Journal articles on the topic 'Sperm membrane'
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1
Reis, L. S. L. S., A. A. Ramos, A. S. Camargos, and E. Oba. "Integrity of the plasma membrane, the acrosomal membrane, and the mitochondrial membrane potential of sperm in Nelore bulls from puberty to sexual maturity." Arquivo Brasileiro de Medicina Veterinária e Zootecnia 68, no. 3 (June 2016): 620–28. http://dx.doi.org/10.1590/1678-4162-8748.
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ABSTRACT This study evaluated the plasma membrane integrity, acrosomal membrane integrity, and mitochondrial membrane potential of Nelore bull sperm from early puberty to early sexual maturity and their associations with sperm motility and vigor, the mass motility of the spermatozoa (wave motion), scrotal circumference, and testosterone. Sixty Nelore bulls aged 18 to 19 months were divided into four lots (n=15 bulls/lot) and evaluated over 280 days. Semen samples, collected every 56 days by electroejaculation, were evaluated soon after collection for motility, vigor and wave motion under an optical microscope. Sperm membrane integrity, acrosomal integrity, and mitochondrial activity were evaluated under a fluorescent microscope using probe association (FITC-PSA, PI, JC-1, H342). The sperm were classified into eight integrity categories depending on whether they exhibited intact or damaged membranes, an intact or damaged acrosomal membrane, and high or low mitochondrial potential. The results show that bulls have a low amount of sperm with intact membranes at puberty, and the sperm show low motility, vigor, and wave motion; however, in bulls at early sexual maturity, the integrity of the sperm membrane increased significantly. The rate of sperm membrane damage was negatively correlated with motility, vigor, wave motion, and testosterone in the bulls, and a positive correlation existed between sperm plasma membrane integrity and scrotal circumference. The integrity of the acrosomal membrane was not influenced by puberty. During puberty and into early sexual maturity, bulls show low sperm mitochondrial potential, but when bulls reached sexual maturity, high membrane integrity with high mitochondrial potential was evident.
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McCulloh, D. H., and E. L. Chambers. "Fusion of membranes during fertilization. Increases of the sea urchin egg's membrane capacitance and membrane conductance at the site of contact with the sperm." Journal of General Physiology 99, no. 2 (February 1, 1992): 137–75. http://dx.doi.org/10.1085/jgp.99.2.137.
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The early events of fertilization that precede and cause activation of an egg have not been fully elucidated. The earliest electrophysiological change in the sea urchin egg is a sperm-evoked increase of the egg's membrane conductance. The resulting depolarization facilitates entry of the fertilizing sperm and precludes the entry of supernumerary sperm. The sequence of the increase in the egg's membrane conductance, gamete membrane fusion, egg activation, and sperm entry, including causal relationships between these events, are not known. This study reports the use of whole egg voltage clamp and loose patch clamp to monitor simultaneously changes of membrane conductance and capacitance at the site of sperm-egg contact. Measurements were made during sperm-egg interactions where sperm entry readily proceeded or was precluded by maintaining the egg's membrane potential either at large, negative values or at positive values. Whenever the sperm evoked an increase of the egg's membrane conductance, that increase initiated abruptly, was localized to the site of sperm attachment, and was accompanied by a simultaneous abrupt increase of the membrane capacitance. This increase of capacitance indicated the establishment of electrical continuity between gametes (possibly fusion of the gametes' plasma membranes). If sperm entry was blocked by large negative membrane potentials, the capacitance cut off rapidly and simultaneously with a decrease of the membrane conductance, indicating that electrical continuity between gametes was disrupted. When sperm entry was precluded by positive membrane potentials, neither conductance nor capacitance increased, indicating that sperm entry was halted before the fusion of membranes. A second, smooth increase of capacitance was associated with the exocytosis of cortical granules near the sperm in eggs that were activated. Electrical continuity between the gametes always preceded activation of the egg, but transient electrical continuity between the gametes alone was not always sufficient to induce activation.
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Miller, Jr, R. R., F. Beranek, A. L. Anderson, S. D. Johnston, and B. Nixon. "Plasma and acrosomal membrane lipid content of saltwater crocodile spermatozoa." Reproduction, Fertility and Development 33, no. 9 (2021): 596. http://dx.doi.org/10.1071/rd21007.
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This study describes the chemical lipid composition of the sperm plasma and acrosomal membranes of the saltwater crocodile Crocodylus porosus with the aim of providing new insights into sperm physiology, particularly that associated with their preservation ex vivo. The specific fatty acid composition of the sperm plasma and acrosomal membranes is documented. The mean (±s.d.) ratio of unsaturated to saturated membrane fatty acids within the plasma membrane was 2.57±0.50, and was determined to be higher than a similar analysis of the lipids found in the acrosomal membrane (0.70±0.10). The saltwater crocodile sperm plasma membrane also contained remarkably high levels of cholesterol (mean (±s.d.) 40.7±4.5 nmol per 106 sperm cells) compared with the spermatozoa of other amniote species that have so far been documented. We suggest that this high cholesterol content could be conferring stability to the crocodile sperm membrane, allowing it to tolerate extreme osmotic fluxes and rapid changes in temperature. Our descriptive analysis now provides those interested in reptile and comparative sperm physiology an improved baseline database for interpreting biochemical changes associated with preservation pathology (e.g. cold shock and cryoinjury), epididymal sperm maturation and capacitation/acrosome reaction.
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Feitosa, Weber Beringui, Marcella Pecora Milazzotto, Renata Simões, Mariana Rovegno, Alessandra Coralo Nicacio, Aníbal Ballarotti Nascimento, José Sérgio Arruda Gonçalves, José Antonio Visintin, and Mayra Elena Ortiz D'Ávila Assumpção. "Bovine sperm cells viability during incubation with or without exogenous DNA." Zygote 17, no. 4 (June 16, 2009): 315–20. http://dx.doi.org/10.1017/s0967199409005449.
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SummaryThe aim of this study was to assess the effect of exogenous DNA and incubation time on the viability of bovine sperm. Sperm were incubated at a concentration of 5 × 106/ml with or without plasmid pEYFP–NUC. Fluorescent probes, propidium iodide/Hoechst 33342, FITC–PSA and JC-1, were used to assess plasma membrane integrity (PMI), acrosome membrane integrity (AMI) and mitochondrial membrane potential (MMP) respectively at 0, 1, 2, 3 and 4 h of incubation. Exogenous DNA addition did not affect sperm viability; however, incubation time was related to sperm deterioration. Simultaneous assessment of PMI, AMI and MMP showed a reduction in the number of sperm with higher viability (integrity of plasma and acrosome membranes and high mitochondrial membrane potential) from 58.7% at 0 h to 7.5% after 4 h of incubation. Lower viability sperm (damaged plasma and acrosome membranes and low mitochondrial membrane potential) increased from 4.6% at 0 h to 25.9% after 4 h of incubation. When PMI, AMI and MMP were assessed separately we noticed a reduction in plasma and acrosome membrane integrity and mitochondrial membrane potential throughout the incubation period. Therefore, exogenous DNA addition does not affect sperm viability, but the viability is reduced by incubation time.
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Belayeva, N. S. "Relation of the male gametes with embryo sec cells. The hypothesis of double fertilization." Acta Societatis Botanicorum Poloniae 50, no. 1-2 (2014): 173–75. http://dx.doi.org/10.5586/asbp.1981.026.
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The sperms one after another get out of synergids. The front sperm gets the first into the egg cell attraction zone and then the sperm comes into contact with egg membrane. At this moment Attraction ceases and the second sperm is led by a current of cytoplasma to the central nucleus. In the egg cell the sperm nucleus is led to the nucleus by cytoplasmic current too. After fertilization the character of cytoplasmic motion changes, because of a cell membrane damage. The presence of the sperm in the female nuclei may also serve as a regulating factor.
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Tubbs, Christopher, and Peter Thomas. "Progestin Signaling through an Olfactory G Protein and Membrane Progestin Receptor-α in Atlantic Croaker Sperm: Potential Role in Induction of Sperm Hypermotility." Endocrinology 150, no. 1 (September 14, 2008): 473–84. http://dx.doi.org/10.1210/en.2008-0512.
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Progestin stimulation of sperm hypermotility remains poorly understood despite having been described in numerous vertebrate species. We show here that progestin stimulation of sperm hypermotility in a teleost, the Atlantic croaker (Micropogonias undulatus) is associated with activation of an olfactory G protein (Golf). Furthermore, we provide evidence that this progestin action is mediated by membrane progestin receptor-α (mPRα). Golf was identified in croaker sperm membranes and was specifically activated after treatment with the progestin 17,20β,21-trihydroxy-4-pregnen-3-one (20β-S). Treatment of sperm membranes with 20β-S caused an increase in cAMP production, which was blocked by pretreatment with cholera toxin and two membrane adenylyl cyclase inhibitors: 2′,5′-dideoxyadenosine and SQ22536. Moreover, preincubation of croaker sperm with 2′,5′-dideoxyadenosine and SQ22536 resulted in a significant inhibition of 20β-S-stimulated hypermotility. Binding of [3H]20β-S to sperm membranes was decreased after pretreatment with GTPγS but not pertussis toxin, suggesting the receptor is coupled to a pertussis toxin-insensitive G protein. Golf and mPRα were coexpressed on the sperm midpiece and flagella and were coimmunoprecipitated from sperm membranes. Finally, expression of mPRα protein on sperm increased after in vivo treatment with LHRH and was associated with increased induction of sperm motility by 20β-S. These results suggest that 20β-S activates mPRα in croaker sperm, which in turn activates Golf and membrane adenylyl cyclase to stimulate sperm hypermotility. Taken together these findings provide a plausible mechanism by which progestins stimulate sperm hypermotility in croaker and provide the first evidence of hormonal activation of Golf in any species. Progestin activation of an olfactory G protein pathway, likely through membrane progestin receptor alpha, is associated with induction of hypermotility in Atlantic croaker sperm.
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Waterhouse, K. E., P. O. Hofmo, A. Tverdal, and R. R. Miller. "Within and between breed differences in freezing tolerance and plasma membrane fatty acid composition of boar sperm." Reproduction 131, no. 5 (May 2006): 887–94. http://dx.doi.org/10.1530/rep.1.01049.
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The response of sperm to cryopreservation and the fertility of frozen–thawed semen varies between species. Besides species differences in sperm physiology, structure and biochemistry, factors such as sperm transport and female reproductive tract anatomy will affect fertility of frozen–thawed semen. Therefore, studying differences in sperm cryotolerance between breeds and individuals instead of between species may reveal sources of variability in sperm cryotolerance. In the present study, the effect of cooling, re-warming and freezing and thawing on plasma membrane and acrosome integrity of sperm within and between Norwegian Landrace and Duroc breeds was studied. Furthermore, the relation between post-thaw survival rate and fatty acid composition of the sperm plasma membranes was investigated. Flow cytometry assessments of plasma membrane and acrosome integrity revealed no significant differences between breeds; however there were significant male-to-male variations within breeds in post-thaw percentages of live sperm (plasma membrane intact). The most abundant fatty acids in the plasma membranes from both breeds were palmitic acid (16:0), stearic acid (18:0), oleic acid (18:1, n-9), docosapentaenoic acid (22:5, n-6) and docosahexaenoic acid (22:6, n-3). The ratio of ∑ 22:5, n-6 and 22:6, n-3/∑ all other membrane fatty acids was significantly related to survival rate (plasma membrane integrity) of sperm for both Norwegian Landrace (correlation coefficient (rs) = 0.64,P< 0.05) and Duroc (rs= 0.67,P< 0.05) boars. In conclusion, male-to-male differences in sperm survival rate after freezing and thawing may be partly related to the amount of long-chain polyunsaturated fatty acids in the sperm plasma membranes.
