Sperm-oviduct interactions: Key factors for sperm survival and maintenance of sperm fertilizing capacity.

BACKGROUND: Although millions or even billions of sperm are deposited in the female genital tract, only very few sperm reach the oocyte, and only one single spermatozoon will successfully fertilize. During the journey of the sperm within the female genital tract, the interactions between spermatozoa and fallopian tube are critical for sperm selection, sperm survival, and maintenance of sperm fertilizing capacity. RESULTS: This review will provide a comprehensive overview of the latest findings regarding sperm transport and behavior of sperm within the oviduct, sperm selection in the oviduct, the formation of the sperm reservoir, and the release of sperm in the presence of the oocyte. It will primarily focus on recent novel insights on sperm-oviduct interactions, which have been obtained by cutting-edge technologies under in vivo or near in vivo conditions. CONCLUSIONS: The comprehensive analysis of the findings to date will elucidate the complex molecular changes in the tubal epithelium, which are induced by the presence of the sperm and will highlight how the epithelial cells of this organ affect transport, behavior, and function of sperm. This knowledge is essential for scientists and clinicians involved in assisted reproductive technologies.

The efficiency of different types of extenders for semen cooling in stallions.

OBJECTIVE: This study was conducted to examine influence of skimmed milk-based extender (SM), INRA 96 extender and BotuSemen Gold extender on parameters of stallions' ejaculate during storage. METHODS: In this study, 14 stallions between 4 and 20 years of age were monitored. Total and progressive motility, viability and morphology of sperm were evaluated at time intervals of 24, 48, and 72 hours after collection. RESULTS: The total motility, progressive motility, and values of sperm with normal morphology were significantly higher in the INRA 96 and BotuSemen Gold extenders than in the SM (p<0.01). The sperm viability differed significantly in all extenders (p<0.01). The highest value of sperm viability was in INRA 96 (64.69%±0.67%) and lowest in SM (59.70%±0.81%). The highest differences occurred at 72 hours of storage. Values of total motility, progressive motility and sperm viability decreased over time (p<0.01). In case of sperm morphology there was no statistically significant decrease between 48- and 72-hour time intervals. CONCLUSION: It can be concluded that the extenders with a chemically defined composition have shown better indicators of insemination capabilities in ejaculates than the SM. The BotuSemen Gold extender is a suitable alternative to the INRA 96, when used within 48 hours; after 72 hours of storage, however, the INRA 96 showed a higher share of viable spermatozoa.

Investigation of Fertilizing Capacity of Zebrafish (Danio rerio) Sperm Exposed to Heavy Metals.

The objective of our study was to investigate the effects of heavy metals on the fertilizing capacity of exposed zebrafish sperm, on embryonic survival, and on occurrence of embryonic deformities following fertilization with exposed sperm. It is important to test heavy metals because they are well-known pollutants. Sperm of externally fertilizing species can get in contact with pollutants found in aquatic environment. Zebrafish sperm, despite its advantages, has seldom been used in in vitro toxicological studies and no reports are available regarding the fertilizing capacity of exposed sperm. Zebrafish sperm was stripped and exposed to concentrations of the tested heavy metals (Zn2+, Cd2+, Cr3+, Cu2+, Ni2+, Hg2+, As3+) for 30 or 120 minutes. Calculated half-maximal effective concentration (EC50) values do not differ significantly from those calculated for motility for any of the tested heavy metals, which means fertilization rate can indicate the toxicity of the given substance following exposure of sperm. Thus, its application as in vitro toxicological end point is reasonable. The survival of embryos and embryonic development have not been affected by the exposure of spermatozoa, which means all alterations in spermatozoa caused by heavy metals have been expressed before 24 hours post fertilization.

Combined in vitro fertilization and culture (IVF/IVC) in mouse for reprotoxicity assessment of xenobiotic exposure.

Litter size and other conventional measures in rodents are common end-points in the assessment of xenobiotics for reprotoxic effects. However, since litter size may be normal despite reduced semen quality, we established and tested a mouse in vitro fertilization/in vitro culture (IVF/IVC) system to assess other aspects of reprotoxicity of xenobiotic exposure. Two pesticides, vinclozolin (V) and chlormequat (C), were added to feed in low (40 and 900 ppm, respectively) and high (300 and 2700 ppm, respectively) doses and compared to control (nil pesticide). Exposed males were used for natural mating to evaluate litter size and then used for IVF/IVC and sperm evaluation. The IVF/IVC system detected significant adverse effect of high dose of vinclozolin on blastocyst formation, which was not detected by conventional measures such as litter size or sperm motility and viability. We conclude that assessment based on IVF/IVC measures may complement litter size and other conventional end-points.

