Publications (24)80.08 Total impact
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Article: Correlation Between the Sperm Motility and the Adenylate Cyclase Activity in Infertile Men/Über die Beziehungen zwischen der Motilität der Spermatozoon und der Adenylcyclase‐Aktivität bei unfruchtbaren Männern
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ABSTRACT: The relationship between the sperm motility and the adenylate cyclase activity in spermatozoa from patients with male infertility was investigated. Cyclic AMP contents were measured from 34 semen specimens and adenylate cyclase activity were assayed from 50 specimens. All semen specimens were collected from infertile men at our clinic by masturbation after 5 days sexual abstinence. Spermatozoa were washed and concentrated according to the method described by Harrison. The cAMP contents and the adenylate cyclase activity of spermatozoa were determined by radioimmunoassay.Positive correlations were found among the cAMP contents, the adenylate cyclase activity in human spermatozoa and the sperm motility.This result suggests that the sperm motility is regulated by the adenylate cyclase activity via cAMP and that poor sperm motility observed in infertile men is partially caused by the impairment of adenylate cyclase system.Zusammenfassung: Es wird eine experimentelle Arbeit über etwaige Beziehungen zwischen Spermatozoenmotilität und Adenylcyclase-Aktivität vorgelegt. Hierzu wurden die Werte für cAMP bei 34 Spermaproben und für Adenylcyclase-Aktivität (ACA) bei 50 Spermaproben gemessen. Alle Proben stammten aus der Infertilitäts-Klinik von unfruchtbaren Männern, sie wurden per masturbationem gewonnen bei einer sexuellen Abstinenz von 5 Tagen. Die Spermatozoen wurden gewaschen und konzentriert nach der Methode von Harrison. Die cAMP-Werte und die ACA wurden radioimmunologisch bestimmt. Es ergaben sich positive Korrelationen zwischen den cAMP-Werten und der ACA in menschlichen Spermatozoen und der Spermatozoen-Motilität. Die Ergebnisse legen es nahe, daß die Spermatozoen-Motilität durch ACA via cAMP reguliert wird und daß eine schlechte Motilität der Spermatozoen bei Unfruchtbarkeit des Mannes wenigstens teilweise verursacht ist durch eine Störung im Adenyl-Cyclase-System.Andrologia 04/2009; 21(5):437 - 440. · 1.55 Impact Factor -
Article: Inhibition of adenylylcyclase activity in mouse cerebellum membranes upon hydrolysis of triacylglycerols by triacylglycerol lipase, but not phospholipids by phospholipase A(2).
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ABSTRACT: We previously showed that arachidonic acid and related unsaturated free fatty acids (U-FFAs) inhibit the activity of adenylylcyclase in brain membranes of mice. The level of U-FFAs elevates when the hydrolysis of triacylglycerols (TAGs) and phospholipids is promoted. In this study, we examined whether activation of triacylglycerol lipase (TAG lipase) and phospholipase A(2) (PLA(2)) results in the inhibition of adenylylcyclase activity in cerebellum membranes of mice. Incubation of Intralipos with TAG lipase in the presence of membranes mainly released oleic acid and linoleic acid and caused > or =95% inhibition of adenylylcyclase activity. In contrast, PLA(2), though releasing substantial amounts of U-FFAs, increased the enzymatic activity. To account for this difference, we examined how by-products formed in U-FFA release by TAG lipase and PLA(2) operated on the arachidonic acid-induced inhibition. Lysophosphatidylcholne and some other lysophospholipids, produced by PLA(2), enhanced the adenylylcyclase activity and attenuated the inhibitory effect of arachidonic acid. On the other hand, no such effects were found with by-products of TAG lipase-mediated lipolysis. Rather, monoacylglycerols having U-FFAs, possibly formed by TAG lipase, potentiated the arachidonic acid-induced inhibition of adenylylcyclase. Bovine serum albumin, added into the mixture for the pretreatment of membranes with TAG lipase, prevented the inhibition of adenylylcyclase. These results indicate that by-products formed in U-FFA release have a crucial role for the U-FFA's action on adenylylcyclase and that U-FFAs released from TAG are an inhibitor of adenylylcyclase. It may be that albumin in plasma, and thus FFA-binding proteins within cells, are of importance in protecting adenylylcyclase upon U-FFA release.Archives of Biochemistry and Biophysics 10/2001; 393(1):123-31. · 2.93 Impact Factor -
Article: Inhibition of adenylyl cyclase activity in brain membrane fractions by arachidonic acid and related unsaturated fatty acids.
