Association of male factors with recurrent pregnancy loss.

The role of male factors in recurrent pregnancy loss (RPL) is receiving increased attention since sperm quantity and quality, male genetic mutations, as well as epigenetic modifications, have all been associated with RPL. A growing number of studies have been published on the relationship between male factors and RPL; however, these reports are limited due to small sample sizes, inconsistent inclusion criteria, and detection methods. Herein, we investigate the effects of several male factors on RPL from a genetic and non-genetic perspective to aid clinicians in determining the etiology and optimal treatment strategy for patients with RPL.

Assessment of hormonal status in male infertility. An update.

BACKGROUND AND AIMS: The prevalence and incidence of infertility are increasing worldwide; they are associated with a significant economic and social impact. Infertility is defined as the inability to achieve pregnancy after 12 months or more of regular unprotected sex. In recent times, the male factor has gained importance and currently it contributes to approximately 50% of infertility cases. Multiple etiologies are stated, such as metabolic, anatomical, genetic or even idiopathic causes; however, the main cause is semen abnormalities. The aim of this manuscript is to provide a complete review of hormonal assessment of male infertility, as well as to review the physiology and pathophysiology related to the male gonadal axis. METHODS: This study is a narrative abstract carried out on basis of systematic bibliographic review, using articles indexed in PubMed/Medline, Scopus, Embase and Scielo, which were published during the last 20 years. RESULTS: The cornerstone of the evaluation of the hormonal status is semen analysis. Clinicians must rule out hypogonadism in those patients who present oligospermia and azoospermia, by determining levels of testosterone and gonadotropins, which provide the functionality status of the hypothalamic-pituitary-testicular axis. Evaluation of the adrenal, thyroid, and lactotroph axis are indicated in those patients with central hypogonadism. CONCLUSIONS: Despite advances in the diagnosis of male infertility, some causes are not fully understood, therefore, it is crucial to perform a timely hormonal evaluation of the male factor in infertile couples, in order to provide adequate treatment and improve fertility rates.

Irisin Improves High-Fat Diet-Induced Sexual Dysfunction in Obese Male Rats.

INTRODUCTION: Obesity is known to cause sexual dysfunction including erectile dysfunction and poor semen quality. Lifestyle modifications such as exercise have increasingly been more recognized to lower the likelihood of having sexual dysfunction or infertility in obese men. In this context, as an exercise-mimetic hormone, irisin has a potential to improve obesity-related reproductive dysfunctions. We aimed to elucidate possible effects of irisin on high-fat diet (HFD)-induced reproductive dysfunction in obese male rats. METHODS: Rats were divided into four groups: vehicle, irisin, obese, and obese + irisin. The rats in obese and obese+irisin groups were fed with HFD (60% kcal fat) pellets for 12 weeks to induce obesity, and then obesity-induced sexual dysfunction was confirmed by the sexual behavior test (SBT). Irisin and obese+irisin groups received irisin (100 ng/kg/day) infusion by an s.c. osmotic minipump for 4 weeks after HFD-induced obesity was formed. RESULTS: Irisin did improve a number of measures of copulation, including penile erection, ejaculation, and sexual performance, and also improved sperm morphology and motility and decreased fat-induced testicular damage. It decreased serum leptin levels. On the other hand, irisin did not affect serum luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone. It also increased gene expression of tyrosine hydroxylase (TH) and adrenoceptor alpha 1A (ADRA1A) in the medial preoptic area (mPOA) and nucleus accumbens (NAc). CONCLUSION: Irisin provided a marked enhancement of HFD-induced decrease in libido, potency, sexual performance, and erection in SBT. Taken together, our results emphasize that irisin has a restorative and improver role in HFD-induced reproductive dysfunctions in obese male rats.

Differential impact of four sperm preparation techniques on sperm motility, morphology, DNA fragmentation, acrosome status, oxidative stress, and mitochondrial activity: A prospective study.