Calibration of sperm concentration for in vitro fertilization in a mouse reprotoxicity model.

Xenobiotics, such as chemicals and pesticides, may result in adverse effects on reproduction in human and animals. Using in-vitro embryo production as a testing system reveals details of fertilization (IVF) and early embryonic development (IVC). The aim of our study was to perform a systematical calibration of sperm concentration in an IVF/IVC system, using an outbred mouse strain, and further determine the sperm concentration that furnishes a sensitive assessment of sperm fertilizing capacity in relation to reprotoxic evaluations. By performing breakpoint analysis, the results revealed a maximum two-cell percentage (51%, 95% CI: 38 to 69%) at 3.6 × 104 sperm/ml (95% CI: 2.1 × 104 to 6.1 × 104). For future application of the IVF/IVC system, a sperm concentration lower than this breakpoint concentration is required to be within the responsive range for determining sperm fertilizing capacity. We conclude that a relatively low sperm concentration (2.5 × 104 sperm/ml) is a precondition in a mouse IVF/IVC system in order to detect potential reprotoxic effects on sperm fertilizing capacity. Our study illustrates that a systematic approach is necessary for validation and appropriate use of such in-vitro system used for reproductive toxicity testing.

Effect of high-fat and vitamin D deficient diet on rat sperm quality and fertility.

Vitamin D (VD) is involved in many functions of the reproductive system male. The intake of diets high-fat and vitamin D deficiency (VD-) can cause morphological and physiological changes in testis that relate to infertility in the male. However, its effects on sperm quality and in vivo fertility have been little studied. This study analyzed the effects of fat and VD on sperm quality and in vivo fertility in sperm of Sprague-Dawley rats. The rats were divided into four groups: G1: control diet with vitamin D (C/VD+), G2: control diet without vitamin D (C/VD-), G3: high-fat diet with vitamin D (HF/VD+) and G4: high-fat diet without vitamin D (HF/VD-) and were fed for 3 months. Adipose tissue weight and plasma glucose were determined. Sperm quality was analyzed for motility (MO), mitochondrial function and fertilizing capacity. The intake of a high-fat diet caused a significant increase in body weight of rats (P < 0.05). There were fat-by-VD interaction effects (P < 0.05) on MO and MMP. MO and MMP were greater (P < 0.05) in G1 (54 ±â€¯5.5% and 60.5 ±â€¯2.6%) than in G3 and G4 (20 ±â€¯6.0% and 27.7 ±â€¯3.6), whereas G2 (36.7 ±â€¯8.9% and 30.7 ±â€¯4.2%) was intermediate. There was no fat-by-VD interaction for fertilizing capacity. However, fertilizing capacity was greater (P > 0.05) in animals receiving control diet (70 ±â€¯21%); than in animals receiving high-fat diet (20 ±â€¯11%). Our results demonstrated that the high-fat diet and VD- contribute to the decrease in sperm quality (MO, MMP) and consequently could decrease the fertilizing capacity.

Effect of dilution in sperm maturation media and time of storage on sperm motility and fertilizing capacity of cryopreserved semen of sex-reversed female rainbow trout.

Masculinized females, also called neomales or sex-reversed females have a male phenotype but retain the female genotype (XX). Therefore, all spermatozoa produced in their functional testes carry an X chromosome, which is desired for the production of all-female rainbow trout populations. Semen of sex-reversed female rainbow trout is of low quality and in vitro maturation is required, which includes dilution of sperm suspensions with specially formulated maturation solutions. The aim of this study was to determine the effect of dilution in different maturation media on sperm quality (sperm motility characteristics and fertilizing capacity) of frozen/thawed sperm of sex-reversed female rainbow trout. The effect of time of post-thaw storage (0, 15, 60 and 120min) on semen quality was also tested. Sperm motility parameters and fertilization rate at the eyed and hatching stages were assessed for post-thaw semen diluted in different media. The cryopreservation procedure resulted in high post-thaw sperm motility of about 57% and did not differ from fresh semen. Unexpectedly, maturation media decreased sperm activation capacity immediately after dilution; however, sperm motility increased over time. Fertilization rates of frozen/thawed semen were high (71-87%) and did not differ significantly between experimental variants at any of tested periods of storage. Our results demonstrated that the effect of the maturation media on frozen/thawed sperm is different from that of fresh sperm. The progressive increase in post-thaw sperm motility in maturation media can potentially be applied to routine hatchery practice.

Cofilin is correlated with sperm quality and influences sperm fertilizing capacity in humans.

