Centrifugation Force and Time Alter CASA Parameters and Oxidative Status of Cryopreserved Stallion Sperm

Marzano, Giuseppina; Moscatelli, Natalina; Giacomo, Mariangela Di; Martino, Nicola Antonio; Lacalandra, Giovanni Michele; Dell’Aquila, Maria Elena; Maruccio, Giuseppe; Primiceri, Elisabetta; Chiriaco, Maria; Zara, Vincenzo; Ferramosca, Alessandra

doi:10.3390/biology9020022

Cited by 9 publications

(10 citation statements)

References 59 publications

(82 reference statements)

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“…However; the mechanical actions of centrifugation and spermatozoa compaction at the bottom could reduce sperm quality [ 48 ]. We observed that TM and velocities of spermatozoa declined after sperm washing, in agreement with Marzano et al [ 49 ] in equine sperm. In a previous study, we observed a beneficial effect of sperm washing on motility in goats [ 18 ] without a significant deleterious effect of the centrifugation process, in agreement with others [ 47 ].…”

Section: Discussionsupporting

confidence: 93%

Effect of Sperm Concentration and Storage Temperature on Goat Spermatozoa during Liquid Storage

Sadeghi

Gallego

García-Colomer

et al. 2020

Biology

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The use of cooled semen is relatively common in goats. There are a number of advantages of cooled semen doses, including easier handling of artificial insemination (AI) doses, transport, more AI doses per ejaculate, and higher fertility rates in comparison with frozen AI doses. However, cooled semen has a short shelf life. The objective of this study was to examine the effect of temperature and sperm concentration on the in vitro sperm quality during liquid storage for 48 h, including sperm motility and kinetics, response to oxidation, mitochondrial membrane potential (MMP) and DNA fragmentation in goats. Three experiments were performed. In the first, the effects of liquid preservation of semen at different temperatures (5 °C or 17 °C), durations (0, 24 and 48 h) and sperm concentrations (250 × 106 sperm/mL (1:2 dilution rate), 166.7 × 106 sperm/mL (1:3 dilution rate) or 50 × 106 sperm/mL (1:10 dilution rate)) on sperm motility and kinetics were studied. In the second experiment, the effect of temperature, sperm washing and concentration on sperm motility and DNA fragmentation was studied. Finally, the effect of sperm concentration and duration of storage at 5 °C on sperm motility, response to oxidative stress and MMP was examined. We found that refrigerated liquid storage of goat sperm impaired sperm quality, such as motility, MMP and response to oxidation, as storage time increased; however, sperm DNA fragmentation index was not significantly affected. Liquid storage at 5 °C preserved higher total motility than at 17 °C. Moreover, we observed that the reduction of sperm concentration below 500 × 106 sperm/mL did not seem to improve the quality of spermatozoa conserved in milk-based extender in the conditions tested.

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Section: Discussionsupporting

confidence: 93%

Effect of Sperm Concentration and Storage Temperature on Goat Spermatozoa during Liquid Storage

Sadeghi

Gallego

García-Colomer

et al. 2020

Biology

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“…However, there seems to be a general absence of research on the impact of centrifugation on any mammalian cells. In fact, most of the scant existing literature stems from the medical fields andrology and urology [ 33 , 34 , 35 ]. Interestingly, results of those studies on sperm cells are quite similar to our data on neutrophils: Marzano et al were able to show, that with increasing g -force and duration of centrifugation, sperm cells suffered a major loss in motility compared to non-centrifuged samples [ 34 ].…”

Section: Discussionmentioning

confidence: 99%

“…In fact, most of the scant existing literature stems from the medical fields andrology and urology [ 33 , 34 , 35 ]. Interestingly, results of those studies on sperm cells are quite similar to our data on neutrophils: Marzano et al were able to show, that with increasing g -force and duration of centrifugation, sperm cells suffered a major loss in motility compared to non-centrifuged samples [ 34 ]. Furthermore, Zini et al, as well as Marzano et al, observed a significant decrease in DNA integrity in centrifuged sperm cells in comparison to controls [ 34 , 35 ].…”

