Impact of glycosylation on the unimpaired functions of the sperm / 대한생식의학회지

One of the key factors of early development is the specification of competence between the oocyte and the sperm, which occurs during gametogenesis. However, the starting point, growth, and maturation for acquiring competence during spermatogenesis and oogenesis in mammals are very different. Spermatogenesis includes spermiogenesis, but such a metamorphosis is not observed during oogenesis. Glycosylation, a ubiquitous modification, is a preliminary requisite for distribution of the structural and functional components of spermatids for metamorphosis. In addition, glycosylation using epididymal or female genital secretory glycans is an important process for the sperm maturation, the acquisition of the potential for fertilization, and the acceleration of early embryo development. However, nonemzymatic unexpected covalent bonding of a carbohydrate and malglycosylation can result in falling fertility rates as shown in the diabetic male. So far, glycosylation during spermatogenesis and the dynamics of the plasma membrane in the process of capacitation and fertilization have been evaluated, and a powerful role of glycosylation in spermatogenesis and early development is also suggested by structural bioinformatics, functional genomics, and functional proteomics. Further understanding of glycosylation is needed to provide a better understanding of fertilization and embryo development and for the development of new diagnostic and therapeutic tools for infertility.

Structural modification of the black bengal buck (Capra hircus) acrosome during post-testicular maturation of spermatozoa / Modificación estructural del acrosoma de la cabra black bengal (Capra hircus) durante la maduración de los espermatozoides

The present study was conducted to measure various biometric parameters of intact/normal acrosomes (AC) collected respectively from caput, corpus and cauda epididymis and vas deferens of Black Bengal buck. Giemsa stained acrosomes were measured after camera lucida drawings. Observations revealed dimensional characters of the acrosomal cap diminished gradually and significantly (p<0.01, p<0.05) during spermatozoa maturation phases in the different regions of the excurrent duct. Shape and size of the AC were also found to be influenced significantly (p<0.01, p<0.05) by the age and body weight of the animals. The structural modification along with decrease in the morphology of the AC reflected one of the maturational indexes of the male gametes in Black Bengal buck.

El presente estudio se realizó para medir diversos parámetros biométricos del acrosoma (AC) intacto/normal recogido desde la cabeza, cuerpo y cola del epidídimo y vas deferens de la Cabra Black Bengal. Los AC teñidos con Giemsa fueron medidos después de la captura con cámara lúcida. Las observaciones revelaron caracteres dimensionales del capuchón acrosomal que disminuyeron gradualmente y de manera significativa (p <0,01, p <0,05) durante fases de maduración espermatática en las diferentes regiones del conducto. La forma y tamaño del AC también fueron influenciados de manera significativa (p <0,01, p <0,05) por la edad y el peso corporal de los animales. La modificación estructural junto con los cambios morfológicos del AC refleja uno de los índices de maduración de los gametos masculinos de la Cabra Black Bengal.

Human testis-expressed sequence 101 is limitedly distributed in germinal epithelium of testis and disappears in seminoma

BACKGROUND: Testis-expressed sequence 101 (TEX101) was found to be highly expressed in testis and involved in acrosome reaction in previous studies. Recently, the metastasis suppressor function of TEX101 in cancer was disclosed, but the comprehensive investigation of its expression has rarely been reported. In this study, the expression features of TEX101 in normal human organs and seminoma were systematically analyzed. RESULTS: Immunohistochemistry demonstrated intense staining of TEX101 in human testis tissues; however, its expression in 27 other types of normal human organs, including the ovary, was negligible. Higher expression of TEX101 was observed in the spermatocytes and spermatids of the testis, but relatively lower staining was detected in spermatogonia. Western blotting showed a single TEX101 band of 38 kDa in human testis, but it did not correspond to the predicted molecular weight of its mature form at 21 KDa. Furthermore, we examined seminoma tissues by immunohistochemistry and found that none of the 36 samples expressed TEX101. CONCLUSIONS: Our data confirmed TEX101 to be a testis protein that could be related to the maturation process of male germ cells. The lack of TEX101 in seminoma indicated its potential role in tumor progression. This characteristic expression of TEX101 could provide a valuable reference for understanding its biological functions.

