HYALURONIC ACID INCREASES MOTILITY/INTRACELLULAR CA 2+ CONCENTRATION IN HUMAN SPERM IN VITRO

This study investigated the mechanisms of the stimulatory effect of hyaluronic acid on motility in human sperm in vitro. A method, involving the measurement of forward progression through an agarose gel, was used to measure sperm motility quantitatively. Changes in intracellular Ca 2+ concentrations in sperm were detected using the fluorescent dye Fluo-3. The effects of hyaluronic acid (6.5, 65, 650 ng/mL) and nifedipine (32 nM) on sperm motility were investigated. The effects of hyaluronic acid, nifedipine (32 nM), A23187 (32 µM), and a monoclonal antibody to human CD44 (1 µg/mL) on changes in intracellular CA 2+ concentrations were investigated. Hyaluronic acid significantly ( p <. 008) stimulated sperm motility and this was partially inhibited by nifedipine.A23187 significantly ( p <. 005) increased intracellular CA 2+ concentrations. Hyaluronic acid significantly ( p <. 04) increased intracellular Ca 2+ concentrations and this was inhibited by nifedipine and a monoclonal antibody to human CD44. Hyaluronic acid stimulated human sperm motility by increasing intracellular Ca 2+ concentration, partially via an influx of extracellular Ca 2+.     

单侧输尿管梗阻对肾脏水通道蛋白表达的影响及临床意义

目的 探讨单侧输尿管梗阻解除前后肾脏水通道蛋白（AQP）变化与尿液浓缩功能障碍之间的联系.方法 56只体重180～200 g成年雄性SD大鼠随机分为实验组（n=48只）和对照组（n=8只）.实验组大鼠通过手术建立单侧输尿管梗阻模型并于相应时间点解除梗阻,对照组接受假手术.于相应时间点检测尿液渗透压改变,处死动物取出肾脏后通过Western blot检测梗阻前后AQP1、AQP2蛋白表达的变化,免疫组化观察蛋白表达部位.结果 梗阻后1、2 d及解除梗阻后1d,患侧AQP1、AQP2的表达随时间延长进行性下降,解除梗阻后7d其表达有所回升,AQP2于梗阻解除后14 d内恢复正常水平,而AQP1则在梗阻解除后21 d内达到正常水平.同时AQPs的变化趋势与患侧尿液渗透压改变趋势相同.梗阻解除前AQP2分布于集合管主细胞的胞浆及管周膜,解除梗阻后其表达更多的集中于管周膜.结论 AQP表达下降是造成梗阻性肾积水中尿液浓缩功能障碍的重要原因,同时梗阻解除后AQP2接受ADH的调节.     

单侧输尿管梗阻对肾脏水通道蛋白表达的影响及临床意义

目的 探讨单侧输尿管梗阻解除前后肾脏水通道蛋白(AQP)变化与尿液浓缩功能障碍之间的联系.方法 56只体重180～200 g成年雄性SD大鼠随机分为实验组(n=48只)和对照组(n=8只).实验组大鼠通过手术建立单侧输尿管梗阻模型并于相应时间点解除梗阻,对照组接受假手术.于相应时间点检测尿液渗透压改变,处死动物取出肾脏后通过Western blot检测梗阻前后AQP1、AQP2蛋白表达的变化,免疫组化观察蛋白表达部位.结果 梗阻后1、2 d及解除梗阻后1d,患侧AQP1、AQP2的表达随时间延长进行性下降,解除梗阻后7d其表达有所回升,AQP2于梗阻解除后14 d内恢复正常水平,而AQP1则在梗阻解除后21 d内达到正常水平.同时AQPs的变化趋势与患侧尿液渗透压改变趋势相同.梗阻解除前AQP2分布于集合管主细胞的胞浆及管周膜,解除梗阻后其表达更多的集中于管周膜.结论 AQP表达下降是造成梗阻性肾积水中尿液浓缩功能障碍的重要原因,同时梗阻解除后AQP2接受ADH的调节.     

