大鼠精子在附睾成熟中精子膜变化的研究

我们运用Ｐｅｒｃｏｌｌ离心技术对ＳＤ大鼠附睾头、体、尾各段的精子分离纯化后，再用硫代巴比妥酸法、酶法、ＳＤＳ－ＰＡＧＬ电泳技术等对精手膜依次进行唾液酸、甘油－３－磷酸胆碱（ＧＰＣ）及蛋白质含量变化的检测。结果显示：精子膜唾液酸、ＧＰＣ量不断降低且有显著统计学意义（Ｐ＜０．０１）。附睾头、体、尾各段精子膜唾液酸和ＧＰＣ量分别为１０．１８±２．８２、８．４２±３．０７、７．８３±２．７９μｇ／１０８精子；１１２．３１±２８．１４、１０９．３３±３７．１６、７４．５０±２５．１３ｎｍｏｌ／１０８精子（－ｘ±ｓ）。膜蛋白变化主要是由大分子蛋白转变成较小分工的蛋白。大鼠精子附睾转运中精子膜的变化与精子成熟具有十分密切的关系。     

碳酸氢盐在小鼠精子体外获能和顶体反应中的作用

检验了精子获能和透明带（ＺＰ）及孕酮激发顶体反应是否需要ＨＣＯ３／ＣＯ２。小鼠精子分别在改良的Ｔｙｒｏｄｅ（ｍＴ－Ｂ２５，含２５ｍｍｏｌ／ＬＨＣＯ３／ＣＯ２）或ｍＴ－Ｈ（无ＨＣＯ３Ｈ／ＣＯ２，含２０ｍｍｏｌ／ＬＨｅｐｅｓ）中预培养９０ｍｉｎ后，以２步７０％和３５％ｐｅｒｃｏｌｌ／ｍＴ－Ｈ洗涤精子，并将精子重新悬浮于ｍＴ－Ｂ２５，ｍＴ－Ｂ１５ｍＴ－ＢＨ和ｍＴ－Ｈ中，用ＩＺＰ／μｌ或１５μｍｏｌ／Ｌ孕酮激发精子顶体反应。在上述ｍＴ诸培养基中，“Ｂ”精子（以ＣＴＣ荧光染色法确定）发生率为６１％～６７％；顶体反应均可达到４１．０士１．４％～４８．０±１．４％。表示精子一旦在ＨＣＯ３／Ｃ０２环境中获能，顶体反应就可在无ＨＣＯ３／ＣＯ２环境中发生。然而，精子预先在ｍＴ－Ｈ中培养，与上述相同方法处理精子，“Ｂ”型精子（２２％～３３％）明显低于前者，对ＺＰ或孕酮的刺激不起反应（１４．Ｏ士３．６～２４．７士０．６％），甚至将精子重新悬浮于ｍＴ－Ｂ２５中也不发生反应（２２．０±９．５％），表示这些精子并未获能。上述结果表明小鼠精子获能依赖于ＨＣＯ３／ＣＯ２，但顶体反应则否。     

抗精浆免疫抑制物抗体的抗生育机理研究

作者进行了抗精浆免疫抑制物抗体（ＳＰＩＭ－Ａｂ）的补体结合功能测定（ＣＦＡ）和ＳＰＩＭ－Ａｂ对精子凝集、制动、穿透力和杀精子影响的研究。结果表明，ＳＰＩＭ－Ａｂ阳性不育病人（ｎ＝２９）ＣＦＡ值为６．４７±１．５５ｋＵ／Ｌ，明显低于ＳＰＩＭ－Ａｂ阴性病人（ｎ＝７ｌ，８．１１±１．６２ｋＵ／Ｌ）和对照组（ｎ＝３０，８．６０±１．８０ｋＵ／Ｌ），Ｐ＜０．０１。经ＳＰＩＭ－Ａｂ阳性血清和ＳＰＩＭ－Ａｂ阴性血清作用后，两组精子凝集、制动和死精子百分率分别为４１．４％和２１．１％、６９．０％和１６．９％、６２．１％和４．２％，精子穿透高度分别为３１．６±１３．０ｍｍ和３８．Ｏ±１２．９ｍｍ，经统计学处理差异显著（Ｐ＜０．０５～ｐ＜０．０１）。提示ＳＰＩＭ－Ａｂ能够以抗原抗体复合物形式激活补体，其对精子凝集、制动、杀伤和穿透力的影响，可能是干扰生育的重要原因之一。     

