| 1. |
- Bergqvist, Ann-Sofi, et al.
(författare)
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Single Layer Centrifugation of Stallion Spermatozoa through Androcoll (TM)-E does not Adversely Affect their Capacitation-Like Status, as Measured by CTC Staining
- 2011
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Ingår i: Reproduction in domestic animals. - : Blackwell Publishing. - 0936-6768 .- 1439-0531. ; 46:1, s. e74-e78
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Tidskriftsartikel (refereegranskat)abstract
- Contents This study was designed to evaluate the effect of single layer centrifugation (SLC) and subsequent cold storage on stallion sperm capacitation-like status and acrosome reaction. Three stallions were included in the study, with three ejaculates per stallion. The samples were examined 4, 24 and 72 h after collection, extension and SLC, with storage at 6 degrees C. Sperm capacitation-like status was investigated using the fluorescent dye chlortetracycline (CTC). There was no difference in capacitation-like status between colloid-selected and non-selected spermatozoa. Sperm motility decreased significantly during cold storage, whereas the proportion of apparently capacitated spermatozoa increased. There was no change in the proportion of acrosome-reacted spermatozoa. In conclusion, SLC through Androcoll (TM)-E does not adversely affect the capacitation-like status of stallion spermatozoa, although it did increase with time during cold storage.
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| 2. |
- Johannisson, Anders, et al.
(författare)
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Colloidal centrifugation with Androcoll-E (TM) prolongs stallion sperm motility, viability and chromatin integrity
- 2009
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Ingår i: Animal Reproduction Science. - : Elsevier Masson. - 0378-4320 .- 1873-2232. ; 116:1-2, s. 119-128
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Forskningsöversikt (refereegranskat)abstract
- The objective was to investigate the changes in stallion sperm quality (sperm motility, viability, membrane integrity and chromatin integrity) occurring during cool storage, and to study the effect of sperm selection by single layer colloidal centrifugation on these parameters of sperm quality. Spermatozoa from 3 stallions (10 ejaculates, 3-4 per stallion) were selected by centrifugation through a single layer of colloid (SLC). The resulting sperm preparations and the control samples (extended but unselected semen samples) were stored at 5 degrees C for 48 h. Assessments of sperm quality, such as sperm motility, viability (SYBR-14/PI staining), membrane stability (Annexin-V/PI staining) and chromatin integrity, were performed on aliquots of the selected sperm preparations and unselected samples on the day of collection (3 h) and after 24 and 48 h of storage. In the SLC-selected sperm samples, sperm motility, sperm viability, proportions of spermatozoa with normal morphology and with intact chromatin were significantly better than in unselected samples (motility: 77 +/- 4% vs. 64 +/- 8% at 3 h; P less than 0.001; viability: 79.5 +/- 9% vs. 64.7 +/- 9%, P less than 0.001: normal morphology 89 +/- 6% vs. 69 9%; chromatin integrity DFI 11.3 +/- 5% vs. 22.1 +/- 10%). Membrane stability, however, was not different in the SLC-selected and unselected samples (74.6 +/- 8% vs. 69.3 +/- 8%). The deterioration seen in sperm quality in the unselected samples was prevented by SLC, so that sperm viability, membrane stability and chromatin integrity were unchanged in the selected samples by 48 h compared to 3 h (Pless than0.001), whereas the unselected samples were significantly worse by 48 h (Pless than0.001). Furthermore, it should be possible to send an aliquot of a normal insemination dose (i.e. unselected spermatozoa) overnight to a reference laboratory for analysis of both plasma membrane and chromatin integrity. In conclusion, centrifugation of stallion spermatozoa through a single layer of colloid is a useful technique for selecting the best spermatozoa from an ejaculate and, moreover, sperm quality is maintained during storage. (C) 2009 Elsevier B.V. All rights reserved.
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| 3. |
- Kumaresan, A., et al.
(författare)
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Quantification of kinetic changes in protein tyrosine phosphorylation and cytosolic Ca2+ concentration in boar spermatozoa during cryopreservation
- 2012
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Ingår i: Reproduction, Fertility and Development. - 1031-3613 .- 1448-5990. ; 24:4, s. 531-542
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Tidskriftsartikel (refereegranskat)abstract
- Protein tyrosine phosphorylation in sperm is associated with capacitation in several mammalian species. Although tyrosine phosphorylated proteins have been demonstrated in cryopreserved sperm, indicating capacitation-like changes during cryopreservation, these changes have not yet been quantified objectively. We monitored tyrosine phosphorylation, intracellular calcium and sperm kinematics throughout the cryopreservation process, and studied the relationships among them in boar spermatozoa. Sperm kinetics changed significantly during cryopreservation: curvilinear velocity, average path velocity and straight line velocity all decreased significantly (P < 0.05). While the percentage of sperm with high intracellular calcium declined (P < 0.05), global phosphorylation increased significantly (P < 0.01). Specifically, cooling to 5 degrees C induced phosphorylation in the spermatozoa. After cooling, a 32-kDa protein not observed in fresh semen appeared and was consistently present throughout the cryopreservation process. While the level of expression of this phosphoprotein decreased after addition of the second extender, frozen-thawed spermatozoa showed an increased expression. The proportion of sperm cells with phosphorylation in the acrosomal area also increased significantly (P < 0.05) during cryopreservation, indicating that phosphorylation might be associated with capacitation-like changes. These results provide the first quantitative evidence of dynamic changes in the subpopulation of boar spermatozoa undergoing tyrosine phosphorylation during cryopreservation.
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| 4. |
- Morrell, Jane, et al.
(författare)
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Colloidal Centrifugation of Stallion Semen: Changes in Sperm Motility, Velocity, and Chromatin Integrity during Storage
- 2009
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Ingår i: Journal of Equine Veterinary Science. - : WB Saunders. - 0737-0806 .- 1542-7412. ; 29:1, s. 24-32
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Tidskriftsartikel (refereegranskat)abstract
- The current study investigated the changes in sperm quality (motility, velocity, and chromatin integrity) occurring during storage at room temperature or 5 degrees C for up to 48 hours in spermatozoa after extension or single-layer centrifugation (SLC) throught Androcoll-E. In unselected samples, all parameters of sperm quality deteriorated significantly during storage (P less than .01), although the deterioration was faster at room temperature (22-30 degrees C) than for cool storage (P less than .01). The SLC-selected spermatozoa had higher motility, velocity, and chromatin integrity than the overall unselected population (motility: selected 85 +/- 10%, unselected 56 +/- 13%; P less than .001; velocity: selected 85.1 +/- 13 mu m/second, unselected 63.5 +/- 15 mu m/second; P less than .001; and DFI selected 12.2 +/- 4.8 mu m/second; unselected 23.6 +/- 7.4 mu m/second; P less than .001). Furthermore, sperm quality did not deteriorate with storage in the SLC-selected samples, either at room temperature (22-30 degrees C for 24 hours) or cooled to 4 degrees C (for at least 48 hours), whereas a significant deterioration in sperm quality was observed in the unselected sperm samples (P less than .01). Thus, room temperature storage of SLC-selected spermatozoa may be an option for insemination doses from stallions whose spermatozoa do not tolerate cooling. In addition, a new sperm analyzer, the Qualisperm, showed good correlation with subjective motility assessment (r = 0.8, P less than .001), was user-friendly, and provided a reasonable volume of data. This instrument may be a useful adjunct to sperm quality assessment at the study.
