Large-headed Spermatozoa Research Articles

Sperm characteristics in normal and abnormal ejaculates are differently influenced by the length of ejaculatory abstinence.

No association between the length of ejaculatory abstinence (LEA) and semen characteristics has been confirmed. A short LEA has been linked to improved sperm characteristics and a higher pregnancy rate, but its negative influence on sperm chromatin maturity and longevity may adversely affect reproductive outcomes. We sought to determine the influence of LEA on (i) semen parameters in normozoospermic and abnormal ejaculates; and (ii) the outcomes of sperm-preparation methods in a large number of subfertile men undergoing infertility workups. This retrospective registry-based cohort study analyzed the data of 10,674 ejaculates from 7972 subfertile men, who were then segregated into normozoospermic, oligozoospermic, asthenozoospermic, and oligo-asthenozoospermic cohorts. Variations in semen characteristics and post-wash outcomes were studied between four LEA intervals across 0-15 days. An age-adjusted analysis of covariance (ANCOVA) model linked significant increases in ejaculate volume, sperm concentration (except in the oligozoospermic cohort), and total sperm count to an increased LEA (p<0.05). LEA was negatively associated with motility (except in the asthenozoospermic cohort) and vitality (p<0.05). Large-headed spermatozoa were less common with an increased LEA only in the oligo-asthenozoospermic cohort (p<0.05). In the normozoospermic cohort, a longer LEA led to fewer spermatozoa with amorphous heads but more spermatozoa with tapered heads and cytoplasmic droplets (p<0.05). LEA extension resulted in greater sperm DNA fragmentation in the abnormal cohort (p<0.01). The post-wash sperm concentration and total motile sperm count were significantly improved with a longer LEA in the normozoospermic cohort (p<0.05). Considering the findings in this study and existing literature, a generalized recommendation for long LEA has no clinical value. The LEA should be individualized based on the ejaculate profile and the need for specific clinical intervention.

Two frequent loss-of-function mutations in Aurora Kinase C gene in Algerian infertile men with macrozoospermia.

Macrozoospermia is associated with severe male infertility. To date, the only gene implicated in this phenotype is the Aurora Kinase C gene. We report in this work the genetic screening of AURKC mutations in 34 patients with macrozoospermia among 3,536 Algerian infertile men. Nineteen patients (56%) were homozygotes for the c.144delC mutation, eight (23.52%) homozygotes for the c.744C>G (p.Y248*) mutation and two (5.88%) compound heterozygotes. No AURKC mutation was identified in five patients (14.7%). Interestingly and although it is generally accepted that nearly all positive mutated AURKC patients have close to 100% large-head spermatozoa, our results showed that 11 patients with AURKC mutations (32.35%) had large-headed spermatozoa lower than 70% (7 with c.144delC and 4 with p.Y248*), and no mutation was found in 2 patients who had >70% of macrocephalic spermatozoa. Twenty ICSI attempts were performed before genetic screening resulting in 39 embryos but no pregnancy was obtained. The sequencing of AURKC exons 3 and 6 is appropriate as a first-line genetic exploration in these patients to avoid unsuccessful ICSI attempts. A percentage of large head spermatozoa beyond 25% and a percentage of multiflagellar spermatozoa beyond 10% are predictive of a positive mutation diagnosis.

Is intracouple assisted reproductive technology an option for men with large-headed spermatozoa? A literature review and a decision guide proposal.

Although the presence of spermatozoa with an abnormally large head is rare, it is associated with low fertility or even total infertility. We reviewed the literature on assisted reproductive technology (ART) strategies and outcomes for men with large-headed spermatozoa. We also discuss additional analyses that can usefully characterize sperm defects and help with the choice between intra-couple ART and insemination with donor sperm. Lastly, we propose a classification for cases of large-headed spermatozoa.

