两种精液处理方法对精液参数及冷冻效果的比较分析

目的 比较两种精子处理方法的优劣.方法 55例精液标本随机分为直接离心组(30例)和加精子洗涤液离心组(25例),进行精液处理后并且冷冻,观察其复苏效果,所有样本均进行精液常规分析.结果 直接离心组精子密度由46.52×106/ml提高到86.50×106/ml,而前向运动精子(a+b)级百分率则由58.00%下降到44.33%;加精子洗涤液离心组精子密度由44.44×106/ml提高到85.96×106/ml,而前向运动精子(a+b)级百分率则由57.76%下降到46.48%,两组间均无显著性差异(P>0.05).对两组处理后的精液进行冷冻,复苏后得到其前向运动精子复苏率均在40%左右,两组间差异无统计学意义(P>0.05).结论 两种处理方法具有相近的浓缩效果和前向运动精子复苏率,直接离心法则更为简便且节省成本.     

肌酸磷酸对冷冻复合后精子的活率和存活率的影响

肌酸磷酸和肌酸磷酸激酶在精子的能量再生和转移中发挥重要作用。西方研究了２０例冷冻复苏后精液标本，在起始时间，０时间，６０分钟，１２０分钟和１８０分钟采用计算机辅助精液分析仪和荧光分子探针方法测定精子的活率与存活率。实验结果表明，加ＣＰ的实验组与ＨＴＦ对照组，在６０分钟采用２０分钟和１８０分钟进行比较，发现精子活率与存活率ＣＰ实验组明显高于ＨＴＦ对照组，两组间有极显著性差异，提示ＣＰ有延长精子活动的     

冷冻前平衡处理对人精子冻融后相关参数的影响

目的:研究冻前平衡处理对冷冻精子复苏率的影响,优化一步熏蒸法,以寻找人类精子冷冻的最优方法。方法:使用50例自愿供精者的正常精液标本,同1例供精者的精液一式3份,分别采用不平衡直接冷冻法、室温平衡10 min后冷冻与4℃平衡10 min后冷冻的方法进行处理。复苏后用计算机辅助精液分析(CASA)系统分析3组标本冷冻前后精子运动参数的变化,并检测各组标本精子的畸形率和存活率。结果:4℃平衡组的前向运动精子复苏率[(61.88±16.94)%]显著高于未平衡组[(48.61±16.44)%]和室温平衡组[(49.41±13.77)%](P<0.05),而室温平衡组与未平衡组前向运动精子复苏率则无显著性差异。3组标本经冷冻复苏后其精子畸形率和存活率无显著性差异。结论:冻前4℃平衡10 min的冷冻前处理方法操作简单,实验条件易控,对提高精子复苏率有重要意义。     

肌酸磷酸对冷冻复苏后精子的活率和存活率的影响

肌酸磷酸和肌酸磷酸激酶在精子的能量再生和转移中发挥重要作用。本文研究了２０例冷冻复苏后精液标本，在起始时间、０时间、６０分钟、１２０分钟和１８０分钟采用计算机辅助精液分析仪和荧光分子探针方法测定精子的活率与存活率。实验结果表明，加ＣＰ的实验组与ＨＴＦ对照组，在６０分钟、１２０分钟和１８０分钟进行比较，发现精子活率与存活率ＣＰ实验组明显高于ＨＴＦ对照组，两组间有极显著性差异（Ｐ＜０．０１），提示ＣＰ有延长精子活动的时间及存活时间，此外，延缓精子活率和存活率的快速下降     

冷冻保护剂与精液比例对冷冻复苏后精子活动力的影响

目的:比较冷冻保护剂与精浆的不同比例对人类精子冷冻复苏后活动力的影响。方法:将57例志愿者的精液标本分别采用冷冻保护剂∶精液=1∶1和冷冻保护剂∶精液=1∶3的体积比进行冷冻,比较精子复苏后的前向活动率和总活动率。结果:冷冻前的前向运动精子百分率和总活动率分别为(58.60±5.57)%和(66.17±5.24)%。采用冷冻保护剂∶精液=1∶1比例进行冷冻复苏后的前向运动精子百分率和总活动率分别为(40.53±8.97)%和(51.23±9.30)%;采用冷冻保护剂∶精液=1∶3比例进行冷冻复苏后的前向运动精子百分率和总活动率分别为(44.70±8.67)%和(51.50±7.40)%。冷冻前后精子的前向运动百分率和总活动率差异有显著性(P<0.05)。两种不同比例的保护剂相比冷冻后前向运动精子百分率差异有显著性(P<0.05),而总活动率差异没有统计学意义。结论:冷冻对精子活动力损伤明显,冷冻保护剂∶精液=1∶3比例较冷冻保护剂∶精液=1∶1比例可提高冷冻后前向运动精子百分率。     

