精子DNA荧光分析与两种常规精液分析方法的比较

目的 :比较计算机辅助的DNA荧光染色精子分析系统 (简称荧光CASA)与两种常规精液分析方法。　方法 :随机选择 2 2例男性不育患者 ,采用荧光CASA、灰度CASA和人工技术分析精液 ,以荧光CASA检测的精子密度为基础 ,分别与常规精液分析及灰度CASA的结果进行比较。　结果 :荧光CASA检测可以区别非精子颗粒及死活精子。与荧光CASA分析方法获得的精子密度相比 ,常规精液分析和灰度CASA法获得的精子密度差值绝对值的平均值分别为 (9.2 3± 8.0 1)× 10 6/ml和 (10 .2 7± 6 .2 2 )× 10 6/ml,检测精子密度误差的百分率分别为 (4 9.0 6±4 9.87) %和 (4 3.39± 2 5 .5 6 ) %。　结论 :荧光CASA更符合最新WHO男性不育实验室诊断标准的要求 ,推荐在临床工作中使用DNA荧光染色精子分析系统来分析精子特性 ,尤其是精子密度分析。     

生育与不育男性精子核DNA成熟度的比较研究

通过对１９对生育夫妇和７２对不育夫妇丈夫精液中绿色吖淀橙（ＡＯ）荧光精子（成熟精子）比率分布情况的比较分析，探讨了绿色ＡＯ荧光精子百分率与精液参数及精子授精能力之间的关系。在１～３年随访中，通过人工授精、其他治疗或在非治疗期，５４对绿色ＡＯ荧光精子≥５０％的夫妇有１８对怀孕，而１８对绿色ＡＯ荧光精子＜５０％的夫妇无１例怀孕。结果还显示了绿色ＡＯ荧光精子百分率与精液参数无关。认为该项指标从精子核ＤＮＡ成熟状态角度反映了单凭精液分析所不能反映的男性生育力。精液中绿色ＡＯ荧光精子＜５０％，是低生育力精液的评估界限     

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.     

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.     

JC-1单标法流式细胞术检测精子线粒体膜电位的研究

目的:探讨应用荧光染料JC-1单色标记法进行流式细胞术检测精子线粒体膜电位(MMP)的可行性及其临床意义。方法:收集63例男性精液标本,分为生育组(n=31)和不育组(n=32)。通过计算机辅助精液分析系统进行精液常规分析。精液标本洗涤处理后用JC-1染色后上流式细胞仪分析,用发橙红色荧光精子百分率(JC-1+%)表示MMP正常精子的比例。结果:生育组精子JC-1+%为(75.89±15.69)%,显著高于不育组[(54.04±22.21)%,P=0.000]。63例标本中,JC-1+%与精子活动率呈显著正相关(r=0.610,P=0.000),与(a+b)级精子百分率呈显著正相关(r=0.614,P=0.000),与d级精子百分率呈显著负相关(r=-0.504,P=0.000)。JC-1+%与已建立的罗丹明/碘化吡啶双染法检测结果(Rh123+/PI-%)呈显著正相关(r=0.938,P=0.000)。结论:应用流式细胞术JC-1单标法检测精子MMP具有可行性,精子JC-1+%可作为男性不育的辅助诊断指标。     

Comparison of the penetration ability of human spermatozoa into bovine cervical mucus and zona-free hamster eggs

In vitro bovine cervical mucus (BCM) penetration tests, sperm penetration assays (SPA) using zona-free hamster eggs, and routine semen analyses were performed on a total of 136 freshly collected semen samples from men who were seen at an infertility clinic. The correlations between bovine cervical mucus penetration and other semen parameters were the percent motile spermatozoa (r = 0.48), progressive motility grade (r = 0.44), sperm count (X 10(6)/ml) (r = 0.47), the percent normal morphology (r = 0.32) and the percent eggs penetrated (r = 0.46) (P less than 0.0001 for each correlation coefficient). When known fertile (n = 32) and infertile (n = 18) groups were tested, positive mucus penetration was associated 75% correctly and positive egg penetration was associated 90% correctly to clinical status. The mucus test had no false-negative results and the SPA had no false-positive results in these groups. It appears, then, that the mucus test and sperm penetration assay, although contributing different elements of data to an infertility evaluation, are both useful adjuncts to a semen analysis.     