Teratozoospermia: spotlight on the main genetic actors in the human

Male infertility affects >20 million men worldwide and represents a major health concern. Although multifactorial, male infertility has a strong genetic basis which has so far not been extensively studied. Recent studies of consanguineous families and of small cohorts of phenotypically homogeneous patients have however allowed the identification of a number of autosomal recessive causes of teratozoospermia. Homozygous mutations of aurora kinase C (AURKC) were first described to be responsible for most cases of macrozoospermia. Other genes defects have later been identified in spermatogenesis associated 16 (SPATA16) and dpy-19-like 2 (DPY19L2) in patients with globozoospermia and more recently in dynein, axonemal, heavy chain 1 (DNAH1) in a heterogeneous group of patients presenting with flagellar abnormalities previously described as dysplasia of the fibrous sheath or short/stump tail syndromes, which we propose to call multiple morphological abnormalities of the flagella (MMAF). A comprehensive review of the scientific literature available in PubMed/Medline was conducted for studies on human genetics, experimental models and physiopathology related to teratozoospermia in particular globozoospermia, large headed spermatozoa and flagellar abnormalities. The search included all articles with an English abstract available online before September 2014. Molecular studies of numerous unrelated patients with globozoospermia and large-headed spermatozoa confirmed that mutations in DPY19L2 and AURKC are mainly responsible for their respective pathological phenotype. In globozoospermia, the deletion of the totality of the DPY19L2 gene represents ∼ 81% of the pathological alleles but point mutations affecting the protein function have also been described. In macrozoospermia only two recurrent mutations were identified in AURKC, accounting for almost all the pathological alleles, raising the possibility of a putative positive selection of heterozygous individuals. The recent identification of DNAH1 mutations in a proportion of patients with MMAF is promising but emphasizes that this phenotype is genetically heterogeneous. Moreover, the identification of mutations in a dynein strengthens the emerging point of view that MMAF may be a phenotypic variation of the classical forms of primary ciliary dyskinesia. Based on data from human and animal models, the MMAF phenotype seems to be favored by defects directly or indirectly affecting the central pair of axonemal microtubules of the sperm flagella. The studies described here provide valuable information regarding the genetic and molecular defects causing infertility, to improve our understanding of the physiopathology of teratozoospermia while giving a detailed characterization of specific features of spermatogenesis. Furthermore, these findings have a significant influence on the diagnostic strategy for teratozoospermic patients allowing the clinician to provide the patient with informed genetic counseling, to adopt the best course of treatment and to develop personalized medicine directly targeting the defective gene products.

Comprehensive investigation in patients affected by sperm macrocephaly and globozoospermia.

The aim of this study was to provide a comprehensive genetic/phenotypic characterization of subjects suffering infertility owing to sperm macrocephaly (n = 3) or globozoospermia (n = 9) and to investigate whether the patients' genetic status was correlated with the alteration of various sperm parameters. AURKC was sequenced in case of sperm macrocephaly while the DPY19L2 status has been analyzed by multiple approaches including a novel qPCR-based copy number assay in case of globozoospermia. Globozoospermic patients were also analyzed for SPACA1, a novel candidate gene herein tested for the first time in humans. The effect of the patients' genetic status was interrogated by implementing the molecular screening with the characterization of several sperm parameters: (i) routine sperm analysis, integrated with transmission electron microscopy; (ii) sperm fluorescent in situ hybridization (FISH) analysis; (iii) sperm DNA fragmentation (DF) analysis. Moreover, for the first time, we performed microsatellite instability analysis as a marker of genome instability in men with sperm macrocephaly and globozoospermia. Finally, artificial reproductive technology (ART) history has been reported for those patients who underwent the treatment. Macrocephalic patients had an AURKC mutation and >89% tetraploid, highly fragmented spermatozoa. DPY19L2 was mutated in all patients with >80% globozoospermia: the two homozygous deleted men and the compound heterozygous showed the severest phenotype (90-100%). The newly developed qPCR method was fully validated and has the potential of detecting also yet undiscovered deletions. DPY19L2 status is unlikely related to FISH anomalies and DF, although globozoospermic men showed a higher disomy rate and DF compared with internal reference values. No patient was mutated for SPACA1. Our data support the general agreement on the negative correlation between macro/globozoospermia and conventional intracytoplasmic sperm injection outcomes. Microsatellites were stable in all patients analyzed. The comprehensive picture provided on these severe phenotypes causing infertility is of relevance in the management of patients undergoing ART.