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| 5. |
- Morrell, Jane, et al.
(författare)
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Effect of osmolarity and density of colloid formulations on the outcome of SLC-selection of stallion spermatozoa
- 2011
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Ingår i: ISRN Veterinary Science. - : Hindawi Limited. - 2090-4452 .- 2090-4460. ; 2011, s. 1-5
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Tidskriftsartikel (refereegranskat)abstract
- The osmolarity and density of colloids used to prepare spermatozoa for assisted reproduction may affect sperm quality in the resultant preparation. In this study, two osmolarities of Androcoll-E for single-layer or density gradient centrifugation of stallion spermatozoa were compared: “normal” (320 mOsm) or “high” (345 mOsm). Mean yields for the two centrifugation techniques did not differ between treatments or osmolarities (single layer centrifugation:30.19±16.9×106and25.8±18.5×106spermatozoa; density gradient centrifugation:31.84±19.7×106and26.46±20.0×106spermatozoa respectively for the two osmolarities). However, use of the high osmolarity colloid for single layer centrifugation increased the proportion of morphologically normal spermatozoa (P<.05). Therefore, increasing the osmolarity of the colloid formulation may be beneficial for processing ejaculates containing a high proportion of abnormal spermatozoa by SLC. Reducing the density of the colloid used for the SLC substantially increased the yield of motile spermatozoa compared to the normal density colloid (mean ± SD:72.6±28.9×106versus28.9±24.7×106), while also prolonging sperm survival by 24 hours compared to the uncentrifuged ejaculate. This increased yield may render Single Layer Centrifugation practical for use in the field.
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| 6. |
- Morrell, Jane, et al.
(författare)
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Morphology and chromatin integrity of stallion spermatozoa prepared by density gradient and single layer centrifugation through silica colloids
- 2009
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Ingår i: Reproduction in Domestic Animals. - : Wiley. - 0936-6768 .- 1439-0531. ; 44, s. 512-517
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Tidskriftsartikel (refereegranskat)abstract
- The objective was to investigate whether it is possible to improve the quality of stallion semen, with respect to sperm morphology and chromatin integrity, both of which have been linked to fertility, using either density gradient centrifugation (DGC) or a new method, hereby named single layer centrifugation (SLC). The two methods of colloidal centrifugation were evaluated using 38 ejaculates from 10 stallions. Sperm morphology, subjective motility and sperm chromatin integrity were compared in uncentrifuged samples and in centrifuged sperm preparations. The proportion of morphologically normal spermatozoa varied between stallions (p < 0.001) and was increased by both methods of colloidal centrifugation (median value before centrifugation 67.5%; after SLC 78%; after DGC 77%; p < 0.001). The incidence of certain abnormalities was reduced, e.g. proximal cytoplasmic droplets were reduced from 12.9% to 8.8% (p < 0.001), and mid-piece defects from 5.3% to 1.4% (p < 0.05). Similarly, sperm motility and chromatin integrity were significantly improved (p < 0.001), with no difference between the two centrifugation methods. Centrifugation through colloids can enrich the proportions of stallion spermatozoa with normal morphology and normal chromatin structure in sperm preparations. The new method, SLC, was as effective as DGC in selecting motile stallion spermatozoa with normal morphology and intact chromatin. SLC, being simpler to use than DGC, would be appropriate for routine use by stud personnel to improve stallion sperm quality in insemination doses.
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| 7. |
- Morrell, Jane, et al.
(författare)
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Single layer centrifugation of stallion spermatozoa improves sperm quality compared with sperm washing
- 2010
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Ingår i: Reproductive BioMedicine Online. - : Elsevier. - 1472-6483 .- 1472-6491. ; 21:3, s. 429-436
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Tidskriftsartikel (refereegranskat)abstract
- This study compared the effect on semen quality of different handling methods used in the preparation of stallion semen doses for artificial insemination. The three methods were (i) extending the ejaculate to 30-50 x 10(6)/ml, (ii) single layer centrifugation (SLC) and (iii) sperm washing (centrifugation without a colloid). An additional treatment was to add seminal plasma (SP) in various proportions to some SLC preparations. The resulting samples were evaluated for sperm motility by computer assisted sperm analysis, membrane integrity using the Nucleocounter SP-100 and chromatin integrity by the sperm chromatin structure assay. SLC samples consistently had better sperm quality than the extended samples. Sperm washing did not confer any beneficial effect compared with the extended samples and these samples had significantly worse sperm quality than the SLC samples (motility, P less than 0.01; viability, P less than 0.001). There was no evidence to suggest that adding SP to the SLC samples could enhance sperm motility for more than a few hours. Longer term cold storage of spermatozoa in the presence of small concentrations of SP resulted in a reduction in total motility and progressive motility compared with SLC alone. High concentrations of SP were detrimental to sperm survival. (C) 2010, Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.
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| 8. |
- Morrell, Jane, et al.
(författare)
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Single-layer centrifugation with Androcoll-E can be scaled up to allow large volumes of stallion ejaculate to be processed easily
- 2009
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Ingår i: Theriogenology. - : Elsevier. - 0093-691X .- 1879-3231. ; 72:6, s. 879-884
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Tidskriftsartikel (refereegranskat)abstract
- The objective of the current study was to optimize the volumes of Androcoll-E and sperm sample used in various sizes of centrifuge tube to scale up single-layer centrifugation (SLC) for routine use in the field. Although sperm suspensions of equivalent quality were produced using Androcoll-E in small and large tubes, the sperm yield was much lower in the latter (P less than 0.001). In contrast, in 200-mL tubes (XL), the yields were approximately 25% higher than those for the small tubes. An increased volume (4.5 mL) of extended ejaculate in small tubes (SLC-Inc) or 15 to 18 mL extended ejaculate on 15 mL of colloid of a reduced density, Androcoll-E-Large (SLC-Large), in 50-mL tubes were both found to give similar yields of motile spermatozoa as that of the SLC-Small method (SLC-Small, 49.7 +/- 18.6%; SLC-Inc, 53.3 +/- 17.1%; SLC-Large, 44.9 +/- 18.3%) and were found to be equivalent in quality (motility: 88.0 +/- 8.8%, 84.0 +/- 3.5%, 90.0 +/- 5.4%; normal morphology: 69.4 +/- 17.0%, 69.4 +/- 12.7%, 63.9 +/- 15.6%; viability: 78 +/- 16.7%, 83.8 +/- 12.5%, 80.05 +/- 14.6%; DNA fragmentation index: 14.7 +/- 10.9%, 12.8 +/- 8.1%, 11.6 +/- 7.6%, respectively). The processing of an "average" stallion Equus caballus ejaculate in approximately twenty-seven 10-mL tubes (SLC-Inc) or eight 50-mL tubes (SLC-Large) is feasible, the latter being considered more practical for on-stud use. (C) 2009 Elsevier Inc. All fights reserved.
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| 9. |
- Ortega-Ferrusola, Cristina, et al.