Section: Discussionmentioning

confidence: 99%

Paralytic Impact of Centrifugation on Human Neutrophils

2022

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Centrifugation is a common step in most of the popular protocols for the isolation of neutrophils from whole blood. Inconsistent results from previous studies on neutrophils may originate from an underestimation of the centrifugation effect, as in consequence impaired, not native cells, being investigated. We hypothesize, that centrifugation significantly impairs major neutrophil functions. However, there is no data yet whether the application of g-force itself or the product of g-force and duration of centrifugation (=“g-time”) defines the impact on neutrophils. Neutrophils were isolated from whole blood via centrifugation with different g-times and subsequently analyzed via live cell imaging for migration, as well as via flow cytometry for oxidative burst and surface antigen expression. Chemotactic migration was significantly reduced with increasing g-time. Oxidative burst decreased likewise the higher the g-time applied. Expression of CD11b was no longer upregulated in response to an n-formylmethionine-leucyl-phenylalanine (fMLP) stimulus in neutrophils having experienced high g-time during the isolation process. We conclude that centrifugation “paralyzes” neutrophils in the form of a significant decrease in functionality. Future investigations on neutrophil granulocytes should reduce the g-time load as far as possible.

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“…We tested a 200× g force, not previously described for cryopreserved stallion sperm and its effect on motility. As previously reported, we found that total sperm motility was negatively affected by centrifugation times longer than 2 min, even at lower g-forces [65]. In this work, we have used samples that were frozen poorly (<35% motility post-thawing) [67], suggesting that a better outcome could be possible using samples with better post-thaw motility.…”

Section: Discussionmentioning

confidence: 75%

“…Moreover, several sperm selection protocols used during ARTs require multiple centrifugations. It was previously described that different conditions of centrifugation alter motility and oxidative status, and consequently increase the DNA damage of cryopreserved stallion sperm [ 65 , 66 ]. We tested a 200× g force, not previously described for cryopreserved stallion sperm and its effect on motility.…”

Section: Discussionmentioning

confidence: 99%

Effects of In Vitro Interactions of Oviduct Epithelial Cells with Frozen–Thawed Stallion Spermatozoa on Their Motility, Viability and Capacitation Status

Gimeno

Bariani

Laiz-Quiroga

et al. 2021

Animals

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Cryopreservation by negatively affecting sperm quality decreases the efficiency of assisted reproduction techniques (ARTs). Thus, we first evaluated sperm motility at different conditions for the manipulation of equine cryopreserved spermatozoa. Higher motility was observed when spermatozoa were incubated for 30 min at 30 × 106/mL compared to lower concentrations (p < 0.05) and when a short centrifugation at 200× g was performed (p < 0.05). Moreover, because sperm suitable for oocyte fertilization is released from oviduct epithelial cells (OECs), in response to the capacitation process, we established an in vitro OEC culture model to select a sperm population with potential fertilizing capacity in this species. We demonstrated E-cadherin and cytokeratin expression in cultures of OECs obtained. When sperm–OEC cocultures were performed, the attached spermatozoa were motile and presented an intact acrosome, suggesting a selection by the oviductal model. When co-cultures were incubated in capacitating conditions a greater number of alive (p < 0.05), capacitated (p < 0.05), with progressive motility (p < 0.05) and with the intact acrosome sperm population was observed (p < 0.05) suggesting that the sperm population released from OECs in vitro presents potential fertilizing capacity. Improvements in handling and selection of cryopreserved sperm would improve efficiencies in ARTs allowing the use of a population of higher-quality sperm.

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Centrifugation Force and Time Alter CASA Parameters and Oxidative Status of Cryopreserved Stallion Sperm

Cited by 9 publications

References 59 publications

Effect of Sperm Concentration and Storage Temperature on Goat Spermatozoa during Liquid Storage

Effect of Sperm Concentration and Storage Temperature on Goat Spermatozoa during Liquid Storage

Paralytic Impact of Centrifugation on Human Neutrophils

Effects of In Vitro Interactions of Oviduct Epithelial Cells with Frozen–Thawed Stallion Spermatozoa on Their Motility, Viability and Capacitation Status

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