BACKGROUND: Suboptimal human semen handling in vitro may induce sperm damage. However, the effects of semen swim-up, pellet swim-up, density gradient, and density gradient followed by SU on sperm motility, morphology, DNA fragmentation, acrosome reaction, intracellular reactive oxygen species, and mitochondrial activity were not fully understood. OBJECTIVES: To study the impact of four sperm preparation techniques on sperm functional parameters. MATERIALS AND METHODS: This study was conducted on 60 infertile men with a minimum sperm concentration of 20 × 106 /ml and total sperm motility of ≥30%. Each raw semen sample was divided into four aliquots. Each aliquot was prepared by one of the tested techniques. Various sperm characteristics were assessed before and after sperm preparation. RESULTS: Density gradient and density gradient followed by SU resulted in significantly higher DNA fragmentation percentages compared with semen swim-up (p < 0.001 and p < 0.001, respectively) and pellet swim-up (p < 0.001 and p < 0.001, respectively). Significantly higher percentages of spermatozoa with intact acrosome were detected in semen swim-up (p < 0.001) and pellet swim-up (p < 0.001) compared with raw semen. The percentage of reactive oxygen species-positive spermatozoa was significantly higher after pellet swim-up (p < 0.001), density gradient (p < 0.001), and density gradient followed by SU (p < 0.001) than raw semen. In addition, the percentages of 100% stained midpiece (active mitochondria) were significantly higher in semen swim-up (p < 0.001) and pellet swim-up (p < 0.001) compared with raw semen. DISCUSSION AND CONCLUSION: To the best of our knowledge, this is the first report comparing the impact of these techniques on various sperm functional parameters. Semen swim-up was more effective than density gradient in selecting better spermatozoa in terms of DNA integrity, reactive oxygen species levels, acrosome status, and mitochondrial activity. Randomized clinical trials comparing these four techniques are required to test their impact on embryo development and pregnancy outcomes.

El glifosato afecta negativamente a los espermatozoides humanos: evidencia in vitro / Glyphosate Negatively Affects Human Sperm: in vitro evidence

Determinar el efecto del glifosato sobre la calidad seminal en humanos a través de una revisión sistemática. Se realizó una búsqueda de la literatura en dos bases de datos (Pubmed y Scopus) de los terminos glyphosate/gliphosate/Roundup/yerbimat y sperm/"male reproductive tract". Los criterios de selección permitieron analizar tres artículos sobre el efecto del glifosato sobre los espermatoozides humanos. Se observó un efecto de negativo del glifosato sobre los espermatoozides humanos (movilidad, actividad mitocondrial y DNA). Aunque son pocos los artículos que evalúan el efecto del glifosfato sobre la calidad seminal, todos concluyen en sus repercusiones negativas. Investigaciones sobre el tema se requieren especialmente ahora que el Gobierno Nacional pretende implementar su uso

To determine the effect of glyphosate on seminal quality in humans through a systematic review. Two databases (Pubmed and Scopus) were searched for potentially relevant publications about glyphosate/gliphosate/Roundup/yerbimat and sperm/"male reproductive tract". Selection criteria allowed analyzing three articles on the effect of glyphosate on human spermatozoa. An adverse effect of glyphosate was observed on human spermatozoa (mobility, mitochondrial activity, and DNA). Although few articles evaluate the effect of glyphosate on seminal quality, all conclude in its negative repercussions. Research on the subject is especially needed now that the National Government intends to implement its use.

Fragmentación del ADN espermático e infertilidad masculina / Fragmentation of sperm DNA and male infertility

La integridad del ADN de los espermatozoides, es un indicador importante de la fertilidad, se utiliza como una variable adicional para evaluar, junto al espermograma, la calidad de una muestra seminal. En el presente trabajo se describen algunos aspectos de interés relacionados con la fragmentación del ADN espermático, cuya etiología es multifactorial y está relacionada con factores intrínsicos, como el empaquetamiento anormal de la cromatina durante la espermiogénesis, la apoptosis defectuosa antes de la eyaculación y la producción excesiva de especies reactivas del oxígeno; y con factores extrínsecos, como son los cambios en los hábitos de vida, la exposición a agentes tóxicos, así como la edad avanzada. La fragmentación está asociada al deterioro de las variables seminales, además afecta la fertilidad natural y la realizada por tratamientos de reproducción asistida, por lo que la implementación en los laboratorios de Andrología, de la tecnología para detectar si el ADN de los espermatozoides está íntegro o fragmentado, incorpora un nuevo conocimiento en el estudio de los hombres con trastornos de fertilidad, lo que contribuye a mejorar el diagnóstico y pronóstico de la infertilidad masculina. Para realizar este trabajo se revisaron 122 artículos, de los cuales 84 cumplieron con los criterios de calidad esperados. La búsqueda se realizó a través de los buscadores habituales(AU)

