Characterizing the glycocalyx of poultry spermatozoa: I. Identification and distribution of carbohydrate residues using flow cytometry and epifluorescence microscopy

The aim of the present work was to use a battery of lectins to 1) delineate the carbohydrate content of sperm glycocalyx in the turkey and chicken using flow cytometry analysis, and 2) evaluate the distribution of existing sugars over the sperm plasma membrane surface with epifluorescent microscopy. Carbohydrate groups (corresponding lectins) that were investigated included galactose (GS-I, Jacalin, RCA-I, PNA), glucose and/or mannose (Con A, PSA, GNA), N-acetyl-glucosamine (GS-II, s-WGA, STA), N-acetyl-galactosamine (SBA, WFA), fucose (Lotus, UEA-I), sialic acid (LFA, LPA), and N-acetyl-lactosamine (ECA). Spermatozoa were assessed before and after treatment with neuraminidase to remove sialic acid. Mean fluorescence intensity (MnFI) was used as indicator of lectin binding for flow cytometry analysis. Nontreated spermatozoa from both species showed high MnFI when incubated with RCA-I, Con A, LFA, and LPA, as did chicken spermatozoa incubated with s-WGA. Neuraminidase treatment increased the MnFI for most lectins except LFA and LPA, as expected. Differences in MnFI between species included higher values for s-WGA and ECA in chicken spermatozoa and for WFA in turkey spermatozoa. Microscopy revealed segregation of some sugar residues into membrane-specific domains; however, the 2 staining techniques (cell suspension vs fixed preparation) differed in identifying lectin binding patterns, with fixed preparations yielding a high degree of nonspecific binding. We conclude that 1) the glycocalyx of turkey and chicken spermatozoa contains a diversity of carbohydrate groups, 2) these residues are extensively masked by sialic acid, 3) the glycocalyx composition is species-specific, and 4) some glycoconjugates appear to be segregated into membrane-specific domains. Characterization of the poultry sperm glycocalyx is the first step in identifying the physiological impact of semen storage on sperm function.     

Localization of specific carbohydrate configurations in the hemophilic synovial membrane

Fluorescein isothiocyanate-conjugated lectins include: concanavalin A (Con-A), Dolichos biflorus agglutinin (DBA), Griffonia simplicifolia-I (GS-I), Griffonia simplicifolia-II (GS-II), Arachis hypogaea agglutinin (PNA), Maclura pomifera agglutinin (MPA), Ricinus communis agglutinin-I (RCA-I), Glycine max agglutinin (SBA), Ulex europaeus agglutinin-I (UEA-I), and wheat germ agglutinin (WGA). These lectins were used to histochemically demonstrate lectin bindings on hemophilic synovial membrane. GS-I (galactose and galactosamine specific) and SBA (galactosamine specific) was shown to bind strongly in the cytoplasm of the lining cells. Lymphocytes and mast cells were largely bound by Con-A (mannose and glucose specific) and GS-II (glucosamine specific) positive. UEA-1 (fucose specific) was shown to bind specifically to synovial vascular endothelial cells. Lectin histochemistry is a useful method for classification of the cells on the synovial membrane.     

Liquid semen storage in elephants (Elephas maximus and Loxodonta africana): species differences and storage optimization

