Mammalian sperm must undergo a series of biochemical and physiological modifications, collectively called capacitation, in the female reproductive tract prior to the acrosome reaction （AR）. The mechanisms of these m...Mammalian sperm must undergo a series of biochemical and physiological modifications, collectively called capacitation, in the female reproductive tract prior to the acrosome reaction （AR）. The mechanisms of these modifications are not well characterized though protein kinases were shown to be involved in the regulation of intracellular Ca2＋ during both capacitation and the AR. In the present review, we summarize some of the signaling events that are involved in capacitation. During the capacitation process, phosphatidyl-inositol-3-kinase （PI3K） is phosphorylated/activated via a protein kinase A （PKA）-dependent cascade, and downregulated by protein kinase C a （PKCa）. PKCa is active at the beginning of capacitation, resulting in PI3K inactivation. During capacitation, PKCa as well as PP172 is degraded by a PKA-dependent mechanism, allowing the activation of PI3K. The activation of PKA during capacitation depends mainly on cyclic adenosine monophosphate （cAMP） produced by the bicarbonate-dependent soluble adenylyl cyclase. This activation of PKA leads to an increase in actin polymerization, an essential process for the development of hyperactivated motility, which is necessary for successful fertilization. Actin polymerization is mediated by PIP2 in two ways： first, PIP2 acts as a cofactor for phospholipase D （PLD） activation, and second, as a molecule that binds and inhibits actin-severing proteins such as gelsolin. Tyrosine phosphorylation of gelsolin during capacitation by Src family kinase （SFK） is also important for its inactivation. Prior to the AR, gelsolin is released from PIP2 and undergoes dephosphorylation/activation, resulting in fast F-actin depolymerization, leading to the AR.展开更多
Aim： To investigate the in vitro effects of insulin and leptin on human sperm motility, viability, acrosome reaction and nitric oxide （NO） production. Methods： Washed human spermatozoa from normozoospermic donors ...Aim： To investigate the in vitro effects of insulin and leptin on human sperm motility, viability, acrosome reaction and nitric oxide （NO） production. Methods： Washed human spermatozoa from normozoospermic donors were treated with insulin （10 μIU） and leptin （10 nmol）. Insulin and leptin effects were blocked by inhibition of their intracellular effector, phosphotidylinositol 3-kinase （PI3K）, by wortmannin （10 μmol） 30 min prior to insulin and leptin being given. Computer-assisted semen analysis was used to assess motility after 1, 2 and 3 h of incubation. Viability was assessed by fluorescence-activated cell sorting using propidium iodide as a fluorescent probe. Acrosome-reacted cells were observed under a fluorescent microscope using fluorescein-isothiocyanate-Pisum sativum agglutinin as a probe. NO was measured after treating the sperm with 4,5-diaminofluorescein-2/diacetate （DAF-2/DA） and analyzed by fluorescence-activated cell sorting. Results： Insulin and leptin significantly increased total motility, progressive motility and acrosome reaction, as well as NO production. Conclusion： This study showed the in vitro beneficial effects of insulin and leptin on human sperm function. These hormones could play a role in enhancing the fertilization capacity of human spermatozoa.展开更多
We developed tests of sperm-oocyte interaction:sperm-zona binding,zona-induced acrosome reaction,sperm-zona penetration and sperm-oolemma binding,using oocytes which failed to fertilise in clinical in vitro fertilizat...We developed tests of sperm-oocyte interaction:sperm-zona binding,zona-induced acrosome reaction,sperm-zona penetration and sperm-oolemma binding,using oocytes which failed to fertilise in clinical in vitro fertilization(IVF).Although oocyte defects contribute to failure of sperm oocyte interaction,rarely are all oocytes from one wom-an affected.Low or zero fertilization in standard IVFwas usually caused by sperm abnormalities.Poor sperm-zona pel-lucida binding was frequently associated with failure of standard IVF and obvious defects of sperm motility or morpholo-gy.The size and shape of the acrosome is particularly important for sperm binding to the oocyte.The proportion ofacrosome intact sperm in the insemination medium was related to the IVF rote.Inducing the acrosome reaction with acalcium ionophore reduced sperm-zona binding.Blocking acrosome dispersal with an acrosin inhibitor prevented sperm-zona penetration.Sperm-zona penetration was even more highly related to IVF rates than was sperm-zona binding.Some patients had low or zero fertilization rates with standard IVF but normal sperm by conventional tests and normalsperm-zona binding.Few of their sperm underwent the acrosome reaction on the surface of the zona and none penetrat-ed the zona.In contrast,fertilization and pregnancy rates were high with intmcytoplasmic sperm injection.We call thiscondition defective zona pellucida induced acrosome reaction.Discovery of the nature of the abnormalities in the signaltmnsduction and effector pathways of the human zona pellucida induced acrosome reaction should result in simpler testsand treatments for the patients and also provide new leads for contraceptive development.展开更多
During mammalian fertilisation, the zona pellucida （ZP） matrix surrounding the oocyte is responsible for the binding of the spermatozoa to the oocyte and induction of the acrosome reaction （AR） in the ZP-bound spe...During mammalian fertilisation, the zona pellucida （ZP） matrix surrounding the oocyte is responsible for the binding of the spermatozoa to the oocyte and induction of the acrosome reaction （AR） in the ZP-bound spermatozoon. The AR is crucial for the penetration of the ZP matrix by spermatozoa. The ZP matrix in mice is composed of three glycoproteins designated ZP1, ZP2 and ZP3, whereas in humans, it is composed of four （ZP1, ZP2, ZP3 and ZP4）. ZP3 acts as the putative primary sperm receptor and is responsible for AR induction in mice, whereas in humans （in addition to ZP3）, ZP1 and ZP4 also induce the AR. The ability of ZP3 to induce the AR resides in its C-terminal fragment. O-linked glycans are critical for the murine ZP3-mediated AR. However, N-linked glycans of human ZP1, ZP3 and ZP4 have important roles in the induction of the AR. Studies with pharmacological inhibitors showed that the ZP3-induced AR involves the activation of the Gi-coupled receptor pathway, whereas ZP1- and ZP4-mediated ARs are independent of this pathway. The ZP3-induced AR involves the activation of T-type voltage-operated calcium channels （VOCCs）, whereas ZP1- and ZP4-induced ARs involve both T- and L-type VOCCs. To conclude, in mice, ZP3 is primarily responsible for the binding of capacitated spermatozoa to the ZP matrix and induction of the AR, whereas in humans （in addition to ZP3）, ZP1 and ZP4 also participate in these stages of fertilisation.展开更多