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Aguas, A. P., and P. Pinto da Silva. "The acrosomal membrane of boar sperm: a Golgi-derived membrane poor in glycoconjugates." Journal of Cell Biology 100, no. 2 (February 1, 1985): 528–34. http://dx.doi.org/10.1083/jcb.100.2.528.
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The acrosome is a large secretory vesicle of the sperm head that carries enzymes responsible for the digestion of the oocyte's investments. The event leads to sperm penetration and allows fertilization to occur. Release of the acrosomal enzymes is mediated by the interaction between sperm acrosomal and plasma membranes (acrosome reaction). Biochemical characterization of the acrosomal membrane has been restrained by a lack of methods to isolate uncontaminated fractions of the membrane. Here, we use new methods to expose the membrane to in situ cytochemical labeling by lectin-gold complexes. We study the topology and relative density of glycoconjugates both across and along the plane of the acrosomal membrane of boar sperm. Detachment of the plasma membrane from glutaraldehyde-fixed cells exposed the cytoplasmic surface of the acrosome to the lectin markers; freeze-fractured halves of the acrosomal membrane were marked by "fracture-label" (Aguas, A. P., and P. Pinto da Silva, 1983, J. Cell Biol. 97:1356-1364). We show that the cytoplasmic surface of the intact acrosome is devoid of binding sites for both concanavalin A (Con A) and wheat germ agglutinin (WGA). By contrast, it contains a high density of neuraminidase-resistant anionic sites detected by cationic ferritin. On freeze-fractured sperm, the receptors for Con A partitioned with the exoplasmic membrane half of the acrosomal membrane. The Con A-binding glycoconjugates were accumulated on the equatorial segment of the membrane. A low density of WGA receptors, as well as of intramembrane particles, was found on the freeze-fracture halves of the acrosomal membrane. The plasma membrane displayed, in the same preparations, a high density of receptors for both Con A and WGA. We conclude that the acrosome is limited by a membrane poor in glycoconjugates, which are exclusively exposed on the exoplasmic side of the bilayer. Regionalization of Con A receptors on the acrosome shows that sperm intracellular membranes, like the sperm surface, express domain distribution of glycocomponents.
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Spungin, B., I. Margalit, and H. Breitbart. "Sperm exocytosis reconstructed in a cell-free system: evidence for the involvement of phospholipase C and actin filaments in membrane fusion." Journal of Cell Science 108, no. 6 (June 1, 1995): 2525–35. http://dx.doi.org/10.1242/jcs.108.6.2525.
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We used a cell-free system to study membrane fusion during sperm exocytosis (acrosome reaction). Extracted bovine sperm plasma and outer acrosomal membranes were labeled with chlorophyll a or DCY, respectively. The occurrence of membrane fusion is indicated by the ability of the probes to diffuse from one membrane species to another which is revealed by resonance energy transfer between the two probes. We have previously shown using this system that the requirement of capacitation for sperm exocytosis is retained in cell-free membrane fusion, and that the pH and calcium dependence of the cell-free fusion mimics those of exocytosis in intact cells. In the present report we further characterize the fusion of sperm membranes which we observe in our assay. Phosphoproteins and phospholipases were found to be involved in the membrane fusion step of sperm exocytosis. Protein kinases, phosphatases, and Gi-like proteins, while involved in exocytosis in intact cells, are not involved specifically in the membrane fusion step of exocytosis. The role of membrane bound F-actin in regulating membrane fusion was also studied using fluorescently labeled phalloidin. The results show that cortical F-actin has two roles in regulating sperm exocytosis. One is to form a scaffolding to hold phospholipase C at the membrane. It also functions as a physical barrier to membrane fusion which is removed by the increases in intracellular calcium and pH which precede fusion.
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Tarvis, K. M., P. H. Purdy, and J. K. Graham. "51 CHANGING ROOSTER SPERM MEMBRANES TO FACILITATE CRYOPRESERVATION." Reproduction, Fertility and Development 24, no. 1 (2012): 137. http://dx.doi.org/10.1071/rdv24n1ab51.
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Cryopreservation damages rooster sperm membranes. Part of this damage is due to membrane transitioning from the fluid to the gel state as temperature is reduced. Some of this damage may be prevented by increasing membrane fluidity at low temperatures by incorporating cholesterol or unsaturated lipids into the membrane. Different concentrations of cholesterol-loaded cyclodextrins (CLC) and lipid-loaded cyclodextrins (LLC) containing 1,2-dilinoleoyl-sn-glycero-3-phosphocholine; 1,2-dilinoleoyl-rac-glycerol; and 1,2-dilinolenoyl-sn-glycero-3-phosphocholine were added to rooster sperm to determine if they improved cryopreservation. Osmotic stresses when cryoprotectants (CPA) are added to the cells before freezing and when the CPA are removed from cells after thawing also cause membrane damage. To minimize this damage, low molecular weight CPA with high membrane permeability were tested to determine their effectiveness for cryopreserving sperm. Rooster semen was collected from several birds, pooled and diluted to 800 million sperm mL–1 at 5°C in Lake's Low Temperature diluent (LLT). Sperm were treated with either LLC (0.25, 0.5, 1, 1.5, 2, 4 and 6 mg mL–1) or CLC (0.5, 1 and 2 mg mL–1) for 30 min. The sperm were diluted 1:1 with LLT containing 18% CPA, resulting in final CPA concentrations of 9%. The CPA tested were glycerol (G), methylacetamide (MA), dimethylformamide (DMF), methylformamide (MF) and ethylene glycol (EG). The sperm were frozen in liquid nitrogen vapor and stored in liquid nitrogen. Straws were thawed in 5°C water and sperm motility and membrane integrity analysed immediately. Sperm motility was measured using computer-assisted sperm analysis (CASA) and membrane integrity was analysed by flow cytometry using propidium iodide to detect cells with damaged membranes. Data were analysed by ANOVA and means separated using Student–Newman–Keuls multiple comparison test. Addition of LLC and CLC did improve sperm cryosurvival rates (P > 0.05). Using G as the CPA resulted in higher percentages of motile (54%) and viable (58%) sperm than MA (47 and 52%; P < 0.05), whereas DMF, EG and MF resulted in less than 45% motile cells (P < 0.05). In conclusion, altering sperm membrane composition using CLC and LLC did not improve post-thaw motility or viability in rooster sperm. Although MA did not protect the rooster sperm from cryodamage as effectively as G, future assays will need to determine the fertilizing capacity of sperm frozen using these CPA. We thank CSU-CVBMS for funding support.
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Wysokińska, Anna, and Dorota Szablicka. "Integrity of Sperm Cell Membrane in the Semen of Crossbred and Purebred Boars during Storage at 17 °C: Heterosis Effects." Animals 11, no. 12 (November 25, 2021): 3373. http://dx.doi.org/10.3390/ani11123373.
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The aim of the study was to assess changes in the integrity of sperm cell membranes during the storage of semen collected from Duroc × Pietrain crossbred boars and purebred boars of the component breeds. To compare the cell membrane integrity of sperm heads in crossbred and purebred boars, heterosis effects were estimated. The study was conducted on 48 ejaculates collected from Duroc × Pietrain crossbred boars and from purebred Duroc and Pietrain boars used for artificial insemination. Microscope slides were prepared from each ejaculate for the evaluation of the cell membrane integrity of the sperm, at 1, 24, 48, 72, and 96 h after collection of the ejaculate. Diluted ejaculates were stored at 17 °C. Sperm membrane integrity was analysed by two methods: SYBR-14/PI and eosin–nigrosin. Our results showed that the cell membrane integrity of sperm heads changed with storage time, but the extent of the changes varied depending on the genetic group of boars. The semen of Duroc × Pietrain crossbreds was clearly seen to be less sensitive to storage conditions than that of boars of the parent breeds, which was confirmed by the calculated heterosis effects. The percentage of sperm with an intact cell membrane was higher in crossbred boars than in purebred boars (p ≤ 0.05). In addition, significantly fewer moribund sperm spermatozoa and spermatozoa with a damaged cell membrane were observed in crossbred boars (p ≤ 0.05). In the semen of purebred Duroc and Pietrain boars, the cell membrane integrity of the sperm should be assessed more often during storage than in the semen of Duroc × Pietrain crossbred boars. This study provides valuable information for the development and implementation of semen quality monitoring in crossbred boars and boars of the parent breeds during storage at 17 °C with respect to the cell membrane structure of sperm heads. The evaluation methods used effectively identify damage to the cell membranes of the sperm during semen storage.
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Betancourt, Miguel, Yvonne Ducolomb, Irma Jiménez, Eduardo Casas, Edmundo Bonilla, and Trish Berger. "Sperm plasma membrane receptors for the porcine oocyte plasma membrane." Zygote 6, no. 2 (May 1998): 155–58. http://dx.doi.org/10.1017/s0967199498000082.
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In vitro fertilisation (IVF) was used to assess the ability of solubilised sperm plasma membrane (PM) proteins to inhibit the interaction of intact gametes. This is a first step before evaluating the ability of individual isolated proteins to competitively inhibit sperm-oocyte interaction as part of the process of studying the molecular events of fertilisation. Porcine oocytes were aspirated from ovaries, matured for 48 h in Medium 199, and the zona pellucida (ZP) was removed by exposure to acid Tyrode's solution. ZP-free matured oocytes were exposed to 200–800 μg/ml sperm PM protein for 1 h prior to insemination and during gamete co-incubation. Twenty-four hours after insemination with 5 × 105 capacitated sperm/ml, the oocytes were fixed, stained and examined. Sperm PM protein clearly inhibited IVF in a concentration-dependent manner (r = −0.87). The inhibition index (I50%), representing the sperm PM protein concentration necessary to inhibit IVF to 50% of the control value, was 310 µg/ml. These results demonstrate that solubilised sperm PM protein inhibits the interaction of intact gametes as one might expect for receptor-ligand interactions. Furthermore, the complement of sperm PM proteins appeared maximally effective at a calculated concentration of 690 µm/ml, providing a foundation for further studies with individual proteins.
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Hartwig, F. P., F. P. Lisboa, G. A. Monteiro, R. R. D. Maziero, M. A. Alvarenga, F. O. Papa, and J. A. Dell'aqua. "22 EFFECT OF CHOLESTEROL ADDITION TO EQUINE SPERM MEMBRANE ON FERTILITY." Reproduction, Fertility and Development 25, no. 1 (2013): 158. http://dx.doi.org/10.1071/rdv25n1ab22.