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ABSTRACT: Pretreatment of mouse brain membranes with arachidonic acid (AA) and related unsaturated fatty acids at 30 degrees C for 10 min decreased basal activity and isoproterenol/guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S)- and forskolin-stimulated activities of adenylyl cyclase to a level less than 5% of control. The presence of the carboxyl group on the fatty acids was essential for the inhibition, because no such inhibition was found with ethyl arachidonate or AA attached to diacylglycerols and phospholipids. The AA-mediated inhibition was observed when the activity was measured in the presence of Mn2+ or forskolin and was insensitive to pertussis toxin or guanosine 5'-O-(2-thiodiphosphate) (GDPbetaS), indicating a mechanism independent of GTP-binding proteins. In addition, the fact that stimulators of the adenylyl cyclase catalytic unit, ATP, GTP gamma S and forskolin, when present during pretreatment, attenuate the inhibitory effect of AA may suggest that the catalytic unit is a target of AA. Bovine serum albumin suppressed the inhibition when present in the mixtures for pretreatment, but could not restore the adenylyl cyclase activity that had been reduced by AA, indicating an irreversible inhibition by AA. The effect of AA was found to be additive to P-site-mediated inhibition. The present study suggests the existence of another mechanism of regulation of adenylyl cyclase by unsaturated fatty acids.Archives of Biochemistry and Biophysics 06/2001; 389(1):68-76. · 2.93 Impact Factor -
Article: Purification and properties of major alpha-D-mannosidase in the luminal fluid of porcine epididymis.
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ABSTRACT: A lysosomal type alpha-D-mannosidase was successfully purified by DEAE-Sephacel, Red-Amicon and Superdex 200 column chromatographies from porcine cauda epididymal fluid. The purified enzyme consisted of 63 and 51 kDa subunits at equimolar amounts. It cleaved alpha1-2 linked mannosyl residues and less but significantly cleaved alpha1-3 and alpha1-6 linked mannosyl residues in the high-mannose oligosaccharides. The optimal pH to hydrolyze oligosaccharide was in the acidic pH range (pH 3.5 approximately 4.0). Total alpha-D-mannosidase activities in the porcine epididymal fluid increased from proximal to distal caput epididymis, which maintained to cauda epididymis. At least two kinds of alpha-D-mannosidase (lysosomal type enzyme and 135 kDa alpha-D-mannosidase (MAN2B2)) were contained in the porcine epididymal fluid. The activity of the lysosomal type enzyme is much higher than MAN2B2 at the physiological pH. These results suggest that the lysosomal type alpha-D-mannosidase is the predominantly active enzyme in the luminal fluid of porcine epididymis and that it participates in the glycoprotein modification on the sperm surface during epididymal transit.Biochimica et Biophysica Acta 08/1999; 1432(2):382-92. · 4.66 Impact Factor -
Article: A porcine homolog of the major secretory protein of human epididymis, HE1, specifically binds cholesterol.