精子运动轨迹图像分析影响因素的探讨

报告了正常生育男性春夏秋冬的精子运动平均速度的正常值范围分别为２４．６±２．１，３０．９±３．０，３１．２±２．７和２３．５±２．２μｍ／ｓ。探讨了季节（温度）、生育年龄、禁欲时间等因素对精子运动轨迹图像分析的影响。结果表明，冬春与夏秋季节对精子运动轨迹有明显的影响（Ｐ＜０．０１）；春季与冬季，夏季与秋季精子运动速度无显著差异；生育年龄和禁欲时间对精子运动轨迹无影响。实验表明，男性在生育力旺盛期间（２１～４８岁），睾丸的功能是相对稳定的，人类的生殖也无季节性变化。     

精子运动轨迹图像分析影响因素的探讨

本文报告了正常生育男性春夏秋冬的精子运动平均速度的正常值范围分别为２４．６±２．１，３０．９±３．０，３１．２±２．７，和２３．５±２．２μｍ／ｓ。探讨了季节（温度）、生育年龄、禁欲时间等因素对精子运动轨迹图像分析的影响。结果表明，冬春与夏秋季节对精子运动轨迹有明显的影响（Ｐ＜０．０１）；春季与冬季，夏季与秋季精子运动速度无显著差异；生育年龄和禁欲时间对精子运动轨迹无影响。实验表明男性在生育力旺盛期间（２１～４８岁），睾丸的功能是相对稳定的，人类的生殖也无季节性变化。     

精浆超氧化物歧化酶与解脲支原体感染的关系

目的：观察解脲支原体（ＵＵ）感染与精浆超氧化物歧化酶（ＳＯＤ）活性的关系。　方法：应用精子活体染色技术和ＳＯＤ活性检测试剂盒,分别检测３６ 例正常生育男性（正常对照组）和８５例ＵＵ 感染不育男性（ＵＵ 感染组）的精子存活率与精浆ＳＯＤ活性。　结果：在正常对照组和ＵＵ 感染组中,精子存活率分别为（７８．２±７．７）％ 和（５３．６±８．４）％ ,差异极其显著（Ｐ＜ ０．０１）;精浆ＳＯＤ活性分别为（７６２．７±１３０．４）和（６４５．４±１３２．５） ＮＧ／ｍ ｌ,差异显著（Ｐ＜ ０．０５）。　结论：ＵＵ 感染可引起精浆ＳＯＤ活性降低,精子膜发生过氧化损伤,影响精子的存活率和活动力。在对ＵＵ 感染病人进行抗感染治疗的同时,辅以清除氧自由基和提高ＳＯＤ活性的物质,有利于病人恢复正常的生育能力。     

氯胺酮用于学龄前儿童静脉滴注的药代动力学

利用反相高效液相色谱法，对８例３．５￣６．０岁学龄前儿童恒速静脉滴注０．０５％的氯胺酮３ｍｇ·ｋｇ＾－１·ｈ＾－１，测定其血中氯胺酮浓度，得出各项药代动力学参数ｔ１／２α＝０．１４１６±０．０４２３ｈ，ｔ１／２β＝２．８３５１±０．４４２７ｈ，Ｃｌ＝０．３３７０±０．０５８３Ｌ·ｋｇ＾－１·ｈ＾－１，Ｖｄ＝０．６６２０±０．２８５４Ｌ／ｋｇ，ＡＵＣ＝９．２０６２±１．００１９ｍｇ／Ｌ·ｈ，房室模型     

精子与宫颈粘液相互作用在预测宫颈性不育的价值

为了解精子与宫颈粘液相互作用预测生育的价值，对２７例不明原因不孕妇女进行性交后试验（ＰＣＴ）和精子穿透试验（ＳＭＰＴ）。１年后按有无妊娠分为妊娠组（ｎ＝１４）和非妊娠组（ｎ＝１３）。两组宫颈粘液评分无显著性差异（Ｐ＞０．０５）；妊娠组在性交后２和４小时精子数（１９．３±３．３及２２．２±４．４个／Ｈｐ）明显高于非妊娠组（５．０±２２及２．９±１．４个／Ｈｐ）（Ｐ＜０．０１）；妊娠组ＰＣＴ正常率（７１％）明显高于非妊娠组（０％），（Ｐ＜０．０１）；ＳＭＰＴ结果与ＰＣＴ结果基本一致。提示ＰＣＴ是有价值的预测生育指标，ＰＣＴ正常可排除男方因素及宫颈性不孕，ＰＣＴ异常则可选择人工授精提高妊娠率     

