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Sperm Cells (sperm + cell)
Selected AbstractsVom Samentier zur Samenzelle: Die Neudeutung der Zeugung im 19.BERICHTE ZUR WISSENSCHAFTSGESCHICHTE, Issue 3 2009Jahrhundert Zeugungstheorien; Vererbung; Samentiere; Samenzellen; Physiologie des 19. Jahrhunderts; Reproduktionstechnologien; Körper- und Geschlechtergeschichte; Mikro- und Makrogeschichte Abstract From Spermatic Animalcules to Sperm Cells: The Reconceptualization of Generation in the 19th Century. At the end of the 18th and still at the beginning of the 19th century most naturalists considered spermatic animalcules to be parasites of the seminal fluid that played no role in procreation. This view was progressively questioned by 19th century physiologists. They gradually redefined the spermatic animals as (cellular) products of the male organism, as agents of fertilization and bearers of the male heredity material. This article discusses this change from two different perspectives: on a microhistorical level, it analyzes the experimental research of the naturalist Lazzaro Spallanzani (1729,1799) and of the physiologist Albert Kölliker (1817,1905) in order to show how spermatozoa were turned into a new epistemic object of biology , the sperm cell. Further, it asks how the role of the reconceptualization of spermatic animalcules affected the long-term transformations that gave rise of our modern understanding of heredity, generation and the sexed body. By combining these two perspectives, the article aims to connect historiographies that are often kept separate: the macrohistorical narratives about gender and the body in the modern age and the microhistorical studies of biomedical practices and objects. [source] Toxicity of tributyltin and triphenyltin to early life-stages of Paracentrotus lividus (Echinodermata: Echinoidea)ENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 4 2002Alessandra Arizzi Novelli Abstract Sperm cell and embryo toxicity tests using the Mediterranean sea urchin Paracentrotus lividus were performed to assess the toxicity of tributyltin chloride, bis(tributyltin)oxide, triphenyltin acetate, and triphenyltin hydroxide. Toxicity values (mean effective concentration [EC50]) ranged from 2.97 to 18.5 ,g/L for sperm cells and from 1.11 to 2.62 ,g/L for embryos. For sperm cells, the toxicity of the two tributyl compounds was significantly greater than that of two triphenyl compounds; for embryos, the triphenyl compounds appeared to be more toxic. Study of embryotoxic effects highlighted closely concentration-dependent damages, the most sensitive stages corresponding to the crucial phases of differentiation (gastrula and prisma). Both EC50 and no-observed-effect concentration values for the four organotin compounds are similar to those reported in the literature for early life stages of other marine organisms. [source] MODULARITY OF THE ANGIOSPERM FEMALE GAMETOPHYTE AND ITS BEARING ON THE EARLY EVOLUTION OF ENDOSPERM IN FLOWERING PLANTSEVOLUTION, Issue 2 2003William E. Friedman Abstract The monosporic seven-celled/eight-nucleate Polygonumtype female gametophyte has long served as a focal point for discussion of the origin and subsequent evolution of the angiosperm female gametophyte. In Polygonumtype female gametophytes, two haploid female nuclei are incorporated into the central cell, and fusion of a sperm cell with the binucleate central cell produces a triploid endosperm with a complement of two maternal and one paternal genomes, characteristic of most angiosperms. We document the development of a four-celled/four-nucleate female gametophyte in Nuphar polysepala (Engelm.) and infer its presence in many other ancient lineages of angiosperms. The central cell of the female gametophyte in these taxa contains only one haploid nucleus; thus endosperm is diploid and has a ratio of one maternal to one paternal genome. Based on comparisons among flowering plants, we conclude that the angiosperm female gametophyte is constructed of modular developmental subunits. Each module is characterized by a common developmental pattern: (1) positioning of a single nucleus within a cytoplasmic domain (pole) of the female gametophyte; (2) two free-nuclear mitoses to yield four nuclei within that domain; and (3) partitioning of three uninucleate cells adjacent to the pole such that the fourth nucleus is confined to the central region of the female gametophyte (central cell). Within the basal angiosperm lineages Nymphaeales and Illiciales, female gametophytes are characterized by a single developmental module that produces a four-celled/four-nucleate structure with a haploid uninucleate central cell. A second pattern, typical of Amborella and the overwhelming majority of eumagnoliids, monocots, and eudicots, involves the early establishment of two developmental modules that produce a seven-celled/eight-nucleate female gametophyte with two haploid nuclei in the central cell. Comparative analysis of onto-genetic sequences suggests that the seven-celled female gametophyte (two modules) evolved by duplication and ectopic expression of an ancestral Nuphar- like developmental module within the chalazal domain of the female gametophyte. These analyses indicate that the first angiosperm female gametophytes were composed of a single developmental module, which upon double fertilization yielded a diploid endosperm. Early in angiosperm history this basic module was duplicated, and resulted in a seven-celled/eight-nucleate female gametophyte, which yielded a triploid endosperm with the characteristic 2:1 maternal to paternal genome ratio. [source] Sperm ultrastructure of the spider crab Maja brachydactyla (Decapoda: Brachyura)JOURNAL OF MORPHOLOGY, Issue 4 2010Carles G. Simeó Abstract This study describes the morphology of the sperm cell of Maja brachydactyla, with emphasis on localizing actin and tubulin. The spermatozoon of M. brachydactyla is similar in appearance and organization to other brachyuran spermatozoa. The spermatozoon is a globular cell composed of a central acrosome, which is surrounded by a thin layer of cytoplasm and a cup-shaped nucleus with four radiating lateral arms. The acrosome is a subspheroidal vesicle composed of three concentric zones surrounded by a capsule. The acrosome is apically covered by an operculum. The perforatorium penetrates the center of the acrosome and has granular material partially composed of actin. The cytoplasm contains one centriole in the subacrosomal region. A cytoplasmic