共查询到20条相似文献,搜索用时 15 毫秒
1.
Incubation of washed human spermatozoa in the presence of 6 mM histidine induces a release of about 75% of the zinc bound to the cells. Zinc release induced by the presence of histidine was accompanied by: 1) a significant increase in the utilization of exogenous 14C-labelled glucose, which is reflected in an increase in the production of 14CO2, 2) a small but significant decrease (−14%) in the utilization of fructose when this sugar is added as exogenous substrate, and 3) a highly significant decrease in the endogenous sperm phospholipids (>30%), an effect which is not inhibited by the addition of exogenous substrates. This behavior of human spermatozoa resembles that previously described for invertebrate spermatozoa and seems to be related to the regulation of energy metabolism and probably to sperm capacitation. 相似文献
2.
R D Raffaniello R A Wapnir 《Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine (New York, N.Y.)》1989,192(3):219-224
The luminal phase of zinc intestinal absorption has not been well characterized. This study was intended to elucidate the possible role of low molecular weight (LMW) ligands in zinc intestinal transport in an isolated rat enterocyte system. Under these in vitro conditions, zinc uptake by the isolated enterocytes was rapid, leveling off within 1 min. Kinetic analysis revealed that both a mediated and diffusion component were involved in zinc uptake in the absence of LMW ligands by the cells. For the mediated component of zinc transport, the Kt and Vmax were 64.1 microM and 13.9 nmol/20 sec/mg protein, respectively. Zinc uptake was not affected by the addition of metabolic inhibitors. In the presence of histidine or cysteine (2:1 ligand:zinc molar ratio), zinc uptake was greatly reduced and occurred solely via mediated transport. Zinc uptake was also significantly decreased upon the addition of EDTA to the assay media. Other amino acids tested had no effect on zinc uptake by the cells. Albumin markedly reduced zinc uptake by the cells. Histidine and other potential LMW ligands were unable to facilitate albumin-inhibited zinc uptake. The results of this study suggest that the intestinal absorption of zinc may not be effected in the form of chelates with LMW ligands. Amino acids such as histidine and cysteine significantly reduce the uptake of the metal by isolated rat enterocytes, making questionable their putative role as necessary vehicles in the luminal phase of zinc absorption. 相似文献
3.
4.
The effects of zinc on human sperm motility, fertilizing capacity (as assessed by penetration of human spermatozoa into the zona pellucida-free hamster oocyte), and nuclear chromatin decondensation were investigated using spermatozoa from four fertile donors. Both sperm motility and the penetration of sperm into zona-free hamster ova were consistently impaired in media containing 1,000 μM zinc. Spermatozoa from one man were similarly affected at a concentration of 500 μM zinc, but no adverse effects were noted at this zinc concentration in experiments with other donors. Since decreased fertilizing capacity in response to zinc was always accompanied by a significant decline in both the percentage of motile cells and mean swimming speeds, it appears that all of these results reflect a general toxic effect on the cells. At lower concentrations (125–250 μM), zinc had no effect on human sperm motility nor their ability to undergo capacitation and penetrate zona-free hamster ova in vitro. For some donors, zinc (125–500 μM) stimulated both the attachment of spermatozoa to the hamster vitellus and the incorporation of spermatozoa into the hamster ooplasm. The decondensation of human sperm nuclear chromatin in sodium dodecyl sulfate was largely inhibited when zinc was added to the medium, but no significant changes in nuclear stability were apparent after capacitation in zinc-free medium. We conclude that zinc, when present in subtoxic concentrations, does not adversely affect the ability of human spermatozoa to undergo capacitation and penetrate zona-free hamster ova in vitro. 相似文献
5.
