整合素系统对人精子顶体反应的作用

目的：观察整合素系统与人精子顶体反应的关系。方法：采用整合素VLA—5的特异性配体纤连蛋白(Fn)及整合素配体的活性片断RGD肽作用于人精子lh后，用三色法行顶体染色，采用了常规显微镜下肉眼计数和图像处理软件进行分析两种方法，观察Fn和RGD作用前后顶体反应(AR)率的变化。结果：常规显微镜下肉眼计数发现，100nM的Fn、200nM的Fn、250mg/L的RGD肽和500mg/L的RGD肽作用于人精子后AR率较作用前有明显增高；运用图像分析软件分析发现，100nW的Fn、200nM的Fn、250mg/L的RGD肽和500mg／L的RGD肽作用于人精子后AR率也有显性升高。以上P均＜0.05。结论：人精子膜上的整合素参与了人精子的顶体反应。     

考马期亮蓝染色法检测人精子顶体反应的评价

为建立一种检测人检测人精子顶体反应的简便实用的新技术，用３．５％高氧酸水溶液本制０．０５％考马斯亮蓝染色液浸染人精子３０ｍｉｎ，顶体完整者顶体区染成紫蓝色，顶体反应者则不梁。对获能前后和钙离子载体Ａ２３１８７诱导顶体反应的精子进行染色并与经典的顶体反应检测技术ＰＳＡ法对照，两种方法检测顶体反应率无显著差异。     

孕酮对正常生育男性精子头部胞浆内游离钙离子浓度（[Ca^2＋]i）的影响

目的：定量分析生育男性获能后单个精子的[Ca^2 ]i基础水平以及10μM孕酮刺激后的[Ca^2 ]i动态改变。方法：生育健康男子10例，手淫法留取精液。精液液化后，应用上游技术分离活动良好的精子，之后将精子悬液置于3‘70C培养2h使之获能。获能后精子移人Petri培养皿，每皿加入终浓度为8．85μmoL／L的钙荧光探针Fluo-3／AM，370C避光孵育40min。     

甲氧滴滴涕对大鼠精子顶体反应的影响

目的:探讨甲氧滴滴涕(methoxychlor,MXC)对雄性大鼠精子顶体反应的影响。方法:在体外培养的精子悬液中加入不同浓度的MXC和孕酮处理,通过精子考马斯亮蓝染色,镜检计数顶体反应型精子的百分率。结果:与空白组比较,单纯经MXC处理组的顶体反应型精子百分率明显增加,另加孕酮的各处理组顶体反应型精子的百分率增加尤为明显,MXC终浓度≤6.25μg/ml的处理组顶体反应型精子百分率不增加。结论:MXC可促进大鼠精子发生顶体反应且与其剂量有关,当MXC终浓度≤6.25μg/ml不诱发顶体反应。MXC能加强孕酮诱发精子顶体反应的作用。     

考马斯亮蓝染色法检测人精子顶体反应的评价

为建立一种检测人精子顶体反应的简便实用的新技术，用３．５％高氯酸水溶液配制０．０５％考马斯亮蓝（Ｒ２５０）染色液浸染人精子３０ｍｉｎ，顶体完整者顶体区染成紫蓝色，顶体反应者则不染。对获能前后和钙离子载体Ａ２３１８７诱导顶体反应的精子进行染色并与经典的顶体反应检测技术ＰＳＡ法对照，两种方法检测顶体反应率无显著差异。方法学检验证明新技术结果可靠。本法操作简便、经济，普通显微镜即可检测，易于推广，克服了原有技术复杂昂贵的缺点，具有研究和临床诊断的良好价值。     

白色念珠菌性阴道炎对人精子顶体反应影响机制的初步探讨

目的探讨白色念珠菌性阴道炎对人精子顶体反应（AR）的影响机制。方法以ELISA和硝酸还原酶法分别检测阴道分泌物中的IL-8与NO,采用BAEE/ADH法测定顶体酶的活性,并通过FITC-PSA法检测AR。结果无症状的白色念珠菌携带者阴道分泌物中IL-8及NO水平无显著变化,但在白色念珠菌性阴道炎患者阴道分泌物中,其水平显著升高,且二者水平呈正相关;IL-8对精子顶体酶活性和AR均无影响;低浓度的NO可诱导精子顶体酶活性,同时促进AR,而高浓度的NO对其起抑制作用。结论白色念珠菌性阴道炎患者阴道分泌物IL-8和NO水平升高,可直接或/和间接抑制精子顶体酶活性和AR,其中高浓度的NO起着直接的抑制作用,而IL-8可能间接参与了此作用。     

