Cauda Epididymidis Research Articles

Association of segmentation of the epididymal interstitium with segmented tubule function in rats and mice

The epithelium of the epididymal tubule has different biological functions in different regions of the tubule. Each region is further organized into lobules or intra-regional segments surrounded by connective tissue septa (CTS). Epididymal segmentation has received little direct attention, yet there is considerable evidence that expression of mRNA and protein often begins or ends precisely at the CTS border of a segment. How such 'on-off' regulation occurs coincident with the passing of the tubule from one segment to the next is unknown. This study examined the segmentation of epididymides in rats and mice. The average adult Sprague-Dawley rat and C57BL/6 mouse caput, corpus and cauda epididymides has seven, two and four, and three, one and two segments, respectively. The apoptosis response of the caput epididymal epithelium to deprivation of lumicrine factors 24 h after efferent duct ligation in rats and the epididymal expression of a marker protein, beta-galactosidase, in mice were segmented precisely. This validated both at a general response and at a specific protein level that many epididymal functions are regulated within segments. Blue dextran (molecular weight 20000) and erythrocine red (molecular weight 880) dyes infused into the interstitial space of specific segments by micropuncture were retained by the CTS of the segments. In similar micropuncture experiments, [(3)H]H(2)O (molecular weight 18) was able to diffuse into an adjacent segment relatively freely whereas [(14)C]polyethylene glycol (molecular weight 4000) could not. These studies indicate that the interstitium of intra-regional segments is organized into different physiological compartments and that these compartments play a role in regulating the epididymal epithelium.

Sodium-inorganic phosphate cotransporter NaPi-IIb in the epididymis and its potential role in male fertility studied in a transgenic mouse model.

Analysis by cDNA microarrays showed that in the murine epididymis, NaPi-IIb was the predominantly expressed epithelial isoform of the sodium-inorganic phosphate cotransporter and was markedly overexpressed in the proximal region in the infertile knockout (KO) compared to the fertile heterozygous (HET) c-ros transgenic mouse. The apparent up-regulation in the KO mouse confirmed by Northern and Western blot analyses could be explained by the absence of NaPi-IIb from the initial segment of the HET epididymis, as revealed by immunohistochemistry, and its presence on the epithelial brush border throughout the proximal epididymis of KO mice, where differentiation of the initial segment fails to occur. Both NaPi-IIb mRNA and protein were scarce or absent from the cauda epididymidis of both genotypes. A high content of inorganic phosphate was measured enzymatically in the HET cauda luminal fluid, with a 27% decrease in the KO mice. This decrease, presumably from a greater reabsorption of inorganic phosphate, particularly in the initial part of the KO epididymis, may disturb the normal process of sperm maturation in these infertile males. By contrast, no apparent consequences were observed for the transport of Na+ and Ca2+, the concentrations of which (approximately 26 mM and approximately 30 microM, respectively) were measured by microelectrodes to be identical in the caudal fluid from both genotypes.

Effect of epididymis handling conditions on the quality of ram spermatozoa recovered post-mortem

Post-mortem spermatozoa recovery is an important technique for obtaining germplasm reserves from genetically valuable animals or endangered species. However, there are many factors that influence the outcome of this technique. We have studied the effect of the interval between animal’s death and sperm recovery (0, 24 or 48 h) on the quality and freezability of ram spermatozoa from cauda epididymidis. Storage temperature of epididymes (room temperature or 5 °C) was also analysed. Spermatozoa were diluted with Tes-Tris-Fructose solution supplemented with egg yolk (10%) and glycerol (4%), and frozen using a programmable biofreezer (−20 °C/min). Pre-freeze and post-thaw sperm samples showed viable spermatozoa up to 48 h after the animal’s death, although their quality declined significantly as post-mortem storage time increased. Epididymes sperm stored at 5 °C showed better motility and a lower percentage of abnormal forms than epididymes stored at room temperature after 24 and 48 h. The fertilizing ability of cauda epididymis ram spermatozoa obtained at 0 and 24 h after the animal’s death is similar to that of ejaculated spermatozoa. Therefore, a good protocol for post-mortem semen collection in rams when epididymal spermatozoa cannot be collected inmediately, is to preserve the epididymes at 5 °C and process the samples in the first 24 h after the animal’s death.

Postnatal development and testosterone dependence of a rat epididymal protein identified by neonatal tolerization.