流式细胞术在精子功能检测中的应用

目的:研究流式细胞术在精子功能评价中的作用。方法:对104例男性不育患者和10例正常生育男性的精液标本进行精液量、精子密度、活动率、畸形率等精液常规分析(CASA法),对精子存活率、染色质结构和线粒体膜电位等进行流式细胞术分析,并将检测结果通过SAS软件进行统计学分析。结果:成组U检验结果表明:不育患者的精子与正常生育男性精子相比除了具有密度低(U=2.51,P=0.014 3)、活动率差(U=3.44,P=0.001)、畸形率高(U=-5.88,P<0.000 1)等特点外,经流式细胞术测定精子结果表明不育男性的精子质膜完整率(U=4.72,P<0.000 1)、线粒体膜电位正常率(U=-2.53,P=0.030 9)和染色质完整率(αt:U=-3.82,P=0.000 3;SDαt:U=-3.98,P=0.000 1;COMPαt:U=-3.57,P=0.000 5)均显著低于正常生育男性。多元逐步回归分析结果表明:精子质膜完整性与精子活动率(t=1.66,P=0.101 6)、精子的线粒体膜电位与精子活动率(t=3.33,P=0.001 4)、精子染色质的完整性与精子活动率(t=3.24,P=0.001 9)均呈正相关,而精子的线粒体膜电位与精子颈、尾部的畸形率呈负相关(t=-3.44,P=0.001)。结论:流式细胞仪测定对精子质量评定意义显著,对男性不育的诊断和治疗也具有较高的参考价值。     

不育病人精浆胆固醇酯转运蛋白的检测

目的 :检测不育病人精浆胆固醇酯转运蛋白 (cholesterolestertransferprotein ,CETP)的含量 ,并探讨其与不育的关系。　方法 :随机选择 163例不育病人及 15例生育男性 ,行精液常规分析及精浆CETP浓度测定 ,其中5 5例不育病人同时测定了血清CETP含量。　结果 :不育病人及生育男性精浆CETP含量分别为 ( 2 .2 1± 1.2 3 )μg/L和 ( 1.40± 0 .45 ) μg/L ,两组间差异无显著性 (P >0 .0 5 ) ;在不育病人中 ,无精子症组 (n =2 9)、少弱精子症组(n =5 8)、少精子症组 (n =15 )、弱精子症组 (n =44 )及正常精子症组 (n =17)间CETP含量差异也无显著性 (P >0 .0 5 )。精浆平均CETP含量仅占血清的 1/ 10 0 0 ,不育病人精浆及其血清CETP含量间并无相关性 (r =0 .0 0 9,P>0 .0 5 )。　结论 :精浆CETP含量极低 ,与精子密度、活率的改变无关 ,可能保证了精子在未进入女性生殖道前膜结构与功能的完整性     

Postoperative changes of sperm chromatin heterogeneity, using acridine orange staining, in varicocele patients

The aim of this study was to evaluate postoperative changes in sperm chromatin heterogeneity in varicocele patients. In 15 infertile patients with varicocele, sperm parameters including concentration, motility, and morphology were evaluated before and after surgical correction of varicocele. Sperm motion analysis using computer-assisted semen analyzer (CASA) was also performed. To analyze the sperm nuclear proteins, the acridine orange staining method was used. On semen analysis, sperm concentration and motility significantly increased after surgery (p = 0.002, p = 0.003, respectively), although sperm morphology was unaltered postoperatively. CASA parameters, including velocity, linearity, amplitude of lateral head displacement and beat cross frequency were unaltered postoperatively. On the other hand, acridine orange staining significantly increased postoperatively (p = 0.002). Varicocele influences the sperm chromatin condition, as well as sperm concentration and motility.     

