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Query: UNIPROT:P56851 (epididymal)
 11,273 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The relationship between thiol-disulfide status and acridine orange fluorescence of testicular, epididymal, and ejaculated spermatozoa in several mammalian species was investigated. Spermatozoa were fixed with acetic alcohol, stained with acridine orange, and examined with a fluorescence microscope. The majority of the nuclei of testicular spermatozoa of the hamster, mouse, and rabbit exhibited red acridine orange fluorescence. The proportion of sperm nuclei with red acridine orange fluorescence decreased as the spermatozoa descended the epididymis. Red acridine orange fluorescence was replaced by green acridine orange fluorescence. The site in the epididymis where 100% of the nuclei exhibited green fluorescence was the distal caput in the mouse, the corpus in the rabbit, and the proximal cauda in the hamster. In semen samples from men with proven fertility, normal semen parameters, or both, about 60% to 90% of the nuclei exhibited green acridine orange fluorescence. The proportion of sperm nuclei exhibiting green acridine orange fluorescence was higher in the spermatozoa pellet (containing highly motile spermatozoa) obtained by centrifugation through a Percoll gradient. From experiments using disulfide-reducing, thiol-oxidizing and thiol-detecting agents, we concluded that sperm nuclei fluoresce red when they are treated with acid while their DNA-associated protamines are poor in disulfides. Under such conditions, DNA is vulnerable to denaturation. Acridine orange binds to denatured (single-stranded) DNA as aggregates and emits red fluorescence. In contrast, when sperm nuclei are treated with acid while their DNA-associated protamines are rich in disulfides, DNA is resistant to denaturation. Acridine orange binds to native (double-stranded) DNA as a monomer and emits green fluorescence.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Thiol-disulfide status and acridine orange fluorescence of mammalian sperm nuclei. 139 37

Flow cytometric measurements were made on acridine orange (AO) and 7-diethylamino-3-(4'-maleimidylphenyl)-4-methyl-coumarin (CPM)-stained epididymal- and vas deferens-derived spermatozoal nuclei to follow the course of chromatin condensation and oxidation of free sulfhydryl groups, respectively, during passage through mouse and rat posttesticular reproductive tracts. Alterations of mouse and rat spermatozoal chromatin during transition from a testicular elongated spermatids to epididymal caput spermatozoa resulted in a threefold loss of DNA stainability with AO. Passage of spermatozoa from the caput to corpus epididymis was accompanied by an approximate 15% loss of DNA stainability, which was maintained at that level throughout passage into the vas deferens. AO stainability of epididymal spermatozoal nuclei was generally independent of -SH group stainability. CPM stainability of rat spermatozoal nuclei free -SH groups was 83%, 18%, and 11% of caput spermatozoal values for corpus, cauda epididymis, and vas deferens, respectively. Comparable values for mice were 69%, 20%, and 18%. CPM stainability was relatively homogeneous for these mouse and rat reproductive tract regions, except mouse corpus epididymis spermatozoal nuclei stained very heterogeneously. Rat spermatozoa detained by ligature up to 7 days in the caput, corpus, and cauda epididymi had CPM staining values equal to or below those of normal vas spermatozoa, indicating that disulfide (S-S) bonding is intrinsic to the spermatozoa and is independent of the epididymal environment. These data suggest that chromatin condensation and loss of spermatozoal DNA stainability during passage from the testis to the vas deferens are independent of S-S bonding.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Flow cytometric analysis of rodent epididymal spermatozoal chromatin condensation and loss of free sulfhydryl groups. 248 18

