The impact of cushioned centrifugation protocols on semen quality of stallions.
Abstract: The objective was to determine if decreased cushion-fluid volume and increased sperm number during centrifugation, or if sperm concentration of extended semen following centrifugation, affected stallion sperm quality. Three ejaculates from each of three stallions were subjected to cushioned centrifugation (1,000g for 20 min). Cushion-fluid volume was set at 1 or 3.5 ml, and sperm number per centrifuge tube was set 1 billion or 3 billion. Following centrifugation, sperm pellets were resuspended in semen extender containing 20% seminal plasma (v/v) with sperm concentrations of 25 or 250 million/mL. Sperm recovery rate among centrifugation treatment groups was compared. Motion characteristics, plasma membrane intactness (SMI), and DNA quality (COMPαt) of sperm were compared among treatment groups and uncentrifuged controls immediately following centrifugation (Time 0 h) and following 24 h of cooled storage (Time 24 h). Centrifugation treatment did not affect sperm recovery rate (P > 0.05). At Time 0 h, no differences in experimental end points were detected between cushion-fluid volumes tested (P > 0.05). Values for percent total sperm motility, percent progressive sperm motility, and track straightness were similar between sperm-number treatments subjected to centrifugation (P > 0.05). At Time 24 h, values for all experimental endpoints were similar between centrifugation treatments for cushion volume per tube, and between centrifugation treatments for sperm number per tube (P > 0.05). Centrifugation treatments and control treatments were similar for five of six variables tested (P > 0.05). Sperm storage concentrations of 25 × 10(6) and 250 × 10(6)/mL yielded similar values for percent total sperm motility, percent progressive sperm motility, percent SMI, and percent COMPαt (P > 0.05). A storage concentration of 250 × 10(6) sperm/ml yielded higher values for curvilinear velocity, and lower values for straightness, than all other groups (P < 0.05). In conclusion, centrifugation with as little as 1 ml of cushion fluid and a sperm number of up to 3 × 10(9) sperm in 50-ml conical-bottom centrifuge tubes had no detrimental effect on initial or cool-stored sperm quality. Additionally, storage of centrifuged sperm at a concentration of 250 × 10(6)/mL with 20% seminal plasma (v/v) did not have a detrimental effect on percentages of motile or progressively motile sperm, or sperm DNA quality.
Copyright © 2012 Elsevier Inc. All rights reserved.
Publication Date: 2011-12-20 PubMed ID: 22192395DOI: 10.1016/j.theriogenology.2011.10.031Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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The study investigates whether different techniques of cushioned centrifugation affect the quality of stallion sperm, concluding that varying the cushion-fluid volume and sperm number does not have any detrimental effect on the sperm quality, whether initially observed or after cooled storage.
Introduction and Purpose
- This research focused on understanding whether variations in cushioned centrifugation techniques affected stallion sperm quality. It aimed to determine the effect of changing the cushion-fluid volume and sperm number during centrifugation, as well as varying sperm concentration in extended semen post-centrifugation.
Methods
- Various centrifugation conditions were created for the experiments. For cushion-fluid volume, either one or 3.5 milliliters was used; regarding sperm number, either 1 billion or 3 billion counts were determined per centrifuge tube.
- After centrifugation, sperm pellets were dismissed, with the resulting sperm concentrations at 25 million/mL or 250 million/mL.
- The experiments carefully observed the sperm recovery rate, motion characteristics, plasmatic membrane integrity (SMI), and DNA quality (COMPαt).
- The researchers analyzed these factors post-centrifugation (Time 0 h) and after 24 hours into cooled storage (Time 24 h) and compared them to control groups that weren’t subjected to centrifugation.
Results
- Across all differing centrifugation conditions, the sperm recovery rate was unaffected.
- At Time 0 h, metrics such as total sperm motility, progressive sperm motility, and track straightness were similar across different cushion-fluid volumes.
- Even after 24 hours of cooled storage, experimental endpoints remained similar across different centrifugation conditions.
- Five of the six variables tested produced similar outcomes between centrifugation and controls.
- Sperm storage concentrations of 25×10(6) and 250×10(6)/mL produced comparable results through various measures.
- However, the storage concentration of 250×10(6) sperm/ml presented higher curvilinear velocity values and lower straightness values than other groups.
Conclusion
- The study concluded that centrifugation techniques using a cushion-fluid volume as low as 1 ml and a sperm number up to 3×10(9) in 50-ml conical-bottom centrifuge tubes did not adversely impact the quality of stallion sperm, neither immediately post-centrifugation nor after 24 hours of cooled storage.
- Moreover, keeping the centrifuged sperm concentrated at 250 × 10(6)/mL with 20% seminal plasma didn’t have any detrimental effects on the sperm’s motility, progressive motility, or DNA quality.
Cite This Article
APA
Bliss SB, Voge JL, Hayden SS, Teague SR, Brinsko SP, Love CC, Blanchard TL, Varner DD.
(2011).
The impact of cushioned centrifugation protocols on semen quality of stallions.
Theriogenology, 77(6), 1232-1239.
https://doi.org/10.1016/j.theriogenology.2011.10.031 Publication
Researcher Affiliations
- Department of Large Animal Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, Texas, USA.
MeSH Terms
- Animals
- Centrifugation / methods
- Centrifugation / veterinary
- Horses
- Male
- Semen Analysis / veterinary
- Spermatozoa / physiology
- Spermatozoa / ultrastructure
Citations
This article has been cited 6 times.- Monaco D, Lacalandra GM, Ansar Z, Trerotoli P, Mulligan BP, Osman TK. The Effect of Cushioned Centrifugation, with and without Enzymatic Reduction of Viscosity, on the Motility Pattern and Kinematic Parameters of Dromedary Camel Bull Spermatozoa. Animals (Basel) 2023 Aug 22;13(17).
- Podico G, Spencer KM, Magalhaes HB, Canisso IF. Semen Quality of the First and Second Ejaculates Collected from Breeding Inactive Stallions after Cooling and Freezing. Vet Sci 2023 Feb 21;10(3).
- Lago-Alvarez Y, Podico G, Segabinazzi LG, Cunha LL, Barbosa L, Arnold CE, Lima FS, King LT, McLean AK, Canisso IF. Donkey Epididymal Transport for Semen Cooling and Freezing. Animals (Basel) 2020 Nov 25;10(12).
- Alamaary MS, Haron AW, Ali M, Hiew MWH, Adamu L, Peter ID. Effects of four extenders on the quality of frozen semen in Arabian stallions. Vet World 2019 Jan;12(1):34-40.
- Gloria A, Carluccio A, Wegher L, Robbe D, Befacchia G, Contri A. Single and double layer centrifugation improve the quality of cryopreserved bovine sperm from poor quality ejaculates. J Anim Sci Biotechnol 2016;7:30.
- Sun W, Guan Q, Wen J, Zhang Q, Yang W, Zhang B, Cui W, Zou Z, Yu Y. Calcium- and integrin-binding protein-1 is down-regulated in the sperm of patients with oligoasthenozoospermia : CIB1 expression in patients with oligoasthenozoospermia. J Assist Reprod Genet 2014 May;31(5):541-7.
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