ring encircles the acrosome in the subapical region of the cell and contains the structures-organelles complex (SO-complex), which is composed of a membrane system, mitochondria with few cristae, and microtubules. In the nucleus, slightly condensed chromatin extends along the lateral arms, in which no microtubules have been observed. Chromatin fibers aggregate in certain areas and are often associated with the SO-complex. During the acrosomal reaction, the acrosome could provide support for the penetration of the sperm nucleus, the SO-complex could serve as an anchor point for chromatin, and the lateral arms could play an important role triggering the acrosomal reaction, while slightly decondensed chromatin may be necessary for the deformation of the nucleus. J. Morphol., 2010. © 2009 Wiley-Liss, Inc. [source] Expression of hck-tr, a truncated form of the src-related tyrosine kinase hck, in bovine spermatozoa and testisMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 5 2008Louis-Jean Bordeleau Abstract In bull testicular haploid germ cells, an mRNA encoding for hck was detected in addition to another one encoding for hck-tr, a truncated form of the tyrosine kinase hck. As the transcripts were expressed in spermatids, we tried to determine whether hck-tr is present in mature bovine spermatozoa. Two polyclonal antibodies were produced against peptides specific to the N- and C-terminal portions of the truncated protein. Western blot analyses confirmed the presence of hck-tr in total protein extracts of ejaculated bull spermatozoa, and sub-cellular fractionation experiments suggest its presence in both head and flagellum. The truncated protein appears tightly associated with cytoskeletal elements as it could be extracted only with SDS under reducing conditions. When assessed by indirect immunofluorescence, hck-tr was mostly localized at the acrosomal area of the sperm cell and a similar localization was observed on demembranated spermatozoa. Immunohistochemical studies on testis sections revealed protein expression in spermatocytes as well as in round and elongating spermatids. The results presented in this study clearly show the presence of mRNAs encoding for hck and hck-tr in testicular germ cells; hck-tr being translated during spermatogenesis and expressed on mature ejaculated bull spermatozoa. Mol. Reprod. Dev. 75: 828,837, 2008. © 2007 Wiley-Liss, Inc. [source] Novel human testis-specific cDNA: Molecular cloning, expression and immunobiological effects of the recombinant proteinMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 1 2001Ramasamy Santhanam Abstract A differential display-polymerase chain reaction was employed to obtain a testis-specific cDNA fragment. On screening the human testis-,gt10-cDNA library with testis-specific cDNA fragment, a novel cDNA encoding for a sperm antigen, designated TSA-1, was obtained. It has a novel open reading frame (ORF) of 471 base pairs encoding for 156 amino acids. The computer generated translated protein has a calculated molecular mass of 17.4 kDa and contains a potential N-glycosylation site at amino acids 122,124. The hydrophilicity analysis of the amino acid sequence suggested that this protein is a membrane-anchored peptide. Extensive analysis for tissue-specificity by Northern blots and RT-PCR-Southern blot procedures using various human tissues indicated that TSA-1 was specifically expressed only in the human testis. Based on the results of in vitro transcription and translation experiments, the TSA-1 (ORF) was subcloned into pGEX-6P-3 vector and expressed using the glutathione S -transferase gene fusion system. Antibodies (Ab) against the purified recombinant protein specifically recognized the ,17 kDa recombinant TSA-1, and a ,24 kDa band in human sperm extract in the Western blot procedure. The recombinant TSA-1 Ab recognized the acrosomal, equatorial, mid-piece, and tail regions of human sperm cell in indirect immunofluorescence, bound to live human sperm in the immunobeads binding technique (IBT) and caused a significant concentration-dependent inhibition of human sperm acrosome reaction. These findings indicate that the novel sperm-specific recombinant TSA-1 has a role in sperm function and may have applications in the development of a contraceptive vaccine, and in the specific diagnosis and treatment of male infertility. Mol. Reprod. Dev. 60: 1,12, 2001. © 2001 Wiley-Liss, Inc. [source] Novel features of Equisetum arvense spermatozoids: insights into pteridophyte evolutionNEW PHYTOLOGIST, Issue 1 2002K. S. Renzaglia Summary ,,To characterize structural diversity within Equisetum and among pteridophytes, architectural features of the sperm cell are described here in a second subgenus of Equisetum, a divergent basal group in the fern clade. ,,Transmission electron microscopy observations of prereleased spermatozoids of Equisetum arvense were correlated with three-dimensional scanning electron microscopy images of swimming cells. ,,The mature spermatozoid completes a helix of approximately 2.5 revolutions. At the cell anterior is a complex multilayered locomotory apparatus with staggered flagella. Mitochondria (elongated,rounded) are aggregated near the locomotory apparatus and organelles extend along the cell length. The spline contains up to 300 microtubules and wraps in part around the long cylindrical nucleus. In swimming sperm cells, the anterior of the cell remains tightly coiled while the posterior relaxes and extends in a trailing fashion. ,,Spermatozoids of Equisetum arvense are smaller than those of Equisetum hyemale but structurally similar, except for nuclear shape. Conservation of cellular features suggests recent radiation of the genus. Equisetum spermatozoids share several critical features with ferns, including Psilotum, and support monophyly of a fern,Equisetum assemblage. Entry of the male gametes of Equisetum in their entirety into the archegonial venters indicates possible biparental inheritance of chloroplast and mitochondrial genomes. [source] Vom Samentier zur Samenzelle: Die Neudeutung der Zeugung im 19.BERICHTE ZUR WISSENSCHAFTSGESCHICHTE, Issue 3 2009Jahrhundert Zeugungstheorien; Vererbung; Samentiere; Samenzellen; Physiologie des 19. Jahrhunderts; Reproduktionstechnologien; Körper- und Geschlechtergeschichte; Mikro- und Makrogeschichte Abstract From Spermatic Animalcules to Sperm Cells: The Reconceptualization of Generation in the 19th Century. At the end of the 18th and still at the beginning of the 19th century