(författare)
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Effect of Different Extenders and Seminal Plasma on the Susceptibility of Equine Spermatozoa to Lipid Peroxidation After Single-Layer Centrifugation, Through Androcoll-E
- 2011
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Ingår i: Journal of Equine Veterinary Science. - : Elsevier Science B. V., Amsterdam. - 0737-0806 .- 1542-7412. ; 31:7, s. 411-416
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Tidskriftsartikel (refereegranskat)abstract
- his study was conducted in an attempt to see whether single-layer centrifugation (SLC) increases the susceptibility of stallion spermatozoa to lipid peroxidation (LPO), in different extenders after removing all seminal plasma (SP). The susceptibility of stallion spermatozoa to LPO was studied before and after SLC. Each ejaculate was split, and aliquots extended with one of the three different extenders: INRA 96, Kenneys, or Equipro, and stored for 24 hours at 5 degrees C (i). From the extended samples, an aliquot was kept as a control and the other was subjected to SLC through Androcoll-E. The selected spermatozoa were re-suspended in the appropriate extenders, without (ii) or with (iii) addition of 50% (v/v) pooled homologous SP for 24 hours at 5 degrees C. Using ferrous sulfate as pro-oxidant, the susceptibility for LPO was flow-cytometrically assessed using the probe Bodipy(581/591)-C(11). Sperm motility, monitored with a Qualisperm motility analyzer, increased after SLC treatment (P andlt; .001). No significant correlations were found between motility and induced LPO with ferrous sulfate. The SP and extenders, per se, did not have a significant protective effect against LPO, but the interaction between SP and Kenney increased the susceptibility to LPO. However, the selected spermatozoa through Androcoll-E and the subsequent dilution in INRA had a significant protective effect against LPO (P andlt; .05), especially when the oxidative insults were higher (80 mu M).
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| 10. |
- Revay, T., et al.
(författare)
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Macrocephaly in Bull Spermatozoa Is Associated with Nuclear Vacuoles, Diploidy and Alteration of Chromatin Condensation
- 2009
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Ingår i: Cytogenetic and Genome Research. - : S. Karger. - 1424-8581 .- 1424-859X. ; 126:1-2, s. 202-209
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Tidskriftsartikel (refereegranskat)abstract
- Spermatozoa from 2 dairy AI (artificial insemination) bulls (A and B), identified by their abnormal spermiogram with cells depicting frequent macrocephaly, double tails and nuclear vacuoles, were case-investigated and compared to normal spermatozoa from a control AI sire (C). Head sizes were measured and morphological abnormalities scored using brightfield and differential interference contrast microscopy. The degree of sperm maturation and of resistance to acid-induced DNA denaturation in situ were determined after uploading of acridine orange using flow cytometry of 5,000 cells/sample. Nuclear fragmentation, i.e. the ratio of red to total (red + green) fluorescence, reached 7.1% and 31% in bulls A and B, compared to 2% in bull C. The proportion of immature spermatozoa, i.e. those with incomplete histone-protamine exchange and depicting higher green fluorescence compared to the main population of the control bull, reached 9.54% in A and 7.75% in B, compared to only 0.47% in the control. In the second part of this study the previously unknown chromosomal constitution of large-headed spermatozoa of bull A was investigated by fluorescence in situ hybridization using an X-Y painting probe set. The 7.5% XY-bearing cells and the presence of diploid spermatozoa detected by flow cytometry indicate a meiotic arrest in the first division in bull A, becoming the first proven case of association of macrocephaly and M1 diploidy. The diverse approaches used for the investigation of spermatozoal DNA provide insights into the etiology of macrocephaly. Copyright (C) 2009 S. Karger AG, Basel
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| 11. |
- Saravia-Ramos, Fernando, et al.
(författare)
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Exposure to the seminal plasma of different portions of the boar ejaculate modulates the survival of spermatozoa cryopreserved in MiniFlatPacks
- 2009
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Ingår i: Theriogenology. - : Elsevier. - 0093-691X .- 1879-3231. ; 71:4, s. 662-675
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Tidskriftsartikel (refereegranskat)abstract
- Spermatozoa present in the first collectable 10 mL of the sperm-rich fraction (SRF) of the boar ejaculate (portion 1, PI I have higher documented viability during and after cryopreservation than spermatozoa in the rest of the ejaculate (portion 2, P2). probably in relation to different features of the surrounding seminal plasma (SP). In the present study. We investigated whether the SP from these ejaculate portions (SP1 or SP2) was able to differently influence sperm viability and chromatin structure of the P1- or P2-contained spermatozoa from individual boars primarily or secondarily exposed (e.g., following cleansing and re-exposure) to pooled SP1 or SP2 from the same males during 60 min. Spermatozoa were subjected to controlled cooling and thawing in MiniFlatPacks (MFPs) and examined for motility (using computer-assisted sperm analysis, CASA) at selected stages of processing Moreover, sperm plasma membrane intactness (investigated using, SYBR-14/propidium iodide, PI), plasma membrane architecture (examined using Annexin-V-PI staining), and chromatin (deoxyribonucleic acid, DNA) integrity (tested using sperm chromatin structure assay, SCSA) were assessed post-thaw (PT). A higher proportion of PI spermatozoa than of P2 spermatozoa incubated in their native SP portion were confirmed to be motile from collection to PT. When P1 spermatozoa were cleansed from their original SP and re-exposed to pooled P2-SP. sperm kinematics deteriorated from extension to PT. By contrast, cleansed P2 spermatozoa increased motility to P1 levels, especially PT when re-exposed to pooled P1-SP. Such differential effects on motility were not clearly accompanied by biologically related modifications of sperm membrane or chromatin structure. This influence of the SP on sperm kinematics was not sire-dependent and it Was presumably related to different concentrations or either SP proteins or bicarbonate in the different ejaculate portions. (C) 2009 Elsevier Inc. All rights reserved.
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| 12. |
- Sharoare Hossain, Md, et al.
(författare)
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Flow cytometry for the assessment of animal sperm integrity and functionality: state of the art
- 2011
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Ingår i: ASIAN JOURNAL OF ANDROLOGY. - : Blackwell Publishing Ltd. - 1008-682X .- 1745-7262. ; 13:3, s. 406-419
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Forskningsöversikt (refereegranskat)abstract
- Flow cytometry is now a recognized methodology within animal spermatology, and has moved from being a research tool to become routine in the assessment of animal semen destined to breeding. The availability of bench-top flow cytometers and of newer and versatile markers for cell structure and function had allowed the instrumentation to measure more sperm parameters, from viability to reactiveness when exposed to exogenous stimuli, and to increase our capabilities to sort spermatozoa for potential fertilizing capacity, or chromosomal sex. The present review summarizes the state of the art regarding flow cytometry applied to animal andrology, albeit keeping an open comparative intent. It critically evaluates the present and future capabilities of flow cytometry for the diagnostics of potential fertility and for the development of current reproductive technologies such as sperm freezing, sperm selection and sperm sorting. The flow cytometry methods will probably further revolutionize our understanding of the sperm physiology and their functionality, and will undoubtedly extend its application in isolating many uncharacterized features of spermatozoa. However, continuous follow-up of the methods is a necessity owing to technical developments and the complexity of mapping spermatozoa.
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| 13. |
- Siqueira, A P, et al.