Sperm macrocephaly syndrome in a patient without AURKC mutations and with a history of recurrent miscarriage

This paper reports a case of recurrent miscarriage in a patient affected by a variant phenotype of sperm macrocephaly syndrome (SMS). SMS is usually related to specific sperm characteristics (large head, multiple tail) and homozygous mutations in the aurora kinase C gene (AURKC). However, the present case observed large-headed spermatozoa with no flagellar abnormalities and no mutations detectable by AURKC sequencing. Furthermore, the patient had repeatedly conceived by intracytoplasmic sperm injection, but pregnancy always aborted. This study performed morphological analysis (Papanicolau staining), annexin V/propidium iodide staining, sperm chromatin structure assay (SCSA), fluorescence in-situ hybridization (FISH) and transmission electron microscopy. This study observed large-headed, mono-tailed, mono-centriolar spermatozoa characterized by abnormal chromatin and swollen mitochondria. SCSA revealed a high ratio of late apoptotic cells with fairly intact amount of DNA. The FISH analysis showed 100% disomy rate. As far as is known, this is the first study to include gene sequencing, TEM, cytogenetic analysis and sperm DNA fragmentation in a case of SMS and also to report recurrent miscarriage related to this specific condition. SMS may be associated with important abnormalities of the sperm subcellular structure and with disomy even in the absence of mutations in the AURKC coding sequence.Sperm macrocephaly syndrome (SMS) is a rare condition that affects spermatozoa and is related to infertility. It is characterized by a specific phenotype of large-headed, multi-tailed spermatozoa with an abnormal chromosomal status. A very few pregnancies have been obtained so far in SMS patients by means of IVF procedures. We present a case of SMS that differs from the classical syndrome as we observed large-headed spermatozoa without tail abnormalities. The affected patient had achieved three pregnancies following IVF, but all aborted. We carried out a detailed examination of the patient’s spermatozoa – morphological, cytogenetic, DNA fragmentation and ultrastructural analysis – and we observed that his spermatozoa are characterized by a large head whose texture appears apoptotic, a single tail and a midpiece whose mitochondria appear swollen. The DNA content within the spermatozoa was altered, as well as the chromosomal status, suggesting that some error must have occurred during spermatogenesis. Interestingly, the genetic sequencing of the specific gene usually related to SMS syndrome (AURKC) revealed no mutations in our patient, suggesting that other genes may be involved in determining this syndrome. As far as is known, this is the first study in which spermatozoa of a SMS patient have been observed using morphological analysis, ultrastructural analysis, cytogenetic analysis and sperm DNA fragmentation analysis together. Moreover, it is believed that this is first report of recurrent miscarriage due to this specific syndrome.

Identification of a new recurrent Aurora kinase C mutation in both European and African men with macrozoospermia

