PubMed Health⌕ Search

Biomedical subjects

S Blottner

Publications and source records attributed to S Blottner.

34 records · Page 2Linked to original sources
PubMed1994

Enrichment of bovine X and Y spermatozoa by free-flow electrophoresis.

Conditions of separation of bovine spermatozoa according to their differences in electrophoretic mobility were studied using free-flow electrophoresis. Distribution of spermatozoa in the electric field depends on composition of buffer system, field intensities and temperature. Conditions are described for reproducible separation of anodically migrating spermatozoa into two distinct peaks both for cold-immobilized (10-13 degrees C) and for motile cells (25 degrees C). Electrophoresis with the buffer used at 25 degrees C and field intensities of 70-100 V/cm provided high percentages of motile spermatozoa (50-90%) in all fractions. However, the sperm distribution across the fractions was the result of superimposed passive electrophoretic motion towards the anode and the active galvanotactic migration towards the cathode. Separation of X- and Y-spermatozoa was verified by quantification of Y-spermatozoa by means of both in situ hybridization, using Y-specific DNA fragments, and the Y-specific fluorescence staining (f-body test). Y-spermatozoa could be enriched in anodic fractions (112 and 144% relative to control). X-spermatozoa were enriched in cathodic fractions (80 and 68% Y-spermatozoa, as compared to control).

Animals↗
PubMed1991

Quantification of Y chromosome bearing spermatozoa of cattle using in situ hybridization.

An easy assay for quantification of Y chromosome-bearing sperm (Y-sperm) is needed, especially to monitor sperm separation techniques. In the present study a tritiated bovine male-specific DNA fragment was tested for identification of Y-sperm by in situ hybridization. A protocol for in situ hybridization to bovine sperm was developed and used to study the proportion of Y-sperm of 12 bulls. The usefulness of the method in optimization of sperm separation procedures is illustrated through analysis of fractions of sperm separated by Percoll density gradient centrifugation.

Animals↗
PubMed1990

Individual differences in capacitation of bull spermatozoa by heparin in vitro: relationship to fertility.

Several parameters of motility and integrity of frozen-thawed spermatozoa are compared with the ability of selected motile, intact spermatozoa to acrosome reaction induced by 0.005% hyamin. Between semen donors there exist distinct individual differences; however only the induced acrosome reaction after heparin treatment showed a significant correlation with fertility. For 12 bulls the nonreturn rates from 334 to 559 services correlated significantly with induced acrosome reaction (r=0.607). The in vitro fertilization of 53 to 93 tubal bovine oocytes with frozen-thawed spermatozoa from five bulls yielded a correlation of r=0.621 with the rate of induced acrosome reaction. The different capacity levels of heparin-treated spermatozoa to undergo acrosome reaction appears to correspond to the varying intensity and kinetics of heparin-mediated head-to-head aggregation of motile cells. The applied functional parameters could be used for the selection of bulls with low fertility in artificial insemination programs, and for spermatozoa donors for in vitro fertilization.

Journal Article↗
PubMed1990

[The effect of cumulus oophorus on the fertilization in vitro of mature oocytes of cattle].

The effect of the cumulus oophorus on fertilisation was investigated by 478 in vitro maturated bovine oocytes. Culturing revealed that the amount of defects on the pellucid zone and in ooplasm of cumulus-free oocytes was higher than that in oocytes cultured in the cumulus-oocyte complex (38 percent or 77/203 versus eleven percent or 30/275). Insemination of the cumulus-oocyte complex (COC) seemed to increase the fertilisation capability of bovine oocytes (63 percent or 70/111), though its penetration rate was only 45 percent (111/245) and was thus below that of cumulus-free oocytes (55 percent or 69/126). Use of COC is recommended as basic material for in vitro maturation and fertilisation.

Animals↗
PubMed1989

[The detection of in vitro capacitation of bull sperm by heparin treatment].

The capacitating effect of heparin upon spermatozoa from original and deep-frozen semen was characterised, using new methods for detection of inducible acrosomal reaction and heparin-mediated sperm aggregation, and was compared with frequently used capacitation by media of high ion strength. Heparin treatment was undertaken also by means of two culture media, "defined medium" (DM) and TCM 199, with 10% fetal calf serum. Higher motility was maintained by means of 10 I.U. of heparin/ml (= 77 micrograms/ml) which also proved helpful in achieving higher capability of inducible acrosomal reaction, as compared to pretreatment, using media of high ion strength. This applied to both fresh and deep-frozen sperm. The highest level of inducible acrosomal reaction was achieved after 2 hours of heparin action on fresh sperm and 30 minutes of action on deep-frozen sperm. That highest value was at its maximum, when TCM with 10% fetal calf serum had been used. This was the medium, after all, in which photometrically recorded aggregation of motile spermatozoa was at its fastest rate, reaching its maximum after about 60 minutes. The photometrically recorded activated motility of spermatozoa occurred more frequently in TCM, as compared to DM. Preparation of bull sperm in TCM 199 with fetal calf serum and heparin may be recommended as an effective and time-saving method for in vitro capacitation.