(författare)
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Quality of boar spermatozoa from the sperm-peak portion of the ejaculate after simplified freezing in MiniFlatpacks compared to the remaining spermatozoa of the sperm-rich fraction
- 2011
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Ingår i: THERIOGENOLOGY. - : Elsevier Science B.V., Amsterdam.. - 0093-691X .- 1879-3231. ; 75:7, s. 1175-1184
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Tidskriftsartikel (refereegranskat)abstract
- Boar sperm viability post-thaw differs depending on the ejaculate fraction used, with spermatozoa present in the first 10 mL of the sperm-rich fraction (SRF) (portion 1, P1, sperm-peak portion) displaying the best cryosurvival in vitro compared with that of spermatozoa from the rest of the ejaculate (portion 2 of the SRF plus the post-spermatic fraction), even when using simplified freezing routines. This viability apparently relates to the specific profile of seminal plasma in P1 (i.e., glycoprotein and bicarbonate concentrations, and pH). However, spermatozoa from PI have not been compared with spermatozoa from the rest of the SRF (SRF P1, usually 30-40 mL of the SRF), which is routinely used for freezing. We compared P1 with SRF P I in terms of sperm kinematics (using the QualiSperm (TM) system), while membrane integrity (SYBR-14/PI), acrosome integrity (FITC PNA/PI), and sperm membrane stability (Annexin-V) were explored using flow cytomety. As well, total protein concentration and the proteomics of the seminal plasma (SP) of both portions of the SRF were studied using two-dimensional electrophoresis (2DE), mass fingerprinting (MALDI-TOF), and collision-induced dissociation tandem mass spectrometry (CID-MS/MS) on selected peptides. The SRF portions were collected weekly from four mature boars (4-5 replicates per boar, sperm concentration: P1, 1.86 +/- 0.20; SRF P1, 1.25 +/- 0.14 x 10(9) spz/mL) and processed using a quick freezing method in MiniFlatPacks. Post-thaw sperm motility reached 50%, without differences between SRF portions, but with clear inter-boar variation. Neither plasma membrane nor acrosome integrity differed (ns) between fractions. These results indicate that there are no differences in cryosurvival after quick freezing of boar spermatozoa derived from either of the two SRF portions. While P1 and SRF-P1 clearly differed in relative total protein contents, as expected, they displayed very similar protein profiles as assessed using 2DE and mass spectrometry (tryptic peptide mass fingerprint analysis and CID-MS/MS), indicating a similar emission of epididymal protein content.
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| 14. |
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| 15. |
- Nagy, Szabolcs, et al.
(författare)
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Evaluation of domestic animal sperm head morphology via flow cytometric DNA labelling and pulse shape analysis using bull and stallion spermatozoa as model species
- 2023
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Ingår i: Reproduction in domestic animals. - : John Wiley & Sons. - 0936-6768 .- 1439-0531. ; 58:11, s. 1569-1575
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Tidskriftsartikel (refereegranskat)abstract
- The aim of the present study was to test a rapid, robust flow cytometric technique for the detection of sperm head abnormalities of domestic bulls and stallions. The so-called PulSA approach detects the pulse profiles of propidium-iodide labelled spermatozoa. In the first experiment, species-specific threshold values were established on sperm samples that were tested for sperm head abnormalities with a classic visual morphology analysis. In the second experiment, serial mixtures of bull and stallion spermatozoa mimicking different percentages of sperm head abnormalities were analysed. Non-metric multidimensional scaling showed a clear separation between the normal and mixed samples. The PulSA approach may be a useful tool in identifying sub- or infertile breeding males as well as in studying the evolutionary aspects of sperm morphology and morphometry.
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| 16. |
- Jansson, Anna, et al.
(författare)
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Plasma aldosterone concentration and cardiovascular response to low sodium intake in horses in training
- 2010
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Ingår i: Equine Veterinary Journal. - : Wiley. - 0425-1644 .- 2042-3306. ; 42, s. 329-334
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Tidskriftsartikel (refereegranskat)abstract
- P>Reason for performing the study:Horses in training lose large amounts of sodium but little is known about the cardiovascular response to low sodium intake.Objectives:To investigate the effect of low sodium intake on plasma aldosterone (pAldo) concentrations and the cardiovascular system of athletic horses, and to identify markers of low sodium intake.Methods:Seven Standardbred geldings in training (trained twice a week) were randomly offered a standardised diet supplemented (NaS, 58 mg Na/kg bwt) and not supplemented (NaN, 3 mg Na/kg bwt) with NaCl for 5 weeks in a changeover design. Blood samples were taken once a week and in Week 5, before and following an exercise test until 22.30 h and analysed for blood sodium (bNa), total plasma protein (TPP), pAldo, troponin I and packed cell volume (PCV). Blood pressure (BP) was measured and pulse wave recorded at rest with high definition oscillometric-technique (HDO). ECG and echocardiography were recorded. Water intake was measured before and on the day of exercise and voluntary saline intake was measured for 2 days after each period. Faecal samples were taken weekly and analysed for sodium and potassium content.Results:The pAldo and the PCV was higher in NaN compared to NaS. There were no differences between diets in BP, ECG, plasma troponin I and echocardiogram but HDO pulse amplitude tended to be smaller on diet NaN. Water intake was lower on diet NaN and saline intake higher. The response to exercise in bNa, pAldo, PCV and TPP was different on the 2 diets. Faecal potassium/sodium ratio was higher on NaN than on NaS.Conclusion:This study shows that 5 weeks of low sodium intake increased plasma aldosterone concentration and PCV but no alterations in heart function was observed. Faecal potassium/sodium ratio could be used to assess sodium status in horses.
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| 17. |
- Johannisson, Anders
(författare)
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Bronchial microdialysis of cytokines in the epithelial lining fluid in experimental intestinal ischemia and reperfusion before onset of manifest lung injury
- 2010
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Ingår i: Shock. - 1073-2322 .- 1540-0514. ; 34, s. 517-524
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Tidskriftsartikel (refereegranskat)abstract
- Today, there is no continuous monitoring of the bronchial epithelial lining fluid. This study used microdialysis as a method of continuous monitoring of early lung cytokine response secondary to intestinal ischemia-reperfusion in pigs. The authors aimed to examine bronchial microdialysis for continuous monitoring of IL-1 beta, TNF-alpha, IL-8, and fluorescein isothiocyanate Dextran 4,000 Da (FD-4). The superior mesenteric artery was cross-clamped for 120 min followed by 240 min of reperfusion (ischemia group, n = 8). Four sham-operated pigs served as controls. The pigs were anesthetized and normoventilated (peak inspiratory pressure, <20 cm H(2)O; positive end-expiratory pressure, 7 cm H(2)O). Samples from bronchial and luminal intestinal and arterial microdialysis catheters (flow-rate of 1 mu L/min) were collected during reperfusion in 60-min fractions. Samples were analyzed for TNF-alpha, IL-1 beta, IL-8, and FD-4. Data are presented as median (interquartile range). A lung biopsy was collected at the end of the experiment. During reperfusion, there was an increase in bronchial concentrations of both IL-8 (3.70 [1.47-8.93] ng/mL per h vs. controls, 0.61 [0.47-0.91] ng/mL per h; P < 0.001) and IL-1 beta (0.32 [0.05-0.56] ng/mL per h vs. controls, 0.07 [0.04-0.10] ng/mL per h; P = 0.008). In the intestinal lumen, IL-8 was increased in the ischemia group (6.33 [3.13-9.23] ng/mL per h vs. controls, 0.89 [0.21-1.86] ng/mL per h; P < 0.001). The FD-4 did not differ between groups. Pulmonary vascular resistance and pulmonary shunt increased versus controls. During reperfusion, PaO(2)/FiO(2) ratio decreased in the ischemia group. Histology was normal in both groups. Bronchial microdialysis detects altered levels of cytokines in the epithelial lining fluid and can be used for continuous monitoring of the immediate local lung cytokine response secondary to intestinal ischemia-reperfusion.