Sperm DNA´s integrity is an important indicator of fertility and it is used as an additional variable to evaluate the quality of a seminal sample, together with the spermogram. This paper describes some interesting aspects related to the fragmentation of sperm DNA, whose etiology is multifactorial and is related to intrinsic factors, such as the abnormal packing of chromatin during spermiogenesis, defective apoptosis before ejaculation, and the excessive production of oxygen´s reactive species; and with extrinsic factors, such as changes in life habits, exposure to toxic agents, as well as aging. Fragmentation is associated with the deterioration of seminal variables, it also affects natural fertility and that produced by assisted reproduction treatments, so the implementation in the andrology laboratories of the technology to detect if the DNA of the sperm is intact or fragmented incorporates a new knowledge in the study of men with fertility disorders, which contributes to improve the diagnosis and prognosis of male infertility. To carry out this work, 122 articles were reviewed, of which 84 met the expected quality criteria. The search was made through the usual search engines(AU)

Fragmentación del ADN espermático e infertilidad masculina / Fragmentation of sperm DNA and male infertility

La integridad del ADN de los espermatozoides, es un indicador importante de la fertilidad, se utiliza como una variable adicional para evaluar, junto al espermograma, la calidad de una muestra seminal. En el presente trabajo se describen algunos aspectos de interés relacionados con la fragmentación del ADN espermático, cuya etiología es multifactorial y está relacionada con factores intrínsicos, como el empaquetamiento anormal de la cromatina durante la espermiogénesis, la apoptosis defectuosa antes de la eyaculación y la producción excesiva de especies reactivas del oxígeno; y con factores extrínsecos, como son los cambios en los hábitos de vida, la exposición a agentes tóxicos, así como la edad avanzada. La fragmentación está asociada al deterioro de las variables seminales, además afecta la fertilidad natural y la realizada por tratamientos de reproducción asistida, por lo que la implementación en los laboratorios de Andrología, de la tecnología para detectar si el ADN de los espermatozoides está íntegro o fragmentado, incorpora un nuevo conocimiento en el estudio de los hombres con trastornos de fertilidad, lo que contribuye a mejorar el diagnóstico y pronóstico de la infertilidad masculina. Para realizar este trabajo se revisaron 122 artículos, de los cuales 84 cumplieron con los criterios de calidad esperados. La búsqueda se realizó a través de los buscadores habituales(AU)

Sperm DNA´s integrity is an important indicator of fertility and it is used as an additional variable to evaluate the quality of a seminal sample, together with the spermogram. This paper describes some interesting aspects related to the fragmentation of sperm DNA, whose etiology is multifactorial and is related to intrinsic factors, such as the abnormal packing of chromatin during spermiogenesis, defective apoptosis before ejaculation, and the excessive production of oxygen´s reactive species; and with extrinsic factors, such as changes in life habits, exposure to toxic agents, as well as aging. Fragmentation is associated with the deterioration of seminal variables, it also affects natural fertility and that produced by assisted reproduction treatments, so the implementation in the andrology laboratories of the technology to detect if the DNA of the sperm is intact or fragmented incorporates a new knowledge in the study of men with fertility disorders, which contributes to improve the diagnosis and prognosis of male infertility. To carry out this work, 122 articles were reviewed, of which 84 met the expected quality criteria. The search was made through the usual search engines(AU)

Leucocitos seminales y calidad espermática de hombres en estudio de infertilidad / Seminal leukocytes and sperm quality in men involved in infertility study