Artificial insemination plays a key role in the genetic management of elephants in zoos. Because freshly extended semen is typically used for artificial insemination in elephants, it has become imperative to optimize conditions for liquid storage and semen transport. The objectives of this study were to examine the interactions between different extenders and storage temperatures on sperm total motility, progressive motility, and acrosomal integrity in Asian (Elephas maximus) and African (Loxodonta africana) elephants. Ejaculates were collected by rectal massage, diluted using a split-sample technique in 5 semen extenders: TL-Hepes (HEP), Modena (MOD), Biladyl (BIL), TEST refrigeration medium (TES), and INRA96 (INR), maintained at 35°C, 22°C, or 4°C. At 0, 4, 6, 12, and 24 hours, aliquots were removed and assessed for sperm total motility, progressive motility, and acrosomal integrity. After 24 hours of storage, African elephant spermatozoa exhibited greater longevity and higher values in sperm quality parameters compared with those of Asian elephants. In both species, semen storage at 35°C resulted in a sharp decline in all sperm quality parameters after 4 hours of storage, whereas storage at 22°C and 4°C facilitated sperm survival. In Asian elephants, MOD and HEP were most detrimental, whereas BIL, TES, and INR maintained motility up to 12 hours when spermatozoa were cooled to 22°Cor4°C. In African elephants, there were no differences among extenders. All media maintained good sperm quality parameters at 22°C or 4°C. However, although MOD, BIL, and INR were most effective at lower temperatures, HEP and TES maintained sperm motility at all storage temperatures. This study demonstrated sperm sensitivity to components of various semen extenders and storage temperatures and offers recommendations for semen extender choices for liquid semen storage for both Asian and African elephants.     

Kinematic changes during the cryopreservation of boar spermatozoa

The present study evaluates the effect that various steps of a conventional cycle of cryopreservation have on the patterns of movement exhibited by boar spermatozoa. Sperm-rich ejaculate fractions collected from 24 mature fertile boars (1 ejaculate per boar) were cryopreserved following a standard freeze-thaw procedure with 0.5-mL plastic straws. Overall sperm motility and the individual kinematic parameters of motile spermatozoa (assessed by the computer-aided sperm analysis system Sperm Class Analyzer [SCA]) were recorded in 5 steps of the cryopreservation procedure. These steps were as follows: 1) at the time that the fresh semen was extended, 2) at 17 degrees C, after sperm concentration by centrifugation and re-extension of the pellet with lactose-egg yolk extender; 3) at 5 degrees C, after added freezing extender; 4) at the time that thawed semen was held in a water bath at 37 degrees C for 30 minutes; and 5) at the time that thawed semen was held in a water bath at 37 degrees C for 150 minutes. Data from individual motile spermatozoa, defined by 7 kinematic parameters (curvilinear velocity [VCL], straight-line velocity [VSL], average path velocity [VAP], linearity [LIN], straightness [STR], mean amplitude of lateral head displacement [ALH], and beat cross frequency [BCF]), were analyzed using a pattern analysis technique (PATN) to identify and quantify populations and subpopulations of motile sperm within the semen samples. After the first cluster analysis, 3 motile sperm populations (P) were identified (P1: progressive and/or vigorous cells [90.4%], P2: poorly progressive cells [8.3%], and P3: nonprogressive cells [1.3%]). These populations remained constant (P > .05) throughout the 5-step cryopreservation procedure. A second PATN was carried out within the P1 sperm population, which identified 3 sperm subpopulations (sP) (eg, sP1: cells with progressive and vigorous movement [58.7%], sP2: progressive cells only [24.6%], and sP3: vigorous cells only, hyperactive-like [16.7%]). Although the relative frequency of these 3 subpopulations varied among ejaculates (boars), there was no interaction with any cryopreservation step we examined. Whereas sP1 remained constant (P > .05), sP2 and sP3 varied significantly (P < .05) through the cryopreservation procedure, with the increase in sP3 after centrifugation at 17 degrees C and during cooling at 5 degrees C considered particularly relevant. In conclusion, the present study confirms the heterogeneity of sperm movement patterns in boar semen, patterns that vary through the cryopreservation procedure, especially after removal of the seminal plasma by centrifugation and subsequent extension at 17 degrees C and after the slow cooling at 5 degrees C, when obvious increases in hyperactivated movement appeared. The vast majority of spermatozoa, those exhibiting progressive and vigorous movement, remained constant during the cryopreservation procedure, although the proportion differed among boars.     

