Acrosin Activity Research Articles

Relation of human sperm acrosin activity and fertilization in vitro.

The aim of the study described here was to determine the possible contribution of the acrosin activity test to routine semen analysis in enhancing the precision of the prognosis of IVF success in a group of patients in which the contribution of the egg factor to infertility was ruled out (20 cases) compared to a control IVF group (39 cases). Semen analysis, acrosin activity and acrosome ultrastructure were determined for all semen samples. The group with high fertilization rates was comprised of normozoospermic patients while the group with low fertilization rates was comprised of astheno-teratozoospermic patients. The mean acrosin level of the positive IVF group was significantly higher than that of the negative group (51.7 +/- 33.2 and 28.6 +/- 13.7, respectively). Two parameters: per cent motile spermatozoa and acrosin level, were found to have a significant positive correlation with subsequent successful IVF (r = 0.36, P < 0.006; r = 0.37, P < 0.004, respectively); and agenesis of the acrosome was found to have a significant negative correlation (r = -0.33, P < 0.01). The ability of these parameters to correctly predict fertilization success was 59%, with 5% false positive, among which 15.4% was predicted solely by the acrosin level (above 54 microIU 10(6) cells-1) and 23% solely by per cent motile spermatozoa (above 50%). Abnormalities of the acrosome ultrastructure did not contribute further to the correct classification. The apparent clinical benefit of the acrosin level test is discussed.

Adenosine 5'-triphosphate (ATP) concentration and acrosin activity in human spermatozoa. Their relation with the sperm penetration assay (SPA).

Fertility of men depends on the quality of semen. The aim of the present paper is to determine both the acrosin activity by radioimmunoassay and ATP concentration by bioluminescence in human spermatozoa, and evaluate these results in those samples with normal or low sperm penetration according to SPA test. Ejaculates obtained from 42 untreated men, were studied one hour after the obtention. These materials were divided into two groups:20 human semen samples with "in vitro" potentiality to penetrate zone-free hamster ova, between 15% to 98% and 22 human semen with SPA test between 0% to 14%. When we compare the group with normal penetration response vs that group with low or absent penetration ones, a significant decrease of ATP and acrosin concentrations was observed (P less than 0.001). Nevertheless no significant difference was observed in relation with percentage of motility, volume (ml), sperm concentration (10(6)/ml), percent of quick progressive spermatozoa and number of gametes capable of migrating into the medium layer (10(6)), between the group with low or absent penetration test against that one with normal zona-free hamster egg test.

Adenosine 5 triphosphate (ATP) content and acrosin activity in polyzoospermic subjects.

The aim of this paper was to measure the concentration of spermatic ATP and acrosin in a group of patients with polyzoospermia, and to compare these values with those found in a group of normozoospermic men. The general characteristics of the semen, the ATP and the acrosin, were measured in a group of 46 men with polyzoospermia, who consulted in our clinic for sterility. The results were compared with those of 50 normozoospermic men in the control group. Statistically, no significant difference was found between the volume, motility and morphology of the samples in both groups; however, the concentration of spermatic ATP and acrosin decreased significantly in a group of polyzoospermic patients. This could be due to a functional defect of the spermatozoa in this group of patients.

Relationship of Sperm Acrosin Activity to Semen and Clinical Parameters in Infertile Patients/Die Beziehung der Akrosinaktivität zu verschiedenen Sperma- und klinischen Parametern bei infertilen Patienten

Acrosin activity (aa) was routinely measured in 189 infertile men who attended the hospital between March 1984 and January 1985. The evaluation of the male included semen analysis, microbial screening, postcoital testing (PCT), in vitro sperm penetration test (SPMT), and screening for anti-sperm antibodies in serum. The strongest positive correlations of aa were found for sperm motility, count, morphology, vitality and volume. In presence of a poor PCT result, aa was significantly reduced, even in patients with good motility in native semen. A similar result was obtained with the SPMT: reduced sperm density and poor sperm motility in the capillary after 2 hours were significantly correlated to reduced aa. Smokers exhibited lower aa in presence of normal sperm count and motility. In the infertile group aa was significantly lower than in the group of men whose spouses conceived. On the basis of our results, the routine determination of aa is not necessary, but it provides additional prognostic information in couples with unexplained infertility.

