Radicaux libres et spermatozoïdes humains: physiologie et physiopathologie

The role of reactive oxygen species in the physiopathology of human sperm function has been emphasized in recent years. Their production in semen has been associated with loss of motility, decreased capacity for spermoocyte fusion and loss of fertility. In semen preparations, there are two major sources of reactive oxygen species: leucocytes and spermatozoa themselve. It has been proposed that reactive oxygen species production by human spermatozoa was dependent upon a membrane-bound NADPH oxidase or a mitochondrial diaphorase. Hydrogen peroxide produced by the dismutation of superoxide anion has been recognized as the most toxic oxidizing species for human spermatozoa. Owing to their high content of polyunsaturated fatty acids, it has been proposed that lipid peroxidation of the sperm plasma membrane is largely responsible for defective sperm function. Reactive oxygen species also affect the sperm axoneme as a result of ATP depletion, inhibit mitochondrial functions, and synthesis of DNA, RNA and proteins, produce cytoskeletal modifications and inhibit sperm-oocyte fusion. Human spermatozoa possess enzymatic defence systems such as superoxide dismutase, glutathion peroxidas/reductase and catalase to counteract the toxic effects induced by reactive oxygen species. Correlations have been reported between their effectiveness and the duration of sperm motility. If the excessive production of reactive oxygen species is detrimental for human spermatozoa, they could also participate in the physiological function of the spermatozoa when present at low concentrations. Indeed, reactive oxygen species have been shown to be involved in the activation of several enzymes. Furthermore, sperm capacitation, acrosome reaction and sperm-zona interaction would be enhanced by reactive oxygen species.     

Estrogènes et spermatozoïdes

Aromatase is the terminal enzyme responsible for estrogen biosynthesis; it is present in the endoplasmic reticulum membrane of steroidogenic cells in vertebrates. This enzyme functions with the ubiquitous reductase as the electron donor. The aromatase gene is unique and its expression is regulated in a tissue and more precisely in a cell-specific fashion via the alternative use of several promoters located in the first exons. This enzymatic complex is generally involved in development, reproduction, sexual differentiation and behaviour, but also in bone and lipid metabolism, brain functions and diseases such as breast and testicular tumors. The aromatase gene expression and its transduction in a fully active protein in testicular somatic cells and germ cells together with the widespread distribution of estrogen receptors (ERα & β) in the testis and the genital tract of the male, are clearly in favor of a physiological role for estrogens in the spermatogenesis processings especialy in sperm maturation. Therefore, we begin to understand the physiopathological roles of the estrogens in males indeed, the aromatase deficiency is associated, with severe bone maturation problems and sterility in man. Conversely, it is also obvious that estrogens in excess are responsible of the impaired spermatogenesis. These female hormones (or the ratio androgens/estrogens) do play a physiological role in the development and maintenance of male gonadal functions and obviously, several steps are concerned especially the sperm production and maturation.     

Chimiotaxonomie des lycopodiales: distribution des alcaloïdes

The distribution of lysine-derived alkaloids in the genus Lycopodium (s.l.) supports the separation of the following taxa, Huperzia (= Urostachys), Lycopodiella s.l. (= Lepidotis, excl. L. volubile and L. deuterodensum) from Lycopodium s. str. Within the latter, the Fastigiatum group and the Complanata section (= Diphasiastrum) can be distinguished. The results obtained are in good agreement with the classification proposed by Wilce.     