Can we identify new sequence variants in the aurora kinase C gene (AURKC) of patients with macrozoospermia and establish a genotype-phenotype correlation? We identified a new non-sense mutation, p.Y248*, that represents 13% of all mutant alleles. There was no difference in the phenotype of individuals carrying this new mutation versus the initially described and main mutation c.144delC. The absence of a functional AURKC gene causes primary infertility in men by blocking the first meiotic division and leading to the production of tetraploid large-headed spermatozoa. We previously demonstrated that most affected men were of North African origin and carried a homozygous truncating mutation (c.144delC). This is a retrospective study carried out on patients consulting for infertility and described as having >5% large-headed spermatozoa. A total of 87 patients are presented here, 43 patients were published previously and 44 are new patients recruited between January 2008 and December 2011. All patients consulted for primary infertility in fertility clinics in France (n = 44), Tunisia (n = 30), Morocco (n = 9) or Algeria (n = 4). Sperm analysis was carried out in the recruiting fertility clinics and all molecular analyses were performed at Grenoble teaching hospital. DNA was extracted from blood or saliva and the seven AURKC exons were sequenced. RT-PCR was carried out on transcripts extracted from leukocytes from one patient homozygous for p.Y248*. Microsatellite analysis was performed on all p.Y248* patients to evaluate the age of this new mutation. We identified a new non-sense mutation, p.Y248*, in 10 unrelated individuals of European (n = 4) and North African origin (n = 6). We show that this new variant represents 13% of all mutant alleles and that the initially described c.144delC variant accounts for almost all of the remaining mutated alleles (85.5%). No mutated transcripts could be detected by RT-PCR suggesting a specific degradation of the mutant transcripts by non-sense mediated mRNA decay. A rare variant located in the 3' untranslated region was found to strictly co-segregate with p.Y248*, demonstrating a founding effect. Microsatellite analysis confirmed this linkage and allowed us to estimate a mutational age of between 925 and 1325 years, predating the c.144delC variant predicted by the same method to have arisen 250-650 years ago. Patients with no identified AURKC mutation (n = 15) have significantly improved parameters in terms of vitality and concentration of normal spermatozoa, and a decreased rate of spermatozoa with a large head and multiple flagella (P < 0.001). Despite adherence to the World Health Organization guidelines, large variations in most characteristic sperm parameters were observed, even for patients with the same homozygous mutation. We believe that is mainly related to inter-laboratory variability in sperm parameter scoring. This prevented us from establishing clear-cut values to indicate a need for molecular analysis of patients with macrozoospermia. This study confirms yet again the importance of AURKC mutations in the aetiology of macrozoospermia. Although a large majority of patients are of North African origin, we have now identified European patients carrying a new non-sense mutation indicating that a diagnosis of absence of a functional AURKC gene should not be ruled out for non-Magrebian individuals. Indirect evidence indicates that AURKC might be playing a role in the meiotic spindle assembly checkpoint (SAC) during meiosis. We postulate that heterozygous men might have a more relaxed SAC leading to a more abundant sperm production and a reproductive advantage. This could be the reason for the rapid accumulation of the two AURKC mutations we observe in North African individuals. None of the authors have any competing interest. This work is part of the project 'Identification and Characterization of Genes Involved in Infertility (ICG2I)' funded by the programme GENOPAT 2009 from the French Research Agency (ANR).

Inverse correlation between chromatin condensation and sperm head size in a case of enlarged sperm heads