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Artificial insemination with cooled-shipped semen has been widely used in horse breeding. However, some stallions, referred to as poor coolers, present abrupt fertility decrease when their semen is processed, cooled, and transported. Cholesterol incorporation into sperm membranes improves the quality of cryopreserved semen by increasing the sperm membrane stability and fluidity at low temperatures. Despite the beneficial effect of cholesterol addition on sperm quality, studies demonstrate that the presence of large amounts of cholesterol in the plasma membrane interferes with the physiological process of sperm capacitation and is detrimental to frozen equine sperm fertility. The aim of this study was to assess the fertility of cooled semen from good-cooler and poor-cooler stallions after adding cholesterol to sperm membranes. Two stallions were used and classified as good cooler (n = 1) and poor cooler (n = 1) based on the ability to maintain sperm progressive motility after 24 h of cooling at 5°C. For classification of the stallions, the fertility history was also taken into account through the results of pregnancy per cycle using inseminations with cooled semen (<50% for poor cooler and >70% for good cooler). Ejaculates of these stallions were subjected to 2 treatments: control (CON) and cholesterol (CLC). In the CON group, the semen was extended to 30 × 106 sperms mL–1 with skim milk-based extender (BotuSemen™). In the CLC group, the semen was extended as in the CON group plus 0.25 µL/1 × 106 sperms of 6.1 mM cholesterol-loaded cyclodextrin was added. Afterwards, both treatments were cooled at 5°C for 24 h. To test the fertility of poor-cooler and good-cooler stallions, 2 cycles from 25 mares and 2 cycles from 10 mares were respectively used. For both stallions, randomly for each mare, the inseminations were performed by alternating both treatments. If the mare was first inseminated with the CLC treatment, in the next cycle the CON treatment was used and vice versa. After 24 h of ovulation induction, the inseminations were done in the uterine body with 1 × 109 viable cells. Statistical analyses were performed using the Fisher exact test and significance was set at P < 0.05. For the poor cooler, the CON treatment presented 44% pregnancy/cycle compared to 76% for the CLC treatment (11/25a v. 19/25b). For the good cooler, both treatments presented 80% (8/10) pregnancy/cycle. The results suggest that the fertility capability of stallions is not affected by incorporation of cholesterol-loaded cyclodextrin to the sperm plasma membrane. Additionally, the utilization of cholesterol-loaded cyclodextrin may be an option to enable the utilization of cooled-shipped semen from poor cooler stallions for AI programs.
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Kasai, T., K. Hoshi, and R. Yanagimachi. "Effect of sperm immobilisation and demembranation on the oocyte activation rate in the mouse." Zygote 7, no. 3 (August 1999): 187–93. http://dx.doi.org/10.1017/s0967199499000568.
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To analyse the effect of the state of the sperm plasma membrane on oocyte activation rate following intracytoplasmic sperm injection (ICSI), three types of human and mouse spermatozoa (intact, immobilised and Triton X-100 treated) were individually injected into mouse oocytes. At 30, 60 and 120 min after injection, maternal chromosomes and sperm nuclei within oocytes were examined. Following human sperm injection, the fastest and the most efficient oocyte activation and sperm head decondensation occurred when the spermatozoa were treated with Triton X-100. Intact spermatozoa were the least effective in activating oocytes. Thus, the rate of mouse oocyte activation following human sperm injection is greatly influenced by the state of the sperm plasma membrane during injection. When mouse spermatozoa were injected into mouse oocytes, the rates of oocyte activation and sperm head decondensation within activated oocytes were the same irrespective of the type of sperm treatment prior to injection. We witnessed that live human spermatozoa injected into moue oocytes often kept moving very actively within the ooplasm for more than 60 min, whereas motile mouse spermatozoa usually became immotile within 20 min after injection into the ooplasm. In 0.002% Triton X-100 solution, mouse spermatozoa are immobilised faster than human spermatozoa. These facts seem to suggest that human sperm plasma membranes are physically and biochemically more stable than those of mouse spermatozoa. Perhaps the physical and chemical properties of the sperm plasma membrane vary from species to species. For those species whose spermatozoa have ‘stable’ plasma membranes, prior removal or ‘damage’ of sperm plasma membranes would increase the success rate of ICSI.
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Spungin, B., and H. Breitbart. "Calcium mobilization and influx during sperm exocytosis." Journal of Cell Science 109, no. 7 (July 1, 1996): 1947–55. http://dx.doi.org/10.1242/jcs.109.7.1947.
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We have previously shown that two intracellular events which occur during capacitation of bovine sperm are the formation of actin filaments on the plasma and outer acrosomal membranes and the attachment of a PIP2-specific phospholipase C (PLC) to this membrane bound F-actin. This PLC plays an essential role in sperm exocytosis (acrosome reaction). In the present report, we further elucidated the role of this PLC using a PIP2-specific PLC of bacterial origin. This PLC is different from the endogenous sperm PLC in that it is calcium independent and not inhibited by neomycin. Here we report using bovine sperm that this bacterial PLC can restore actin release from extracted membranes as well as membrane fusion in a cell-free assay when the endogenous PLC is inhibited by neomycin. The sperm PLC requires 2 microM calcium for half maximal activation, while half maximal actin release from extracted plasma membranes occurs at 80 microM. Extracted sperm membranes were examined for calcium pumps and channels. Sperm plasma membranes were found to possess a thapsigargin insensitive calcium pump and calcium channels which are opened by phosphorylation by protein kinase C. The acrosomal membrane possesses a calcium pump which is inhibited by thapsigargin and calcium channels which are opened by cAMP. These observations are discussed in terms of a model of acrosomal exocytosis which involves a calcium rise that occurs in two stages resulting from calcium mobilization from internal stores followed by influx of extracellular calcium.
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Kan, F. W., and Y. Lin. "Topographical distribution of phospholipids in boar sperm plasma and intracellular membranes as revealed by freeze-fracture cytochemistry." Journal of Histochemistry & Cytochemistry 44, no. 7 (July 1996): 687–701. http://dx.doi.org/10.1177/44.7.8675990.
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We used fracture-label and label-fracture cytochemistry in conjunction with the phospholipase A2-colloidal gold (PLA2-CG) technique to study the distribution of phospholipids in ejaculated boar spermatozoa. These techniques provide visualization of the topographical distribution of phospholipids in freeze-fractured sperm membranes in a three-dimensional view. In various freeze-fractured boar sperm membranes and crossfractured cytoplasmic structures, quantitative analysis revealed that the nuclear envelope membranes and the nuclear content possessed the highest labeling density of PLA2-CG. Moderate labeling was detected over acrosomal membranes, especially the inner acrosomal membrane. Replicas of both protoplasmic and exoplasmic fracture faces of the plasma membrane of boar sperm head showed a relatively low density of PLA2-CG labeling. Moreover, a differential distribution of phospholipids was seen over the protoplasmic face of the plasma membrane domains of the sperm head, which showed the highest concentration of gold particles in the postacrosomal region, followed by the equatorial segment and the anterior acrosome region. The PLA2-CG labeling densities over the post-acrosomal region and the equatorial segment were significantly higher than that over the anterior acrosome region. In the flagellum, an intense labeling was also seen over crossfractured mitochondria, dense fibers, and fibrous sheath. The protoplasmic fracture face of the plasma membrane over the middle piece, the annulus, and the principal piece was moderately labeled by PLA2-CG. No significant difference in mean labeling density of PLA2-CG was detected among the three membrane domains. In label-fracture preparations, exoplasmic halves of the plasma membrane of the head and the middle piece of the tail were uniformly labeled with PLA2-CG. However, the annulus and principal piece were devoid of PLA2-CG binding sites. These results indicate that differential distribution of phospholipids associated with the boar sperm membranes may reflect phospholipid composition of membrane domains characteristic of special physiological functions.
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Tulsiani, D. R., M. D. Skudlarek, and M. C. Orgebin-Crist. "Novel alpha-D-mannosidase of rat sperm plasma membranes: characterization and potential role in sperm-egg interactions." Journal of Cell Biology 109, no. 3 (September 1, 1989): 1257–67. http://dx.doi.org/10.1083/jcb.109.3.1257.
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During the course of a study of glycoprotein processing mannosidases in the rat epididymis, we have made an intriguing discovery regarding the presence of a novel alpha-D-mannosidase on the rat sperm plasma membranes. Unlike the sperm acrosomal "acid" mannosidase which has a pH optimum of 4.4, the newly discovered alpha-D-mannosidase has a pH optimum of 6.2, and 6.5 when assayed in sperm plasma membranes and intact spermatozoa, respectively. In addition, the two enzymes show different substrate specificity. The acrosomal alpha-D-mannosidase is active mainly towards synthetic substrate, p-nitrophenyl alpha-D-mannopyranoside, whereas the sperm plasma membrane alpha-D-mannosidase shows activity mainly towards mannose-containing oligosaccharides. Evidence is presented which suggest that the sperm plasma membrane alpha-D-mannosidase is different from several processing mannosidases previously characterized from the rat liver. The newly discovered alpha-D-mannosidase appears to be an intrinsic plasma membrane component, since washing of the purified membranes with buffered 0.4 M NaCl did not release the enzyme in soluble form. The enzyme requires nonionic detergent (Triton X-100) for complete solubilization. The enzyme is activated by Co2+ and Mn2+. However, Cu2+ and Zn2+ are potent inhibitors of the sperm plasma membrane alpha-D-mannosidase. At a concentration of 0.1 mM, these divalent cations caused nearly complete inactivation of the sperm enzyme. In addition methyl-alpha-D-mannoside, methyl-alpha-D-glucoside, mannose, 2-deoxy-D-glucose, and D-mannosamine are inhibitors of the sperm surface alpha-D-mannosidase. The physiological role of the newly discovered enzyme is not yet known. Several published reports in three species, including the rat, suggest that the sperm surface alpha-D-mannosidase may have a role in binding to mannose-containing saccharides presumably present on the zona pellucida.
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Frolikova, Michaela, Eliska Valaskova, Jiri Cerny, Audrey Lumeau, Natasa Sebkova, Veronika Palenikova, Noemi Sanches-Hernandez, Alzbeta Pohlova, Pavla Manaskova-Postlerova, and Katerina Dvorakova-Hortova. "Addressing the Compartmentalization of Specific Integrin Heterodimers in Mouse Sperm." International Journal of Molecular Sciences 20, no. 5 (February 26, 2019): 1004. http://dx.doi.org/10.3390/ijms20051004.
