Study of a group of 484 fertile men Part I: Distribution of semen characteristics

The semen characteristics studied were the sperm count, semen volume, morphology and pre-freeze and post-thaw motility. Two categories of fertile men were investigated: semen donor candidates for artificial insemination and pre-vasectomy subjects. Since mean values for each variable in the two series were similar, they could be considered as a single group of 484 fertile men. Only those subjects whose ejaculates were obtained after an abstinence of 5 days or less were retained. The distribution, mean and percentiles were determined for each variable. The 10th and 90th percentiles for sperm count, percentage of motile forms and percentage of normal cells were respectively 25 and 180 million per ml, 60% and 80% and 50% and 75% The three variables, sperm count, semen volume and total number of spermatozoa which were dependent on abstinence were analysed in the same manner for 3 days of abstinence. The group studied seemed to be as representative and as well defined as possible.     

Kruger strict morphology and post-thaw progressive motility in cryopreserved human spermatozoa

The purpose of this prospective study was to evaluate Kruger strict morphology and conventional semen analysis in predicting cryosurvival and the progressive motility recovery rate of frozen spermatozoa. Our study included 56 semen samples with >10 million spermatozoa per ejaculate. The main outcome measures were conventional semen analysis, strict morphology analysis by the Kruger method, cryosurvival rate and post-thaw sperm motility. A significant reduction in sperm motility after cryopreservation was demonstrated. The freeze-thawing process caused a 66% reduction in rapid progressive motile spermatozoa, a 45% reduction in slow progressive motile spermatozoa and a 2% reduction in nonprogressive motile spermatozoa. The cryosurvival and progressive motility recovery rates were not correlated with parameters of conventional semen analysis, such as sperm concentration, motility, WHO morphology and total motile count, but the progressive motility recovery rate was significantly correlated with the percentage of spermatozoa exhibiting Kruger normal morphology (P = 0.028). The recovery rate of rapidly progressive motility was profoundly decreased compared with slow progressive motility following the frozen-thaw procedure of semen. Kruger strict morphology assessment was a better predictor of the progressive motility recovery rate following the freezing-thaw procedure than parameters of conventional semen analysis.     

Nonsperm cells in human semen and their relationship with semen parameters

The prevalence and clinical significance of leukocytes (WBC) and immature germ cells in semen is currently a matter of controversy. The aim of this work was to assess the prevalence of leukocytospermia in semen samples from Venezuelan men and its possible effects on sperm parameters. The concentration of WBC and round cells (RC) was evaluated in 118 semen samples from 19 fertile subjects (group 1), 62 infertile patients (group II), and 37 men with varicocele (group III). Semen WBC concentration was assessed by peroxidase assay. Twenty-six (22%) of the total samples had more than 10 WBC/mL semen. Twenty of the infertile men had leukocytospermia (32%) compared with 16% in the fertile group and 8% in the varicocele group. Semen RC concentration was lower than 5 x 10(6)/mL in all groups but, in groups II and III was significantly higher compared with group I. Infertile men had the highest WBC concentration. WBC concentration was negatively correlated with progressive motility, percentage of morphologically normal sperm, and hypoosmotic swelling test in infertile men but not in the varicocele group. In this group a negative correlation was obtained between immature germ cells and normal sperm morphology. The data show that leukcytospermia occurs frequently in infertile patients and is associated with poor semen quality parameters. In contrast, in men with varicocele, the increased number of immature germ cells might play a pivotal role in the pathogenesis of abnormal spermatozoa.     