To bind and fertilize the egg, the spermatozoon should undergo few biochemical and motility changes in the female reproductive tract collectively called capacitation. The capacitated spermatozoon binds to the egg zona...To bind and fertilize the egg, the spermatozoon should undergo few biochemical and motility changes in the female reproductive tract collectively called capacitation. The capacitated spermatozoon binds to the egg zona pellucida, and then undergoes the acmsome reaction （AR）, which allows its penetration into the egg. The mechanisms regulating sperm capacitation and the AR are not completely understood. In the present review, we summarize some data regarding the role and regulation of the epidermal growth factor receptor （EGFR） in these processes. In the capacitation process, the EGFR is partially activated by protein kinase A （PKA）, resulting in phospholipase D （PLD） activation and actin polymerization. Protein kinase C alpha （PKCα）, which is already activated at the beginning of the capacitation, also participates in PLD activation. Further activation of the EGFR at the end of the capacitation enhances intracellular Ca2＋ concentration leading to F-actin breakdown and allows the AR to take place. Under in vivoconditions, the EGFR can be directly activated by its known ligand epidermal growth factor （EGF）, and indirectly by activating PKA or by transactivation mediated by G protein-coupled receptors （GPCRs） activation or by ouabain. Under physiological conditions, sperm PKA is activated mainly by bicarbonate, which activates the soluble adenylyl cyclase to produce cyclic adenosine monophosphate （cAMP）, the activator of PKA. The GPCR activators angiotensin II or Ivsoohosphatidic acid, as well as ouabain and EGF are phvsioloeical comoonents oresent in the female reoroductive tract.展开更多
Egg water was used to induce acrosome reaction in mature sperm of Fenneropenaeus chinensis .Transmission electron microscope and SDS PAGE were used to study the ultrastructure and protein changes of the sperm in ...Egg water was used to induce acrosome reaction in mature sperm of Fenneropenaeus chinensis .Transmission electron microscope and SDS PAGE were used to study the ultrastructure and protein changes of the sperm in F.chinensis ,which has undergone acrosome reaction.The results demonstrated that the sperms from female thelycum could be induced acrosome reaction in vitro by egg water,which was a kind of egg jelly released from oocyte into seawater during oocyte activation.More than fifty percent of sperm finished acrosome reaction during the first 30 min, in vitro .The whole event consists of the retraction of the spike and acrosome exocytosis.Spike was retracted and fused to acrosome cap followed by the release of acrosome granules.When the original egg water was diluted 5 and 10 times,The diluted egg water also have the ability to induce sperm acrosome reaction,but the acrosome reaction rate is much lower than that of the original egg water in the same time.Capillary zone electrophoresis was used to detect the biochemical components of egg water.Egg water was composed of two different components,which were also demonstrated by SDS PAGE.The molecular weights of the two kinds of egg jellies are both about 200kDa.Gelatin substrate SDS PAGE results did not show any hydrolytic enzyme activity in egg water.After acrosome reaction,many sorts of proteins in sperm are degraded.When the reacted sperms were examined with gelatin substrate SDS PAGE,there were six major peptide bands with hydrolytic enzyme activity were detected.Their molecular weights are 200kDa,130 kDa,66 kDa,53 kDa,48 kDa and 41 kDa respectively.These hydrolases cannot be detected in the sperms before acrosome reaction.The acrosome reaction of Chinese shrimp, F.chinensis is much different to that of the sperm in the shrimp, Sicyonia ingentis ,which the acrosome reaction was clearly studied.There is not filament formation during the acrosome reaction in Chinese shrimp.The time of exocytosis in the sperm of Chinese shrimp is also much longer than that of the sperm of Sicyonia ingentis .展开更多
The fertilization process is the net result of a complex sequence of events that collectively result in the fusion of theopposite gametes. The male gamete undergoes continuous morphological and biochemical modificatio...The fertilization process is the net result of a complex sequence of events that collectively result in the fusion of theopposite gametes. The male gamete undergoes continuous morphological and biochemical modifications during spermdevelopment in the testis (spermatogenesis), maturation in the epididymis, and capacitation in the female reproductivetract. Only the capacitated spermatozoa are able to recognize and bind to the bioactive glycan residue(s) on the ovum'sextracellular coat, the zona pellucida (ZP). Sperm-zona binding in the mouse and several other species is believed totake place in two stages. First, capacitated (acrosome-intact) spermatozoa loosely and reversibly adhere to the zona-in-tact ovum. In the second stage tight irreversible binding occurs. Both types of bindings are attributed to the presence ofglycan- binding proteins (receptors) on the sperm plasma membrane and their complementary bioactive glycan units(ligands) on the surface of the ZP. The carbohydrate-mediated adhesion event initiates a signal transduction cascade re-sulting in the exocytosis of acrosomal contents. This step is believed to be prerequisite which allows the hyperactivatedacrosome-reacted spermatozoa to penetrate the ZP and fertilize the ovum. This review focuses on the role of carbohy-drate residues in sperm-ovum interaction, and triggering of the acrosome reaction. I have attempted to discuss extensiveprogress that has been made to enhance our understanding of the well programmed multiple molecular events necessaryfor successful fertilization. This review will identify these events, and discuss the functional significance of carbohy-drates in these events.展开更多
Aim： To investigate the possible functions of human sperm membrane protein （hSMP-1） in the process of fertilization. Methods： A 576-bp cDNA fragment of HSD-1 gene coding for the extracellular domain of hSMP-1 was ...Aim： To investigate the possible functions of human sperm membrane protein （hSMP-1） in the process of fertilization. Methods： A 576-bp cDNA fragment of HSD-1 gene coding for the extracellular domain of hSMP-1 was cloned and expressed. The localization of this protein on human and mouse sperm was determined by indirect immunofluorescent staining by using anti-recombinant hSMP-1 （anti-rhSMP-1） antibodies. Sperm acrosome reaction and spermzona pellucida （ZP） binding assay were carried out in 10-week-old BALB/c mice. Results： Recombinant hSMP-1 was successfully cloned and expressed. The expression of the native protein was limited on the acrosome of human and mouse sperm. Treatment of anti-rhSMP-1 antibodies significantly decreased the average number of sperms bound to each egg. Meanwhile, the percentage of acrosome reaction was decreased in comparison to pre-immune control after treatment with anti-rhSMP-1 （P 〈 0.05）. Conclusion： The results suggest that anti-rhSMP-1 antibody inhibited mouse acrosome reaction and sperm-ZP binding.展开更多