Viability and fecundity of human semen specimens cryostored and transported at 5 degrees C using the Bio-Tranz shipping

This study was designed to assess the viability and fecundity of semen stored at 5 degrees C for 24 hours using the Bio-Tranz shipping system. Semen specimens were assessed for motility and sperm membrane integrity at the time of collection and 24 hours after storage in the Bio-Tranz. In group 1 (n = 61), specimens were diluted in TYB, processed and used for intrauterine insemination (IUI), leaving an aliquot for storage for 24 hours in the Bio-Tranz. In group 2 (n = 67), specimens were diluted in TYB, stored for 24 hours in the Bio-Tranz and then processed and used for IUI. In both groups, the total motile sperm used for IUI was similar and the women that underwent IUI were standardized for ovulation prediction and time of insemination. The overall sperm characteristics between the two groups were within normal range. Significant decreases were noted in sperm motility and membrane integrity in both groups after storage. Similar pregnancy rates were obtained between the two patient populations. The use of the Bio-Tranz shipper is extremely convenient for patients requiring semen evaluation, cryostorage or IUI and other assisted reproductive technologies.     

TEST-egg yolk buffer storage increases the capacity of human sperm to penetrate hamster eggs in vitro

A total of 85 semen samples from infertility clinic patients were examined to study the effect of storage at 4 degrees C in TES-Tris (TEST)-egg yolk buffer for 24 h on the penetrating capacity of sperm in the zona-free hamster egg penetration assay (HEPA). The mean sperm penetration rate and the fertilization index increased significantly after storage in TEST-egg yolk buffer. Only five out of the 85 samples (5.9%) failed to show any improvement in sperm penetration rate after cold storage. The sperm penetration rate before cold storage showed no significant correlations with routine semen characteristics, semen ATP concentration or the functional integrity of sperm membranes as measured by the hypo-osmotic swelling technique. Significant but low correlations were observed between sperm penetration rate after cold storage and the following semen parameters: sperm count, % motility, total number of motile sperm, % normal sperm morphology, total number of normal sperm, semen ATP concentration and sperm penetration rate before cold storage. Partial correlation analysis revealed that the positive correlation between semen ATP concentration and sperm penetration rate after cold storage was not a direct relationship but was due to the correlation with sperm count. The combination of sperm penetration rate before cold storage, sperm count and % normal sperm morphology accounted for 26.2% of the variation in sperm penetration rate after cold storage by stepwise multiple regression analysis, while sperm penetration rate before cold storage alone explained 13.5% of the variation. The results indicate that TEST-egg yolk buffer treatment can enhance sperm penetration rate in vitro and may be useful in the treatment of impaired sperm fertility.(ABSTRACT TRUNCATED AT 250 WORDS)     

Novel association between sperm deformity index and oxidative stress-induced DNA damage in infertile male patients

Aim: To investigate the impact of abnormal sperm morphology using the sperm deformity index (SDI) on reactive oxygen species (ROS) production and its correlation with sperm DNA damage. Methods: Semen samples were collected from men undergoing infertility screening (n=7) and healthy donors (n=6). Mature spermatozoa were isolated and incubated with 5mmol/L β-nicotinamide adenine dinucleotide phosphate (NADPH) for up to 24h to induce ROS. Sperm morphology was evaluated using strict Tygerberg‘s criteria and the SDI. ROS levels and DNA damage were assessed using chemiluminescence and terminal deoxynucleotidyl transferase-mediated fluoresceindUTP nick end labeling (TUNEL) assays, respectively. Results: SDI values (median [interquartiles]) were higher in patients than donors (2 [1.8,2.1] vs. 1.53 [1.52, 1.58], P=0.008). Aliquots treated with NADPH showed higher ROS levels (1.22 [0.30, 1.87] vs. 0.39 [0.10, 0.57], P=0.03) and higher incidence of DNA damage than those not treated (10 [4.69, 24.85] vs. 3.85 [2.58, 5.10], P=0.008). Higher DNA damage was also seen following 24h of incubation in patients compared to donors. SDI correlated with the percentage increase in sperm DNA damage following incubation for 24h in samples treated with NADPH (r=0.7, P=0.008) and controls (r=0.58, P=0.04).Conclusion: SDI may be a useful tool in identifying potential infertile males with abnormal prevalence of oxidative stress (OS)-induced DNA damage. NADPH plays a role in ROS-mediated sperm DNA damage, which appears to be more evident in infertile patients with semen samples containing a high incidence of morphologically abnormalspermatozoa.     