Kanaya E Watanabe K Nakajima N Okada K Shimura Y 《The Journal of biological chemistry》2001,276(10):7383-7390
The FILAMENTOUS FLOWER gene from Arabidopsis thaliana is a member of a gene family whose role is to specify abaxial cell fate in lateral organs. Analysis of the amino-terminal region of the FILAMENTOUS FLOWER protein suggests that seven cysteine residues at positions 14, 26, 30, 33, 54, 56, and 57, and two histidine residues at positions 18 and 24 contribute to a putative zinc finger motif, Cys-X(3)-His-X(5)-His-X-Cys-X(3)-Cys-X(2)-Cys-X(20)-Cys-X-Cys-Cys. Zinc determination experiments revealed that the FILAMENTOUS FLOWER protein binds two zinc ions per molecule. Chemical modification was required to release one zinc ion, whereas the other was released spontaneously or more rapidly in the presence of metallochromic indicator. The loss of a zinc ion and the subsequent structural change of the zinc finger domain were correlated with the multimerization of the FILAMENTOUS FLOWER protein. A cysteine residue at position 56 in the FILAMENTOUS FLOWER protein potentially interferes with zinc ligation within the zinc finger and causes this zinc release. In support of this, substitution of the Cys(56) by alanine suppressed both the zinc release and the multimerization of the FILAMENTOUS FLOWER protein. Deletion analysis showed that the region between positions 45 and 107 functions in the intermolecular contacts between FILAMENTOUS FLOWER proteins. This region corresponds to the carboxyl-terminal half of the zinc finger domain and the following hydrophobic region containing two putative alpha-helices. Our results suggest that the FILAMENTOUS FLOWER protein forms a range of different conformers. This attribute may lead to a greater degree of functional flexibility that is central to its role as an abaxial cell fate regulator. 相似文献
6.
Prostasomes are particular lipid vesicles secreted by the human prostate and found in the semen. No specific action has yet been attributed to prostasomes, but they appear to act at various levels. For example, prostasomes enhance sperm motility in vitro and participate in the immunomodulation properties of seminal plasma. Excessive production of reactive oxygen species (ROS) in human semen has a negative influence on the functional capacities of spermatozoa. The presence of leukocytes in semen is associated with increased production of ROS that can be harmful to sperm cells, under certain conditions. Previous results tend to suggest a possible role of prostasomes on ROS production in human semen. After reviewing the literature concerning the structural and functional characteristics of prostasomes and the role of ROS in human semen, we report our results concerning the influence of prostasomes on ROS production and the consequences on semen. We have demonstrated that prostasomes exert an antioxidant function in human semen. This function is effective both on polymorphonuclear neutrophils and on sperm cells. The mechanism of action of prostasomes is unusual, as they act on ROS production mainly on the plasma membranes of neutrophils. They induce a decrease of NADPH-oxidase activity associated with rigidification of the plasma membrane. Prostasomes protect the functional capacities of spermatozoa during an oxidative stress created by the presence of NADPH in the incubation medium. 相似文献
7.
The detection and the isolation of a zinc-protein from the secretion of the rat dorsolateral prostate is described. The purification procedure, based on gel filtration and cationic exchange chromatography, allowed to separate a minor protein (Mr approximately 66,000) from free zinc ions and other secretory components. Two zinc ions were estimated to be associated with one molecule of isolated protein. The zinc-protein was labelled with 125I and then incubated at 37 degrees C with spermatozoa from rat epididymal cauda. Time-dependent in vitro binding of the radioactive protein to sperm cells was demonstrated. This binding was not affected by the presence of proteins from the seminal vesicle during the incubation, while it was blocked in the presence of an excess of unlabelled zinc-protein. After binding, the labelled spermatozoa were treated with a buffer containing 0.5% sodium deoxycholate and 40 mM EDTA; only very small amounts of label were removed from the cells, thus suggesting that the zinc-proteins were kept on the plasma membrane by interactions which do not involve merely hydrophobic bonds. 相似文献
8.
Mounir Ajina Amira Zairi Soumaya Mougou Samira Ibalaa Radhouène Naiefer Ghaya Merdassi Mariem Mehdi Khaled El Heni Ali Saad 《Andrologie》2005,15(1):53-61
The spermicidal efficacy of synthetic peptides, dermaseptin (DS1) and (DS4), was studied under in vitro conditions using human spermatozoa. The data showed that sperm motility was inhibited with various concentrations of dermaseptins at different intervals ranging from 2 to 240 min. The effective 100% inhibitory concentration (EC100) of DS4 sperm immobilization assay was equal to 50 mg/ml at 30 min, while the EC 100 of DS1 was equal to 100 mg/ml. The presence of 0.1% of chelating agent, EDTA, reduced the EC100 of DS4 to 5 mg/ml, while less than a twofold enhancement in DS 1 activity was observed in combination with EDTA. The action of dermaseptins on sperm motility was observed to be dose-dependent. Addition of pentoxifylline, which is known to enhance sperm motility, and Ca2+, which is a key element for sperm movement, did not prevent the spermicidal action of dermaseptins. 相似文献
9.