Sterols are essential components of the cell membrane lipid bilayer that include molecules such as cholesterol and desmosterol, which are significantly found in the spermatozoa of several animal species. However, the ...Sterols are essential components of the cell membrane lipid bilayer that include molecules such as cholesterol and desmosterol, which are significantly found in the spermatozoa of several animal species. However, the presence of desmosterol in rabbit semen has never been investigated. The aims of this study were to characterize the sterol composition of subfractions of ejaculated rabbit semen and evaluate the in vitro effects of sterol on the spermatozoa acrosome reaction and motility. Two sterols, occurring prevalently in the free form （94.3%）, were identified in whole semen collected from 10 fertile New Zealand White rabbits, specifically desmosterol （58.5% of total sterols） and cholesterol （35.9% of total sterols）. Desmosterol was the predominant sterol found in all subfractions of rabbit semen, varying from 56.7% （in the prostatic secretory granules, PSGs） to 63.8% （in the seminal plasma）. Spermatozoa contained an intermediate proportion of desmosterol （59.8%）, which was asymmetrically distributed between the heads （52.0% of the total content of sterols） and the tails （81.8%）. Results showed that both desmosterol and cholesterol can be transferred from the PSGs to the spermatozoa and are equally effective in inhibiting in vitro spermatozoa capacitation at a concentration higher than 1 mg L^-1. In contrast, neither desmosterol nor cholesterol had a significant effect on spermatozoa motility. Thus, it was concluded that, the various fractions of rabbit seminal fluid differ from each other in sterol composition and quantity, probably due to their different functional properties, and these fractions may undergo significant sterol changes depending on the stage of spermatozoa capacitation.展开更多
Aim： To investigate if the phytoestrogen, genistein, affects essential functions of cryopreserved bovine spermatozoa. Methods： The effect of genistein upon motility was assessed by computer-assisted motion analysis....Aim： To investigate if the phytoestrogen, genistein, affects essential functions of cryopreserved bovine spermatozoa. Methods： The effect of genistein upon motility was assessed by computer-assisted motion analysis. Hemizona assay was performed to detect the ability of spermatozoa binding to the zona pellucida. The inducibility of the acrosome reaction using progesterone and ZP3-6 peptide was analysed by fluorescein-conjugated Pisum sativum agglutinin （FITC-PSA）/Hoechst 33258 double staining. Capacitation after incubation with genistein was assessed by the chlortetracycline （CTC） assay. Immunoblots showed the pattern of protein tyrosine phosphorylation of cryopreserved bovine spermatozoa. Results： Immunodetection of tyrosine-phosphorylated proteins showed that genistein did not affect tyrosine phosphorylation in cryopreserved bovine spermatozoa. However, genistein significantly reduced the progesterone- and ZP3-6 peptide-mediated induction of the acrosome reaction and led to a dose-dependent inhibition of sperm-zona pellucida binding; while sperm motility and capacitation were not affected by this phytoestrogen, as indicated by computer-assisted sperm motion analysis and the CTC assay, respectively. Conclusion： Our results suggest that in cryopreserved bovine spermatozoa, genistein affects a protein tyrosine phosphorylation-independent signal transduction pathway that is involved in sperm capacitation, the acrosome reaction and sperm-zona pellucida binding.展开更多
The acrosome reaction （AR）, an absolute requirement for spermatozoa and egg fusion, requires the influx of Ca2＋ into the spermatozoa through voltage-dependent Ca2＋ channels and store-operated channels. Maitotoxin ...The acrosome reaction （AR）, an absolute requirement for spermatozoa and egg fusion, requires the influx of Ca2＋ into the spermatozoa through voltage-dependent Ca2＋ channels and store-operated channels. Maitotoxin （MTx）, a Ca2＋-mobilizing agent, has been shown to be a potent inducer of the mouse sperm AR, with a pharmacology similar to that of the zona pellucida （ZP）, possibly suggesting a common pathway for both inducers. Using recombinant human ZP3 （rhZP3）, mouse ZP and two MTx channel blockers （U73122 and U73343）, we investigated and compared the MTx- and ZP-induced ARs in human and mouse spermatozoa. Herein, we report that MTx induced AR and elevated intracellular Ca2＋ （[Ca2＋]~） in human spermatozoa, both of which were blocked by U73122 and U73343. These two compounds also inhibited the MTx-induced AR in mouse spermatozoa. In disagreement with our previous proposal, the AR triggered by rhZP3 or mouse ZP was not blocked by U73343, indicating that in human and mouse spermatozoa, the AR induction by the physiological ligands or by MTx occurred through distinct pathways. U73122, but not U73343 （inactive analogue）, can block phospholipase C （PLC）. Another PLC inhibitor, edelfosine, also blocked the rhZP3- and ZP-induced ARs. These findings confirmed the participation of a PLC-dependent signalling pathway in human and mouse zona protein-induced AR. Notably, edelfosine also inhibited the MTx-induced mouse sperm AR but not that of the human, suggesting that toxin-induced AR is PLC-dependent in mice and PLC-independent in humans.展开更多
Capacitation and acrosome reaction are important prerequisites of the fertilization process. Capacitation is a highlycomplex phenomenon occurring in the female genital tract, rendering the spermatozoa capable of bindi...Capacitation and acrosome reaction are important prerequisites of the fertilization process. Capacitation is a highlycomplex phenomenon occurring in the female genital tract, rendering the spermatozoa capable of binding and fusionwith the oocyte. During capacitation various biochemical and biophysical changes occur in the spermatozoa and thespermatozoal membranes. Ions and ion channels also play important roles in governing the process of capacitation bychanging the fluxes of different ions which in turn controls various characteristics of capacitated spermatozoa. Alongwith the mobilization of ions the generation of free radicals and efflux of cholesterol also plays an important role in thecapacitation state of the spermatozoa. The generation of free radical and efflux of cholesterol change the mechano-dynamic properties of the membrane by oxidation of the polyunsaturated lipids and by generating the cholesterol freepatches. The process of capacitation renders the spermatozoa responsive to the inducers of the acrosome reaction. Theglycoprotein zona pellucida 3 (ZP3) of the egg coat zona pellucida is the potent physiological stimulator of the acro-some reaction; progesterone, a major compoent of the follicular fluid, is also an induce of the acrosome reaction.The inducers of the acrosome reaction cause the activation of the various ion-channels leading to high influxes of calci-um, sodium and bicarbonate. The efflux of cholesterol during the process of capacitation alters the permeablity of themembrane to the ions and generate areas which are prone to fusion and vesculation process during the acrosome reac-tion. Ths review focuses mainly on effects of the ion and ion-channels, free radicals, and membrane fluidity changesduring the process of capacitation and acrosome reaction. (Asian J Androl 1999 Sep; 1: 95-107)展开更多