Animals↗
PubMed1989

[Characterizing the in vitro capacitation of motile bull sperm by the hyamine-induced acrosome reaction].

Described in this paper is a method to characterize capacitation of bull spermatozoa. Involved is the action of 0.005 per cent of hyamine 2389 on selected motile cells at 37 degrees C for 30 minutes to detect the number of spermatozoa capable of acrosomal reaction. The latter amount results from the percentual difference between acrosome-free spermatozoa prior to hyamine action and those after hyamine action. Acrosome-free spermatozoa can be easily identified by means of simple staining under optical light microscopy. The value thus obtained is related to the amount of spermatozoa in progressively motile in the unselected original suspension and is called "Percentage Acrosome Reaction Induction" (% ARI). The ARI capacitation-specific acts: Only small amounts of acrosome-free spermatozoa are recordable among incapacitated spermatozoa or from spermatozoa incubated in the presence of decapacitation factors (1.9 or 2.3 per cent, respectively). On the other hand, clearly increased values of 17.6 ot 26.1 per cent are obtainable from treatments with capacitation effect (high ionic strength or addition of heparin). The method is claimed to be suitable for adequate choice of conditions for improved capacitation and in vitro fertilization as well as for elucidation of specific individual differences in capacitation.

Acrosome↗
PubMed1989

[A simple method for the quantitative determination of sperm aggregation].

Described in this paper are 2 methodological variants for photometric recording of sperm aggregation. Heparin was used to induce aggregation. One of the working principles was related to aggregation-associated alteration of turbidity in an agitated cell suspension, measured by the KZM-1 coagulation-time meter, in analogy to measurement of thrombocyte aggregation. The 2nd variant was based on variation of cloudiness due to sedimentation of aggregates in a non-agitated suspension. Both methods provided equally valid information. They characterised both intensity of aggregation as a combined effect of the number and size of aggregates as well as the rate of aggregation. Photometric recording has proved to be an objectivated method for quantitative assessment of aggregation. Its use is proposed for studies into capacitation or immunological response of spermatozoa.

Animals↗
PubMedDate not supplied

Influence of environmental cadmium on testicular proliferation in roe deer.

The influence of cadmium on spermatogenesis in roe deer was studied, comparing animals from areas with high (n = 37) and low (n = 23) cadmium exposure. Testes were evaluated during pre-rut (May) and rut (July/August). During these periods the cadmium accumulation (mg/kg kidney) in polluted regions averaged 3.41 and 3.52 in comparison with 1.61 and 1.81 in controls. Cadmium accumulation was positively correlated with testis weight during both seasons. In May the proliferation (units of tissue polypeptid specific antigen/g parenchyma) was lower in animals with higher cadmium (24.02 compared with 78.20; P < 0.01). During the rut, testicular proliferation and spermatozoa/g testis as well as apoptosis showed no significant changes with increased cadmium contamination. The results suggest delayed proliferation during the pre-rutting period in animals with high cadmium exposure, but other indications of effects on the testis were not significant.

Animals↗
PubMedDate not supplied

Influence of chlorocholinechloride-treated wheat on selected in vitro fertility parameters in male mice.

The aim of this study was to evaluate the influence of feeding with food and water containing chlorocholinechloride (CCC) on the fertility of male mice in a two-generation study. For this purpose the number of testicular spermatozoa and the relative proportion of primary and secondary spermatocytes involved in spermatogenesis were measured. Furthermore, the fertility of epididymal spermatozoa from tested male mice was investigated in a special in-vitro fertilization system. The experimental food was composed of CCC-treated wheat in the first experiment and CCC-free wheat and water mixed with pure CCC in the second experiment. The CCC residue content in the treated food and water was 0.21 mg/kg and 0.2 mg/L, respectively. Under the influence of feeding with CCC-treated wheat (Experiment 1) the fertilization and cleavage rates of oocytes incubated with spermatozoa from CCC-fed mice were reduced: the fertilization rate 65.1% vs. 21.1% and the cleavage rate 51.9% vs. 20.3%, p < 0.01 (control feeding vs. CCC feeding, respectively). Feeding of sperm donors with pure CCC mixed with untreated wheat pellets or water (Experiment 2) led to a reduction in the fertilization and cleavage rate (control: 60.8%, 32.4%; CCC-food: 29.8%, 12.1%; CCC-water: 30.1%, 10.2%; CCC-food/water: 36.6%, 12.5%; p < 0.01, respectively). The normal course of spermatogenesis was unchanged after the exposure to CCC. Testicular weight, the number of spermatozoa, and the proportion of haploid, diploid, and tetraploid testicular cells were not influenced. However, the functional competence of epididymal spermatozoa from CCC-fed donors was reduced, resulting in a significantly diminished fertilization and cleavage rate in vitro. The results suggest that CCC could interfere with epididymal protein secretion and the process of sperm maturation during passage through the epididymis.