Introducción: los leucocitos en exceso en el semen humano pueden estar asociados a la disminución de la calidad espermática. La elevada incidencia de leucocitos en el semen de hombres que acuden a nuestro servicio, pudiera estar vinculada al deterioro de las variables de calidad espermática, y vincularse con problemas de infertilidad masculina. Objetivo: determinar la frecuencia de leucocitospermia en el semen de hombres que consultan por infertilidad, e identificar si existe asociación entre la presencia de leucocitospermia y alteraciones en las variables de calidad del semen. Métodos: se realizó un estudio descriptivo transversal, en el cual se incluyeron 136 hombres, con edades entre 20 y 45 años, que acudieron al examen de calidad del semen, procedentes de las consultas de atención a parejas en estudio de infertilidad, del Instituto Nacional de Endocrinología, en La Habana, y que no refirieron síntomas de infección genitourinaria clínicamente activa, ni causas demostrables que afectaran la calidad seminal. El análisis del semen, para determinar las variables seminales, se realizó siguiendo los lineamientos de la OMS, incluyendo la técnica de peroxidasa para determinar la concentración de leucocitos seminales. Se consideraron los aspectos éticos en la investigación y los resultados se analizaron mediante la aplicación de la prueba de chi cuadrado. Un valor de p< 0,05 se consideró estadísticamente significativo.Resultados: de las 136 muestras de semen evaluadas, se encontraron leucocitospermia en 31 (22,7 por ciento). Todos los indicadores de calidad seminal fueron menores en presencia de leucocitospermia, aunque solo se obtuvo una diferencia significativa en relación con la concentración espermática (p< 0,05). Conclusiones: la frecuencia de leucocitospermia en la muestra estudiada es prevalente y asociada con un deterioro estadísticamente significativo de la concentración espermática(AU)

Introduction: excessive amount of leukocytes in the human semen can be related to lower sperm quality. The high incidence of leukocytes in semen from males who went to our service could be linked to deterioration of the sperm quality variables and to male infertility problems.Objective: to determine the frequency of leukocytospermia in the semen of men who visited our infertility service and to identify the possible association between the leukocytospermia and altered quality variables of semen. Methods: a cross-sectional and descriptive study of 136 men aged 20 to 45 years, who were performed the semen quality exam and came from the service provided to couples under infertility study in the National Institute of Endocrinology in Havana. These men had not stated any symptom of clinically active urogenital infection and no proven causes that might affect the semen quality. The semen analysis to determine the seminal variables complied with the WHO guidelines including the peroxidase technique to estimate the seminal leukocyte concentrations. The ethical aspects of research were respected whereas the Chi-square test served to analyze the results; the p< 0.05 was considered as statistically significant. Results: of 136 evaluated semen samples, leukocytospermia was found in 31 (22.7 percent). All the seminal quality indicators were lower in leukocytospermia, although the significant difference was only found in the sperm concentration (p< 0.05). Conclusions: the frequency of leukocytospermia in the studied sample is prevalent and associated to statistically significant deterioration of the sperm concentration(AU)

Leucocitos seminales y calidad espermática de hombres en estudio de infertilidad / Seminal leukocytes and sperm quality in men involved in infertility study

Introducción: los leucocitos en exceso en el semen humano pueden estar asociados a la disminución de la calidad espermática. La elevada incidencia de leucocitos en el semen de hombres que acuden a nuestro servicio, pudiera estar vinculada al deterioro de las variables de calidad espermática, y vincularse con problemas de infertilidad masculina. Objetivo: determinar la frecuencia de leucocitospermia en el semen de hombres que consultan por infertilidad, e identificar si existe asociación entre la presencia de leucocitospermia y alteraciones en las variables de calidad del semen. Métodos: se realizó un estudio descriptivo transversal, en el cual se incluyeron 136 hombres, con edades entre 20 y 45 años, que acudieron al examen de calidad del semen, procedentes de las consultas de atención a parejas en estudio de infertilidad, del Instituto Nacional de Endocrinología, en La Habana, y que no refirieron síntomas de infección genitourinaria clínicamente activa, ni causas demostrables que afectaran la calidad seminal. El análisis del semen, para determinar las variables seminales, se realizó siguiendo los lineamientos de la OMS, incluyendo la técnica de peroxidasa para determinar la concentración de leucocitos seminales. Se consideraron los aspectos éticos en la investigación y los resultados se analizaron mediante la aplicación de la prueba de chi cuadrado. Un valor de p< 0,05 se consideró estadísticamente significativo. Resultados: de las 136 muestras de semen evaluadas, se encontraron leucocitospermia en 31 (22,7 por ciento). Todos los indicadores de calidad seminal fueron menores en presencia de leucocitospermia, aunque solo se obtuvo una diferencia significativa en relación con la concentración espermática (p< 0,05). Conclusiones: la frecuencia de leucocitospermia en la muestra estudiada es prevalente y asociada con un deterioro estadísticamente significativo de la concentración espermática(AU)