脑室注射γ-氨基丁酸对雄性大鼠下丘脑促性腺激素释放激素含量的影响

本研究观察了不同剂量γ－氨基丁酸（ＧＡＢＡ）对雄性大鼠下丘脑促性腺激素释放激素（ＧｎＲＨ）含量的作用。结果表明，脑室注射５、５０、２５０及５００ｍｍｏｌ／ＬＧＡＢＡ各为２０μｌ，４０分钟后下丘脑ＧｎＲＨ含量分别为３．５７±０．５８、３．７５±０．６６、４．６３±０．６３及５．０７±０．５９ｎｇ／１０ｍｇ湿重组织，均显著高于对照组下丘脑ＧｎＲＨ含量（１．７６±０．２１ｎｇ／１０ｍｇ湿重组织，Ｐ＜０．０５）。脑室注射２５０ｍｍｏｌ／ＬＧＡＢＡ２０μｌ后２０、４０、６０、１２０及２４０分钟下丘脑ＧｎＲＨ含量依次为２．２４±０．２９、４．６３±０．６３、５．１１±０．３８、４．２０±０．４０及３．９８±０．９１ｎｇ／１０ｍｇ湿重组织。ＧＡＢＡ受体拮抗剂荷包牡丹碱（ｂｉｃｕｃｕｌｉｎｅ，Ｂｉｃ）可阻断ＧＡＢＡ对下丘脑ＧｎＲＨ含量的促进作用。     

人重组γ-干扰素抗生育效应及其机理研究

本研究以新西兰品系实验兔为动物模型,观察人重组－γ干扰素（ｈｒＩＦＮ－γ）的抗生育效应并探讨其作用机理。实验结果：ｈｒＩＦＮ－γ能降低兔胚泡的着床率,对照组的着床率为８０％,ｈｒＩＦＮ－γ２．５、５．０和１０万ＩＵ３组的着床率分别为３０％、２７％和２７％。检测血清中雌二醇（Ｅ２）水平,对照组为１５７．９±３１．８ｐｍｏｌ／Ｌ,ｈｒＩＦＮ－γ各组均有降低趋势,分别为１０７．３±１９．２、１０６．３±２１．７和１０４．９±１９．８０ｐｍｏｌ／Ｌ;血清孕酮（Ｐ）水平,对照组为５０．５９±７．６２ｎｍｏｌ／Ｌ,ｈｒＩＦＮ－γ各组分别为１０．２１±３．１８、６．２３±１．６４和４．８７±０．２６ｎｍｏｌ／Ｌ。在对卵巢和子宫组织的实验病理学的系统观察发现,ｈｒＩＦＮ－γ剂量为１０万ＩＵ时,呈明显的黄体退化、子宫内膜的发育及内膜上皮和腺体的分泌均被抑制。研究表明ｈｒＩＦＮ－γ对孕兔有一定的抗生育效应,其抗孕作用的主要靶器官可能是卵巢,从而影响子宫内环境,使不利于妊娠     

Two-dimensional gel electrophoretic analysis of rat sperm membrane interaction with cauda epididymal fluid

Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) was used to analyze the polypeptide composition of rat cauda epididymal fluid, blood serum and membrane-enriched fractions of caput, corpus, and cauda epididymal spermatozoa. Several polypeptides were found in both cauda fluid and blood serum, and in both cauda fluid and epididymal spermatozoa. Prominent cauda epididymal fluid polypeptides that were associated with caput, corpus, and cauda sperm membranes were 32 and 33 kDa. Passage of spermatozoa from the caput to the cauda epididymidis was characterized by the loss of three glycopolypeptides of 32, 30 and 29 kDa, and by the addition of a 37-kDa glycopolypeptide. Incubation of intact caput, corpus and cauda spermatozoa with cauda epididymal fluid revealed major changes in the polypeptide maps of the incubation fluid and the membrane-enriched fractions of caput and corpus, but not cauda spermatozoa. The incubation of cauda fluid with caput and corpus sperm cells was characterized by a loss of several polypeptides and the addition of a 24-kDa glycopolypeptide. The most striking change in spermatozoa incubated with cauda epididymal fluid was the addition of two glycopolypeptides of 32 and 33 kDa to the polypeptide maps of caput sperm cells. These data demonstrate that rat spermatozoa undergo surface modifications during epididymal maturation and that these modifications can be influenced by epididymal fluid.     

