Jacques Auger

World Health Organization reference values for human semen characteristics

BACKGROUND Semen quality is taken as a surrogate measure of male fecundity in clinical andrology, male fertility, reproductive toxicology, epidemiology and pregnancy risk assessments. Reference intervals for values of semen parameters from a fertile population could provide data from which prognosis of fertility or diagnosis of infertility can be extrapolated. METHODS Semen samples from over 4500 men in 14 countries on four continents were obtained from retrospective and prospective analyses on fertile men, men of unknown fertility status and men selected as normozoospermic. Men whose partners had a time-to-pregnancy (TTP) of < or =12 months were chosen as individuals to provide reference distributions for semen parameters. Distributions were also generated for a population assumed to represent the general population. RESULTS The following one-sided lower reference limits, the fifth centiles (with 95th percent confidence intervals), were generated from men whose partners had TTP < or = 12 months: semen volume, 1.5 ml (1.4-1.7); total sperm number, 39 million per ejaculate (33-46); sperm concentration, 15 million per ml (12-16); vitality, 58% live (55-63); progressive motility, 32% (31-34); total (progressive + non-progressive) motility, 40% (38-42); morphologically normal forms, 4.0% (3.0-4.0). Semen quality of the reference population was superior to that of the men from the general population and normozoospermic men. CONCLUSIONS The data represent sound reference distributions of semen characteristics of fertile men in a number of countries. They provide an appropriate tool in conjunction with clinical data to evaluate a patients semen quality and prospects for fertility.

Decline in Semen Quality among Fertile Men in Paris during the Past 20 Years

Background Several studies have suggested a population-wide decline in the quality of semen over the past 50 years, but clear evidence of decreasing semen quality in recent decades is lacking. Methods From 1973 through 1992 we measured the volume of seminal fluid, the sperm concentration, and the percentages of motile and morphologically normal spermatozoa in 1351 healthy fertile men. The data on the semen samples were collected at one sperm bank in Paris. The data in each calendar year were analyzed as a function of the year of donation, the age of each patient, the year of birth, and the duration of sexual abstinence before semen collection. Results There was no change in semen volume during the study period. The mean concentration of sperm decreased by 2.1 percent per year, from 89 ×106 per milliliter in 1973 to 60×106 per milliliter in 1992 (P<0.001). During the same period the percentages of motile and normal spermatozoa decreased by 0.6 percent and 0.5 percent per year, respectively (both P<0.001). ...

Semen quality and male reproductive health: the controversy about human sperm concentration decline

Concern about the effect of environmental changes on male reproductive health has grown in recent years to become a major preoccupation in some developed countries. A possible decline in human sperm concentration was suggested in the early seventies following studies in the US. In 1992 a metaanalysis of 61 articles published by Carlsen et al. concluded that the mean sperm count of healthy men had declined by 1% per year over the previous 50 years. From 1995 and onwards, some retrospective, longitudinal analyses of the sperm count of fertile or infertile men contradicted this while others did not. The demonstration of a geographical variation in sperm concentration, between and within countries or regions, appears to be less controversial. The amplitude of the difference observed cannot only be explained by methodological or confounding factors, and must to some extent be attributed to ethnic, genetic or environmental factors. As many of the published studies suffer from imprecision regarding the description of population characteristics and confounding factors, and were not designed with controlled and standardised methodology, the debate remains open. Prospective studies in well‐defined cohorts of men in various populations are required to evaluate the potential effect of external factors on male reproductive health. These studies should not be limited to the analysis of sperm concentration, as this may not be the best biomarker of testis function and human fertility.

Chronic Dietary Exposure to a Low-Dose Mixture of Genistein and Vinclozolin Modifies the Reproductive Axis, Testis Transcriptome, and Fertility