两种精子分析系统的比较研究

目的:评估自动化精子质量分析仪SQA-V与计算机辅助精液分析(CASA)系统之间各主要参数的差异性及其在不育和生育男性精液质量分析中的应用。方法:用SQA-V和CASA系统分别检测12例正常生育者和73例不育患者新鲜精液标本,分别分析精子密度、精子活动率、活动精子浓度、精子活动力指数、精子头侧向位移、鞭打频率、精子曲线运动速度、精子前向运动速度、平均路径速度、直线性及前向性运动速度等参数,针对两者进行相关性分析。结果:不育组和生育组男性各项指标间存在显著差异,两种分析系统检测的精子密度之间(r=0.58,P<0.01)、活动精子浓度之间(r=0.75,P<0.01)、平均精子速度(SQA-V)和精子路径速度(CASA)之间(r=0.59,P<0.01)具有较好的一致性,SQA-V的活动精子指数(SMI)和CASA的前向性、曲线运动速度、精子运动的直线性、前向运动速度、平均路径速度、鞭打频率等参数具有显著相关性(P<0.05)。结论:SQA-V和CASA系统比较,在各主要参数上具有较好的一致性,能够反映临床不同组群患者精液的差异性。     

无症状不育病人精液抗沙眼衣原体IgG、IgM类别抗体的检测

目的 :探讨无症状不育病人精液抗沙眼衣原体 (Chlamydiatrachomatis,CT) IgG、 IgM抗体检测的临床意义。　方法 :随机选择 116例无生殖道感染症状的不育病人及 18例生育男性 ,作精液常规参数分析 ,然后分离精浆 ,测定其抗CT IgG、 IgM抗体。 　结果 :①不育病人精浆抗CT IgG、 IgM抗体阳性率分别为13 .8% ( 16/ 116)和3 .4% ( 4/ 116) ,而生育男性分别为 11.1% ( 2 / 18)和 0 ,两组相比差异均无显著性 (P均 >0 .0 5 )。② 116例不育病人中有 2 2例为无精子症。其余 94例病人中 ,精子密度异常组的精浆抗CT IgG、 IgM抗体阳性率分别为2 1.4% ( 6/2 8)和 7.1% ( 2 / 2 8) ,精子密度正常组则较低 ,分别为 12 .1% ( 8/ 66)和 3 .0 % ( 2 / 66) ,但差异均无显著性 (P均 >0 .0 5 ) ;精子活率正常组和异常组之间的精浆抗CT IgG、 IgM抗体阳性率也无明显差异 (P均 >0 .0 5 )。③ 116例不育病人中 3 0例CT阳性 ( 2 5 .9% )。　结论 :无症状不育病人的精浆抗CT IgG、 IgM抗体阳性率与生育男性相似 ,与其精液参数改变无关 ,不能作为CT感染的辅助指标     

Evaluation of Hamilton-Thorn automated semen analysis system.

Sperm concentration and percentage motility values generated by the HT M-2030 system (CASA) were compared with those obtained by subjective semen analysis (SSA). Three calibrations arbitrarily designated as A, M, and D for gates and discriminators and three calibrations arbitrarily designated LI, MI, and HI for default pixel count (DPC) and default intensity (DI) were evaluated. The best correlation between CASA and SSA was observed utilizing M calibration (n = 114) with which sperm concentration was +0.3 x 10(6)/mL (r = .96) and motility was -6.3% (r = .89) compared to the values obtained by SSA. It was found that 35.9% of sperm concentration values and 34.2% of sperm motility values were within 10% of the values obtained by SSA. When sperm concentration was between 50 and 100 x 10(6)/mL the difference in motility was reduced (-3.2%) while the difference in sperm concentration was reversed (-2.6 x 10(6)/mL). LI calibration (DPC = 4, DI = 86) gave acceptable results with M calibration for sperm concentration (+2.1 x 10(6)/mL) and motility (-6.9%) compared to the values obtained by SSA. In the presence of sperm clumping, the difference between CASA and SSA was reversed for sperm concentration (+0.56 x 10(6)/mL for normal samples vs. -2.2 x 10(6)/mL for samples with clumping) and was reduced for sperm motility (-7.14% vs. -4.55%, respectively). HT M-2030 under proper calibration can be used as a rapid, objective, and reliable alternative to conventional semen analysis in routine and for research purposes.     