most naturalists considered spermatic animalcules to be parasites of the seminal fluid that played no role in procreation. This view was progressively questioned by 19th century physiologists. They gradually redefined the spermatic animals as (cellular) products of the male organism, as agents of fertilization and bearers of the male heredity material. This article discusses this change from two different perspectives: on a microhistorical level, it analyzes the experimental research of the naturalist Lazzaro Spallanzani (1729,1799) and of the physiologist Albert Kölliker (1817,1905) in order to show how spermatozoa were turned into a new epistemic object of biology , the sperm cell. Further, it asks how the role of the reconceptualization of spermatic animalcules affected the long-term transformations that gave rise of our modern understanding of heredity, generation and the sexed body. By combining these two perspectives, the article aims to connect historiographies that are often kept separate: the macrohistorical narratives about gender and the body in the modern age and the microhistorical studies of biomedical practices and objects. [source] The geometry and motion of nematode sperm cellsCYTOSKELETON, Issue 6 2009Evgeny Demekhin Abstract The nematode sperm cell crawls by recycling major sperm protein (MSP) from dimers into subfilaments, filaments, and filament complexes, as a result of thermal writhing in the presence of hydrophobic patches. Polymerization near leading edges of the cell intercolates MSP dimers onto the tips of growing filament complexes, forcing them against the cell boundary, and extending the cytoskeleton in the direction of motion. Strong adhesive forces attach the cell to the substrate in the forward part of the lamellipod, while depolymerization in the rearward part of the cell breaks down the cytoskeleton, contracting the lamellipod and pulling the cell body forward. The movement of these cells, then, is caused by coordinated protrusive, adhesive and contractile forces, spatially separated across the lamellipod. This paper considers a phenomenological model that tracks discrete elements of the cytoskeleton in curvilinear coordinates. The pseudo-two dimensional model primarily considers protrusion and rotation of the cell, along with the evolution of the cell boundary. General assumptions are that pH levels within the lamellipod regulate protrusion, contraction and adhesion, and that growth of the cytoskeleton, over time, is perpendicular to the evolving cell boundary. The model follows the growth and contraction of a discrete number of MSP fiber complexes, since they appear to be the principle contributors for force generation in cell boundary protrusion and contraction, and the backbone for the dynamic geometry and motion. Cell Motil. Cytoskeleton 2009. © 2009 Wiley-Liss, Inc. [source] Toxicity of tributyltin and triphenyltin to early life-stages of Paracentrotus lividus (Echinodermata: Echinoidea)ENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 4 2002Alessandra Arizzi Novelli Abstract Sperm cell and embryo toxicity tests using the Mediterranean sea urchin Paracentrotus lividus were performed to assess the toxicity of tributyltin chloride, bis(tributyltin)oxide, triphenyltin acetate, and triphenyltin hydroxide. Toxicity values (mean effective concentration [EC50]) ranged from 2.97 to 18.5 ,g/L for sperm cells and from 1.11 to 2.62 ,g/L for embryos. For sperm cells, the toxicity of the two tributyl compounds was significantly greater than that of two triphenyl compounds; for embryos, the triphenyl compounds appeared to be more toxic. Study of embryotoxic effects highlighted closely concentration-dependent damages, the most sensitive stages corresponding to the crucial phases of differentiation (gastrula and prisma). Both EC50 and no-observed-effect concentration values for the four organotin compounds are similar to those reported in the literature for early life stages of other marine organisms. [source] Specific Fab fragments recovered by phage display technique recognizing human spermatozoaINTERNATIONAL JOURNAL OF ANDROLOGY, Issue 5 2009Dorota Fiszer Summary Human hybridoma cell lines are often unstable and loose ability for antibody production. Sometimes, they show low and varying levels of heavy and light chains synthesis. Therefore it is reasonable to preserve generated specificities of light and heavy chains by cloning them to phagemid vector and creating phage display library. The aim of this study was to construct phage display library of Fab fragments recognizing sperm surface antigens. The source of mRNA constituted seven hybridoma cell lines producing antisperm antibodies which was proved by ELISA, and agglutination test as well as by inhibition of sperm to penetrate hamster oocytes. Fragments of cDNA encoding ,/, and , chains were cloned into pComb3HSS phagemid vector and amplified in XL-1Blue. The library was panned against whole unfixed sperm cells. Three positive clones selected after fourth round of panning showed heavy chain belonging to VH4 family, two of them (G28, K61) possessed lambda chain from VL2 family and one (H43) kappa chain from VK1 family. As these Fabs revealed similarities to antibodies against some proteins involved in sperm motility and cell fusion it can be suggested that these Fabs may be a cause of infertility. Finally, we proved that it is feasible to preserve specificities produced by human hybridomas using phage display technique and we recovered some Fabs which may be of diagnostic and research value, and may also have some value for contraceptive vaccine. [source] A new test for immunological infertility: an ELISA based on prostasomesINTERNATIONAL JOURNAL OF ANDROLOGY, Issue 3 2004L. Carlsson Summary Antisperm antibodies (ASA) are present in patients with immunological infertility, but the antigens are poorly characterized. Prostasomes adhere to sperm cells and are recognized as antigens for ASA. This investigation aimed to study the prevalence of antiprostasome antibodies in ASA-classified sera. We studied the reactivity of ASA-positive sera from 116 immunoinfertile patients. Ninety-seven per cent (113 of 116) of the patients' sera contained IgG antibodies against seminal prostasomes. Accordingly, prostasomes are one of the major targets for ASA. An enzyme-linked immunosorbent assay based on prostasomes is simpler to perform than ASA tests presently in use. It is also easier to achieve reproducible and standardized results. [source] Interaction