Background The reproductive consequences and mechanisms of action of chronic exposure to low-dose endocrine disruptors are poorly understood. Objective We assessed the effects of a continuous, low-dose exposure to a phytoestrogen (genistein) and/or an antiandrogenic food contaminant (vinclozolin) on the male reproductive tract and fertility. Methods Male rats were exposed by gavage to genistein and vinclozolin from conception to adulthood, alone or in combination, at low doses (1 mg/kg/day) or higher doses (10 and 30 mg/kg/day). We studied a number of standard reproductive toxicology end points and also assessed testicular mRNA expression profiles using long-oligonucleotide microarrays. Results The low-dose mixture and high-dose vinclozolin produced the most significant alterations in adults: decreased sperm counts, reduced sperm motion parameters, decreased litter sizes, and increased post implantation loss. Testicular mRNA expression profiles for these exposure conditions were strongly correlated. Functional clustering indicated that many of the genes induced belong to the “neuroactive ligand-receptor interactions” family encompassing several hormonally related actors (e.g., follicle-stimulating hormone and its receptor). All exposure conditions decreased the levels of mRNAs involved in ribosome function, indicating probable decreased protein production. Conclusions Our study shows that chronic exposure to a mixture of a dose of a phytoestrogen equivalent to that in the human diet and a low dose—albeit not environmental—of a common anti-androgenic food contaminant may seriously affect the male reproductive tract and fertility.

Standardisation de la classification morphologique des spermatozoïdes humains selon la méthode de David modifiée

RésuméIl est maintenant bien établi que le pourcentage de spermatozoïdes normaux et de certaines anomalies spécifiques des spermatozoïdes a une valeur pronostiquein vivo etin vitro. Aussi, le spermocytogramme représente un temps indispensable de l’analyse du sperme humain. Malheureusement, cette analyse, simple à première vue, présente de réelles difficultés avec pour conséquence une fiabilité très relative des résultats d’un laboratoire à l’autre. Aussi, les procédures employées, de la réalisation des frottis au rendu des résultats, doivent faire l’objet d’une standardisation. La lecture des lames colorées de préférence au Shorr est faite à l’objectif ×100 à immersion. La méthode de David modifiée pour le classement des anomalies morphologiques des spermatozoïdes humains a été adoptée en France par la grande majorité des laboratoires publics et privés. Elle permet le classement de 1°) sept anomalies de la tête: têtes allongées, amincies, microcéphales, macrocéphales, multiples, présentant un acrosome anormal ou absent, présentant une base (région post-acrosomique) anormale, 2°) trois anomalies de la pièce intermédiaire: reste cytoplasmique, grêle, angulée et 3°) cinq anomalies de la pièce principale: absente, écourtée, de calibre irrégulier, enroulée et multiple. La grande originalité de la méthode est de recenser l’ensemble des anomalies associées par spermatozoïde anormal grâce à un système à entrées multiples. Comme il n’y a aucune raison objective de considérer plus une anomalie qu’une autre, il est impératif de décrire toutes les anomalies observées pour chaque spermatozoïde anormal. L’index d’anomalies multiples ou IAM, application directe de ce système n’est autre que le rapport du nombre total d’anomalies recensées au nombre total de spermatozoïdes anormaux. L’IAM, indicateur du nombre moyen d’anomalies associées par spermatozoïde anormal, présente une valeur pronostique de la fertilité et I’OMS recommande son utilisation. La présente mise au point s’inscrivant dans la démarche nécessaire d’assurance de qualité en biologie de la reproduction est focalisée sur une redéfinition précise des anomalies, les justifications ultrastructurales et fonctionnelles et la pratique du classement des spermatozoïdes humains typiques et atypiques selon la méthode de David modifiée.AbstractIt has now been clearly established that the percentage of morphologically normal sperm and the level of several specific anomalies of spermatozoa have a prognostic valuein vivo andin vitro. Assessment of human sperm morphology therefore represents an essential step in routine semen analysis. Smears, preferably stained by the Shorr technique, are examined at a final magnification of ×1000 (×100 objective with oil immersion and ×10 ocular lens). David’s method for classification of morphological anomalies of human spermatozoa was proposed in the mid-seventies and was modified in the early nineties to account for all of the defects known to interfere with normal sperm functions. This method has been adopted in France by the vast majority of public and private laboratories. It allows classification of seven anomalies of the head: tapered, thin, microcephalous, macrocephalous, multiple, abnormal or absent acrosome, abnormal postacrosome, three anomalies of the midpiece: cytoplasmic droplet, thin, bent, and five anomalies of the principal piece: absent, short, irregular, coiled and multiple. The originality of the method is to account for all combinations of anomalies found for each abnormal sperm by means of a multiple entry system. Since there is no objective reason to favour certain anomalies over others, all anomalies found for each abnormal sperm should be recorded using a systematic procedure: description of the head, followed by the midpiece, and lastly the principal piece. The multiple anomalies index or MAI is calculated. This index, which is a direct application of the multiple entry system, corresponds to the mean number of anomalies per abnormal sperm (the ratio of the total number of anomalies to the number of abnormal sperm). Several studies have indicated the prognostic value of MAI, and calculation of the MAI is recommended in the WHO manual on semen analysis. Unfortunately, the morphological assessment of human spermatozoa which appears simple at a first glance, raises a number of real difficulties: this microscopic analysis is highly subjective by nature and no uniform recommendations for staining technique and classification have yet been defined. Internal and external quality control schemes have revealed the marked intra- and interindividual variability of the results, which in turn have important consequences on the reliability of the analysis and the management of infertile couples. All steps of the procedure, from preparing the smear to writing the examination report must therefore be standardised. The present article focuses on standardisation of the classification step using David’s method. Precise descriptions of the anomalies found by conventional microscopy and their ultrastructural substratum are presented, together with practical recommendations for the classification of sperm defects and calculation of the MAI.