冰箱冷冻法和程序降温仪冷冻法对人类精液冷冻复苏后精子参数的影响

<正>近些年,随着更多的精子库在国内得到卫生部的批准而成立,精液的冷冻技术也有了很大的提高,除了世界卫生组织《人类精液检查与处理实验室手册》(第5版)推荐的冷冻方法外[1],还衍生出了更多的其他精液冷冻方法[2-3]。程序降温仪由于能够精确控制液氮蒸气注入冷冻室的速率,降温速率精确,仍然为首选的冷冻方法。但由于人们对精子库认识的加深和理解,更多的捐精志愿者来到精子库进行精液的捐献,而仅靠程序降温仪已经不能满足     

精子冻存技术及上游法精子处理技术对精子DNA完整性的影响

目的通过对冷冻前后及精子处理前后精子DNA完整性的比较,探讨冷冻技术、冷冻时间及上游法精子处理技术对精子DNA完整性的影响。方法（1）精液常规检测正常的患者30例,手淫法取精,精液液化混匀后分4份,分别用于精子染色质扩散（SCD）实验检测精子DNA完整性、上游法处理精液、以及两组冻存实验（不加保护剂直接冻存的为冻存1组;添加蛋黄葡萄糖保护剂冻存的为冻存2组）;（2）上游法处理后的精液一部分用于检测精子动力和形态,一部分用于精子DNA完整性检测;（3）冻存1组分别于冻存第7天和第90天解冻,SCD实验检测精子DNA完整性;（4）冻存2组分别于第7天和第90天解冻,0．1ml用于精子DNA完整性检测,剩余精液应用上游法处理,检测上游处理前后精子DNA的完整性。结果（1）冻存1组中,冻存90d后解冻的精子DNA损伤率[（25．6土7．3）％]显著高于冻存7d者[（22．4±7．4）％]（P〈0．05）,且均显著高于新鲜精液的精子DNA损伤率[（20．6±7．3）％]（P〈O．05）;冻存2组中,冻存90d后精子DNA损伤率显著高于冻存7d者[（25．9±7．2）％VS．（23．6土7．8）％]（P〈O．05）,且均显著高于新鲜精液（P〈0．05）;而冻存7d和90d后,两个冻存组间比较,精子DNA损伤率均无显著差异（P〉0．05）。（2）新鲜精液经上游法处理后,精子DNA损伤率由处理前的（20．6±7．3）％降为（6．4±2．5）％（P〈O．05）;冻存2组中精液冻存7d和90d后复苏上游法处理后,精子DNA损伤率较未经上游法处理者均显著降低[分别为（9．38±2．8）％VS．（23．6±7．8）％和（9．7±2．6）％VS．（25．9±7．2）％]（P％0．05）。结论冻存对精子DNA有损伤,冻存时间对于精子DNA完整性有影响。不添加保护剂直接冻存和添加保护剂对精子DNA完整性的影响无显著差异。不论是新鲜精液还是冻存复苏精液,上游法处理并不会增加精子DNA的损伤,且有利于筛选出具有更好DNA完整性的精子。     

精子上游处理技术的改进和影响因素的评价

本文介绍一种改进的精子上游处理技术,用以提高活动精子的回收率,并对其影响因素进行了分析。共选择精子密度正常的精子活力低下症22例进行该上游处理,活动精子回收率为50.07±23.4％。研究发现不同精子密度、精子运动速度、精液粘稠度等因素可影响活动精子回收率。作者认为该法设备操作简单,可复性强,便于在人工授精中推广应用。     

不育男性精子染色质结构与精液常规参数的关系

目的 研究不育男性精子染色质结构参数与精液常规参数的关系.方法 采集36例男性不育患者和18例健康对照者的精液标本,用TUNEL法检测精子DNA碎片化,CMA3染色法检测精子染色质包装质量,按照世界卫生组织标准(1999)检测精液常规参数.结果 不育组的精子TUNEL阳性率和CMA3阳性率均显著高于健康对照组,差异具有统计学意义(P＜0.05),精液常规参数正常的不育男性精子TUNEL阳性率和CMA3阳性率也显著高于健康对照组,差异具有统计学意义(P＜0.05),精子TUNEL阳性率和CMA3阳性率与精液常规参数之间具有显著的相关性(P＜0.05).结论 不育男性的精子染色质结构检测结果与健康男性存在显著差异,精子染色质结构检测可提供反映男性生育能力的信息.     