Exposure of 100-d old rats to 1,3-dinitrobenzene (m-DNB) at dosages up to 48 mg/kg resulted in disruption of spermatogenesis as measured by flow cytometry (FCM) of acridine orange-stained sperm and testis cells. One day (d 1) after a single exposure to 48 mg/kg m-DNB, FCM measurements of caput epididymal fluid cells demonstrated the presence of testicular germinal epithelial cells apparently sloughed off into the epididymis. Also, at d 1 after the same exposure, a decrease in pachytene spermatocytes was observed. By d 16 after exposure to 32 or 48 mg/kg, testicular damage was evidenced by an alteration of cell type ratios in FCM-analyzed populations of testicular cells. Extensive recovery of cell type ratios occurred by d 32. At d 16, dosages of 32 and 48 mg/kg caused alterations of sperm chromatin structure as determined by the flow cytometric sperm chromatin structure assay (SCSA); 48 mg/kg caused alterations at both d 16 and d 32. Exposure to m-DNB caused a dose response increase in percent sperm head morphology abnormalities (%ABN) assessed in cauda epididymal and vas sperm. A slightly higher correlation existed between dose and SCSA alpha t values (d 16, .78; p less than .01) than between dose and %ABN (d 16, .70; p less than .01). Also, a higher correlation existed between standard deviation of alpha t (SD alpha t) values and %ABN (.97; p less than .01) than between dose and %ABN (.70; p less than .01). This study demonstrated rapid and unique FCM procedures originally derived for reproductive toxicology studies in mice to be equally useful for studies in rats.
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PMID:Flow cytometric analysis of effects of 1,3-dinitrobenzene on rat spermatogenesis. 277 50

Multiparameter flow cytometry (FCM) measurements were made on acridine orange (AO)-stained mouse testicular cells and epididymal sperm cells to determine the effects of varying dosages of thiotepa (0-5 mg/kg ip daily X 5 days) on spermatogenesis at 7, 28, and 67 days after the last exposure (ALE). FCM multiparameter measurements included DNA stainability vs RNA content, peak amplitude vs integrated area of DNA fluorescent signal, and double-stranded DNA vs single-stranded DNA. Thiotepa exhibited dramatic damaging effects on the kinetics and/or cell kill of seven testicular cell types measured by dual-parameter flow cytometry. At 7 days ALE, one 4N cell type, likely the pachytene spermatocyte, was absent from the testes, and another was reduced by about 70%. By 28 days ALE, most of the germ cells were absent from the seminiferous tubules, and by 67 days ALE the testes were undergoing recovery of spermatogenesis with only half of the seminiferous tubules repopulated after treatment with 5.0 mg/kg. The dual parameters of DNA stainability vs RNA content provided better resolution of testicular cell types into distinct populations than the peak vs area processing of the green fluorescent signal of AO-stained cells. Dosage of thiotepa was significantly related to percentage of sperm head morphological abnormalities assayed by light microscopy. Utilizing the metachromatic properties of acridine orange, FCM measurements of the amount of single-stranded DNA induced within acid-stressed whole sperm or heat-stressed nuclei detected alterations of chromatin structure at the same minimal effective dose required to increase abnormal sperm head morphology. Epididymal sperm isolated from mice exposed to some concentrations of thiotepa had an increased percentage of free heads and tails. DNA in free heads denatured in situ to a greater extent than DNA in intact sperm.
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PMID:Toxicity of thiotepa on mouse spermatogenesis as determined by dual-parameter flow cytometry. 308 Aug 21

The effects of hydralazine (1-hydrazinophthalazine), an antihypertensive drug, on mammalian cell growth, viability, and differentiation were assessed using Friend leukemia cells, Chinese hamster ovary cells, human lymphocytes, and rat lymphocytes, testicular germ cells, and epididymal sperm. Cultured cells in exponential phase growth were more susceptible to hydralazine cytotoxicity than stationary phase (G0) cells. Growth inhibition was associated with a dose-related slowdown of cell progression through S phase and was observed prior to a decrease of cell viability. At high drug concentrations, progression in all phases of the cell cycle was partially or totally inhibited. Hydralazine did not have an effect on the proliferation and differentiation of testicular germ cells in spontaneously hypertensive rats receiving 0-90 mg/kg/day (up to 20 times the dose used in humans) of hydralazine for a 12-week period. Hydralazine-exposed, histone-containing somatic cells and protamine-containing sperm cells failed to show any alterations in stainability with a DNA-intercalating dye nor in the susceptibility of nuclear DNA to undergo acid-induced denaturation in situ. The data suggest that hydralazine causes a dose-related suppression of mammalian cell growth with S phase appearing to be the most susceptible to hydralazine cytotoxicity. Furthermore, the interaction of hydralazine with chromatin at concentrations leading to antigenicity did not inhibit DNA staining with the intercalating dye acridine orange, suggesting that the drug does not competitively intercalate at a detectable level. Association of hydralazine with chromatin did not cause a detectable level of stabilization or destabilization of the DNA to denaturation in situ.
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PMID:In vivo and in vitro effects of hydralazine on cellular growth, differentiation, and chromatin structure. 335 68