非阻塞性输精管滤过装置节育术的精浆中性α-糖苷酶的检测

本文采用ＷＨＯ推荐的改良精浆中性α－糖苷酶的测定方法测定输精管滤过装置节育术（ＩＶＤ组）和输精管结扎术（结扎组）的精浆中性α－糖苷酶活性。术前两组精浆均测出中性α－糖苷酶活性：ＩＶＤ组为４３．５０±２９．０１ｍＵ／每次射精（ｘ±ｓ）；结扎组为４７．８１±３１．２０（ｘ±ｓ）ｍＵ／每次射精；两组间无显著差异（Ｐ＞０．０５）。术后６、１２个月ＩＶＤ组分别有９１．５７％和７９．１７％的精浆测出中性α－糖苷酶活性；而结扎组仅有４．４８％和４．２４％的精浆测出中性α－糖苷酶活性；两组间均有非常显著的差异（Ｐ＜０．００１）。结果提示：ＩＶＤ组术后一年内其大部分受试对象的精浆中仍有附睾液存在。     

计算机辅助分析人、家兔、大鼠和小鼠附睾精子运动能力

本研究应用计算机辅助精子分析（ＣＡＳＡ）定量分析了人、家兔、大鼠和小鼠精子附睾成熟过程中，精子运动能力的发生和发展。同时对这几种动物和人进行了系统分析和比较。结果表明：在不同种属之间，其运动的发生和发展具有一定的差异；各种不同种属动物精子在各自附睾成熟过程中，其运动能力的两个方面参数，运动速度和运动方式的发展是不平行的；附睾尾部精子的运动能力（包括运动速度和直线程度）最强。     

Contribution of epididymal factors to sperm maturation and storage

Summary.  In fertile men, the majority of epididymal spermatozoa acquire the potential to fertilize (assessed with sperm function assays) on passage into the corpus and cauda regions of the epididymis. Although secretions of the epididymal epithelium are clearly important for sperm maturation and survival, their role in this process has yet to be fully determined. Alterations in epididymal sperm membranes may result from the incorporation of protein, sugar and lipid determinants. Most probably, factors of epididymal origin act in concert with constitutional changes to spermatozoa, which together permit full sperm function. Epididymal spermatozoa incubated with epididymal epithelial cell cultures can undergo some maturation in vitro , which can lead to the development of sperm fertilizing capacity. Co-incubations of human sperm with epididymal epithelial cultures, at 37 °C with medium replenished every other day, led to 50% of spermatozoa retaining motility after 8 days. In one case, a few spermatozoa survived for 17 days, the inherent maximal survival time of human spermatozoa in situ. An important aspect of coculture experiments is that close interactions between spermatozoa and epithelial cells can be examined in detail. This coculture technique may yield important information related to epididymal sperm maturation and storage.     

Thyroid gland and epididymal function in rats. II. Sperm motile efficiency

The epididymis is controlled by the endocrine system and plays an active role in the process of spermatic maturation. In order to extend these studies, our investigations were focused on the relationship between the hypofunction of the thyroid gland and the motility of the caput, corpus, and cauda epididymal spermatozoa. The hypofunction was associated with decreasing sperm motility efficiency. In thyroidectomized rats injected with T-3, no sperm motility changes were observed.     

