1 American Hospital, Istanbul, Turkey, 2 Reproductive Medicine, SU/Sahlgrenska, Gothenburg and 3 NidaCon International, Gothenburg, Sweden
4 To whom correspondence should be addressed. e-mail: paul{at}nidacon.com
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Abstract |
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Key words: hyaluronate/ICSI/PVP/slowing sperm motility
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Introduction |
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Previously, the only products commercially available for slowing sperm motility contained a synthetic plastic, polyvinylpyrrolidone (PVP). However, some PVP is injected into the oocyte along with the sperm and, as the PVP cannot diffuse out and is not digestible by lysosomal enzymes, it will remain in the oocyte for a prolonged period (Jean et al., 2001). To avoid any potential damaging effects of PVP, techniques have been developed which do not necessitate slowing sperm motility (Harari et al., 1995
; Jean et al., 1996
; 1997; Butler and Masson, 1997
; Hlinka et al, 1998
; Tsai et al., 2000
). However, not everybody is able to crush the tails of sperm moving at their normal velocity.
Therefore, a physiological alternative to PVP has been sought for reducing sperm motility to facilitate capturing sperm for ICSI. Ideally the product should possess the following characteristics: (i) be sufficiently viscous, to slow sperm motility enough to aspirate them into an ICSI pipette; (ii) be sufficiently fluid, to facilitate ease of aspirating and dispensing the sperm and a little of the liquid into, and from, the pipette; (iii) be able to prevent sperm sticking to either the plastic culture dish or the glass ICSI pipette; and (iv) have no deleterious effects on post-ICSI zygote development.
This study was designed to evaluate SpermCatchTM (NidaCon International, Gothenburg, Sweden), a viscous liquid containing hyaluronate and human serum albumin, as a potential substitute for PVP. Since hyaluronate and human serum albumin are found naturally in the mammalian reproductive tract, this product may prove to be a physiological alternative to PVP which can be used in the clinic to reduce sperm motility. SpermCatch is manufactured according to current Good Manufacturing Practice (cGMP) by NidaCon International AB, an ISO-registered company, and is approved by medical authorities.
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Materials and methods |
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The sperm were slowed by exposure to SpermCatch, immobilized by nicking their tail and drawn into an ICSI pipette. The ease with which the sperm could be caught, and drawn into, or expelled from, the pipette was assessed subjectively by workers accustomed to using PVP.
In a separate study, oocytes were injected with sperm exposed to either the hyaluronate or the PVP-containing product. The oocytes were from different patients, and were randomly assigned to the media that was used to slow sperm motility. Post-ICSI rates of fertilization and cleavage were compared between the two groups, and embryos were graded according to the following classification; G1 embryos: embryos with no fragmentation and equal sized homogeneous blastomeres; G2 embryos: embryos with <20% fragmentation and/or equal sized blastomeres; G3 embryos: embryos with 2050% fragmentation and/or unequal sized or granular necrotic blastomeres; G4 embryos: embryos with >50% fragmentation.
All the sperm samples in the ICSI study came from patients with male factor infertility.
Mean velocities for each group were compared using ANOVA (Campbell, 1974). Significance was accepted where P < 0.05.
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Results |
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CASA was performed on sperm exposed to either the hyaluronate-containing medium or the PVP product (ICSI-100). These sperm had mean curvilinear velocities, mean velocity of the average path and mean straight line velocities which were significantly slower (P < 0.001 for all parameters) than those of control sperm diluted in sperm maintenance medium (Table I).
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Discussion |
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Some embryologists, concerned about the potential adverse effects of PVP, have developed modifed techniques to capture sperm without slowing sperm motility (Harari et al., 1995; Hlinka et al., 1998
; Tsai et al., 2000
). Hlinka et al. (1998
) attached the sperm to the end of the injection needle by suction before crushing the tail. They claim that even fast-moving sperm can be caught by this method, with the added advantage that since the sperm is never aspirated into the needle, a minimal volume of fluid is expelled during sperm deposition in the oocyte (Hlinka et al., 1998
). Others have reported crushing the sperm tail before aspirating the sperm into the tip of the ICSI pipette (Harari et al., 1995
; Tsai et al., 2000
). Jean et al. (2001
) noted that training was sufficient to overcome any initial difficulties experienced in catching the sperm in the absence of PVP.
Although avoiding the use of exogenous substances altogether is probably the most desirable situation, sperm are not easy to tail-nick or catch without slowing their motility. Therefore, a physiological alternative to PVP is required. The new product, SpermCatch, which contains sodium hyaluronate, slowed sperm sufficiently for them to be caught, was easy to handle in the ICSI pipette and prevented sperm from sticking to the culture dish or to the pipette. Although sperm motility was somewhat faster in the hyaluronate-containing product than in the PVP-containing product, which necessitated some slight adaptations to the handling techniques on the part of the operator, it was not difficult to learn to use this product. Furthermore, since there was no difference in zygote development after ICSI, it would appear that the hyaluronate-containing product could be used in preference to PVP-containing products without detriment to the patients chances of becoming pregnant.
The observation that hyaluronic acid can modulate sperm motility is not new; for example, Zimmerman et al. (1994) reported that exposure of sperm to hyaluronic acid at 1 mg/ml caused a decrease in straight line velocity, amplitude of lateral head deviation and mean angular deviation compared with non-exposed sperm. Moreover, the suggestion that hyaluronic acid could be used to slow sperm motility prior to ICSI was advocated by Barak et al. (1999; 2001
). These authors reported similar fertilization rates and pregnancy rates after ICSI with sperm exposed to either hyaluronic acid or PVP and recommended the use of hyaluronic acid as a physiological replacement for PVP.
The results reported here concur with those of Barak et al. (1999; 2001), in that sperm motility was effectively slowed by the hyaluronate-containing product, and zygote development after ICSI was not impaired. Furthermore, the hyaluronate-containing product was easy to handle in the ICSI pipette and prevented sperm from sticking to the culture dish or to the pipette. Therefore, it is concluded that SpermCatch represents an effective, physiological alternative to PVP for modulating sperm motility prior to aspirating a single sperm into an ICSI pipette.
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FOOTNOTES |
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References |
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