正常生育力和精子活力低下不育者精子运动计算机辅助分析

用精子活力分析仪对７３名正常生育力男性射出精子进行了精子动力学指标分析；用上述指标分析了２００例精子活力低下不育男性射出精子动力学指标及各项运动参数间相互关系；用己酮可可碱作用精子活力低下不育男性射出精子，观察精子各项运动参数改变。结果：（１）正常生育力组，前向运动精子在总运动精子中比例与精子活动率间不呈相关性；随精子活动率提高，快速相和慢速相精子分布分别逐渐增加和减少，精子ＶＡＰ、ＶＣＬ、ＶＳＬ、ＢＣＦ、ＡＬＨ增加，ＬＩＮ和ＳＴＲ降低。（２）精子活力低下不育组，随精子活动率下降，主要表现在运动速度下降，与正常组比较，精子前向运动能力和运动速度均有明显下降（Ｐ＜０．０５），但ＬＩＮ和ＳＴＲ不管在组内还是与正常组相比均无明显差别（Ｐ＞０．０５），反映出这种前向运动能力的减低是由速度下降引起，速度分布变化以中速相明显。（３）己酮可可碱能明显提高精子活力低下不育者精子的活动率，前向运动率和精子运动快速相的分布；对精子ＬＩＮ，ＳＴＲ影响不明显。     

Comparison of the Sperm Quality Analyzer IIC variables with the computer-aided sperm analysis estimates

Sperm Quality Analyzer (SQA) IIC, an upgrade version, is an inexpensive device and provides a quantitative estimation of sperm motility, whereas the use of computer-aided sperm analysis (CASA) provides high precision and provision of quantitative data on sperm kinetics. The aim of the present study was to evaluate if the SQA IIC variables correlated with the CASA estimates. Semen quality analysis of 71 fresh semen samples was performed using SQA IIC and CASA. Total sperm concentration, percentage of progressively motile sperm, percentage of normal morphology, motile sperm concentration, sperm motility index (SMI) and functional sperm count (FSC) determinations were performed using SQA IIC. Sperm concentration, sperm motility, and sperm motion variables including amplitude of lateral head displacement (ALH), beat cross frequency (BCF), curvilinear velocity (VCL), straight line velocity (VSL), average path velocity (VAP), linearity (LIN=VSL/VCL), and straightness (STR=VSL/VAP) were evaluated simultaneously on the same semen samples using CASA. The sperm characteristics were compared between SQA IIC and CASA. There were significant correlations of sperm concentration (r=0.634, p < 0.0001), sperm motility (r=0.697, p < 0.0001), and motile sperm concentration (r=0.757, p < 0.0001) between the two devices. Both SMI and FSC significantly correlated with eight CASA estimates, including sperm concentration, sperm motility, motile sperm concentration, ALH, VCL, VSL, VAP, and Rapid. SQA IIC is simple and easy to use. Moreover, the SQA IIC variables well correlated with the CASA estimates. As a screening test for semen quality, SQA IIC is considered as useful in the management of male infertility.     

Human sperm movement assessed with the Hamilton-Thorn motility analyzer and in vitro fertilization

Summary. The relationship between sperm movement characteristics obtained by computerized analysis and the in vitro fertilization rates of human oocytes was studied. In 144 consecutive in vitro fertilization treatments a sample of prepared semen was analysed by a Hamilton-Thorn Motility Analyzer. In addition a visual estimation of sperm count and motility was made. Significant correlations with the fertilization rate were found for all visual parameters. Of the computerized measurements, the mean velocities of motile spermatozoa and the concentration of motile cells were significantly correlated. The average path velocity correlated best ( r = 0.42, P < 0.001). There was no relationship between the percentage of motile sperm showing hyperactivated movement and the fertilization rate. A forward stepwise logistic regression analysis selected the following variables of predictive value for fertilization: average path velocity, male factor infertility as indication for in vitro fertilization, motility and concentration, as measured by the Hamilton-Thorn analyzer. A logistic regression model to predict the cases with low (< 0.2) or high fertilization rates, included the average path velocity as a significant variable and classified the samples with 90% overall accuracy. In conclusion: movement characteristics of spermatozoa in culture medium, especially the average path velocity are of prognostic value in prediction of human oocyte fertilization rates.     