Time-related decline in sperm motility patterns in men with cytotoxic antibodies

Sperm motility patterns in semen from 10 fertile nonautoimmune men (fertile control group) and 33 infertile men with various titers of cytotoxic sperm antibodies in their seminal plasma (group 1: titers less than or equal to 16, n = 6; group 2: titers 64 to 512, n = 12; group 3: titers greater than or equal to 1024, n = 15) were evaluated every 2 hours for 12 hours and finally at 24 hours. A significant decline in sperm swimming speed and linearity was observed at 6 hours in semen from 27 infertile men with sperm antibodies. Beginning at 8 hours, semen from sperm antibody-positive men in group 2 showed a significant decline in percentage motile sperm (p less than 0.001) compared to the fertile controls. The percentage motility in semen of donors in groups 1 and 3 was significantly lower than that in semen of fertile donors at 10 hours (p less than 0.05), 12 hours (p less than 0.01), and 24 hours (p less than 0.001). The mean velocity in groups 2 and 3 was significantly less than that in fertile controls at 10 and 12 hours (p less than 0.05). The linearity of sperm motility started to decline 4 hours after semen samples were obtained from sperm antibody-positive groups 2 and 3 in contrast to sperm antibody-negative fertile or infertile groups (p less than 0.05). It is concluded that the presence of cytotoxic sperm antibodies in the seminal plasma adversely affects sperm linearity as early as 4 hours after semen collection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lectin Histochemical Study of Lipopigments Present in the Cerebellum of Solanum fastigiatum var. fastigiatum Intoxicated Cattle

This study was carried out to investigate the pattern of lectin binding in the cerebellum of calves poisoned with Solanum fastigiatum var. fastigiatum. For the experimental reproduction of the illness, S. fastigiatum var. fastigiatum was collected from farms where the intoxication occurs. The dried ground plant was administered to two 1‐year‐old cattle by a ruminal cannula. The animals received 5 g/kg b.w. daily, 5 days a week, during periods of 107 and 140 days. After these periods the animals were bled to death. For the histological study, transverse sections of the cerebellum were used. Paraffin‐embedded sections were incubated with the following biotinylated lectins with different specificity: Concanavalia ensiformis (Con‐A), Glycine max (SBA), Dolichos biflorus (DBA), Ulex europeus‐I (UEA‐I), Triticum vulgaris (WGA), succynyl‐WGA (sWGA), Arachis hypogaea (PNA), Ricinus communis‐I (RCA‐I) and Bandeirea simplicifolia‐I (BS‐I). Avidin–biotin–peroxidase complex was applied as a detection system. Purkinje cells showed vacuolation in the pericaryon. The stored material present in the cells reacted strongly with the following lectins: Con‐A, sWGA, WGA and RCA‐I. An irregular affinity was observed with PNA and DBA. The lectin‐binding pattern was compatible with a glycolipid storage disease.     

Cryopreservation of rhesus monkey (Macaca mulatta) epididymal spermatozoa before and after refrigerated storage

