Enzymatic treatment of spermatozoa with a trypsin solution, SpermSolute: improved motility and enhanced ATP concentration

Figenschau, Yngve; Bertheussen, Kjell

doi:10.1046/j.1365-2605.1999.00180.x

Cited by 9 publications

(3 citation statements)

References 8 publications

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“…Several other enzymes like α-chymotrypsin, lysozyme or hyaluronidase have also been described for this purpose. Recently, a special trypsin-based dissolving solution, SpermSolute, has been reported [297]. However, as these enzymes can also damage spermatozoa, special care should be taken to remove the enzymes by washing procedures as soon as liquefaction is completed.…”

Section: In Vitro Treatment Of Spermatozoamentioning

confidence: 99%

Sperm preparation for ART

Henkel

Schill

2003

Reprod Biol Endocrinol

415

175

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The onset of clinical assisted reproduction, a quarter of a century ago, required the isolation of motile spermatozoa. As the indication of assisted reproduction shifted from mere gynaecological indications to andrological indications during the years, this urged andrological research to understand the physiology of male germ cell better and develop more sophisticated techniques to separate functional spermatozoa from those that are immotile, have poor morphology or are not capable to fertilize oocytes. Initially, starting from simple washing of spermatozoa, separation techniques, based on different principles like migration, filtration or density gradient centrifugation evolved. The most simple and cheapest is the conventional swim-up procedure. A more sophisticated and most gentle migration method is migration-sedimentation. However, its yield is relatively small and the technique is therefore normally only limited to ejaculates with a high number of motile spermatozoa. Recently, however, the method was also successfully used to isolate spermatozoa for intracytoplasmic sperm injection (ICSI). Sperm separation methods that yield a higher number of motile spermatozoa are glass wool filtration or density gradient centrifugation with different media. Since Percoll® as a density medium was removed from the market in 1996 for clinical use in the human because of its risk of contamination with endotoxins, other media like IxaPrep®, Nycodenz, SilSelect®, PureSperm® or Isolate® were developed in order to replace Percoll®. Today, an array of different methods is available and the selection depends on the quality of the ejaculates, which also includes production of reactive oxygen species (ROS) by spermatozoa and leukocytes. Ejaculates with ROS production should not be separated by means of conventional swim-up, as this can severely damage the spermatozoa. In order to protect the male germ cells from the influence of ROS and to stimulate their motility to increase the yield, a number of substances can be added to the ejaculate or the separation medium. Caffeine, pentoxifylline and 2-deoxyadenosine are substances that were used to stimulate motility. Recent approaches to stimulate spermatozoa include bicarbonate, metal chelators or platelet-activating factor (PAF). While the use of PAF already resulted in pregnancies in intrauterine insemination, the suitability of the other substances for the clinical use still needs to be tested. Finally, the isolation of functional spermatozoa from highly viscous ejaculates is a special challenge and can be performed enzymatically to liquefy the ejaculate. The older method, by which the ejaculate is forcefully aspirated through a narrow-gauge needle, should be abandoned as it can severely damage spermatozoa, thus resulting in immotile sperm.

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Section: In Vitro Treatment Of Spermatozoamentioning

confidence: 99%

Sperm preparation for ART

Henkel

Schill

2003

Reprod Biol Endocrinol

415

175

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“…, 1994; Lanzafame et al. , 1994; Minhas & Ripps, 1996; Figenschau & Bertheussen, 1999; Terriou et al. , 2000).…”

Section: Introductionmentioning

confidence: 99%

Phosphatidylinositol 3‐kinase inhibition enhances human sperm motility and sperm–zona pellucida binding

et al. 2004

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Various signalling pathways are involved in the regulation of sperm motility, capacitation, acrosome reaction and sperm-zona binding. Recent data pointed out an important role for phosphatidylinositol 3-kinase (PI3K) in human sperm motility. However, no study as of yet has been carried out to determine the effect of sperm treatment with the PI3K inhibitor LY294002 on other sperm parameters. In the present study, we investigated the role of PI3K on human sperm motility, acrosome reaction and sperm-oocyte binding by using this inhibitor. We demonstrate that in vitro incubation of washed unselected spermatozoa with LY294002 increased the percentage motility and progressive motility in asthenozoospermia patients as evaluated by computer-aided sperm analysis. The compound furthermore did not influence the acrosome reaction, whilst it (further) slightly enhanced sperm-oocyte binding. Our results therefore imply that PI3K negatively affects sperm motility and oocyte binding and might suggest a possible therapeutic role for PI3K inhibitors in the treatment regime for asthenozoospermia.

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“…Loskutoff et al (2005) reported that DGC of semen with trypsin added to density layers had no detrimental effects on sperm motility or viability [14]. Furthermore, trypsin treatment of human sperm, prior to swim up, resulted in increased motility and adenosine triphosphate (ATP) concentration [37]. Westhoff and Kamp (1997) suggested that the increased ATP concentration could be attributed to the activation of the glycolytic enzyme glyceraldehyde triphosphate dehydrogenase (GADPH) by trypsin [38], explaining the improved motility.…”

Section: Discussionmentioning

confidence: 99%

Treatment of human sperm with serine protease during density gradient centrifugation

Fourie

Loskutoff²,

Huyser

2012

J Assist Reprod Genet

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Purpose Seminal pathogens can bind specifically or nonspecifically to spermatozoa, rendering semen decontamination procedures ineffective, whereby vertical or horizontal transmission of the infection could occur. Serine proteases have been demonstrated to effectively inactivate viruses and to break pathogen-sperm bonds. However, the addition of a protease to density gradient layers during semen processing could negatively impact on sperm parameters. This study investigated the effect of the addition of a recombinant, human-sequence protease (rhProtease) on sperm parameters during density gradient centrifugation. Methods (i) Pooled semen samples (n09) were split and processed by density gradient centrifugation, with the top density layers supplemented, or non-supplemented with rhProtease at three different concentrations (diluted 2, 10 and 20 times). Sperm parameters were then analysed by flow cytometry and computer-assisted semen analyses.(ii) Semen samples (n05) were split and similarly processed using PureSperm® Pro, with rhProtease in the 40 % density gradient layer, or standard PureSperm® not supplemented with rhProtease (Nidacon, International) respectively. The Hemizona assay was then utilized to compare sperm-zona binding post processing.Results Evaluation of sperm parameters indicated that rhProtease did not, at any of the tested concentrations, have an impact on (i) mitochondrial membrane potential, vitality, motility, or (ii) zona binding potential. Conclusion We report that the addition of rhProtease to density gradients is a non-detrimental approach that could improve the effectiveness of semen processing for the elimination of seminal pathogens, and benefit assisted reproduction outcome.

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Enzymatic treatment of spermatozoa with a trypsin solution, SpermSolute: improved motility and enhanced ATP concentration

Cited by 9 publications

References 8 publications

Sperm preparation for ART

Sperm preparation for ART

Phosphatidylinositol 3‐kinase inhibition enhances human sperm motility and sperm–zona pellucida binding

Treatment of human sperm with serine protease during density gradient centrifugation

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