Introduction: excessive amount of leukocytes in the human semen can be related to lower sperm quality. The high incidence of leukocytes in semen from males who went to our service could be linked to deterioration of the sperm quality variables and to male infertility problems. Objective: to determine the frequency of leukocytospermia in the semen of men who visited our infertility service and to identify the possible association between the leukocytospermia and altered quality variables of semen. Methods: a cross-sectional and descriptive study of 136 men aged 20 to 45 years, who were performed the semen quality exam and came from the service provided to couples under infertility study in the National Institute of Endocrinology in Havana. These men had not stated any symptom of clinically active urogenital infection and no proven causes that might affect the semen quality. The semen analysis to determine the seminal variables complied with the WHO guidelines including the peroxidase technique to estimate the seminal leukocyte concentrations. The ethical aspects of research were respected whereas the Chi-square test served to analyze the results; the p< 0.05 was considered as statistically significant. Results: of 136 evaluated semen samples, leukocytospermia was found in 31 (22.7 percent). All the seminal quality indicators were lower in leukocytospermia, although the significant difference was only found in the sperm concentration (p< 0.05). Conclusions: the frequency of leukocytospermia in the studied sample is prevalent and associated to statistically significant deterioration of the sperm concentration(AU)

Use of suboptimal sperm increases the risk of aneuploidy of the sex chromosomes in preimplantation blastocyst embryos.

OBJECTIVE: To compare autosomal and sex chromosome aneuploidy rates of embryos derived from sperm with abnormal and normal parameters. DESIGN: Retrospective cohort study. SETTING: Assisted reproduction center. PATIENT(S): Three thousand eight hundred thirty-five embryos generated from 629 couples undergoing IVF. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Incidence of aneuploidy in the trophectoderm of blastocyst embryos derived from standard IVF embryos and intracytoplasmic (ICSI) males with normal and oligozoospermic semen samples, in couples with donor eggs (mean maternal age, 25.0 years) and their own eggs (mean maternal age, 35.4 years). RESULT(S): The rate of sex chromosome aneuploidy was significantly (around threefold) higher in the oligozoospermic group compared with in both control groups (standard vs. ICSI insemination). This applied whether donor (young) or own (older) eggs were used. Significant differences were seen in the oligozoospermic samples for autosomes 1, 2, 11 (own eggs), and 18 (donor eggs) compared with both control groups; however, no significant difference was seen between each of the treatment groups for the overall rate of autosomal aneuploidy. No significant differences were seen between the two control groups (normozoospermic males, standard vs. ICSI insemination) in either of the egg group types for any chromosome pairs. CONCLUSION(S): Severe male factor infertility is associated with a significant increase in the occurrence of sex chromosome abnormalities in blastocyst embryos compared with in embryos derived from normal semen samples. Aneuploidy rates in embryos derived from sperm with normal parameters were not significantly different whether ICSI or standard insemination was used to achieve fertilization. These results highlight severe male factor infertility as a possible referral category for preimplantation comprehensive chromosomal screening.

In situ viability detection assays induce heat-shock protein 70 expression in spermatozoa without affecting the chromatin integrity.