Spermatozoa should undergo a series of biochemical modifications in female reproduction tract, which is collectively called sperm capacitation. The capacitated spermatozoa can bind to the egg zona pellucida, resulting in the occurrence of acrosome reaction which enabled spermatozoa penetrate into the egg. The formation of actin plays an important role in these processes. Actin polymerized during sperm capacitation, but the polymers dispersed before acrosome reaction. In this study, we take our focus on actin-binding protein, cofilin. Our results showed that the % and intensity of sperm expressing cofilin in normal sperm were significantly higher than in abnormal sperm, and the sperm expressing cofilin was correlated with sperm quality. Furthermore, treatment with anti-cofilin antibody increased the percentage of sperm capacitation and inhibited progesterone- or A23187- induced acrosome reaction in a dose-dependent manner. The presence of 100 ng/mL anti-cofilin antibodies markedly blocked the sperm penetration of zona-free hamster eggs. Besides, immunofluorescence results revealed that cofilin was colocalized with F-actin in the midpiece of spermatozoa; however, phospho-cofilin was expressed in the tail rather than in the midpiece of spermatozoa, which was not colocalized with F-actin in spermatozoa. Moreover, western blot revealed that phospho-cofilin increased in sperm capacitation, and the total cofilin and cofilin in insoluble fraction increased in acrosome reaction; immunofluorescence results showed that the amount of cofilin in acrosome increased in sperm capacitation. In conclusion, our study revealed that cofilin expression in human sperm is correlated with sperm quality and the alterations of cofilin and phospho-cofilin in fertilization affects sperm capacitation, acrosome reaction, and spermatozoa-oocyte fusion.

Factors affecting sperm fertilizing capacity in men infected with HIV.

Studies on the sperm-fertilizing capacity of HIV-seropositive men show conflicting results for reasons that are not yet clear. The aim of this study was to investigate the effects and relationships of some factors such as patient age, CD4(+) cells count, fathering offspring, concomitant sexually transmitted diseases (STD), and receipt of highly active anti-retroviral therapy (HAART) on sperm fertilizing capacity. Semen samples were collected from 33 HIV-seropositive men. Data on the above factors were acquired from a self-designed questionnaire. Computer-assisted sperm analysis, a hypo-osmotic swelling, and zona-free hamster oocyte penetration tests were performed according to criteria of the World Health Organization. CD4(+) cells in peripheral blood were examined using a flow cytometric (FCM) analyzer. Sperm vitality, sperm motility (grades a + b), total sperm motility, and sperm penetration rates were significantly higher in patients whose CD4(+) counts were ≧350/µl than in those whose CD4(+) counts were <350/µl (P < 0.05), and the parameters mentioned above were also significantly correlated with CD4(+) cell number (all P < 0.05). Significant differences in total sperm count and sperm tail swelling rate between patients co-infected with STD and without STD were observed (P < 0.05). Sperm penetration rate in patients receiving HAART was significantly higher than in those not receiving HAART (P < 0.05). Blood CD4(+) cell counts are an important indicator for evaluating sperm fertilizing capacity of HIV-seropositive men. After receiving HAART, the sperm penetration rate of HIV-seropositive men can be improved.

Técnicas para el análisis de la fertilidad potencial del semen equino / Techniques for analyzing the potential fertility of stallion semen / Métodos de avaliação do potencial de fertilidade do sêmen equino

Para optimizar los resultados se debe determinar la fertilidad potencial del semen equino usado en procedimientos de reproducción asistida, tales como la inseminación artificial. Sin embargo, los procesos convencionales de evaluación seminal pueden dar un bajo valor predictivo de la tasa de gestación. Por tal motivo se han desarrollado nuevos métodos de análisis seminal, incorporado diversos recursos tecnológicos y mejorado las técnicas existentes, y en muchos casos adaptándolas a la especie equina. El desarrollo de los sistemas de análisis de semen asistido por computador (CASA) ha permitido que la evaluación espermática sea más objetiva y precisa, incluyendo la determinación de nuevas variables con valor diagnóstico. El hallazgo de una amplia variedad de fluorocromos y de compuestos conjugados a sondas fluorescentes y el desarrollo de diferentes tecnologías para visualizar y cuantificar la fluorescencia de la célula y sus compartimentos permite un análisis más completo de los espermatozoides. La aplicación de ensayos o técnicas que utilizan oocitos de otras especies o incluso partes de la zona pelúcida ha favorecido el diagnóstico más certero de la verdadera capacidad fecundante de los espermatozoides equinos. El objetivo de esta revisión es ofrecer y analizar información sobre los métodos convencionales y recientes empleados para evaluar la fertilidad potencial del semen equino.