Reproductive cycle of the neotropical cichlid yellow peacock bass cichla kelberi: a novel pattern of testicular development

The present study describes the testicular maturation phases (associating the germ cells development and the morphological changes suffered by the germinal epithelium along the whole year), and the testicular morphology in the yellow peacock bass Cichla kelberi, relating it to other species. For this purpose, 78 specimens were studied according conventional techniques of light microscope. The testes in C. kelberi were classified as unrestricted spermatogonial lobular, an apomorphic characteristic in the recent groups of Teleost. Furthermore, were defined five testicular maturation phases: Preparatory phase; Early Germinal Epithelium Development; Mid Germinal Epithelium Development; Late Germinal Epithelium Development and; Regression. Similar classifications were described to other species indicating that the testicular classifications based on this propose, can be applied to lots of fishes. However, besides it similarity, the testicular reproductive cycle of C. kelberi follows a different pattern in the Regression phase, on which the gonadal restructuration and the spermatogonial proliferation gathers at the same time. So, the testes in C. kelberi never return to the Preparatory phase to start a new reproductive cycle, being this one present only at the first reproductive cycle in this species. This fact also explains the absence of individuals totally spent after their first reproductive cycle.

O presente estudo descreve as fases de maturação testicular (associando o desenvolvimento das células germinativas e as alterações morfológicas sofridas pelo epitélio germinativo ao longo do ano), e a morfologia testicular do tucunaré amarelo Cichla kelberi, relacionando a outras espécies. Com este propósito, 78 indivíduos foram estudados de acordo com técnicas convencionais para microscopia de luz. Os testículos em C. kelberi foram classificados como lobular espermatogonial irrestrito, uma característica apomórfica encontrada nos grupos recentes dos teleósteos. Além disso, cinco fases de maturação testicular foram definidas para C. kelberi: Fase Preparatória; Desenvolvimento Inicial do epitélio germinativo; Desenvolvimento Intermediário do epitélio; Desenvolvimento Final do epitélio germinativo e Regressão. Classificações similares foram descritas para outras espécies, indicando que a classificação gonadal baseada nesta proposta, pode ser aplicada a muitas espécies de peixes. Entretanto, apesar desta similaridade, o ciclo reprodutivo testicular de C. kelberi segue um padrão diferente na fase de Regressão, na qual a reestruturação gonadal e a proliferação espermatogonial ocorrem ao mesmo tempo. Deste modo, os testículos em C. kelberi nunca retornam à fase Preparatória para começar um novo ciclo reprodutivo nesta espécie. Este fato também explica a ausência de indivíduos totalmente esgotados após seu primeiro ciclo reprodutivo.

Intracytoplasmic spermatid injection and in vitro maturation: fact or fiction?

Intracytoplasmic injection with testicular spermatozoa has become a routine treatment in fertility clinics. Spermatozoa can be recovered in half of patients with nonobstructive azoospermia. The use of immature germ cells for intracytoplasmic injection has been proposed for cases in which no spermatozoa can be retrieved. However, there are low pregnancy rates following intracytoplasmic injection using round spermatids from men with no elongated spermatids or spermatozoa in their testes. The in vitro culture of immature germ cells to more mature stages has been proposed as a means to improve this poor outcome. Several years after the introduction of intracytoplasmic injection with elongating and round spermatids, uncertainty remains as to whether this approach can be considered a safe treatment option. This review outlines the clinical and scientific data regarding intracytoplasmic injection using immature germ cells and in vitro matured germ cells.

[Evaluation of sperm quality, maturation and DNA integrity in adult mice treated with sulpiride]

Use of certain antipsychotic drugs has severe effects on fertility in males. Hypothalamus and hypophysial impressions and changes in plasma hormones concentration like prolactin, LH and FSH can affect sperm production. In this study, we investigated the effects of sulpiride on sperm quality, maturation and DNA damage. Twenty for adult male mice [age: 6-8 weeks] were divided into three groups. The treatment group received 40 mg/kg sulpiride solution and the control sham group was given carrier of the drug intraperitoneally [IP] daily for 45 days but the control group received nothing. Finally, all the mice were sacrificed by cervical dislocation and their cauda epididymis were removed surgically. The excised specimens were placed in 1 ml HTF medium and incubated for 30 min in CO2 incubator to allow the spermatozoa to swim out. Later, sperm count, motility and viability were analyzed. Additionally, sperm chromatin quality and DNA integrity were assessed by aniline blue and acridine orange staining. Significant decrease in sperm motility and count were observed in the treatment group while the number of abnormal sperm increased as compared with the other two groups. Sperm viability and DNA maturation showed significant reduction and the rate of DNA damage increased in comparison with the control sham and the control groups [P<0.05]. The study showed that sulpiride has negative effects on sperm parameters in treated animals and in some cases it could cause secondary infertility