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ABSTRACT: A porcine homolog of the major secretory protein of human epididymis, HE1, was for the first time purified from the porcine cauda epididymal fluid. The HE1 homolog was secreted into the epididymal fluid as a 19-kDa glycoprotein, whose sugar moiety was gradually processed to form a 16-kDa protein during transit through the epididymis. The HE1 homolog mRNA was detected only in the caput and corpus epididymis among the porcine tissues examined. The purified HE1 homolog specifically bound cholesterol with high affinity (Kd=2. 3 microM). The binding stoichiometry was determined to be 0.94 mol/mol, suggesting that 1 mol of cholesterol binds to 1 mol of the protein. It was also found that the HE1 homolog is a major cholesterol-binding protein in the porcine epididymal fluid. The possibility that the HE1 homolog is involved in the regulation of the lipid composition of the sperm membranes during the maturation in epididymis is discussed.Biochimica et Biophysica Acta 07/1999; 1438(3):377-87. · 4.66 Impact Factor -
Article: Role of epididymal secretory proteins in sperm maturation with particular reference to the boar.
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ABSTRACT: This review considers the role of proteins secreted by the epididymis on post-testicular sperm maturation and storage. Two-dimensional gels show that 150 to 200 proteins are secreted into the epididymal lumen. Most are secreted in relatively small amounts; in rams, for example, fewer than ten contribute 90% of the total secretion and only two contribute 52% of the total protein secreted. Most of the proteins are confined to specific regions of the epididymis. The changing pattern of protein secretion along the epididymis corresponds to change in surface protein on spermatozoa, but no epididymal proteins have been identified that appear to be directly involved in modifying the sperm membrane. Most of the major proteins that have been identified seem to be playing a homeostatic role in maintaining the epididymal milieu for spermatozoa.Journal of reproduction and fertility. Supplement 02/1998; 53:99-107. -
Article: High concentration of glucose causes impairment of the function of the glutathione redox cycle in human vascular smooth muscle cells.
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ABSTRACT: We demonstrated that high glucose reduced H2O2 scavenge activity in human vascular smooth muscle cells. In the cells exposed to high glucose, the intracellular glutathione content decreased, although the NADPH content was unchanged. The rate of uptake of cystine, which is a rate-limiting precursor of the glutathione synthesis, decreased in the high glucose group compared with the control group. These decreases were shown to be dependent on glucose concentration. It was suggested that high glucose causes impairment of the function of the glutathione redox cycle in human vascular smooth muscle cells, resulting in reduced H2O2 scavenge activity.FEBS Letters 02/1998; 421(1):19-22. · 3.54 Impact Factor -
Article: Stage-specific expression of a mouse homologue of the porcine 135kDa alpha-D-mannosidase (MAN2B2) in type A spermatogonia.
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ABSTRACT: The cDNA encoding a mouse homologue of porcine epididymis-specific 135kDa alpha-D-mannosidase (MAN2B2, D28521) was cloned from the mouse testis cDNA library. It was found that 1018 amino acids were coded in its open reading frame, and 62% of the amino acid sequence was identical to that of porcine MAN2B2. In the adult mouse, testis contained higher amounts of mRNA encoding the MAN2B2 homologue than the epididymis, though porcine MAN2B2 was mainly expressed in the narrow region between the caput and corpus epididymis. mRNA of the mouse MAN2B2 homologue was localized exclusively in spermatogonia in the testis. It was specifically expressed in type A spermatogonia at stages IX-XI of spermatogenesis and was detected there until the cell developed into type B spermatogonia. We conclude that the expression of the MAN2B2 homologue can serve as a good marker for the late stages of type A spermatogonia and may have an important role to play in the early step of spermatogenesis in mice.Biochemical and Biophysical Research Communications 01/1998; 241(2):439-45. · 2.48 Impact Factor -
Article: Direct evidence for the secretion of lactoferrin and its binding to sperm in the porcine epididymis.
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ABSTRACT: Lactoferrin has been for the first time purified from the porcine cauda epididymal fluid as a 70 kDa protein. Both Western and Northern blot analyses show that lactoferrin is synthesized in the regions from the distal caput to the cauda epididymis and secreted into the luminal fluid. Lactoferrin is first secreted as a 75 kDa glycoprotein and its carbohydrate moieties are gradually digested to form 70 kDa protein in the cauda epididymis. Lactoferrin has already bound to the surface of the epididymal sperm because the anti-lactoferrin antiserum induces the mature sperm tail-to-tail agglutination. These results strongly suggest new physiological functions of lactoferrin on the sperm maturation in the epididymis.Molecular Reproduction and Development 09/1997; 47(4):490-6. · 2.53 Impact Factor -
Article: Molecular cloning and characterization of the epididymis-specific glutathione peroxidase-like protein secreted in the porcine epididymal fluid.