A rat epididymal protein of 27 kDa was identified using neonatal tolerization. This study reports the production and characterization of a polyclonal antiserum to this protein. ELISA was used to demonstrate that this antiserum reacts strongly with epididymal sperm proteins, but has little or no reactivity with testicular proteins. Western blot analysis revealed that this polyclonal antiserum recognized a 27 kDa protein extracted from the corpus epididymidis as well as from spermatozoa from the corpus and cauda epididymides, and immunostaining revealed the presence of the protein in the corpus to cauda epididymides. Stronger reactivity was observed in the supranuclear region and stereocilla of principal cells of the corpus epididymidis and in the luminal content of the corpus and cauda epididymides. The testicular section showed no reactivity. Treatment with the antiserum resulted in time- and dose-dependent agglutination of rat spermatozoa. By indirect immunofluorescence, the antiserum localized proteins in the mid-piece region of rat spermatozoa. Studies were carried out to determine the age at which the protein first became apparent during postnatal development. The protein was expressed from day 40 onwards, as demonstrated by western blot analysis. The androgen regulation of this protein was ascertained by castration and supplementation studies. Expression of this protein showed a decline starting at day 14 after castration and by day 21 the protein was absent; however, androgen replacement resulted in the reappearance of the protein. The results of these studies indicate that the protein identified is specific to the epididymis, and is regulated by development and androgens. The importance of epididymis-specific proteins that are regulated by androgens in sperm maturation is discussed, and the need to ascertain the sequence of the protein and clone the cognate gene is indicated.

Resorption of the element zinc from spermatozoa by the epididymal epithelium.

In this study, elimination of the element zinc from spermatozoa during epididymal maturation was investigated. Testes and epididymides from 40 bulls were collected; epididymal fluid was flushed, pooled, labelled with 0.5 MBq 65Zn2+ per sample and proteins were separated on a Sephacryl S-200 HR and zinc chelate column chromatography. To follow the resorption of zinc in the epididymal epithelial lining, an autometallographic technique (AMG) was performed in tissue from caput, corpus, cauda and vas deferens. The results showed a zinc-binding protein fraction with an apparent molecular weight of 150-160 kDa, which was enriched after chelate column chromatography. Specific labelling of 65Zn was about five times higher in the caput than in the cauda epididymidis. AMG revealed no detectable zinc in the caput, but a significant increase of zinc resorption from the corpus to the cauda and vas deferens. Controls showed that the detectable zinc was located within the principal cells. In conclusion, our study proves that zinc present in the sperm flagellum starts to be mobilized in the caput epididymidis and is resorbed by the epididymal epithelium as from the corpus. This zinc elimination is a mandatory step in sperm maturation to obtain motility.

Effects of caloric restriction on gene expression along the epididymis of the Brown Norway rat during aging

Aging in the Brown Norway (BN) rat, is associated with changes in the male reproductive tract. In the epididymis, the site of sperm maturation and storage, aging has profound effects on the histology, biochemistry and gene expression of the tissue. Caloric restriction consistently slows aging and maintains health in aging animals. The goal of this study was to assess the effects of caloric restriction on aging-related changes in gene expression in the BN rat epididymis using cDNA microarrays. Caloric restriction attenuated or reversed age-related gene expression changes throughout the epididymis. In the distal regions of the tissue (corpus and cauda epididymidis), greater than 80% of the gene expression decreases were attenuated by caloric restriction. In the most proximal region of the tissue (initial segment), caloric restriction affected the expression of genes associated with lipid and carbohydrate metabolism. In all regions of the epididymal epithelium, caloric restriction had a dramatic effect on the age-related decreased expression of genes associated with protein synthesis and mitochondrial function. These results indicate that caloric restriction lessens age-related mitochondrial dysfunction and improves protein synthetic capability throughout the epididymis.