between leucocytes and human spermatozoa influencing reactive oxygen intermediates releaseINTERNATIONAL JOURNAL OF ANDROLOGY, Issue 2 2004Monika Fr Summary The relationship between the presence of white blood cells (WBCs) and the fertilizing potential of human semen is still an open question. It is well known that the presence of leucocytes in human semen can be related to the production of reactive oxygen intermediates (ROI). Semen samples were obtained from 15 normozoospermic men and leucocytes were isolated from heparinized blood drawn from 15 volunteers. Lucigenin and luminol-mediated chemiluminescence assays were used to determine reactive oxygen species (ROS) generation by non-activated or activated leucocytes through 12-myristate-13-acetate or N-formyl-methionyl-leucyl-phenyalanine (FMLP) before the addition of spermatozoa isolated by swim-up or Percoll procedures. All spermatozoal fractions used in this study were characterized by defining their motility, morphology and viability. The levels of ROS formation by non-activated as well as stimulated leucocytes were significantly decreased after addition of swim-up separated spermatozoa (p < 0.01). The ability to inhibit the basal chemiluminescence was of lower degree for spermatozoa isolated from 90% Percoll fractions than for swim-up sperm. However, addition of sperm cells from 47% Percoll fraction was found to increase both lucigenin and luminol signals. Moreover, the determined ROI levels changed depending on the type of inducing factor used for oxidative burst. Then, spermatozoa selected by swim-up procedure although with only slightly higher viability and morphology than sperm obtained from 90% Percoll fraction clearly exhibited much higher capacity to inhibit ROI secretion by receptor-stimulated leucocytes (FMLP-activation) than Percoll fractionated sperm. Such results may indicate that within normal semen may exist sperm subpopulations with different biochemical mechanisms controlling the interaction between spermatozoa and contaminating leucocytes. When ROI levels contained in normozoospermic semen are dependent on the WBCs activation, it seems that spermatozoa with preserved normal functional competence are able to defend themselves against leucocytes-derived ROI. Also for normozoospermic ejaculates, swim-up sperm may improve semen antioxidant characteristics when comparing with Percoll (90%) separated sperm. It may help for optimal sperm preparation when assisting to infertility treatment. [source] Enzymatic and immunochemical evaluation of phospholipid hydroperoxide glutathione peroxidase (PHGPx) in testes and epididymal spermatozoa of rats of different agesINTERNATIONAL JOURNAL OF ANDROLOGY, Issue 2 2002Federica Tramer Selenium (Se) and selenoproteins such as glutathione peroxidases are necessary for the proper development and fertilizing capacity of sperm cells. Phospholipid hydroperoxide glutathione peroxidase (PHGPx, E.C. 1.11.1.12) is a monomeric seleno-enzyme present in different mammalian tissues in soluble and bound form. Its function, like the other glutathione peroxidases, was originally viewed as a protective role against hydroperoxides, but direct and indirect evidence indicates that it has additional regulatory roles. PHGPx is present in testis cells and sperm cells, and its appearance is hormone regulated. We present here biochemical data, which clearly indicate that the enzyme specific activity in rat is age-dependent during the life-span monitored (from 36 to 365 days), with a maximum at 3 months of age in the testis germ cells and at 6 months of age in the isolated epididymal sperm cells. Western blotting and immunocytochemical analysis by means of anti-PHGPx antibodies show the different distribution and the strong binding of PHGPx in the testes and sperm cell subcellular compartments (nucleus, acrosome, mitochondria and residual bodies) of rats of different age. The presence of the protein exhibits in the testis cells a pattern different from that of the catalytic activity, with a maximum at 6 months of age. The subcellular distribution of PHGPx is qualitatively, but not quantitatively, unchanged during ageing. These different behaviours are compared and discussed. [source] Bacillus subtilis and B. mojavensis strains connected to food poisoning produce the heat stable toxin amylosinJOURNAL OF APPLIED MICROBIOLOGY, Issue 6 2009C. Apetroaie-Constantin Abstract Aim:, To screen and characterize toxic, heat-stable substances produced by food borne strains from Bacillus subtilis group. Methods and Results:, Using the boar sperm motility inhibition assay, six isolates from two outbreaks, out of the 94 isolates from 26 foods, were found to produce ethanol-soluble heat-stable substances that were toxic to sperm cells by depleting the mitochondrial membrane potentials. The toxic isolates were identified as Bacillus subtilis and B mojavensis. Colon carcinoma cells (Caco-2) were used to model the contact with the human digestive tract. The extract of B. subtilis F 2564/96 depolarized the mitochondria in intact Caco-2 cells similarly as in sperm cells. The substance responsible for these effects was purified using HPLC and identified by electron spray ionization ion trap mass spectrometry analysis as amylosin. The temperature requirement for amylosin production was 21,37°C for B. subtilis and 11,21°C for B. mojavensis. Both species produced amylosin in air as well as in 7,8% CO2 with 8,9% O2. Conclusions:, Food borne illness related strains of B. subtilis and B. mojavensis, produced the heat-stable toxin amylosin. Significance and Impact of the Study:, This is the first report that suggests a role for the heat-stable, ion-channel forming toxin amylosin, as a virulence factor in food borne Bacillus. [source] Localizations of intracellular calcium and Ca2+ -ATPase in hamster spermatogenic cells and spermatozoaMICROSCOPY RESEARCH AND TECHNIQUE, Issue 8 2006H.L. Feng Abstract Calcium plays a predominant role regulating many functional processes of spermatogenesis and fertilization. The purpose of the present study is to define the exact location of calcium as well as examine the role it plays during spermatogenesis and sperm capacitation. Testes and epididymides were obtained from adult healthy male hamsters. Spermatozoa were incubated with modified Tyrode's medium up to 4 h at 37°C for sperm capacitation in vitro. Samples of the testes and sperm cells were analyzed by cytochemical techniques to determine the location of calcium and