Molecular Markers in Sperm Analysis

Payan-Carreira, Rita; Borges, Paulo; Mir, Fernando; Fontbonne, Alain

doi:10.5772/52231

Cited by 6 publications

(6 citation statements)

References 48 publications

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“…The TUNEL assay is one of the most common methods used to assess sperm DNA fragmentation [ 8 , 11 ]. Several kinds of TUNEL assay kits are available commercially, which are cost effective and standardized by the manufacturers.…”

Section: Discussionmentioning

confidence: 99%

“…Sperm DNA strand breaks occur in each ejaculate and can also be induced by ROS [ 9 , 10 ]. Therefore, the TUNEL assay may be the most appropriate method for assessing sperm DNA strand breaks [ 8 , 11 ]. Furthermore, infertile patients whose spermatozoa had a high percentage of DNA damage as detected by TUNEL assay showed significantly lower conception rates [ 12 ].…”

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confidence: 99%

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Evaluation of sperm DNA damage in bulls by TUNEL assay as a parameter of semen quality

Takeda

Uchiyama

Kinukawa

et al. 2015

Journal of Reproduction and Development

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Sperm DNA damage affects the conception rate resulting from human assisted reproduction technology. The objective of this study was to adapt the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay to provide a quality parameter for bull semen based on the detection of sperm DNA damage. Fresh semen was collected from two Japanese Black bulls (A, B) several times over the course of a year, and the percentage of TUNEL-positive spermatozoa (sperm TUNEL index) was determined. Individual differences in semen were detected using the sperm TUNEL index in these bulls (P < 0.01). The sperm TUNEL index of cryopreserved semen obtained from test-mated Japanese Black (n = 30, including two bulls with a conception rate lower than 10%) and Holstein (n = 34) bulls were analyzed. The average sperm TUNEL index and conception rate resulting from artificial insemination (AI) were 4.7% and 55.7% for Japanese Black, and 4.9% and 39.5% for Holstein, respectively. A weak negative correlation between sperm TUNEL index and conception rate was observed in Holstein bulls (P < 0.05). Semen samples from six bulls with more than 10% sperm TUNEL index were studied, and these samples showed low sperm viability. However, semen resulting in a very low conception rate did not have a high sperm TUNEL index. Although it would be difficult to predict a low conception rate resulting from AI using the sperm TUNEL index alone, the index can be used as an additional parameter to provide a more comprehensive description of semen quality.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Evaluation of sperm DNA damage in bulls by TUNEL assay as a parameter of semen quality

Takeda

Uchiyama

Kinukawa

et al. 2015

Journal of Reproduction and Development

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“…Sperm velocity and beat frequency estimated using computer-assisted sperm analysis has been used in several species (Yang and Tiersch 2011), and in Atlantic salmon sperm curved- and straight-line velocity showed correlated with post-thaw sperm motility and fertility (Figueroa et al 2016). In addition, other cellular-level analyses, such as plasma membrane integrity, mitochondria membrane integrity, and acrosome integrity assessed by flow cytometry (Daly and Tiersch 2011), and molecular level assays such as DNA integrity (Gandini et al 2006) and DNA markers (Payan-Carreira et al 2013), were also investigated for correlation analysis with post-thaw sperm viability.…”

Section: Discussionmentioning

confidence: 99%

A Strategy for Sperm Cryopreservation of Atlantic Salmon, Salmo salar, for Remote Commercial‐scale High‐throughput Processing

Yang

Buchanan³

et al. 2017

J World Aquaculture Soc

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Sperm cryopreservation is an essential tool for long-term storage of genetic resources for aquaculture fishes. The goal of this study was to develop an efficient and streamlined protocol for high-throughput processing for sperm cryopreservation in Atlantic salmon, . The objectives were to evaluate: 1) osmolality of blood serum for determining extender osmolality; 2) effects of extenders for fresh sperm dilution and refrigerated storage; 3) effects of methanol and dimethyl sulfoxide (DMSO) on fresh sperm motility, and 4) motility and fertility after thawing. In this study, sperm samples were collected at a hatchery site in Canada, and shipped to a freezing site located 2200 miles (3550 km) away in the United States. Evaluation of three extenders indicated that Mounib solution was suitable for diluting dry sperm for sample processing. Ten percent of methanol or DMSO was less toxic to sperm cells than was 15% within 30 min. Further testing with methanol at 5, 10, and 15%, and sperm solution:extender dilutions (v:v) of 1:1, 1:3, 1:19 (at concentrations of ~5×10; 3×10, and 1×10 cells/mL) indicated that methanol at 5% and 10% showed less toxicity to fresh sperm within 1 hr at sperm: extender dilutions of 1:1 and 1:3. Post-thaw motility of sperm cryopreserved with 10% methanol was significantly higher than that with 10% DMSO, and fertility reflected those results (0-1% in DMSO vs. 38-55% in methanol). Further evaluation of sperm cryopreservation with 10 and 15% methanol at sperm dilution ratios of 1:1, 1:3, 1:19 indicated post-thaw motility in 10% methanol was significantly higher than that in 15% methanol, and post-thaw fertility in 10% methanol at 1:1 and 1:3 dilution ratios had fertilization rates similar to that of fresh sperm controls. Sperm samples from 12 males cryopreserved with 10% methanol showed male-to-male variation in post-thaw motility (0-36%). Overall, a simplified standard protocol was established for cryopreservation of shipped sperm of Atlantic salmon using extender without egg yolk and yielded satisfactory post-thaw motility and fertilization rates. This procedure can be readily adopted by aquaculture facilities to take advantage of high-throughput cryopreservation capabilities at remote service centers. Most importantly, this approach lays the groundwork for an alternative commercial model for commercial-scale production, quality control and development of industrial standards. Control of male variability and sperm quality remain important considerations for future work.