S Mack A K Bhattacharyya C Joyce H van der Ven L J Zaneveld 《Biology of reproduction》1983,28(5):1032-1042
The effect of in vitro capacitation (events that occur before the acrosome reaction) on the acrosomal enzymes of human spermatozoa was determined. Capacitation of human spermatozoa was assessed by their ability to penetrate denuded hamster oocytes. The activities of a number of enzymes commonly associated with the sperm acrosome, including nonzymogen acrosin, proacrosin, inhibitor-bound acrosin, hyaluronidase, acid phosphatase, beta-glucuronidase, beta-glucosidase, beta-N-acetylglucosaminidase, beta-galactosidase and beta-N-acetylgalactosaminidase were assessed. With the exception of acid phosphatase, no alteration in enzyme activity occurred after 4 h of incubating the spermatozoa under capacitation conditions although gamete fusion took place. The acid phosphatase levels decreased twofold, presumably due to the loss of seminal (prostatic acid phosphatase that loosely adheres to spermatozoa. After 8 h of capacitation, a large decrease in sperm enzyme levels took place only in the case of hyaluronidase, although small decreases were also noted in total acrosin, proacrosin and inhibited acrosin. No new electrophoretically migrating forms of acrosin were observed. Decreases in total acrosin and proacrosin, but not in inhibited acrosin, also occurred when spermatozoa were incubated under noncapacitating conditions for 8 h, indicating that capacitation may specifically cause the release of some acrosin inhibitor from human spermatozoa. It is concluded that, with the possible exception of hyaluronidase, the in vitro capacitation of human spermatozoa does not cause a major change in its acrosomal enzyme content so that these hydrolases are fully present before the acrosome reaction takes place during gamete fusion. Serum albumin appears to protect against the loss of some of these enzymes since the activity of several glycosidases was significantly reduced when the spermatozoa were incubated for 8 h in human serum albumin-free medium. 相似文献
10.
Immunolocalization of CRES (Cystatin-related epididymal spermatogenic) protein in the acrosomes of mouse spermatozoa. 总被引:5,自引:0,他引:5
The CRES (cystatin-related epididymal spermatogenic) protein is a member of the cystatin superfamily of cysteine protease inhibitors and exhibits highly restricted expression in the reproductive tract. We have previously shown that CRES protein is present in elongating spermatids in the testis and is synthesized and secreted by the proximal caput epididymal epithelium. The presence of CRES protein in developing germ cells and in the luminal fluid surrounding maturing spermatozoa prompted us to examine whether CRES protein is associated with spermatozoa. In the studies presented, indirect immunofluorescence, immunogold electron microscopy, and Western blot analysis demonstrated that CRES protein is localized in sperm acrosomes and is released during the acrosome reaction. Interestingly, while the 19- and 14-kDa CRES proteins were present in testicular and proximal caput epididymal spermatozoa, the 14-kDa CRES protein was the predominant form present in mid-caput to cauda epididymal spermatozoa. Furthermore, following the ionophore-induced acrosome reaction, CRES protein localization was similar to that of proacrosin/acrosin in that it was detected in the soluble fraction as well as associated with the acrosome-reacted spermatozoa. The presence of CRES protein in the sperm acrosome, a site of high hydrolytic and proteolytic activity, suggests that CRES may play a role in the regulation of intraacrosomal protein processing or may be involved in fertilization. 相似文献
11.