GLUTATHIONE-RELATED ENZYMES IN CELL CULTURES FROM DIFFERENT REGIONS OF HUMAN EPIDIDYMIS

Protection of maturing sperm from potential endogenous or exogenous harmful substances during their transit throughout the epididymis is a critical event. The authors studied the activity of gamma-glutamyl transpeptidase (GGT) and glutathione S-transferase (GST), and glutathione (GSH) levels in epithelial cell cultures from human caput, corpus, and cauda epididymides. Tissue was obtained from patients undergoing therapeutic orchidectomy for prostatic cancer. Enzymatic activity was measured in conditioned media and cellular fractions. Androgen influence was also evaluated. Both enzymatic activities were found in cellular homogenates and conditioned media from cultures of all epididymal regions. GGT activity was highest in cultures from cauda epididymis, both in conditioned media and cell fractions, while GST activity did not show regional differences in conditioned media, but exhibited higher activity in cell homogenates from cauda cultures than those obtained from corpus and caput epididymis. GSH level showed no regional difference in cell homogenates and it could not be detected in conditioned media by the method used. Presence of different concentrations of dihydrotestosterone (DHT) had no influence neither on the enzymatic activities nor GSH concentration. The results indicate that GGT and GST are present along the human epididymis and a fraction or isoform of these enzymes might be secreted to the luminal fluid to play a detoxificative role in sperm maturation.

Gene expression is differentially regulated in the epididymis after orchidectomy.

The epididymis is the site for the transport, maturation, and storage of spermatozoa. Regulation of epididymal structure and function is highly dependent on the ipsilateral testis. At the molecular level, however, few studies have been undertaken to determine which genes are expressed in the epididymis under testicular regulation. The goal of this study was to identify genes for which expression is regulated after orchidectomy, both throughout the epididymis and in a segment-specific manner. Microarrays spotted with 474 rat cDNAs were used to examine gene expression changes over the first 7 d post orchidectomy in the initial segment, caput, corpus, and cauda epididymidis of the adult Brown Norway rat. Using k-means cluster analysis, we show that four patterns of gene expression are activated in each epididymal segment over the first week following orchidectomy. Transient up-regulation of gene expression in the epididymis after orchidectomy is described for the first time. Potential androgen-repressed genes, including Gpx-1, show increased expression in the epididymis after orchidectomy. Several glutathione-S-transferases and calcium-binding proteins decline throughout the epididymis after orchidectomy, indicating that these may be novel androgen-regulated epididymal genes. Other genes coding for metabolism-associated proteins, transporters, and alpha-1 acid glycoprotein show segment-specific regulation in the epididymis after orchidectomy. Finally, we describe the expression of the previously uncharacterized heat shock proteins, and apoptosis-associated genes in the epididymis after orchidectomy. Thus, gene expression in the epididymis is differentially affected over time after orchidectomy. These results provide novel insight into androgen-dependent and segment-specific epididymal function.

2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) induces oxidative stress in the epididymis and epididymal sperm of adult rats.

2,3,7,8-Tetrachlorodibenzo- p-dioxin (TCDD) is one of the most potent environmental contaminants, which has been shown to induce oxidative stress in testis and epididymal sperm of rats. However, the nature and mechanism of action of TCDD on the epididymis is not clear. The aim of the present study was to investigate whether induction of oxidative stress in epididymal sperm was direct effect of TCDD on epididymis. In the present studies, TCDD (0.1, 1.0 and 10 micro g/kg body weight per day) was administered orally to rats for 4 days. Twenty-four hours after the last treatment the animals were killed using anesthetic ether. Both epididymides were dissected out and epididymal sperm were collected by cutting the epididymides into small pieces in Ham's F-12 medium at 35 degrees C. The epididymal sperm and caput, corpus and cauda epididymides were homogenized and used for biochemical studies. Epididymal sperm counts did not decrease in the rats treated with TCDD. Administration of TCDD increased the production of reactive oxygen species such as hydrogen peroxide while the activities of antioxidant enzymes superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase were found to be decreased in the epididymal sperm as well as in cauda epididymides. Lipid peroxidation also increased in the epididymal sperm and in the various regions of the epididymides after exposure to TCDD. The results indicated that TCDD induces oxidative stress in the epididymis and epididymal sperm by decreasing the antioxidant enzymes through induction of reactive oxygen species. Thus, the adverse effects of TCDD on the epididymal sperm were due to direct effect of TCDD on epididymis.

Motility activation and second messenger signalling in spermatozoa from rat cauda epididymidis.