手动或计算机辅助方法评估经间歇性阴囊隔离的公羊的精子活力(英文)

目的:研究用计算机辅助的精液分析(CASA)技术辅助测量得到的精子运动特征是否比手动检测得到的活动的、快速的或前进的细胞百分比能更敏感地指示精液质量下降。方法:6只公羊每天进行16 h 间歇性阴囊隔离,持续21天,对其中2只在次年进行每天12 h 的间歇性阴囊隔离,持续28天。收集其精液并立即进行 CASA 评估,然后冷冻或在30℃或5℃的条件下储藏,用于再次评估。结果:间歇性阴囊隔离导致运动的、快速的和前进的精子百分比下降,与冷冻—解冻或储存在30℃或5℃下的情况相同。只有当运动精子的百分比下降时,通过 CASA 测定的精子能动性(精子侧摆幅度、平均路径速度、平均前进速度和轨迹速度)才降低。冷冻—解冻或液态储存导致从阴囊隔离公羊的精液中有运动能力和快速的精子百分比较之对照组下降幅度更大,但是仅在有运动能力的精子百分比下降时才影响精子的运动特征。结论:间歇性阴囊隔离不仅影响新鲜收集的精子运动,而且影响其抵抗外界储存压力的能力。而通过 CASA技术获得的其他关于运动特征的数据仅是精液质量下降时运动细胞百分比的一个敏感指示。     

Comparison of two sperm preparation techniques using automated sperm motion analysis: migration sedimentation versus swim-up

Migration sedimentation and spermatozoa swim-up techniques were used for obtaining spermatozoa from the semen samples of 39 infertile men. Concentration, percentage of motile sperm, velocity, linearity, and motility index of the sperm preparations obtained by both methods were compared using the CellSoft automated sperm motion analyzer. The mean velocity of the spermatozoa obtained after the migration sedimentation technique was significantly higher than that with swim-up technique. Since it is not necessary to centrifuge spermatozoa with the migration sedimentation technique, this method may be more desirable than other techniques using centrifugation.     

Advanced methods for evaluation of sperm quality

Summary.  Routine semen analysis includes measurements of sperm concentration, motility, and morphology. In our study, three additional tests were evaluated in relation to fertilization rate in an in vitro fertilization program: the freezing and hypo-osmotic swelling tests that evaluate the functional integrity and stability of the sperm membrane under extreme osmotic conditions, and migration sedimentation test that isolates high-quality motile sperm cells. The study was performed on semen delivered by men of couples treated at the In Vitro Fertilization Unit, and men who served as semen donors (fresh or cryopreserved semen). No correlation was found between the sperm fertilization rate, and the decrease in motility percent following the freezing-thawing process. Thus, the freezing test cannot be used to predict semen fertilization capacity. The hypo-osmotic swelling test was applied on semen given by a similar population of men. While the hypo-osmotic swelling test values carried out with fresh semen was found to have good correlation with fertilization, no correlation was found when frozen thawed semen was used. Post-migration sedimentation test sperm characteristics, and especially the recovery rate of the motile sperm, were significantly better in in vitro fertilization cycles with fertilizations. Both the hypo-osmotic swelling test and migration sedimentation test can assist in evaluating semen quality, judged by the fertilization rate in an in vitro fertilization program.     

Comparison of measurements of human sperm motility characteristics by the automated CellSoft system and time-exposure photomicrography

Human sperm motility characteristics in 28 semen samples with sperm concentrations less than 40 x 10(6) ml-1, as determined by the World Health Organization manual analysis (WHO, 1987), were measured by the automated CellSoft semen analyser (Cryo Resources Ltd, New York, NY, USA) using different system parameter settings (Mortimer & Mortimer, 1988a). The results were compared with those obtained by time-exposure photomicrographic (TEP) analysis. It was found that the settings of the minimum video frame rate and the threshold velocity used to distinguish motile from immotile sperm by the automated CellSoft system had a significant influence on measurements of percentage motility but not on linear velocity. At the five different parameter settings used in the present study, the automated CellSoft system gave significantly lower mean values for percentage motility in comparison with the WHO manual and TEP analyses. Measurements for linear velocity between the automated CellSoft system and TEP analyses were found not to be significantly different in these defined semen samples.     

