Dataset Information

Comparative characteristics between calyculin A-induced and thimerosal-induced hyperactivation of cryopreserved bovine spermatozoa.

ABSTRACT: This study aimed to characterize calyculin A (CL-A)-induced and thimerosal-induced hyperactivation of cryopreserved bovine spermatozoa. Hyperactivation was effectively induced by treating with 10 nM CL-A for 60 min in the presence of cyclic AMP analogs, extracellular Ca²⁺, and albumin or with 12.5 µM thimerosal briefly in the absence of these capacitation-supporting factors. Majority of the spermatozoa exhibiting CL-A-induced hyperactivation were characterized by the 3-dimensional helical movement with head rotation, higher degree of flagellar curvature, and faster beating of the flagella than those exhibiting thimerosal-induced hyperactivation of the 2-dimensional planar movement without head rotation. The CL-A-induced hyperactivation was linked to the activation of cAMP/protein phosphorylation-dependent signaling cascades and to the decreased activity of glycogen synthase kinase-3α (GSK-3α). In contrast, the thimerosal-induced hyperactivation was suppressed by pretreatment with CL-A and cyclic AMP analogs in the absence of CaCl₂ to activate cAMP/protein phosphorylation-dependent signaling cascades. Additionally, the intracellular Ca²⁺ level in live sperm flagella was significantly higher in the CL-A-treated samples than in the thimerosal-treated samples. These results indicate that CL-A-induced hyperactivation of cryopreserved bovine spermatozoa is an extracellular Ca²⁺-dependent type with the 3-dimensional helical movement, which can be regulated not only by the activation of cAMP/protein phosphorylation-dependent signaling cascades, leading to a large enhancement of the intracellular Ca²⁺ level, but also by the reduction in GSK-3α activity. Considering the different characteristics of thimerosal-induced hyperactivation, our results suggest that the diversity of sperm hyperactivation arises from different combinations of flagellar bending and head rotation.

SUBMITTER: Miyamoto N

PROVIDER: S-EPMC10267582 | biostudies-literature | 2023 Jun

REPOSITORIES: biostudies-literature

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Comparative characteristics between calyculin A-induced and thimerosal-induced hyperactivation of cryopreserved bovine spermatozoa.

Miyamoto Natsuko N Ohya Akihiro A Duritahala Sakase Mitsuhiro M Harayama Hiroshi H

The Journal of reproduction and development 20230418 3

This study aimed to characterize calyculin A (CL-A)-induced and thimerosal-induced hyperactivation of cryopreserved bovine spermatozoa. Hyperactivation was effectively induced by treating with 10 nM CL-A for 60 min in the presence of cyclic AMP analogs, extracellular Ca<sup>2+</sup>, and albumin or with 12.5 µM thimerosal briefly in the absence of these capacitation-supporting factors. Majority of the spermatozoa exhibiting CL-A-induced hyperactivation were characterized by the 3-dimensional hel ...[more]

