The effect of swim-up purification and incubation of cells on sperm viability in dogs of different ages

Abstract

ABSTRACT:The influence of selected semen extenders on the motility of frozen-thawed dog spermatozoa has been clearly demonstrated in several studies, although there are no reports indicating the effect of swim-up purification on sperm viability in this species of mammals. Therefore, this study was aimed at investigating phosphatidylserine (PS) externalization and necrosis in sperm after variable lengths of time of in vitro incubation after swim-up purification. Dog semen samples were collected from (i) ten dogs aged six months to 1.5 year, (ii) ten dogs aged six to eight years, and (iii) ten dogs aged 11 to 13 years. A flow cytometric method was employed to evaluate dog sperm viability in animals of different age groups after employment of a swim-up (SU) purification technique. After SU spermatozoa were incubated for 15, 30, 45 and 60 min in Sperm-TALP medium. We observed an increase in the number of viable sperm (double negatives) after 15 min of incubation compared to sperm undergoing PS externalization and late necrotic sperms (P < 0.001) in each group of dogs. We also found a higher number of early necrotic sperm after 60 min of in vitro incubation (P < 0.001). The amounts of late necrotic sperm and cells with PS externalization were similar among animals of different age groups. We show for the first time that most viable sperm are recovered after an in vitro incubation step of 15 min (control samples in this study) because as the time of incubation increases so does the number of degenerated or damaged cells. The higher number of early necrotic cells after 60 min of in vitro incubation may be a special feature of this species and may result from the induction of necrosis in the sperm. This knowledge may be used in future experiments for the preparation of spermatozoa following in vitro fertilization in dogs.

Differences in preservation of canine chilled semen using simple sperm washing, single-layer centrifugation and modified swim-up preparation techniques

“…In the last decade, a wide variety of sperm selection methods has been developed, which vary greatly in the recovery rate and sperm quality (motility, morphology, chromatin integrity, viability and acrosome integrity) of the samples obtained (Morrell and Rodriguez-Martinez 2011). The techniques that have been used to prepare canine spermatozoa for assisted reproductive techniques (ART) include sperm washing (Rijsselaere et al 2002), swim-up (Bukowska et al 2011;Sa ´nchez et al 2011), density-gradient centrifugation (Kim et al 2010;Dorado et al 2011aDorado et al , 2011bPhillips et al 2012), single-layer centrifugation (SLC; Morrell et al 2008;Dorado et al 2013aDorado et al , 2013bMorrell et al 2013) and glass-wool filtration (Kim et al 2010).…”

“…The sperm migration or swim-up (SU) method is commonly used for the isolation of highly-motile spermatozoa from the ejaculate (Rodriguez-Martinez et al 1997) and can be used for selecting dog spermatozoa to be used in sperm vitrification (Sa ´nchez et al 2011). It has also been reported that the sperm viability in fresh dog semen samples is improved by the SU technique (Bukowska et al 2011). However, the standard SU procedure involves centrifugation steps that induce a decline in motility of spermatozoa (Alvarez et al 1993).…”

Theinfluence of acetyl-L-carnitine on the reproductive system of the Wistar male rats after electromagnetic exposure from mobile telephone

“…Из эпидидимиса выделяли сперматозоиды, количество которых подсчитывали в камере Горяева [18], определяли их жизнеспособность [19], индекс DFI (фрагментация ДНК) [20], число апоптотических и некротических сперматозоидов по [21] и содержание фруктозы в семенных пузырьках [22].…”

Some factors influencing canine sperm motility

“…Sperm motility is one of the most important parameters analyzed in a breeding soundness evaluation because it reflects the ability of sperm to migrate through the female reproductive tract (Bukowska et al, 2011). Sperm motility is also an essential parameter to consider when evaluating the efficiency of extenders, cryoprotectants, and freezing protocols (Rijsselaere et al, 2005).…”