Spermatozoïdes,environnement et prévention

Over the last fivty years, the risk factors for infertility have increased substantially, particularly those due to the environment. Spermatogenesis and spermatozoa can be affected by physical (ionizing radiation microwaves, heat, cryopreservation) or chemical agents (antimitotics drugs, antibiotics, tranquillizers, insecticides, pesticides, industrial solvants, some heavy metals, alcohol, cannabis etc.). Some natural factors, as stress or paternal age (ageing or very youthful age relatively to about thirty) also seems to affect spermatogenesis and, particularly, the age can be joined with the previous ones. On the whole, these factors are able to decrease the male fertility through some changes about the concentration, the motility or the morphology of spermatozoa and so it is possible to describe populations subject to the risk. Moreover, these spermatogenetic changes can lead abnormalities in progeny. For instance, some antimitotic drugs as cyclophosphamide, when administrated to the male rat, lead malformations or functional anomalies as behavioral troubles. The industrial solvents lead a decrease of the birth weight and the cannabis leads an increase of the ante-or post-natal death. Moreover, the change of the paternal spermatogenesis caused by cannabis can be found again in the male progeny. The problem is similar with the lead, the benzodiazepines and the alcohol. Concerning the physical factors, some authors have shown that the children born from radiation exposed fathers presented an increase of the probability of leukemia. In animal, the postimplantation loss is increased when the father is irradiated or subjected to heat before mating. Finally, the paternal ageing is responsible for new dominant autosomic mutations. Moreover, in animal and man, paternal ageing and, in man, very youthful age, also seems responsible for a gradual lowering in the level of progency cerebral functions. On the whole, these data should lead to an preventive attitude which would be more effective before about thirty years of age than after this period.     

Distribution et activités des alcaloïdes défensifs des Coccinellidae

From a survey of 30 species and varieties of ladybugs the presence of alkaloids appears to be correlated with the existence of aposematic colour and not with being carnivorous or phytophagous. The alkaloids described until now all belong to the Coccinellini and are closely related, but other types of bases have been detected in some genera. The observed distributions are in agreement with the modern taxonomy of the family.Ladybug alkaloids constitute an effective defence against ants, Myrmica rubra, and quails, Coturnix coturnix, but all the beetles containing alkaloids do not possess the same degree of protection. Individual quail react differently towards moderately protected species.The bioassay used for the first isolation of coccinelin is described. The repulsive activities of aqueous solutions of coccinellin and convergin towards ants have been compared.     

Morphologie des spermatozoïdes

Spermatozoa morphology is one of the qualitative characteristics of spermatogenesis. However, because of both the variations in the definition of normal morphology and the existence of different kinds of sperm abnormalities as well as the use of various techniques of morphology assessment, such a parameter is poorly used in usual laboratory work. Morphological sperm anomalies can be from testicular or post-testicular origines, while the latter is still unproved. The causes of such anomalies are either from genetic origines, but in these cases any spermatozoa demonstrate this anomaly, or due to an endogenous factor with varicocele the most usually quoted but unproved pathology, But exogenous factors, either chemical such as drugs and pesticides or physical such as heat, are also responsible for morphological sperm anomalies. Analysis of sperm morphology is indicative of both the testicular health status (in cases of occupational exposure to chemical or physical toxics) and the fertility potential since morphology is correlated to sperm motility and involved in fertilization through the acrosome reaction.     

Anticorps anti-spermatozoïdes: techniques de dépistage et interprétation des résultats

Since the first publication on the detection of sperm-agglutinating antibodies in infertile men, multiple assays have been described. The most useful tests are able to detect antibodies bound to the sperm membrane of motile spermatozoa. The immunobeads test (IBT) is considered to be the most advantageous in terms of its sensitivity, the low incidence of false-positive results, and its ability to localize antibodies of different immunoglobulin classes on the sperm surface. The IBT assay can be used in parallel with the mixed antiglobulin reaction (MAR), to detect sperm-associated antibodies in the ejaculates of infertile men, the most rational way to test for antisperm antibodies (ASA) in males. In view of the high level of agreement between the two assays, MAR, the easier of the two, may be used as a first step in the detection of these antibodies. A positive MAR must be confirmed by IBT, as this assay is more specific for the detection of IgA antibodies. The clinical significance of sperm-associated antibodies is usually established according to the proportion of motile spermatozoa coated with immunobeads, its class and its localization on the sperm surface. However, binding of immunobeads does not provide any information about the antigens against which the antibodies are directed. As the functional effects of sperm-associated antibodies may vary as a function of their antigenic specificities, other assays, using purified fertilization related antigens, are necessary to establish, for each individual, the specific impact of the antibodies on the fertilization process. The indirect IBT assay has recently become the most widely used test to detect the various classes of ASA in serum and cervical mucus of infertile women and in the serum and seminal fluid of infertile men, in combination with the direct assay described above. However, in most laboratories, it is performed with only one dilution of the biological fluid tested, usually a low dilution, so that antibody levels of no significance for fertility could be detected. This may explain a recente debate (Human Reprod, 1999) on the significance of ASA as a cause of infertility. At present, and in the absence of standardized assays able to identify the antigens involved in each individual immune reaction, antibody assays, as detected by the IBT assay, in the serum and/or genital secretions of infertile subjects might provide useful clinical guidance.     