The influence of pentoxifylline on motility and viability of spermatozoa from normozoospermic semen samples

In order to evaluate the effects of pentoxifylline on sperm motility and longevity, a controlled in-vitro study was conducted on normozoospermic donor semen samples using the Cellsoft automated system for sperm motility analysis. After incubation and selection, pentoxifylline was found to improve the recovery of spermatozoa and to increase their velocity. In the subgroup of progressively motile spermatozoa, curvilinear velocity was also enhanced. It is concluded that pentoxifylline has an effect on the vigour, but not on the pattern, of sperm motion. Pentoxifylline did not improve the motility characteristics of senescent spermatozoa in normozoospermic sperm samples. Sperm survival, as shown by supra-vital staining, and motility longevity both decreased with time after pentoxifylline treatment.     

Cryopreservation of human semen. Comparison of cryopreservatives, sources of variability, and prediction of post-thaw survival.

Human semen was cryopreserved using Human Sperm Preservation Medium, TEST-Yolk buffer, or glycerol alone. Sperm characteristics for each specimen were measured before and after freezing to determine which cryopreservative resulted in better cryosurvival and recovery of motile sperm. Sperm frozen in Human Sperm Preservation Medium had a significantly better recovery of all semen parameters (motility, velocity, and recovery) than either TEST-Yolk or glycerol alone. Statistical analyses also were done to examine the variability between and within donor semen specimens. Differences between donors, between specimens, and measurements within donors all contributed to variability of sperm characteristics. Specimen-to-specimen variability for a given donor represented 12% to 47% of the total variability, whereas processing and measurement variability represented 12% to 41%. Donors also varied in the ability of their sperm to tolerate freezing. There was a relationship between motile count after dilution with cryopreservative and post-thaw motile count. This relationship allows the prediction of poor-thaw survival before freezing a specimen.     

Usefulness of sperm quality analyzer-V (SQA-V) for the assessment of sperm quality in infertile men

The aim of this study was to evaluate the correlation of new of Sperm Quality Analyzer (SQA-V) with the computer-assisted sperm analysis (CASA) and manual semen analysis estimates. One hundred five fresh semen samples were analyzed using SQA-V and CASA and manual semen analysis. Significant correlations of sperm concentration (p < 0.0001), sperm motility (p < 0.0001), and normal morphology (p < 0.0001) were observed between SQA-V variables and manual semen analysis estimates. There also were significant correlations of sperm concentration (p < 0.0001), sperm motility (p < 0.0001), and sperm velocity (p = 0.0235) between SQA-V variables and CASA estimates. Meanwhile, it did not correlate with amplitude of lateral head displacement, beat cross frequency, lineality assessed by CASA. The value of the sperm concentration and the sperm motility measured by SQA-V showed high correlations with the value of those measured by CASA and manual semen analysis. In addition, velocity and sperm morphology may also be evaluated to some extent using SQA-V.     

Standardisation of a novel sperm banking kit – NextGen® – to preserve sperm parameters during shipment

Many male patients diagnosed with cancer are within their reproductive years. These men are advised to freeze their spermatozoa prior to the start of cancer treatment. Very often, sperm banking facilities may not be readily available and patients may be required to travel to distant sperm bank centres. Our objective was to design and standardise a remote home shipping sperm kit that allows patients to collect a semen sample at home and ship it overnight to a sperm bank. A total of 21 semen samples and two transport media (refrigeration media and human tubal fluid) and five different combinations of ice packs were tested for maintaining desired shipping temperature. Ten semen samples were assessed for pre‐ and post‐shipment changes in sperm motility, membrane integrity, total motile spermatozoa and recovery of motile spermatozoa. Even though motility, membrane integrity and total motile spermatozoa declined both in samples examined under simulated shipped conditions and in overnight‐shipped samples, the observed motility and total motile spermatozoa were adequate for use with assisted reproductive techniques. Using refrigeration media, cooling sleeve and ice packs, adequate sperm motility can be maintained utilising NextGen^® kit and these spermatozoa can be used for procreation utilising ART techniques such as intracytoplasmic sperm injection.     

