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Significance of regional difference in ion concentrations in lizard, Hemidactylus flaviviridis (Rüppell): assessment of ionic influence on sperm motility in vitro.

A gradual increase in the concentration of Ca2+ from anterior to the posterior region was observed when mono- and divalent cations were estimated in different segments of the epididymis in wall lizard. Na+ and K+ levels increased from anterior to middle segment but declined significantly in the posterior segment. However, no significant difference in the levels of Mg2+ was observed in various segments. To study the influence of mono- and divalent cations on sperm motility in vitro, the spermatozoa from posterior region of the epididymis were incubated in medium with varying concentrations of Na+, K+, Ca2+ and Mg2+. Spermatozoa were non-motile when suspended in Na+-free medium. Addition of NaCl induced the acquisition of sperm motility in a concentration-dependent manner. Further, amiloride, a Na+-influx blocker, markedly reduced the Na+-induced forward progressive motility. Unlike Na+, the presence of K+ or Ca2+ in the incubation medium reduced the motility of spermatozoa even at very low concentrations. The inhibitory effect of Ca2+ decreased when nifedipine, a Ca2+-influx blocker, was added to the medium. Mg2+ at high concentrations only was able to reduce the forward progressive motility.

Animals↗

Reactive oxygen species released by activated neutrophils, but not by deficient spermatozoa, are sufficient to affect normal sperm motility.

OBJECTIVES: To determine whether reactive oxygen species (ROS) produced by abnormal or deficient spermatozoa and polymorphonuclear leukocytes (PMN, activated or not) can affect normal sperm motility. To determine what level of ROS is detrimental to spermatozoa. SUBJECTS: Patients consulting the Infertility Clinic at the Royal Victoria Hospital and healthy volunteers. METHODS: Normal spermatozoa (not producing ROS) were incubated with ROS-producing spermatozoa or PMN (activated or not), and motility was analyzed using a computer-aided sperm analysis system. The proportion of ROS produced by spermatozoa and released extracellularly was estimated by the decrease in the chemiluminescence observed in the presence of catalase and superoxide dismutase. RESULTS: There was no consistent effect on motility when ROS-producing and non-ROS-producing spermatozoa were mixed. This lack of effect could be due to the relatively low level of ROS produced by spermatozoa as well as the fact that only a third of these ROS are released outside spermatozoa. To cause a low but significant decrease (15 +/- 4%) in sperm motility after a 5-hour incubation, the level of ROS imposed on spermatozoa must be equivalent to ROS released by 1 x 10(6) activated PMN/mL. CONCLUSIONS: The production of ROS by deficient spermatozoa is low and of no consequence to the motility of normal spermatozoa present in the same sperm preparation. However, 1 x 10(6) activated PMN/mL, with a 1,000-fold higher ROS production, have detrimental effects on the motility of normal washed spermatozoa.

Extracellular Space↗

The automated analysis of rat sperm motility following subchronic epichlorohydrin administration: methodologic and statistical considerations.

The automated analysis of sperm motion endpoints is potentially useful in identifying male reproductive toxicants and ultimately in predicting fertility in humans. The present study was designed to evaluate the automated analysis of rat sperm motility characteristics following subchronic administration of epichlorohydrin. This type of validation is a prerequisite for inclusion of sperm motion measurements in the process of reproductive risk assessment. In the present studies videotapes were made of cauda epididymal spermatozoa from Long-Evans rats, both untreated and treated with epichlorohydrin. From analysis of videotapes of control epididymal spermatozoa, the relationship of various sperm motion endpoints and settings of the CellSoft computer-assisted sperm motion analysis system (Cryo Resources, Ltd., New York, NY) is described. Optimal settings of the system for analysis of rat spermatozoa are detailed. Employing data from both control and epichlorohydrin-treated animals, a statistical methodology is described that evaluates: (1) the distributions of CellSoft generated sperm motion endpoints, (2) the correlations between these endpoints, and (3) techniques for detection of dose-related effects.

Animals↗

Comparative effects of (+)-propranolol and nonoxynol-9 on human sperm motility in-vitro.