Observation des spermatozoïdes au fort grossissement (MSOME): intérêt et perspectives

Motile Sperm Organelle Morphology Examination (MSOME) constitutes a real improvement in ART management and outcome, as it allows detection of specific sperm anomalies on living cells, which cannot be detected by routine analysis. MSOME applied to the selection of sperm injected into the oocyte is called IMSI (Intracytoplasmic Morphologically Selected sperm Injection) and is associated with a considerable improvement of implantation, clinical pregnancy and delivery rates. A high-power microscope (X 2,000 to X 10,000) and video enhancement system are necessary and technical limitations are related to cryptozoospermia and/or severe teratozoospermia. Compared to routine sperm morphology assessment, MSOME allows the detection of subtle cephalic anomalies, such as vacuoles. These vacuoles seem to have a deleterious effect on fertilization and embryo developmentin vitro. These observations have led to a detailed classification of anomalies and this morphological diagnosis on living sperm demonstrates that most sperm selected for conventional ICSI at X 400 are actually abnormal on MSOME at X 6,000 or more. In addition to the very good results obtained in IMSI, this new approach opens up interesting prospects concerning the relationship between the phenotype of the injected sperm and its fertilization capacity and embryo development. In terms of diagnosis, MSOME could be used to select and study homogeneous groups of normal sperm or homogeneous groups of sperm exhibiting the same well defined anomaly. Such studies, associated with fine analysis of injected sperm and follow-up of each oocyte and each embryo, should provide more information about the relationship between sperm structure and function and should help to define the relevant indications for IMSI and the choice of spermatozoa to be injected.     

Evaluation morphologique des spermatozoïdes

Semen analysis, as defined by World Health Organisation (WHO), is a fundamental step in the work-up of infertile couples. Spermatozoa morphology has been recognised as the best predictive factor in natural fertility, and in intrauterine insemination and classicalin vitro fertilisation. Ultrastructural spermatozoa abnormalities are the only sperm alterations likely to influence the outcome of ICSI. However, these abnormalities cannot be detected by conventional microscopy (¥100 or ¥200–400). Bartoov et al. (2002) developed a real-time spermatozoa observation system using a ¥6600 magnification called MSOME (Magnification Motile Sperm Morphology Examination). Spermatozoa abnormalities detected with this technique are vacuoles localised on spermatozoa heads with variable number, size and site (nucleus or acrosome). Spermatozoa evaluation using MSOME could be performed to predict the probability of fertilisation by these spermatozoa either spontaneously or after assisted reproductive technology.     

Élastographie et cancers thyroïdiens : certitudes et perspectives

Thyroid nodule is very frequent. On the other hand, thyroid cancer is rare and the prognosis is very good. The efficiency of specialized ultrasound examination associate with fine-needle cytology is well known but some problems remain. In some cases, the value of the technique is at fault (multinodular goiter and indeterminate cytology). Strain elastography with quantification is a well-tested technique. ShearWave elastography gives us the real value of stiffness of the nodule. Is it efficient in the delicate diagnoses? Lymph-nodes elastography will probably help us in the follow-up of thyroid cancers after initial treatment.     