Recently, there has been an increased interest in preservation of epididymal sperm as a potential source of material for genetic resource banking; however, cryopreservation of epididymal sperm from the rhesus monkey has not been explored. This study evaluated the effect of prolonged refrigerated storage of the intact cauda epididymides at various conditions on the postthaw motility of rhesus monkey epididymal spermatozoa, and also tested whether altering cryoprotectants and cooling methods could improve post-thaw motility for epididymal sperm after refrigerated storage. Motility before freezing decreased significantly after refrigerated storage (0 degrees C) for a period of 24 or 48 hours. Although postthaw motility was not significantly different after 24 hours of refrigerated storage, epididymides stored at a higher temperature (4 degrees C-10 degrees C) yielded better results, but postthaw motility still decreased significantly after 48 hours of refrigerated storage at 4 degrees C. Comparisons of glycerol and ethylene glycol at 3% and 6% revealed similar postthaw motility. However, consistently high postthaw motility was obtained with 3% glycerol throughout all freezing trials regardless of whether samples were collected fresh or after refrigerated storage for 24 or 48 hours. Cooling at a higher rate of 220 degrees C/min was found to yield better postthaw motility than the slower rate of 29 degrees C/min. Thawing time duration was evaluated, and a minimum of 30 seconds was required for thawing 0.25-mL straws containing 50-microL semen samples. An overall average of 42% postthaw motility was obtained for rhesus monkey epididymal sperm packed in 3% glycerol and cooled after 24 or 48 hours refrigerated storage. These postthaw motility results for epididymal sperm indicate that this method should be practical for use in preserving epididymal sperm, even if tissue must be shipped from sites remote from the cryopreservation laboratory.     

Capacitated and acrosome reacted spermatozoa of goat (Capra indicus): a fluorescence and electron microscopic study

Summary. Plasma membrane alterations accompanying in vitro capacitation and acrosome reaction of goat spermatozoa were studied using lectin labelling, scanning electron microscopy, and freeze-fracture methods. Fluorescein isothi-ocyanate linked lectins namely; Canavalia ensiformis (ConA), Maclura pomifera (MPA), Arachis hypogaea (PNA), Glycine max (SBA) and Triticum vulgaris (WGA) agglutinin were used to examine the distribution of surface carbohydrates during these two events. The head and the sperm tail reveal altered lectin labelling features after capacitation and acrosome reaction. After capacitation the surface coat components for MPA, SBA, and WGA are shed from the spermatozoon head. ConA receptors on the head are retained after capacitation but are partially shed in the acrosome reacted spermatozoa. SBA receptor sites appear on the sperm tail of the capacitated spermatozoa. Unusual morphological changes attending capacitation involve the sperm tail-end which develops a novel entity, which we have termed 'spatula'. The 'spatula' shows strong binding with ConA and WGA only. In the acrosome reacted spermatozoa the spatulated tail-end unwinds with a concomitant loss of lectin labelling. Highly ordered membrane particles, 'ladders' of the middle piece of the epididymal sperm tail, disappear and IMP clearings appear on the middle piece and in the spatulated ends of the capacitated spermatozoa. But in the acrosome reacted sperm IMPs reappear and are randomly disposed on the middle-piece and are clustered in small patches on the principal-piece. IMP free areas appear on the plasma membrane covering the acrosome and the outer acrosomal membrane (OAM) of the capacitated spermatozoa. The plasma membrane and OAM fuse at multiple foci and appear as acrosomal 'ghosts' which remain associated with the sperm head even after acrosome reaction.     

A study on occupational exposure to petrochemicals and smoking on seminal quality

A cross-sectional study of 68 petrochemical workers (23 had never smoked [E/NS], 45 were current smokers [E/S]) and 130 subjects with no known history of exposure to petrochemicals (49 had never smoked [NE/NS], 81 were current smokers [NE/S]) was conducted to assess the effect of occupational exposure to petrochemicals and smoking on semen quality. In-person interviews revealed occupational history, smoking habit, and lifestyle. Semen parameters such as volume, viability, sperm forward progression rate, sperm density, and total sperm count were determined for all subjects. The results show that the E/NS workers had a lower sperm forward progression rate (P < .05) compared with controls (NE/NS). Individuals in the NE/S group showed a significant inverse relationship between years smoked and sperm density (r = -.24, P < .05). The data also revealed that cigarette smokers who had worked in a petrochemical plant had significantly poorer quality semen, including sperm density, total sperm count, and forward progression rate, compared with the control (NE/NS) group (P < .01). Furthermore, there was a significant inverse correlation between combined exposure and smoking years, and sperm density (r = -.28, P < .05). These findings suggest that occupational exposure to petrochemical compounds may aggravate the adverse effect that smoking has on semen quality.     