Untersuchungen zur Akrosin- und Humanspermatrypsininhibitor-aktivität im menschlichen Sperma

Report concerning the sperm examination of 825 patients of the Dept. for Andrology. The acrosin as well as the human sperma trypsin inhibitor activity were investigated. The results were related to sperm density. It was found that acrosin activity was significantly dependent on sperm density. For the human sperma trypsin inhibitor activity this relationship could not be established, although lower values were found by sperm deficiency and higher values by high sperm density.

Sperm acrosin: liberation from the acrosome and activity of the free proteinase in the presence of nonoxinol-9.

The effects of the non-ionic detergent nonoxinol-9 and a vaginal contraceptive containing nonoxinol-9 on the liberation of the cellular proteinase acrosin from human spermatozoa and on the activity of the solubilized proteinase were investigated. Detachment of acrosin from the acrosome was observed in the presence of 0.01 to 1% nonoxinol-9 as assessed by activity assays and the gelatin substrate film technique. Activity of the free proteinase was modulated by nonoxinol-9 in a biphasic manner, i.e. lower concentrations of the detergent (0.001 to 0.1%) stimulated the activity of acrosin up to 180%, while higher concentrations of the detergent (1%) resulted in an extensive inhibition of acrosin (30% residual activity). It is demonstrated that local intravaginal nonoxinol-9 concentrations of 1% equalling the post-coital concentration of the intravaginal suppository Patentex oval when properly applied will result in an effective inhibition of the liberated sperm proteinase acrosin thus preventing unspecific proteolysis of the vaginal epithelia.

Bovine epididymal sperm proacrosin-acrosin system: quantification and partial characterization

Several studies suggest that acrosin, an acrosomal trypsin-like serine proteinase, plays a role in fertilization. The enzyme is present in an enzymatically inactive precursor form, called proacrosin and is believed to be converted to the enzymatically active form(s) through one/multiple physiological event(s) prior to the sperm penetration of the zona pellucida. Although, the proacrosin-acrosin system of several species has been well documented, the study of the enzyme system in bovine caput and cauda epididymis (where the maturation of spermatozoa occurs) has not been characterized. The present study demonstrates the quantification and partial characterization of the proacrosin-acrosin proteinase system in unpurified acrosomal extracts of bovine caput and cauda epididymal sperm. Proacrosin activation followed the sigmoidal type of activation curve. Activation experiments demonstrate that almost 80-90% of this protein exists in zymogen (proacrosin) form either in ejaculated or caput and cauda epididymal spermatozoa. Time-course activation studies showed that the zymogen in isolated spermatozoa was completely converted to active non-zymogen form in 3 and 5 h after removal from the cauda and caput regions, respectively, at pH 8.0 at 25 degrees C. This conversion was markedly inhibited by calcium in a dose dependent manner and the inhibition was reversible. On the other hand, calcium has a stimulatory effect on the hydrolytic activity of acrosin. These studies reveal that the proacrosin-acrosin system can be identified in crude extracts of bull epididymal and ejaculated sperm.

Indikationen zur Akrosin-Bestimmung

Assessment of acrosin activity for evaluation of acrosomal functions and the fertilizing capacity of spermatozoa is a suitable method within the diagnostic workup of male sterility in selected cases. Indications for determination of acrosin activity are cases with teratozoospermia and polyzoospermia. In addition, demonstration of sufficient amounts of acrosin activity in idiopathic sterility or in cases with an unknown male sterility factor and before in vitro fertilization may exclude severe disturbances of the sperm acrosome.

Biochemical and genetic investigation of round-headed spermatozoa in infertile men including two brothers and their father.