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| 18. |
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| 19. |
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| 20. |
- Johannisson, Anders, et al.
(författare)
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Single Layer Centrifugation with Androcoll-P can be scaled-up to process larger volumes of boar semen
- 2011
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Ingår i: ISRN Veterinary Science. - : Hindawi Limited. - 2090-4452 .- 2090-4460. ; 2011
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Tidskriftsartikel (refereegranskat)abstract
- The objective of this study was to scale-up the procedure for Single Layer Centrifugation (SLC) through AndrocollTM-P, as a preliminary step towords processing the whole ejaculate. The first experiment compared Single Layer Centrifugation using 4.5 mL and 15 mL extended ejaculate (SLC-4.5 and SLC-15, resp.), assessing sperm quality by objective motility analysis, morphology, viability, and the production of reactive oxygen species (ROS). In the second experiment, SLC-4.5 was compared to Single Layer Centrifugation with 25 mL extended ejaculate (SLC-25) using motility analysis and morphology. In both experiments, normal morphology and linear motility were significantly higher in the SLC-selected samples than in the uncentrifuged controls (P<.001), whereas total motility and membrane integrity were unchanged. Although ROS production was higher in the SLC-selected samples than in the controls (P<.01), this might have been due to the presence of antioxidants in seminal plasma in the latter. In conclusion, there was no difference in sperm quality between SLC-4.5 and SLC-15 samples, or between SLC-4.5 and SLC-25 samples, indicating that the SLC method can be scaled-up successfully.
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| 21. |
- Johannisson, Anders, et al.
(författare)
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Spermatozoa in the sperm-peak-fraction of the boar ejaculate show a lower flow of Ca2+ under capacitation conditions post-thaw which might account for their higher membrane stability after cryopreservation
- 2011
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Ingår i: Animal Reproduction Science. - : Elsevier BV. - 0378-4320 .- 1873-2232. ; 128, s. 37-44
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Tidskriftsartikel (refereegranskat)abstract
- Boar spermatozoa collected in the ejaculate sperm peak-portion (P1, first 10 mL of the sperm-rich fraction, SRF), had shown a higher resilience to freezing and thawing compared to spermatozoa from the rest of the ejaculate (2nd portion of the SRF plus the post-sperm-rich fraction, PSRF), even when using a simplified freezing technique, as long as spermatozoa were incubated in their own seminal plasma (SP). This experiment studied the stability of P1- and SRF-P1 boar spermatozoa frozen in MiniFlatPacks (MFP), post-thaw, using flow cytometry. Since spermatozoa from either portion showed similar cryosurvival and low proportions of unstable membranes (<3%, annexin-V/propidium iodide staining), and only a tendency for SRF-P1 live spermatozoa to depict acrosome exocytosis (FITC-PNA/PI/H33342); they were explored for Ca2+ contents using a Fluo-4 probe under in vitro capacitating conditions (mBO+ medium), as well they were tested for their ability to sustain a short Ca2+-ionophore (A23187) in vitro challenge. The proportions of live spermatozoa depicting high Ca2+-levels were initially <2% but increased over incubation time, particularly in SRF-P1(P < 0.05), while proportions of live spermatozoa with low Ca2+-levels were basically constant over incubation time (similar to 11-14%), for either portion. Incubation in capacitation medium did not modify the proportions of low-Ca2+ but dramatically increased the proportions of high-Ca2+ spermatozoa (P < 0.001) already after 15 min exposure, highest for SRF-P1 spermatozoa. While the proportion of live spermatozoa with intact acrosome was significantly decreased among SRF-P1 (P < 0.001), that of P1-spermatozoa remained unchanged, probably owing to the lowest relative content of cytosolic Ca2+. The results suggest that spermatozoa in the P1-portion are more resilient to express acrosome exocytosis post-thaw compared to those bathing in the rest of the SRF-fraction when cryopreserved using a simplified technique, in MFPs. (C) 2011 Elsevier B.V. All rights reserved.
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| 22. |
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| 23. |
- Johannisson, Anders, et al.
(författare)
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The Association of the Presence of Seminal Plasma and Its Components with Sperm Longevity in Fractionated Stallion Ejaculates
- 2011
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Ingår i: Reproduction in Domestic Animals. - : Wiley. - 0936-6768 .- 1439-0531. ; 46, s. 1073-1081
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Tidskriftsartikel (refereegranskat)abstract
- Semen jets in the stallion's ejaculate differ in both the biochemical composition of seminal plasma (SP) and in sperm concentration, forming sperm-rich fractions (HIGH) and sperm-poor fractions (LOW). This study examined (i) the association of the levels of selected components of SP [alkaline phosphatase (AP), acid phosphatase (ACP), Na(+), Cl(-), K(+), Ca, Mg and prostaglandin E(2)] with semen quality, sperm longevity and fertility; (ii) the effects of the presence of SP on sperm survival during cooled storage; (iii) the differences in the composition of SP between sperm-rich and sperm-poor ejaculatory fractions; (iv) the differences between these fractions in the effects of SP on sperm longevity. The levels of AP, ACP and Na(+) were higher in HIGH compared with LOW, whereas higher concentrations of Ca, K(+) and Mg were found in LOW than in HIGH. None of the measured components were correlated to the first cycle pregnancy rate. The presence of SP during cooled storage caused DNA degradation, but total and progressive sperm motility was not affected. Sperm-rich fractions stored with low levels of SP maintained higher level of DNA integrity compared with sperm-rich fractions in higher levels of SP, or sperm-poor fractions irrespective of the proportion of SP. The concentration of K(+) in LOW may give some indication on the maintenance of total sperm motility during cooled storage.
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| 24. |
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| 25. |
- Kumaresan, Arumugam, et al.
