Changes in Hyaluronidase, Acrosin, and N-Acetylhexosaminidase Activities of Dog Sperm after Incubation.

Abstract

ABSTRACT. Hyaluronidase, acrosin and N-acetylhexosaminidase activities were examined in sperm collected from 12 beagle dogs and in culture medium after 0.5 hr and 7 hr of sperm incubation. The activities of the three enzymes were significantly higher at 7 hr than at 0.5 hr (P<0.05, 0.01), and the increases were associated with sperm capacitation. It was considered that the three enzymes in the dog sperm are related to fertilization by reason of the findings of the release of these enzymes from the sperm into the medium after 7 hr of incubation.-KEY WORDS: canine sperm, capacitation, enzyme.J. Vet. Med. Sci. 61(2): 183-184, 1999 The mean (±S.E.) values of %HA and %AR after 7 hr of sperm incubation were 56.0 ± 4.6% and 28.3 ± 1%, respectively (Fig. 1). The activities of HR, AS and AH were significantly higher after 7 hr of incubation than after 0.5 hr (P<0.05, 0.01) ( Table 1). After 7 hr of incubation all three enzyme activities were detected in the medium following removal of the sperm ( [14] are required for sperm to penetrate through the oocyte cumulus cell matrix, and these two enzymes are assumed to be required for penetration of the zona pellucida [2,3], in addition to AS. AS activity in non-capacitated stallion sperm has been reported to be very low after ejaculation, but its activity is very high in capacitated sperm after incubation [2]. In the present study, the activities of HR, AS and AH in dog sperm increased markedly after 7 hr of incubation, compared with 0.5 hr, and the percentages of capacitated sperm after 7 hr of incubation were significantly higher than after 0.5 hr. Thus it was concluded that the three enzyme activities increased in association with sperm capacitation.HR, AS and AH have been found to be released from the head of hamster [14] and human [13] sperm not only during the acrosome reaction but before it. The results of the present study do not demonstrate the release of the three enzymes from acrosome-intact sperm in the dog. However, release of these enzymes from dog sperm into the medium was observed. The acrosome reaction of dog sperm has Table 1. Changes in hyaluronidase, acrosin and N-acetylhexosaminidase activities (mean ± S.E.) in spermatozoa collected from 12 beagle dogs and in canine capacitation medium (CCM) after 0.5 and 7 hr incubation Hyaluronidase Acrosin N-acetylhexosaminidase (p mol/min/mg or ml) (n mol/min/mg or ml) (p mol/min/mg or ml) 0.5 hr 7 hr 0.5 hr 7 hr 0.5 hr 7 hr It is well known that hyaluronidase (HR) [1,3] and acrosin (AS) [4,9] located within sperm acrosomes are very important enzymes for fertilization, and it has also been reported that N-acetylhexosaminidase (AH) in sperm is associated with penetration of the oocyte [2,14]. However, since there have been few reports on these enzymes in dog sperm [5], the authors examined the activities of HR, AS and AH in incubated dog sperm and in the culture medium.The quality of the semen collected by digital manipulation from 12 normal beagle dogs, age of 2-6 years, was evaluated, and the sperm were washed wit...

Acrosin release and acrosin activity during incubation in capacitating media using fresh and frozen-thawed dog sperm

“…However, fresh spermatozoa retained their acrosin activity over time, since the presence and the sizes of gelatin halos increased throughout incubation periods. This increasing gelatin digestion observed in fresh samples agrees well with the results of Kawakami et al, (1999), who concluded that acrosin activity in fresh dog sperm, measured with light spectrophotometry, increased in association with sperm capacitation.…”

Influence of Canine Cumulus Oophorus on Homologous Sperm Motility.

“…Since almost all canine sperm that penetrate cumuli have an acrosome cap, acrosome-intact dog sperm, but not acrosome-reacted sperm, may be able to penetrate the cumuli as in hamster sperm [37] and pig sperm [22]. The authors have reported that canine sperm release hyaluronidase, an enzyme that hydrolyzes the hyaluronic acid component of the cumulus intercellular matrix [18], and the acrosome reaction of canine sperm is induced by homologous ZP [20], the same as in mouse [12], hamster [27], pig [4] and bull [23]. In the present study, it was shown that hyperactivated movement of canine sperm is induced by homogenated cumuli, as in hamster sperm [13].…”

“…These results suggest that canine hyperactivated sperm with intact acrosome can penetrate homologous cumuli and that the sperm are able to pass through the cumulus because the hyperactivated movement is maintained by hyaluronic acid secreted by the cumulus cells. Therefore, the hyperactivated movement of sperm that have penetrated a cumulus may be maintained by hyaluronic acid, which is also the main component of the cumulus intercellular matrix in many species [1,10,25,28].Sperm penetration into cumuli in many species [24] is made possible by the release of hyaluronidase, an enzyme that hydrolyzes hyaluronic acid, from the sperm head, and the authors have reported that canine sperm also release hyaluronidase [18]. It has been discovered that hamster acrosome-reacted sperm can not penetrate cumuli [5,7] and that the acrosome reaction is induced by homologous zona pellucida (ZP) [27], but there have been no reports on detailed investigation of the relationship between canine sperm and homologous cumuli during fertilization.…”

“…Sperm penetration into cumuli in many species [24] is made possible by the release of hyaluronidase, an enzyme that hydrolyzes hyaluronic acid, from the sperm head, and the authors have reported that canine sperm also release hyaluronidase [18]. It has been discovered that hamster acrosome-reacted sperm can not penetrate cumuli [5,7] and that the acrosome reaction is induced by homologous zona pellucida (ZP) [27], but there have been no reports on detailed investigation of the relationship between canine sperm and homologous cumuli during fertilization.…”

Induction of Dog Sperm Capacitation by Glycosaminoglycans and Glycosaminoglycan Amounts of Oviductal and Uterine Fluids in Bitches.

“…The authors had previously found that hyaluronidase and Nacetylhexosaminidase, the enzymes which hydrolyze GAGs, are present in dog sperm and that they are released during capacitation [14]. Thus, the GAGs in the fluid secreted by the genital organs of bitches may serve to accelerate the capacitation of dog sperm.…”