Ca2+ -ATPase and the percentage of acrosome reactions under light and electron microscopy. The data showed that (1) Sertoli cells exhibited numerous calcium precipitates as large, round, electron-dense bodies distributed throughout the cytoplasm and the mitochondrial matrix. Fine calcium precipitates existed in fewer numbers in the intracellular storage sites of spermatogonia and primary spermatocytes, in sharp distinction to secondary spermatocyte and spermatids, which showed an abundance of large and round calcium precipitates, especially in the mitochondrial matrix of spermatids. More calcium deposits were distributed in the plasma membrane (PM), acrosome membrane, and matrices of the acrosome and mitochondria following capacitation; (2) Ca2+ -ATPase was found in the endoplasmic reticulum system and PM of noncapacitated spermatozoa as well as Sertoli cells. Capacitated spermatozoa showed a weak signal. These results suggest that the presence of calcium in spermatogenic cells might play a role in cell growth and differentiation during spermatogenesis. The Ca2+ -ATPase function may be inhibited during capacitation, leading to an increase in acrosomal calcium level and triggering of acrosomal exocytosis. Microsc. Res. Tech., 2006. © 2006 Wiley-Liss, Inc. [source] Sperm binding properties and secretory activity of the bovine oviduct immediately before and after ovulationMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 1 2008Edita Sostaric Abstract The possibility that differences in hormonal regimes between the two oviducts in the cow around ovulation affects secretory activity of the oviduct epithelial cells and/or sperm,oviduct binding was studied. Oviducts were collected immediately after slaughter at 6 hr before to 5 hr after timed ovulation of 14 normally cyclic cows that had been inseminated (n,=,6) or not (n,=,8) and material obtained from the same cows was processed in three ways. First, in vivo, after artificial insemination of the cows, low numbers of sperm cells (approx. 15 per oviduct) were found within the entire oviducts as observed by scanning electron microscopy (SEM). Almost all sperm were located in the isthmus and then only on ciliated cells and showed without exception fully matured, intact morphology. Secretory activity of noninseminated oviduct epithelia was induced after ovulation which was most predominant in the pockets of the ipsi-lateral ampulla compared to the contra-lateral ampulla (P,<,0.01). Second, ex vivo, explants dissected from oviducts of the noniseminated cows were incubated with sperm. In all cases, the sperm bound to the explants in a similar pattern as observed in vivo and this binding was strictly fucose-dependent. The main difference with in vivo experiments was the high numbers of sperm bound at any site of the oviduct (,3,000 cells per mm2) indicating the high sperm binding capacity of the oviduct epithelia. Ovulation induced a striking drop in sperm binding capacity in the oviducts and was most pronounced in the isthmus (,1,300 cells per mm2; P,<,0.001) and to a lesser extent in the ampulla (,2,000 cells per mm2, P,<,0.01). Third, in vitro, pieces of tissue dissected from oviducts of the noninseminated cows were cultured to mono-layers. Culturing epithelial cells resulted in loss of their normal morphological appearance. In all cases, the sperm binding capacity in monolayers was very low (<50 cells per mm2) when compared to corresponding explants (P,<,0.0001). Sperm binding to monolayers originating from the isthmus (<25 cells per mm2) was lower than in those from the ampulla (40,50 cells per mm2; P,<,0.01) and remained similar after ovulation. In all three approaches, no significant differences were found in sperm,oviduct binding characteristics and sperm-distribution in the ipsi- versus contra-lateral oviducts. This indicates, that systemic endocrine changes around ovulation rather than specific oviduct changes at the ipsi,lateral oviduct induce secretion in oviduct epithelial cells, and thus induce sperm release. Mol. Reprod. Dev. 75: 60,74, 2008. © 2007 Wiley-Liss, Inc. [source] Stage-dependent Dishevelled-1 expression during mouse spermatogenesis suggests a role in regulating spermatid morphological changesMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 6 2006Pengpeng Ma Abstract Dishevelled (Dsh in Drosophila or DVL in mice) is a member of the highly conserved Wg/Wnt signaling pathway, which regulates important processes such as cell proliferation, polarity, and specification of cell fate. Three orthologous genes of Dishevelled (Dvl-1, Dvl-2, and Dvl-3) have been found in both humans and mice. They play pivotal roles in regulating cell morphology and a variety of changes in cell behaviors. In the present study, we show that the expression of Dvl-1 is stage-dependent during mouse spermatogenesis, although Dvl-2 and Dvl-3 show relative consistent expression. The expression of Dvl-1 mRNA first appears in pachytene spermatocytes, increases in round and elongating spermatids, and then turns to an undetectable level in mature sperm cells. Analyses of immunohistochemistry and immunofluorescence staining show that DVL-1 is present diffusely in the cytoplasm of pachytene spermatocytes and exhibits mainly a vesicular pattern and perinuclear distribution and a weak diffusely cytoplasmic signal in round and elongating spermatids. The vesicular pattern of DVL-1 has been observed by previous studies in somatic cells, and suggested to play roles in signal transduction. Immunoprecipitation experiments show that DVL-1 coimmunprecipitates with spermatogenic cells ,-actin rather than ,-tubulin. These results indicate that DVL-1 may be involved in spermatid morphological changes during mouse spermiogenesis through mediating signal transduction and/or regulating actin cytoskeleton organization. Mol. Reprod. Dev. © 2006 Wiley-Liss, Inc. [source] Lipid peroxide formation in relation to membrane stability of fresh and frozen thawed stallion spermatozoaMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 2 2005D.M. Neild Abstract In this study we used a new method to detect reactive oxygen species (ROS) induced damage at the level of the sperm plasma membrane in fresh and frozen-thawed stallion sperm. Lipid peroxidation (LPO) in sperm cells was assessed by a fluorescent assay involving the labeling of stallion sperm with the LPO reporter probe C11-BODIPY581/591. The peroxidation dependent spectral emission shift of this membrane probe could be localized using inverted spectral