Sperm characteristics and accessory sex gland functions in HIV-infected men

A comparative study was carried out in the andrology clinic, Parirenyatwa Hospital, Harare, Zimbabwe, to investigate the sperm characteristics and accessory sex gland functions in HIV-infected individuals. Sixty-two patients with infertility problems who attended the clinic were requested to donate semen and blood after consent was obtained. HIV antibodies in paired semen and blood samples, sperm morphology, sperm count, sperm motility, seminal leucocytes, seminal fructose, seminal neutral alpha-glucosidase, and citric acid were analyzed. Nine out of 31 blood samples tested positive, while 21 out of 62 semen samples were positive for HIV. Leucocytospermia was associated with HIV-seropositive men (p < .01). The accessory sex gland function, as evaluated by biochemical markers, was not affected in HIV-seropositive men. HIV causes impairment of sperm motility by activating seminal leucocytes, which in turn induce oxidative stress on the sperm. Leucocytospermia is almost always present in HIV-seropositive men.     

Reassessment of the accuracy of traditional sperm characteristics and adenosine triphosphate (ATP) in estimating the fertilizing potential of human semen in vivo

Receiver operating characteristic curves and accuracy parameters were computed for traditional sperm characteristics (concentration, motility, morphology) and the number of peroxidase negative cells, and the concentration of adenosine triphosphate (ATP) in semen from populations of fertile and infertile men, and men who achieved a pregnancy after varicocele treatment. The percentage and concentration per millilitre of spermatozoa with rapid linear progressive motility, and the ATP concentration, provided the best discrimination between fertile and treated fertile from infertile men. The misclassification rate was higher for sperm morphology, total progressive motility and viability, whereas sperm concentration and the total sperm count per ejaculate had the worst discriminating power. The number of peroxidase negative cells per 100 spermatozoa was highly specific in identifying men who achieved pregnancy after varicocele treatment. The lower limit of normality of sperm characteristics was remarkably different between fertile men and men achieving pregnancy after treatment or during infertility work-up.     

Diagnostic value of sperm function tests and routine semen analyses in fertile and infertile men

The results of routine semen analyses, the zona-free hamster oocyte penetration test, the hypoosmotic swelling test, and semen adenosine triphosphate levels were studied in 66 fertile and 130 infertile men. Multivariate discriminant analysis demonstrated that routine semen parameters including semen volume, sperm count, percent sperm motility, and percent normal spermatozoa in combination could predict the fertility of these patients with 70.4% accuracy. Of the three sperm function tests evaluated, the zona-free hamster oocyte penetration test and the hypoosmotic swelling test were selected by the multivariate discriminant analysis as variables capable of providing significant information on the fertility status of the patients. However, the addition of the results of these two tests to the routine semen analysis did not significantly improve the predictability of fertility. The overall correct prediction rate was 77.6% after incorporation of the results of these two sperm function tests. In this group of subjects, the presently available sperm function tests did not predict the fertility status of a patient with a high degree of accuracy.     

上海地区正常生育力男性精液参考值初探

目的:回顾分析上海地区志愿捐精者与正常生育力男性精液分析各项主要参数的分布特征,比较两组男性精液质量的差别,探讨上海地区男性精液参数的正常参考值下限。方法:2010年10月至2011年7月上海市人类精子库招募正常生育力男性41例,健康捐精者100例,按《世界卫生组织人类精液检查与处理实验室手册》(第5版)进行精液常规检测,评估精液体积、精子浓度、前向运动(PR)精子百分率、精子总数和PR精子总数的均值,标准差,并进行t检验。同时统计正常生育力组上述各参数的分布,得出精液特征参数的正常参考值下限。结果:健康捐精组与正常生育力组精液常规各项主要参数(精液体积、精子浓度、PR精子百分率、精子总数、PR精子总数)间差异无统计学意义(P<0.05)。上海地区正常生育力男性精液参考值下限(P<0.05)为:浓度≥27.3×106/ml、PR≥8.1%、体积≥0.82 ml、精子总数≥44.73×106/1次射精、PR精子总数≥24.68×106/1次射精。结论:在评估男性生育力时,精子总数和PR精子总数可能是比精子浓度、精液体积和PR精子百分数更具参考价值的评价指标。     

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.     