Soy, phyto-oestrogens and male reproductive function: a review.

There is growing interest in the possible health threat posed by the effects of endocrine disruptors on reproduction. Soy and soy-derived products contain isoflavones that mimic the actions of oestrogens and may exert adverse effects on male fertility. The purpose of this review was to examine the evidence regarding the potential detrimental effects of soy and phyto-oestrogens on male reproductive function and fertility in humans and animals. Overall, there are some indications that phyto-oestrogens, alone or in combination with other endocrine disruptors, may alter reproductive hormones, spermatogenesis, sperm capacitation and fertility. However, these results must be interpreted with care, as a result of the paucity of human studies and as numerous reports did not reveal any adverse effects on male reproductive physiology. Further investigation is needed before a firm conclusion can be drawn. In the meantime, caution would suggest that perinatal phyto-oestrogen exposure, such as that found in infants feeding on soy-based formula, should be avoided.

Impact of chemotherapy and radiotherapy for testicular germ cell tumors on spermatogenesis and sperm DNA: a multicenter prospective study from the CECOS network

OBJECTIVE To determine the consequences of adjuvant testicular germ cell tumor treatment (TGCT) on sperm characteristics and sperm DNA, and to evaluate the predictors of sperm recovery. DESIGN Multicenter prospective longitudinal study of patients analyzed before treatment and after 3, 6, 12, and 24 months. SETTING University hospitals. PATIENT(S) One hundred twenty-nine volunteer TGCT patients and a control group of 257 fertile men. INTERVENTION(S) Routine semen analyses, sperm DNA, and chromatin assessments. MAIN OUTCOME MEASURE(S) Comparisons of mean sperm characteristics before and after treatment, with sperm recovery analyzed by the Kaplan-Meier method. RESULT(S) The quantitative and qualitative sperm characteristics decreased after treatment, with lowest values at 3 and 6 months and with variations according to treatment type. The mean total sperm count recovered to pretreatment values at 12 months after treatment after two or fewer bleomycin, etoposide, and cisplatin (BEP) cycles, but not after radiotherapy or more than two BEP cycles. Only the treatment modalities and pretreatment sperm production were related to recovery of the World Health Organization reference sperm values. An increased proportion of patients had elevated high sperm DNA stainability at 6 months after radiotherapy. CONCLUSION(S) Adjuvant treatments for testicular germ cell tumor have drastic effects on spermatogenesis and sperm chromatin quality. These new data on both the recovery period according to treatment modalities and the post-treatment chromatin status of sperm are useful tools for counseling patients wishing to conceive.

Flow Cytometric Sorting of Living, Highly Motile Human Spermatozoa Based on Evaluation of Their Mitochondrial Activity

We investigated the applicability of flow cytometric (FCM) sorting to select, with no deleterious effects, fractions of living, highly motile spermatozoa after staining with rhodamine 123 (Rh123) and propidium iodide (PI) for assessment of their mitochondrial activity and viability, respectively. Sperm cells were subjected to FCM sorting according to their Rh123 fluorescence intensity, and computer-aided sperm analysis (CASA) for percentage motility and movement characteristic measurements was carried out on the entire sperm populations and on the Rh123-positive (Rh123+) sorted fractions. A first experiment on five sperm samples from fertile donors pre-selected by either swim-up or simplified Percoll gradient indicated no detrimental effect of the FCM sorting procedure because: (a) the numbers of Rh123+ motile sperm were not decreased by FCM sorting; (b) data on the sorted fractions showed a tendency (not significant) for an increase in movement parameters rather than a drop; and (c) a significant decrease in the percentage of PI-positive (PI+) sperm (13% vs 3%; p < 0.05) was measured. A second experiment was performed on sperm samples from four patients, only washed and re-suspended in B2 medium. This demonstrated a significant increase in some characteristics of movement quality related to a substantial and selective immobilization of the less motile sperm. The significant drop in the percentage of PI+ sperm after FCM sorting (p < 0.01) was less pronounced than after FCM sorting of pre-selected sperm (12% vs 3%, respectively), indicating a lethal effect of FCM sorting on a small proportion of presumably moribund sperm. These preliminary data indicate a differential effect of FCM sorting on sperm according to their function characteristics and suggest the potential importance of these methods for the characterization in vitro of sperm subpopulations on the basis of functional criteria.