Predictive value of CASA parameters in IUI with frozen donor sperm

The objective of this study was to determine if characteristics of sperm motion determined by computer-aided semen analysis (CASA) after thawing and preparation on discontinuous gradient could predict pregnancy outcome after intrauterine insemination (IUI) from frozen donor sperm. A retrospective analysis of 100 non-selected women undergoing 171 consecutive donor insemination cycles was conducted between January 2006 and April 2007. Semen samples from all donors were analysed after thawing and density gradient preparation. Women who became pregnant and those who did not were comparable in terms of age, ovarian stimulation regimen and indication of IUI with donor semen. Pregnancy rate per cycle was 21.8%, and pregnancy occurred after 2.5 IUI cycles on average. Motility parameters of sperm measured by CASA (VAP, VCL, VSL, LIN, STR, and ALH) and total spermatozoa concentration after preparation on discontinuous gradient showed no difference in both groups. Progressive and total motile spermatozoa concentration, as well as progressive and total motile percentages was significantly higher in pregnancy group. The receiver operating characteristic (ROC) curve analysis showed that total motile percentage >17% and motile concentration >0.9 × 10⁶/mL best predicted pregnancy. In a multivariate analysis, only total motility percentage was able to predict pregnancy. Sperm motility parameters of frozen-thawed prepared donor sperm obtained by CASA do not seem to predict pregnancy in IUI cycles. Total motile and progressive percentages and concentrations remain the best prognostic elements for pregnancy in IUI with donor semen.     

Temperature controlled centrifugation improves sperm retrieval

Sperm retrieval techniques form an integral part of the assisted reproductive programme. The success of sperm separation is measured by the number of motile sperm retrieved from a given semen sample. The study aimed to evaluate the effect of temperature during sperm preparation events on the number and percentage motile sperm retrieved following a double wash swim-up procedure. Thirty semen samples were obtained from 10 normozoospermic donors. After collection samples were divided into two aliquots, one aliquot was placed in an incubator at 34 °C, while the second aliquot was left at room temperature (25 °C). Sperm motility assessments were recorded with a computer assisted sperm analyser. Motile sperm fractions were retrieved from the semen samples following a double wash swim-up technique. Two tubes were prepared for each experiment. Tubes were placed in two different centrifuges: (i) SpermFuge (Shivani Industries, India) with temperature centrifuge control (34 °C) and (ii) Sigma with no temperature control facilities. Both centrifuges were set at 484 g for 5 min. Following the second wash, sperm pellets were layered with culture medium, and sperm was allowed to swim up. Supernatants were removed and analysed for sperm concentration and motility values. Percentage motile sperm was transformed to ARCSIN values and results of the two centrifugation methods at 34 °C and room temperature were compared with Mann-Whitney test for independent samples. The mean sperm concentration retrieved at 34 °C was 43.8 ± 50 (SpermFuge) and 32.7 ± 21 (Sigma) (P < 0.05), compared to retrieved concentration at room temperature namely, 30.9 ± 33 (SpermFuge) and 30.6 ± 17 (Sigma) (P ≥ 0.05). The mean percentage motile sperm at 34 °C was 64.0 ± 19 (SpermFuge) and 44.2 ± 24 (Sigma) (P = 0.02), while at room temperature the percentage motile sperm was 54.7 ± 17 (SpermFuge) compared to 46.5 ± 14 (Sigma) (P ≥ 0.05). Centrifuge temperature and incubation temperature significantly influenced the percentage retrieved motile sperm. The use of temperature-controlled sperm preparation might have clinical value for men with poor sperm motility values.     