(författare)
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The effect of oviductal fluid on protein tyrosine phosphorylation in cryopreserved boar spermatozoa differs with the freezing method
- 2012
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Ingår i: Theriogenology. - : Elsevier BV. - 0093-691X .- 1879-3231. ; 77, s. 588-599
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Tidskriftsartikel (refereegranskat)abstract
- Sperm capacitation takes place in the oviduct and protein tyrosine phosphorylation of sperm proteins is a crucial step in capacitation and acquisition of fertilizing potential. Cryopreserved spermatozoa show altered expression of protein tyrosine phosphorylation in the oviduct. The present study compared two freezing methods (conventional-conventional freezing (CF) and simplified-simplified freezing (SF) methods) for their effect on the ability of boar spermatozoa to undergo protein tyrosine phosphorylation in response to oviductal fluid (ODF). Cryopreserveel boar-spermatozoa were incubated with pre- and post-ovulatory ODF for 6 h at 38 degrees C under 5% CO(2). Aliquots of sperm samples were taken at hourly intervals and analyzed for kinematics and protein tyrosine phosphorylation. Global protein tyrosine phosphorylation in spermatozoa was measured using flow cytometry and different patterns of phosphorylation were assessed using confocal microscopy. Immediately after thawing, no significant difference was observed in post-thaw sperm motility, velocity and global tyrosine phosphorylation between the two methods of freezing although the freezing method significantly (P < 0.05) influenced the effect of oviductal fluid on these parameters during incubation. While spermatozoa frozen by the CF method showed a significantly higher (P < 0.001) proportion of phosphorylation in response to preovulatory ODF during incubation, spermatozoa frozen by the SF method did not elicit such significant response as there was no significant difference in the proportion of tyrosine phosphorylated spermatozoa between treatments at any given time during incubation. If the CF method was used, the proportion of spermatozoa displaying either tail or full sperm phosphorylation increased in response to both preovulatory (EODF) and postovulatory oviductal fluid. However, if the SF method was used, a significant increase in these patterns was noticed only in the EODF treated group. The present study demonstrates that preovulatory isthmic ODF induce tyrosine phosphorylation in a higher proportion of boar spermatozoa compared to the post-ovulatory fluid and that the method of freezing significantly influences the response of post-thaw spermatozoa to porcine ODF. (C) 2012 Elsevier Inc. All rights reserved.
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| 26. |
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| 27. |
- Madej, Malgorzata, et al.
(författare)
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The Effect of Boar Seminal Plasma on the Release of Prostaglandins and Interleukin-6 by Porcine Endometrial and Cervical Cells and Bovine Endometrial Cells
- 2012
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Ingår i: Reproduction in Domestic Animals. - : Wiley. - 0936-6768 .- 1439-0531. ; 47, s. 113-124
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Tidskriftsartikel (refereegranskat)abstract
- Artificial insemination (AI) of sows results in a significant elevation of prostaglandin F2a metabolite (PGFM) levels in peripheral plasma, whereas in mated sows such elevation is not seen. The aim of this study was to investigate whether boar seminal plasma (SP) has any effect on the release of PGFM, prostaglandin F2a (PGF2a), prostaglandin E2 (PGE2) or interleukin-6 (IL-6) by in vitro cultured porcine endometrial (epithelial pUE and stromal pUS), cervical (pCE and pCS) and bovine endometrial epithelial cells (bUE). This study shows that boar SP inhibits the release of PGFM, PGF2a and PGE2 by porcine endometrial and cervical cells and bovine endometrial cells after 3 and 24 h incubation. Boar SP stimulated IL-6 release by pUE, pUS and even bUE after 3 h incubation. Tumour necrosis factor a (TNFa) stimulated the release of IL-6 by pUS only after 24 h incubation, but in the presence of boar SP, this stimulation was attenuated. The overall results from these in vitro studies give us possibility to understand the difference in prostaglandin response between mated and inseminated sows. Furthermore, we demonstrated that frozen-stored epithelial and stromal cells from pig endometrium, as well as from the cervix are suitable for studying the effect of SP on the release of prostaglandins. The only prerequisite is to incubate these thawed cells with arachidonic acid as a source for the synthesis of prostaglandins. A similar effect of boar SP on porcine and bUE cells may suggest inter-species reactivity.
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| 28. |
- Morrell, Jane, et al.
(författare)
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Processing stored stallion semen doses by Single Layer Centrifugation
- 2011
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Ingår i: Theriogenology. - : Elsevier BV. - 0093-691X .- 1879-3231. ; 76, s. 1424-1432
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Tidskriftsartikel (refereegranskat)abstract
- The purpose of this study was to determine if the quality of stored stallion semen doses could be enhanced by the scaled-up version of Single Layer Centrifugation using Androcoll-E-Large. Three semen doses from each of fifteen stallions were transported overnight to the Swedish University of Agricultural Sciences (SLU) for processing 24 h after semen collection. Sperm quality in the resulting SLC-selected samples was significantly improved compared to the uncentrifuged samples: mean progressive motility was increased by 8% on the day of processing (P < 0.001) and by 13% after 24 h cold storage (P < 0.001), normal morphology was increased by 4% (P < 0.01), whereas mean %DFT was decreased by 2% (P < 0.001). When these SLC-selected samples were compared retrospectively to fresh samples processed by SLC with Androcoll-E Small, sperm quality was found to be similar, although it was not maintained for as long in the sperm samples stored before SLC. These results suggest an additional option for improving sperm quality in stallion semen doses for artificial insemination. (C) 2011 Elsevier Inc. All rights reserved.
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| 29. |
- Morrell, Jane, et al.
(författare)
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Restoration of seminal plasma to stallion spermatozoa selected by colloid centrifugation increases sperm progressive motility but is detrimental to chromatin integrity
- 2012
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Ingår i: Theriogenology. - : Elsevier BV. - 0093-691X .- 1879-3231. ; 78, s. 345-352
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Tidskriftsartikel (refereegranskat)abstract
- There is controversy about whether the presence of some seminal plasma (SP) in an equine insemination dose is necessary for promoting fertility. A new technique for improving stallion sperm quality, single layer centrifugation (SLC) using a species-specific colloid, Androcoll-E, selects a sperm subpopulation that is highly motile with normal morphology, intact membranes and good chromatin integrity from the rest of the ejaculate and removes SP. The present study was designed to investigate the effect of restoring homologous SP (5% and 10%) on the progressive motility, velocity, and chromatin integrity of SLC-selected stallion spermatozoa in 44 semen samples over time. Sperm progressive motility (P < 0.01) and the proportion with class A velocity (>50 mu m/sec) were increased in samples where SP was restored, whereas the proportion with class B velocity (10 to 50 mu m/sec) was decreased compared with SLC samples. However, after 24 h cold storage of treated samples, progressive motility was not different for the SP-treated groups compared with SLC, whereas chromatin damage DNA fragmentation index (%DFI) was higher. In contrast, adding SP to untreated 24 h-stored SLC samples did not affect progressive motility although it did increase the proportion of spermatozoa with class A velocity. There was individual variation between stallions whether 5% or 10% SP produced a greater increase in progressive motility. In conclusion, 5% to 10% SP can be added back to SLC-selected samples if considered necessary to optimize fertility. However, it should be added immediately before insemination rather than before storage of the sperm dose, to benefit from the transient increase in sperm progressive motility and avoid increased chromatin damage. (c) 2012 Elsevier Inc. All rights reserved.
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| 30. |
- Morrell, Jane, et al.
(författare)
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Sperm selection using single layer centrifugation prior to cryopreservation can increase thawed sperm quality in stallions
- 2011
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Ingår i: Equine Veterinary Journal. - : Wiley. - 0425-1644 .- 2042-3306. ; 43, s. 35-41
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Tidskriftsartikel (refereegranskat)abstract
- Reasons for performing study: The increasing use of modern reproductive techniques in human medicine has led to a higher demand for isolation of motile sperm. Several of these isolation techniques have been adapted for veterinary use and can be applied for the selection of a superior sperm sample from stallion semen. Until recently a major disadvantage of such isolation techniques was the limitation in sperm volume that could be handled. Androcoll-E had been shown to be successful for processing large volumes of equine semen but there are few data to substantiate the potential beneficial effect of freezing an Androcoll-E selected equine sperm sample to obtain higher quality following thawing.Objectives and methods: In this study, the effect of Androcoll-E treatment of sperm prior to cryopreservation was compared with cushioned centrifugation using ejaculates from 8 different stallions selected because they were known to have semen of differing quality following freezing.Results: Androcoll-E treatment increased measures of semen quality prior to freezing. However, Androcoll-E treatment reduced the yield of sperm following centrifugation when compared with the cushion centrifuged control group (50.9 +/- 14.2% vs. 97.1 +/- 9.0%, respectively). Quality analysis following thawing showed an overall improved sperm quality for Androcoll-E treated samples and average post thaw progressive motility (PM) was 41.6% compared with 30.5% for the cushion centrifuged group.Conclusions and potential relevance: Androcoll-E can be used with good results to select a superior sperm population prior to cryopreservation, in order to produce good-quality frozen thawed semen.