Serum inhibin-b in fertile men is strongly correlated with low but not high sperm counts: a coordinated study of 1,797 European and US men

OBJECTIVE To describe associations between serum inhibin-b and sperm counts, adjusted for effect of time of blood sampling, in larger cohorts than have been previously reported. DESIGN Cross-sectional studies of spermatogenesis markers. SETTING Four European and four US centers. PATIENT(S) Fertile men (1,797) were included and examined from October 1996-February 2005. INTERVENTION(S) The study was observational and therefore without any intervention. MAIN OUTCOME MEASURE(S) Associations between inhibin-b and semen variables controlled for time of blood sampling and other covariates. RESULT(S) Inhibin-b decreased about 2.00% per hour from 8 am-12 pm and then about 3.25% per hour from 12 pm-4 pm. There was a strong positive association between inhibin-b levels less than 150 pg/mL and both sperm concentration and total sperm count (slopes of the regression lines were β=0.011 and β=0.013 for natural logarithm-transformed sperm concentration and total sperm count, respectively). For inhibin-b levels of 150-300 pg/mL the associations were not as steep (β=0.002), but still significant. For inhibin-b levels more than 300 pg/mL there was little association to the sperm counts. Neither sperm motility nor morphology was significantly related to inhibin-b level in any group. CONCLUSION(S) Serum inhibin-b levels decrease nonlinearly during the daytime, and are positively correlated with sperm counts, but the predictive power is best when inhibin-b is low.

Impact of lymphoma treatments on spermatogenesis and sperm deoxyribonucleic acid: a multicenter prospective study from the CECOS network

OBJECTIVE To determine consequences of lymphoma treatments on sperm characteristics and sperm DNA, and to evaluate predictors of sperm recovery. DESIGN Multicenter prospective longitudinal study of patients analyzed before treatment and after 3, 6, 12, and 24 months. SETTING University hospitals. PATIENT(S) Seventy-five Hodgkin lymphoma and non-Hodgkin lymphoma patients and a control group of 257 fertile men. INTERVENTION(S) Semen analyses, and sperm DNA and chromatin assessments. MAIN OUTCOME MEASURE(S) Comparisons of sperm characteristics before and after treatment. RESULT(S) Patients already had altered sperm characteristics before lymphoma treatment, with no identified risk factor. Sperm count, total sperm count, motility, and vitality decreased after treatment, with lowest values at 3 and 6 months. Twelve months after treatment, mean sperm count recovered to pretreatment values after doxorubicin, bleomycin, vinblastine, darcarbacine (ABVD) or ABVD+radiotherapy, but not after doxorubicin, cyclophosphamide, vincristine, prednisone (CHOP) or mechlorethamine, oncovin, procarbazine, prednisone (MOPP) chemotherapies. It was noteworthy that 7% of patients remained azoospermic at 24 months. After 24 months, Kaplan-Meier estimates showed that more than 90% of patients will recover normal sperm count after ABVD or ABVD+radiotherapy vs. 61% for CHOP chemotherapies. In multivariate analyses including diagnosis and treatment protocol, only pretreatment total sperm count was related to recovery. Compared with a control group, lymphoma patients had higher sperm chromatin alterations and DNA fragmentation before any treatment. After treatment, DNA fragmentation assessed by TUNEL assay and sperm chromatin structure assay decreased from 3 and 6 months, respectively, while remaining higher than in the control group during follow-up. CONCLUSION(S) Lymphoma patients had altered sperm DNA and chromatin before treatment. Lymphoma treatment had damaging effects on spermatogenesis. These data on both the recovery period according to treatment modalities and the pre- and post-treatment chromatin status of sperm are useful tools for counseling patients wishing to conceive.