Temporal decreases in sperm motility: which patients should have motility checked at both 1 and 2 hours after collection?

A decrease in sperm motility, and thus total motile sperm count (TMSC), over a period of hours might have clinical implications in counseling couples considering intrauterine insemination (IUI), in vitro fertilization (IVF), and intracytoplasmic sperm injection (ICSI). The objective of this study was to identify patients with decreases in sperm motility from 1 to 2 hours after collection and examine predictive relationships with semen analysis parameters. Between 2001 and 2005, 2313 semen samples were analyzed. Sperm motility was evaluated at both 1 and 2 hours after time of collection. Relevant seminal parameters were compared between patients, with a decrease in 1-hour to 2-hour motility (n = 384) compared with those that showed no change (n = 1929). The same analysis was performed in a subset of patients with a TMSC between 10 and 40 million. In the total patient population, only 16% (384/2313) demonstrated a decrease in 1-hour to 2-hour motility. In patients displaying a decrease in the 1-2-hour motility, sperm concentration (33.5 vs 79 million/mL, P < .0001) and percent normal morphology (7% vs 8%, P < .0001) were significantly lower. Additionally, a significantly higher incidence of 1-2-hour motility decrease was seen in patients with midpiece anomalies (33.3% vs 15.9%, P = .01). Within the subpopulation of 10-40 million TMSC, the only statistically significant difference was in patients with midpiece anomalies (80.0% vs 28.2%, P = .02) who demonstrated a higher incidence of the 1-2-hour motility decrease. Overall, patients with a TMSC between 10 and 40 million showed a significantly higher incidence of 1-2-hour motility decrease compared with the rest of the patient population (29.0% vs 14.6%, P < .0001). Because decreases in 1-2-hour sperm motility affect only a small portion of patients, it is not necessary to check 2-hour motility on all patients. However, because patients with a TMSC between 10 and 40 million were significantly more likely to show a decrease in sperm motility-a decrease that could have possible clinical implications in couples deciding between IUI, IVF, or ICSI--checking 2-hour sperm motility should be considered in this population.     

Effects of ejaculation-to-analysis delay on levels of markers of epididymal and accessory sex gland functions and sperm motility

This study aimed to examine the association between the interval from ejaculation to analysis and epididymal and accessory sex gland function in relation to sperm motility. Ejaculates from 1079 men assessed for infertility were analyzed according to World Health Organization guidelines. Biochemical markers were measured in semen to assess the function of the epididymis (neutral alpha-glucosidase [NAG]), prostate (prostate-specific antigen [PSA] and zinc), and seminal vesicles (fructose). Three groups were defined according to time from ejaculation to analysis: G(< or =30) (24-30 minutes), G(31-60) (31-60 minutes), and G(>60) (63-180 minutes). The proportion of progressively motile sperm was significantly lower in G(>60) than in G(< or =30) (mean difference, 8.0%; 95% confidence interval [CI], 2.0%-13%) or G(31-60) (mean difference, 6.0%; 95% CI, 1.0%-12%). The proportion of rapid progressive sperm motility was significantly higher in G(< or =30) compared with G(31-60) (mean difference, 3.0%; 95% CI, 1.0%-5.0%) and G(>60) (mean difference, 6.0%; 95% CI, 1.0%-10%). Sperm morphology and viability did not vary significantly between the groups. However, PSA levels in G(>60) were 29% and 31% significantly lower than in G(< or =30) (95% CI, 3.0%-54%) and G(31-60) (95% CI, 7.0%-58%), respectively. Moreover, men in G(>60) had 29% and 17% significantly lower zinc compared with those in G(< or =30) (95% CI, 4.0%-69%) and G(31-60) (95% CI, 4.0%-64%), respectively. Levels of NAG and fructose did not differ significantly between the groups. There were negative associations between the ejaculation-to-analysis interval and sperm motility and levels of PSA and zinc. In male infertility assessments, semen analysis should be performed within 60 minutes of ejaculation.     