Dual-parameter (DNA, RNA) flow cytometry (FCM) measurements were made on testicular and epididymal sperm cells isolated from mice exposed by oral gavage to 0, 250, 500, or 1000 mg/kg X 5 d of the fungicide methylbenzimidazol-2-yl carbamate (MBC), which is known to bind with tubulin subunits and inhibit polymerization and microtubule formation. Effects of exposure to MBC were measured at 7, 24, and 39 d posttreatment. MBC had no effect on body weights, but testis weights and sperm parameters were altered, with few exceptions, only at the highest exposure level. Testis weights were reduced by about 25% at 7 and 24 d after exposure; recovery was observed by 39 d after treatment. FCM measurements of testicular cells showed relative percentages of certain testicular populations (round, elongating, and elongated spermatids) were different from the control pattern 7 and 24 d after treatment. The mean percent of cauda epididymal sperm head morphology abnormalities and the susceptibility of the nuclear DNA to denaturation were both elevated at 7, 24, and 39 d after exposure to 1000 mg/kg. The level of denaturation was determined by FCM measurements of the metachromatic shift in acridine orange (AO) stained sperm nuclei from green (native DNA) to red (single-stranded DNA) fluorescence and quantitated by the expression alpha t[red/(red + green] fluorescence. These data demonstrate that spermatogenesis is sensitive to high-dose MBC exposure resulting in an altered ratio of testicular cell types present, abnormal sperm head morphology, and an altered sperm chromatin structure.
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PMID:Effects of the fungicide methyl-benzimidazol-2-yl carbamate (MBC) on mouse germ cells as determined by flow cytometry. 356 Feb 61

The effects of the mutagenic agent ethylnitrosourea (ENU) on spermatogenic function and sperm chromatin structure were studied by flow cytometry and the results compared with sperm head morphology measurements. Groups of mice received daily exposures ranging from 0 to 75 mg/kg body weight X 5 days and were sacrificed 28 days later. Fresh testicular cell suspensions and epididymal sperm were stained with acridine orange (AO) and measured by flow cytometry. Sperm nuclei were isolated, fixed, rehydrated, and then either subjected to thermal stress or not prior to staining with AO. Body weights were unaffected by the chemical exposure while the testicular weights were reduced by about 50%. Two-parameter (DNA, RNA) flow cytometry measurements showed a dose-response relationship in the loss of certain cell types, particularly the elongated spermatids, from the testes of treated animals. Flow cytometric analysis of both heat-stressed and non-heat-stressed nuclei showed a relationship between dosage and the coefficient of variation of alpha t [red/(red + green fluorescence)] measurements of AO stained nuclei, thereby demonstrating that alterations of chromatin structure occurred in response to ENU. Enzymatic digestions with RNAse, DNAse, and nuclease S1 suggest that the increase in red fluorescence is due to an increase of single-stranded DNA induced by heat or acid treatment of chemically altered chromatin structure. The lowest daily dosage used (5 mg/kg) caused no significant changes in ratios of testicular cell types, a questionable increase in abnormal sperm head morphology and a detectable change in chromatin structure expressed as alpha t. This report shows that our technique for assaying sperm nuclear chromatin structure appears to have the same level of sensitivity to ENU induced nuclear alterations as the sperm head morphology test.
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PMID:Flow cytometric analysis of mouse spermatogenic function following exposure to ethylnitrosourea. 399 39