confocal microscopy and quantified on living and deteriorated sperm cells using flow cytometry. Mass spectrometric analysis of the main endogenous lipid class, phosphatidylcholine (PC), was carried out to determine the formation of hydroxy- and hydroperoxyphosphatidylcholine in fresh sperm cells. Peroxidation as reported by the fluorescent probe corresponded with the presence of hydroxy- and hydroperoxyphosphatidylcholine in the sperm membranes, which are early stage products of LPO. This allowed us to correlate endogenous LPO with localization of this process in the living sperm cells. In absence of peroxidation inducers, only relatively little peroxidation was noted in fresh sperm cells whereas some mid-piece specific probe oxidation was noted for frozen-thawed sperm cells. After induction of peroxidation in fresh and frozen-thawed sperm cells with the 0.1 mM of lipid soluble ROS tert -butylhydrogen peroxide (t -BUT) intense probe oxidation was produced in the mid-piece, whereas the probe remained intact in the sperm head, demonstrating antioxidant activity in the head of fresh sperm cells. At higher levels of t -BUT, probe peroxidation was also noted for the sperm head followed by a loss of membranes there. Frozen-thawed sperm were more vulnerable to t -BUT than fresh sperm. The potential importance of the new assays for sperm assessments is discussed. © 2005 Wiley-Liss, Inc. [source] Specific localization of transcription factors in the chromatin of mouse mature spermatozoaMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 1 2001Carmine Pittoggi Abstract We previously characterized a nuclease-hypersensitive fraction of mouse sperm chromatin, which is organized in a typical nucleosomal structure. A partial genomic library was constructed with the DNA from the nuclease-hypersensitive chromatin, which revealed a high content in retroposon/retroviral DNA sequences. Here we report that the cloned nuclease-hypersensitive DNA also contains clusters of potential sites for transcription factors: among those, binding sites for Oct-1, Oct-4, TBP, Ets-1, and C/EBP are most abundant. This observation prompted us to ask whether mature spermatozoa contain the corresponding protein factors. Indirect immunofluorescence experiments show that all analyzed factors are indeed present in the sperm heads. Moreover, transcription factors are associated with the nuclease-hypersensitive chromatin of spermatozoa, as endogenous nucleases that degrade the hypersensitive fraction also cause the concomitant release of transcription factors from sperm cells into the medium. Band-shift assays with proteins extracted from the supernatant, and immunofluorescence analysis of sperm pellets, indicate that transcription factors are largely recovered in the supernatant while being absent or poorly retained in spermatozoa. The possible involvement of these factors in early embryogenesis is discussed. Mol. Reprod. Dev. 60: 97,106, 2001. © 2001 Wiley-Liss, Inc. [source] Novel features of Equisetum arvense spermatozoids: insights into pteridophyte evolutionNEW PHYTOLOGIST, Issue 1 2002K. S. Renzaglia Summary ,,To characterize structural diversity within Equisetum and among pteridophytes, architectural features of the sperm cell are described here in a second subgenus of Equisetum, a divergent basal group in the fern clade. ,,Transmission electron microscopy observations of prereleased spermatozoids of Equisetum arvense were correlated with three-dimensional scanning electron microscopy images of swimming cells. ,,The mature spermatozoid completes a helix of approximately 2.5 revolutions. At the cell anterior is a complex multilayered locomotory apparatus with staggered flagella. Mitochondria (elongated,rounded) are aggregated near the locomotory apparatus and organelles extend along the cell length. The spline contains up to 300 microtubules and wraps in part around the long cylindrical nucleus. In swimming sperm cells, the anterior of the cell remains tightly coiled while the posterior relaxes and extends in a trailing fashion. ,,Spermatozoids of Equisetum arvense are smaller than those of Equisetum hyemale but structurally similar, except for nuclear shape. Conservation of cellular features suggests recent radiation of the genus. Equisetum spermatozoids share several critical features with ferns, including Psilotum, and support monophyly of a fern,Equisetum assemblage. Entry of the male gametes of Equisetum in their entirety into the archegonial venters indicates possible biparental inheritance of chloroplast and mitochondrial genomes. [source] Effect of Mucuna urens (horse eye bean) on the gonads of male guinea-pigsPHYTOTHERAPY RESEARCH, Issue 2 2001Paul Udoh Abstract The effect of Mucuna urens (seeds) on the gonads and sex accessory glands of male guinea-pigs was investigated. Sexually mature guinea-pigs of proven fertility were administered orally with 70,mg/kg and 140,mg/kg body weight of crude extract daily for 8 weeks respectively. Phytochemical screening of the seeds revealed the presence of alkaloids. No death or weight loss were observed during the duration of treatment. No pregnancy occurred in females mated with the treated males. Histological observations at high dose (140,mg/kg) showed complete degeneration of sperm in the testicular tubules. In some tubules, the acrosomal cap of the sperm cells was separated from the nuclei which underwent colour changes. In some tubules only the tails were left in the lumen. The spermatids, primary and secondary spermatocytes showed pycnosis while the morphology of spermatogonia and germinal epithelium appeared normal. Some epididymides were devoid of sperm while others contained degenerated spermatozoa and cell debris. In the prostate gland there was collapse of the villi and reduction of secretion in both the prostate and seminal vesicles. At low doses (70,mg/kg), there was spermatogenic arrest at spermatid stage. These observations have shown that M. urens is a potential male antifertility agent. Copyright © 2001 John Wiley & Sons, Ltd. [source] Resolving a genetic paradox throughout preimplantation genetic diagnosis for autosomal dominant severe congenital neutropeniaPRENATAL DIAGNOSIS, Issue 3 2010Mira Malcov Abstract Objective Severe congenital neutropenia is an inherited disease characterized by low peripheral blood neutrophils, amenable to bone marrow transplantation. Genetic analysis in the family here described detected a ELA2 splice-site mutation in the affected child and also in