Acrosin and the outer acrosomal membrane (OAM) were studied in the spermatozoa of 9 infertile patients who differed in the number of round-headed spermatozoa between 14 and 71% in their ejaculates. These sperm components were also investigated in two infertile brothers who exhibited exclusively round-headed spermatozoa in their ejaculates, and in their fertile father. It turned out that round-headed spermatozoa lack both acrosin and the OAM as studied by indirect immunofluorescent and immunoperoxidase staining technique, gelatinolysis tests and by acrosin activity measurements. The normally shaped spermatozoa of 6 of the 9 infertile patients were found to be positive for acrosin and the OAM as expected, but in the remaining three patients even these spermatozoa were abnormal; in one patient they were unstainable for acrosin and in two patients they were unstainable both for acrosin and the OAM. These results have been confirmed by studies with the gelatinolysis test. The father of the two brothers with exclusively acrosomeless spermatozoa had more than 94% of normally shaped spermatozoa. However, only 10% of these spermatozoa were acrosin positive and only 30% were positive for the OAM. On the basis of these results we postulate that the mode of inheritance of the round-headed spermatozoa syndrome is polygenic rather than monogenic as suggested by previous authors.

Acrosin and trypsin inhibitor activity in the human ejaculate following testosterone treatment in cases of autoagglutination of spermatozoa.

3 men showing autoagglutination of spermatozoa were given 250 mg depot injections of testosterone enanthate once or twice weekly for 6 weeks then 250 mg every 2 weeks once sperm density had been reduced for a total dosage of 2000-3500 mg. Acrosin activity fell along with sperm density and trypsin-inhibitor activity showed a concomitant rise. Autoagglutination disappeared. Some wives became pregnant under these conditions.

Human sperm acrosin activity with relation to semen parameters and acrosomal ultrastructure

It was suggested that although not related to the standard semen parameters the level of the acrosin enzyme system is related to the fertility potential in men. Recently a simple clinical assay for total acrosin level was recommended for routine semen analysis. The improved clinical assay was analysed on the freshly liquified semen of 198 Asthenoteratozoospermic men and compared with the routine semen parameters including biochemical data and the ultrastructure of the acrosome. Only the sum of the per cent of live, motile, and normal-shaped spermatozoa had positive significant and reasonably high correlation with the acrosin level (r = 0.382, P < 0.0001). Each characteristic exhibits significant but low (< 0.35) correlation. Similarly negative significant and reasonably high correlation was obtained between the acrosin level and the sum of the principle acrosomal malformations observed by TEM (r = 0.396, P < 0.0001) while lower negative correlation was found only with agenesis or loss of the acrosome. Acrosin levels below 8.1 microIU 10(-6) cells were obtained in 4 specimens with above 80% round-form associated with more than 95% of agenesis of the acrosome. The possible significance of the low correlation obtained between the acrosin levels and seminal plasma zinc levels, malformations in the acrosomal equatorial region, and the presence of white blood cells is also discussed. We concluded that the acrosin activity reflects an aspect of male fertility which is not diagnosed by the routine semen analysis or by the ultrastructure of the acrosome, and is therefore a useful diagnostic sperm parameter.

Low acrosin activity in polyzoospermia.

Compared to normozoospermic men total sperm acrosin activity is considerably decreased (-59,1%) in patients with polyzoospermia indicating a functional defect of the proteolytic potential of the sperm acrosome, which may be responsible for the reduced fertility observed in this group of patients.

Ram proacrosin. A simple method for isolation of proacrosin free of inhibitors, and proacrosin autoactivation studies.

Two partially purified proacrosin forms have been obtained from acid extracts of ram ejaculated spermatozoa by Sephadex G-100 column chromatography in 0.001 M HCl. The form obtained from the extracts of whole ejaculates with apparently higher molecular mass was free of any acrosin inhibitor and was used for autoactivation studies. The autoactivation followed a classical S-shaped activation curve and was a two-step process. The presence of calcium ions slowed down the autoactivation process and stabilized the active acrosin formed. The activity of latter rapidly decreased when the pH of the fully activated mixture was adjusted to pH 3. The activity was not restored by re-adjusting the pH to 8. A proacrosin with apparently lower molecular mass was isolated from the extract of washed spermatozoa. Both proacrosins, however, showed the same molecular mass Mr approximately 58 000 daltons, when examined by gel filtration in 0.1 M NaCl (pH 3).