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Integrins are transmembrane cell receptors involved in two crucial mechanisms for successful fertilization, namely, mammalian intracellular signaling and cell adhesion. Integrins α6β4, α3β1 and α6β1 are three major laminin receptors expressed on the surface of mammalian cells including gametes, and the presence of individual integrin subunits α3, α6, β1 and β4 has been previously detected in mammalian sperm. However, to date, proof of the existence of individual heterodimer pairs in sperm and their detailed localization is missing. The major conclusion of this study is evidence that the β4 integrin subunit is expressed in mouse sperm and that it pairs with subunit α6; additionally, there is a detailed identification of integrin heterodimer pairs across individual membranes in an intact mouse sperm head. We also demonstrate the existence of β4 integrin mRNAs in round spermatids and spermatogonia by q-RT-PCR, which was further supported by sequencing the PCR products. Using super-resolution microscopy accompanied by colocalization analysis, we located integrin subunits as follows: α6/β4-inner apical acrosomal membrane and equatorial segment; α3, α6/β1, β4-plasma membrane overlaying the apical acrosome; and α3/β1-outer acrosomal membrane. The existence of α6β4, α3β1 and α6β1 heterodimers was further confirmed by proximity ligation assay (PLA). In conclusion, we delivered detailed characterization of α3, α6, β1 and β4 integrin subunits, showing their presence in distinct compartments of the intact mouse sperm head. Moreover, we identified sperm-specific localization for heterodimers α6β4, α3β1 and α6β1, and their membrane compartmentalization and the presented data show a complexity of membranes overlaying specialized microdomain structures in the sperm head. Their different protein compositions of these individual membrane rafts may play a specialized role, based on their involvement in sperm-epithelium and sperm-egg interaction.
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Hamilton, Thais Rose dos Santos, Camilla Mota Mendes, Letícia Signori de Castro, Patrícia Monken de Assis, Adriano Felipe Perez Siqueira, Juliana de Carvalho Delgado, Marcelo Demarchi Goissis, et al. "Evaluation of Lasting Effects of Heat Stress on Sperm Profile and Oxidative Status of Ram Semen and Epididymal Sperm." Oxidative Medicine and Cellular Longevity 2016 (2016): 1–12. http://dx.doi.org/10.1155/2016/1687657.
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Higher temperatures lead to an increase of testicular metabolism that results in spermatic damage. Oxidative stress is the main factor responsible for testicular damage caused by heat stress. The aim of this study was to evaluate lasting effects of heat stress on ejaculated sperm and immediate or long-term effects of heat stress on epididymal sperm. We observed decrease in motility and mass motility of ejaculated sperm, as well as an increase in the percentages of sperm showing major and minor defects, damaged plasma and acrosome membranes, and a decrease in the percentage of sperm with high mitochondrial membrane potential in the treated group until one spermatic cycle. An increased enzymatic activity of glutathione peroxidase and an increase of stressed cells were observed in ejaculated sperm of the treated group. A decrease in the percentage of epididymal sperm with high mitochondrial membrane potential was observed in the treated group. However, when comparing immediate and long-term effects, we observed an increase in the percentage of sperm with low mitochondrial membrane potential. In conclusion, testicular heat stress induced oxidative stress that led to rescuable alterations after one spermatic cycle in ejaculated sperm and also after 30 days in epididymal sperm.
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AVILÉS, Manuel, Irene ABASCAL, José Angel MARTÍNEZ-MENÁRGUEZ, María Teresa CASTELLS, Sheri R. SKALABAN, José BALLESTA, and Jack A. ALHADEFF. "Immunocytochemical localization and biochemical characterization of a novel plasma membrane-associated, neutral pH optimum α-l-fucosidase from rat testis and epididymal spermatozoa." Biochemical Journal 318, no. 3 (September 15, 1996): 821–31. http://dx.doi.org/10.1042/bj3180821.
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1. Immunocytochemical and biochemical techniques have been used to localize and characterize a novel plasma membrane-associated, neutral-pH-optimum α-l-fucosidase from rat spermatozoa. Light and electron microscopy specifically localized the fucosidase on the plasma membrane of the convex region of the principal segment of testicular and cauda epididymal sperm heads. Immunoreactivity for α-l-fucosidase was also detected in the Golgi apparatus of spermatocytes and spermatids but no immunoreactivity was observed in the acrosome. 2. Fractionation of epididymal sperm homogenates indicated that over 90% of the α-l-fucosidase activity was associated with the 48000 g pellet. This pellet-associated activity could be solubilized with 0.5 M NaCl but not with 0.5% Triton X-100, suggesting that fucosidase is peripherally associated with membranes. Sucrose-density-gradient centrifugation of sperm homogenates indicated that fucosidase was enriched in the plasma membrane-enriched fraction. Analysis of α-l-fucosidase on intact epididymal sperm indicated that the enzyme was active, displayed linear kinetics and had a pH–activity curve (with an optimum near 7) which was comparable to that of fucosidase from epididymal sperm extracts. These results further suggest that fucosidase is associated with plasma membranes, and that its active site is accessible to fucoconjugates. Evidence that most of the fucosidase is associated with the exterior of the plasma membrane came from studies in which intact sperm had fucosidase activity comparable to that of sperm sonicates, and from studies in which approx. 90% of the fucosidase activity on intact sperm could be released from the sperm by gentle shaking with 0.5 M NaCl. Isoelectric focusing indicated that the NaCl-solubilized epididymal sperm fucosidase appears to have one major and one minor isoform with pIs near 7.2 and 5.2, respectively. SDS/PAGE and Western blotting indicated that the NaCl-solubilized extract of epididymal sperm contains two protein bands of 54 and 50 kDa which were highly immunoreactive with the IgG fraction of anti-fucosidase antibodies. Although the function of the novel sperm fucosidase is not known, its specific localization to the plasma membrane of the region of the rat sperm head involved in sperm–egg binding and its high enzymic activity at neutral pH on intact sperm suggest that this enzyme may have a role in sperm–egg interactions.
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Kurz, Anke, Dagmar Viertel, Andreas Herrmann, and Karin Müller. "Localization of phosphatidylserine in boar sperm cell membranes during capacitation and acrosome reaction." Reproduction 130, no. 5 (November 2005): 615–26. http://dx.doi.org/10.1530/rep.1.00561.
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One of the essential properties of mammalian, including sperm, plasma membranes is a stable transversal lipid asymmetry with the aminophospholipids, phosphatidylserine (PS) and phosphatidylethanolamine (PE), typically in the inner, cytoplasmic leaflet. The maintenance of this nonrandom lipid distribution is important for the homeostasis of the cell. To clarify the relevance of lipid asymmetry to sperm function, we have studied the localization of PS in boar sperm cell membranes. By using labeled annexin V as a marker for PS and propidium iodide (PI) as a stain for nonviable cells in conjunction with different methods (flow cytometry, fluorescence and electron microscopy), we have assessed the surface exposure of PS in viable cells during sperm genesis, that is, before and during capacitation as well as after acrosome reaction. An approach was set up to address also the presence of PS in the outer acrosome membrane. The results show that PS is localized in the cytoplasmic leaflet of the plasma membrane as well as on the outer acrosome membrane. Our results further indicate the cytoplasmic localization of PS in the postacrosomal region. During capacitation and acrosome reaction of spermatozoa, PS does not become exposed on the outer surface of the viable cells. Only in a subpopulation of PI-positive sperm cells does PS became accessible upon capacitation. The stable cytoplasmic localization of PS in the plasma membrane, as well as in the outer acrosome membrane, is assumed to be essential for a proper genesis of sperm cells during capacitation and acrosome reaction.
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Oldenhof, Harriëtte, Anna Heutelbeck, Anne-Kathrin Blässe, Heinrich Bollwein, Gunilla Martinsson, Willem F. Wolkers, and Harald Sieme. "Tolerance of spermatozoa to hypotonic stress: role of membrane fluidity and correlation with cryosurvival." Reproduction, Fertility and Development 27, no. 2 (2015): 285. http://dx.doi.org/10.1071/rd13177.
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The aim of this study was to evaluate inter-individual variability in osmotic properties of stallion spermatozoa and its correlation with cryosurvival. In addition, temperature dependency of hypo-osmotic tolerance and membrane fluidity were studied. Stallion sperm membranes exhibited good resistance towards hypotonic stress in the 15–30°C temperature range, whereas membrane stability was found to be decreased at 4 and 37°C. Bull spermatozoa showed greater hypo-osmotic tolerance compared with stallion spermatozoa, especially at temperatures above 30°C, which coincided with decreased membrane fluidity of bovine spermatozoa in this temperature range. The critical osmolality at 22°C, at which half of the sperm population survived exposure to hypotonic saline solution, was found to vary between 55 and 170 mOsm kg–1 among different stallions. Clear correlations were found for pre- versus post-freeze sperm motility and membrane integrity. Pre-freeze percentages of membrane-intact spermatozoa after exposure to hypotonic stress showed a weak correlation with sperm motility after cryopreservation. This correlation, however, was not found when data were corrected for initial numbers of membrane-intact spermatozoa in the sample. We thus conclude that studies on pre-freeze tolerance towards hypotonic stress cannot be used to predict sperm cryosurvival rates for individual stallions.
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KRAWCZYK, ALEKSANDRA, and JADWIGA JAWORSKA-ADAMU. "Activation of sperm in the female reproductive tract in mammals." Medycyna Weterynaryjna 76, no. 09 (2020): 6445–2020. http://dx.doi.org/10.21521/mw.6445.
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The formation of a new diploidal organism is preceded by a series of mutual interactions of haploidal gametes. This process is very complicated and requires the prior activation of reproductive cells. Male gametes eventually mature in the female reproductive tract, acquiring mobility and fertilization. This process takes place in two stages. Sperms are first capacitated. This phenomenon is reversible and leads to structural, cytophysiological and biochemical changes in the sperm plasma membrane as well as to the sperm hyperactivation. Then, due to the contact with the zona pellucida of the oocyte, the irreversible acrosome reaction occurs. This process involves the fusion of the sperm plasma membrane with the outer membrane of the acrosome, the release of enzymes and exposure of the inner acrosome membrane. This enables sperm to penetrate towards the perivitelline space and oolemma. Contact with the oocyte initiates a series of interactions leading to egg activation and the fusion of gametes. Each of these stages involves many different factors that result in the recognition, attraction and adhesion of reproductive cells. Knowledge about the activation mechanisms can improve the effectiveness of supported and controlled reproduction techniques.
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Mendeluk, Gabriela Ruth, Mariano Isaac Cohen, Carla Ferreri, and Chryssostomos Chatgilialoglu. "Nutrition and Reproductive Health: Sperm versus Erythrocyte Lipidomic Profile andω-3 Intake." Journal of Nutrition and Metabolism 2015 (2015): 1–8. http://dx.doi.org/10.1155/2015/670526.
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Fatty acid analyses of sperm and erythrocyte cell membrane phospholipids in idiopathic infertile patients evidenced that erythrocyte contents of EPA, DHA, omega-6–omega-3 ratio and arachidonic acid provide a mathematical correspondence for the prediction of EPA level in sperm cells. The erythrocyte lipidomic profile of patients was significantly altered, with signatures of typical Western pattern dietary habits and no fish intake. A supplementation with nutritional levels of EPA and DHA and antioxidants was then performed for 3 months, with the follow-up of both erythrocyte and sperm cell membranes composition as well as conventional sperm parameters. Some significant changes were found in the lipidomic membrane profile of erythrocyte but not in sperm cells, which correspondently did not show significant parameter ameliorations. This is the first report indicating that membrane lipids of different tissues do not equally metabolize the fatty acid elements upon supplementation. Molecular diagnostic tools are necessary to understand the cell metabolic turnover and monitor the success of nutraceuticals for personalized treatments.