This study examined molecular mechanisms involved in the activation of motility in spermatozoa from the cauda epididymidis of rats. A 1.05-fold dilution of semen from the cauda epididymidis with 300 mmol sucrose l(-1) did not activate motility in spermatozoa. Addition of dibutyryl cAMP, pentoxifylline or Ca(2+) to the sucrose activated motility in the short term (<30-60 min). A fivefold dilution of semen from the cauda epididymidis with a modified Tyrode's medium (BWW) activated and sustained vigorous motility that could not be attenuated with kinase inhibitors. This motility was associated with a transient increase in intracellular cAMP during the first 60 s of activation. Lower motility was activated in Ca(2+)-deficient media but this was not associated with an increase in cAMP. A fivefold dilution with plasma from the cauda epididymidis did not activate motility. The addition of Ca(2+) to the sucrose induced an increase in cAMP of similar duration but lower magnitude to that associated with dilution in BWW. The results from this study indicate that the cAMP and Ca(2+) signal transduction pathways are involved in activation of sperm motility, and that the increase in intracellular cAMP in rat spermatozoa from the cauda epididymidis undergoing motility activation is Ca(2+)-dependent. This is the first study to report a Ca(2+)-dependent increase in cAMP associated with motility activation in immotile mammalian spermatozoa. In light of these data, a model is proposed whereby cAMP and Ca(2+) act as synarchic messengers, initiating a signal transduction cascade, which is independent of protein kinase A-mediated phosphorylation of flagella proteins in immotile spermatozoa from the cauda epididymidis.

Studies on sperm storage in the vas deferens of the spinifex hopping mouse (Notomys alexis).

The cauda epididymidis, with its relatively cool temperature (32-35 degrees C), is considered to be the main site of sperm storage in male mammals. However, in the adult male spinifex hopping mouse, Notomys alexis, similar numbers of spermatozoa are found in the vas deferens to those in the cauda epididymidis. The present study shows that, unlike in the laboratory mouse in which spermatozoa of the vas deferens are found mainly in the epididymal region of the duct, spermatozoa in the hopping mouse are localized mainly to the middle and urethral regions of the vas deferens which lies in the inguinal and lower abdominal region of the body cavity. After ligation of the vas deferens close to its connection with the epididymis, many spermatozoa in the vas deferens retain the potential for motility for up to 2 weeks, indicating that the viability of spermatozoa is not compromised by being restricted to core body temperature. This urethral region of the vas deferens, in which spermatozoa reside, has a highly divergent structural organization compared with that of common laboratory rodents in which there is an expanded lumen with a network of epithelial folds. Ultrastructural observations of the cells lining the duct indicate that there are not any marked differences in morphology compared with the cells lining the duct in common laboratory murids, but the infoldings of the vas deferens of the hopping mouse are highly vascular which might facilitate supply of oxygen and nutrients to the spermatozoa residing in the lumen.

Motility activation and second messenger signalling in spermatozoa from rat cauda epididymidis

Motility activation and second messenger signalling in spermatozoa from rat cauda epididymidis

Principal cells in the cauda epididymidis resorb zinc eliminated from spermatozoa

Principal cells in the cauda epididymidis resorb zinc eliminated from spermatozoa

ILA 147 immunoreactivity of the bull spermatozoa membrane during epididymal maturation

A cytochemical quantitative study was carried out to detect immunostaining of bull spermatozoa during epididymal maturation using an ILA 147 monoclonal antibody and a standard immunoperoxidase method. This antibody recognizes a bovine panleukocyte determinant. Microdensitometric measurements were made on spermatozoa collected from different sites of the male genital tract (caput, corpus and cauda epididymidis and ductus deferens). On the basis of ILA 147 staining, at least 5 subpopulations of sperm cells from each site of the genital tract were found. These subpopulations showed: 1) immunostaining in the acrosome domain and in the cytoplasmic droplet in a proximal position; 2) immunostaining in the acrosome domain and in the distal cytoplasmic droplet; 3) immunostaining in the acrosomal region and lack of a cytoplasmic droplet; 4) immunostaining only in the cytoplasmic droplet; 5) lack of a cytoplasmic droplet and absence of staining of the head. The possible relationship between the presence of ILA 147 on the sperm head and the maturation process was evaluated, and we suggest that the most significant changes in ILA 147 expression occur in the corpus epididymidis. The absence of immunocytochemical staining in some spermatozoa may be related to plasma membrane damage due to spontaneous peroxidative damage of lipids.

Detection of ClC-3 and ClC-5 in epididymal epithelium: immunofluorescence and RT-PCR after LCM.

