conceive, body mass index (BMI), family history of infertility and living environment. In cases, total sperm motility and the percentage of morphologically normal spermatozoa were significantly decreased. No difference was found between cases and controls in sperm DNA fragmentation or chromatin integrity. In cases, spermatozoa with aneuploidy, hyperhaploidy and chromosome 18 disomy were significantly increased.

Conclusions

This prospective case-control study is one of the largest to examine environmental factors, sperm characteristics, sperm DNA fragmentation and chromatin, and chromosome anomalies in spermatozoa in relation to unexplained recurrent pregnancy loss. The originality of our study lies in the comprehensive andrological examination and search for risk factors and fertility history. Further studies are needed to confirm the links between unexplained RPL and a male family history of infertility or miscarriages. The increased sperm aneuploidy observed in unexplained RPL supports a male etiology. These data pave the way for further studies to demonstrate the value of preimplantation genetic screening in men with increased sperm aneuploidy whose partners experience unexplained RPL.