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Yang, Kang, Na Wang, Hai-Tao Guo, Jing-Ran Wang, Huan-Huan Sun, Liang-Zhen Sun, Shun-Li Yue, and Jia-Bo Zhou. "Effect of L-carnitine on sperm quality during liquid storage of boar semen." Asian-Australasian Journal of Animal Sciences 33, no. 11 (November 1, 2020): 1763–69. http://dx.doi.org/10.5713/ajas.19.0455.
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Objective: This study was conducted to investigate the effect of L-carnitine on the pig semen characteristics during storage.Methods: Spermatozoa samples were examined for spermatozoa quality and then randomly divided into 5 groups: 0 (control), 12.5, 25, 50, and 100 mM L-carnitine. Sperm motility, plasma membrane integrity and antioxidant parameters (total reactive oxygen species, total antioxidant capacity, and malondialdehyde) were evaluated after 0, 3, 5, and 10 day cooledstorage at 17°C. Moreover, ATP content, mitochondria activity as well as sperm-binding and in vitro fertilizing ability of preserved boar sperm were also investigated.Results: Supplementation with 50 mM L-carnitine could effectively maintain boar sperm quality parameters such as sperm motility and membrane integrity. Besides, we found that L-carnitine had positive effects on boar sperm quality mainly through improving antioxidant capacities and enhancing ATP content and mitochondria activity. Interestingly, by assessing the effect of L-carnitine on sperm fertility and developmental potential, we discovered that the extender containing L-carnitine could improve sperm quality and increase the number of sperms bounding to zona pellucida, without improving in vitro fertility and development potential.Conclusion: These findings suggested that the proper addition of L-carnitine to the semen extender improved boar sperm quality during liquid storage at 17°C.
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De Toni, Luca, Iva Sabovic, Vincenzo De Filippis, Laura Acquasaliente, Daniele Peterle, Diego Guidolin, Stefania Sut, Andrea Di Nisio, Carlo Foresta, and Andrea Garolla. "Sperm Cholesterol Content Modifies Sperm Function and TRPV1-Mediated Sperm Migration." International Journal of Molecular Sciences 22, no. 6 (March 18, 2021): 3126. http://dx.doi.org/10.3390/ijms22063126.
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Transient receptor potential channels-vanilloid receptor 1 (TRPV1) regulates thermotaxis in sperm-oriented motility. We investigated the role of membrane cholesterol (Chol) on TRPV1-mediated human sperm migration. Semen samples were obtained from five normozoospemic healthy volunteers. Sperm membrane Chol content, quantified by liquid chromatography-mass spectrometry, was modified by incubating cells with 2-hydroxypropyl-ß-cyclodextrin (CD) or the complex between CD and Chol (CD:Chol). The effect on sperm migration on a 10 μM capsaicin gradient (CPS), a TRPV1 agonist, was then investigated. Motility parameters were evaluated by Sperm Class Analyser. Intracellular calcium concentration and acrosome reaction were measured by staining with calcium orange and FITC-conjugated anti-CD46 antibody, respectively. TRPV1-Chol interaction was modelled by computational molecular-modelling (MM). CD and CD:Chol, respectively, reduced and increased membrane Chol content in a dose-dependent manner, resulting in a dose-dependent increase and reduction of sperm migration in a CPS gradient. MM confirmed a specific interaction of Chol with a TRPV1 domain that appeared precluded to the Chol epimer epicholesterol (Epi-Chol). Accordingly, CD:Epi-Chol was significantly less efficient than CD:Chol, in reducing sperm migration under CPS gradient. Chol inhibits TRPV1-mediated sperm function by directly interacting with a consensus sequence of the receptor.
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Ariyanto, Kurnia Bagus, Lilis Khotijah, Dewi Apri Astuti, Raden Iis Arifiantini, and Jean-Baptise Menassol. "Semen Quality of Garut Rams feed by Different Protein Sources and Their Implementation Potential in Small Farms of West Java." Jurnal Agripet 20, no. 1 (April 1, 2020): 47–55. http://dx.doi.org/10.17969/agripet.v20i1.15391.
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ABSTRACT. Maggot Hermetia illucens (Maggot Black Soldier Fly, MBSF) is an alternative protein source besides soybean meal (SBM) which may be used as a feed for improving the quality of semen particularly in Garut rams to support prolific nature. The aims of this study were to analyzed and compare the impact of different protein sources in feed on semen quality of Garut rams, and to assess the prediction ability of Garut rams to serve ewe in small-scale breeders in West Java, Indonesia. This study was conducted using a completely randomized design with 3 treatments and 4 replications, consisted of Brachiaria humidicola (BH) grass and T1 (concentrate contains 20% of SBM), T2 (concentrate contains 10% of SBM and 10% of MBSF), and T3 (concentrate contains 20% of MBSF). The parameters measured were feed consumption, semen quality (macroscopic and microscopic characteristics), also a potential ability of rams to serve ewe. The results showed there were no significant effect on protein consumption, semen volume, semen pH, semen color and consistency, sperm mass movement, sperm motility, sperm concentration, sperm morphology, and prediction potential ability to serve ewe. However, the result showed a significant effect (P0.05) on sperm viability and sperm plasma membrane integrity. Sperm plasma membrane integrity of ram feed with T3 was better than T1 and T2 (P0.05). The prediction potential ability rams to serve ewes on MBSF treatment was 38 heads, while in T1 and T2 were 43 and 57 heads, respectively. In conclusion, MBSF can be an alternative source of protein besides SBM to improve the semen quality of Garut rams. ABSTRAK. Maggot Hermetia illucens (Maggot Black Soldier Fly; MBSF) adalah sumber protein alternatif selain bungkil kedelai (SBM) yang dapat dipergunakan sebagai pakan untuk memperbaiki kualitas semen terutama pada domba Garut untuk mendukung sifat prolifik. Tujuan penelitian ini adalah menganalisis dan membandingkan dampak pemberian sumber protein berbeda terhadap kualitas semen domba Garut dan untuk menilai kemampuan domba Garut pejantan dalam melayani betina pada peternakan rakyat di Jawa Barat, Indonesia. Penelitian ini menggunakan Rancangan Acak Lengkap (RAL) dengan 3 perlakuan dan 4 ulangan yang terdiri dari rumput Brachiaria humidicola (BH) dan T1 (konsentrat mengandung 20% SBM), T2 (konsentrat mengandung 10% SBM dan 10% MBSF), dan T3 (konsentrat mengandung 20% MBSF). Parameter yang diukur adalah konsumsi pakan, karakteristik semen (makroskopis dan mikroskopis) serta potensi domba jantan melayani betina. Hasil Penelitian menunjukkan tidak ada perbedaan signifikan pada konsumsi protein pakan, volume semen, pH semen, warna dan konsistensi semen, gerakan massa sperma, motilitas sperma, konsentrasi sperma, morfologi sperma, dan prediksi potensi pejantan dalam melayani betina. Namun, hasil penelitian menunjukkan terdapat perbedaan (P0.05) pada viabilitas sperma dan membran plasma utuh sperma. Membran plasma utuh pada perlakuan T3 lebih baik dibandingkan perlakuan T1 dan T2 (P0.05). Prediksi potensi betina terlayani dari pejantan yang diberi pakan MBSF adalah 38 ekor, sedangkan yang diberi SBM dan kombinasinya adalah 43 dan 57 ekor. Kesimpulan penelitian ini adalah MBSF dapat menjadi alternatif sumber protein selain bungkil kedelai dalam memperbaiki kualitas sperma domba Garut.
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Hemachand, Tummala, Bagavathi Gopalakrishnan, Dinakar M. Salunke, Satish M. Totey, and Chandrima Shaha. "Sperm plasma-membrane-associated glutathione S-transferases as gamete recognition molecules." Journal of Cell Science 115, no. 10 (May 15, 2002): 2053–65. http://dx.doi.org/10.1242/jcs.115.10.2053.
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Glutathione S-transferases (GSTs) are enzymes that detoxify electrophilic compounds. Earlier studies from our laboratory showed that anti-GST antibodies interfered with the fertilising ability of spermatozoa from Capra hircus (goat) in vitro, suggesting that GSTs are localised at the cell surface. In this study, we provide evidence for the presence of GSTs of 24 kDa on the sperm plasma membrane attached by non-covalent interactions. The GST activity associated with the spermatozoal plasma membrane was significantly higher than the activity present in the plasma membranes of brain cells,hepatocytes, spleenocytes and ventriculocytes. Analysis of GST isoforms demonstrates the presence of GST Pi and Mu on the sperm plasma membranes. Both isoforms were able to bind to solubilised as well as intact zona pellucida(ZP) through their N-terminal regions but failed to bind to ZP once the oocytes were fertilised. Solubilised goat ZP separates into three components,one of which, the ZP3-like component, bound to sperm GSTs. High concentrations of anti-GST antibodies or solubilised ZP led to aggregation of sperm GSTs,resulting in the release of acrosin. In contrast, inhibition of sperm GST binding to ZP, by saturation of binding sites for sperm GSTs on the solubilised ZP using peptides designed from the N-terminii of GST Pi or Mu or blocking of binding sites for ZP on sperm GSTs with antibodies raised against the N-terminal GST peptides, inhibited essential prefertilisation changes in sperm. These data therefore demonstrate the strategic location of catalytically active defensive enzymes on the sperm surface that also act as zona-binding proteins. Therefore, sperm-surface GSTs serve as bifunctional molecules in a transcriptionally inactive cell whose requirement for cellular defense and economy of molecules that it can carry is greater than that of any somatic cell type.
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Lopez, L. C., and B. D. Shur. "Redistribution of mouse sperm surface galactosyltransferase after the acrosome reaction." Journal of Cell Biology 105, no. 4 (October 1, 1987): 1663–70. http://dx.doi.org/10.1083/jcb.105.4.1663.
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Gamete recognition in the mouse is mediated by galactosyltransferase (GalTase) on the sperm surface, which binds to its appropriate glycoside substrate in the egg zona pellucida (Lopez, L. C., E. M. Bayna, D. Litoff, N. L. Shaper, J. H. Shaper, and B. D. Shur, 1985, J. Cell Biol., 101:1501-1510). GalTase has been localized by indirect immunofluorescence to the dorsal surface of the anterior sperm head overlying the intact acrosome. Sperm binding to the zona pellucida triggers induction of the acrosome reaction, an exocytotic event that results in vesiculation and release of the outer acrosomal and overlying plasma membranes. Consequently, we examined the fate of sperm surface GalTase after the acrosome reaction. Contrary to our expectations, surface GalTase is not lost during the acrosome reaction despite the loss of its membrane domain. Rather, double-label indirect immunofluorescence assays show that GalTase is redistributed to the lateral surface of the sperm, coincident with the acrosome reaction. This apparent redistribution of GalTase was confirmed by direct enzymatic assays, which show that 90% of sperm GalTase activity is retained during the acrosome reaction. No GalTase activity is detectable on plasma membrane vesicles released during the acrosome reaction. In contrast, removal of plasma membranes by nitrogen cavitation releases GalTase activity from the sperm surface, showing that GalTase redistribution requires a physiological acrosome reaction. The selective redistribution of GalTase to a new membrane domain from one that is lost during the acrosome reaction suggests that GalTase is repositioned for some additional function after initial sperm-zona binding.
