Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P56851 (epididymal)
 11,273 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The toxicities of theobromine and cocoa extract on the reproductive tract of male rats were compared in the present study. A cocoa powder extract containing 117 mg theobromine/g extract was prepared using 85% boiling methanol. Sprague-Dawley rats were weighed and dosed daily for 31 days with vehicle, 250 mg/kg theobromine, 2.14 g/kg cocoa extract (117 mg theobromine/g extract), or 0.43 g/kg cocoa extract by oral gavage. The animals were sacrificed on day 32. One testis and epididymis were removed and weighed. The epididymis was saved for the determination of epididymal sperm reserves. The remaining testis was fixed by whole body glutaraldehyde perfusion and processed for morphologic examination. A decrease in body weight gain and epididymal weights were observed in theobromine and high-dose cocoa-extract-treated groups. Theobromine and high-dose cocoa extract caused vacuolation within the Sertoli cell, abnormally shaped spermatids, and failed release of late spermatids in treated animals. Most of the vacuolations were found in the earlier and middle stage seminiferous tubules (stages I to VIII). However, the frequency of some parameters of testis alterations were significantly lower in the high-dose cocoa-extract-treated group compared to the theobromine-treated group. These data demonstrate the ability of a cocoa extract containing theobromine to alter testis structure in a similar pattern but with reduced intensity compared to that observed after oral exposure to pure theobromine.
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PMID:Reproductive toxicity of theobromine and cocoa extract in male rats. 152 Oct 8

The murine monoclonal antibody H316 reacts with a cell-surface antigen of human trophoblast, leukocytes, certain epithelia, and several malignant cell types. We have found that the H316 antibody also recognizes an antigen synthesized by pre- and post-meiotic human testicular germ cells and is expressed in the acrosomal region of methanol-fixed testicular, epididymal, and ejaculated sperm. The antigen is poorly expressed on the surface of fresh ejaculated motile sperm, but is detectable on most viable sperm after a 6-h incubation in medium containing human serum albumin (HSA), or 60-min incubation with the calcium ionophore A23187 (both treatments induce sperm acrosomal changes termed capacitation and acrosome reaction). We found that antigen recognized by H316 is immunoprecipitated as a single, broad 50 kDa band from radiolabeled ionophore-treated sperm extracts and that preincubation of HSA-capacitated sperm with this antibody causes a moderate, but significant, inhibition of hamster egg penetration. These data indicate that the antigen recognized by the H316 monoclonal antibody is synthesized by testicular germ cells and is surface-expressed on capacitated/acrosome-reacted sperm populations. Its potential as a human sperm acrosome reaction marker, and possible biological role in sperm-egg or sperm-lymphocyte interactions, warrants further investigation.
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PMID:Trophoblast/leukocyte-common antigen is expressed by human testicular germ cells and appears on the surface of acrosome-reacted sperm. 252 14

Using a partially purified enzyme preparation obtained from hamster epididymis, a simple assay has been developed to measure the sulfurylation of dehydroisoandrosterone (DHA) and desmosterol in the presence of 3'-phosphoadenosine 5'-phospho[35S]sulfate [( 35S]PAPS). After stopping the enzymatic reaction with methanol and KCl, the 35S-labelled steroid sulfates are readily extracted into an organic phase. Optimal conditions for the sulfurylation of the two steroids were compared; optimum pH is 8.7 for DHA and 9.8 for desmosterol. Sulfoconjugation of desmosterol increases with magnesium concentrations up to 6 mM, while 40 mM concentrations of the divalent ion are required for the optimal sulfurylation of DHA. Maximum sulfurylation of these steroids requires the presence of 15 mM cysteine. Michaelis-Menten kinetics are observed with DHA which has an apparent Km of 32 microM, while desmosterol inhibits sulfotransferase activity at high concentrations. Saturation of the enzyme with PAPS results in an allosteric behaviour. Only the 3 beta-hydroxyl function of the steroid nucleus appears to be an appropriate sulfate acceptor for the epididymal hydroxysteroid sulfotransferase.
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PMID:The assay and partial characterization of 3 beta-hydroxysteroid sulfotransferase of the hamster epididymis. 315 30

A monoclonal antibody generated against hamster epididymal spermatozoa and recognizing an antigen within the acrosome was used in conjunction with FITC-antimouse immunoglobulin as a marker of the human acrosome during sperm development, capacitation, and the acrosome reaction. The specificity of binding of the monoclonal antibody was assessed using immunolocalization by epi-fluorescence and electron microscopy. Immunofluorescence revealed that antibody bound over the entire anterior acrosome in hamster and human spermatozoa. Ultrastructural localization indicated that antigen was predominantly present on the inner face of the outer acrosomal membrane and within the acrosomal content. Qualitative specificity was studied using a highly purified preparation of hamster acrosomes in an enzyme-linked immunosorbent assay. Since the antibody rapidly visualized human acrosomes, it was used to detect abnormal acrosome morphology of mature spermatozoa and to mark spermatids present in the ejaculate. During incubation in capacitating medium, changes in the immunofluorescence of live or methanol fixed spermatozoa were correlated with incubation interval and the ability of spermatozoa to fuse with zona-free hamster oocytes. Spermatozoa bound to zona-free hamster oocytes displayed no fluorescence, confirming that acrosome loss occurred before spermatozoa attached to the vitellus.
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PMID:Visualization and characterization of the acrosome reaction of human spermatozoa by immunolocalization with monoclonal antibody. 333 57