Administration, Oral↗
PubMedDate not supplied

Fluorescence body of bovine spermatozoa represents Y-chromosome.

The fluorescence body test would be an inexpensive method of quantifying the Y-chromosomes of mammalian spermatozoa, and could be used to monitor the enrichment of X- and Y-spermatozoa. However, a definitive proof that the fluorescence bodies are markers of the Y-chromosome, is available only for humans and gorillas. In the study described here it is demonstrated for the first time in cattle that the fluorescence bodies of spermatozoa represent the Y-chromosomes. The evidence is furnished by a combination of the fluorescence staining (quinacrine mustard) with the in-situ-hybridization using bovine male-specific DNA fragments. The incidence of f-bodies corresponded with autoradiographic grains caused by the tritiated Y-specific DNA in the examined 144 single cells (75, 5, and 88% agreement).

Animals↗
PubMedDate not supplied

Y-chromosome-specific fluorescence (f-body) of poorly decondensed bovine spermatozoa.

An account is given in this paper of a method of identifying the fluorescence body (f-body) as a marker of the Y-chromosome. Also covered by this method is poor decondensation of spermatozoal nuclei when exposed to action of 1.25% of papain, 0.155% of DTE, and 0.025% of DMSO. Quinacrine mustard was used as fluorescent stain, its final concentration being 0.0025%. For staining, spermatozoa were suspended for 1-5 h. Average f-body frequency accounted for 41.0 +/- 5.1% in 35 ejaculates from 22 bulls. The overall variation coefficient amounted to 12.4% and thus was higher than each of the single values individually recorded from six bulls which were involved with three or four ejaculates (3.6-7.0%). F-bodies could not be detected by the method generally used on human spermatozoa. The applicability of the f-body test to quantification of Y spermatozoa in experimental separation of androspermatozoa and gynospermatozoa is discussed and is demonstrated by an example.

Animals↗
PubMedDate not supplied

Chromatin condensation in cat spermatozoa during epididymal transit as studied by aniline blue and acridine orange staining.

Chromatin stability and DNA-resistance to acidic denaturation was evaluated by acidic aniline blue and acridine orange staining of cat sperm from different regions of the epididymis. The results were related to conventional sperm parameters. The percentage of aniline blue-stained spermatozoa (persisting histones) decreased significantly from the caput to the cauda region (31.8% and 7.8%, respectively; P<0.0001). The percentage of stained heads of cauda sperm was much lower in populations of morphologically normal forms than in those with abnormal forms (4.1% and 13.8%, respectively, P<0.0001). Among spermatozoa with abnormalities, the percentage of stained heads was significantly higher in cells with head abnormalities than in sperm with only tail abnormalities (87.1% and 10.3%, respectively; P>0.0001). With acridine orange fluorescence staining, 86.5% of cauda epididymal region (51.1%) to the cauda epididymal region (86.5%). The percentage of cauda epididymal sperm with normal condensed chromatin was neither linked to testicular sperm count, motility nor to age of the cats. The parameter of chromatin condensation and stability can be a valuable index of sperm quality, reflecting the possible disorders of spermatogenesis and epididymal sperm maturation, frequently observed in feline species.

Acridine Orange↗
PubMedDate not supplied

[Species specific motility patterns of hyperactivated mammalian spermatozoa and quantitative analysis of the hyperactivation of bull spermatozoa].

Movement tracks of spermatozoa of human, lion, tiger, cow, pig and sheep are recorded by dark field photography (fluid layer thickness 16.7 microns, exposure time 1 s). Comparison before and after capacitation by an incubation of 2 h in modified tissue culture medium TCM 199 with 10% fetal calf serum resulted in two quite different patterns of hyperactivated spermatozoa: 1) Tracks are broadened due to enlarged lateral head displacement or radius of rotating head movements respectively (tiger, lion) and show beside that a markedly increase in erratic motility (human). 2) In the studied species of artiodactyla, cow, pig and sheep, a qualitative new, panicle-like pattern arised as a result of superposition of spermatozoa head pendular movements around the axis of forward motility and the other one around the head axis. This new type of tracks allows a simple quantitative analysis of hyperactivation of bull spermatozoa, first described in this report. The method is applied to investigations on efficacy of capacitation media and provides evidence for high individual differences of semen donors in capacitation success.

Animals↗