Relationship between hypoosmotic swelling test, semen analysis, and zona-free hamster ovum test

The hypoosmotic swelling test is a simple test for measuring the functional competence of the human sperm membrane. Fifty-four patients with idiopathic infertility were assessed by hypoosmotic swelling test and the results were compared with those of routine semen analysis and zona-free hamster ovum human sperm penetration test (ZSPT). Semen samples with abnormal semen parameters had lower percentage of swollen sperm in comparison with those with normal semen parameters. A positive correlation was observed between sperm concentration and sperm swelling (r = .50, p less than .05). A strong positive correlation was observed between the percentage of sperm motility and the percentage of swollen sperm (r = 0.60, p less than .01), and between motile sperm concentration and sperm swelling (r = .62, p less than .01). On the other hand, sperm swelling correlated only weakly with the percentage of sperm penetration. The results indicate that the hypoosmotic swelling test appears to evaluate different functional qualities of sperm than ZSPT.     

Accuracy of the normal sperm morphology value by Sperm Quality Analyzer IIC: comparison with the strict criteria

The aim of the present study is to investigate the accuracy of the normal sperm morphology value by Sperm Quality Analyzer IIC (SQA IIC), which was developed to provide a rapid and low-cost quantitative evaluation of semen quality. Normal sperm morphology was assessed using SQA IIC in comparison with that by the strict criteria in 62 semen samples. Normal sperm morphology value by SQA IIC was based on the studies of three traditional sperm parameters from over 4000 fresh, untreated semen samples, while the strict criteria was based on the method by Kruger et al. The mean +/- SD of percent normal morphology by SQA IIC and the strict criteria were 37.6 +/- 10.9% (range 15-52) and 19.9 +/- 8.2 (range 1-34), respectively. There was a significant correlation of the sperm morphology assessment between the two methods (r=0.454, p < 0.001). Using the cut-off value of >30% normal morphology by SQA IIC, the positive predictive value and the negative predictive value of the 'normal' strict criteria were 79.6% (39/49) and 46.2% (6/13), respectively. These results indicate that SQA IIC might be used as an initial screening test for the evaluation of sperm morphology. However, sperm morphological assessment by the strict criteria should be performed in order to make decisions in planning strategies for the treatment of infertile couples.     

不育患者精浆α-1,4糖苷酶活性与精液参数的关系

目的　探讨精浆 α-1 ,4糖苷酶活性与精液参数之间的关系。　方法　分光光度比色法测定精浆 α-1 ,4糖苷酶活性及进行精液常规分析。　结果　 2 90 2例男性不育者精浆α-1 ,4糖苷酶活性异常率为 3 8.87%。该酶活性与精子密度、精子活率、a,b级精子活力和顶体酶活性呈显著正相关 ( r分别为 0 .460、0 .1 2 2、0 .0 86和 0 .2 3 0 ,P均 <0 .0 0 1 ) ,而与精液量、精液 p H、液化时间和畸形精子率无显著相关 ( P>0 .0 5)。 α-1 ,4糖苷酶活性正常组精子密度、活率、a,b级精子活力和精子顶体酶活性均明显高于 α-1 ,4糖苷酶活性异常组 ( P<0 .0 0 1 )。以常规精液分析法 ( RSA)主要参数正常与否分成的两组间α-1 ,4糖苷酶活性差异有显著性 ( P<0 .0 0 1 )。　结论　α-1 ,4糖苷酶活性对精子密度、活率、a,b级精子活力和顶体酶活性均有明显影响 ,对精液量、精液 p H、液化时间和畸形精子率无显著影响     