Among sperm morphology abnormalities, macrocephalic and large-headed spermatozoa are commonly associated with a low chance of pregnancy, mainly in relation to meiotic abnormalities during spermatogenesis. Here is reported the case of a patient with 98% of spermatozoa showing abnormal morphology, many having enlarged heads (47%). Sperm-head measurement, sperm fluorescent in situ hybridization analysis, sperm chromatin decondensation and molecular biology were performed. Fifty-six percent of the sperm displayed a large head (length >4.7 μm and width >3.2 μm), and the mean sperm-head area was 15.8 ± 3.8 μm 2 (9.7 ± 1.5 μm 2 and 9.3 ± 1.4 μm 2 for two controls). Normal chromosomal content was found in 97% of the cells and no aurea kinase C-gene mutation was found. Mean sperm chromatin decondensation rate was 46%, 64% for large-head forms and 10% for other forms. This is, as far as is known, one of the first cases of semen with enlarged-head spermatozoa linked to sperm chromatin condensation dysfunction with no major meiotic dysfunction. The study centre advised the couple to undergo intracytoplasmic sperm injection with the patient’s spermatozoa. Among sperm morphology abnormalities, macrocephalic and large-headed spermatozoa are commonly associated with a low chance of pregnancy, mainly in relation to meiotic abnormalities during spermatogenesis. We report the case of a 51-year-old referred to our centre for a 3-year history of secondary infertility who had 98% of spermatozoa with abnormal morphology and many with enlarged sperm heads (47%). Sperm head measurements, sperm fluorescent in situ hybridization analysis, and sperm chromatin decondensation were performed to propose the best treatment for this couple. In contrast with previous studies indicating a high abnormal chromosome content correlated with the percentage of macrocephalic spermatozoa, 97% of the spermatozoa in this case had a normal content for chromosomes analysed. Aniline blue staining showed a high rate of decondensed spermatozoa (46%), compared with controls (19% and 20%). We showed that the mean sperm head area for decondensed sperm heads was greater ( P < 0.0001) than that for condensed sperm heads, and an increased rate ( P < 0.0001) of decondensed sperm heads was found for macrocephalic spermatozoa (>14 μm 2), compared with normal sperm heads; respectively 64% and 10%. Because during the ICSI procedure the operator select apparently normal sperm form (mainly with normal condensed head), we advised, in this particular case, the couple to undergo ICSI with the patient’s spermatozoa as they had a good prognosis compared with previous data published for macrocephalic spermatozoa.

A new AURKC mutation causing macrozoospermia: implications for human spermatogenesis and clinical diagnosis

The presence of close to 100% large-headed multi-tailed spermatozoa in the ejaculate has been described as a rare phenotype of male infertility with a very poor prognosis. We demonstrated previously that most cases were caused by a homozygous mutation (c.144delC) in the Aurora Kinase C gene (AURKC) leading to the absence or the production of a non-functional protein. AURKC deficiency in these patients blocked meiosis and resulted in the production of tetraploid spermatozoa unsuitable for fertilization. We describe here the study of two brothers presenting with large-headed spermatozoa. Molecular analysis of the AURKC gene was carried out in two brothers presenting with a typical large-headed spermatozoa phenotype. Both affected brothers were heterozygous for the c.144delC mutation. After complete sequencing of the gene a new heterozygous variant, c.436-2A>G, was identified in both patients. This mutation is located in the acceptor consensus splice site of exon 5. AURKC transcripts were extracted from one of the patient's leukocytes and reverse transcription polymerase chain reaction could be realized showing the presence of a truncated transcript indicating that c.436-2A>G leads to the skipping of exon 5. These results indicate that AURKC molecular analysis of patients with large-headed spermatozoa should not be stopped in the absence of a homozygous recurrent mutation on exon 3 but complete sequence analysis should be performed. This diagnosis is important as the identification of AURKC mutations in patients indicates that all spermatozoa will be chromosomally abnormal and that ICSI should not be attempted.

Study of aneuploidy rate and sperm DNA fragmentation in large-headed, multiple-tailed spermatozoa

The aim of this study was to analyse the meiotic segregation and DNA fragmentation rates in ejaculated spermatozoa of Tunisian men who presented the macrocephalic sperm head syndrome and to compare the results with those from 20 fertile men with normal semen profiles. Sperm DNA fragmentation was evaluated by the terminal desoxynucleotidyl transferase-mediated deoxyuridine triphosphate biotin nick-end labelling assay. Fluorescence in situ hybridisation for chromosomes X, Y and 18 was performed for the study of meiotic segregation. Despite a normal blood karyotype, patients with large-headed spermatozoa showed a significantly higher incidence of sperm chromosomal abnormalities compared with the control group. For all the patients, tetraploidy, triploidy and diploidy were the most observed abnormalities. A very high level of DNA fragmentation was shown for these patients. In conclusion, our results demonstrated that patients with large-headed, multiple-tailed spermatozoa had significantly higher incidence of sperm chromosomal abnormalities and very high level of DNA fragmentation. So intracytoplasmic sperm injection should not be recommended to these patients, not only because of its low chances of success rate but also because of its high genetic risk.