抗人精子甘露糖受体抗血清对精子穿透实验的影响

目的 研究人精子甘露糖受体(MR)在精卵融合中的作用.方法 用亲合层析法分离纯化的人精子甘露糖受体来免疫Balb/c小鼠得到抗MR抗血清;用抗血清处理顶体反应后的人精子,并进行去透明带金黄地鼠卵的精子穿透实验(SPA),观察其体外受精情况.结果 抗MR抗血清可抑制人精子与去透明带金黄地鼠卵的结合与穿透,使卵子的受精率下降,具剂量依赖性.结论 人精子MR与精卵膜的识别与融合有关.     

精子获能的评价

人及哺乳动物精子在离开生殖道时，还不能立即与卵子受精。它必须在雌性生殖道内经历一段成熟过程，才能获得受精能力。1951年著名华裔科学家张明觉（M．CChang）博士和C．R．Austin分别发现了这一生理现象。翌年，Austin将这一现象称之为精子获能（capacitmion）。精子获能是一系列的生理生化反应过程步骤大致分为：     

Fertilization and early embryology: Oscillations in intracellular free calcium induced by spermatozoa in human oocytes at fertilization

Calcium has an important role in the events of egg activationand early preimplantation development. We investigated changesin intracellular calcium concentration in human oocytes at fertilizationusing the calcium-sensitive photoprotein aequorin. Oocytes weredonated for research by patients undergoing in-vitro fertilizationtreatment in the Department of Obstetrics and Gynaecology. Cumuluscells, and in some cases zonae pellucidae, were removed by appropriateenzyme treatment. Single oocytes were micro-injected with aequorinand incubated in a chamber perfused with pre-equilibrated culturemedium in a photomultiplier system. Eleven zona-intact and 15zona-free oocytes were incubated with sperm, and oocytes fromeach group were incubated without sperm as controls. Dramatictransient increases in intracellular free calcium concentrationwere recorded in three zona-intact and seven zona-free oocytes,thought to be the first direct measurements of intracellularchanges in human oocytes at fertilization. The amplitude (upto 2.5 µM), duration (120 s) and frequency (every 10–35min) of these transients were similar in zona-intact and zona-freeoocytes. They resemble those recorded in mouse oocytes, whichmay therefore be a suitable model for biochemical events athuman fertilization.     

Regulation of cyclic adenosine monophosphate synthesis in human ejaculated spermatozoa. II. The role of calcium and bicarbonate ions on the activation of adenylyl cyclase.

In this study, we have applied our previous data describing the experimental conditions necessary for expression of cyclic adenosine monophosphate (cAMP)-synthesizing adenylyl cyclase in human ejaculated spermatozoa, to investigate the direct effects of calcium (Ca2+) and bicarbonate (HCO3-) upon activation of the enzyme in vitro. We report that the effects of Ca2+ and HCO3- were significantly dependent on the status of the enzyme activity. Thus, at a near saturating (10 mM) concentration of MnCl2 giving high enzyme activity, addition of less than 10 mM HCO3- did not affect adenylyl cyclase activity and higher concentrations inhibited the enzyme, with 50 mM HCO3- reducing the activity by 33%. Also, addition of less than 20 mM CaCl2 alone or in combination with 10 mM HCO3- did not significantly change the enzyme activity. In great contrast, enzyme activation was highly responsive to Ca2+ and HCO3- when MnCl2 was present at a concentration giving submaximal enzyme activity. Thus, at 2 mM MnCl2, adenylyl cyclase was markedly increased by CaCl2 concentrations between 10 and 100 mM. The activation was further enhanced by increasing concentrations of HCO3-, with 50 mM HCO3- giving the highest activity at 50-100 mM CaCl2. Activation by CaCl2 was also observed in the absence of added MnCl2, being significantly greater than basal activity at 10 mM CaCl2 and maximal at 100 mM CaCl2.(ABSTRACT TRUNCATED AT 250 WORDS)     