Objective: To explore the effect of NO on the spontaneous acrosome reaction in antisperm antibody (AsAb) positive rat spermatozoa. Methods: The rat model of AsAb was set up by artificial immunization. The level of AsA...Objective: To explore the effect of NO on the spontaneous acrosome reaction in antisperm antibody (AsAb) positive rat spermatozoa. Methods: The rat model of AsAb was set up by artificial immunization. The level of AsAb in blood serum was determined by TAT and ELISA. Rat spermatozoa was visualized by staining the acrosome with Coomassie brilliant blue. The NO concentration in rat spermatozoa was assayed by HPLC. Results: The percentage of acrosome reaction, NO concentration, superoxide dismutase (SOD) and Na^+-K^+ATPase activity in AsAb positive rat spermatozoa were significantly decreased compared with the control group. Low dose of NO (SNP 10^(-9)～10^(-8) mol/L) increased the percentage of acrosome reaction and SOD activity, but had no effect on Na^+-K^+ATPase activity. High dose of NO (SNP 10^(-6)～10^(-4) mol/L) decreased the three items. Conclusion: The decrease in acrosome reaction in positive AsAb rat spermatozoa might be related to a decrease in NO and increase in O_2. (the SOD activity was decreased) in sperm. A low dose of NO might increase the percentage of acrosome reaction in AsAb positive rat by inhibiting the superoxide, while a high dose of NO was harmful to sperm function. [Reprod Contracep (in Chinese) 2002; 22: 203]展开更多
Aim: To study the roles of tumor necrosis factor alpha (TNF-α) on the sperm acrosin activity and acrosome reaction. Methods: The sperm acrosin activity was tested by the method of BAEE/ ADH Unity and the acrosome rea...Aim: To study the roles of tumor necrosis factor alpha (TNF-α) on the sperm acrosin activity and acrosome reaction. Methods: The sperm acrosin activity was tested by the method of BAEE/ ADH Unity and the acrosome reaction by the Triple-stain technique. Results: TNF-α decreased the sperm acrosin activity and acrosome reaction (P<0.01; P<0.01, respectively); it also inhibited the Ca2+-ATPase activity and SOD activity in sperm (P< 0.05; P<0.001, respectively), but increased the NOS activity and the amount of NO in sperm (P<0.001; P<0.001, respectively). While it had a less significant effect on the activity of Na+-K+-ATPase (P>0.05). Conclusion: TNF-α inhibits the sperm acrosin activity and acrosome reaction.展开更多
The acrosome reaction of sperm was induced by calcium ionophore A 23187.The letionship between some ions and acrosome reaction by removing Na+ from the medium.or by adding angtagonist of K+.TEA chloride,or antagonist ...The acrosome reaction of sperm was induced by calcium ionophore A 23187.The letionship between some ions and acrosome reaction by removing Na+ from the medium.or by adding angtagonist of K+.TEA chloride,or antagonist of Ca++,verapamil,or anlagonist of Na+-K+-ATPase,acetyl strophanthithidin is studied.The results show that Na+,H,Ca++ and Na+ pump are necessary.for acrosome reaclion of human sperm.The Ca++ might not enter the sperms through the channel of Ca++.展开更多
No Con A receptor site was found on the intact plasma membranes of mammalian sperms before capacitation.After capacitation, however, the plasma membranes sloughed off, exposing the outer acrosomal membrane which is ri...No Con A receptor site was found on the intact plasma membranes of mammalian sperms before capacitation.After capacitation, however, the plasma membranes sloughed off, exposing the outer acrosomal membrane which is rich in Con A receptor sites.The vesicles formed during acrosome reaction were also found to bc rich in Con A receptor sites, suggesting their origin from outer acrosomal membranes.With completion of acrosome reaction, only inner acrosomal membrane was left in which no Con A receptor sites could be demonstrated.Also no Con A receptor site was found on egg plasma membrane throughout fertilization.It was thus shown that different membranes possess different properties.展开更多
The biological characteristics of crustacean spermatozoa is important for the artificial reproduction and genetic breeding. With reference to the latest studies and related materials, this paper reviewed the research ...The biological characteristics of crustacean spermatozoa is important for the artificial reproduction and genetic breeding. With reference to the latest studies and related materials, this paper reviewed the research progress in the biological characteristics of crustacean spermatozoa, such as morphological structure of sperm, spermatogenesis, sperm viability, preservation in vitro and acrosome reaction et al. The prospects of the research field have also been anticipated.展开更多
A membrane protein was isolated from mouse sperm heads that had undergone acrosomal reaction induced by C2+ ionophore, A 23187, which, with a molecular weight of 77.6 kd, shows capability to block egg-sperm fusion. As...A membrane protein was isolated from mouse sperm heads that had undergone acrosomal reaction induced by C2+ ionophore, A 23187, which, with a molecular weight of 77.6 kd, shows capability to block egg-sperm fusion. As revealed by analysis usintg isotopic markers, this protein is one of the chief membrane proteins of inner acrosomal membrane or the outer membrane of equatorial segment and Post-acrosomal region; treatment of mouse sperms with 0.6 μg/ml of the Purified protein for 30 minutes reduced the sperm-egg fusion index by 51%.The above results led us to the conclusion that the protein is an active participant in sperm-egg fusion. The possible existence of sperm receptor on egg plasma membrane was discussed.展开更多