To differentiate dead spermatozoa from viable but immotile spermatozoa, several techniques are being used during ICSI. As processed spermatozoa from poor-quality ejaculate are confronted with a higher risk of experiencing stress on exposure to altered osmotic conditions or chemicals, this study was undertaken to determine the expression of stress response gene Hsp70 and chromatin integrity in spermatozoa subjected to in situ viability assays such as hypo-osmotic swelling (HOS) test, modified hypo-osmotic swelling (M-HOS) test and pentoxifylline in 25 fresh and frozen-thawed asthenozoospermic ejaculates. RT-PCR and immunofluorescence detection of Hsp70 were performed to elucidate the expression and localisation of Hsp70 in spermatozoa, whereas DNA fragmentation analysis was performed by sperm chromatin dispersion assay. Exposure of fresh and frozen-thawed asthenozoospermic spermatozoa to M-HOS and pentoxifylline significantly increased Hsp70 expression as evidenced by increased RNA expression and immunolocalisation of Hsp70 protein in sperm head (P < 0.05-0.001). However, chromatin integrity was not significantly affected in any groups until 6 h of post-exposure time period. Our results suggest that conventional HOS may be preferred for the in situ detection of the viability as there was no immediate stress response and chromatin instability in the exposed spermatozoa.

Motile sperm organelle morphology evaluation-selected globozoospermic human sperm with an acrosomal bud exhibits novel patterns and higher levels of phospholipase C zeta.

STUDY QUESTION: Does motile sperm organelle morphology examination (MSOME) affect levels and localization patterns of the oocyte activation factor phospholipase C zeta (PLCζ) in globozoospermic sperm with and without an acrosomal bud? SUMMARY ANSWER: MSOME identified round-headed globozoospermic sperm with increased levels of PLCζ relative to sperm from the same sample that did not undergo MSOME, and identified novel patterns of PLCζ localization in sperm exhibiting an acrosomal bud. WHAT IS KNOWN ALREADY: Absence or reduction in the level of PLCζ in the sperm head, abnormal localization patterning, or defective functional ability as a result of PLCζ gene mutation, have been linked to certain types of human male factor infertility in which oocyte activation is deficient. It has been determined that a subpopulation of sperm (1%) from a patient exhibiting 100% globozoospermia presented with an acrosome bud upon MSOME. A cycle of intracytoplasmic morphologically selected sperm injection, carried out with sperm exhibiting an acrosomal bud led to pregnancy and birth of a healthy baby boy, without the use of assisted oocyte activation (AOA). STUDY DESIGN, SIZE, DURATION: Immunofluorescent analysis of PLCζ in globozoospermic sperm from three patients, before and after MSOME. PARTICIPANTS/MATERIALS, SETTING, METHODS: Quantitative immunofluorescence was used to investigate PLCζ levels and localization patterns in individual sperm (n = 1 patient) identified by MSOME and isolated by micromanipulation, and presenting with and without the acrosomal bud. A secondary aim was to investigate levels and localization patterns of PLCζ in sperm before and after MSOME from two other globozoospermic men. MAIN RESULTS AND THE ROLE OF CHANCE: Non-globozoospermic control sperm exhibited characteristic localization patterns of PLCζ immunofluorescence. Completely round-headed globozoospermic sperm from patients 1-3 were either devoid of PLCζ immunofluorescence, or exhibited an abnormal, punctate, pattern of PLCζ localization. PLCζ immunofluorescence in sperm exhibiting an acrosomal bud was observed in the midpiece with varying fluorescent intensity and was detected in 28.5% of such sperm. The majority of sperm with an acrosomal bud (43.0%) exhibited punctate patterns of PLCζ localization within the sperm head. A further 28.5% of sperm exhibited PLCζ in both the head and the midpiece. Total levels of PLCζ, and the proportions of sperm exhibiting PLCζ immunoreactivity, showed significant variance (P ≤ 0.05) amongst control [45.8 arbitrary units (a.u.) and 95.7%, respectively], non-MSOME-selected (25.9 a.u. and 46.1%, respectively) and MSOME-selected globozoospermic sperm (33.4 a.u. and 65.0%, respectively). Total levels of PLCζ immunofluorescence, and proportions of sperm exhibiting PLCζ immunoreactivity, in control sperm was significantly higher (P≤ 0.05) compared with non-MSOME-selected sperm, but not significantly different from MSOME-selected sperm. LIMITATIONS, REASONS FOR CAUTION: The low numbers of sperm analysed may not be ideal for conclusive statistical analysis. Evaluation of the effects of MSOME on morphologically normal sperm would confirm conclusions. WIDER IMPLICATIONS OF THE FINDINGS: The present findings provide hope for the future treatment of globozoospermia without the need for AOA, and provide further evidence for the clinical application of PLCζ as a therapeutic and prognostic tool. STUDY FUNDING/COMPETING INTEREST(S): The research described herein was funded by the Nuffield Department of Obstetrics and Gynaecology, University of Oxford. The authors report no conflict of interest.