Hyperparasitisme facultatif de parasitoïdes en cours de développement par les femelles des ectoparasitoïdes Eupelmus vuilleti et Eupelmus orientalis Craw

In a community of three ectoparasitoids, Dinarmus basalis, Eupelmus vuilleti and E. orientalis, the host Callosobruchus maculatus parasitised 48 h before by D. basalis, is accepted by E. vuilleti females after they have eliminated the eggs and neonatal larvae of D. basalis. This ovicidal and larvicidal behaviour enables E. vuilleti to develop on C. maculatus instead of D. basalis. E. vuilleti females are able to parasitise the L5 larval stage and the pupa of D. basalis: their larvae therefore feed at the expense of the developing parasitoid. This trophic level is that of hyperparasitism. However, E. vuilleti females rarely practise hyperparasitism on their own L5 larvae and on those of E. orientalis. This behaviour reveals a high behavioural plasticity enabled by intra- and interspecific recognition of parasitoids used as hosts. Hyperparasitism activity in E. orientalis females is higher than that in E. vuilleti females since they hyperparasitise host parasitoids more frequently without preferential species choice. However, E. vuilleti seems to be free from competitive pressure with E. orientalis, as the former penetrates deeply into a grain store contaminated with C. maculatus in contrast to E. orientalis females, which remain on the surface from where they escaped.     

Principes de préparation et de congélation des spermatozoïdes testiculaires humains

The advantages and feasibility of human testicular spermatozoa cryoconservation for intracytoplasmic sperm injection (ICSI) have now been clearly demonstrated. However, the freezing protocol is based on empirical knowledge obtained from freezing of ejaculated spermatozoa. Testicular spermatozoa may not be fully mature gametes and may also be retrieved in only limited quantities. Little research has been conducted to determine whether they have the same cryobiological requirements as ejaculated spermatozoa. A better understanding of their cryobiological features and assessment of possible subcellular changes after thawing would help to optimize testicular preparations for cryopreservation (whole biopsies, seminiferous tubules, shredded suspension, single spermatozoa, etc.), freezing-thawing procedure, freezing media, and storage. Finally, there is a growing need for welldefined criteria (nuclear quality, etc.) to evaluate the tolerance of testicular spermatozoa to freezing-thawing procedure for ICSI     

La vitalité des spermatozoïdes

Most of the numerous techniques used to assess sperm viability only have research applications, while only two classical tests, i.e. eosin-Y and hypo-osmotic swelling test (HOST), are currently used in routine sperm analysis to determine the percentage of viable sperm. A viability rate below 50% of living sperm defines necrozoospermia, a condition whose clinical significance is fairly difficult to assess as the mechanisms of sperm cell death are still poorly understood. However, even when a precise cause for necrozoospermia cannot be identified, abnormal viability requires further andrological investigations with particular emphasis on clinical and laboratory signs of chronic infection of the male reproductive tract. Intracytoplasmic sperm injection (ICSI) can yield very good pregnancy rates, even in couples with the most severe forms of male infertility. However, when no motile sperm are available after sperm preparation, the outcome of ICSI is seriously impaired, probably because of a high risk of injecting dead sperm. In these patients, sperm viability could therefore be assessed by the hypo-osmotic swelling test in order to select only viable sperm for ICSI. However, the long incubation time of sperm in the hypo-osmotic solution, as recommended in the classical HOST procedure, has been shown to be detrimental to the spermatozoa. A single sperm test able to assess the viability of each individual spermatozoon within microdroplets covered by mineral oil therefore seems to be preferable. This selection procedure is less suitable in the case of immotile frozen-thawed sperm, as viability does not appear to be reliably predicted by HOST in cryopreserved sperm. Examination of sperm viability now also evaluates programmed cell death or apoptosis, as apoptotic alterations can be detected in spermatozoa by several techniques. The percentage of apoptotic sperm is correlated with deficient sperm parameters and poor outcome of assisted reproductive techniques. More effective selection procedures are therefore needed in order to identify spermatozoa not only with intact membranes but also with an intact genome to be used for ICSI.