Using the modified transmembrane migration method to measure sperm motility, it was shown that the surfactant nonoxynol-9 alone, was twice as potent as (+)-propranolol alone as a spermicidal agent. Addition of (+)-propranolol to nonoxynol-9 shifted the dose-response curve to the left of the curves for either component alone, and a surprising synergistic action was evident. These observations may form the basis for the development of a new advantageous topical contraceptive combination product.

Dose-Response Relationship, Drug↗

A technique for selecting and concentrating the motile sperm from semen in oligozoospermia.

Using an albumen-filled column, it was possible to separate highly motile sperm from autoagglutinated, dead and immotile sperm, as well as from other seminal debris. Motility was increased from a mean of 45.4 to 74.3% (P less than 0.001), although the sperm density fell from a mean of 28.2 million per ml to 9.7 million per ml (P less than 0.001). When these selected highly concentional AlH had been repeatedly tried without success, 3 pregnancies were achieved in 18 women (16.7%).

Cell Separation↗

Complementary effects of propranolol and nonoxynol-9 upon human sperm motility.

The inhibitory effects of nonoxynol-9, DL- and D-propranolol upon human sperm motility were determined in vitro. All three compounds were capable of causing complete cessation of sperm movement. However, greater efficacy was achieved using combinations of nonoxynol-9 and propranolol, suggesting a complementary interaction between these compounds. Investigations of the mechanism of action of propranolol revealed that an influx of calcium accompanied the loss of motility. However, since incubation in the absence of calcium enhanced the spermicidal effects of this compound, it was concluded that this calcium influx did not constitute the primary means by which motility was disrupted. Low doses of propranolol, which did not affect motility, were found to inhibit the capacity of human spermatozoa for sperm-oocyte fusion.

Calcium↗

The effect of pentoxifylline on sperm motility, oocyte fertilization, embryo quality, and pregnancy outcome in an in vitro fertilization program.

OBJECTIVE: To study the effect of pentoxifylline on sperm motility, oocyte fertilization, embryo cleavage, and quality as well as pregnancy outcome on asthenospermic patients participating in an IVF program. DESIGN: Prospective randomized study. SETTING: Private IVF unit. PATIENTS: Ninety-seven couples, 24 of whom were repeating IVF. Two semen specimens were obtained from each patient and each specimen was divided equally into two parts, nontreated (control semen) and pentoxifylline-treated (treated semen). MAIN OUTCOME MEASURE: Sperm progressive motility, oocyte fertilization. RESULTS: Overall and progressive motility did not differ significantly between the two semen specimens. There was a significant increase in the progressive motality of the pentoxifylline-treated semen compared with control semen. No significant difference was noticed between control and treated semen in fertilization rate, cleavage rate, embryo quality, and pregnancy rate. The percentage of patients who fertilized only with control semen (9.3%) was not significantly different from that of patients who fertilized only with treated semen (10.3%). Couples who were repeating IVF did not show significant difference in fertilization between the present study and previous attempts. CONCLUSION: Our results showed that although the sperm progressive motility is improved after pentoxifylline treatment, it is doubtful whether this effect is of any clinical significance.

Cleavage Stage, Ovum↗

ADP-ribosylation of Rho proteins inhibits sperm motility.

The highly homologous Rho proteins RhoA, RhoB and RhoC are low-molecular-mass GTP-binding proteins. They are selectively ADP-ribosylated by Clostridium botulinum ADP-ribosyltransferase C3 (C3 exoenzyme). The biological function of the Rho proteins is still unclear; there is evidence that they are involved in the regulation of the filamental network of cells. Here we report that C3 exoenzyme-like toxins ADP-ribosylate small GTP-binding proteins in bovine spermatozoa and inhibit sperm motility. These findings indicate that Rho proteins which reportedly regulate the microfilament system are basically involved in sperm motility.

ADP Ribose Transferases↗

Nitric oxide synthase and nitrite production in human spermatozoa: evidence that endogenous nitric oxide is beneficial to sperm motility.