Confirmation de la forte prévalence au Maroc de la mutation homozygote c.144delC du gène aurora kinase C (AURKC) dans les tératozoospermies avec spermatozoïdes macrocéphales

To confirm the recurrent character of the mutation c.144delC in the aurora kinase C (AURKC) gene in Morocco and determine the indication and the interest of the research of this anomaly. We looked for the mutation c.144delC in the AURKC gene in 18 infertile Moroccan patients. They were seen in medical genetic consultation within the framework of the preparatory assessment for medically-assisted procreation. Genomic DNA was extracted from 5 ml of EDTA-blood. The gene AURKC exon 3 was amplified by PCR then sequenced by using the Big-Dye Terminator V3.1 kit and an ABI Prism 310 Genetic Analyzer (Applied Biosystems). All the patients who had a typical phenotype with high rates of large-headed spermatozoa were homozygous for the mutation c.144delC in the AURKC gene. We confirm in this study the research interest of the recurrent mutation c.144delC in the gene AURKC in male infertility with high rates of large-headed spermatozoa. This molecular analysis avoids to many infertile couples unnecessary expenses and succession failures in case of use of medically-assisted procreation (MAP).

Macrocephaly in Bull Spermatozoa Is Associated with Nuclear Vacuoles, Diploidy and Alteration of Chromatin Condensation

Spermatozoa from 2 dairy AI (artificial insemination) bulls (A and B), identified by their abnormal spermiogram with cells depicting frequent macrocephaly, double tails and nuclear vacuoles, were case-investigated and compared to normal spermatozoa from a control AI sire (C). Head sizes were measured and morphological abnormalities scored using brightfield and differential interference contrast microscopy. The degree of sperm maturation and of resistance to acid-induced DNA denaturation in situ were determined after uploading of acridine orange using flow cytometry of 5,000 cells/sample. Nuclear fragmentation, i.e. the ratio of red to total (red + green) fluorescence, reached 7.1% and 31% in bulls A and B, compared to 2% in bull C. The proportion of immature spermatozoa, i.e. those with incomplete histone-protamine exchange and depicting higher green fluorescence compared to the main population of the control bull, reached 9.54% in A and 7.75% in B, compared to only 0.47% in the control. In the second part of this study the previously unknown chromosomal constitution of large-headed spermatozoa of bull A was investigated by fluorescence in situ hybridization using an X-Y painting probe set. The 7.5% XY-bearing cells and the presence of diploid spermatozoa detected by flow cytometry indicate a meiotic arrest in the first division in bull A, becoming the first proven case of association of macrocephaly and M1 diploidy. The diverse approaches used for the investigation of spermatozoal DNA provide insights into the etiology of macrocephaly.

Rôle d’aurora kinase C ( AURKC) dans la reproduction humaine

Infertility concerns at least 70 million couples worldwide. An important proportion of cases is believed to have a genetic component, yet few causal genes have been identified so far. Hundreds of genes are probably involved in spermatogenesis and oogenesis and this genetic heterogeneity has so far hindered the identification of genes causing infertility in the human. Careful morphological examination of spermatozoa can provide cues to identify homogeneous cohorts of patients likely to have the same genetic defect. We studied a cohort of North-Africans patients with a rare phenotype of large-headed spermatozoa. Using a homozygosity mapping strategy, we could map the morbid gene and we identified the same homozygous mutation (c.144delC) in the aurora kinase C gene ( AURKC) of all patients studied initially. We then genotyped a total of 62 patients. All who had a typical phenotype with close to 100% large-headed spermatozoa were homozygously mutated ( n = 34), whereas no AURKC mutations were detected in the others. A carrier frequency of 1/50 was established from individuals from the Maghrebian population, indicating that 1 in 10,000 men from North-African can be expected to present this form of infertility, a frequency comparable to that of Y-microdeletions, thus far the only known recurrent genetic event altering spermatogenesis. Then we demonstrated by flow cytometry that all spermatozoa have in fact a homogeneous 4C. We recommend the realisation of a molecular diagnosis to all patients with large-headed spermatozoa. ICSI is formally contraindicated for all homozygous patients who can have recourse to donor sperm or adoption. One cannot be as categorical for the patients not harbouring an AURKC mutation.