The effect of bacterial contamination of semen on sperm chromatin integrity and standard semen parameters in men from infertile couples

A considerable proportion of male factor infertility cases are associated with inflammatory processes. The most common sexually transmissible agents causing sexually transmitted diseases in industrial countries are Chlamydia trachomatis, genital Ureaplasma and Mycoplasma spp. This study was undertaken to investigate whether these bacterial contaminants in semen affect sperm quality parameters and particularly DNA integrity (detected by sperm chromatin structure assay) in males from infertile couples (n = 293). The results showed that semen contaminations with the investigated bacterial species were not associated with sperm DNA fragmentation. However, contaminations with Mycoplasma spp. and C. trachomatis were associated with decreased sperm concentrations. Total sperm numbers in contaminated semen samples tended to be decreased, but not significantly. Mycoplasma had the highest adverse effect on sperm quality (concentration, motility, morphology and DNA condensation). Antibiotic therapy of the selected 47 men was successful in 55%, but semen quality parameters did not improve at least up to 3 months after the therapy. The presence of pathogenic bacteria in semen is primarily associated with low sperm production. Our data showed that Mycoplasma spp. contamination of semen had the highest adverse effect on sperm quality. Sperm chromatin integrity assessed by the presence of DNA breaks was not disturbed.     

Chemiluminescence in semen of infertile men

Summary. Luminol-dependent chemiluminescence (CL) can be used to determine the production of reactive oxygen species (ROS) by cells. Enhanced formation of ROS in human semen was reported to be of pathological significance for a disturbed sperm function. To investigate incidence of elevated CL-signals in semen samples and their correlation to conventional semen parameters, CL-signals in the semen of both 49 consecutive infertile men and 20 controls were measured. Semen was analysed according to WHO-criteria including bovine mucus-penetration- and water-test. A CL-signal of 1.5 × 10⁵ counts min⁻¹/2 × 10⁶ spermatozoa was considered to be the upper normal limit. The CL in infertile men's semen was elevated with statistically significant differences in oligozoospermia patients/controls ( P < 0.0001) and normozoo-spermia patients/controls ( P < 0.05). In the group with elevated CL-signals, a higher percentage of spermatozoa with a pathologic morphology was detected ( P = 0.05). In the groups with pathologic results of eosin- and water-tests, the CL-counts were elevated ( P < 0.006; P < 0.03). The spermatozoa motility in the group with elevated CL-counts was significantly reduced after 4 h ( P < 0.05). The CL-signals correlated inversely with the results of the bovine mucus-penetration-test ( r = -0.67, P < 0.0001). In conclusion, semen samples of 28% of our patients showed elevated CL-signals; these were associated with pathological results of membrane integrity-tests. The negative correlation of CL with the results of Penetrak®-test reflects its importance to depict the functional capacity of spermatozoa.     

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.     

精浆弹性蛋白酶与精液常规参数及精子DNA完整性的相关性研究

目的:观察男性不育患者精浆弹性蛋白酶水平与精液质量及精子DNA完整性之间的关系,探讨生殖道炎症隐性感染对男性不育的影响。方法:选择202例男性不育患者依照精浆弹性蛋白酶实验结果并结合其他生殖道感染炎症标志物将男性不育分为确认感染组和隐性感染组,选择同期因女方因素不孕前来检查的生育力评估正常,排除生殖道炎症及其他影响生育的致病因素的健康男性32例作为对照组。统计分析不育组与正常对照组、不育组中确认感染组与隐性感染组、不育组中弹性蛋白酶水平正常者与升高者之间精浆弹性蛋白酶水平与精液质量及精子DNA完整性之间的差异。精子浓度及活力用计算机辅助精子分析系统检测,精子形态检查采用Diff-Quik染色法,精子DNA完整性分析用精子染色质扩散法(SCD),精浆弹性蛋白酶用酶联免疫吸附法。结果:正常对照组和不育组的正常形态精子百分率分别为(11.9±9.2)%和(4.1±3.3)%、精子DNA碎片指数(DFI)分别为(10.9±8.7)%和(21.3±12.7)%,精浆弹性蛋白酶分别为(220.9±73.1)ng/ml和(1687.3±1621.2)ng/ml,前向运动精子百分率(PR)分别为(49.7±10.8)%和(19.1±10.3)%,两组相互比较各指标均具有统计学差异(P均0.05)。不育组中弹性蛋白酶升高者较弹性蛋白酶正常者,PR显著降低,DFI显著升高,差异均具有统计学意义(P0.05)。隐性感染和确证感染患者间精液量、精子浓度、PR、正常形态精子百分率和DFI的差异均无统计学意义(P均0.05)。结论:生殖道炎症感染无论是显性或者是隐性有可能导致DNA损伤程度增加,从而影响男性生育。     

LDH isoenzymes in semen of infertile men

LDH isoenzymes (LDH1, LDH2, LDH3, LDH4, LDH5, and LDHx) were evaluated in 68 infertile men and in a control of 10 fertile men. No statistically significant difference was found in the value of the 5 isoenzymes (LDH 1-5) in the different subgroups of infertile men and in the comparison of the infertile men with the control group. LDHx was detected in the infertile men, regardless of the cause of infertility, as long as the sperm concentration was greater than or equal to 16 million/ml, LDHx was not detected not only in all azoospermic men but, also, in those infertile men whose sperm count was less than or equal to 15 million/ml. There is a positive relationship (p less than 0.001) between LDHx and sperm concentration, but there is no such relationship between LDHx and % motility.     