The aim of this study was to investigate the presence of nitric oxide synthase (NOS) and the production of nitric oxide (NO) by human spermatozoa. Immunoreactivity was examined using a polyclonal antibody raised against porcine cerebellar nitric oxide synthase and monoclonal endothelial (eNOS) and brain (bNOS) antibodies. Using each antibody, NOS was observed localized in the head and midpiece regions of the spermatozoon. Immunofluorescence observed for eNOS and bNOS was more intense in normozoospermic samples. Sperm motility was assessed by computer-assisted semen analysis (CASA) in the presence and absence of NG-nitro-L-arginine methyl ester (L-NAME; 10(-5)M), and NO synthesis inhibitor or tumour necrosis factor (TNF)-alpha (20 IU/ml), a superoxide inducer. In the presence of L-NAME, percentage progressive motility, average path velocity (VAP), straight line velocity (VSL) and curvilinear velocity (VCL) were significantly reduced after 30 min. Sperm viability was not decreased by TNF alpha or L-NAME. The accumulation of nitrite (the stable end-product of the NOS/NO pathway) by spermatozoa was measured using the Griess reaction. After 8 h, nitrite concentrations were lower in asthenozoospermic compared to normozoospermic samples. In the presence of TNF alpha, nitrite accumulation was significantly reduced in normozoospermic samples. We conclude that NOS is present in human spermatozoa and that eNOS and bNOS are abundant in normozoospermic samples. Nitric oxide (at endogenous concentrations) appears to be necessary for adequate sperm motility.

Animals↗

Sperm motility hyperactivation facilitates penetration of the hamster zona pellucida.

These experiments were conducted to determine whether or not sperm motility hyperactivation facilities penetration of the zona pellucida of the oocyte. Two approaches were used. For the first, hamster sperm were incubated for 4.0-4.25 h in a capacitating medium that contained either 2.9 or 25.0 mM sodium bicarbonate. In these media, sperm became equally capacitated as evidenced by their ability to undergo the acrosome reaction when exposed to lysophosphatidyl choline or intact zonae pellucidae; however, sperm became hyperactivated only in the medium containing 25.0 mM bicarbonate. When these sperm were added to cumulus-free oocytes in vitro, only 2 of 88 oocytes were penetrated by sperm preincubated in 2.9 mM bicarbonate, while 31 of 86 oocytes were penetrated by sperm in 25.0 mM bicarbonate. It was found that equal numbers of sperm were bound to the oocytes and that equal numbers were acrosome-reacted on the surface of the zonae in the two media. For the second approach, sperm were incubated in a capacitating medium containing 25 mM bicarbonate. When > 70% were hyperactivated, aliquots were added to three sets of oocytes. After 10 min had been allowed for sperm to attach and acrosome-react, inhibitors of hyperactivation were added and the sperm and oocytes were incubated for an additional 20 min before fixation and examination for zona penetration. In the dishes treated with the inhibitors verapamil or Cd2+, 1 of 42 and 0 of 42 oocytes were penetrated, respectively, compared with 25 of 40 in controls. Therefore, it appears that hyperactivation facilitates penetration of the hamster zona pellucida.

Acrosome↗

Changes in sperm motility, morphology and concentration induced by the swim-up procedure.

Fifteen semen samples from normal donors were washed twice in Earle's medium by centrifugation and allowed to swim up into overlayered Earle's medium. Aliquots of 0.2 ml were drawn before and after the swim-up, for assessment of sperm motility, morphology and concentration. The percentage of sperm showing progressive motility improved by an average of 54.02 +/- 45.97%, and sperm with the normal morphology increased by 63.8 +/- 31.01%. Head and tail abnormalities decreased by 47.17 +/- 21.3% and 89.37 +/- 16.1% respectively. There was no statistical change in acrosomal abnormalities. Sperm concentration was reduced by 89.02 +/- 7.4%. It is concluded that the standard swim-up technique resulted in an overall improvement in sperm quality, but that the technique was not effective in reducing the acrosomal abnormalities.

Humans↗

Evaluation of human sperm motility by means of transmembrane migration method and computer assisted semen analysis: a comparison study.