Ascorbic Acid, Catalase and Chlorpromazine Reduce Cryopreservation-induced Damages to Crossbred Bull Spermatozoa

The present study evaluated the effectiveness of ascorbic acid, catalase, chlorpromazine and their combinations in reducing the cryodamages to crossbred bull (Bos taurus x Bos indicus) spermatozoa. A total of 32 ejaculates (eight each from four bulls) were diluted in Tris-citric acid-fructose-egg yolk-glycerol extender. Each ejaculate was split into six parts (five treatment and one control). Treatment groups included 10 mm ascorbic acid, 0.1 mm chlorpromazine, 200 IU/ml catalase, 10 mm ascorbic acid + 0.1 mm chlorpromazine or 200 IU/ml catalase + 0.1 mm chlorpromazine in the extender. Fluorescent probes (Fluorescein isothiocyanate--Pisum sativum agglutinin + Propidium iodide) were used for the assessment of spermatozoa viability and acrosomal status. The proportion of acrosome intact live (AIL), acrosome intact dead, acrosome reacted live and acrosome reacted dead sperm was assessed in fresh, equilibrated and frozen-thawed semen. The functional status of the sperm was assessed using hypo-osmotic sperm swelling test (HOSST). Activities of acrosin and hyaluronidase enzyme were also determined. Lipid peroxidation level was assayed based on the melonaldehyde (MDA) production. In cryopreserved semen, the values of AIL spermatozoa, HOSST response, hyaluronidase and acrosin activity were reduced by 53%, 47%, 34% and 54%, respectively from their initial values in fresh semen. However, MDA level was threefold higher in the frozen-thawed sperm compared with fresh sperm. Significant (p < 0.05) improvement in motility, viability, HOSST response, retention of hyaluonidase and acrosin and reduction in MDA was recorded in ascorbic acid, catalase, ascorbic acid + chlorpromazine and catalase + chlorpromazine incorporated groups. The percentage of AIL sperm was significantly (p < 0.05) higher in ascorbic acid, catalase and ascorbic acid + chlorpromazine incorporated groups compared with the control. Chlorpromazine alone did not improve the post-thaw semen quality but when combined with either ascorbic acid or catalase, improvement in semen quality was noticed. It was inferred that incorporation of ascorbic acid, catalase and ascorbic acid + chlorpromazine in semen extender improved the post-thaw semen quality in crossbred bulls.

Leptin and leptin receptor in pig spermatozoa: evidence of their involvement in sperm capacitation and survival

Several studies have recently investigated the role of leptin, the adipocyte-secreted hormone, in the growth and reproduction of rodents, humans, and domestic animals. The present study was designed to explore the expression of leptin and its receptor in pig spermatozoa. Successful Western blot evidenced a 16 kDa band for leptin and six isoforms, ranging from 120 to 40 kDa, for the leptin receptor. Both leptin and leptin receptor were interestingly located at sperm acrosomal level, suggesting their involvement in the oocyte fertilization events. In fact, both capacitation indexes and acrosin activity were enhanced by leptin, and these effects were reduced by the anti-leptin receptor antibody. Afterwards, we investigated the main transduction pathways regulated by the hormone. Our results showed that, in pig sperm, leptin can trigger the signal transducer and activator of transcription 3, a classical component of cytokine signal transduction pathways, whose expression has not been previously reported in male gamete; in addition it was found constitutively activated. Besides, leptin was able to induce the activation of phosphatidylinositol phosphate kinase 3 and MAP kinase pathways as well as of BCL2, a known antiapoptotic protein. These data address to a role of leptin and its receptor on pig sperm survival. The presence of leptin and its receptor in pig sperm suggests that they, through an autocrine short loop, may induce signal transduction and molecular changes associated with sperm capacitation and survival.

Assessment of released acrosin activity as a measurement of the sperm acrosome reaction

To develop a method for assessing sperm function by measuring released acrosin activity during the acrosome reaction (AR). Human semen samples were obtained from 24 healthy donors with proven fertility after 3-7 days of sexual abstinence. After collection, samples were liquefied for 30 min at room temperature. Standard semen parameters were evaluated according to World Health Organization (WHO) criteria. Calcium ionophore A23187 and progesterone (P4) were used to stimulate the sperm to undergo AR. After treatment, sperm were incubated with the supravital dye Hoechst33258, fixed in a glutaraldehyde-phosphate-buffered saline solution, and the acrosomal status was determined by fluorescence microscopy with fluorescein isothiocyanate-labeled Pisum sativum agglutinin (FITC-PSA). The percentage of sperm undergoing AR (AR%) was compared to sperm acrosin activities as assessed by spectrocolorimetry. The correlation between AR% and acrosin activity was determined by statistical analysis. The AR% and released acrosin activity were both markedly increased with A23187 and P4 stimulation. Sperm motility and viability were significantly higher after stimulation with P4 versus stimulation with A23187 (P < 0.001). There was a significant positive correlation between released acrosin activity and AR% determined by FITC-PSA staining (r=0.916, P < 0.001). Spectrocolorimetric measurement of released acrosin activity might serve as a reasonable alternative method to evaluate AR.

