Routine chemicals and reagents were obtained from Nakarai Chemical Co. Section snippets Media and seminal plasma To evaluate this freezing and thawing method, we analyzed frozen-thawed sperm motility, capacitation and acrosomal status in vitro and fertilization competence in vitro and in vivo. Second, the sperm was frozen using a conventional method, and the cryopreserved sperm was thawed in a medium containing seminal plasma collected from a boar with proven high reproductive performance. First, seminal plasma was removed from the sperm by centrifugation immediately after semen collection. Consequently, we designed the following novel freezing and thawing method to preserve sperm from boars exhibiting poor sperm freezability. Thus, we hypothesized that the presence of seminal plasma during freezing is not a critical determinant of post-thaw sperm quality in boars with good freezability, but has deleterious effects in those with poor freezability, and that seminal plasma is essential to maintain fertility competence during thawing. Additionally, it has been reported that seminal plasma is required to protect sperm against a spontaneous capacitation-like reaction during the thawing process. However, frozen-thawed epididymal porcine sperm does not have high in vivo fertilization competence. Removing seminal plasma immediately after semen collection is one of the available techniques to improve motility of frozen-thawed sperm and in vitro fertility competence in miniature pigs. Although the poor freezability of boar sperm is potentially affected by genetic background, we speculated that the effects of seminal plasma on sperm after ejaculation may be another factor. In addition, the freezability of epididymal sperm is higher than that of ejaculated sperm in boars. In a study in which semen was collected as two portions, the sperm-rich fraction and the post sperm rich fraction, in several boars there were significant differences between fractions in post-thaw sperm motility. Additionally, they identified molecular markers linked to genes controlling semen freezability by amplified restriction fragment length polymorphism technology, suggesting that the freezability of boar sperm was affected by genetic factors. categorized sperm head morphology by Fourier descriptors, and detected a significant association between the head morphology of fresh spermatozoa and the motility of sperm after freezing and thawing. However, it is well known that the quality of frozen-thawed spermatozoa and the conception success rate are also dependent on unique characteristics of individual boars,. Recently, we and other groups reported a high conception rate (70–80%) by AI using boar spermatozoa cryopreserved using a modification of previously described cryopreservation methods and through the development of a novel sperm infusion method. In conclusion, our novel cryopreservation and thawing method increased the success of AI with frozen-thawed porcine semen, particularly from boars with poor post-thaw semen quality.Ĭryopreservation of boar spermatozoa offers an effective means of long-term storage of important genetic material. 18.4 ± 2.8%, P < 0.01) and expression of a 15 kDa tyrosine-phosphorylated protein (capacitation marker) in thawed sperm relative to controls the addition of 10% (v/v) seminal plasma to the thawing solution significantly suppressed both changes and increased conception rate to AI (70% vs. Freezing sperm without seminal plasma increased both loss of the acrosome cap (37.5 ± 1.6% vs. Removal of seminal plasma did not significantly affect post-thaw sperm motility in good freezability boars however, in boars with poor freezability, it increased post-thaw motility relative to control sperm cooled with seminal plasma (64.5 ± 3.4% vs. To determine the effects of seminal plasma during and after cyopreservation on post-thaw sperm functions in semen from poor freezability boars, seminal plasma was removed immediately after collection, and sperm was subjected to cooling and freezing.
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