| 1. |
- Balao da Silva, C. M., et al.
(författare)
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Effect of Overnight Staining on the Quality of Flow Cytometric Sorted Stallion Sperm: Comparison with Tradtitional Protocols
- 2014
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Ingår i: Reproduction in domestic animals. - : Wiley. - 0936-6768 .- 1439-0531. ; 49:6, s. 1021-1027
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Tidskriftsartikel (refereegranskat)abstract
- Flow cytometry is considered the only reliable method for the separation of X and Y chromosome bearing spermatozoa in equines. The MoFlo SX DP sorter is highly efficient, allowing the production of foals of the desired sex. However, to achieve acceptable pregnancy rates the currently used protocol requires working with fresh semen obtained close to, or at, the sorting facility. An alternative protocol was tested during two consecutive breeding seasons. Fresh stallion semen was cooled for 20 h, during which staining with Hoechst 33342 took place. On the following day, this sample was flow sorted and compared with spermatozoa from the same ejaculate that had been sexed on the previous day. All sperm parameters evaluated remained unchanged when fresh sorted and refrigerated sorted semen were compared. Pre-sorting storage at 5 degrees C did not alter sperm velocities nor kinetics, viability or membrane permeability, production of reactive oxygen species, mitochondrial membrane potential or DNA fragmentation index of the sorted sample. The findings open for the possibility of using semen from stallions housed far from the sorting facilities. Processed and stained sperm could be shipped refrigerated on the previous day, sorted and inseminated on the next day.
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| 2. |
- Balao da Silva, C M., et al.
(författare)
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Sex sorting increases the permeability of the membrane of stallion spermatozoa
- 2013
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Ingår i: Animal Reproduction Science. - : Elsevier Masson. - 0378-4320 .- 1873-2232. ; 138:3-4, s. 241-251
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Tidskriftsartikel (refereegranskat)abstract
- At present, the only repeatable means of selecting the sex of offspring is the Beltsville semen sorting technology using flow cytometry (FC). This technology has reached commercial status in the bovine industry and substantial advances have occurred recently in swine and ovine species. In the equine species, however, the technology is not as well developed. To better understand the changes induced in stallion spermatozoa during the sorting procedure, pooled sperm samples were sorted: sperm motility and kinematics were assessed using computer assisted sperm analysis, sperm membrane integrity was assessed using the YoPro-1 assay, while plasmalemmal stability and lipid architecture were assessed using Merocyanine 540/SYTOX green and Annexin-V, respectively. Lipid peroxidation was also investigated with the probe Bodipy(581/591)-C11. All assays were performed shortly after collection, after incubation and after sex sorting using FC. In order to characterize potential molecular mechanisms implicated in sperm damage, an apoptosis protein antibody dot plot array analysis was performed before and after sorting. While the percentage of total motile sperm remained unchanged, sex sorting reduced the percentages of progressive motile spermatozoa and of rapid spermatozoa as well as curvilinear velocity (VCL). Sperm membranes responded to sorting with an increase in the percentage of YoPro-1 positive cells, suggesting the sorted spermatozoa had a reduced energy status that was confirmed by measuring intracellular ATP content.
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| 3. |
- Ortega Ferrusola, C., et al.
(författare)
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Stallion spermatozoa surviving freezing and thawing experience membrane depolarization and increased intracellular Na
- 2017
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Ingår i: Andrology. - : WILEY. - 2047-2919 .- 2047-2927. ; 5:6, s. 1174-1182
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Tidskriftsartikel (refereegranskat)abstract
- In order to gain insight of the modifications that freezing and thawing cause to the surviving population of spermatozoa, changes in the potential of the plasma membrane (Em) and intracellular Na+ content of stallion spermatozoa were investigated using flow cytometry. Moreover, caspase 3 activity was also investigated and the functionality of the Na+-K+ ATPase pump was investigated before and after freezing and thawing. Cryopreservation caused a significant (pamp;lt;0.001) increase in the subpopulation of spermatozoa with depolarized sperm membranes, concomitantly with an increase (pamp;lt;0.05) in intracellular Na+. These changes occurred in relation to activation of caspase 3 (pamp;lt;0.001). Cryopreservation reduced the activity of the Na-K+ pump and inhibition of the Na+-K+ ATPase pump with ouabain-induced caspase 3 activation. It is concluded that inactivation of Na+-K+ ATPase occurs during cryopreservation, an inhibition that could play a role explaining the accelerated senescence of the surviving population of spermatozoa.