Defining the "normal" postejaculate urinalysis

Although sperm have been shown to be present in the postejaculate urinalysis (PEU) of both fertile and infertile men, the number of sperm present in the PEU of the general population has never been well defined. The objective of this study was to describe the semen and PEU findings in both the general and infertile population, in order to develop a better appreciation for "normal." Infertile men (n = 77) and control subjects (n = 71) were prospectively recruited. Exclusion criteria included azoospermia and medications known to affect ejaculation. All men underwent a history, physical examination, semen analysis, and PEU. The urine was split into 2 containers: PEU1, the initial voided urine, and PEU2, the remaining voided urine. Parametric statistical methods were applied for data analysis to compare sperm concentrations in each sample of semen and urine between the 2 groups of men. Controls had higher average semen volume (3.3 ± 1.6 vs 2.0 ± 1.4 mL, P < .001) and sperm concentrations (112 million vs 56.2 million, P = .011), compared with infertile men. The presence of sperm in urine was common in both groups, but more prevalent among infertile men (98.7% vs 88.7%, P = .012), in whom it comprised a greater proportion of the total sperm count (46% vs 24%, P = .022). The majority of sperm present in PEU were seen in PEU1 of both controls (69%) and infertile men (88%). An association was noted between severe oligospermia (<5 million/mL) with low semen volume (<0.5 mL), and significant sperm counts in PEU (<5 million). Although infertile men tend to have a higher proportion of their total sperm in the urine compared with control, there is a large degree of overlap between the 2 populations, making it difficult to identify a specific threshold to define a positive test. Interpretation of a PEU should be directed by whether the number of sperm in the urine could affect subsequent management.     

A critical method of evaluating tests for male infertility

Considerable uncertainty surrounds the selection of test values that separate infertile from fertile men in the evaluation of male infertility. We herein describe an objective method of determining these values, referred to as threshold values, for different infertility tests. Using test results from fertile men threshold values were chosen such that 96 per cent of the semen samples from the fertile men were scored as fertile. These threshold values then were used to evaluate 100 semen samples from 74 men presenting for evaluation of infertility. Using this method we constructed infertility profiles on each of the 100 semen samples presented for infertility evaluation and found that the zona pellucida-free hamster egg penetration test (a measure of a spermatozoon's ability to undergo capacitation and penetrate an egg) identified 66 per cent of these samples as infertile, while multiple exposure photomicrography (a quantitative measure of sperm motility) identified 54 per cent of these samples as infertile. This compares with results from routine semen analyses using the same method, which identified none of the samples as infertile by sperm motility grade, 1 per cent by semen pH, 4 per cent by the percentage of motile sperm, 7 per cent by the total count of motile sperm, 10 per cent by the total sperm count, 11 per cent by the semen leukocyte concentration, 12 per cent by the concentration of motile sperm, 13 per cent by ejaculate volume, 16 per cent by sperm concentration and 27 per cent by sperm morphology. This method of analyzing infertility test results provides insight into the potential causes of male infertility and offers a critical approach towards understanding the complex problem of male fertility dysfunction.     

Laboratory methods for the diagnosis of asthenozoospermia

Human sperm samples from 46 men were evaluated for standard semen parameters by two trained technicians, according to WHO criteria. A computer-aided semen analyzer (CASA, HTM-IVOS) was employed. The overall mean coefficient of variation for the 2 participants and the 46 samples studied was 17% for the percentage of progressive motile spermatozoa (r = 0.77). Twenty-nine (63%) samples were classified as asthenozoospermia by both methods. From the studied samples, 12 (26%) were classified as asthenozoospermia by the subjective method and 2 (4.3%) by CASA (p =.01). The two methods do not provide directly comparable data.     

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)     

溶脲脲原体感染对男性不育患者精子质膜完整性的影响

目的:应用荧光染料SYBR-14/碘化丙锭(PI)双标法流式细胞术检测溶脲脲原体(Uu)感染对男性不育患者精子质膜完整性的影响。方法:收集63例男性精液标本,分为Uu感染组(n=32)和正常对照组(n=31)。通过计算机辅助精液分析系统进行精液常规分析,培养法进行Uu检测。精液标本经PBS洗涤处理后用荧光染料SYBR-14/PI双染后上流式细胞仪分析,用发绿色荧光精子百分率(SYBR-14+/PI-%)表示质膜完整精子的比例,检测精子质膜完整性。结果:Uu感染组精子质膜完整性[(45.14±10.69)%versus(72.68±9.91)%]和(a+b)级精子百分率[(23.29±8.81)%versus(46.32±9.54)%]均显著低于正常对照组(P<0.01),两组间精液量、pH值、精子浓度差异无显著性(P>0.05)。结论:Uu感染引起精子质膜完整性下降,可能是导致男性不育的重要原因之一。