In order to optimize the results of assisted reproductive procedures such as artificial insemination, the potential fertility of stallion semen should be determined. However, conventional processes of seminal evaluation can give an low predictive value of the pregnancy rate. For this reason, the new methods of semen analysis incorporate several technological resources and improve the existing techniques. In many cases those techniques have been adapted to the equine species. The development of the computer-assisted semen analysis (CASA) has allowed a more objective and accurate sperm evaluation, including the identification of new variables with diagnostic value. The finding of a wide variety of fluorochromes and compounds conjugated with fluorescent probes, and the development of various technologies to visualize and quantify the fluorescence emmited by the cell and cell compartments allows a more complete sperm analysis. The application of tests or techniques using oocytes from other species or even parts of the zona pellucida has favored a more accurate diagnosis of the true sperm fertilizing capacity in horses. The aim of this review is to provide and analyze information on recent conventional methods used to assess the potential fertility of stallion semen.

Determinar a fertilidade potencial do sêmen equino utilizado em procedimentos de reprodução assistida como a inseminação artificial, é fundamental para aperfeiçoar seus resultados. Embora, os procedimentos convencionais de avaliação seminal poderiam ter um valor preditivo reduzido das taxas de gestação que se podem obter a partir da utilização deste. Por tal motivo, tem se desenvolvido novos métodos de analise seminal, incorporando diversos recursos tecnológicos e melhorando as técnicas existentes, e em muitas ocasiões adaptando-lhas a espécie equina. O desenvolvimento dos sistemas de analise de sêmen assistido por computador (CASA), tem permitido que a avaliação espermática fosse mais objetiva e precisa, incluindo a determinação de novas variáveis com valor diagnostica. Tem se encontrado uma ampla variedade de fluorocromos e de compostos conjugados a sondas fluorescentes, e o desenvolvimento de diferentes tecnologias para visualizar e quantificar a fluorescência emitida pela célula e seus compartimentos, o qual tem permitido um analise mais ampla das características dos espermatozoides. A aplicação de ensaios ou técnicas que utilizam ovócitos de outras espécies ou incluso partes da zona pelúcida, tem favorecido dispor de sistemas de diagnostico mais certeiros da verdadeira capacidade fecundante dos espermatozoides equinos. Esta revisão tem como objetivo, recopilar e analisar informação referente aos métodos convencionais e recentes, que poderiam ser empregados para a avaliação da fertilidade potencial do sêmen equino.

Influence of Sperm Fertilizing Capacity on Embryonic Development and Pregnancy in in Vitro Fertilization / 대한불임학회지

OBJECTIVES: To assess the fertilizing capacity using sperm penetration assay (SPA) to predict the outcome of the in vitro fertilization-embryo transfer (iVF-ET) outcome. MATERIALS AND METHODS: Semen samples were provided by 129 patients undergoing iVF. We attempted to correlate the extent of sperm penetration under enhanced SPA protocol with the results of fertilization, cleavage, preimplantation embryo development, and pregnancy. RESULTS: Univariate analysis demonstrated a statistically significant correlation between fertilizing capacity and motility, kinetics, fertilization, cleavage and embryo development, and pregnancy rate. By logistic regression analysis, fertilizing capacity was found to be the only variable that was statistically significant with respect to pregnancy rate. Fertilizing capacity, cleavage rate and pregnant rate were significantly higher in pregnant group. However, the fertilization rates was comparable with both group. CONCLUSIONS: Lower fertilizing capacity could denote a poorer prognosis for establishing a pregnancy, even after satisfactory fertilization rate is achieved.

A Case of Male Infertility, who Induced Natural Pregnancy with Treatment using Hochu-ekki-to and Keishi-bukuryo-gan After In Vitro Fertilization Failure / 日本東洋医学雑誌

The authors report a case of primary male infertility with Grade I varicocele that responded well to Kampo therapy. The duration of infertility was 8 years. During that time, the authors tried AIH (artificial insemination with husband's semen) 26 times and HIT (hysteroscopic insemination into tube) twice, but both these measures failed. IVF-ET (in vitro fertilization-embryo transter) was then attempted twice. In the first attempt, the authors succeeded in fertilization and cleavage of one of ten extracted eggs, but implantation failed to occur. In the second trial, none of the five eggs extracted were successfully fertilized. The authors then prescribed a combination of Hochuekki-to and Keishibukuryo-gan. Three months later, natural pregnancy occurred that resulted in the birth of a healthy boy by natural delivery. Although the authors have sometimes seen cases of natural pregnancy after the failure of IVF-ET, none of such cases suffered from reduced fertility. Male infertility is frequently involved with cases of decreased fertility. In such cases, success is rarely achieved with ordinary IVF-ET. For this reason, the science of microscopic fertilization is developing rapidly. There is currently no remedy for low fertility. In this type of situation, Kampo may help to improve fertility.