his asymptomatic father. The parents requested preimplantation genetic diagnosis (PGD), coupled with HLA matching, to obtain a suitable bone marrow donor for the affected child. Methods A PGD protocol was developed, based on multiplex nested PCR for direct analysis of the ELA2 mutation, flanking polymorphic markers and HLA typing. Results The amplification efficiency of the mutation was > 90% in single leukocytes from the affected child but only 67% in the father. Analysis of single haploid sperm cells from the father demonstrated three different sperm-cell populations: (1) sperm cells harboring the ELA2 mutation on the ,affected' haplotype, (2) sperm cells without the ELA2 mutation on the ,normal' haplotype, and (3) sperm cells without the ELA2 mutation on the ,affected' haplotype. Conclusion These data demonstrate that the ELA2 mutation in the father occurred de novo during his embryonic development, resulting in somatic as well as germ-line mosaicism. This conclusion was also taken into consideration when PGD was performed. Copyright © 2010 John Wiley & Sons, Ltd. [source] Prenatal diagnosis of citrullinemia and argininosuccinic aciduria: evidence for a transmission ratio distortion in citrullinemiaPRENATAL DIAGNOSIS, Issue 3 2006Wim J. Kleijer Abstract Background In the course of 25 years, we have experienced a high rate of affected fetuses in the prenatal diagnosis of citrullinemia. Methods and Results Ninety-one pregnancies at 1 in 4 risk were tested; 36 were diagnosed as affected (39.5%; P = 0.0015). The high rate of positive diagnoses was found both after chorionic villus sampling (24/68 = 35.3%) and amniocentesis (12/23 = 52.2%) despite the completely different and independent techniques used. Using exactly the same (indirect) enzyme assay for argininosuccinic aciduria on chorionic villi and a similar method on amniotic fluid, the expected rate of affected fetuses was found: 13/53 = 24.5%. Technical and genetic causes for the unexpected results were excluded by confirmatory studies performed on independent fetal material, which was available for 27 of the 36 fetuses affected with citrullinemia. Biochemical confirmation was obtained in the 27 cases, whereas in 18 fetuses homozygosity or compound heterozygosity for disease-causing mutations were retrospectively demonstrated in the stored fetal cells. Conclusion The results suggest the occurrence of preferential transmission of the mutant allele. An explanation for this phenomenon may be found in a protective role of argininosuccinic acid synthetase deficiency in mutant sperm cells against the possibly detrimental or apoptotic effect of nitric oxide produced normally from arginine by nitric oxide synthase. Copyright © 2006 John Wiley & Sons, Ltd. [source] Effect of Ram Age on Structural and Functional Competence of Frozen,Thawed Spermatozoa in Dairy SheepREPRODUCTION IN DOMESTIC ANIMALS, Issue 4 2010AG Lymberopoulos Contents The objectives of this study were to investigate the influence of ram age on structural and functional competence of frozen,thawed spermatozoa and to test the hypothesis that increasing number of sperm bound to the zona pellucida in vitro was associated with decreasing in vivo fertility of frozen semen. Rams were allocated into two groups. Each group consisted of five rams aged either 1,2 years (young) or 4,5 years (mature). Three successive ejaculates were collected from each ram using an artificial vagina. Only ejaculates of , 2.5 × 109 sperm/ml and 80% sperm progressive motility were pooled per ram, diluted with Bioxcell® medium and frozen in 0.25 ml straws. The end points of post-thawing semen evaluation were computer-assisted cell motility analysis, sperm capacitation (chlortetracycline assay), simultaneous assessment of plasma membrane integrity, mitochondrial membrane potential and condensation status of nucleus, per-cell analysis of lipid peroxidation using C11-BODIPY581/591, sperm-hemizona binding (HZB) ability and sperm fertility after laparoscopic insemination of ewes (n = 114) in the progestagen-synchronized oestrus. The results showed that mature rams had significantly lower values of sperm hyperactivated motility and peroxidized sperm, higher percentages of live non-capacitated sperm and sperm cells with intact plasma membrane, functional mitochondria and condensed chromatin, as well as, greater lambing rate and ewe prolificacy. Sperm HZB binding ability was higher (p < 0.05) for young than for mature rams. Significant correlations were found between number of spermatozoa bound to the zona pellucida and semen fertility (r = ,0.63 to ,0.71). In conclusion, mature rams have better semen quality and in vivo fertility than young rams. Cryocapacitation can be involved in decreasing ram semen fertility as evidenced by the high number of spermatozoa bound to the zona pellucida in vitro. [source] Dynamic expression of Krüppel-like factor 4 (Klf4), a target of transcription factor AP-2, during murine mid-embryogenesisTHE ANATOMICAL RECORD : ADVANCES IN INTEGRATIVE ANATOMY AND EVOLUTIONARY BIOLOGY, Issue 2 2003Julia Ehlermann Abstract Krüppel-like factor 4 (Klf4) belongs to the family of transcription factors that are thought to be involved in the regulation of epithelial and germ cell differentiation, based on their expression in postproliferative cells of the skin, gut, and testes. Gene ablation experiments suggest that Klf4 plays a role in keratinocyte differentiation, since mice lacking Klf4 fail to establish proper barrier function and, as a consequence, die postnatally due to dehydration. Recent studies have shown that Klf4 is also expressed in postnatal male mice, in postmeiotic sperm cells undergoing terminal differentiation into sperm cells. However, prior to the current study, the expression pattern of Klf4 during early and mid-embryogenesis had not been examined. Here we demonstrate that Klf4 transcripts can be detected from embryonic day 4.5 (E4.5) on in the developing conceptus, and that Klf4 expression before E10 is restricted to extraembryonic tissues. The embryo proper displays a highly dynamic and changing Klf4 signal from E10 of murine development on. In addition to being expressed in a stripe of mesenchymal cells extending from the forelimb bud rostrally over the branchial arches to the developing eye, Klf4 is also expressed in the mesenchyme surrounding the nasal pit at day E11.5. In addition, Klf4 has been detected in the apical ectodermal ridge and adjacent mesenchymal cells in the limb buds, and in mesenchymal