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SACCHI, L., S. CORONA, M. CASIRAGHI, and C. BANDI. "Does fertilization in the filarial nematode Dirofilaria immitis occur through endocytosis of spermatozoa?" Parasitology 124, no. 1 (January 2002): 87–95. http://dx.doi.org/10.1017/s0031182001008964.
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Information on the ultrastructural details of fertilization in filarial nematodes are still unavailable. Here we report new data on this process in Dirofilaria immits, the heartworm of dogs and cats. Electron microscopy allowed us to observe oocytes engulfing spermatozoa through an endocytosis-like process. We also observed spermatozoa inside the oocytes which still possessed their plasma membrane and which were clearly enveloped by a further membrane, likely derived from the endocytosis process. At this stage, at the interface between the sperm membrane and the endocytotic membrane (vacuolar space), we observed flocculent material in the proximity of the membranous organelles (MOs) of the sperm. In the proximity of the MOs, we also observed the enlargement of the vacuolar space. Other images showed the dissolution of the sperm membrane, and the release of nuclear masses and organelles in the egg cytoplasm. We did not observe the fusion of lysosomes to the endocytotic vacuoles. In addition, the lysis of the sperm organelles has never been observed inside the vacuoles containing the whole sperm. Thus we suggest that the degradation of the endocytotic and sperm plasma membranes is determined by material released by the MOs. Since we did not observe the entry of sperm into the oocytes by other mechanisms, we also suggest that endocytosis is the normal process used by the spermatozoon to get into the egg cytoplasm in D. immitis. Finally, during our observations of the seminal receptacle we did not observe any structure in the spermatozoa which could be interpreted as an intracellular bacterium. This is consistent with previous results indicating that the bacterium Wolbachia in filarial nematodes is not transmitted through the sperm.
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Prasinou, Paraskevi, Ippolito De Amicis, Isa Fusaro, Roberta Bucci, Damiano Cavallini, Salvatore Parrillo, Maurizio Caputo, Alessandro Gramenzi, and Augusto Carluccio. "The Lipidomics of Spermatozoa and Red Blood Cells Membrane Profile of Martina Franca Donkey: Preliminary Evaluation." Animals 13, no. 1 (December 20, 2022): 8. http://dx.doi.org/10.3390/ani13010008.
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Fatty acid-based lipidomic analysis has been widely used to evaluate health status in human medicine as well as in the veterinary field. In equine species, there has been a developing interest in fertility and sperm quality. Fatty acids, being the principal components of the membranes, play an active role in the regulation of the metabolic activities, and their role on spermiogenesis seems to be of great importance for the resulting quality of the sperm and, thus, fertility. With the application of widely used lipidomic techniques, the aim of this study was to evaluate: a) the fatty acid content of the spermatozoa’s membranes of 26 healthy male Martina Franca donkeys and its possible correlation with sperm parameters, and b) the evaluation of the composition of the red blood cells’ membrane. PUFA omega-6 are the principal components (40.38%) of the total PUFA content (47.79%) in both types of cells; however, DPA is the predominant one on the spermatozoa’s membrane (27.57%) but is not present in the erythrocyte’s membrane. Spermatozoa’s motility (%) is positively correlated with stearic acid and EPA, and progressive motility (%), with oleic acid. These findings offer information on the composition of both types of cells’ membranes in healthy male MF donkeys and reflect the metabolic transformations of the spermatozoa’s membrane during the maturation period, providing a better perception of the role of fatty acids in sperm parameters and fertility.
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Sasanami, Tomohiro, Norio Yoshizaki, Hideo Dohra, and Hideo Kubo. "Sperm acrosin is responsible for the sperm binding to the egg envelope during fertilization in Japanese quail (Coturnix japonica)." REPRODUCTION 142, no. 2 (August 2011): 267–76. http://dx.doi.org/10.1530/rep-11-0120.
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An antibody library against quail sperm plasma membrane components was established and a mAb, which strongly inhibits sperm perforations of the perivitelline membrane (PVM) was obtained from the library. The antigen molecule of the mAb showed an apparent molecular weight of 45 kDa, and was distributed both on the surface and in the acrosomal matrix of the sperm head. Periodate oxidation revealed that the epitope of the antigen includes a sugar moiety. Tandem mass spectrometry analysis of the antigen revealed that the mAb recognizes sperm acrosin. When sodium dodecyl sulfate-solubilized PVM immobilized on a polyvinylidene difluoride membrane was incubated with sperm plasma membrane lysates, the sperm acrosin was detected on the PVM immobilized on the membrane, indicating that the sperm acrosin interacts with the components of PVM. Indeed, the mAb effectively inhibited the binding of acrosome-intact sperm to the PVM. These results indicate that the 45 kDa sperm acrosin is involved in the binding of sperm to the PVM in fertilization of Japanese quail.
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Zhuang, Dazhong, Xiangfen Song, Guojun Hu, Qingyuan Sun, and Dayuan Chen. "Sperm antigen MSH27 participates in sperm-egg membrane fusion." Chinese Science Bulletin 44, no. 16 (August 1999): 1483–89. http://dx.doi.org/10.1007/bf03183568.
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FRIEND, DANIEL S. "Sperm Maturation: Membrane Domain Boundaries." Annals of the New York Academy of Sciences 567, no. 1 Viral Oncogen (August 1989): 208–21. http://dx.doi.org/10.1111/j.1749-6632.1989.tb16472.x.
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Lagares, M. A., R. Petzoldt, H. Sieme, and E. Klug. "Assessing equine sperm-membrane integrity." Andrologia 32, no. 3 (June 2000): 163–67. http://dx.doi.org/10.1046/j.1439-0272.2000.00351.x.
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WANG, L. F., Y. C. YAN, and S. S. KOIDE. "Immunobiology of Sperm Membrane Protiens." Development, Growth and Differentiation 28, s1 (August 1986): 31–32. http://dx.doi.org/10.1111/j.1440-169x.1986.28s1_31.x.
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Lagares, M. A., R. Petzoldt, H. Sieme, and E. Klug. "Assessing equine sperm-membrane integrity." Andrologia 32, no. 3 (April 24, 2009): 163–67. http://dx.doi.org/10.1111/j.1439-0272.2000.tb02881.x.
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Moran, S. P., T. Chi, M. S. Prucha, H. R. Engelhardt, A. Yuksel, and A. W. S. Chan. "61 TRANSGENIC HUNTINGTON'S DISEASE MONKEY SPERM HAS A LOWER CRYOTOLERANCE." Reproduction, Fertility and Development 26, no. 1 (2014): 144. http://dx.doi.org/10.1071/rdv26n1ab61.
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Cryopreservation is an important tool routinely used for preserving sperm for artificial reproductive technologies (ART), as well as genetic preservation of unique animal models. The cryopreservation process is harsh and detrimental to the fragile gametes, and damage to the sperm is not only known, but inevitable. This study presents new data in which sperm from 3 transgenic Huntington's disease (HD) monkeys (rhesus macaques) are compared with 3 wild-type (WT) rhesus sperm donors. Currently, there are no data comparing HD versus WT sperm viability and cryotolerance in humans. The goal of this study was to investigate differences between fresh and frozen semen by quantitative analysis on sperm viability based on (1) motility, (2) membrane integrity, and (3) acrosome integrity. Sperm motility was determined by visual evaluation. Membrane and acrosome integrity were assessed simultaneously by Hoechst 33342, propidium iodide (PI), and fluorescein isothiocyanate-peanut agglutinin (FITC-PNA) triple staining. Sperm viability analysis was divided into 3 groups: (1) fresh HD versus fresh WT, (2) fresh versus cryopreserved-thawed WT, (3) and fresh HD versus cryopreserved-thawed HD sperm. Interestingly, fresh HD sperm had a lower percentage of membrane-damaged cells (38.57 ± 3.15) compared with WT (49.67 ± 3.56; P < 0.03). However, after cryopreservation and subsequent thawing, HD sperm had a significantly higher percentage increase in damaged membranes than WT sperm (27.91 ± 2.93 v. 8.27 ± 8.28; P < 0.001), respectively. No significant difference in acrosome damage between groups was identified in either fresh or cryopreserved sperm populations. Motility significantly declined in both cryopreserved populations [HD: 89.7 to 43.4% (P < 0.001) and WT: 90.0 to 45.6% (P < 0.001)]. There was no significant difference between either freeze-thawed group. These data illustrate that HD sperm have a lower cryotolerance than WT sperm. Our findings suggest that the optimization of the HD sperm cryopreservation method and investigation on biochemical differences (e.g. membrane lipid composition) are necessary to improve post-thaw survival. This in turn is important for the establishment of a sperm cryobank and future derivation of a unique animal model such as HD monkey. Our study also suggests that HD monkey could be a useful model for optimizing cryopreservation method for HD patients.
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Yeste, Marc, Sandra Recuero, Carolina Maside, Albert Salas-Huetos, Sergi Bonet, and Elisabeth Pinart. "Blocking NHE Channels Reduces the Ability of In Vitro Capacitated Mammalian Sperm to Respond to Progesterone Stimulus." International Journal of Molecular Sciences 22, no. 23 (November 23, 2021): 12646. http://dx.doi.org/10.3390/ijms222312646.
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Few data exist about the presence and physiological role of Na+/H+ exchangers (NHEs) in the plasma membrane of mammalian sperm. In addition, the involvement of these channels in the ability of sperm to undergo capacitation and acrosomal reaction has not been investigated in any mammalian species. In the present study, we addressed whether these channels are implicated in these two sperm events using the pig as a model. We also confirmed the presence of NHE1 channels in the plasma membrane of ejaculated sperm by immunofluorescence and immunoblotting. The function of NHE channels during in vitro capacitation was analyzed by incubating sperm samples in capacitating medium for 300 min in the absence or presence of a specific blocker (DMA; 5-(N,N-dimethyl)-amiloride) at different concentrations (1, 5, and 10 µM); acrosome exocytosis was triggered by adding progesterone after 240 min of incubation. Sperm motility and kinematics, integrity of plasma and acrosome membranes, membrane lipid disorder, intracellular calcium and reactive oxygen species (ROS) levels, and mitochondrial membrane potential (MMP) were evaluated after 0, 60, 120, 180, 240, 250, 270, and 300 min of incubation. NHE1 localized in the connecting and terminal pieces of the flagellum and in the equatorial region of the sperm head and was found to have a molecular weight of 75 kDa. During the first 240 min of incubation, i.e., before the addition of progesterone, blocked and control samples did not differ significantly in any of the parameters analyzed. However, from 250 min of incubation, samples treated with DMA showed significant alterations in total motility and the amplitude of lateral head displacement (ALH), acrosomal integrity, membrane lipid disorder, and MMP. In conclusion, while NHE channels are not involved in the sperm ability to undergo capacitation, they could be essential for triggering acrosome exocytosis and hypermotility after progesterone stimulus.