Epithelial cells of the epididymis and vas deferens establish an optimum luminal environment in which spermatozoa mature and are stored. This is achieved by active transepithelial transport of various ions including Cl(-) and H(+). We investigated the localization of three closely related members of the ClC family, ClC-3, ClC-4, and ClC-5, in the epididymis and vas deferens. RT-PCR using mRNA isolated by laser capture microdissection (LCM)-detected ClC-3 and ClC-5 transcripts but did not detect any ClC-4-specific transcript. Western blot and immunofluorescence analysis demonstrated that ClC-3 and ClC-5 proteins are present in all regions of the epididymis and in the vas deferens. ClC-5 is expressed exclusively in H(+)-ATPase-rich cells (narrow and clear cells). Confocal microscopy showed that ClC-5 partially colocalizes with the H(+)-ATPase in the subapical pole of clear cells. ClC-3 is strongly expressed in the apical membrane of principal cells of the caput epididymidis and the vas deferens and is less abundant in principal cells of the body and cauda epididymidis. These findings are consistent with a potential role for ClC-3 in transepithelial chloride transport by principal cells and for ClC-5 in the acidification of H(+)-ATPase-containing vesicles in narrow and clear cells. ClC-5 might facilitate endosome trafficking in the epididymis, as has been proposed in the kidney.

Glutathione-related enzymes in cell cultures from different regions of human epididymis.

Protection of maturing sperm from potential endogenous or exogenous harmful substances during their transit throughout the epididymis is a critical event. The authors studied the activity of gamma-glutamyl transpeptidase (GGT) and glutathione S-transferase (GST), and glutathione (GSH) levels in epithelial cell cultures from human caput, corpus, and cauda epididymides. Tissue was obtained from patients undergoing therapeutic orchidectomy for prostatic cancer. Enzymatic activity was measured in conditioned media and cellular fractions. Androgen influence was also evaluated. Both enzymatic activities were found in cellular homogenates and conditioned media from cultures of all epididymal regions. GGT activity was highest in cultures from cauda epididymis, both in conditioned media and cell fractions, while GST activity did not show regional differences in conditioned media, but exhibited higher activity in cell homogenates from cauda cultures than those obtained from corpus and caput epididymis. GSH level showed no regional difference in cell homogenates and it could not be detected in conditioned media by the method used. Presence of different concentrations of dihydrotestosterone (DHT) had no influence neither on the enzymatic activities nor GSH concentration. The results indicate that GGT and GST are present along the human epididymis and a fraction or isoform of these enzymes might be secreted to the luminal fluid to play a detoxificative role in sperm maturation.

52. Phytoestrogen exposure reduces fertility of male rats

Phytoestrogens are plant-derived compounds able to bind to and activate oestrogen receptors α (ERα) and β (ERβ). Exposure to phytoestrogens, in particular soy, through diet is very common. ERα and ERβ are present throughout the male reproductive tract, but the exact role of oestrogen in male reproductive biology is unclear. Male and female Wistar rats used for this study were offspring of female rats maintained on a low soy diet (containing 112 μg/g isoflavanoid) prior to conception through to weaning. After weaning, the juvenile rats were fed the same low soy diet into adulthood. Six adult male rats were transferred to a high soy diet (containing 465 μg/g isoflavanoid) (n = 6); the remaining male rats were continued on the low soy control diet (n = 8). On days 3, 6, 12 and 25 following the commencement of the high soy diet, the male rats were housed overnight with prooestrus female rats (1:1). The female rats were housed separately until parturition. The size and sex ratio of the litters were recorded. After the final mating, the male rats were killed and the epididymides were removed. Sperm counts were performed on the initial segment, caput, corpus and cauda of one epididymis from each rat. Sperm counts showed fewer sperm in the initial segment (P<0.05), corpus (P<0.05) and cauda (P<0.01) epididymides of high soy rats, as compared to the low soy rats. The litter sizes for the treatment groups showed an exposure-dependent response. The litter sizes of the day 3 (P<0.01) and day 6 (P<0.05) high soy groups was significantly lower than the low soy group, while litter sizes of the day 12 and 25 groups were not significantly different to low soy fed animals. The sex ratios of the litters from both groups were not significantly different. In conclusion, short-term exposure to high phytoestrogen levels reduces male fertility. The mechanisms involved in these changes are being investigated.

Mediation of contraction in rat cauda epididymidis by alpha-adrenoceptors.