Campos VF Amaral MG Seixas FK Pouey JL Selau LP Dellagostin OA Deschamps JC Collares T 《Animal reproduction science》2011,126(1-2):136-141
Sperm from different species shows biological differences, determining the success or failure of the sperm-mediated gene transfer (SMGT) technique. There is evidence that exogenous DNA uptake by the spermatozoa is a species-specific and highly regulated phenomenon. Problems involving SMGT procedures might be related to activation of defenses in spermatozoa and in seminal plasma such as DNase enzymes. The objective in the present study was to transfect South American catfish spermatozoa after seminal plasma removal. Seminal plasma had a strong DNase activity that is reduced after sperm washes in isosmotic solution, in which Western blot analysis demonstrated a reduction in the DNase content after washes and Southern blot evaluations show the presence of plasmid after sperm washes. The seminal plasma DNase digests exogenous DNA in a few minutes and has an optimal activity at 43°C. Also, EDTA at 30 mM concentration inhibits the DNase activity. Using PCR the pEGFP vector was internalized by sperm cells even at lesser concentrations (5-40 ng/10(6) spermatozoa) without motility loss after seminal plasma removal. Conversely, using greater pEGFP concentrations (100 ng/10(6) spermatozoa), there were no motile cells, suggesting toxicity of exogenous DNA for sperm cells. These results are interpreted to provide information that can improve the protocol for generation of transgenic South American catfish. 相似文献
12.
13.
Release of mature bundles of spermatozoa from the testis into the vas deferens is a critical but poorly understood step in male insect reproduction. In moths, the release of sperm bundles is controlled by a circadian clock which imposes a temporal gate on the daily exit of bundles through the terminal epithelium-a layer of specialized epithelial cells separating testis follicles from the vas deferens. The sequence of cellular events associated with the daily cycle of sperm release was investigated by scanning and transmission electron microscopy. In the hours preceding sperm release, there is a solid barrier between the testis and the vas deferens formed by the interdigitation of cytoplasmic processes of adjacent terminal epithelial cells. At the beginning of the sperm release cycle, sperm bundles protrude through this barrier while the terminal epithelial cells change their shape and position relative to the bundles. Subsequently, the cyst cells enveloping the sperm bundles break down and spermatozoa move out of the testis through the exit channels formed between the epithelial cells. Afterwards, cyst cell remnants and other cellular debris are released into the vas deferens lumen, and the epithelial barrier is reconstructed due to phagocytic activity of its cells. These data provide a foundation on which to build an understanding of the cellular mechanisms of clock-controlled sperm release in insects. 相似文献
14.
Involvement of zinc in the regulation of pHi, motility, and acrosome reactions in sea urchin sperm 总被引:3,自引:2,他引:1
下载免费PDF全文
![点击此处可从《The Journal of cell biology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
D L Clapper J A Davis P J Lamothe C Patton D Epel 《The Journal of cell biology》1985,100(6):1817-1824
When sperm of Strongylocentrotus purpuratus or Lytechinus pictus are diluted into seawater, motility is initiated; and when exposed to egg jelly, an acrosome reaction is induced. In the presence of a variety of structurally different metal chelators (0.1-1 mM EDTA, EGTA, phenanthroline, dipyridyl, cysteine, or dithiothreitol), motility initiation is delayed and the acrosome reaction is inhibited. Of the metals detected in the sperm of these two species, very low levels of Zn+2 (0.1 microM free Zn+2) uniquely prevent this chelator inhibition. L. pictus sperm concentrate 65Zn+2 from seawater, and EDTA removes 50% of the accumulated 65Zn+2 by 5 min. Since both sperm motility and acrosome reactions are in part regulated by intracellular pH (pHi), the effect of chelators on the sperm pHi was examined by using the fluorescent pH sensitive probe, 9-aminoacridine, EDTA depresses sperm pHi in both species, and 0.1 microM free Zn+2 reverses this pHi depression. When sperm are diluted into media that contain chelators, both NH4Cl and monensin (a Na+/H+ ionophore) increase the sperm pHi and reverse the chelator inhibition of sperm motility and acrosome reactions. The results of this study are consistent with the involvement of a trace metal (probably zinc) in the pHi regulation of sea urchin sperm and indicate a likely mechanism for the previously observed effects of chelators on sperm motility and acrosome reactions. 相似文献
15.