Hypospermatogenesis and spermatozoa maturation arrest in rats induced by mobile phone radiation

To determine the morphological changes induced by mobile phone radiation in the testis of Wistar albino rats. Cohort study. Department of Physiology, College of Medicine, King Saud University, Riyadh, Saudi Arabia, from April 2007 to June 2008. Forty male Wistar albino rats were divided in three groups. First group of eight served as the control. The second group [group B, n=16] was exposed to mobile phone radiation for 30 minutes/ day and the third group [group C, n=16] was exposed to mobile phone radiation for 60 minutes/day for a total period of 3 months. Morphological changes in the testes induced by mobile phone radiation for 60 minutes/day caused 18.75% hypospermatogenesis and 18.75% maturation arrest in the testis of albino rats compared to matched controls. However, no abnormal findings were observed in albino rats that were exposed to mobile phone radiation for 30 minutes/day for a total period of 3 months. Long-term exposure to mobile phone radiation can cause hypospermatogenesis and maturation arrest in the spermatozoa in the testis of Wistar albino rats

Estandarización del manejo y la criopreservación de semen de hembras masculinizadas de trucha arco iris (Oncorhynchus mykiss) / Padronizar a gestão ea criopreservação de sêmen de fêmeas de truta arco-íris (Oncorhynchus mykiss) sob masculinização / Standardization of handling and freezing sperm from masculinized females of rainbow trout (Oncorhynchus mykiss)

La demanda de líneas monosexo hembras en la producción de trucha ha incrementado significativamente en los últimos años, por lo que se han desarrollado tecnologías para estandarizar este proceso como el uso de semen de hembras genéticas sometidas a reversión sexual. El objetivo de la presente investigación fue estandarizar la maduración in vitro y la criopreservación de semen de hembras masculinizadas (neomachos XX) de trucha arco iris (Oncorhynchus mykiss) como estrategia para producir descendencias 100% hembras de reproductores colombianos. Para la obtención del semen los neomachos fueron sacrificados y el semen recuperado fue sometido a proceso de maduración con plasma seminal de machos normales o plasma seminal artificial. Para la criopreservación del semen se probaron los crioprotectores dimetilsulfóxido 10% y metanol 10%. En el experimento se evaluron la movilidad post maduración y post descongelación y la fertilidad del semen. El proceso de maduración tuvo un efecto significativo sobre el porcentaje de movilidad (p<0.05), al obtener los mejores valores con el plasma seminal artificial (55 ± 10.4% espermatozoides móviles). Los mayores valores de movilidad postdescongelación (29.9 ± 13.3%) al igual que la mejor fertilidad (26.33 ± 7.53%), se registraron con el protocolo de dimetilsulfóxido al 10%.

The demand of monosex female stocks in production of trout has significantly increased during the past years, which has led to develop new technologies to standardize this process. The usage of semen of genetic females submitted to sexual reversion is a good choice. The objective of this research was to develop a methodology to mature in vitro and cryopreserved semen of sex-reversed rainbow trout (Oncorhynchus mykiss) females as strategy to produce lineage 100% Colombian trout female. The semen was directly obtained from the gonads after its surgical extraction of the slaughtered individuals, later it was submitted to maturation process implementing seminal plasma of normal males and artificial plasma. The semen was cryopreserved in two extender dimetyhyl sulfoxide 10% and methanol 10%. Postmaturation, postcriopreservation movility and sperm fertility were evaluated. �Maturation process had a significative effect on movility, the highest movility was obtained with artificial seminal plasma (55 ± 10.4 %). Highest post criopreservation movility (29.9 ± 13.3%) and highest fertility rates (26.33 ± 7.53 %) were obtained with dimetyhyl sulfoxide 10%.