Subtypes of alpha-adrenoceptors responsible for contractions of the rat cauda epididymidis were studied in vivo by micropuncture using a servo-nulling pressure transducer system. Administration of both non-selective and selective alpha-adrenoceptor agonists in doses of 1-40 microg noradrenaline kg(-1) body weight (BW), 1-100 microg clonidine kg(-1) BW, or 100-800 mg methoxamine kg(-1) BW enhanced contractions of the proximal cauda epididymidis in a dose-response manner. The potency of the agonists were noradrenaline > or = clonidine>methoxamine. Pre-treatments with selective alpha(1)-adrenoceptor antagonist (prazosin) and alpha(2)-adrenoceptor antagonist (yohimbine) at the doses of 400 and 800 microg kg(-1) BW, respectively, had very little effect on spontaneous contractions, but effectively blocked the responses to the maximal doses of methoxamine and clonidine. The responses could not be explained by the systemic effects of agonists and antagonists. The results suggest that contraction of the proximal cauda epididymidis of rats is mediated by both alpha(1)- and alpha(2)-adrenoceptors. The latter appears to be more abundant.

Mediation of contraction in rat cauda epididymidis by alpha-adrenoceptors

Subtypes of alpha-adrenoceptors responsible for contractions of the rat cauda epididymidis were studied in vivo by micropuncture using a servo-nulling pressure transducer system. Administration of both non-selective and selective alpha-adrenoceptor agonists in doses of 1-40 microg noradrenaline kg(-1) body weight (BW), 1-100 microg clonidine kg(-1) BW, or 100-800 mg methoxamine kg(-1) BW enhanced contractions of the proximal cauda epididymidis in a dose-response manner. The potency of the agonists were noradrenaline > or = clonidine>methoxamine. Pre-treatments with selective alpha(1)-adrenoceptor antagonist (prazosin) and alpha(2)-adrenoceptor antagonist (yohimbine) at the doses of 400 and 800 microg kg(-1) BW, respectively, had very little effect on spontaneous contractions, but effectively blocked the responses to the maximal doses of methoxamine and clonidine. The responses could not be explained by the systemic effects of agonists and antagonists. The results suggest that contraction of the proximal cauda epididymidis of rats is mediated by both alpha(1)- and alpha(2)-adrenoceptors. The latter appears to be more abundant.

Functional role of hepatocyte growth factor receptor during sperm maturation.

Mammalian spermatozoa acquire motility and fertilizing capacity during their transit through the epididymis. Hepatocyte growth factor (HGF) is a pleiotropic cytokine with potent motogenic capacities that has been identified in different organs, including the mammalian male genital tract. In mice, HGF is present in the testis and, in large amounts, in the distal part of the epididymis. In prepuberal rats, we have demonstrated that HGF is synthesized by the peritubular myoid cells and in men, HGF is present in significant quantities in seminal plasma. It has been suggested that in mice, HGF has a role in initiating sperm motility, whereas in men, no significant correlations between HGF concentration and sperm motility have been found. In the present paper we report that in rats, HGF receptor, c-met, is expressed in testicular and epididymal spermatozoa. Through immunocytochemistry, we have found that c-met is exclusively localized on the head in testicular sperm. A different localization of c-met has been found in sperm isolated from caput and cauda epididymidis. Cells isolated from epididymal caput show a c-met localization exclusively restricted to the head in most cells. In a minority of caput epididymis spermatozoa the receptor is localized both in the cell head and along the flagellum. Spermatozoa isolated from the epididymal cauda were quite homogeneous, showing the receptor localized along the entire cell surface. We also report that HGF is synthesized and secreted by the rat epididymis as indicated by the scatter effect of epididymal cell homogenate and culture medium on MDCK cells. To clarify whether HGF is involved in the acquisition of sperm motility in the epididymis, its maintenance, or both, spermatozoa isolated from caput epididymidis have been cultured in medium alone or supplemented with HGF. The results obtained indicated that HGF has a positive effect on the maintenance of sperm motility which, in the absence of HGF, significantly decreases during the first hour of culture, whereas it is maintained for at least 3 hours when HGF is present in the culture medium. We also report that HGF does not influence spermatozoa viability as indicated by the cytometrical analysis of propidium iodide-labeled sperm; an equal number of dead cells appeared in control and in HGF-treated preparations. In conclusion, our data strongly support the hypothesis that HGF positively influences sperm motility maintenance during sperm transit through the epididymis, indicating that c-met receptor and its ligand, HGF, have a role in male fertility.