Effects of semen processing on the generation of reactive oxygen species and mitochondrial membrane potential of human spermatozoa.

This study was designed to evaluate the effects of semen processing on the generation of intracellular reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) in spermatozoa, and to develop reliable indexes for the evaluation of sperm quality during sperm preparation. Swim-up and density gradient centrifugation methods were used to separate semen in oligoasthenoteratozoospermia (OAT), leucocytospermia (LC) and normozoospermia groups. Levels of ROS and MMP were measured by flow cytometry. Before preparation, the patients with abnormal semen parameters had a lower MMP and higher ROS, and there was a negative correlation between MMP and ROS. The levels of MMP and ROS increased significantly, especially ROS produced by swim-up. A significant difference was found between the correlation of MMP and total normal motile sperm count after preparation in the OAT group. The level of ROS was associated with the amount of white blood cells in the LC group. The MMP can be used as an objective index to evaluate the sperm quality of OAT patients, and the combination of MMP and ROS can be used to assess the efficiency of sperm preparation in LC patients. These findings can guide selection of the ideal sperm separation technique for different sperm samples.

Clearance after vasectomy with a single semen sample containing < than 100 000 immotile sperm/mL: analysis of 1073 patients.

OBJECTIVE: To evaluate the safety and efficacy of a new semen analysis protocol after vasectomy, where clearance is given to patients who provide a single semen sample with <100 000 immotile sperm/mL at > or = 3 months after vasectomy. PATIENTS AND METHODS: Between 1 July 2005 and 31 March 2008, 1073 men provided a first semen sample at > or = 3 months after vasectomy. Semen was first evaluated on a wet-slide preparation. Those samples with no ('azoospermia') or sporadic immotile spermatozoa could be cleared without further analysis. Samples with motile sperm were immediately labelled as potentially fertile, while those with a significant number of immotile sperm were re-analysed using a Neubauer haemocytometer. All samples with <100 000 immotile sperm/mL were cleared. RESULTS: Of men providing semen at 3 months after vasectomy, 96% could be cleared. No sperm were seen ('azoospermia') in 51.3% of samples, and 44.7% of samples contained <100 000 immotile sperm. No paternity has been reported in the cleared group after a follow-up of at least 1 year. CONCLUSIONS: A protocol stipulating that patients can be cleared after a single semen sample containing <100 000 immotile sperm/mL at > or = 3 months after vasectomy is safe and dramatically reduces the number of men who cannot be cleared at 3 months after vasectomy.

Semen preparation methods and sperm apoptosis: swim-up versus gradient-density centrifugation technique.

OBJECTIVE: To compare the effects of density-gradient centrifugation and swim-up on sperm apoptosis by using a multiparameter flow cytometric method. DESIGN: Autocontrolled split-sample study. SETTING: Tertiary infertility center. PATIENT(S): Sixty-two male partners of couples undergoing infertility investigations. INTERVENTION(S): Each sample was analyzed both before and after semen preparation by optical microscopy and by flow cytometry. MAIN OUTCOME MEASURE(S): Percentage of viable, apoptotic, and necrotic sperm and recovery rate of total motile, progressive motile, and viable sperm before and after the two sperm preparation methods. RESULT(S): Compared with the original semen, the mean percentages of apoptotic and necrotic sperm were significantly lower after both sperm preparation methods. The mean percentage of viable sperm was significantly higher after swim-up compared with gradient centrifugation. The recovery rates of total motile, progressive motile, and viable sperm were significantly higher using gradient centrifugation compared with swim-up. The viable sperm percentage and the progressive sperm motility were significant predictors for negative difference between the two methods in terms of viable sperm percentage after preparation. CONCLUSION(S): Both sperm preparation methods allow obtaining a sperm population with a low percentage of apoptotic sperm. Therefore, the risk of using apoptotic sperm for clinical treatment seems to be rather low. The choice of method will depend on whether IVF/ICSI or intrauterine insemination is to be performed.