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Bianchi Rodrigues Alves, M., A. Furugen Cesar de Andrade, R. Paes de Arruda, L. Batissaco, R. Lançoni, M. Andrade Torres, G. Mouro Ravagnani, et al. "265 TESTICULAR DEGENERATION AFFECTED PLASMA, ACROSOMAL AND MITOCHONDRIAL MEMBRANE INTEGRITY, AND DNA FRAGMENTATION IN RAM SPERMATOZOA." Reproduction, Fertility and Development 27, no. 1 (2015): 222. http://dx.doi.org/10.1071/rdv27n1ab265.
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Testicular degeneration, an important cause of male infertility, adversely affects sperm motility and morphology. However, few studies describe effects on integrity of plasma and acrosomal membranes, mitochondrial membrane potential, and DNA fragmentation; therefore, they were evaluated in the present study. Testicular degeneration was induced in 17 White Dorper rams (scrotal insulation for 72 h). Semen was collected (artificial vagina) twice before insulation and twice thereafter (15-day intervals between post-insulation collections). Sperm motility and morphology were analysed by SCA software (Sperm Class Analyser®, MICROPTIC®, Barcelona, Spain) and differential interference contrast microscopy (DIC, model 80i, Nikon, Tokyo, Japan), respectively. Membrane integrity and potential were assessed with fluorescent probes: Hoescht 33342, propidium iodide, FITC-PSA, and JC-1 (Celeghini et al. 2010 Arq. Bras. Med. Vet. Zootec. 62, 536–543) and imaged with fluorescence microscopy (Nikon Model 80i, Nikon, Tokyo, Japan). Fragmentation of DNA was evaluated with a Halomax® kit (Halotech® DNA, Madrid, Spain). Data were analysed with Statview software (Stat View 1998, SAS Institute Inc., Cary, NC, USA). Data obtained from the periods (before × after insulation) were evaluated by analysis of variance (ANOVA) and means were compared using Tukey's test. Total motility (before: 87.53 ± 1.21%; after: 46.53 ± 4.46%) and progressive motility (before: 58.64 ± 2.00%; after: 31.33 ± 3.82%) were reduced (P < 0.01) by scrotal insulation, as were sperm major defects (before: 10.64 ± 1.65%; after: 54.30 ± 3.67%) and total defects (before: 20.50 ± 2.40%; after: 63.85 ± 3.41%; P < 0.0001). Sperm with intact plasma and acrosomal membranes and high mitochondrial potential (PIAIH) decreased (P < 0.0001) after insulation. In that regard, 53.19 ± 2.20 and 28.48 ± 3.48% of sperm were classified as PIAIH before v. after insulation, respectively. Furthermore, plasma membrane integrity, acrosome membrane integrity, and high mitochondrial potential were assessed independently. The quantity of plasma membrane integrity cells (before: 62.01 ± 2.07%; after: 33.92 ± 3.94%), acrosome membrane integrity cells (before: 57.17 ± 2.30%; after: 31.47 ± 3.77%), and high mitochondrial potential cells (before: 85.72 ± 1.42%; after: 57.28 ± 3.12%) were also reduced (P < 0.0001) after insulation. Likewise, DNA integrity decreased (P = 0.002) from 98.87 ± 0.26% before insulation to 91.88 ± 2.6% afterward. In conclusion, sperm plasma and acrosomal membrane integrity, mitochondrial membrane potential, and DNA fragmentation were adversely affected by testicular degeneration in rams induced by scrotal insulation.Research was supported by FAPESP process 2012/00040-0 and 2011/16744-3.
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Gadella, B. M., M. Lopes-Cardozo, L. M. van Golde, B. Colenbrander, and T. W. Gadella. "Glycolipid migration from the apical to the equatorial subdomains of the sperm head plasma membrane precedes the acrosome reaction. Evidence for a primary capacitation event in boar spermatozoa." Journal of Cell Science 108, no. 3 (March 1, 1995): 935–46. http://dx.doi.org/10.1242/jcs.108.3.935.
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In order to extend the static information of immunolabelling sulphogalactolipids in fixed boar spermatozoa, a fluorescent sulphogalactolipid analogue, galactose(3-sulphate)-beta 1–1′[(N-lissamine rhodaminyl)-12-aminodode-canoyl]-sphingosine, was incorporated into plasma membranes of living spermatozoa and its lateral distribution over the sperm head was studied. The fluorescent lipid was enriched in the apical ridge subdomain of freshly ejaculated sperm cells. After sperm binding to the zona pellucida the lipid redistributed to the equatorial segment of the sperm surface. A similar shift occurred during capacitation in vitro with 2 mM CaCl2 or with 4% (w/v) bovine serum albumin. The desulphated derivative galactose-beta 1–1′[(N-lissamine rhodaminyl)-12-aminododecanoyl]-sphingosine was also incorporated into the plasma membrane of freshly ejaculated sperm cells and clearly stained the apical ridge subdomain and the (pre)-equatorial subdomains of the sperm heads. The desulphogalactolipid analogue showed a slightly faster migration to the equatorial segment of the sperm plasma membrane than did its sulphated counterpart. The measured fluorescence intensity distributions correlated linearly with the spatial probe distribution, which was checked by fluorescence lifetime imaging microscopy. The observed migration of the incorporated glycolipids precedes the acrosome reaction and is one of the underlying molecular events likely to be important in the process of sperm capacitation. The results of this study suggest that lipid phase segregation is an important driving force for the organization of the sperm head plasma membrane into subdomains.
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Wysokińska, Anna, and Stanislaw Kondracki. "Assessment of changes in sperm cell membrane integrity occurring during the storage of semen from genetically different males using two diagnostic methods." Canadian Journal of Animal Science 94, no. 4 (December 2014): 601–6. http://dx.doi.org/10.4141/cjas2013-095.
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Wysokińska, A. and Kondracki, S. 2014. Assessment of changes in sperm cell membrane integrity occurring during the storage of semen from genetically different males using two diagnostic methods. Can. J. Anim. Sci. 94: 601–606. The present study was carried out to assess changes in sperm cell membrane integrity occurring during the storage of semen collected from genetically different domestic male pigs. The study was aimed at assessing differences in the course of changes in the integrity of cell membranes in spermatozoa produced by males with different degrees of genetic diversity (pure-bred males, two-breed hybrids and multi-breed crosses) and testing the usefulness of two methods of sperm cell membrane integrity evaluation, based on material collected from genetically different males. The experiments were conducted on 56 ejaculates collected from 28 domestic male pigs. The examination of sperm cell membrane integrity was performed three times for each ejaculate, i.e., after 1 h, after 24 h and after 48 h from collection. The preparations for analysing cell membrane integrity were made using two methods: the SYBR 14/PI method and the eosin–nigrosin method. It was found that both SYBR 14/PI and eosin–nigrosin staining methods make it possible to successfully assess the integrity of the plasma membrane of domestic pig sperm cells under in vitro conditions. Hybrid pig spermatozoa, especially those from multi-breed crosses, better retain the integrity of their plasmalemmas than the spermatozoa of pure-bred boars. The ejaculates of Hypor cross-breed boars assessed after 1, 24 and 48 h of storage contain more spermatozoa with intact cell membranes than the ejaculates of pure-bred Duroc and Pietrain boars. The ejaculates of Hypor boars also show fewer decaying spermatozoa than those produced by pure-bred boars.
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Mortillo, S., and P. M. Wassarman. "Differential binding of gold-labeled zona pellucida glycoproteins mZP2 and mZP3 to mouse sperm membrane compartments." Development 113, no. 1 (September 1, 1991): 141–49. http://dx.doi.org/10.1242/dev.113.1.141.
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Egg zona pellucida glycoproteins mZP3 and mZP2 serve as primary and secondary sperm receptors, respectively, during initial stages of fertilization in mice [Wassarman (1988) A. Rev. Biochem. 57, 415–442]. These receptors interact with complementary egg-binding proteins (EBPs) located on the sperm surface to support species-specific gamete adhesion. Results of whole-mount autoradiographic experiments suggest that purified egg mZP3 and mZP2 bind preferentially to acrosome-intact (AI) and acrosome-reacted (AR) sperm heads, respectively [Bleil and Wassarman (1986) J. Cell Biol. 102, 1363–1371]. Here, we used purified egg mZP2, egg mZP3 and fetuin, which were coupled directly to colloidal gold (‘gold-probes’), to examine binding of these glycoproteins to membrane compartments of AI and AR sperm by transmission electron microscopy. mZP3 gold-probes were found associated primarily with plasma membrane overlying the acrosomal and post-acrosomal regions of AI sperm heads. They were also found associated with plasma membrane overlying the post-acrosomal region of AR sperm heads. mZP2 gold-probes were found associated primarily with inner acrosomal membrane of AR sperm heads, although some gold was associated with outer acrosomal membrane of AI sperm that had holes in plasma membrane overlying the acrosome. Fetuin gold-probes, used to assess background levels of binding, were bound at relatively low levels to plasma membrane and inner acrosomal membrane of AI and AR sperm, respectively. None of the gold-probes exhibited significant binding to sperm tails, or to red blood cells and residual bodies present in sperm preparations. These results provide further evidence that mZP2 and mZP3 bind preferentially to heads of AR and AI sperm, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
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Yuan, Ruiyong, Paul Primakoff, and Diana G. Myles. "A Role for the Disintegrin Domain of Cyritestin, a Sperm Surface Protein Belonging to the ADAM Family, in Mouse Sperm–Egg Plasma Membrane Adhesion and Fusion." Journal of Cell Biology 137, no. 1 (April 7, 1997): 105–12. http://dx.doi.org/10.1083/jcb.137.1.105.
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Sperm–egg plasma membrane fusion is preceded by sperm adhesion to the egg plasma membrane. Cell–cell adhesion frequently involves multiple adhesion molecules on the adhering cells. One sperm surface protein with a role in sperm–egg plasma membrane adhesion is fertilin, a transmembrane heterodimer (α and β subunits). Fertilin α and β are the first identified members of a new family of membrane proteins that each has the following domains: pro-, metalloprotease, disintegrin, cysteine-rich, EGF-like, transmembrane, and cytoplasmic domain. This protein family has been named ADAM because all members contain a disintegrin and metalloprotease domain. Previous studies indicate that the disintegrin domain of fertilin β functions in sperm–egg adhesion leading to fusion. Full length cDNA clones have been isolated for five ADAMs expressed in mouse testis: fertilin α, fertilin β, cyritestin, ADAM 4, and ADAM 5. The presence of the disintegrin domain, a known integrin ligand, suggests that like fertilin β, other testis ADAMs could be involved in sperm adhesion to the egg membrane. We tested peptide mimetics from the predicted binding sites in the disintegrin domains of the five testis-expressed ADAMs in a sperm–egg plasma membrane adhesion and fusion assay. The active site peptide from cyritestin strongly inhibited (80–90%) sperm adhesion and fusion and was a more potent inhibitor than the fertilin β active site peptide. Antibodies generated against the active site region of either cyritestin or fertilin β also strongly inhibited (80–90%) both sperm–egg adhesion and fusion. Characterization of these two ADAM family members showed that they are both processed during sperm maturation and present on mature sperm. Indirect immunofluorescence on live, acrosome-reacted sperm using antibodies against either cyritestin or fertilin β showed staining of the equatorial region, a region of the sperm membrane that participates in the early steps of membrane fusion. Collectively, these data indicate that a second ADAM family member, cyritestin, functions with fertilin β in sperm–egg plasma membrane adhesion leading to fusion.