Aim: To produce biologically active recombinant human (rh) ZP proteins in a human cell for use in sperm function tests. Methods: The human embryonic kidney cell line 293T was employed to produce rhZP1, rhZP2 and rhZP3...Aim: To produce biologically active recombinant human (rh) ZP proteins in a human cell for use in sperm function tests. Methods: The human embryonic kidney cell line 293T was employed to produce rhZP1, rhZP2 and rhZP3 proteins individually and together by co-expression. Presence of these proteins in the culture medium and cell lysate was assessed by Western blotting analysis. The effect of the recombinant proteins on the human AR was assessed. Results: RhZP2 and rhZP3 were secreted into the culture medium, whereas rhZPl was found only in the cell lysate. Interestingly, when all zona pellucida proteins were co-expressed in the same cells, rhZPl was also secreted into the culture medium. However, despite the presence of all three ZP proteins in sufficient concentration and evidence of heavy glycosylation on gel electrophoresis, biological activity to induce the AR was not observed. Conclusion: RhZP1, rhZP2 and rhZP3 were successfully expressed in the human embryonic kidney cell line 293T. It appears that an interaction amongst these proteins may be required for release of rhZPl from the cell. Although this approach is not satisfactory for producing active human ZP proteins, it makes a significant contribution to the understanding of the structural and functional characteristics of the ZP proteins.展开更多
Prior to fertilization sperm has to undergo an activation process known as capaciation,leading to the acrosome reaction.Till now,little is known about the mechanism for preventing premature capacitation in sperm altho...Prior to fertilization sperm has to undergo an activation process known as capaciation,leading to the acrosome reaction.Till now,little is known about the mechanism for preventing premature capacitation in sperm although decapacitation factors from various sources have been thought to be involved.In this study,we report that NYD-SP27,an isoform of phospholipase C Zeta 1(PLCZ1),is localized to the sperm acrosome in mouse and human spermatozoa by immunofluorescence using a specific antibody.Western blot and double staining analyses show NYD-SP27 becomes detached from sperm,as they undergo capacitation and acrosome reaction.The absence of HCO_(3)^(-),a key factor in activating capacitation,from the capacitation-inducing medium prevents the loss of NYD-SP27 from sperm.The anti-NYD-SP27 antibody also prevents the loss of NYD-SP27 from sperm,reduced the number of capacitated sperm,inhibited the acrosome reaction induced by ATP and progesterone,and inhibited agonist-induced PLC-coupled Ca^(2+)mobilization in sperm,which can be mimicked by the PLC inhibitor,U73122.These data strongly suggest that NYD-SP27 is a physiological inhibitor of PLC that acts as an intrinsic decapacitation factor in sperm to prevent premature capacitation and acrosome reaction.展开更多
文摘Mammalian sperm must undergo a series of biochemical and physiological modifications, collectively called capacitation, in the female reproductive tract prior to the acrosome reaction （AR）. The mechanisms of these modifications are not well characterized though protein kinases were shown to be involved in the regulation of intracellular Ca2＋ during both capacitation and the AR. In the present review, we summarize some of the signaling events that are involved in capacitation. During the capacitation process, phosphatidyl-inositol-3-kinase （PI3K） is phosphorylated/activated via a protein kinase A （PKA）-dependent cascade, and downregulated by protein kinase C a （PKCa）. PKCa is active at the beginning of capacitation, resulting in PI3K inactivation. During capacitation, PKCa as well as PP172 is degraded by a PKA-dependent mechanism, allowing the activation of PI3K. The activation of PKA during capacitation depends mainly on cyclic adenosine monophosphate （cAMP） produced by the bicarbonate-dependent soluble adenylyl cyclase. This activation of PKA leads to an increase in actin polymerization, an essential process for the development of hyperactivated motility, which is necessary for successful fertilization. Actin polymerization is mediated by PIP2 in two ways： first, PIP2 acts as a cofactor for phospholipase D （PLD） activation, and second, as a molecule that binds and inhibits actin-severing proteins such as gelsolin. Tyrosine phosphorylation of gelsolin during capacitation by Src family kinase （SFK） is also important for its inactivation. Prior to the AR, gelsolin is released from PIP2 and undergoes dephosphorylation/activation, resulting in fast F-actin depolymerization, leading to the AR.
文摘Aim： To investigate the in vitro effects of insulin and leptin on human sperm motility, viability, acrosome reaction and nitric oxide （NO） production. Methods： Washed human spermatozoa from normozoospermic donors were treated with insulin （10 μIU） and leptin （10 nmol）. Insulin and leptin effects were blocked by inhibition of their intracellular effector, phosphotidylinositol 3-kinase （PI3K）, by wortmannin （10 μmol） 30 min prior to insulin and leptin being given. Computer-assisted semen analysis was used to assess motility after 1, 2 and 3 h of incubation. Viability was assessed by fluorescence-activated cell sorting using propidium iodide as a fluorescent probe. Acrosome-reacted cells were observed under a fluorescent microscope using fluorescein-isothiocyanate-Pisum sativum agglutinin as a probe. NO was measured after treating the sperm with 4,5-diaminofluorescein-2/diacetate （DAF-2/DA） and analyzed by fluorescence-activated cell sorting. Results： Insulin and leptin significantly increased total motility, progressive motility and acrosome reaction, as well as NO production. Conclusion： This study showed the in vitro beneficial effects of insulin and leptin on human sperm function. These hormones could play a role in enhancing the fertilization capacity of human spermatozoa.
文摘We developed tests of sperm-oocyte interaction:sperm-zona binding,zona-induced acrosome reaction,sperm-zona penetration and sperm-oolemma binding,using oocytes which failed to fertilise in clinical in vitro fertilization(IVF).Although oocyte defects contribute to failure of sperm oocyte interaction,rarely are all oocytes from one wom-an affected.Low or zero fertilization in standard IVFwas usually caused by sperm abnormalities.Poor sperm-zona pel-lucida binding was frequently associated with failure of standard IVF and obvious defects of sperm motility or morpholo-gy.The size and shape of the acrosome is particularly important for sperm binding to the oocyte.The proportion ofacrosome intact sperm in the insemination medium was related to the IVF rote.Inducing the acrosome reaction with acalcium ionophore reduced sperm-zona binding.Blocking acrosome dispersal with an acrosin inhibitor prevented sperm-zona penetration.Sperm-zona penetration was even more highly related to IVF rates than was sperm-zona binding.Some patients had low or zero fertilization rates with standard IVF but normal sperm by conventional tests and normalsperm-zona binding.Few of their sperm underwent the acrosome reaction on the surface of the zona and none penetrat-ed the zona.In contrast,fertilization and pregnancy rates were high with intmcytoplasmic sperm injection.We call thiscondition defective zona pellucida induced acrosome reaction.Discovery of the nature of the abnormalities in the signaltmnsduction and effector pathways of the human zona pellucida induced acrosome reaction should result in simpler testsand treatments for the patients and also provide new leads for contraceptive development.