Monitoring the reactive oxygen species in spermatozoa during liquid storage of boar semen and its correlation with sperm motility,free thiol content and seasonality

Oxidative stress is an important factor affecting the quality of spermatozoa during liquid storage of boar semen; however, monitoring of reactive oxygen species (ROS) that provides direct insight into the oxidative status is not yet attempted. This study aimed to monitor ROS in boar sperm during liquid semen storage to determine its correlation with sperm motility and free thiol (SH) content, and seasonality. Ejaculate was collected from mature Duroc boars in a commercial farm in autumn and spring, diluted in Mulberry III extender, stored at 15°C, and examined daily for sperm ROS level, SH content and motility. The ROS levels in spermatozoa prepared during autumn and spring were constantly low until days 4 and 5 of storage, respectively, which thereafter progressively increased in association with the loss of sperm motility. The increased sperm ROS level correlated with the higher SH level and lower motility, which was accentuated from day 4 of storage and was higher in September, or early autumn. This study indicates that increased sperm ROS levels during liquid storage results in oxidative damage, causing loss of sperm motility, presumably through decreased sperm viability, suggesting that sperm ROS monitoring effectively evaluates the quality of boar semen.     

梯度离心前后精子形态对人工授精妊娠结局的影响

目的:探讨精液处理前后精子形态学变化及其对人工授精结局的影响。方法:回顾性分析185对共228个人工授精周期,精液经密度梯度离心法处理,形态学分析严格按照WHO《人类精液检查与处理实验室手册》第5版标准,比较处理前后精子形态4%和≥4%两组对妊娠结局的影响以及其对女性年龄和不孕年限各组的影响。结果:处理前精子形态4%组,周期妊娠率为14.6%(14/96),处理前精子形态≥4%组,周期妊娠率为12.8%(17/132),无统计学差异(P0.05);处理后精子形态4%组,周期妊娠率为8.7%(6/69),处理后精子形态≥4%组,周期妊娠率为15.7%(25/159),无统计学差异(P0.05)。处理前精子形态4%和≥4%两组在不同女性年龄和不孕年限各组间无差异(P0.05);处理后精子形态4%和≥4%两组,当女性年龄35岁时,妊娠率分别为0(0/39)、14.3%(6/42),差异显著(P0.05);当女性不孕年限5年时,妊娠率分别为0(0/38)、14.0%(6/43),差异有显著性(P0.05)。结论:当处理后精子形态4%,女性年龄35岁或女性不孕年限5年时妊娠率明显降低,应及时行IVF或ICSI治疗。     

Characterizing the reproductive physiology of the male southern black howler monkey, Alouatta caraya

Limited reproductive data are available for any species of howler monkey, including those listed as threatened (Alouatta pigra) and endangered (A. palliata) by the Convention on International Trade in Endangered Species Status (CITES) report. The Southern black howler monkey (A. caraya) is being considered as a model species to develop assisted reproductive technology (ART) for vulnerable howler species. Specific objectives of this study were to evaluate the effect of 1) time of year on ejaculate quality and testosterone concentration, 2) age of male on ejaculate quality, and 3) seminal plasma on sperm longevity in vitro. Three adult (4.5 to 5 years) and 3 subadult (1.5 to 2.5 years) males were evaluated for a 1.5-year period. Semen samples were obtained by electroejaculation, and testosterone levels were monitored by fecal steroid metabolite radioimmunoassay. Males produced coagulum-free ejaculates throughout the year. Likewise, most (4/6) males exhibited constant testosterone levels (3.66 +/- 0.45 ng/g) during the year. Testosterone levels for the remaining 2 males, housed as a bachelor troop, were elevated (43 ng/g) during the months of May and June. Seminal characteristics were similar (P > .05) between age groups. Average semen volume was higher during the summer months (P < .05). Sperm concentrations were highly variable through the year and ranged from 7.0 x 10(6) sperm/mL to 583.0 x 10(6) sperm/mL. Percentages of motile sperm (73% +/- 2.3%) and forward progressive sperm motility (3.3 +/- 0.1), however, were consistent (P > .05) throughout the year. The average pH (8.9 +/- 0.1) and osmolality (356.7 +/- 26.1 mmol/kg) of raw semen also did not vary (P > .05) throughout the year. Ejaculates from subadult males, however, contained more (P < .05) morphologically abnormal spermatozoa than adult ejaculates. In addition, in vitro sperm longevity was poor (<2 hours) for subadult male samples, regardless of the presence or absence of seminal plasma (P > .05). For adult males, seminal plasma was detrimental to sperm longevity; however, spermatozoa survived more than 5 hours in vitro when seminal plasma was removed. Although subadult males produce semen, these ejaculates would not be ideal for further characterization of seminal traits or development of ART for other howler monkey species.     