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ABSTRACT: The epididymis-specific glutathione peroxidase was purified from the porcine cauda epididymal fluid in order to analyze its enzymatic activity and roles in the epididymis. The purified protein was found to consist of four identical 23 kDa subunits. The complementary DNA encoding the 23 kDa subunit was cloned from the cDNA library of the porcine proximal caput epididymis, only where the 23 kDa subunit is expressed. Although the selenocysteine codon (TGA) is contained in the cDNA of the other cytosolic type of glutathione peroxidases, it is replaced by cysteine codon (TGT) in the 23 kDa subunit cDNA, similarly to the results previously obtained for cDNAs encoding the epididymis-specific form of the secreted glutathione peroxidases of mouse, rat and monkey. By the direct analysis of the selenium, the purified protein was proved to contain no selenium atom in the molecule. The activities of the purified epididymis-specific glutathione peroxidase toward hydrogen peroxide or organic hydroperoxides were by far lower than the activity of cytosolic selenium-dependent glutathione peroxidase (less than 0.1%). In addition, the concentration of glutathione in the porcine epididymal fluids was about 20 microM, which is much lower than the optimal concentration for the glutathione peroxidase activity of the purified protein. These results strongly suggest that this protein is enzymatically quiescent at least in the porcine epididymal fluid. An immunocytochemical study showed that this protein was found to bind to the acrosomal region of the epididymal sperm and to disappear during the acrosome reaction. Furthermore, this protein significantly retarded the acrosome reaction induced in vitro. The possibilities have been discussed that it protects sperm from the premature acrosome reaction and maintains sperm fertilizing ability in the epididymis.Biochimica et Biophysica Acta 08/1997; 1336(1):99-109. · 4.66 Impact Factor -
Article: Characterization and identification of proteins secreted in the various regions of the adult boar epididymis.
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ABSTRACT: The synthesis and secretion of proteins by the boar genital tract were studied in vitro by incubating epididymal tissues with [35S]methionine and cysteine. Characterization of the major neosynthesized proteins was performed electrophoretically by one- and two-dimensional PAGE analysis, and an epididymal protein cartography was established. Some of the proteins secreted were found to be unregionalized. Polarization studies of the secretions in the epididymal tubule were carried out by in vitro incubation of isolated tubules, and most of these unregionalized proteins were found not to be secreted in the epididymal lumen. Inside the epididymal lumen, protein secretion was highly regionalized, and electrophoresis analysis detected few proteins secreted at all points along the organ. A total of 146 epididymal proteins, covering 220 spots, were found to be secreted by the epididymis. The distal caput showed the highest number of spots, the lowest number of proteins secreted being found in the proximal caput and cauda. Most of the epididymal proteins analyzed are highly polymorphic in terms of both isoelectric point and molecular mass. The presence and importance of the different compounds in the various regions of the epididymis were established. Several distinct secretory regions of the epididymis can be determined by the presence of major characteristic proteins. The concentrations of a given protein in the fluids of various regions were not related to the respective secretion intensity of that protein. Identification of some major epididymal proteins was accomplished by N-terminal amino microsequencing and by the use of specific antisera. Of the various major proteins, clusterin, glutathione peroxidase, retinol-binding protein, lactoferrin, EP4, beta-N-acetyl-hexosaminidase, alpha-mannosidase, and procathepsin L were identified and localized along the organ. Several polypeptides found in this study remain unidentified.Biology of Reproduction 12/1996; 55(5):956-74. · 4.01 Impact Factor -
Article: Cloning of complementary DNA encoding a 135-kilodalton protein secreted from porcine corpus epididymis and its identification as an epididymis-specific alpha-mannosidase.