Computerized videomicrographic analysis of rat sperm motility

Quantitative methods for the determination of the concentration, percent motility and swimming speed of human and animal spermatozoa can assist in the objective analysis of sperm and semen quality. These parameters are among the most discriminating indicators for both clinical and toxicologic assessments of reproductive function. A computerized videomicrographic analysis system to measure sperm motility characteristics in the Fischer 344 rat was characterized and compared with both manual and semi-automated videomicrographic methods (Blazak et al, 1985). The system compares favorably, both in accuracy and sensitivity, to these more conventional methods. The most variable indicator of potential reproductive function in the Fischer 344 rat is the total sperm count from the cauda epididymidis (coefficient of variation [CV] = 24%), while parameters of sperm motility vary least. These include percentage of motile cells (CV = 15%), curvilinear velocity (CV = 9%) and linearity (CV = 10%), which is a ratio of straight-line to total distance traveled. It was concluded that the computerized system may be useful for routine assessment of changes in sperm quality that may occur in the rat after exposure to toxic drugs or chemicals.     

Freezability and semen parameters in candidates of sperm bank donors: 1992-2010

There has been considerable concern worldwide about possible semen quality deterioration over the last 2 decades. The aim of this study was to evaluate freezability and semen quality of healthy young males during the years 1992-2010. A total of 1211 young (20-32 years old) candidates for sperm bank donation were recruited into the study with no exclusion criteria. They were instructed to observe 2 to 3 days of abstinence from sexual activity, and most of them supplied 2 specimens each. Average values of the various semen parameters, including freezing survival, were calculated for each participant. The change in different semen parameters over years, according to yearly and monthly average temperatures, was evaluated by SAS PROC SURVEYREG analysis. During that period, there were significant increases in motility and vitality percentages, as well as in the percentage of thawed sperm motility. The parameters of volume, concentration, normal morphology, total count, and total motile count showed a significant decrease with years (P < .01). The significant increase in average yearly temperature (P < .004) had limited, nonsignificant association with any of the semen variables. However, average monthly temperature contributed significantly to the trend of semen quality parameters (ie, specimen volume, concentration, percentage of normal morphology, and thawed motility). To the best of our knowledge, this is the first demonstration of the occurrence of an improvement in percent thawed motility over the years, and its significance lies in enabling a higher proportion of sperm bank candidates to be suitable for donation. It is suggested that the global warming phenomenon might have only partial contribution to semen variable changes over the years.     

人类精液冷冻前后精子运动变化的计算机辅助分析

目的 :利用计算机辅助精液分析 (CASA)系统分析冷冻前后人类活动精子的运动参数 ,并观察相关参数的变化规律。　方法 :2 38份精液样本在冷冻前和复苏后均采用CASA分析其运动参数。　结果 :复苏后精子活动率下降 ;除精子鞭打频率 (BCF)外 ,冷冻前与复苏后全部精子运动参数呈显著正相关且差异存在显著性 ;平均路径速度 (VAP)、直线速度 (VSL)、曲线速度 (VCL)下降 ,精子头侧摆幅 (ALH)下降 ,但直线性 (LIN)、前向性 (STR)升高。　结论 :CASA系统可对冻融前后精子运动参数进行客观细致的分析 ,有一定的临床应用价值。     

男性血液和精液中微量元素含量与精子活力的相关性研究

目的:探讨男性血液和精液中微量元素的分布及其与男性精液参数的相关性。方法:采用BH-5100五通道原子吸收仪、YY-1001血铅仪和伟力彩色精子自动分析仪,对113例男性血液和精液进行钙、镁、铜、锌、铁、铅的测定及精液常规分析,以了解微量元素和精液参数的相互关系。结果:①血液与精液中铅、铜、锌、铁含量显著相关,其中以铜的相关系数最大;钙、镁没有相关性。②血液和精液中铅、铜、锌、钙、镁、铁6种微量元素分布存在显著性差异。③精液中钙与精子活率、直线运动精子活率、曲线速度、平均移动角度、活跃精子密度、直线速度、平均路径速度、侧摆幅度等指标显著正相关;铅与精子活率、直线运动精子活率、活跃精子密度、活动精子密度、直线运动精子密度等指标呈显著负相关;镁元素与精子活率、直线运动精子活率、直线运动精子密度呈显著负相关;精液酸碱度与钙、镁、铁3种元素呈显著负相关。④血液中的锌与精子活率和直线运动精子活率呈显著负相关,镁与精子活率、直线运动精子活率、平均路径速度呈显著负相关,铅与侧摆幅度呈显著正相关。结论:不同体液中的微量元素分布和精子的运动功能存在不同的相关性。     