Abnormal semen quality and low serum testosterone in men with inflammatory bowel disease treated for a long time with sulfasalazine.

Our purpose was to study the effects of long-term salazopyrine treatment on male fertility. Seven patients, 21-37 years old, who had been treated for ulcerative colitis with 3 g/day of salazopyrine for more than 6 months, were examined. All their semen samples showed abnormal sperm motility (17% +/- 9.3) and abnormal morphology (36.3% +/- 14.4) with the predominance of large-headed spermatozoa. The endocrine picture and pituitary reserve were evaluated by GnRH and TRH tests. The basal values of testosterone were lower than the normal in our laboratory (greater than 3 ng/ml) and significantly lower (2.4 +/- 0.7 ng/ml, p less than 0.025) than in the control group (fertile men). LH was significantly higher (9 +/- 2 mIU/ml, p less than 0.005) but still within the limits of normal. The responses of FSH and LH to GnRH and of PRL to TRH were normal. The mechanisms by which salazopyrine may affect fertility are discussed.

The Aurora Kinase C c.144delC mutation causes meiosis I arrest in men and is frequent in the North African population

Infertility concerns a minimum of 70 million couples worldwide. An important proportion of cases is believed to have a genetic component, yet few causal genes have been identified so far. In a previous study, we demonstrated that a homozygous mutation (c.144delC) in the Aurora Kinase C (AURKC) gene led to the production of large-headed polyploid multi-flagellar spermatozoa, a primary infertility phenotype mainly observed in North Africans. We now want to estimate the prevalence of the defect, to improve our understanding of AURKC physiopathology in spermatogenesis and assess its implication in oogenesis. A carrier frequency of 1/50 was established from individuals from the Maghrebian general population, comparable to that of Y-microdeletions, thus far the only known recurrent genetic event altering spermatogenesis. A total of 62 patients were genotyped, all who had a typical phenotype with close to 100% large-headed spermatozoa were homozygously mutated (n = 32), whereas no AURKC mutations were detected in the others. Two homozygous females were identified; both were fertile indicating that AURKC is not indispensible in oogenesis. Previous FISH results had showed a great chromosomal heterogeneity in these patient's spermatozoa. We demonstrate here by flow cytometry that all spermatozoa have in fact a homogeneous 4C DNA content and are thus all blocked before the first meiotic division. Our data thus indicate that a functional AURKC protein is necessary for male meiotic cytokinesis while its absence does not impair oogenesis.

Can intracytoplasmic morphologically selected sperm injection be used to select normal-sized sperm heads in infertile patients with macrocephalic sperm head syndrome?

To study the chromosomal content of spermatozoa selected by intracytoplasmic morphologically selected sperm injection (IMSI) in cases of macrocephalic sperm head syndrome. Case report. Obstetrics, gynecology, urology, and reproductive biology departments. Two infertile patients with large-headed spermatozoa. Fluorescence in situ hybridization on selected spermatozoa with normal-sized heads after IMSI selection. Percentages of polyploid, diploid, haploid aneuploid, and normal spermatozoa. Of the six spermatozoa that could be selected, all were haploid but aneuploid. Absence of normal haploid spermatozoa among high magnification-selected spermatozoa contraindicated IMSI for these two patients.