Protein-bound steroids can be separated from free steroids using microcolumns of silica gel coated with an hydrophobic (octadecyl) solid phase. The bound fraction is eluted in the assay buffer, whereas the free fraction is retained quantitatively on the column in the first step and can be recovered in methanol. Both fractions can be quantitated directly (e.g. by liquid scintillation spectrometry when using radioactive ligands) or kept for further analysis (e.g. by TLC, HPLC etc.). Separation of the bound and free fractions is rapid, accurate and reproducible; intra- and inter-assay coefficients of variation are lower than 5 and 10%, respectively. Recovery of radioactive steroids is high (usually over 85%) and can be estimated separately for each sample. Since assay blanks are very low (typically less than 0.1% of input), this new method, which could be termed "hydrophobic interaction chromatography" (HIC), should prove especially useful for the development of sensitive binding assays, particularly in the field of steroid receptors. The HIC method compared well with three methods currently used for steroid binding assays, namely adsorption of unbound steroids on dextran-coated charcoal, gel filtration on Sephadex LH-20 and adsorption of steroid-protein complexes on DEAE-cellulose filters. Examples of application described here include studies on human plasma sex hormone binding globulin (SHBG) and SP2 placental protein (saturation analysis, binding specificity etc.), the separation of antibody-bound steroids in a radioimmunoassay and the estimation of androgen binding to rat epididymal androgen binding protein (rABP). Receptor assays are illustrated by saturation analysis of the mouse uterine oestrogen receptor and of the androgen receptor in the human genital skin.
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PMID:Hydrophobic interaction chromatography (HIC) for the separation of protein-bound and free steroids. Application to binding protein and receptor assays. 409 24

Monoclonal antibodies were prepared against rabbit sperm antigens by fusing P3-X63-Ag8-653 mouse myeloma cells with lymphocytes from Balb/c mice immunized with Tergitol NP-40 detergent-solubilized rabbit epididymal sperm. Ascites fluid from mice injected with two of these hybridomas (8C4.1 and 8C10.5) was negative in immobilization and agglutination methods, however, acrosome positive on methanol fixed sperm and plasma membrane positive on unfixed sperm in indirect immunofluorescence. Insemination of female rabbits with the sperm treated with either of these monoclonal antibodies resulted in significant reduction in fertility as seen by the percentage of 9-day implants/corpora lutea ratio (8C4.1, 25.7%; 8C10.5, 1.9%; and control, 64.7%). Though the antibodies inhibited in vitro binding of the rabbit sperm with zonae pellucidae of rat ova, fertilization in vivo was not affected significantly. The antibodies did not demonstrate antiblastocyst activity by immunofluorescence. Both of these monoclonal antibodies appeared to recognize the same antigen by the SDS-PAGE/Protein Blot enzyme immunobinding procedure. The antigen was of testicular origin and had a molecular weight of approximately 63,000 daltons. It is concluded that these monoclonal antibodies which were organ specific, block post-fertilization fertility by inhibiting some step necessary for viable embryo formation.
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PMID:Inhibition of fertility in rabbits by monoclonal antibodies against sperm. 618 81

The ejaculated porcine spermatozoa were fractionated into the cytosol, membrane, midpiece plus tail (flagella) and head fractions, and their adenylate cyclase activities were measured. About 65% of the total activity was located in the flagella fraction. For all the fractions, Mn2+-dependent adenylate cyclase activity was about 20 times higher than Mg2+-dependent activity, and guanine nucleotides, fluoride, and other reagents tested did not activate adenylate cyclase. The results suggest that the GTP-dependent regulatory subunit is absent in porcine spermatozoa. The porcine seminal plasma was found to stimulate the adenylate cyclase activity in spermatozoa. The stimulating factor in porcine seminal plasma was partially purified by gel filtration and the molecular weight of the factor appeared to be between 200 and 300. The partially purified factor is heat stable and is not inactivated by treatment with Pronase, trypsin, phospholipase A2 or D but is inactivated by acid hydrolysis. It is easily soluble in water, partially soluble in methanol, and insoluble in ethanol, ethyl ether, chloroform, or acetone. The activation of sperm adenylate cyclase by the factor occurred without a time lag. The activating effect was dose-dependent, saturated at high dose, and ascribed to the increase of the maximal velocity (Vmax). The effect of the factor appears to be limited to adenylate cyclase in spermatozoa; the factor activated adenylate cyclase both in porcine and bovine spermatozoa but failed to activate those in other porcine tissues. The factor was shown to activate the enzyme not only in the ejaculated spermatozoa but also in the epididymal sperm. The factor was also found to elevate the cAMP level in the intact porcine spermatozoa. The factor enhanced the motility of corpus and cauda epididymal spermatozoa. These findings indicate the possibility that this factor initiates the spermatozoan motility upon ejaculation through directly activating adenylate cyclase.
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PMID:Activation of spermatozoan adenylate cyclase by a low molecular weight factor in porcine seminal plasma. 663 Feb 19