Nitric oxide interacts with the cAMP pathway to modulate capacitation of human spermatozoa 总被引:7,自引:0,他引:7
Belén Herrero M Chatterjee S Lefièvre L de Lamirande E Gagnon C 《Free radical biology & medicine》2000,29(6):522-536
This study aimed to demonstrate nitric oxide production by human spermatozoa and to characterize the interaction between nitric oxide and cAMP-related pathway in the control of human sperm capacitation and protein tyrosine phosphorylation. Spermatozoa were incubated in Tyrode's medium with or without bovine serum albumin (BSA), and nitric oxide was measured with the spin trap sodium N-methyl-D-glucamine dithiocarbamate. Under noncapacitating conditions, spermatozoa produced low levels of nitric oxide. However, under capacitating conditions, prominent nitric oxide adduct signals were obtained and a time-dependent increase of nitric oxide production was observed. When spermatozoa were incubated in Tyrode+BSA medium with nitric oxide-releasing compounds, intracellular cAMP concentrations increased to levels higher than those of spermatozoa incubated in Tyrode+BSA alone. In contrast, incubation with nitric oxide synthase inhibitors (N(G)-nitro-L-arginine methyl ester or N(G)-monomethyl L-arginine) decreased intracellular sperm cAMP concentrations. The inhibitory effect observed with N(G)-nitro-L-arginine methyl ester on capacitation and tyrosine phosphorylation of two sperm proteins (105, 81 kDa) was overcome by the presence of cAMP analogs or of a phosphodiesterase inhibitor. These results indicate that nitric oxide is produced by capacitating human spermatozoa and that it may act as a cellular messenger by modulating the cAMP pathway involved in capacitation and protein tyrosine phosphorylation. 相似文献
16.
Goat sperm lysate from cauda epididymis was incubated in the presence of [14C]phosphatidyl-choline, -ethanolamine, -inositol and diphosphatidyl-glycerol. The release of [14C]diacylglycerol from only phosphatidyl inositol confirmed the presence of phosphatidyl inositol specific phospholipase C. The enzyme activity was linear up to 1 hr of incubation at a sperm concentration of 1-10 x 10(9). It had pH optimum of 7.5 in a broad range of pH activity profile (pH 6-9). Maximum activity was observed at 8 mM calcium ion concentration. EDTA and EGTA (5 mM) did not inhibit the activity completely. A comparative study on spermatozoa and fluid from caput, corpus and cauda epididymis revealed 6.5-fold increase of activity in spermatozoa and a 4-fold decrease in case of fluid during the epididymal transit. However, the total protein content remained unchanged in fluid and decreased up to the extent of 2.4-fold in spermatozoa during this process. Thirty five percent of the caudal sperm activity was soluble and the rest was associated with head (44%), mid-piece (10%) and tail (10%). 相似文献
17.
The location of the inducible and constitutive forms of tyrosine sulfate sulfohydrolase in Comamonas terrigena was investigated by subjecting resting cells to osmotic shock and to treatment with lysozyme in the presence of EDTA. The bulk of this enzyme present in induced cells was released by these procedures, suggesting that the induced form is cell wall associated. The constitutive form present in noninduced cells was not released under these conditions. Evidence was also presented which suggests that SO4(2-)release by intact cells during exposure to tyrosine sulfate was primarily due to the action of the inducible form of the enzyme. 相似文献
18.
Gil PI Guidobaldi HA Teves ME Uñates DR Sanchez R Giojalas LC 《Animal reproduction science》2008,108(1-2):236-246
The aim of this study was to verify whether cattle spermatozoa respond by chemotaxis to follicular fluid (FF). The experimental conditions were defined to maintain a frozen-thawed sperm population with great motility and capacitation, and lesser sperm agglutination. Several sperm preparation conditions were studied: sperm separation from the seminal plasma by Sephadex column or migration-sedimentation, incubation under capacitating conditions in the presence or absence of a superficial layer of mineral oil, and different pH of the culture medium. The percentage of motile and agglutinated spermatozoa was determined in plate dishes under inverted phase contrast microscope. The percentage of capacitated spermatozoa was calculated as the difference between the percentages of acrosome reacted spermatozoa with and without lysophosphatidylcholine stimulation. The most ideal experimental conditions to evaluate chemotaxis in frozen-thawed cattle spermatozoa were: to separate the cells from the seminal plasma by migration-sedimentation and to incubate them under oil, in culture medium at pH 7.2, for less than 2h. The chemotaxis assays were conducted with spermatozoa treated as mentioned above which were confronted to several dilutions of FF (1:10(3), 1:10(4), 1:10(5), 1:10(6)) in a chemotaxis chamber by videomicroscopy and computer image analysis. A subpopulation of capacitated spermatozoa ( approximately 10%) that responded chemotactically to a concentration gradient generated by FF (1:10(4) to 1:10(5)) was observed. Since cryopreserved spermatozoa are regularly used to artificially inseminate the cows, the sperm chemotactic response towards FF would be potentially used to diagnose the bull sperm sample or to select the spermatozoa in the most functional state. 相似文献
19.