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| 4. |
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| 5. |
- 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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| 6. |
- Abraham, Maria Celina, et al.
(författare)
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Semen collection methods in alpacas
- 2017
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Ingår i: Veterinary Record. - : Wiley. - 0042-4900 .- 2042-7670. ; 180, s. 613-614
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Forskningsöversikt (refereegranskat)abstract
- South American camelids, particularly alpacas, are gaining in popularity outside their native lands. Reproductive biotechnologies, such as artificial insemination or embryo transfer with cryopreserved embryos, are more complicated in these species than many others due to differences in their reproductive physiology compared to the more commonly encountered domestic livestock. This article reviews the methods currently available for obtaining semen or spermatozoa from alpacas and describes some of the problems associated with handling the viscous seminal plasma characteristic of camelids. Possibilities and limitations of reproductive biotechnologies in this species are discussed, and future developments are outlined, especially some new techniques which are currently being developed for use with camel semen and may be transferable to alpacas.
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| 7. |
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| 8. |
- Abraham, Maria Celina, et al.
(författare)
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Testicular length as an indicator of the onset of sperm production in alpacas under Swedish conditions
- 2016
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Ingår i: Acta Veterinaria Scandinavica. - : Springer Science and Business Media LLC. - 0044-605X .- 1751-0147. ; 58
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Tidskriftsartikel (refereegranskat)abstract
- Background The popularity of alpacas (Vicugna pacos) is increasing in Sweden as well as in other countries; however, knowledge about optimal management practices under Swedish conditions is still limited. The wide age range reported when the onset of puberty can occur, between 1 and 3years of age, makes management decisions difficult and may be influenced by the conditions under which the alpacas are kept. The aim of this study was to find out when Swedish alpacas can be expected to start producing sperm, by using testicular length and body condition score as a more precise indirect indicator than age. Results This study suggests that animals with a testicular length ≥3.8cm would be producing sperm; however, if it is crucial to know that there is no sperm production for management purposes, the threshold level for testicular length used to differentiate between sperm-producing and non-sperm producing animals should be ≤1.6cm instead. If only one variable is considered, testicular length appears to better than age alone to predict sperm production. Body condition score together with testicular length explains the individual onset of puberty and better guide management recommendations. Conclusions Using a combination of these parameters (testicular length, body condition score and age) as a tool for decision making for alpaca husbandry under Swedish conditions is suggested.
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| 9. |
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| 10. |
- Al-Essawe, Essraa M, et al.
(författare)
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Addition of seminal plasma to thawed stallion spermatozoa did not repair cryoinjuries
- 2018
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Ingår i: Animal Reproduction Science. - : Elsevier BV. - 0378-4320 .- 1873-2232. ; 196, s. 48-58
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Tidskriftsartikel (refereegranskat)abstract
- Freezing and thawing processes induce structural and functional damage to sperm plasma membranes and internal organelles. Adding seminal plasma (SP) has been found to minimize or repair the cryoinjuries in some species. The objective of this study was to investigate whether adding SP from stallions of known freezability after thawing could repair cryoinjuries. Semen was collected from warmblood stallions (n = 8, three ejaculates/stallion) and processed by Single Layer Centrifugation (SLC) to remove SP prior to freezing. Pooled SP (5%) from bad freezer (BF) or good freezer (GF) stallions was added after thawing. Post-thaw sperm quality was assessed by flow cytometry in terms of chromatin integrity (ßI), membrane integrity, mitochondrial membrane potential (MMP), reactive oxygen species (ROS), and MitoSOX. Sperm kinematics were also assessed by computer-assisted sperm analysis. The ßI was lower in SLC control (C) than in BF or GF (P < 0.0001, P < 0.0003 respectively). The proportion of viable spermatozoa with intact cell membranes was higher in C than in SP treated groups (C vs. BF, P = 0.02; C vs GF, P = 0.05). There were fewer spermatozoa with low MMP and more with high MMP for C than GF (P = 0.006). The spermatozoa treated with SP from good freezers produced more ROS than when treated with SP from bad freezers (P = 0.007). Motility parameters were not affected by adding SP. In conclusion, adding SP after thawing does not have a beneficial effect on sperm quality, suggesting an inability to repair stallion sperm cryoinjuries, regardless of whether the SP originated from stallions semen, which has good or bad quality after thawing.
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