A procura de linhas monosexo fêmeas na produção de trutas tem aumentado significativamente nos últimos anos, de modo tecnologias foram desenvolvidas com a finalidade de padronizar este processo como o uso do esperma de genética feminina submetido a reversão sexual. O objectivo do presente inquérito foi para uniformizar a maturação in vitro e criopreservação de sêmen masculinização de fêmeas (neomachos XX) trutas arco-íris (Oncorhynchus mykiss) como uma estratégia para produzir descendentes de 100% do sexo feminino dos jogadores colombianos. Para a obtenção do esperma neomachos foram mortas e sêmen foi recuperado submetida a maturação processo normal de plasma seminal plasma seminal masculina ou artificiais. Para a criopreservação de sêmen foi testado crioprotectores dimethylsulphoxide 10% e 10% de metanol. O experimento foi evaluron mobilidade pós maturação e pós descongelamento e fertilidade do sêmen. O processo de maturação teve um efeito significativo sobre a porcentagem de mobilidade (p<0.05), para obter os melhores valores com artificial seminal plasma (55 ± 10.4% esperma). Os maiores valores de mobilidade postdescongelación (29.9 ± 13.3%), bem como a melhor fertilidade (26.33 ± 7.53%), foram registrados com o protocolo de dimethylsulphoxide a 10%.

Testicular biopsies--histomorphologic patterns in male infertility.

The objective of the present study is to observe the spectrum of histopathological changes in the testicular biopsies of infertile men and to assess if a bilateral biopsy is required to reveal the pathology of infertility in every case or a unilateral biopsy would suffice. Thirty testicular biopsies (21 bilateral and 9 unilateral) were studied from 30 infertile men. The patterns of testicular damage seen in the present study were maturation arrest followed by hypospermatogenesis, Sertoli-cell only syndrome, tubular hyalinisation and one case was associated with normal histology. Comparing the histopathological findings in bilateral biopsies, it was seen that a unilateral biopsy would suffice to reveal the pathology in most instances and a bilateral biopsy is needed only when there is appreciable difference in the size of the testes.

Supramolecular organization of the sperm plasma membrane during maturation and capacitation / 亚洲男科学杂志(英文版)

<p><b>AIM</b>In the present study, a variety of high resolution microscopy techniques were used to visualize the organization and motion of lipids and proteins in the sperm's plasma membrane. We have addressed questions such as the presence of diffusion barriers, confinement of molecules to specific surface domains, polarized diffusion and the role of cholesterol in regulating lipid rafts and signal transduction during capacitation.</p><p><b>METHODS</b>Atomic force microscopy identified a novel region (EqSS) within the equatorial segment of bovine, porcine and ovine spermatozoa that was enriched in constitutively phosphorylated proteins. The EqSS was assembled during epididymal maturation. Fluorescence imaging techniques were then used to follow molecular diffusion on the sperm head.</p><p><b>RESULTS</b>Single lipid molecules were freely exchangeable throughout the plasma membrane and showed no evidence for confinement within domains. Large lipid aggregates, however, did not cross over the boundary between the post-acrosome and equatorial segment suggesting the presence of a molecular filter between these two domains.</p><p><b>CONCLUSION</b>A small reduction in membrane cholesterol enlarges or increases lipid rafts concomitant with phosphorylation of intracellular proteins. Excessive removal of cholesterol, however, disorganizes rafts with a cessation of phosphorylation. These techniques are forcing a revision of long-held views on how lipids and proteins in sperm membranes are assembled into larger complexes that mediate recognition and fusion with the egg.</p>

Extracellular quality control in the epididymis / 亚洲男科学杂志(英文版)

The epididymal lumen represents a unique extracellular environment because of the active sperm maturation process that takes place within its confines. Although much focus has been placed on the interaction of epididymal secretory proteins with spermatozoa in the lumen, very little is known regarding how the complex epididymal milieu as a whole is maintained, including mechanisms to prevent or control proteins that may not stay in their native folded state following secretion. Because some misfolded proteins can form cytotoxic aggregate structures known as amyloid, it is likely that control/surveillance mechanisms exist within the epididymis to protect against this process and allow sperm maturation to occur. To study protein aggregation and to identify extracellular quality control mechanisms in the epididymis, we used the cystatin family of cysteine protease inhibitors, including cystatin-related epididymal spermatogenic and cystatin C as molecular models because both proteins have inherent properties to aggregate and form amyloid. In this chapter, we present a brief summary of protein aggregation by the amyloid pathway based on what is known from other organ systems and describe quality control mechanisms that exist intracellularly to control protein misfolding and aggregation. We then present a summary of our studies of cystatin-related epididymal spermatogenic (CRES) oligomerization within the epididymal lumen, including studies suggesting that transglutaminase cross-linking may be one mechanism of extracellular quality control within the epididymis.