Osycka-Salut C Gervasi MG Pereyra E Cella M Ribeiro ML Franchi AM Perez-Martinez S 《PloS one》2012,7(2):e30671
Mammalian spermatozoa are not able to fertilize an egg immediately upon ejaculation. They acquire this ability during their transit through the female genital tract in a process known as capacitation. The mammalian oviduct acts as a functional sperm reservoir providing a suitable environment that allows the maintenance of sperm fertilization competence until ovulation occurs. After ovulation, spermatozoa are gradually released from the oviductal reservoir in the caudal isthmus and ascend to the site of fertilization. Capacitating-related changes in sperm plasma membrane seem to be responsible for sperm release from oviductal epithelium. Anandamide is a lipid mediator that participates in the regulation of several female and male reproductive functions. Previously we have demonstrated that anandamide was capable to release spermatozoa from oviductal epithelia by induction of sperm capacitation in bovines. In the present work we studied whether anandamide might exert its effect by activating the nitric oxide (NO) pathway since this molecule has been described as a capacitating agent in spermatozoa from different species. First, we demonstrated that 1 μM NOC-18, a NO donor, and 10 mM L-Arginine, NO synthase substrate, induced the release of spermatozoa from the oviductal epithelia. Then, we observed that the anandamide effect on sperm oviduct interaction was reversed by the addition of 1 μM L-NAME, a NO synthase inhibitor, or 30 μg/ml Hemoglobin, a NO scavenger. We also demonstrated that the induction of bull sperm capacitation by nanomolar concentrations of R(+)-methanandamide or anandamide was inhibited by adding L-NAME or Hemoglobin. To study whether anandamide is able to produce NO, we measured this compound in both sperm and oviductal cells. We observed that anandamide increased the levels of NO in spermatozoa, but not in oviductal cells. These findings suggest that anandamide regulates the sperm release from oviductal epithelia probably by activating the NO pathway during sperm capacitation. 相似文献
20.
Ejaculated boar spermatozoa are vulnerable to cold shock. Prolonged storage of boar spermatozoa at low temperatures reduces survival rate, resulting in a bottleneck for the extension of artificial insemination in pig husbandry. This study evaluated whether alginate microencapsulization processing can improve the longevity of boar spermatozoa stored at 5 degrees C and the fertility of microencapsulated spermatozoa in vivo. Sperm-rich fraction semen from three purebred boars were concentrated and microencapsulated using alginate at 16-18 degrees C, and then were stored at 5 degrees C. Following storage for 1, 3 and 7 days, the microcapsule was taken out to assess sperm release under 37 degrees C incubation with or without 110 rpm stirring. The percentage of sperm released from microcapsules with 110 rpm stirring was higher than without stirring (81 versus 60%) after 24h of incubation. In another experiment, semen was also microencapsulated to evaluate the sperm motility. The motility of spermatozoa was assessed at 10 min, 8, 24, 32, 48, 56 and 72 h following incubation at 37 degrees C for nine consecutive days. The fertility of the free and microencapsulated semen was assessed by inseminating sows, and the reproductive traits (conception rate, farrowing rate, and litter size) were recorded. The motility of encapsulated spermatozoa was significantly higher than that of free semen after 8h incubation at 37 degrees C after storing for over three days (P<0.05). No significant difference existed in conception rate, farrowing rate, and litter size between the microencapsulated and non-encapsulated semen after four days of storage. In conclusion, microencapsulation can increase the longevity of boar spermatozoa and may sustain in vivo ova fertilization ability. 相似文献