Effect of acrosome reaction progress in frozen-thawed boar spermatozoa on the efficiency of in vitro oocyte fertilization

Abstract

ABSTRACT:A good functional status of cryopreserved boar spermatozoa is very important for successful fertilization of porcine oocytes and in vitro embryo production. The purpose of the study was to evaluate the changes in functional status of boar spermatozoa separated from frozen-thawed semen and capacitated in vitro by caffeine. The effect of acrosome reaction development in spermatozoa on the efficiency of oocyte fertilization has been studied in boars A, B and C. Motile spermatozoa were separated by Percoll gradient, untreated (control) or treated with both 1mM and 2mM caffeine, and capacitated or co-cultured with matured oocytes. The motility, viability, chromatin and acrosome integrity, and fertilizing ability of spermatozoa were assessed. The separation significantly increased (P < 0.05) the percentage of viable spermatozoa in all tested boars and percentages of motile and acrosome intact spermatozoa in boars B and C. The capacitation significantly decreased (P < 0.05) the percentages of viable and motile spermatozoa, but after capacitation, the motility and viability were significantly higher (P < 0.05) for the caffeine-treated spermatozoa than for the untreated controls. A fall in the proportion of acrosome-intact spermatozoa was different for each caffeine concentration and each boar, but in all boars, acrosome reaction progress was faster and, similarly, monospermy and the total efficiency of fertilization were significantly higher (P < 0.01) for the spermatozoa treated with 1mM caffeine than for those treated with 2mM caffeine. It can be concluded that there is a potential relationship between the acrosome reaction progress in frozen-thawed boar spermatozoa and the efficiency of fertilization of porcine oocytes. A faster AR induced in spermatozoa by appropriate caffeine treatment resulted in a higher monospermy rate and total efficiency of fertilization. Thus, it is important to test sires before their semen is used for in vitro embryo production. The faster AR induced by 1mM caffeine was more effective in terms of monospermy and total efficiency of fertilization.

Centrifugation on Percoll density gradient enhances motility, membrane integrity andin vitrofertilizing ability of frozen–thawed boar sperm

“…It has been shown that the sperm plasma membrane after cryopreservation is injured and capacitation and/or acrosome reaction is enhanced due to elevated intracellular calcium levels of frozen–thawed sperm (Maxwell & Johnson, 1997; Bailey et al ., 2000). Here, the Percoll separation method enhanced recovery rate of sperm with intact plasma and acrosome membranes and high motility from frozen–thawed boar semen, as reported previously (Martecikova et al ., 2010). However, it has also been reported that there was no significant difference in the percentage of sperm with acrosomes that reacted in humans (Drobnis et al ., 1991), but was loss of acrosomes in bulls (Suzuki et al ., 2003) and boars (Grant et al ., 1994) between Percoll-separated and centrifuged semen.…”

“…Percoll® (a colloidal suspension of polyvinyl pyrrolidone-coated silica particles) density gradients are widely used for the selection of sperm with superior quality by centrifugation from man (Drobnis et al ., 1991), bull (Suzuki et al ., 2003; Oliveira et al ., 2011), boar (Grant et al ., 1994; Suzuki & Nagai, 2003; Martecikova et al ., 2010; Matás et al ., 2011) and ram (Valcárcel et al ., 1996) semen. In the pig, sperm with high motility (Grant et al ., 1994; Martecikova et al ., 2010; Matás et al ., 2011) and intact plasma and acrosomal membranes (Martecikova et al ., 2010) were separated by Percoll gradient centrifugation from fresh or frozen–thawed semen. Furthermore, rates of penetration (Matás et al ., 2011), cleavage (Grant et al ., 1994; Jeong & Yang, 2001) and blastocyst formation (Jeong & Yang, 2001) after IVF and embryo culture were improved by insemination with boar sperm prepared by Percoll gradient centrifugation.…”

Sperm Nuclei Analysis and Nuclear Organization of a Fertile Boar–Pig Hybrid by 2D FISH on Both Total and Motile Sperm Fractions

“…Percoll separation methods (colloidal suspension of polyvinyl pyrrolidone coated silica particles) have been broadly used for in vitro fertilization (IVF) and have been reported to have a positive correlation with sperm quality (induced by motility, viability, normal morphology and fertility) [ 33 , 34 ]. In pigs, spermatozoa separated by Percoll gradient from fresh or frozen–thawed semen presented high motility, intact plasma and acrosomal membranes [ 35 , 36 ]. In this study, the MSF was successfully separated through the dual-layer (75–60%) discontinuous Percoll gradient, showing a significant reduction in the DNA damage considering the %DFI variables, as demonstrated by Larson et al [ 37 ].…”

The Production of Freeze-Dried Egg Yolk Powder and Its Effect on the Quality of Garut Ram Liquid Semen

“…IA spermatozoa was done by diluting the semen using formolsaline solution ,which functions to fix the spermatozoa, and was made into a peripheral smear and dried on a bunsen and stained with 5% (v/v) aniline blue and 0.5% (v/v) crystal violet for 15 seconds (Martecikova et al, 2010). Spermatozoa that still have intact acrosomal were marked with 1/2 to 2/3 of the anterior part of the head darker than the posterior part.…”