Sperm maturation in the epididymis: a new look at an old problem / 亚洲男科学杂志(英文版)

The osmotic challenges facing maturing spermatozoa and their responses to them are discussed in relation to the concept of sperm maturation, defined as the increased ability of more distally recovered epididymal spermatozoa to fertilize eggs when inseminated into the female tract. One explanation could be that the more distal cells are better able to regulate their volume, and reach the oviducts, as a consequence of uptake of epididymal osmolytes. Increased motility, zona binding and oolemma fusion capacities are also acquired within the epididymis and are necessary for those cells that finally arrive at the site of fertilization.

[Evaluating the presence of testis specific transcripts in mature human spermatozoa]

Progress and completion of spermatogenesis is related to simultaneous expression of various genes. Recent studies show that many genes are expressed in the sperm and several RNA copies are present in the mature spermatozoa. Identification of these genes and evaluation of their functions would improve our understanding of the molecular basis of fertilization, early embryo cleavage and the causes of many types of unexplained male infertility. In this study, we investigated the expression of DAZ, PRM1, PRM2, TSGA10, SYCP3 and AKAP4 genes in ejaculated human spermatozoa. Semen samples were collected from men referring to Avicenna Infertility Clinic. Normal semen samples [According to WHO criteria] were subjected to density-gradient centrifugation to specifically recover the pure fraction of motile spermatozoa with normal morphology. Total RNA was extracted from sperm pellets and cDNA was synthesized using RT-PCR. The presence of DAZ, TSGA10, PRM1 and PRM2 cDNAs were evaluated using appropriate primers. Expression of SYCP3 [Testis specific gene] was evaluated by nested RT-PCR. The cDNA synthesized from normal testis tissues was used as positive control. Study on cDNAs showed that DAZ, TSGA10, PRM1 and PRM2 transcripts were present in normal human testis and all of the evaluated mature spermatozoa samples but not AKAP4 or SYCP3 transcripts. According to our previous study, the expression of SYCP3 and AKAP4 genes is started from spermatocyte level in human testis during spermatogenesis process. However, we did not found any transcripts of these genes in mature spermatozoa. It is estimated that mRNAs of TSGA10, PRM1, PRM2 and DAZ and other testis specific genes in spermatozoa may participate in later sperm functions such as fertilization and early embryo cleavage. Therefore, further studies are needed to understand the role of these transcripts in the process of fertilization and early embryo development

Serum concentration of testosterone, epididymal mast cell population and histamine content in relation to sperm count and their motility in albino rats following H2 receptor blocker treatment.

H2 receptor blocker treatment over a period of 2 weeks had been found to cause significant reduction in epididymal tissue mast cell population and tissue histamine content in caput, corpus and cauda regions in albino rats. There was also a highly significant fall of serum testosterone level and sperm count in these segmental fluids collected by micropuncture. The motility of sperms was also greatly reduced and the number of abnormal spermatozoa was found to be increased, the increase being highly significant in the caudal segment. In view of histamine being involved in steroidogenic activity, it appears that reduction in epididymal tissue histamine content following H2 receptor blocker treatment had caused low availability of testosterone to the tissues and thereby the reduction in sperm count and their motility. Increase in number of abnormal sperms particularly in the caudal epididymis is likely to be due to malformation and increased destruction of sperms, because of alteration in epididymal environment due to fall in serum testosterone concentration.

Carnitines and male reproduction / 中华男科学杂志

Carnitines are important conditionally essential nutrients in the organism, with extensive physiological functions, and highly concentrated in the epididymis and sperm. Carnitines play an important role not only in initiating sperm motility, promoting sperm maturation and enhancing sperm fertilizing, but also in regulating Sertoli cell function, protecting sperm against oxidative damage, reducing apoptosis of spermatogenic cell and inhibiting sperm aggregation. Accordingly, the objective of this review is to summarize the multifunctional roles of carnitine in male reproduction.