cells of the developing body wall in trunk areas. These findings suggest that Klf4 plays an important role in regulating cellular proliferation, which underlies the morphogenetic changes that shape the developing embryo. Anat Rec Part A 273A:677,680, 2003. © 2003 Wiley-Liss, Inc. [source] ORIGINAL ARTICLE: Sperm Antibodies, Intra-Acrosomal Sperm Proteins, and Cytokines in Semen in Men from Infertile CouplesAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 3 2009Zdenka Ulcova-Gallova Problem, The aim of this study was to investigate seminal sperm-agglutinating antibodies, intra-acrosomal proteins, sperm head abnormalities, and cytokines (IL-1,, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70 TNF-,, and IFN-,) in men from infertile couples. Method of study, The direct mixed anti-immunoglobulin reaction test for IgG, IgA, and IgE in semen, and immunocytochemical method using monoclonal antibodies and indirect immunofluorescence for the examination of intra-acrosomal proteins in the spermatozoa were used. Cytokines in seminal plasma were determined by multiplex immunoanalytic xMAP (LUMINEX) technology. Results, Sperm-agglutinating antibodies, IgG and IgA, in seminal plasma were found to be more in asthenospermatic and oligoasthenospermatic men than in normospermatic men. Sperm head pathology and very low amounts of acrosomal proteins were frequently detected in pathologic semen samples. Cytokine levels defined as ,high' (based on the 75 percentile for each cytokine in all groups) were obtained especially for IL-8, IL-5, IL-6, and IL-10. The high cellularity in semen was correlated with higher IL-5. Conclusion, Immunologic cause of male infertility is a very important risk factor in the pathogenesis of sperm cells. Sperm autoantibodies and the presence of intra-acrosomal factors must be studied together, cytokines according to accessory cellularity in the semen. [source] Relationship between hyaluronic acid binding assay and outcome in ART: a pilot studyANDROLOGIA, Issue 5 2010M. Nijs Summary The sperm,hyaluronan binding assay (HBA) is a diagnostic kit for assessing sperm maturity, function and fertility. The aim of this prospective cohort pilot study was to evaluate the relationship between HBA and WHO sperm parameters (motility, concentration and detailed morphology) and possible influence of sperm processing on hyaluronic acid binding. A cohort of 68 patients undergoing a first combo in vitro fertilisation/intracytoplasmic sperm injection treatment after failure of three or more intrauterine insemination cycles were included in the study. Outcome measures studied were fertilisation rate, embryo quality, ongoing pregnancy rate and cumulative pregnancy rate. HBA outcome improved after sperm preparation and culture, but was not correlated to detailed sperm morphology, concentration or motility. HBA did not provide additional information for identifying patients with poor or absent fertilisation, although the latter had more immature sperm cells and cells with cytoplasmic retention present in their semen. HBA outcome in the neat sample was significantly correlated with embryo quality, with miscarriage rates and ongoing pregnancy rates in the fresh cycles, but not with the cumulative ongoing pregnancy rate. No threshold value for HBA and outcome in combo IVF/ICSI treatment could be established. The clinical value for HBA in addition to routine semen analysis for this patient population seems limited. [source] Effects of varicocele upon the expression of apoptosis-related proteinsANDROLOGIA, Issue 4 2010F.-W. Chang Summary Varicocele-associated apoptosis has been recognised as a cause of male infertility. Thus, we assessed the expression of somatic apoptosis-related proteins (the typical protein-dependent apoptosis markers) in ejaculated sperm plasma from both patients with varicocele and normal donors. We evaluated the relationships between certain apoptosis-related proteins and normal semen quality. Semen samples were obtained from 25 patients with varicocele and from 10 normal fertile controls. These samples were compared using computer-assisted semen analysis for motion parameters and manual analysis for morphology, and were also assayed for apoptosis-related protein activation including caspase-3, poly-ACP-ribose polymerase (PARP), the Bcl-2 family (Bcl-2, Bak) and p53 by means of immunoblot analysis. PARP, Bak and p53 were expressed substantially more in the sperm cells of the varicocele group when compared with the normal group (P < 0.05). The expression of caspase-3 and Bcl-2 did not appear to differ between these two study groups. An increased expression of PARP, Bak and p53 for varicocele-afflicted individuals indicated an increased participation by these agents in the regulating of apoptosis in the ejaculated semen from patients with varicocele, suggesting that certain protein-development apoptotic mechanisms might originate in the cytoplasmic droplet or within mitochondria of spermatocytes and then might function within the nucleus of the cell. [source] Effect of anion channel blockers on l- arginine action in spermatozoa from asthenospermic menANDROLOGIA, Issue 2 2010S. Srivastava Summary In earlier studies, we have established that l- arginine enhances motility and metabolic rate in spermatozoa of goat, bull and mouse. In the present study this work was extended to human sperm cells obtained from the semen samples of asthenospermic patients, which are characterised by low motility. The metabolic rate was followed by monitoring the glucose consumption (1- 13C glucose as substrate) and the production of lactate in sperm cells, using 13C NMR. The stimulatory effect of l- arginine was neutralised on adding an NO-synthase inhibitor like N, -nitro- l- arginine methyl ester. On the other hand, the inactive d -enantiomorph did not affect the stimulatory effect of l- arginine. This strongly suggests that l- arginine acts through the NO signal pathway. We also demonstrated that the stimulatory effect of l- arginine was inhibited in the presence of anion channel inhibitors like 4-acetamido-4,-isothiocyanostilbene-2,2,-disulphonic acid, 2,4-dinitrophenol and carbonyl cyanide m-chlorophenylhydrazone. Furthermore, bicarbonate supplementation was found to be essential for the action of l- arginine. These observations indicate that l- arginine induces NO synthesis and stimulates motility and metabolism only when an active anion transport system is present. 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