Chromosomal analysis of spermatozoa with normal-sized heads in two infertile patients with macrocephalic sperm head syndrome

To study the chromosomal content of spermatozoa that could be selected for intracytoplasmic sperm injection (ICSI) in cases of macrocephalic sperm head syndrome. Case report. Obstetrics, gynecology, urology, and reproductive biology departments. Two infertile patient candidates for ICSI presenting with total teratozoospermia (100%) with mainly large-headed spermatozoa (91% and 82%, respectively). Fluorescence in situ hybridization with X, Y, 18 centromeric probes on unselected spermatozoa (all migrated spermatozoa) and specifically on selected spermatozoa with normal-sized heads. Percentage of polyploid, diploid, aneuploid, and normal haploid spermatozoa, according to X, Y, 18 chromosome centromeric probes on selected spermatozoa (head size compatible with ICSI). All the nonselected spermatozoa were abnormal, diploid, or polyploid. The rate of normal ploidy (haploid cells) among the selected sperm population was 1 per 28 for patient 1 and 5 per 51 for patient 2. This very low proportion of normal haploid spermatozoa among selected spermatozoa contraindicated ICSI for the two patients. We suggest performance of this selection and analysis before including (or not), in an ICSI program, patients with macrocephalic sperm head syndrome, associated or not with preimplantation genetic diagnosis.

Dynamics of activation of in vitro matured bovine follicular oocytes following combined ethanol and cycloheximide treatment

Among sperm morphology abnormalities, macrocephalic and large-headed spermatozoa are commonly associated with a low chance of pregnancy, mainly in relation to meiotic abnormalities during spermatogenesis. Here is reported the case of a patient with 98% of spermatozoa showing abnormal morphology, many having enlarged heads (47%). Sperm-head measurement, sperm fluorescent in situ hybridization analysis, sperm chromatin decondensation and molecular biology were performed. Fifty-six percent of the sperm displayed a large head (length >4.7 μm and width >3.2 μm), and the mean sperm-head area was 15.8 ± 3.8 μm2 (9.7 ± 1.5 μm2 and 9.3 ± 1.4 μm2 for two controls). Normal chromosomal content was found in 97% of the cells and no aurea kinase C-gene mutation was found. Mean sperm chromatin decondensation rate was 46%, 64% for large-head forms and 10% for other forms. This is, as far as is known, one of the first cases of semen with enlarged-head spermatozoa linked to sperm chromatin condensation dysfunction with no major meiotic dysfunction. The study centre advised the couple to undergo intracytoplasmic sperm injection with the patient’s spermatozoa.Among sperm morphology abnormalities, macrocephalic and large-headed spermatozoa are commonly associated with a low chance of pregnancy, mainly in relation to meiotic abnormalities during spermatogenesis. We report the case of a 51-year-old referred to our centre for a 3-year history of secondary infertility who had 98% of spermatozoa with abnormal morphology and many with enlarged sperm heads (47%). Sperm head measurements, sperm fluorescent in situ hybridization analysis, and sperm chromatin decondensation were performed to propose the best treatment for this couple. In contrast with previous studies indicating a high abnormal chromosome content correlated with the percentage of macrocephalic spermatozoa, 97% of the spermatozoa in this case had a normal content for chromosomes analysed. Aniline blue staining showed a high rate of decondensed spermatozoa (46%), compared with controls (19% and 20%). We showed that the mean sperm head area for decondensed sperm heads was greater (P < 0.0001) than that for condensed sperm heads, and an increased rate (P < 0.0001) of decondensed sperm heads was found for macrocephalic spermatozoa (>14 μm2), compared with normal sperm heads; respectively 64% and 10%. Because during the ICSI procedure the operator select apparently normal sperm form (mainly with normal condensed head), we advised, in this particular case, the couple to undergo ICSI with the patient’s spermatozoa as they had a good prognosis compared with previous data published for macrocephalic spermatozoa.