Caffeine- and kallikrein-induced stimulation of human sperm motility: a comparative study

Stimulation of human sperm motility by caffeine and kallikrein was compared in the same semen material. Caffeine, a cyclic nucleotide phosphodiesterase inhibitor, induced an immediate stimulation of sperm motility the intensity being relatively constant during the first two hours of incubation. Kallikrein, a kinin-releasing proteinase, induced a similar enhancement of total sperm motility, but showed a delayed type of reaction with maximum stimulation at 2 hours of incubation. In contrast to the effect of caffeine lasting some hours, enhancement of sperm motility induced by kallikrein was observed 24 hours. Simultaneous addition of serum (kininogen source) and kallikrein to semen samples led to a sitmulation of total sperm motility with higher mean values than those obtained by caffeine of kallikrein alone. However, the ratio of spermatozoa with very good forward progression was highest during caffeine stimulation. Simultaneous addition of caffeine and kallikrein led to a further improvement of sperm motility which was significantly above that produced by caffeine or kallikrein alone. This observation and the finding of a different response of the spermatozoa of two ejaculates towards caffeine of kallikrein indicate that caffeine (cyclic AMP) interferes quite differently in comparison to kallikrein (kinins) in stimulating sperm motility.     

1985～2008年间我国正常男性精液质量变化分析

目的:研究1985～2008年间国内正常男性精液质量变化趋势。方法:检索历年发表的精子质量方面的文献,系统收集相关资料和数据,利用散点图和直线回归法,分析近24年正常男性精液精子密度、精液量及精子总数的变化。结果:1985～2008年间国内正常男性精液质量参数中精液量没有明显变化(P>0.05)。1985～1994年精子密度和精子总数不存在与时间相关的变化(P>0.05),但自1995年后的14年以来,精子密度和精子总数呈现显著下降趋势(P<0.05),精子密度从1995年81.5×106/ml下降到2008年66.7×106/ml,每年降低1.40%;精子总数从1995年257.2×106下降到2008年的185.9×106,每年下降2.15%。结论:1985～1995年间国内正常男性精液质量未见明显变化,而1995～2008年14年间精子密度和精子总数明显降低,很可能与进入重化工时代引起的日益加重的环境污染有关。     

冻储时间对冷冻精子复苏率的影响

目的 :探讨冻储时间对冷冻精子复苏率的影响 ,以期找到最佳复苏时间。　方法 :取 88份正常供精者标本进行精液常规分析 ,精液用程序降温仪通过三步降温法冻存后置 - 196℃液氮中冻储 ,88份冻存标本随机分成 5组 ,分别于冻存后的 1d(Ⅰ组 ,n =14 )、7d(Ⅱ组 ,n =2 4 )、30d(Ⅲ组 ,n =19)、180d(Ⅳ组 ,n =18)、30 0d(Ⅴ组 ,n =12 )取出 ,37℃水浴复苏 ,再次进行精液常规分析。计算冷冻复苏率。　结果 :5组间的复苏率差异均无显著性 (P均 >0 .0 5 ) ,冻储时间与复苏率之间无相关性 (r=- 0 .0 5 ,P >0 .0 5 )。　结论 :程序降温仪冻存精液标本置 - 196℃液氮后 2 4h即可复苏分析 ,便于人类精子库耐冻实验批量筛查