Influence of sperm processing on the fertilizing capacity and recovery of motile sperm from thawed human semen

Artificial insemination programs now rely almost exclusively on frozen semen preparations as their source of sperm. Unfortunately, several reports indicate that conception rates using frozen-thawed semen are inferior to freshly ejaculated specimens. The present study was designed to investigate the fertilizing capacity of sperm and the recovery of motile sperm from thawed semen following different sperm processing methods. Washed resuspended pellets contained sperm with at least the same fertilizing potential as sperm from swim-up techniques. However, the recovery of motile sperm from the pellets was more than 3 times greater than from the swim-up techniques. Percentage motility, progressive velocity, and amplitude of lateral head displacement were generally higher in the sperm from swim-up techniques than in the sperm from the pellets, despite equivalent fertilizing potentials. It can be argued from these results that washed resuspended sperm preparations provide a significantly greater number of motile sperm without a loss in fertility when compared to sperm from swim-up techniques. Accordingly, this study raises questions about the use of the sperm swim-up as a procedure for processing thawed semen for use in intrauterine insemination in which maximal numbers of motile sperm are required. It also demonstrates the need to identify new methods for processing frozen-thawed specimens for assisted reproductive procedures.     

Repeated vitrification/warming of human sperm gives better results than repeated slow programmable freezing

In this study, we compared the effects of repeated freezing/thawing of human sperm by our in-house method of rapid freezing with slow programmable freezing. Sperm samples from 11 normozoospermic subjects were processed through density gradients and divided into three aliquots: non-frozen, rapid freezing and slow programmable freezing. Sperm in the rapid freezing group had better motility and viability than those in the slow freezing group (P<0.01) after the first, second and third cycles of freezing/thawing, but there was no difference in morphology. In the second experiment, rapid freezing was repeated three times in 20 subjects. The samples from each thawing cycle were evaluated for DNA fragmentation using the alkaline comet assay. DNA fragmentation began to increase considerably after the second cycle of freezing/thawing, but to a level that was not clinically important. In the third experiment, rapid freezing was done repeatedly in 10 subjects, until no motile sperm were observed after thawing. The median number of repeated freezing/thawing that yielded no motile sperm was seven (range: 5–8, mean: 6.8). In conclusion, we demonstrated that repeated freezing/thawing of processed semen using our rapid freezing method gave better results than standard slow programmable freezing. This method can help maximize the usage of precious cryopreserved sperm samples in assisted reproduction technology.     

Relationship between morphology and motion characteristics of human spermatozoa in semen and in the swim-up sperm fractions

In this study, the authors evaluated the morphology pattern and motion characteristics of human spermatozoa before and after swim-up separation. Samples were divided into two, morphologically different groups according to the percentage of normal sperm forms assessed by the strict criteria of the Norfolk laboratory: "good"(G) and "poor" (P) prognosis patterns. The percentages of normal forms, slightly abnormal forms, and severe head defects were significantly different in the two groups. Motile characteristics were analyzed by a computerized semen analyzer with constant parameter settings. Before swim-up there were no significant differences in semen volume, percentage of neck and tail defects, concentration, or percentage of motility and linearity, but the mean velocity was higher in group G. After swim-up the percentage of motility, total number of motile cells, and recovery rate were higher for group G, and the incidence of severe head defects correlated negatively with the percentage of cells with a velocity of greater than 80 microns/sec. The results suggest that patients with a high incidence of sperm head defects have impaired original velocity, and swim-up selects for velocity as well as normal forms and motility. Although motility and velocity improved substantially after swim-up, the recovery rate and percentage of motility were significantly lower in the P group.