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| 31. |
- Morrell, Jane, et al.
(författare)
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Stallion Sperm Viability, as Measured by the Nucleocounter SP-100, Is Affected by Extender and Enhanced by Single Layer Centrifugation
- 2010
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Ingår i: Veterinary Medicine International. - : Hindawi Limited. - 2090-8113 .- 2042-0048. ; 2010
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Tidskriftsartikel (refereegranskat)abstract
- On-stud assessment of stallion sperm quality can be problematic. A new instrument, the Nucleocounter SP-100, was validated for measuring stallion sperm concentration and viability. It was subsequently used to evaluate sperm viability in Kenney's extender and INRA96. There was a strong correlation between sperm concentrations measured by the Nucleocounter SP-100 and by the Bürker counting chamber (; ). Similarly, there was a good correlation between sperm viability results from the Nucleocounter SP-100 and flow cytometric results (; ). Sperm viability at 24 hours was significantly better for samples extended in INRA96 than in Kenney's extender (). Furthermore, sperm kinematics were better for stored samples in INRA96 than in Kenney's extender. Single Layer Centrifugation selected spermatozoa that maintained their viability better during storage for 24 hours than the uncentrifuged samples. In conclusion, the type of semen extender used and Single Layer Centrifugation were found to influence both the kinematics and viability of stallion spermatozoa. The Nucelocounter-SP100 was considered to be a useful instrument for rapidly measuring stallion sperm concentration and viability.
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| 32. |
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| 33. |
- Rodriguez, Heriberto, et al.
(författare)
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Single layer centrifugation of stallion spermatozoa consistently selects the most robust spermatozoa from the rest of the ejaculate in a large sample size
- 2010
-
Ingår i: Equine Veterinary Journal. - : Wiley. - 0425-1644 .- 2042-3306. ; 42, s. 579-585
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Tidskriftsartikel (refereegranskat)abstract
- Reasons for performing study: An improvement in sperm quality after single layer centrifugation (SLC) has been seen in previous studies using small sample sizes (for example, n = 10 stallions). There is a need to investigate whether this improvement is repeatable over several breeding seasons with a larger number of stallions (n >= 30 stallions).Objective: To make a retrospective analysis of the results of SLC performed on more than 250 sperm samples (176 ejaculates) from 31 stallions in 3 consecutive breeding seasons.Methods: Sperm quality (motility, proportion of morphologically normal spermatozoa and the proportion of spermatozoa with undamaged chromatin) was assessed before and after SLC.Results: All parameters of sperm quality examined were significantly better in sperm samples after SLC than in their unselected counterparts (P < 0.001 for each parameter). The yield of spermatozoa obtained after SLC was influenced by the type of extender used and also by the concentration of spermatozoa in the original ejaculate, with fewer spermatozoa being recovered when the loading dose contained a high concentration of spermatozoa. The optimal concentration was approximately 100 x 10(6)/ml. Sperm concentration in the samples loaded on to the colloid influenced the sperm yield while the type of semen extender affected sperm quality and survival. Furthermore, the scaled-up SLC method was found to be suitable for use with a range of ejaculates, with similar sperm kinematics being observed for standard and scaled-up preparations.Conclusions: SLC consistently improved the quality of stallion sperm samples from a large number of ejaculates. The method could be scaled-up, allowing larger volumes of ejaculate to be processed easily from a wide range of stallions.
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| 34. |
- Rodriguez, Heriberto, et al.
(författare)
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The physiological roles of the boar ejaculate
- 2009
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Ingår i: Society of Reproduction and Fertility Supplement. - 1747-3403. ; 66, s. 1-21
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Tidskriftsartikel (refereegranskat)abstract
- During ejaculation in the boar, sperm cohorts emitted in epididymal cauda fluid are sequentially exposed and resuspended in different mixtures of accessory sex gland secretion. This paper reviews the relevance of such unevenly composed fractions of seminal plasma (SP) in vivo on sperm transport and sperm function and how this knowledge could benefit boar semen processing for artificial insemination (AI). The firstly ejaculated spermatozoa (first 10 ml of the sperm-rich fraction, SRF [P1]) remain mainly exposed to epididymal cauda fluid and its specific proteins i.e. various lipocalins, including the fertility-related prostaglandin D synthase; than to prostatic and initial vesicular gland secretions. P1-spermatozoa are hence exposed to less bicarbonate, zinc or fructose and mainly to PSP-I spermadhesin; than if they were in the rest of the SRF and the post-SRF (P2). Since the P1-SP is less destabilizing for sperm membrane and chromatin, P1-spermatozoa sustain most in vitro procedures, including cryopreservation, the best. Moreover, ejaculated firstly, the P1-spermatozoa seem also those deposited by the boar as a vanguard cohort, thus becoming overrepresented in the oviductal sperm reservoir (SR). This vanguard SR-entry occurs before the endometrial signalling of SP components (as PSP-I/PSP-II and cytokines) causes a massive influx of the innate defensive PMNs to cleanse the uterus from eventual pathogens, superfluous spermatozoa and the allogeneic SP. The SP also conditions the mucosal immunity of the female genital tract, to tolerate the SR-spermatozoa and the semi-allogeneic conceptus. These in vivo gathered data can be extrapolated into procedures for handling boar spermatozoa in vitro for AI and other biotechnologies, including simplified cryopreservation.
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| 35. |
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| 36. |
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| 37. |
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| 38. |
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| 39. |
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| 40. |
- Deori, Sourabh, et al.
(författare)
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Single Layer Centrifugation with 20% or 30% Porcicoll separates the majority of spermatozoa from a sample without adversely affecting sperm quality
- 2020
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Ingår i: Reproduction in Domestic Animals. - : Wiley. - 0936-6768 .- 1439-0531. ; 55, s. 1337-1342
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Tidskriftsartikel (refereegranskat)abstract
- Centrifugation of boar semen through one layer of 40% colloid (Porcicoll) was previously shown to separate spermatozoa from bacteria without having a detrimental effect on sperm quality. However, some spermatozoa were lost. The purpose of the present study was to determine whether 20% or 30% Porcicoll could be used to recover most of the spermatozoa without impacting on sperm quality. Insemination doses (n = 10) from a commercial boar station were sent to the laboratory at the Swedish University of Agricultural Sciences and processed by Single Layer Centrifugation with 20% and 30% Porcicoll approximately 7 hr after semen collection. The resulting sperm samples and controls were evaluated for sperm quality immediately and again after storage at 16-18 degrees C for 4 and 7 days. Sperm recovery was 94 +/- 18% and 87 +/- 15% for 20% and 30% Porcicoll, respectively (p > .05). Sperm mitochondrial membrane potential and chromatin integrity were unaffected (p > .05). The proportion of live spermatozoa producing superoxide (9 +/- 8%, 7 +/- 6% and 3 +/- 1%;p < .05), and the proportion of spermatozoa with high stainability DNA (0.68 +/- 19%, 0.61 +/- 0.22% and 0.96 +/- 0.23%;p < .05- <0.01), were marginally increased whereas membrane integrity, although high, was lower in the centrifuged samples than in the controls (82 +/- 8%, 83 +/- 5% versus 92 +/- 4%;p < .05). In conclusion, centrifugation through 20% or 30% Porcicoll enables most spermatozoa to be recovered, without having a major effect on sperm quality. These results are encouraging for further studies involving microbiological investigation of the processed samples, and scaling-up to process larger volumes of boar ejaculates.