Induction of capacitation in human spermatozoa in vitro by an endometrial sialic acid-binding protein

A Ca²⁺-dependent sialic acid-binding protein (SABP) of humanendometrium, which specifically bound to human sperm head plasmamembrane in vitro, was found to increase the percentage motilityand acrosome-reacted pattern of uncapacitated spermatozoa. Theprotein was synthesized in the endometrium and secreted intothe uterine fluid. This intra-uterine factor, which is apparentlyadvantageous in vitro in inducing human sperm capacitation,may play a significant role in promoting the postrelease maturationof ejaculated spermatozoa by enhancing ⁴⁵Ca uptake into spermatozoaby a pathway which is insensitive to calcium-channel blockers.However, the ⁴⁵Ca uptake could be enhanced on exposure to thedivalent cation ionophore A23187 and inhibited in the presenceof the calmodulin inhibitor trifluoperazine. The SABP also inducesan increase in intracellular Ca²⁺ in spermatozoa, as seen byFURA-2 AM studies. Furthermore, overlay studies show human SABPto be a Ca²⁺-binding protein. The data presented here suggestthat SABP induces invitro sperm capacitation and the subsequentacrosome reaction by increasing intracellular Ca²⁺ concentration.     

雌激素对钙通道作用的研究进展

雌激素是一种类固醇激素,在体内具有广泛的生物学活性。细胞膜钙通道是介导外钙内流的主要途径,参与了细胞内的很多过程,包括细胞兴奋、增殖、分化和凋亡。大量研究表明,雌激素可通过基因组效应机制影响神经系统、心血管系统以及其他组织细胞钙通道的表达和功能,也可通过非基因组效应机制快速激活或者抑制多种不同钙通道的活性,并且多呈现一种非单调的量效关系,本文就雌激素对钙通道作用的研究进展作一综述。     

The thiol reagent, thimerosal induces intracellular calcium oscillations in mature human oocytes

The aim of this study was to investigate the effects of thimerosalon intracellular calcium in human oocytes related to the stageof nuclear maturity. A total of 20 oocytes from superovulatedwomen undergoing gamete intra-Fallopian transfer (GIFT) werestudied. The calcium-sensitive fluorescent dye Fura-2 was usedto monitor intracellular calcium. Regular oscillations in theconcentration of cytosolic calcium were observed in 13 out of20 oocytes following exposure to thimerosal; five oocytes didnot respond to thimerosal treatment, and spontaneous oscillationsof cytosolic free calcium were recorded in two oocytes. Thimerosalinduced oscillations of intracellular calcium in a significantlyhigher proportion of metaphase II oocytes (10/11) compared withmetaphase I oocytes (3/8; P < 0.2). These findings demonstratethat thimerosal is a potentially useful agent for the studyof the calcium signalling processes in human eggs and suggestthat the underlying cellular mechanisms develop at a relativelylate stage of oocyte maturation.     

The hypoosmotic swelling test and cryosurvival of human spermatozoa

The hypoosmotic swelling test is a simple laboratory test to evaluate the functional integrity of the membrane of human spermatozoa. This test was performed on 83 samples of human semen before cryopreservation to determine whether it has any predictive value for the cryosurvival of human spermatozoa. Stepwise regression analysis demonstrated that conventional sperm characteristics, including the concentration, motility, normal morphology and viability, of pre-freeze semen samples were of limited value in predicting the cryosurvival of human spermatozoa. Further, the hypoosmotic swelling test results from pre-freeze semen samples did not correlate with the post-thaw motility or the survival rate of spermatozoa after cryopreservation.     

Ultrastructural abnormalities of human spermatozoa

The observation of sperm cells in the scanning and electronmicroscope reveals that malformed spermatozoa show either asingle anomaly of each of their structural components—acrosome,nucleus, axoneme and accessory structures—or a combinationof these anomalies. A defined semen profile can be establishedwhen the majority of ejaculated spermatozoa present a predominantlysingle anomaly or the same combination of associated anomalies.Only in these case can there be a relationship between morphologicaldefects and impairment of the fertilizing ability. The validityof the ultrastructural examination of sperm cells depends uponthe data obtained by light microscopy, and quantification ofthe frequencies of the ultrastructural alterations is necessaryto define clearly the limits of abnormality.