文摘During mammalian fertilisation, the zona pellucida （ZP） matrix surrounding the oocyte is responsible for the binding of the spermatozoa to the oocyte and induction of the acrosome reaction （AR） in the ZP-bound spermatozoon. The AR is crucial for the penetration of the ZP matrix by spermatozoa. The ZP matrix in mice is composed of three glycoproteins designated ZP1, ZP2 and ZP3, whereas in humans, it is composed of four （ZP1, ZP2, ZP3 and ZP4）. ZP3 acts as the putative primary sperm receptor and is responsible for AR induction in mice, whereas in humans （in addition to ZP3）, ZP1 and ZP4 also induce the AR. The ability of ZP3 to induce the AR resides in its C-terminal fragment. O-linked glycans are critical for the murine ZP3-mediated AR. However, N-linked glycans of human ZP1, ZP3 and ZP4 have important roles in the induction of the AR. Studies with pharmacological inhibitors showed that the ZP3-induced AR involves the activation of the Gi-coupled receptor pathway, whereas ZP1- and ZP4-mediated ARs are independent of this pathway. The ZP3-induced AR involves the activation of T-type voltage-operated calcium channels （VOCCs）, whereas ZP1- and ZP4-induced ARs involve both T- and L-type VOCCs. To conclude, in mice, ZP3 is primarily responsible for the binding of capacitated spermatozoa to the ZP matrix and induction of the AR, whereas in humans （in addition to ZP3）, ZP1 and ZP4 also participate in these stages of fertilisation.
文摘To bind and fertilize the egg, the spermatozoon should undergo few biochemical and motility changes in the female reproductive tract collectively called capacitation. The capacitated spermatozoon binds to the egg zona pellucida, and then undergoes the acmsome reaction （AR）, which allows its penetration into the egg. The mechanisms regulating sperm capacitation and the AR are not completely understood. In the present review, we summarize some data regarding the role and regulation of the epidermal growth factor receptor （EGFR） in these processes. In the capacitation process, the EGFR is partially activated by protein kinase A （PKA）, resulting in phospholipase D （PLD） activation and actin polymerization. Protein kinase C alpha （PKCα）, which is already activated at the beginning of the capacitation, also participates in PLD activation. Further activation of the EGFR at the end of the capacitation enhances intracellular Ca2＋ concentration leading to F-actin breakdown and allows the AR to take place. Under in vivoconditions, the EGFR can be directly activated by its known ligand epidermal growth factor （EGF）, and indirectly by activating PKA or by transactivation mediated by G protein-coupled receptors （GPCRs） activation or by ouabain. Under physiological conditions, sperm PKA is activated mainly by bicarbonate, which activates the soluble adenylyl cyclase to produce cyclic adenosine monophosphate （cAMP）, the activator of PKA. The GPCR activators angiotensin II or Ivsoohosphatidic acid, as well as ouabain and EGF are phvsioloeical comoonents oresent in the female reoroductive tract.
文摘Egg water was used to induce acrosome reaction in mature sperm of Fenneropenaeus chinensis .Transmission electron microscope and SDS PAGE were used to study the ultrastructure and protein changes of the sperm in F.chinensis ,which has undergone acrosome reaction.The results demonstrated that the sperms from female thelycum could be induced acrosome reaction in vitro by egg water,which was a kind of egg jelly released from oocyte into seawater during oocyte activation.More than fifty percent of sperm finished acrosome reaction during the first 30 min, in vitro .The whole event consists of the retraction of the spike and acrosome exocytosis.Spike was retracted and fused to acrosome cap followed by the release of acrosome granules.When the original egg water was diluted 5 and 10 times,The diluted egg water also have the ability to induce sperm acrosome reaction,but the acrosome reaction rate is much lower than that of the original egg water in the same time.Capillary zone electrophoresis was used to detect the biochemical components of egg water.Egg water was composed of two different components,which were also demonstrated by SDS PAGE.The molecular weights of the two kinds of egg jellies are both about 200kDa.Gelatin substrate SDS PAGE results did not show any hydrolytic enzyme activity in egg water.After acrosome reaction,many sorts of proteins in sperm are degraded.When the reacted sperms were examined with gelatin substrate SDS PAGE,there were six major peptide bands with hydrolytic enzyme activity were detected.Their molecular weights are 200kDa,130 kDa,66 kDa,53 kDa,48 kDa and 41 kDa respectively.These hydrolases cannot be detected in the sperms before acrosome reaction.The acrosome reaction of Chinese shrimp, F.chinensis is much different to that of the sperm in the shrimp, Sicyonia ingentis ,which the acrosome reaction was clearly studied.There is not filament formation during the acrosome reaction in Chinese shrimp.The time of exocytosis in the sperm of Chinese shrimp is also much longer than that of the sperm of Sicyonia ingentis .
基金The work was supported in part by grants HD25869 and HD34041 from the National Institute of Child & Human Development
文摘The fertilization process is the net result of a complex sequence of events that collectively result in the fusion of theopposite gametes. The male gamete undergoes continuous morphological and biochemical modifications during spermdevelopment in the testis (spermatogenesis), maturation in the epididymis, and capacitation in the female reproductivetract. Only the capacitated spermatozoa are able to recognize and bind to the bioactive glycan residue(s) on the ovum'sextracellular coat, the zona pellucida (ZP). Sperm-zona binding in the mouse and several other species is believed totake place in two stages. First, capacitated (acrosome-intact) spermatozoa loosely and reversibly adhere to the zona-in-tact ovum. In the second stage tight irreversible binding occurs. Both types of bindings are attributed to the presence ofglycan- binding proteins (receptors) on the sperm plasma membrane and their complementary bioactive glycan units(ligands) on the surface of the ZP. The carbohydrate-mediated adhesion event initiates a signal transduction cascade re-sulting in the exocytosis of acrosomal contents. This step is believed to be prerequisite which allows the hyperactivatedacrosome-reacted spermatozoa to penetrate the ZP and fertilize the ovum. This review focuses on the role of carbohy-drate residues in sperm-ovum interaction, and triggering of the acrosome reaction. I have attempted to discuss extensiveprogress that has been made to enhance our understanding of the well programmed multiple molecular events necessaryfor successful fertilization. This review will identify these events, and discuss the functional significance of carbohy-drates in these events.
文摘Aim： To investigate the possible functions of human sperm membrane protein （hSMP-1） in the process of fertilization. Methods： A 576-bp cDNA fragment of HSD-1 gene coding for the extracellular domain of hSMP-1 was cloned and expressed. The localization of this protein on human and mouse sperm was determined by indirect immunofluorescent staining by using anti-recombinant hSMP-1 （anti-rhSMP-1） antibodies. Sperm acrosome reaction and spermzona pellucida （ZP） binding assay were carried out in 10-week-old BALB/c mice. Results： Recombinant hSMP-1 was successfully cloned and expressed. The expression of the native protein was limited on the acrosome of human and mouse sperm. Treatment of anti-rhSMP-1 antibodies significantly decreased the average number of sperms bound to each egg. Meanwhile, the percentage of acrosome reaction was decreased in comparison to pre-immune control after treatment with anti-rhSMP-1 （P 〈 0.05）. Conclusion： The results suggest that anti-rhSMP-1 antibody inhibited mouse acrosome reaction and sperm-ZP binding.