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| 41. |
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| 42. |
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| 43. |
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| 44. |
- Abraham, Maria Celina, et al.
(författare)
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Effect of sperm preparation on development of bovine blastocyst in vitro
- 2016
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Ingår i: Zygote. - 0967-1994 .- 1469-8730. ; 24, s. 825-830
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Tidskriftsartikel (refereegranskat)abstract
- colloids has been used to select normal sperm for assisted reproduction in several species. Animal models can sometimes be used as a preliminary step to investigate sperm preparation methods that are potentially of use for human fertility treatments. In this study bovine semen was prepared using three variants of the single-layer centrifugation sperm selection technique (Small, Mini, Mini-EP) with Bovicoll (Androcoll-B). Computer-assisted sperm motility analysis, the hypo-osmotic swelling test, and the sperm chromatin structure assay were performed on unselected (control) and SLC-selected sperm samples. Mini and Mini-EP gave the highest yield of motile spermatozoa, progressive motility and membrane integrity. In vitro fertilization trials were performed to investigate the fertilizing ability of the frozen-thawed bovine spermatozoa selected with Bovicoll. Mini-SLC (single-layer centrifugation) and swim-up (Control) were performed and cleavage rate and blastocyst rate did not differ significantly between groups. As there was a trend to an increased number of cells in blastocysts in the SLC group, the Mini-SLC method is at least as good as swim-up for selecting frozen-thawed bull spermatozoa for in vitro fertilization (IVF). This method could potentially be used to prepare human sperm for assisted reproduction.
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| 45. |
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| 46. |
- Al-Kass, Ziyad, et al.
(författare)
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Variation among stallions in sperm quality after single layer centrifugation
- 2021
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Ingår i: Reproduction in Domestic Animals. - : Wiley. - 0936-6768 .- 1439-0531. ; 56, s. 848-856
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Tidskriftsartikel (refereegranskat)abstract
- Although single layer centrifugation (SLC) selects robust spermatozoa from stallion semen, the effect of individual variation has not been studied in detail. The objective of this study was to determine the variation among stallions in the effects of SLC on sperm quality during cooled storage for up to 48 hr. Semen samples from seven stallions (18 ejaculates) were split, with one portion being used for SLC and the other serving as a control (CON). Sperm quality (kinematics, reactive oxygen species (ROS) production, membrane integrity (MI) and chromatin integrity) were analysed at 0, 24 and 48 hr using computer-assisted sperm analysis and flow cytometry. Sperm quality was better in SLC than in CON at all timepoints, especially chromatin integrity and MI (p < .0001 for both), and some categories of ROS production (e.g. proportion of live hydrogen peroxide negative spermatozoa, p < .0001), but the degree of improvement varied among stallions and type of ROS (p p < .0001). Total and progressive motility were also better in SLC samples than in CON at 24 and 48 hr (p < .0001), although the effect on sperm kinematics varied. The interaction of treatment, time and stallion was not significant. In conclusion, sperm quality was better in SLC samples than in CON, although there was considerable individual variation among stallions. The improvement in sperm quality, particularly in chromatin integrity, was clearly beneficial, and therefore the use of this technique would be warranted for all stallion semen samples.
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| 47. |
- Cojkic, Aleksandar, et al.
(författare)
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Single layer centrifugation as a method for bacterial reduction in bull semen for assisted reproduction
- 2024
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Ingår i: Veterinary Research Communications. - 0165-7380 .- 1573-7446. ; 48, s. 39-48
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Tidskriftsartikel (refereegranskat)abstract
- Semen samples contain bacteria originating from the animal urogenital tract, environment, and/or contamination during semen processing, negatively affecting sperm quality by producing toxins and/or competing for nutrients in extenders. The aims of this study were to evaluate two methods of Single-layer centrifuges (SLC), high and low density colloid, as a method for bacterial removal from bull semen, and to evaluate sperm quality after treatment. In total, semen samples from 20 bulls (3 ejaculates per bull) were used in this study. Bacterial reduction was evaluated by bacterial quantification (colony forming unit - CFU/mL) while bacterial identification was performed by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) after culturing bacteria on blood agar. Sperm motility parameters were evaluated by Computer Assisted Sperm Analyses (CASA), and sperm chromatin structure assay (SCSA) by Flow cytometry. Both, High and Low density SLC reduced number of bacteria significantly (p < 0.001) compared with control. The difference in bacterial count between High and Low SLC was also significant (p < 0.001). Furthermore, High density SLC was successful in removing almost all Bacillus and Proteus spp. Most CASA parameters were significantly improved after both treatments (p < 0.001, p < 0.01, p < 0.05). The Deoxyribonucleic acid (DNA) fragmentation index evaluated by SCSA in High (p < 0.01) and Low (p < 0.05) SLC group differed significantly compared with control. Single-layer centrifugation (SLC) with either a high or a low density colloid is a suitable method for bacterial removal in bull semen.
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| 48. |
- Goodla, Lavanya, et al.
(författare)
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Quality of bull spermatozoa after preparation by single-layer centrifugation
- 2014
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Ingår i: Journal of Dairy Science. - : American Dairy Science Association. - 0022-0302 .- 1525-3198. ; 97, s. 2204-2212
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Tidskriftsartikel (refereegranskat)abstract
- The present study aimed to evaluate the effect of single-layer centrifugation (SLC) through a species-specific colloid (Androcoll-B; patent pending, J. M. Morrell) on bull sperm quality. Computer-assisted sperm analysis of motility and flow cytometric analysis of sperm viability (SYBR-14/propidium iodide staining), chromatin integrity (acridine orange staining), reactive oxygen species production [Hoechst 33258-hydroethidine-2',7'-dichlorodihydrofluorescein diacetate (HO-HE-DCFDA) staining], mitochondrial membrane potential (staining with JC-1 probe), and protein tyrosine phosphorylation (specific antibody staining) were performed on unselected and SLC-selected sperm samples. Single-layer centrifugation of bull spermatozoa resulted in the selection of a sperm population that had high mitochondrial membrane potential, a higher content of phosphorylated protein, and more reactive oxygen species than control samples. Sperm chromatin damage was lower in the SLC samples although sperm viability and motility did not differ between SLC samples and controls. These observations suggest that SLC of bull semen in a soybean-containing extender improved some, but not all, parameters of sperm quality.
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| 49. |
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| 50. |
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