The techniques of Feulgen staining, acridine orange staining, and a sperm chromatin structure assay using acridine orange and flow cytometry were compared for selective examination of bovine sperm nuclei. Twenty frozen semen samples were simultaneously analysed by all three methods. The prevalence of abnormally condensed DNA and its relationship to other semen traits were determined in ejaculates from 70 bulbs presented for routine examination for breeding soundness and in frozen semen from 348 bulls evaluated over five years. A breeding trial with 118 beef heifers using semen from six bulls with different degrees of nuclear abnormalities was performed to assess the importance of the defects with respect to fertility. The results indicate that few spermatozoa with abnormal DNA condensation are found in normal semen, but the incidence increases with disturbance of spermatogenesis. However, high numbers of abnormally condensed nuclei were found in the absence of an increase in other defects. This nuclear defect might be at least partially of epididymal origin; it can lower fertility and can be compensated for by increasing the numbers of normal spermatozoa in the insemination dose. The percentage of abnormally condensed sperm nuclei as detected by Feulgen staining was significantly correlated with that detected by microscopy after acridine orange staining and by the sperm chromatin structure assay. We therefore consider the Feulgen technique to be a valuable tool for assessing the nuclear integrity of bovine spermatozoa.
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PMID:Flow cytometric and microscopic evaluation and effect on fertility of abnormal chromatin condensation in bovine sperm nuclei. 752 53

During passage of hamster spermatozoa through the epididymis their maturation is shown to involve changes in the sperm head, midpiece (mitochondria) and tail. The sum of these changes results in a dramatic increase in the fertilizing potential of the spermatozoa. When comparable numbers of spermatozoa from the caput or corpus epididymis were injected into one uterine horn of mature females, following ovulation induction, and spermatozoa from the cauda epididymis were injected into the contralateral horn, no fertilization was observed with caput epididymal spermatozoa, 1.7% of oocytes were fertilized by corpus epididymal spermatozoa, whereas 79.5% fertilization was obtained with cauda epididymal spermatozoa. Total sperm numbers increased from caput to corpus to cauda [28.3 +/- 12.2, 40.6 +/- 20.8, 144 [corrected] +/- 62 million, respectively]. The percentage of progressively motile spermatozoa increased from 27.9 +/- 6.4 to 33.8 +/- 4.8 to 70 +/- 10.7 during this passage. Viability, measured by exclusion of the dye, propidium iodide, was significantly less in spermatozoa from the cauda than from the proximal or mid-caput epididymis. The percentage of the live cells that were stained intensely by rhodamine-123 (a measure of mitochondrial membrane potential) increased during epididymal passage from 22.8 +/- 7.8% in the proximal caput epididymis to 57.2 +/- 16.5% in the cauda epididymis. Staining with acridine orange (a measure of DNA packaging in the sperm head) indicated an increase in chromatin condensation in cauda epididymal spermatozoa, when compared to those obtained from the caput or corpus.
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PMID:Investigation of epididymal sperm maturation in the golden hamster. 769 51

The testicular regions of male mice were exposed to x-ray doses ranging from 0 to 400 rads. Forty days after exposure the mice were killed and the testes and cauda epididymal sperm removed surgically. Flow cytometric measurements of acridine orange stained testicular samples indicated a repopulation of testicular cell types following x-ray killing of stem cells. Cauda epididymal sperm were analyzed by the sperm chromatin structure assay (SCSA), a flow cytometric measurement of the susceptibility of the sperm nuclear DNA to in situ acid denaturation. The SCSA detected increased susceptibility to DNA denaturation in situ after 12.5 rads of x-ray exposure, with significant increases following 25 rads. Abnormal sperm head morphology was not significantly increased until the testes were exposed to 60 rads of x-rays. These data suggest that the SCSA is currently the most sensitive, non-invasive method of detecting x-ray damage to testicular stem spermatogonia.
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PMID:Effects of X-irradiation on mouse testicular cells and sperm chromatin structure. 787 23


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