Effects of altered epididymal sperm transit time on sperm quality

The epididymal sperm transit time seems to have an important role in the process of sperm maturation, and it seems that alterations to the transit can harm the process. The aim of the present work was to evaluate the influence of altered sperm transit time through the epididymis on sperm parameters and fertility of rats, as well as the role of testosterone in the alterations. Sprague–Dawley adult male rats were randomly assigned to four different groups and were treated for 12 days: (i) 10 μg/rat/day DES, to accelerate the transit; (ii) 6.25 mg/kg/day guanethidine sulphate, to delay the transit; (iii) same treatment as group 1, plus androgen supplementation; (iv) control animals received the vehicles. Guanethidine treatment delayed the sperm transit time through the epididymal cauda, provoking increased sperm reserves in this region. Animals exposed to DES showed an acceleration of sperm transit time in the epididymis, and consequently decreased sperm density in both epididymal regions, the caput-corpus and cauda, and diminished sperm motility. In both cases sperm production was not altered. Testosterone supplementation was able to restore the transit time to values close to normality, as they were higher than in the control rats. The same occurred in relation to sperm motility. Rats exposed to DES presented lower fertility after in utero artificial insemination using sperm collected from the proximal cauda epididymis. Therefore, it was concluded that the acceleration of rat sperm transit time appeared to harm normal sperm maturation, thus decreasing sperm quality and fertility capacity, in an androgen-dependent way.     

Sperm maturation in dogs: sperm profile and enzymatic antioxidant status in ejaculated and epididymal spermatozoa

Spermatozoa become more susceptible to the attack of reactive oxygen species during maturation. To avoid oxidative damage, the epididymis must provide the necessary antioxidant protection. The aim of this study was to compare the canine sperm profile and the enzymatic antioxidant status of the ejaculated fractions and samples collected from the different segments of the epididymis (caput, corpus and cauda). Five adult dogs were used, and after 1–3 weeks, subsequently to bilateral orchiectomy and epididymal storage, sperm samples were collected from the different segments of the epididymis. Samples were evaluated for conventional microscopy and computer‐assisted motility analysis: sperm plasma membrane permeability and the activity of the antioxidant enzymes catalase, glutathione peroxidase (GPx) and superoxide dismutase (SOD). Samples collected from the caput and corpus showed lower values for most of the motility variables evaluated, indicating different levels of immaturity. Catalase activity was observed only in ejaculated samples. Conversely, GPx activity was higher in the cauda epididymidis. Correlations were found between SOD and GPx and SOD and sperm motility in the epididymal cauda and corpus, highlighting the importance of the enzymes for the protection of spermatozoa during the transit along the epididymis.     

大鼠精子在附睾成熟过程中肉毒碱量变化的研究

肉毒碱被认为是与精子在附睾中成熟有关的成熟因子。本文报告测定大鼠附睾头部,体部及尾部精子中的肉毒碱。结果表明精子在循附睾头、体、尾运行过程中,精子中肉毒碱含量逐步增加,在附睾体一头部,附睾尾一头部的精子内的肉毒碱量呈现十分显著性差异。结果提示附睾体部或体一头部交界部位可能是精子在附睾中成熟的关键部位。     

Use of neonatal tolerization and chemical immunosuppression for the production of monoclonal antibodies to maturation-specific sperm surface molecules.

Mammalian sperm acquire functional maturity as they move from the caput to the cauda epididymidis. Changes occur in the protein/glycoprotein composition of the sperm plasma membrane during this time, and may be essential to the maturation process. The production of monoclonal antibody (Mab) probes to the maturation-specific molecules has been difficult since new proteins comprise a minor portion of total membrane proteins. This report describes a protocol for enhancing the production of Mabs to maturation specific molecules. By injecting neonatal mice with caput epididymal sperm plasma membranes, in combination with chemical immunosuppression at adulthood, the mice were made tolerant to the antigens expressed on the caput sperm membranes. Subsequent immunization with cauda epididymal sperm plasma membranes allowed the production of Mabs to the maturation-specific moieties without the necessity for extensive antigen purification procedures. The majority of the resulting Mabs recognize cauda, not caput, epididymal sperm plasma membranes as determined by enzyme-linked immunosorbent assay (ELISA), immunocytochemistry on unfixed cells, and Western blot analyses, even though the protein profile from caput epididymal sperm plasma membranes is very similar to that from cauda membranes. The five Mabs described also recognize cauda fluid antigens, suggesting that the maturational changes on the sperm plasma membranes arise from interactions with the epididymal fluid. Use of the tolerization/immunosuppression protocol has provided Mab tools to assist in the study of sperm maturation during epididymal transit.