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ABSTRACT: In the preceding study (Okamura et al., 1992; Biol Reprod 47:1040-1052) we suggested that a 135-kDa protein secreted by porcine epididymis is involved in the sperm maturation. In this work, we have isolated the cDNA clone coding the 135-kDa protein in an effort to investigate its structure and function. The 135-kDa protein was purified from porcine cauda epididymal fluid. Three oligonucleotide probes were synthesized according to the amino acid sequences of N-termini of the native protein and trypsin-digested peptides. A cDNA clone hybridizing with these three probes was isolated from the cDNA library derived from the porcine proximal corpus epididymis. It encodes a novel protein with 1,006 amino acid residues in an open reading frame. Its overall amino acid sequence was significantly homologous (25.7%) to the alpha-mannosidase precursor of Dictiostelium discoideum (P34098). The 135-kDa protein could digest both p-nitro-phenyl-alpha-D-mannoside and high mannose oligo saccharide (Man8-GlcNAc2), strongly suggesting that it is an alpha-mannosidase homologue. The expression of this protein was specific to porcine and was localized to the very narrow parts of epididymis: the border of the caput and corpus epididymis. This protein may serve as a good marker for the functional differentiation in porcine epididymis. A possible role of this protein in the species-specific sperm-egg interaction is discussed.Molecular Reproduction and Development 11/1995; 42(2):141-8. · 2.53 Impact Factor -
Article: Direct evidence for the elevated synthesis and secretion of procathepsin L in the distal caput epididymis of boar.
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ABSTRACT: The proteins which are secreted from the restricted part of the epididymis are suggested to sustain sperm maturation. In porcine species, as the potential abilities of sperm for movement and fertilization greatly increase in the corpus epididymis, the secretions in both the caput and corpus epididymis seem to be very important for the sperm maturation. In this study, we have directed our attention to the 40 kDa protein which is detected in the fluid of the distal caput epididymis of boar. It was purified from the porcine cauda epididymal fluid and its cDNA was cloned from the cDNA library of the distal caput epididymis. According to the deduced amino acid sequence, the 40 kDa protein has been identified as procathepsin L. Northern blot analysis showed that the procathepsin L mRNA was most abundant in the distal caput epididymis among the tissues as examined. Consistent with the distribution of the procathepsin L mRNA in the epididymis, the activity of procathepsin L was absent in the fluid of the proximal and mid caput epididymis and first appeared in the distal caput epididymal fluid, whose contents gradually decreased with the passage through the epididymis. These results first appeared in the first distal caput epididymis expresses very high levels of procathepsin L and unusually secretes it into the luminal fluid instead of targeting it to lysosomes. It has been also found that the mRNA of PDGF, which is known to enhance cathepsin L expression in the culture cells, is very high in the mid caput epididymis, which just precedes the site of procathepsin L secretion. This result indicates that PDGF directly regulates the locally restricted expression and secretion of procathepsin L in the epididymis, which is one of the possible mechanisms involved in the functional differentiation in the epididymis.Biochimica et Biophysica Acta 11/1995; 1245(2):221-6. · 4.66 Impact Factor -
Article: Forskolin stimulates porcine sperm capacitation by increasing calcium uptake.
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ABSTRACT: Using the fluorescent calcium indicator fura-2, forskolin was found to dose-dependently cause an immediate increase in the concentration of intracellular free calcium of porcine cauda epididymal sperm. This stimulatory effect of forskolin is due to the enhancement of Ca2+ uptake by the verapamil-sensitive transporter on the sperm plasma membrane and results in the promotion of the sperm capacitation and subsequent acrosome reaction.FEBS Letters 03/1993; 316(3):283-6. · 3.54 Impact Factor -
Article: Localization of a maturation-dependent epididymal sperm surface antigen recognized by a monoclonal antibody raised against a 135-kilodalton protein in porcine epididymal fluid.