Effects of varicocelectomy on sperm DNA fragmentation, mitochondrial function, chromatin condensation, and apoptosis

The aim of this study was to evaluate conventional semen parameters (density, morphology, and progressive motility) and the flow-cytometric parameters of DNA fragmentation, mitochondrial membrane potential, phosphatidylserine externalization, and chromatin compactness in patients with varicocele before and after varicocelectomy. Thirty men (26.5 ± 3.2 years old, range 20-32 years) with oligoasthenoteratozoospermia and grade 3 left varicocele were selected (without other causes of male infertility). Each of them underwent sperm analysis and flow cytometric evaluation before and 4 months after subinguinal microsurgical varicocelectomy (SMV). After varicocelectomy, men had significantly higher sperm density, progressive motility, and normal forms compared with baseline. They also had a significantly lower percentage of spermatozoa with low mitochondrial membrane potential. After SMV, they showed a significantly lower percentage of spermatozoa with phosphatidylserine externalization, an early sign of apoptosis. Significantly decreased percentages of spermatozoa with abnormal chromatin compactness and spermatozoa with DNA fragmentation were found after SMV compared with baseline. Subinguinal microsurgical varicocelectomy improves sperm function in oligoasthenoteratozoospermia secondary to grade 3 left varicocele. Improvements are seen in conventional parameters and biofunctional parameters not routinely evaluated.     

Morphology of spermatozoa in infertile men with and without varicocele

This study was carried out to evaluate the morphology of spermatozoa in infertile men with and without varicocele. A series of 285 ejaculates were fully evaluated for seminal volume, sperm count, motility, and morphology, and classified into fertile (165 subjects), infertile without varicocele (93 subjects) and infertile with varicocele (27 subjects). Sperm morphology was classified by multiple entry criteria and recorded as normal, abnormal with head, midpiece, or tail single anomaly or abnormal with simultaneous multiple abnormalities. Semen volume was almost identical in the three groups. Among the infertile men, sperm count was lower in those having a varicocele, but conversely those with varicocele had a higher percentage of motile spermatozoa, higher percentage of spermatozoa with forward movement and higher sperm velocity. There were higher proportions of spermatozoa with abnormal morphology, total number of anomalies, and multiple anomalies in infertile men, both with and without varicocele, than in fertile men. The percentage of abnormal spermatozoa was higher in infertile men with varicocele than in those without varicocele. The pattern of sperm morphology differed between the infertile and the fertile group, and with the presence or absence of varicocele. In the presence of varicocele, only the incidence of elongated (tapered) forms was significantly increased.     

Clinical significance of reactive oxygen species in semen of infertile Indian men

Reactive oxygen species (ROS) levels in semen are believed to play both physiological and pathological roles in male fertility. The study was aimed to find the clinical significance of ROS levels in infertile Indian men. This pilot study included 33 idiopathic infertile men and 18 proven fertile controls. ROS levels in the washed sperm were measured using chemiluminescence assay and expressed as 10⁶ cpm per 20 million spermatozoa. Sperm count, percent sperm motility, and percent normal sperm morphology were found to be significantly ( P  < 0.0001) reduced in infertile men compared with the controls. Median (minimum, maximum range) ROS levels of the infertile group [24.90 (6.89, 44.71)] were found to be significantly ( P  < 0.0001) elevated compared with the fertile controls [0.167(0.15, 2.78)]. No significant correlation was seen between ROS levels and semen parameters. Elevated ROS levels in the idiopathic Indian infertile men may be one of the underlying reasons for impaired fertility. Therefore measurement of seminal ROS levels may be used in Indian infertile men for better understanding of the aetiology and selection of antioxidant regimen in the treatment of male infertility. However, large studies may be urgently warranted to find out the role of antioxidants in ROS elevated Indian infertile men through randomised, controlled clinical study.     