Specialized gene expression and regulation in the epididymis / 中华男科学杂志

The epididymis is a single and highly convoluted tubule system in mammals. The epithelium is the major compartment for epididymal function. Proteins synthesized and secreted by epididymal epithelium provide a special and ever-changing luminal fluid environment for sperm as they progress through the epididymis, which makes sperm achieve motility and ultimately results in sperm functional maturation. Specialized genes expressed in the epididymis have regional-specific characteristics. They are regulated by androgen and/or testicular factors and present spatial and tempel-specialized expression pattern in postnatal development, all these hint that they play important and unique roles in epididymis.

Morphological changes of Sertoli cells during the male reproductive cycle of the teleost Piaractus mesopotamicus (Holmberg, 1887)

Realizou-se uma investigação das mudanças histológicas e ultra-estruturais das células de Sertoli durante o ciclo reprodutivo de machos de Piaractus mesopotamicus. Os resultados mostraram que o desenvolvimento das células de Sertoli está estritamente relacionado à maturação das células gaméticas. Portanto, as células de Sertoli têm algum papel na maturação das células germinativas durante o ciclo reprodutivo dessa espécie, talvez formando um tecido de sustentação para os cistos espermatogênicos em desenvolvimento, ajudando a reorganização testicular para um novo ciclo reprodutivo, além de outras possíveis funções discutidas no texto.

Epididymis as a target for contraception.

Advantage of using a vaccine based on sperm antigens is that it can be used both in males and females as individuals who have antisperm antibodies are usually infertile but otherwise healthy. Several sperm specific antigens identified as prospective candidates for immunocontraception are of testicular origin. For the purpose of immunocontraception it may be desirable not to disrupt spermatogenesis and testicular function. Concept of post testicular maturation of spermatozoa has been very well established. During post testicular voyage spermatozoa undergo a series of complex and sequential events which transforms the immature immotile spermatozoa into mature sperm. Acquisition of functional maturity is necessary for progressive motility, zona pellucida recognition culminating in sperm egg binding. Importance of epididymal maturation is highlighted by the fact that high percentage of male infertility in human originates from the malfunction of the epididymis. The epididymis has also shown to be involved in sperm storage and provides an adequate environment for final maturation of the sperm. It provides a conducive microenvironment by virtue of which the spermatozoa are protected during the storage. In view of this it is imperative that more attention needs to be focused on epididymal antigens. The information obtained will enable us to identify epididymal antigens relevant to fertility and also help in infertility diagnosis.

Study on the male contraceptive based on epididymis / 中华男科学杂志

More and more study on the epididymal function and sperm maturation has shown that epididymis will be one of the best target organs of male contraception, although at present there is not a male contraceptive medicine based on epididymis for clinical practice. The promoting research aspects in epididymal contraception in animal included affecting directly epididymis (such as Sulpasalazine), interfering energy metabolism and sperm mobility (such as Chlorinated Glycerol), altering the internal environment of epididymis (such as copper particles and TW19). The epididymal specific proteins could bring out some new target antigens for immunological contraception, to produce contraceptive vaccine. Some special genes, which expressed distinctively in epididymis such as SC342, bin1, have been cloned and studied on their function. These works would be helpful not only for clinical diagnosis and treatment of epididymitis and male infertility, but also for male contraceptive research and progress.

Epididymal sperm protein P34H and male reproduction / 中华男科学杂志

During epididymal transit, mammalian spermatozoa acquire new surface proteins that are necessary for gamete interaction. P34H, a member of the short-chain dehydrogenase/reductase(SDR) superfamily, is acquired on the acrosomal cap of human spermatozoon during its maturation arising within epididymis. P34H has been shown to be involved in sperm-zona pellucida interaction. Research revealed that the occurrence of low concentration of sperm protein P34H were significant amongst the idiopathic infertile male population and P34H protein could also be considered as a marker of epididymal sperm maturation in human. Therefore the level of sperm protein P34H is proposed to be a auxiliary diagnostic tool for male infertility. This paper reviews the molecular properties and regulation of the expression of P34H and its association with male reproduction.