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ABSTRACT: A specific 135-kDa protein was purified from porcine cauda epididymal fluid. Analysis of its N-terminal amino acid sequence revealed it to be a new protein. Stable clones of hybridomas that produced monoclonal antibodies against the purified 135-kDa protein were established. A clone, B-11, reacting both with epididymal fluid and with sperm plasma membranes was selected and used in this study. Immunoblotting analysis showed that B-11 reacted only with a 135-kDa protein among epididymal fluid proteins. In contrast, B-11 did not recognize a similar 135-kDa sperm protein but did strongly react with a 27-kDa protein among sperm membrane proteins, extracted by NP-40 in the presence of protease inhibitors. B-11 also reacted only with a 27-kDa protein fragment among trypsin digests of the 135-kDa epididymal protein. The 135-kDa protein was first detected, by ELISA or immunoblotting analysis, at the beginning of the corpus epididymis. Maximal levels were reached in the distal corpus and levels were slightly decreased in the cauda epididymis. On the other hand, the surface of caput sperm were found to contain small amounts of antigen(s), the concentration of which gradually increased during epididymal transit. In immunocytochemical studies, the antigen was detectable in the epithelial cells from the initial segment to the corpus of the epididymis but not in the caudal cells. In the lumen, the presence of the 135 kDa protein was apparent in the corpus (at a maximum in the middle and distal corpus) and to a lesser degree in the caudal lumen. The 27-kDa protein was distributed all over the equatorial region of the acrosome of less than 10% of caput epididymal sperm. As sperm passed through the corpus epididymis, the percentage of immunoreactive cells increased and the protein was restricted to specific domains of the sperm head. Thus, on the mature sperm, antigen was localized in a crescent-shaped area of the equatorial segment just behind the anterior part of the acrosome and on the apical rim of the sperm head. This is the first observation of a sperm surface antigen derived from an epididymal protein as a proteolytic fragment that interacts with specific regions of the sperm membrane during the process of spermatozoa maturation.Biology of Reproduction 01/1993; 47(6):1040-52. · 4.01 Impact Factor -
Article: Changes in the nature of calcium transport systems on the porcine sperm plasma membrane during epididymal maturation.
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ABSTRACT: Comparative studies of 45Ca(2+)-transport across the plasma membrane were performed using porcine caput, corpus and cauda epididymal sperm. The Ca(2+)-uptake is dependent on the presence of the substrates for respiration and is sensitive to verapamil. The Ca(2+)-efflux is mediated by both Na(+)-dependent and -independent systems. In the immature sperm in caput epididymis, Na(+)-independent efflux is predominant, but it is gradually replaced by Na(+)-dependent efflux during the epididymal transit. The net activity of Ca2+ accumulation into sperm increases with the epididymal maturation.Biochimica et Biophysica Acta 08/1992; 1108(1):110-4. · 4.66 Impact Factor -
Article: Purification of bicarbonate-sensitive sperm adenylylcyclase by 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid-affinity chromatography.
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ABSTRACT: Anion transport inhibitors, such as SITS (4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid) and heparin, inhibit reversibly the bicarbonate-sensitive adenylylcyclase of porcine sperm plasma membrane. In the light of this, SITS- and heparin-affinity chromatographies were applied in order to purify sperm adenylylcyclase. SITS-Affi-Gel 102 binds proteins extracted from the porcine cauda epididymal sperm plasma membrane by Lubrol-PX, more selectively than heparin-agarose. However, recovery of adenylylcyclase activity is higher when heparin-agarose is used. The hormone-sensitive liver adenylylcyclase, which is less sensitive to bicarbonate than sperm enzyme, has less affinity for these affinity resins than sperm enzyme. Adenylylcyclase can be purified to apparent homogeneity on two-dimensional gel electrophoresis (isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis) from the Lubrol-PX extract of the purified sperm plasma membrane by using SITS-affinity chromatography at the first step of the purification followed by preparative isoelectric focusing and gel filtration. The molecular weight and pI of the purified enzyme are 46,300 and 6.9, respectively. The purified enzyme activity is highly dependent on Mn2+. Bicarbonate activates even the purified enzyme both by decreasing Km and by increasing Vmax.Journal of Biological Chemistry 10/1991; 266(27):17754-9. · 4.77 Impact Factor -
Article: Water insoluble fraction of egg yolk maintains porcine sperm motility by activating adenylate cyclase.