Effect of leukocytospermia on sperm DNA integrity: a negative effect in abnormal semen samples

Controversy exists over levels of DNA integrity in the sperm of fertile and infertile men. In addition, the effect of leukocytospermia on sperm DNA in these 2 groups is unclear. We decided to address these questions by collecting semen samples from men known or presumed to be fertile and men from infertile couples. Samples were analyzed and assessed for sperm concentration, motility, and morphology. Samples failing to meet World Health Organization (WHO) standards in one or more of these parameters were judged abnormal. Samples were then arbitrarily assigned normalized scores in each of the above parameters, and scores were summed to give a normalized value for overall sperm quality. DNA abnormality was determined by an in situ DNA denaturation test with acridine orange and expressed as a percentage of cells with abnormal DNA integrity (ADI). Assessment of 187 samples revealed a moderate inverse correlation between ADI and sperm quality (r =.58), although a large degree of ADI dispersion was observed in abnormal semen samples. The average ADI for normal and abnormal semen samples was 18% +/- 2.8% and 36% +/- 5.8%, respectively, with the threshold of 95% probability set at 30%. When sorted for leukocytospermia, the difference in ADI between normal and abnormal semen groups without leukocytospermia was much smaller (17% +/- 2.2% and 22% +/- 4.6%; P =.023). Leukocytospermia had no significant effect on ADI in the normal semen group (P = .46); however, ADI was more than double the ADI in the abnormal semen group (18% +/- 2.4% and 50% +/- 11%; P < .001). The results of our analysis show that at least 3 factors affect net DNA integrity in leukocytospermic samples that fail to meet WHO standards: 1) primary DNA damage, which is moderately inverse to sperm quality, in particular to sperm concentration; 2) effect of leukocytes increasing primary or provoking potential DNA damage in a cascade-like manner, particularly in sperm with poor morphology and motility; and 3) a decreasing proportion of cells with damaged DNA in semen with the worst quality.     

Cystic fibrosis transmembrane conductance regulator is correlated closely with sperm progressive motility and normal morphology in healthy and fertile men with normal sperm parameters

Cystic fibrosis transmembrane conductance regulator (CFTR) has been demonstrated to be expressed in mature spermatozoa and correlated with sperm quality. Sperm CFTR expression in fertile men is higher than that in infertile men suffering from teratospermia, asthenoteratospermia, asthenospermia and oligospermia, but it is unknown whether CFTR is correlated with sperm parameters when sperm parameters are normal. In this study, 282 healthy and fertile men with normal semen parameters were classified into three age groups, group (I): age group of 20–29 years (98 cases, 27.1 ± 6.2), group (II): age group of 30–39 years (142 cases, 33.7 ± 2.6) and group (III): age group of more than or equal to 40 years (42 cases, 44.1 ± 4.6). Sperm concentration, total count and progressive motility were analysed by computer‐assisted sperm analysis. Sperm morphology was analysed by modified Papanicolaou staining. Sperm CFTR expression was conducted by indirect immunofluorescence staining. There was a significant positive correlation (P < 0.001) between CFTR expression and sperm progressive motility (r = 0.221) and normal morphology (r = 0.202), but there were no correlations between sperm CFTR expression and semen volume, sperm concentration, sperm total count as well as male age (P > 0.05). Our findings show that CFTR expression is associated with sperm progressive motility and normal morphology in healthy and fertile men with normal sperm parameters, but not associated with the number of spermatozoa and male age.     

Correlation between semen parameters and the Hamster Egg Penetration Test (HEPT) among fertile and subfertile men in Singapore

The objective of this retrospective study was to distinguish between fertile and subfertile men based on their semen parameters and hamster egg penetration test (HEPT) outcome. This study involved 110 subfertile men recruited from an infertility clinic and 48 fertile men attending an antenatal clinic in Singapore. The men were required to donate a semen specimen for semen analysis and HEPT assay. The results indicated that the subfertile group had significantly lower normal sperm morphology according to the Tygerberg strict criteria, and lower progressive motility (P < .05). Semen volume, density, HEPT decondensation rate, and sperm penetration index were not significantly different between the 2 groups. Receiver operating characteristic curve analysis indicated that sperm morphology had the highest predictive power of 65.7% with a threshold value of 7%, and progressive motility had a predictive power of 61.8% with a threshold value of 50%. Using the tenth percentile of the fertile population as the cutoff, lower adjusted thresholds of 3% for sperm morphology and 28% for progressive motility were obtained, giving higher positive predictive values of 81.8% and 84.4%, respectively. This study shows that these new cutoff values can be used to screen the general population to identify subfertile men. In contrast, the HEPT proved to be an insensitive and unreliable assay in identifying subfertile males. To our knowledge the comparison of HEPT and semen parameters between subfertile and fertile men has not been previously reported in an Asian population.     