Failed fertilization after ICSI and spermiogenic defects

To evaluate the relationship between late spermiogenic events, including histone-protamine replacement, acrosome integrity, and sperm morphology, with fertilization rate after intracytoplasmic sperm injection (ICSI). Prospective study. Isfahan Fertility and Infertility Center, Royan Institute, Tehran, Iran. Semen samples from 68 infertile couples undergoing ICSI at Isfahan Fertility and Infertility center were assessed during this study. Semen analysis was carried out according to World Health Organization criteria. Protamine deficiency, acrosin activity, sperm morphology, and acrosome size were assessed by chromomycin A3 (CMA3) staining, gelatinolysis test, and Papanicolaou staining (strict criteria), respectively. The correlation between protamine deficiency, sperm morphology, acrosin activity, and acrosome size with each other and fertilization rate were assessed. Percentage CMA3 positivity and mean halo diameter show a significant correlation with fertilization rate. However, no correlation was found between sperm normal morphology and fertilization rate. The mean values of acrosome size and fertilization rate were significantly different when patients were grouped for CMA3 positivity of 40%. Multiple linear regression analysis revealed that only protamine deficiency has direct effect on fertilization rate. Protamine deficiency appears to have a more significant effect on fertilization after ICSI than acrosin activity and semen parameters.

P-835: Relationship between acrosin activity of human spermatozoa and oxidative stress

P-835: Relationship between acrosin activity of human spermatozoa and oxidative stress

Acrosin activity in spermatozoa of infertile Nigerian males

The objective of this study was to evaluate acrosin activity in spermatozoa of infertile Nigerian men and to compare with those of the fertile men. The acrosin activity was evaluated using the Acroscreen reagent kit. The acrosin activity of the fertile men was 40.7±5.2 mIU/10(6) sperm (range 30.1-51.3) and those of the infertile men was 22.4±8.33 mIU/10(6) sperm (range 5.7-39.1). The difference in the mean was statistically significant (P<0.001). There was a subpopulation of the infertile men who had their acrosin activity within normal range of the fertile men, 32.9±1.57 mIU/10(6) sperm. But the difference in the mean was statistically significant (P<0.001). Acrosin activity decreased with increased morphological changes in the spermatozoa. It is concluded that acrosin activity in the infertile Nigerian men is significantly lower than that in the fertile men. Acrosin activity may also be affected by morphological changes in the spermatozoa.

Proacrosin/acrosin quantification as an indicator of acrosomal integrity in fresh and frozen dog spermatozoa

The scope of the present study was to evaluate the presence and activation of proacrosin/acrosin as a tool to determine the acrosomal status of fresh and frozen/thawed dog spermatozoa. Monoclonal antibody C5F11, directed against human acrosin, cross-reacted with dog spermatozoa and labeled the acrosome of both fresh and frozen/thawed dog spermatozoa. Frozen/thawed spermatozoa had a lesser proportion of labeled spermatozoa than fresh spermatozoa ( P < 0.05). When live spermatozoa were labeled with soybean trypsin inhibitor conjugated with Alexa 488 (SBTI-Alexa 488), the proportion of acrosome-labeled fresh spermatozoa was less than frozen/thawed spermatozoa ( P < 0.05). By using Western blots and enzymatic activity, frozen/thawed spermatozoa had a greater proportion of active acrosin than fresh spermatozoa. In addition, beta 1,4-galactosyl-transferase (GalT), a plasma membrane bound protein, remained attached to frozen/thawed spermatozoa. Proacrosin is activated during freezing/thawing of dog spermatozoa, and that proacrosin/acrosin may be a good indicator of acrosomal integrity of frozen/thawed spermatozoa.