A Mr = 32,000 membrane glycoprotein can be uniquely labeled by galactose oxidase/[3H]sodium borohydride on rat caudal, but not caput, epididymal sperm. It has been suggested that this protein is related to a Mr = 32,000 galactose oxidase-sensitive glycoprotein present in rat caudal epididymal fluid. The tritiated membrane glycoprotein was solubilized and its hydrodynamic properties were determined by conventional gel filtration, high performance gel filtration, sedimentation rate determination in linear sucrose gradients prepared in H2O and D2O, and equilibrium isopycnic centrifugations in CsCl. The Stokes radius and sedimentation coefficient were 4.87 +/- 0.07 nm and 1.73 +/- 0.08 S, respectively. The sedimentation profile in CsCl gradients was asymmetric with a major peak occurring at a density of 1.081 g/cm3 (v = 0.92 cm3/g) and a shoulder at 1.108 g/cm3 (v = 0.90 cm3/g). The glycoprotein did not enter a 5 to 20% linear sucrose gradient prepared in D2O and could be extracted from the intact sperm into acidic chloroform:methanol solutions. These data are consistent with a protein which binds substantial amounts of detergent and/or lipid and has exposed hydrophobic regions. Two-dimensional gel electrophoresis indicated that the membrane protein exhibits charge heterogeneity, with the major components having pI values of 5.4 and 4.9. The fluid glycoprotein was monodisperse on two-dimensional gel electrophoresis having a pI of 3.8. Binding studies failed to demonstrate specific binding of the Mr = 32,000 caudal fluid glycoprotein to caput cells. Moreover, "Western blots" of electrophoretically resolved caput and caudal fluid proteins, followed by immunolabeling with antibodies raised against unfractionated caudal fluid, demonstrated the presence of a Mr = 32,000 protein in caudal fluid which was absent from caput epididymal fluid. Using the same technique, it was shown that antibodies raised against caudal fluid proteins did not cross-react with a Mr = 32,000 caudal membrane glycoprotein. Our data do not support the view that the Mr = 32,000 fluid and membrane proteins are identical.
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PMID:Characterization of a maturation-associated glycoprotein on the plasma membrane of rat caudal epididymal sperm. 669 99

A more sensitive assay for protein methylesterase (PME) was developed. The new assay measures the methanol formed from hydrolysis of protein-methyl esters instead of methyl esters remaining on proteins. The formation of methanol is linear with time and enzyme concentration up to 20% of substrate hydrolysis. With this assay, we have detected very low PME activity in rat spermatozoa from cauda epididymidis. However, PME activity in spermatozoa from caput epididymidis was 10-fold higher. In a more detailed study on bull spermatozoa during epididymal maturation, we observed that PME activity was low in testicular spermatozoa, increased in spermatozoa from caput epididymidis to a maximum of 1.29 pmol/mg protein, and progressively decreased to a very low level (0.06 pmol/mg protein) in spermatozoa from cauda epididymidis. The drop in PME activity during epididymal transit parallels the progressive acquisition of motility and fertilizing capacity by spermatozoa.
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PMID:Sensitive assay detects protein methylesterase in spermatozoa: decrease in enzyme activity during epididymal maturation. 673 2

The present report describes in vitro experiments with golden hamster sperm designed to determine whether there is any relationship between sperm phospholipid methylation and capacitation and/or the acrosome reaction. Washed cauda epididymal hamster sperm were incubated in a capacitation medium containing [methyl-3H] methionine. After 0.5, 1.5, 2.5 and 3.5 h of incubation, sperm were extracted with a chloroform:methanol:2 N HCl mixture to extract total phospholipids. Liquid scintillation counting revealed that the methyl-3H-group was incorporated into phospholipids with maximum incorporation at 3.5 h and an increase of 50% between 2.5 and 3.5 h. Uptake of labeled methionine by sperm reached its plateau by 1.5 h of incubation. Some sperm were capacitated by 3.5 h because that is the time at which the rate of acrosome reactions began to increase and because at least 50% of them were able to undergo the acrosome reaction 10 min after the addition of the fusogen lysophophatidylcholine (LPC) at 3.5 h but not at 2.5 h. Homocysteine thiolactone and 3-deazadenosine, inhibitors of transmethylation, inhibited incorporation of methyl-3H into phospholipids at 3.5 h by approximately 90% and also inhibited LPC-induced acrosome reactions by 60%. Separation of methylated sperm phospholipid by thin-layer chromatography demonstrated the presence of 3H-labeled phosphatidyl-N-monomethylethanolamine, phosphatidyl-N,N-dimethylethanolamine, and to a lesser extent phosphatidylcholine. In addition, an unidentified lipid was also highly labeled. These results strongly suggest a positive correlation between phospholipid methylation and capacitation and/or the acrosome reaction of hamster sperm in vitro. Possible mechanisms for phospholipid methylation involvement in these events are discussed.
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PMID:Phospholipid methylation increases during capacitation of golden hamster sperm in vitro. 687 7


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