Abstract
BackgroundDensity gradient centrifugation was reported as a technique of semen preparation in assisted reproductive techniques in humans and animals. This technique was found to be efficient in improving semen quality after harmful techniques such as cryopreservation. Recently a modified technique, single layer centrifugation, was proposed as a technique providing a large amount of high quality spermatozoa, and this treatment was performed before conservation. Single layer centrifugation has been studied prevalently in stallions and in boars, but limited data were available for bulls. Occasionally bulls are known to experience a transient reduction in semen quality, thus techniques that allow improvement in semen quality could be applied in this context. The aim of this study was the evaluation of single layer and double layer centrifugation by the use of iodixanol, compared with conventional centrifugation and non-centrifuged semen, on the sperm characteristics during the cryopreservation process in bulls with normal and poor semen quality.ResultsSingle layer centrifugation and double layer centrifugation both significantly increased the percentage of normal spermatozoa and decreased the percentage of non-sperm cells in poor quality samples, while both were ineffective in those of normal quality. Sperm characteristics in poor quality samples increased after single layer centrifugation and double layer centrifugation, reaching values similar to those recorded in normal samples, and this trend is maintained after equilibration and after cryopreservation. On the other hand, SLC and DLC resulted in a consistent reduction in the spermatozoa recovered, and this resulted in a reduction of the absolute amount of spermatozoa cryopreserved in the normal samples, without a clear improvement in sperm characteristics in this type of sample.ConclusionsThese data suggested that both SLC and DLC could be performed in practice, but their application should be limited to the cases in which the quality of the spermatozoa recovered is more important than the total amount of spermatozoa.
Highlights
Density gradient centrifugation was reported as a technique of semen preparation in assisted reproductive techniques in humans and animals
Samples (1 mL) were stained with 2.4 μmol of propidium iodide (PI) and 5 μg/mL of FITC-conjugated agglutinin derived from pisum sativum (FITC-PSA)
Sperm total and progressive motility, sperm velocities, and sperm morphology were significantly different in samples with normal and poor semen quality (P ≤ 0.05)
Summary
Density gradient centrifugation was reported as a technique of semen preparation in assisted reproductive techniques in humans and animals. Artificial insemination (AI) is generally considered to be the technique that has contributed the most to the improvement of animal reproduction This technique can result in pregnancies only if the insemination dose contains sufficient viable spermatozoa capable of reaching the site of fertilization [1]. Density gradient centrifugation through colloids is one of the procedures recommended by the World Health Organization for the preparation of human spermatozoa used in assisted reproduction [6] In animals, this method has been suggested as a potential means of improving the quality of sperm after cryopreservation [7,8,9]. A modified gradient centrifugation technique, single layer centrifugation (SLC), was proposed to improve the semen quality before conservation and AI [1, 10]
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