文摘Sterols are essential components of the cell membrane lipid bilayer that include molecules such as cholesterol and desmosterol, which are significantly found in the spermatozoa of several animal species. However, the presence of desmosterol in rabbit semen has never been investigated. The aims of this study were to characterize the sterol composition of subfractions of ejaculated rabbit semen and evaluate the in vitro effects of sterol on the spermatozoa acrosome reaction and motility. Two sterols, occurring prevalently in the free form （94.3%）, were identified in whole semen collected from 10 fertile New Zealand White rabbits, specifically desmosterol （58.5% of total sterols） and cholesterol （35.9% of total sterols）. Desmosterol was the predominant sterol found in all subfractions of rabbit semen, varying from 56.7% （in the prostatic secretory granules, PSGs） to 63.8% （in the seminal plasma）. Spermatozoa contained an intermediate proportion of desmosterol （59.8%）, which was asymmetrically distributed between the heads （52.0% of the total content of sterols） and the tails （81.8%）. Results showed that both desmosterol and cholesterol can be transferred from the PSGs to the spermatozoa and are equally effective in inhibiting in vitro spermatozoa capacitation at a concentration higher than 1 mg L^-1. In contrast, neither desmosterol nor cholesterol had a significant effect on spermatozoa motility. Thus, it was concluded that, the various fractions of rabbit seminal fluid differ from each other in sterol composition and quantity, probably due to their different functional properties, and these fractions may undergo significant sterol changes depending on the stage of spermatozoa capacitation.
文摘Aim： To investigate if the phytoestrogen, genistein, affects essential functions of cryopreserved bovine spermatozoa. Methods： The effect of genistein upon motility was assessed by computer-assisted motion analysis. Hemizona assay was performed to detect the ability of spermatozoa binding to the zona pellucida. The inducibility of the acrosome reaction using progesterone and ZP3-6 peptide was analysed by fluorescein-conjugated Pisum sativum agglutinin （FITC-PSA）/Hoechst 33258 double staining. Capacitation after incubation with genistein was assessed by the chlortetracycline （CTC） assay. Immunoblots showed the pattern of protein tyrosine phosphorylation of cryopreserved bovine spermatozoa. Results： Immunodetection of tyrosine-phosphorylated proteins showed that genistein did not affect tyrosine phosphorylation in cryopreserved bovine spermatozoa. However, genistein significantly reduced the progesterone- and ZP3-6 peptide-mediated induction of the acrosome reaction and led to a dose-dependent inhibition of sperm-zona pellucida binding; while sperm motility and capacitation were not affected by this phytoestrogen, as indicated by computer-assisted sperm motion analysis and the CTC assay, respectively. Conclusion： Our results suggest that in cryopreserved bovine spermatozoa, genistein affects a protein tyrosine phosphorylation-independent signal transduction pathway that is involved in sperm capacitation, the acrosome reaction and sperm-zona pellucida binding.
文摘The acrosome reaction （AR）, an absolute requirement for spermatozoa and egg fusion, requires the influx of Ca2＋ into the spermatozoa through voltage-dependent Ca2＋ channels and store-operated channels. Maitotoxin （MTx）, a Ca2＋-mobilizing agent, has been shown to be a potent inducer of the mouse sperm AR, with a pharmacology similar to that of the zona pellucida （ZP）, possibly suggesting a common pathway for both inducers. Using recombinant human ZP3 （rhZP3）, mouse ZP and two MTx channel blockers （U73122 and U73343）, we investigated and compared the MTx- and ZP-induced ARs in human and mouse spermatozoa. Herein, we report that MTx induced AR and elevated intracellular Ca2＋ （[Ca2＋]~） in human spermatozoa, both of which were blocked by U73122 and U73343. These two compounds also inhibited the MTx-induced AR in mouse spermatozoa. In disagreement with our previous proposal, the AR triggered by rhZP3 or mouse ZP was not blocked by U73343, indicating that in human and mouse spermatozoa, the AR induction by the physiological ligands or by MTx occurred through distinct pathways. U73122, but not U73343 （inactive analogue）, can block phospholipase C （PLC）. Another PLC inhibitor, edelfosine, also blocked the rhZP3- and ZP-induced ARs. These findings confirmed the participation of a PLC-dependent signalling pathway in human and mouse zona protein-induced AR. Notably, edelfosine also inhibited the MTx-induced mouse sperm AR but not that of the human, suggesting that toxin-induced AR is PLC-dependent in mice and PLC-independent in humans.
文摘Capacitation and acrosome reaction are important prerequisites of the fertilization process. Capacitation is a highlycomplex phenomenon occurring in the female genital tract, rendering the spermatozoa capable of binding and fusionwith the oocyte. During capacitation various biochemical and biophysical changes occur in the spermatozoa and thespermatozoal membranes. Ions and ion channels also play important roles in governing the process of capacitation bychanging the fluxes of different ions which in turn controls various characteristics of capacitated spermatozoa. Alongwith the mobilization of ions the generation of free radicals and efflux of cholesterol also plays an important role in thecapacitation state of the spermatozoa. The generation of free radical and efflux of cholesterol change the mechano-dynamic properties of the membrane by oxidation of the polyunsaturated lipids and by generating the cholesterol freepatches. The process of capacitation renders the spermatozoa responsive to the inducers of the acrosome reaction. Theglycoprotein zona pellucida 3 (ZP3) of the egg coat zona pellucida is the potent physiological stimulator of the acro-some reaction; progesterone, a major compoent of the follicular fluid, is also an induce of the acrosome reaction.The inducers of the acrosome reaction cause the activation of the various ion-channels leading to high influxes of calci-um, sodium and bicarbonate. The efflux of cholesterol during the process of capacitation alters the permeablity of themembrane to the ions and generate areas which are prone to fusion and vesculation process during the acrosome reac-tion. Ths review focuses mainly on effects of the ion and ion-channels, free radicals, and membrane fluidity changesduring the process of capacitation and acrosome reaction. (Asian J Androl 1999 Sep; 1: 95-107)
文摘Objective: To explore the effect of NO on the spontaneous acrosome reaction in antisperm antibody (AsAb) positive rat spermatozoa. Methods: The rat model of AsAb was set up by artificial immunization. The level of AsAb in blood serum was determined by TAT and ELISA. Rat spermatozoa was visualized by staining the acrosome with Coomassie brilliant blue. The NO concentration in rat spermatozoa was assayed by HPLC. Results: The percentage of acrosome reaction, NO concentration, superoxide dismutase (SOD) and Na^+-K^+ATPase activity in AsAb positive rat spermatozoa were significantly decreased compared with the control group. Low dose of NO (SNP 10^(-9)～10^(-8) mol/L) increased the percentage of acrosome reaction and SOD activity, but had no effect on Na^+-K^+ATPase activity. High dose of NO (SNP 10^(-6)～10^(-4) mol/L) decreased the three items. Conclusion: The decrease in acrosome reaction in positive AsAb rat spermatozoa might be related to a decrease in NO and increase in O_2. (the SOD activity was decreased) in sperm. A low dose of NO might increase the percentage of acrosome reaction in AsAb positive rat by inhibiting the superoxide, while a high dose of NO was harmful to sperm function. [Reprod Contracep (in Chinese) 2002; 22: 203]
文摘Aim: To study the roles of tumor necrosis factor alpha (TNF-α) on the sperm acrosin activity and acrosome reaction. Methods: The sperm acrosin activity was tested by the method of BAEE/ ADH Unity and the acrosome reaction by the Triple-stain technique. Results: TNF-α decreased the sperm acrosin activity and acrosome reaction (P<0.01; P<0.01, respectively); it also inhibited the Ca2+-ATPase activity and SOD activity in sperm (P< 0.05; P<0.001, respectively), but increased the NOS activity and the amount of NO in sperm (P<0.001; P<0.001, respectively). While it had a less significant effect on the activity of Na+-K+-ATPase (P>0.05). Conclusion: TNF-α inhibits the sperm acrosin activity and acrosome reaction.