PMID: 37081666

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Project description:Graphical abstractAbstractSperm cryopreservation is a valuable tool for the long-term preservation of male fertility. Thus, determining the optimal technique for isolating spermatozoa post-thaw is vital to ensure recovery of the highest quality spermatozoa with minimal iatrogenic damage. This not only enhances the chances of successful conception but also reduces the risk of genetic damage in the embryo. To address this issue, human semen samples were cryopreserved using a slow freezing protocol and Quinn's Advantage™ Sperm Freeze medium. The samples were subsequently thawed and subjected to three types of sperm isolation procedures: direct swim-up, density gradient centrifugation, and electrophoretic separation using the Felix™ device. Cryopreservation led to the anticipated loss of sperm motility and vitality in association with increases in lipid peroxidation and DNA damage. Following sperm selection, all three isolation techniques resulted in an increase in sperm motility which was particularly evident with the swim-up and Felix™ procedures. The latter also significantly improved sperm vitality. There were no differences between sperm separation techniques with respect to morphology, and mitochondrial reactive oxygen species generation remained essentially unchanged when cell vitality was taken into account. By contrast, major differences were observed in DNA integrity and lipid aldehyde formation, where Felix™ isolated cells exhibiting significantly less DNA damage than the other isolation procedures as well as lower levels of 4-hydroxynonenal formation. Electrophoretic sperm isolation, therefore, offers significant advantages over alternative separation strategies, in terms of the quality of the gametes isolated and the time taken to achieve the isolation.Lay summaryLong-term storage of sperm is vital to assisted reproductive technology because it permits the preservation of fertility that might be compromised as a result of factors such as chemotherapy or vasectomy. This goal can be achieved via cryopreservation - the freezing of cells to -196°C. When the sperm are subsequently required for conception, they must be carefully separated from the cryopreservation medium in a manner that maximizes the chances of successful conception and minimizes the risk of genetic defects in the offspring. In this paper, three isolation techniques were compared for their ability to separate ideal sperm from semen and media following cryopreservation. It was found that cryopreservation led to lower levels of motility and vitality and created higher levels of DNA and cell membrane damage. Of the three techniques compared, only cells separated on the basis of their size and electric charge (electrophoretic isolation) exhibited significantly lower levels of DNA fragmentation.

Project description:Epididymal spermatozoa obtained post mortem are considered a valuable source of genetic material which is often irrevocably lost. This makes these gametes constitute a key element in protection and restitution programs. The wisent (Bison bonasus, Linnaeus 1758) is a species that survived in zoos after extinction from its natural habitat. This resulted in a narrowing of the genetic pool of the whole population, which is at present derived from only 12 ancestors. Currently, wisent protection programs are aimed at preserving the genetic diversity by establishing a germplasm bank. The objective of this study was to comprehensively characterize the morphology, morphometry and functionality of wisent epididymal spermatozoa and evaluate the effectiveness of their cryopreservation in extender based on Tris buffer and chicken egg yolk. The median total number of spermatozoa obtained from one individual was 1985.0 × 106 (62.5 × 106-7452.0 × 106). These gametes were characterized by median: 40.0% (0.5-70.0%) subjective motility, 69.8% (32.5-90.0%) viability and 54.3% (10.5-83.3%) normal morphology. The sperm head had a median size of 5.0 μm (3.5-6.7 μm) width, 8.5 μm (6.4-11.3 μm) length and 36.9 μm2 (23.7-48.6 μm2) surface area. The viable population of the obtained gametes was characterized by median values 53.2% (4.5-80.3%) of intact sperm membrane, 50.8 (26.0-76.6%) of intact acrosome, 0.4% (0-98.7%) of fragmented chromatin, 5.9% (0.0-88.8%) of cells with high mitochondrial potential and 42.1% (8.3-63.7%) without lipid peroxidation. The viable population of the frozen/thawed gametes was characterized by median values: 18.4% (2.4-57.9%) of intact sperm membrane, 35.1 (11.9-56.7%) of intact acrosome, 0.07% (0-89.2%) of fragmented chromatin, 12.8% (0.0-49.7%) of cells with high mitochondrial potential and 16.3% (2.2-53.6%) without lipid peroxidation. Due to the material originating from a relatively large number of wild individuals, the research presented here contributed to the description of certain species standards for the assessment of wisent epididymal spermatozoa. The presented effect of cryopreservation on these gametes justifies the use of an extender based on Tris buffer with the addition of chicken egg yolk. The obtained effects are satisfactory from the point of view of preserving valuable genetic material and their use in ART.

Dataset Information

Comparative characteristics between calyculin A-induced and thimerosal-induced hyperactivation of cryopreserved bovine spermatozoa.

Publications

Comparative characteristics between calyculin A-induced and thimerosal-induced hyperactivation of cryopreserved bovine spermatozoa.

Similar Datasets

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