Comparing motility parameters with a trans-membrane migration method and a Hamilton-Thorn HTM-2000 computer assisted semen analyzer, we found that trans-membrane migration ratio (TMMR) correlated best with critical motility which indicated the fraction of fastest and straightest sperm in a semen sample. We also found that TMMR correlated better with progressive velocity than with track speed. It is concluded that nonprogressive sperm were not included in the estimation of TMMR and the trans-membrane migration method is most suitable for studying drug effect on straight and rapid sperm motility.

Computers↗

The 12 kD FK 506 binding protein FKBP12 is released in the male reproductive tract and stimulates sperm motility.

BACKGROUND: The 12 kD FK506 binding protein FKBP12 is a cytosolic receptor for the immunosuppressant drugs FK506 and rapamycin. In addition to its critical role in drug-induced T-cell immunosuppression, FKBP12 associates physiologically with ryanodine and inositol 1,4,5-trisphosphate (IP3) receptors, regulating their ability to flux calcium. We investigated a role for FKBP12 in male reproductive physiology on the basis of our identification of extremely high levels of [3H]FK506 binding in male reproductive tissues. MATERIALS AND METHODS: [3H]FK506 binding studies were performed to identify tissues enriched with FK506 binding sites. The abundant [3H]FK506 binding sites identified in the male reproductive tract were localized by [3H]FK506 autoradiography. FK506 affinity chromatography was employed to purify FKBP from epididymal fluid. Anti-FKBP12 Western analysis was used to confirm the identity of the purified FKBP. The binding of exogenous FKBP12 to sperm was evaluated by [32P]FKBP12 binding studies and [33P]FKBP12 autoradiography. The effect of recombinant FKBP12 on sperm motility was investigated using a Hamilton Thorne motility analyzer. RESULTS: Male reproductive tissues contained high levels of [3H]FK506 binding. The localization of [3H]FK506 binding sites to the tubular epithelium of the caput epididymis and the lumen of the cauda and vas deferens suggested that FKBP is released in the male reproductive tract. FKBP12 was purified from epididymal plasma by FK506 affinity chromatography. Radiolabeled FKBP12 specifically bound to immature but not mature sperm. In sperm motility studies, FKBP12-treated caput sperm exhibited double the curvilinear velocity of untreated controls. CONCLUSIONS: High levels of FKBP12 are released in the male reproductive tract and specifically associate with maturing sperm. Recombinant FKBP12 enhances the curvilinear velocity of immature sperm, suggesting a role for FKBP12 in motility initiation. The highest concentrations of soluble FKBP12 in the male reproductive tract occur in the lumen of the vas deferens, a site of sperm storage and the conduit for ejaculated sperm. Preservation of mammalian sperm for reproductive technologies may be optimized by supplementing incubation or storage media with FKBP12.

Animals↗

Effect of number of motile sperms inseminated on reproductive performance of rabbit does.

The relationship between the number of motile sperms inseminated and fertility rate and litter size at birth in rabbits was investigated. A total of 651 artificial inseminations on multiparous rabbit does in different physiological states (lactating or not; sexually receptive or not) were inseminated with a number of motile cells/dose varying from about 1-20 x 10(6). The statistical model computed the following traits: maximum expected value (a), rate of approach to this value (b) and number of motile spermatozoa/dose needed to obtain threshold level (95% of the maximum). The physiological condition of the doe affected the reproductive performance. The sexual receptivity had significant effect during lactation: non-receptive does had the lowest fertility (a: 43.70%; P < 0.05) while no significant differences were found in non-lactating ones (73.80 vs. 89.11%). Also the rate of approach to this maximum was different: non-receptive does showed a higher dependence on spermatozoa number than receptive females (b: 0.69 and 0.58, respectively, in lactating and non-lactating does) and consequently more spermatozoa (13.1 and 11.1 x 10(6) motile/dose) were needed to reach the threshold value. The physiological state of the does did not affect litter size and only 4.1 x 10(6) of motile spermatozoa/dose are needed to reach the threshold value.

Animals↗

The effects of beta-adrenergic drugs on the human sperm motility in vitro. I. The effects of propranolol and isoprenaline.