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ABSTRACT: The decrease in motility of porcine cauda epididymal sperm was less than that of caput epididymal sperm in the medium containing bicarbonate. This may be due to the difference of sensitivity of adenylate cyclase to bicarbonate between mature and immature sperm; activation of mature sperm enzyme by bicarbonate was higher than that of immature sperm. Nondialysable fraction of egg yolk prevented the decrease in motility of immature sperm in the presence of bicarbonate, but it was not effective for the motility of mature sperm under the same condition, because only bicarbonate is sufficient for the maintenance of its motility. In the absence of bicarbonate, both mature and immature sperm required egg yolk to maintain motility. The favorable effect of egg yolk on the motility is ascribed to the enhancement of intracellular cAMP level. Partial fractionation of egg yolk showed that water-insoluble lipoprotein fraction contains factor(s) which activates adenylate cyclase in sperm plasma membrane. This is the first report in which high molecular weight activator of the sperm enzyme was demonstrated.Molecular Reproduction and Development 03/1991; 28(2):136-42. · 2.53 Impact Factor -
Article: Effects of a membrane-bound trypsin-like proteinase and seminal proteinase inhibitors on the bicarbonate-sensitive adenylate cyclase in porcine sperm plasma membranes.
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ABSTRACT: Plasma membranes were purified from flagella of porcine cauda epididymal sperm and proteolytic regulation of bicarbonate-sensitive adenylate cyclase was studied. It was found that the epididymal sperm plasma membrane contained a trypsin-like proteinase which inactivated adenylate cyclase. Bicarbonate activates adenylate cyclase as reported previously, but, at the same time, the anions enhance the inactivation of the enzyme by the membrane-bound trypsin-like proteinase. This phenomenon is not due to the direct activation of the proteinase, but closely related to the activation of adenylate cyclase by bicarbonate. It was also found that seminal proteinase inhibitors blocked the inactivation of adenylate cyclase and maintained the bicarbonate activation of the enzyme at high level. Actually, bicarbonate keeps adenylate cyclase fully active in ejaculated sperm, because membrane-bound proteinase is completely inhibited by the seminal proteinase inhibitors. These results suggest that the interactions between membrane-bound proteinase and seminal proteinase inhibitor are involved in the regulation of the bicarbonate-sensitive adenylate cyclase system.Biochimica et Biophysica Acta 08/1990; 1035(1):83-9. · 4.66 Impact Factor -
Article: The enhancing effects of anion channel blockers on sperm activation by bicarbonate.
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ABSTRACT: We found that anion channel blockers such as phosphotungstate and 4,4'-diisothiocyanatostilbene-2,2'-disulfonate (DIDS) enhanced HCO3(-)-induced activation on porcine epididymal sperm. In the presence of these compounds, HCO3- increased the motility, respiration rate and especially the cAMP content of the sperm to a greater extent than did HCO3- alone. The enhancing effects were not observed in the absence of HCO3-, but were evident when the concentration of HCO3- was low. These compounds did not significantly alter the intracellular pH and did inhibit the adenylate cyclase activity of the sperm plasma membrane. When these compounds were added to sperm homogenate with ATP, the cAMP formed was reduced compared to the control. In addition, these compounds inhibited both the SO4(2-) influx and efflux of the sperm. From these results, we conclude that the anion channel blockers tested principally inhibit the efflux of endogenous HCO3- derived from metabolic CO2, so that HCO3- accumulates intracellularly and stimulates the adenylate cyclase of the sperm.Biochimica et Biophysica Acta 07/1990; 1034(3):326-32. · 4.66 Impact Factor
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1989–2009
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University of Tsukuba
- • Department of Urology
- • Institute of Basic Medical Sciences
Tsukuba, Ibaraki-ken, Japan
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