Effect of freeze–thawing procedure on chromatin stability, morphological alteration and membrane integrity of human spermatozoa in fertile and subfertile men

Cryopreservation is known to impair sperm motility and decrease the fertilization rate by detrimental effects on acrosomal structure and acrosin activity. However, the consequences of cryopreservation on the integrity of the sperm nucleus, chromatin stability and centrosome are less clear. The present study was designed to determine the effect of the freeze-thawing procedure on chromatin condensation (aniline blue staining) and the morphology (strict criteria) and membrane integrity of human spermatozoa. The structural and functional characteristics of the sperm plasma membrane were measured by the eosin-test and hypo-osmotic swelling test which were done separately. Sperm cryopreservation was performed on semen samples from two groups of men classified as fertile (n = 20) and subfertile (n = 72), based on their reproductive history and semen analysis according to WHO guidelines. The mean percentage of condensed chromatin, morphologically normal spermatozoa and membrane integrity in all semen samples investigated (n = 92) decreased significantly (p = 0.0001) after freeze-thawing, in comparison to the value observed prior to freezing. By comparing the semen samples between fertile and subfertile patients, significantly (p = 0.0009) greater damage was demonstrated in the subfertile than in the fertile group. Furthermore, no significant difference was observed between the two groups with regard to the morphological alteration and structural as well as functional damage of the sperm membrane. In conclusion, the freeze-thawing procedure significantly affects chromatin structure and sperm morphology, especially in the head and the tail regions, and this may explain the lower fertilization rate and IVF/ICSI outcome when frozen-thawed spermatozoa are used. In addition, this study demonstrates that chromatin condensation is a sensitive parameter for the evaluation of cryodamage of semen samples from fertile and subfertile patients, though subfertile patients with very poor semen characteristics have yet to be studied. It is therefore recommended that chromatin condensation be used as an additional parameter for the assessment of sperm quality after freeze-thawing.     

Seminal fibronectin in fertile and infertile males

This study aimed to assess seminal plasma fibronectin in fertile and infertile males. Ninety infertile males were investigated; asthenozoospermia (n = 27), asthenoteratozoospermia (n = 30), oligoasthenoteratozoospermia (n = 33) compared with 20 healthy fertile controls. They were subjected to semen analysis, seminal plasma fibronectin estimation by radial immune diffusion, serum testosterone (T) and follicle stimulating hormone (FSH) estimation by ELISA. There was significant increase of seminal plasma fibronectin among different infertile groups compared with the controls. Significant negative correlation was elicited between seminal fibronectin and sperm count, sperm motility grades A, B, A + B, sperm velocity, linear velocity, linearity index, sperm normal forms and serum T. Seminal fibronectin showed significant positive correlation with grade D sperm motility and serum FSH. ROC curve analysis discriminating controls and other infertile groups demonstrated criteria value of < 674 mg l(-1) (sensitivity 100% and specificity 96.4%). It is concluded that increased seminal fibronectin is associated with decreased sperm count and sperm motility.     

Seminal plasma cotinine and insulin-like growth factor-I in idiopathic oligoasthenoteratozoospermic smokers

OBJECTIVE

To assess seminal plasma insulin‐like growth factor‐I (IGF‐I) levels in cigarette smokers with idiopathic oligoasthenoteratozoospermia (iOAT).

PATIENTS AND METHODS

In all, 110 men were divided into fertile healthy non‐smokers, fertile smokers, infertile non‐smokers with iOAT and infertile smokers with iOAT. Semen was analysed, and seminal cotinine and seminal IGF‐I levels estimated.