精索静脉曲张不育患者氧化应激水平和精子DNA完整性及精液参数的相关性分析

目的:分析精索静脉曲张(VC)不育患者氧化应激同DNA完整性及精液参数的关系。方法:采用前瞻性研究,纳入98例患者,根据活性氧水平(ROS)将VC不育患者分为ROS低水平组(54例)及高水平组(44例),分析两组精子DNA碎片指数(DFI)、精子活力、形态等参数的差异,并分析它们之间的相关性。结果:ROS高水平组DFI显著高于ROS低水平组[(34.49±6.05)%vs(27.38±8.10)%,P=0.039]。与ROS低水平组比较,ROS高水平组精子活力[(25.21±18.22)%vs(36.16±22.83)%,P=0.017]、前向运动精子百分率[(16.34±9.22)%vs(23.34±11.53)%,P=0.041]、曲线速率[(20.62±4.38)%vs(27.03±6.21)%,P=0.013]及直线性率[(18.30±7.93)%vs(29.75±8.24)%,P=0.024]均显著降低,而精液白细胞数显著升高[(0.86±0.41)×106/ml vs(0.65±0.15)×106/ml,P=0.022]。Spearman相关性分析表明ROS水平同精液白细胞数(r=0.41,P0.01)及DFI(r=0.21,P=0.006)呈显著正相关,同曲线速率(r=-0.24,P=0.017)、直线性率(r=-0.24,P=0.021)、精子活力(r=-0.31,P=0.002)及前向运动精子百分率(r=-0.41,P=0.012)呈显著负相关。DFI同精子活力(r=-0.29,P0.01)、前向运动精子百分率(r=-0.34,P0.01)呈显著负相关。结论:精浆ROS水平同VC不育患者DFI显著正相关。氧化应激及DNA完整性可影响患者的精子参数。     

少精子症患者精液沉淀物乳酸脱氢酶同工酶凝胶电泳分析

本文运用乳酸脱氢酶同工酶聚丙烯酰胺凝胶电泳分析法，分析比较了少精子症、弱精子症、精液正常者的精液沉淀物乳酸脱氢酶同工酶百分比活性。结果显示：少精子症患者的ＬＤＨ－Ｃ４同工酶百分比活性４６％显著低于精液正常者的７１％（Ｐ＜０．０１）及弱精子症患者的６７％（Ｐ＜０．０１）；相对的，少精子症的ＬＤＨ５同工酶百分比活性１６％显著高于精液正常者的４．８％（Ｐ＜０．０１）和弱精子症的６．３％（Ｐ＜０．０１）。本文认为少精子症者精液沉淀物ＬＤＨＣ４活性降低与其精液中的各类细胞诸如未成熟的精子细胞、白细胞及一些微生物的相对增加有关     

The impact of body mass index on semen parameters in infertile men

This hospital‐based, prospective study was conducted to evaluate the relationship between body mass index (BMI) and various semen parameters in infertile men. A total of 439 men presented for infertility evaluation were assessed by basic infertility evaluation measures including semen analysis and BMI calculation. The main outcome measure was the relationship between BMI groups [BMI: 18.5–24.9 kg/m² (normal weight), 25–29.9 kg/m² (overweight) and ≥30 kg/m² (obese)] and different semen parameters [volume, concentration, motility and morphology]. The mean BMI was 29.67 ± 5.89. Most of patients (82.91%) were overweight or obese. The 3 BMI groups were comparable in semen parameters (P > 0.05). BMI had a negative correlation with various semen parameters. However, this correlation was significant only with sperm concentration (P = 0.035). We concluded that sperm concentration was the only semen parameter which showed significant reduction with higher BMI in infertile men.     

巨噬细胞对不育男子精液质量的影响

本文应用免疫细胞化学方法,研究正常生育男子、不育男子精液中巨噬细胞水平变化,同时,分析精液质量参数.结果显示不育组精液巨噬细胞显著增高;巨噬细胞阳性组与阴性组比较,活动率,活动力,尾部肿胀率均显著降低,畸形率明显增高;尾部肿胀率,畸形率与巨噬细胞含量分别呈负相关和正相关.研究提示在男性不育症患者精液中巨噬细胞有较高发生率,精液巨噬细胞数量变化可影响精液质量,造成男性不育的机理有待进一步研究.