The investigations were carried out on 28 samples of human semen with asthenozoospermia. The experiments were performed in three groups of semen, adding to the first one 0.34 mM of propranolol and 0.025 mM of isoprenaline, to the second one the double dose and to the last one the fivefold dose of the drugs. The lowest concentrations of the drugs applied in our investigations result from the used therapeutic intravenous doses for people. The proportional content of moving sperma was evaluated under the phase-contrast microscope at the 5, 30, 60, 120 and 240 minute of the experiment. The addition of 0.34 mM of propranolol caused a decrease of the sperm motility but only after 60 minutes and for a short period of time. However, the addition of the double and the fivefold doses of the drug led to immediate and significant reduction of the sperm motility. The lowest dose of isoprenaline markedly increased the sperm activity in comparison with the control values but not earlier than 120 minutes after the addition. The further increase in dose showed no effect on the moving sperm percentage increase, merely accelerating this effect.

Adult↗

Expression and localization of prolyl oligopeptidase in mouse testis and its possible involvement in sperm motility.

Prolyl oligopeptidase (POP) expression in mouse testis during sexual maturation was examined. Northern blot analysis showed that POP mRNA expression was highest at 2 weeks of age, and gradually reduced thereafter. However, enzyme activity was almost constant during the examined period. In situ hybridization study revealed a change in the expression site of POP mRNA in testis during sexual maturation. Positive signals were detected in all types of cells in the seminiferous tubules before maturation, and were restricted to spermatids at the spermatogenesis cycle stages I-VIII in adult mice. POP was detected in the insoluble fraction of sperm by Western blot analysis. Immunohistochemical analyses showed that POP is localized in the spermatids at steps 12-16 of spermiogenesis and in the midpiece of the sperm fragellum. It was also found that specific POP inhibitors, poststatin and benzyloxycarbonyl-proline-prolinal, suppressed sperm motility. These results suggest that POP may be involved in meiosis of spermatocytes, differentiation of spermatids, and sperm motility in the mouse.

Aging↗

Effects of pentoxifylline on sperm motility in normogonadotropic asthenozoospermic men.

Forty-seven normogonadotropic men with idiopatic asthenozoospermic were divided at random: group I (N = 22) received placebo and group II (N = 25) received 1200 mg of pentoxifylline/day during 6 months. Semen analysis was performed basal and at 3 and 6 months of the study period. No statistical changes in serum hormone concentration were found, nor in volume, sperm counts, viability, and morphology before and after treatment. Sperm motility increased following pentoxifylline treatment after 3 and 6 months from 25.5 (21.0-30.0) to 35.5 (31.5-39.0) (p < .00001) and to 42.0 (38.0-46.0) (p < .00001), respectively. Although in the placebo control cases some changes were observed in the sperm motility, they were less significant. Furthermore, progressive motility only in grade A increased with pentoxifylline from 2.5 (0.0-6.0) to 12.0 (6.0-19.5) (p < .001) at 3 months and to 22.5 (17.0-26.0) at 6 months (p < .00001). In conclusion, pentoxifylline had an additional effect rather than placebo and was useful treatment in these cases of male factor infertility.

Adult↗

Inhibitory effects of gossypol analogs on human sperm motility.

Aqueous-soluble gossypol Schiff's bases, SP562: bis-8,8'-[(N-(2-(dimethylamino)ethyl]-iminomethylene]- [1,1',6,6',7,7'-hexahydroxy-5,5'-diisopropyl-3,3'-dimethyl-2,2- binaphthalene dihydrochloride; SP563: bis-8,8'-[(N-(2-(diethylamino)ethyl]-iminomethylene]-1,1',6,6',7,7 '- hexahydroxy-5,5'-diisopropyl-3,3'-dimethyl-2,2'-binaphthalene++ + dihydrochloride; and SP564: bis-8,8'-[(N-(2-(diethylamino)propyl]-iminomethylene]- 1,1',6,6',7,7'-hexahydroxy-5,5'-diisopropyl-3,3'-dimethyl-2,2'- binaphthalene dihydrochloride, were investigated for their effects on human sperm motility. SP564, which has the longest alkyl substituent of the Schiff's base, appeared to exert the greatest inhibitory effects on human sperm motility. These inhibitory effects were even greater than those caused by (+/-)gossypol acetic acid at the same concentration.

Dose-Response Relationship, Drug↗