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Xiao, Yao, Hualin Zhang, Sibtain Ahmad, Liya Bai, Xiaomin Wang, Lijun Huo, Xin Zhang, Wengong Li, Xiang Li, and Liguo Yang. "Sperm capacitation combined with removal of the sperm acrosome and plasma membrane enhances paternal nucleus remodelling and early development of bovine androgenetic embryos." Reproduction, Fertility and Development 25, no. 4 (2013): 624. http://dx.doi.org/10.1071/rd12075.
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The androgenetic embryo is a useful model for functional analysis of the paternal genome during embryogenesis. However, few studies have focused on the factors involved in the suppressed developmental competence of such embryos or why sperm cloning-derived androgenetic embryos fail to develop beyond the morula stage in large domestic animals. To overcome this developmental failure, we tried to improve sperm decondensation, as well as to enhance embryonic development by sperm capacitation and removal of the acrosome and plasma membrane before injection of the spermatozoa. Before injection of the spermatozoa, we quantified the effects of sperm capacitation combined with sperm pretreatment on the acrosome and plasma membrane status. We also evaluated sperm decondensation potential, sperm viability and chromatin integrity. Immunostaining data showed that the sperm acrosome and plasma membrane could be more efficiently removed after capacitation. Dithiothreitol-induced sperm decondensation potential was improved with capacitation and removal of the acrosome and plasma membrane. Although most spermatozoa lost viability after pretreatment, their chromatin remained integrated. The patterns of paternal chromatin remodelling within uncleaved androgenetic embryos and the nucleus morphology of cleaved embryos indicated that capacitation combined with membrane disruption could make injected spermatozoa decondense synchronously not only with each other, but also with the developmental pace of the ooplasm. We successfully produced androgenetic blastocysts, and efficiency increased with sperm pretreatment. In conclusion, sperm decondensation and the early development of androgenetic embryos were enhanced with sperm capacitation and removal of the acrosome and plasma membrane prior to sperm injection.
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Boitano, S., and C. K. Omoto. "Membrane hyperpolarization activates trout sperm without an increase in intracellular pH." Journal of Cell Science 98, no. 3 (March 1, 1991): 343–49. http://dx.doi.org/10.1242/jcs.98.3.343.
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Sperm from trout, like other sperm, are immotile in the seminal tract and initiate motility upon dilution into an appropriate fertilizing environment. Trout sperm motility is inhibited by high extracellular [K+] and can be activated by dilution of extracellular [K+]. Activation of trout sperm by the dilution of extracellular [K+] suggests regulation by membrane potential. Using the membrane potential-sensitive fluorescent dye 3,3′-dipropylthiocarbocyanine iodide (diS-C3-(5)) we directly measured the K+ contribution to the membrane potential. Manipulating the membrane potential with Cs+ and the ionophore valinomycin can override K+ regulation. We show that trout sperm can also be activated in the presence of inhibitory [K+] by the addition of divalent cations. Activation by divalent cations is explained by the cations' ability to mask membrane surface potential and thus alter the potential sensed by membrane voltage sensors. Using the surface potential-sensitive dye, 1-anilino-8-naphthosulfonate (ANS), we directly measure the divalent cations' ability to mask surface potential. We propose a model where membrane hyperpolarization is the trigger that initiates the cascade of events leading to trout sperm activation. An increase in intracellular pH has been suggested to be a conserved step in the activation of sperm motility. We show that increasing intracellular pH by procedures that activate sea urchin and mammalian sperm does not activate trout sperm. In contrast, there is a decrease in intracellular pH upon activation of trout sperm motility. Artificially decreasing intracellular pH is not sufficient for activation of motility in trout sperm in an inhibitory [K+]. Thus, unlike some other sperm, changes in intracellular pH do not regulate trout sperm motility.
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Noda, Taichi, Yonggang Lu, Yoshitaka Fujihara, Seiya Oura, Takayuki Koyano, Sumire Kobayashi, Martin M. Matzuk, and Masahito Ikawa. "Sperm proteins SOF1, TMEM95, and SPACA6 are required for sperm−oocyte fusion in mice." Proceedings of the National Academy of Sciences 117, no. 21 (May 11, 2020): 11493–502. http://dx.doi.org/10.1073/pnas.1922650117.
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Sperm−oocyte membrane fusion is one of the most important events for fertilization. So far, IZUMO1 and Fertilization Influencing Membrane Protein (FIMP) on the sperm membrane and CD9 and JUNO (IZUMO1R/FOLR4) on the oocyte membrane have been identified as fusion-required proteins. However, the molecular mechanisms for sperm−oocyte fusion are still unclear. Here, we show that testis-enriched genes, sperm−oocyte fusion required 1 (Sof1/Llcfc1/1700034O15Rik), transmembrane protein 95 (Tmem95), and sperm acrosome associated 6 (Spaca6), encode sperm proteins required for sperm−oocyte fusion in mice. These knockout (KO) spermatozoa carry IZUMO1 but cannot fuse with the oocyte plasma membrane, leading to male sterility. Transgenic mice which expressed mouseSof1,Tmem95,andSpaca6rescued the sterility ofSof1,Tmem95, andSpaca6KO males, respectively. SOF1 and SPACA6 remain in acrosome-reacted spermatozoa, and SPACA6 translocates to the equatorial segment of these spermatozoa. The coexpression of SOF1, TMEM95, and SPACA6 in IZUMO1-expressing cultured cells did not enhance their ability to adhere to the oocyte membrane or allow them to fuse with oocytes. SOF1, TMEM95, and SPACA6 may function cooperatively with IZUMO1 and/or unknown fusogens in sperm−oocyte fusion.
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Sarkar, Madhumita, Gopal C. Majumder, and Tapati Chatterjee. "Goat sperm membrane: lectin-binding sites of sperm surface and lectin affinity chromatography of the mature sperm membrane antigens." Biochimica et Biophysica Acta (BBA) - Biomembranes 1070, no. 1 (November 1991): 198–204. http://dx.doi.org/10.1016/0005-2736(91)90164-4.
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Gadea, J., F. García-Vázquez, and C. Matás. "292CHANGES IN MEMBRANE SULFHYDRYL STATUS OF BOAR SPERMATOZOA BY FREEZING." Reproduction, Fertility and Development 16, no. 2 (2004): 265. http://dx.doi.org/10.1071/rdv16n1ab292.
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The processes of cooling and freezing/thawing produce physical and chemical stresses on the sperm membrane that reduce the viability and fertilizing capacities. The cold shock and freezing of spermatozoa are associated with an oxidative stress, with reactive oxygen species (ROS) generation (Chatterjee et al., 2001, Mol. Reprod. Dev. 60, 498–506) and with a significant reduction of the GSH content (Gadea et al., in press). In the processes of capacitation, fertilization and freezing, qualitative and quantitative changes in protein membrane composition occurs, including changes in distribution of sulphydryl groups on the sperm membrane. The aim of this work was to evaluate the changes in the sulfhydryl groups of proteins from the sperm surface after cooling and freezing procedures as a marker of membrane changes. Ejaculate-rich fractions from three mature Pietrain boars were diluted in Beltsville Thaw Solution (BTS) extender and cooled to 15°C over 2h (control). Thereafter sperm were centrifuged and diluted in lactose/egg-yolk extender cooled to 5°C over 2h and later frozen with glycerol and equex by classic methodology (Westendorf et al., 1975, Dtsch. Tierärztl. Wschr. 82, 261–267). Sperm parameters were measured in extended semen (control) at 0, 1 and 2h after cold shock at 5°C and after freezing-thawing. The structure of the sperm membrane was evaluated with carboxyfluorescein diacetate/propidium iodide (DCF) (Harrison and Vickers, 1990, J. Reprod. Fert. 88, 343–352), and the sulfhydryl status of proteins from spermatozoa surface are evaluated with fluorescent-staining 5-iodoacetamidofluoresceine (5-IAF) and by acrosome integrity (normal apical ridge, NAR). Some seminal parameters to evaluate functionality such as motility (MOT), forward progressive motility (FPM, 0–5), and mitochondria activity with Rhodamine 123(MIT) were also evaluated. Data from 11 freezing batches were analyzed by one-way ANOVA. When ANOVA revealed a significant effect, values were compared by the Tukey test. The freezing process significantly affected all the sperm parameters studied. Motility was negatively affected from the onset of cooling to 5°C. However, DCF, NAR and 5-IAF were only affected after freezing process. Mitochondria activity decreased in the last period of the cooling procedure (2h) and it was lower after freezing. An inverse significant relation was found between 5-IAF and motility, viability, NAR and mitochondria functionality (P<0.01). These results show that freezing damage produces an alteration in the structure of the sperm membranes (DCF, NAR, 5-IAF) and sperm functionality (motility and mitochondrial). However, only motility (MOT and FPM) was affected by cold shock when lactose/egg-yolk extender was used. Previous studies of cold shock with no cryo-protective medium (BTS) showed a marked effect on sulphydryl membrane characteristics (Marco and Gadea, 2003). These preliminary results in the use of 5-IAF in boar semen showed that freezing produces an alteration in the structure of the sperm membranes, which could be detected by simple fluorescent staining. This research was supported by grant AGL 2000-0485-C02-01.
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Arnoult, C., Y. Zeng, and H. M. Florman. "ZP3-dependent activation of sperm cation channels regulates acrosomal secretion during mammalian fertilization." Journal of Cell Biology 134, no. 3 (August 1, 1996): 637–45. http://dx.doi.org/10.1083/jcb.134.3.637.
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The sperm acrosome reaction is a Ca(2+)-dependent secretory event required for fertilization. Adhesion to the egg's zona pellucida promotes Ca2+ influx through voltage-sensitive channels, thereby initiating secretion. We used potentiometric fluorescent probes to determine the role of sperm membrane potential in regulating Ca2+ entry. ZP3, the glycoprotein agonist of the zona pellucida, depolarizes sperm membranes by activating a pertussis toxin-insensitive mechanism with the characteristics of a poorly selective cation channel. ZP3 also activates a pertussis toxin-sensitive pathway that produces a transient rise in internal pH. The concerted effects of depolarization and alkalinization open voltage-sensitive Ca2+ channels. These observations suggest that mammalian sperm utilize membrane potential-dependent signal transduction mechanisms and that a depolarization pathway is an upstream transducing element coupling adhesion to secretion during fertilization.