RESULTS

There were significantly lower seminal IGF‐I levels in the smokers and in men with iOAT than in controls, and in both iOAT groups. Smokers, either fertile or with iOAT had significantly lower levels than in controls in mean semen volume, sperm production index, percentage of motile sperms, rapid linear forward progressive motility, linear velocity and sperm normal forms. Smokers with iOAT had significantly lower levels than non‐smokers with iOAT in mean sperm production index, rapid linear forward progressive motility and linear velocity. In smokers, seminal cotinine was significantly and negatively correlated with both seminal IGF‐I and sperm motility, while seminal IGF‐I was positively correlated with the percentage of motile spermatozoa.

CONCLUSION

Smoking effects on sperm variables could be mediated by decreased seminal IGF‐I.     

Effect of alcohol intake and cigarette smoking on sperm parameters and pregnancy

Much has been published about smoking and alcohol intake influencing male fertility, sperm parameters and reproductive outcome. However, there is no conclusive agreement about the effects of cigarette smoking and alcohol use on these outcomes and thus no generally accepted guidelines. The combined effect of cigarette smoking and alcohol intake, though, has not been rigorously investigated. Because alcohol consumption and smoking are often seen together, this study focuses on the effect of smoking and drinking habits separately and combined on semen parameters, such as volume, sperm count, motility and morphology, and on pregnancy outcome. These suggested toxic effects are studied in a group of subfertile, asthenozoospermic men (<10% motile spermatozoa), compared with a group of ‘proven fertile’, healthy men. The extreme asthenozoospermic group has especially been chosen because of the suspected effect, that is, oxidative stress, on sperm motility. In our study, we found that cigarette smoking and alcohol intake did not differ between the subfertile and fertile group. In conclusion, cigarette smoking and alcohol consumption do not appear to significantly affect sperm parameters, such as volume, sperm count, motility and morphology or pregnancy outcome in our study population.     

Leukocytospermia in male infertility patients in China

Summary. Recent studies have revealed a high prevalence of leukocytospermia (> 1 × 10⁶ white blood cells ml⁻¹ semen) in male infertility patients in the USA and certain European countries, and have implicated white blood cells as a cause of infertility. Since leukocytospermia may often be attributed to male genital-tract infections, its prevalence could vary widely in different populations depending on factors such as sexual practices and the prevalence of sexually transmitted pathogens. In the study described here the incidence of leukocytospermia was determined in a group of 101 male infertility patients and a small reference group of normal fertile men in Beijing, China. Seminal white blood cells (WBC) and WBC sub-populations were enumerated by peroxidase staining and immunohistological assay. Eight out of 101 (7.9%) samples from infertility patients and 0/10 samples from fertile donors were leukocytospermic. The incidence of leukocytospermia in the Chinese infertility patients was considerably lower than the 23% incidence observed in a recent study of infertility patients in the USA using a similar technique. All but one of the patients with leukocytospermia had a poor sperm count and/or poor sperm motility. However, due to the low incidence of leukocytospermia and the small number of patients in this group, a statistically significant association between leukocytospermia and poor semen quality was not attained. The simple peroxidase test correlated well with the more expensive and technically demanding immunohistological assay for detection of white blood cells in semen.     

精子功能检测的研究——Ⅰ.底物膜技术对精子顶体酶的测定

本研究应用健康人和不育症患者的精液各30人份,用底物膜技术检测精子顶体酶,同时进行精子形态学和多次曝光摄片分析。结果表明,在精子头部周围,顶体蛋白酶和透明质酸酶活性区域呈现亮区或晕。不育症患者顶体蛋白酶反应率和反应区的平均直径显著降低(P<0.01),但是透明质酸酶无明显变化(P>0.05).不育症患者精子畸形率高于健康人.不育症患者和健康人精子运动平均速度相似,但直线运动方式的精子在前者明显减少.本文对精子的顶体酶、形态异常和运动等相互关系作了讨论.