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“…It is believed that sperm capacitation is thermally dependent, and the blocking effect is overcome when the sperm returns to a temperature of 37 C [32]. The key factor here is the partial cryopreservation of the cell membrane, in particular, in the area of the acrosomal cover, resulting in changes in intracellular calcium con-Sperm (½10 6 cell per semen dose) with a straight-forward movement depending on the temperature of defrosting and different periods of subsequent incubation: 1 -5 h, 2 -20 h, centration [34]. Even though in cattle sperm motility correlates with fertilizing ability and can be successfully used for its rapid testing, in practice, it is necessary to control the life expectancy of germ cells outside the body.…”

Section: Motility and The Proportion (%) Of Mobile Spermatozoa In Semmentioning

confidence: 99%

Optimal Mode to Thaw Cryopreserved Sperm of Holstein Bull Sires

Комбарова¹,

ЕСКИН²,

Абилов³

et al. 2018

s-h. biol.

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A b s t r a c tAs milk yielding increases, the number of animals with defective sexual cycles and quiet manifestations of hunting grows, and therefore there is a need for semen with high quality. Also, at artificial insemination, after dilution of the native semen, packaging, cryopreservation and thawing, the number of spermatozoa per sperm dose decreases. This also necessitates semen with high quality parameters. One of the key stages of the standard processing of cryopreserved bovine semen prior to artificial insemination is the thawing procedure. Therefore, cryopreservation and thawing procedures should be optimized with regard to the proposed modifications of these methods and the changing reproductive abilities of animals. In our study, we compared the effects of previously recommended thawing protocols and those proposed by us on the safety of spermatozoa of Russian Holstein sires and identified the modes that reliably provide a prolonged positive effect on the quality characteristics of spermatozoa and better preservation of their viability after thawing. The semen of 6-7 year-old Holstein bulls was cryopreserved in polypropylene straws (72 doses) with the use of the IMV Technology equipment (France) and in uncoated pellets (90 doses) on the dry ice plates according to GOST State Standard 26030-2015 and the National Technology for Freezing and Handling Semen of Pedigree Bull Sires. Thus, 162 semen doses 0.25 ml each were analyzed. The sperm motility and movement velocity parameters were assessed immediately and after in vitro post-thawed incubation for 5 to 24 hours at 38 С. The percentage of motile semen was visually assessed with the Olympus microscope CX41 (Japan) at ½150 magnification. Thereafter, in the same samples the motility (%), the number of the highly motile sperm (%), and the sperm velocity (mcm/sec) were automatically measured with the SFA-500 Sperm Analyzer (Biola Company, Ltd, Russia). The dynamic of thawing temperature up to the final value was measured with the Center 304 thermocouple. It has been ascertained that the effects of different procedures used for thawing straws and pellets are quite steady and approximately the same. However, the best results of the male sex cell survival after the long in-vitro incubation period (for 24 hours at the animal body temperature of 38 С) are in the samples thawed in a water bath at 38 С for 10 sec when compared to those for the other methods (p < 0.001).For more than 70-year history of cryopreservation for male germ cells, the method has been widely used both in research programs on biodiversity conservation [1] and for commercial purposes in the creation of sperm banks of different animal species [2] and human [3]. Such a long history of the issue allows researchers to discuss not only the achievements but also the risks of artificial insemination [4,5]. It is believed that the increase in milk production has a neg-

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Molecular Markers in Sperm Analysis

Cited by 6 publications

References 48 publications

Evaluation of sperm DNA damage in bulls by TUNEL assay as a parameter of semen quality

Evaluation of sperm DNA damage in bulls by TUNEL assay as a parameter of semen quality

A Strategy for Sperm Cryopreservation of Atlantic Salmon, Salmo salar, for Remote Commercial‐scale High‐throughput Processing

Optimal Mode to Thaw Cryopreserved Sperm of Holstein Bull Sires

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