文摘The acrosome reaction of sperm was induced by calcium ionophore A 23187.The letionship between some ions and acrosome reaction by removing Na+ from the medium.or by adding angtagonist of K+.TEA chloride,or antagonist of Ca++,verapamil,or anlagonist of Na+-K+-ATPase,acetyl strophanthithidin is studied.The results show that Na+,H,Ca++ and Na+ pump are necessary.for acrosome reaclion of human sperm.The Ca++ might not enter the sperms through the channel of Ca++.
文摘No Con A receptor site was found on the intact plasma membranes of mammalian sperms before capacitation.After capacitation, however, the plasma membranes sloughed off, exposing the outer acrosomal membrane which is rich in Con A receptor sites.The vesicles formed during acrosome reaction were also found to bc rich in Con A receptor sites, suggesting their origin from outer acrosomal membranes.With completion of acrosome reaction, only inner acrosomal membrane was left in which no Con A receptor sites could be demonstrated.Also no Con A receptor site was found on egg plasma membrane throughout fertilization.It was thus shown that different membranes possess different properties.
基金supported by the Jiangsu Province Science and Technology Development Project (Grant No. BN2010026)
文摘The biological characteristics of crustacean spermatozoa is important for the artificial reproduction and genetic breeding. With reference to the latest studies and related materials, this paper reviewed the research progress in the biological characteristics of crustacean spermatozoa, such as morphological structure of sperm, spermatogenesis, sperm viability, preservation in vitro and acrosome reaction et al. The prospects of the research field have also been anticipated.
文摘A membrane protein was isolated from mouse sperm heads that had undergone acrosomal reaction induced by C2+ ionophore, A 23187, which, with a molecular weight of 77.6 kd, shows capability to block egg-sperm fusion. As revealed by analysis usintg isotopic markers, this protein is one of the chief membrane proteins of inner acrosomal membrane or the outer membrane of equatorial segment and Post-acrosomal region; treatment of mouse sperms with 0.6 μg/ml of the Purified protein for 30 minutes reduced the sperm-egg fusion index by 51%.The above results led us to the conclusion that the protein is an active participant in sperm-egg fusion. The possible existence of sperm receptor on egg plasma membrane was discussed.
文摘Aim: To produce biologically active recombinant human (rh) ZP proteins in a human cell for use in sperm function tests. Methods: The human embryonic kidney cell line 293T was employed to produce rhZP1, rhZP2 and rhZP3 proteins individually and together by co-expression. Presence of these proteins in the culture medium and cell lysate was assessed by Western blotting analysis. The effect of the recombinant proteins on the human AR was assessed. Results: RhZP2 and rhZP3 were secreted into the culture medium, whereas rhZPl was found only in the cell lysate. Interestingly, when all zona pellucida proteins were co-expressed in the same cells, rhZPl was also secreted into the culture medium. However, despite the presence of all three ZP proteins in sufficient concentration and evidence of heavy glycosylation on gel electrophoresis, biological activity to induce the AR was not observed. Conclusion: RhZP1, rhZP2 and rhZP3 were successfully expressed in the human embryonic kidney cell line 293T. It appears that an interaction amongst these proteins may be required for release of rhZPl from the cell. Although this approach is not satisfactory for producing active human ZP proteins, it makes a significant contribution to the understanding of the structural and functional characteristics of the ZP proteins.
基金National 973 Project of China(No.2006CB504002),Chinese National Prominent Youth Foundation(No.30425006)Program for Changjiang Scholars and Innovative Research Team in University(PCSIRT)(No.IRT0631)+1 种基金Research Grants Council(RGC)of Hong Kong(No.CUHK4524/05M)Li Ka Shing Institute of Health Sciences and Focused Investment of the Chinese University of Hong Kong,China.
文摘Prior to fertilization sperm has to undergo an activation process known as capaciation,leading to the acrosome reaction.Till now,little is known about the mechanism for preventing premature capacitation in sperm although decapacitation factors from various sources have been thought to be involved.In this study,we report that NYD-SP27,an isoform of phospholipase C Zeta 1(PLCZ1),is localized to the sperm acrosome in mouse and human spermatozoa by immunofluorescence using a specific antibody.Western blot and double staining analyses show NYD-SP27 becomes detached from sperm,as they undergo capacitation and acrosome reaction.The absence of HCO_(3)^(-),a key factor in activating capacitation,from the capacitation-inducing medium prevents the loss of NYD-SP27 from sperm.The anti-NYD-SP27 antibody also prevents the loss of NYD-SP27 from sperm,reduced the number of capacitated sperm,inhibited the acrosome reaction induced by ATP and progesterone,and inhibited agonist-induced PLC-coupled Ca^(2+)mobilization in sperm,which can be mimicked by the PLC inhibitor,U73122.These data strongly suggest that NYD-SP27 is a physiological inhibitor of PLC that acts as an intrinsic decapacitation factor in sperm to prevent premature capacitation and acrosome reaction.