二羟基乙二肟对生殖系统的毒性研究

报道了二羟基乙二肟（ＤＨＧ）高剂量时可诱发小鼠初级精母细胞染色体畸变，小鼠精子畸形率升高，对孕鼠和胚胎有毒作用，可引起孕鼠体重减轻、流产、胚胎死亡，活胎生长发育迟缓，腭裂，胎鼠畸形，下颌缺失等，致畸指数为１２；病理组织检查显示；雄鼠睾丸曲细精管生精细胞层次紊乱，生精细胞减少并可见多核巨精细胞；电镜可见，雄鼠卵巢黄体细胞浆内线粒体肿大，峭减少，基质空泡变，出现髓鞘样结构，这些结果表明高剂量的ＤＨＧ对     

芹菜汁对小鼠精液参数及毒性作用的影响

目的:探讨芹菜汁对小鼠精液参数以及毒性作用的影响,为芹菜的进一步开发利用提供一定依据。方法:将小鼠按体重随机分为7d、14d和28d3个时间组,再将时间组按体重随机分为4组(对照组、低剂量组、中剂量组和高剂量组),每天定时给对照组0.3ml生理盐水灌胃,剂量组给0.3ml不同浓度的芹菜汁灌胃,分别于各时间段结束后处死动物。应用WLJY-9000型伟力彩色精子质量检测系统测定精子运动参数。结果:7d组、14d组中高剂量组和中剂量组精子密度与相应对照组比较差异均有统计学意义(P<0.05)。中剂量组:VCL14d组和28d组分别和7d组比较有统计学意义(P<0.01),VSL7d组和14d组比较有统计学意义(P<0.05);高剂量组:VCL、MAD、WOB7d组和14d组比较均有统计学意义(P<0.05)。结论:芹菜汁能够影响小鼠精子运动参数,降低精子密度。     

丙烯酰胺对小鼠精子的毒性作用

目的研究丙烯酰胺(AA)对小鼠精子的毒性作用,为AA生殖毒性研究提供科学依据。方法将25只7～8周龄体重为25～30g的清洁级雄性昆明小鼠随机分为阴性对照组(双蒸水),AA染毒组(染毒剂量分别为20、40、60mg/kg)和阳性对照组(环磷酰胺),每组5只。除阳性对照组采用腹腔注射染毒外,其余各组均采用经口灌胃染毒,1次/d,连续5d,于首次染毒后14d处死小鼠,观察精子数、活动度与精子形态的变化。结果各剂量AA染毒组精子数均显著低于阴性对照组,差异有统计学意义(P<0.05);中、高剂量AA染毒组精子活动率显著低于阴性对照组,精子畸形率显著高于阴性对照组,差异均有统计学意义(P<0.01)。AA染毒后精子畸形类型主要表现在头部,且以无钩和无定形为主。结论AA对小鼠精子有毒性作用,且存在明显的剂量-效应关系。     

EFFECT OF SEMINAL PLASMA CALCITONIN LEVELS ON SPERM MOTILITY

This study investigated the effect of the seminal and blood plasma calcitonin levels on the sperm motility in idiopathic infertile patients. The number of sperm cells and their motility were evaluated in the spermiograms of 52 idiopathic infertile patients. The levels of seminal plasma calcitonin were studied with double antibody technique using a DPC kit. Fifty-two patients were divided into 2 groups according to the motility rates of sperm and 20 healthy volunteers were assigned to a control group. The difference between the groups was evaluated by using Kruskall-Wallis and Mann-Whitney U tests, and the correlation of seminal and blood calcitonin levels with sperm motility were determined. The difference in motility rates between the 3 groups was statistically significant ( p =. 000, p <. 05). Blood plasma calcitonin levels were in normal ranges in all cases and no significant difference was found among the 3 groups ( &#104 2 = 2.7219, p =. 2589, p >. 05). While sperm motility was correlated with seminal calcitonin levels ( r =. 8581), blood calcitonin levels did not show a correlation with sperm motility rate ( r = -.0265). Moreover, there was no correlation between seminal and blood plasma levels of calcitonin ( r = -.0010). Motility rates decreased in the patients with low seminal calcitonin levels and seminal calcitonin levels had a significant effect on sperm motility.     

EFFECT OF LISNOPRIL,AN ANGIOTENSIN CONVERTING ENZYME (ACE) INHIBITOR ON SPERMATOGENESIS IN RATS

The role of ACE inhibitors (Lisinopril) in reproductive function remains controversial. Some benefits seem to be derivable even in non-hypertensive males with low doses. This study was done using rat model to establish this fact. Male rats were divided into different groups to receive different doses of lisinopril. A control group received no drugs. The mean arterial pressure fell the most with 5 mg of lisinopril. The greatest increase in sperm count and motility was recorded for this same group. This response was dose dependent, falling as the drug dose fell. Lisinopril appeared to, in a dose dependent manner, improve sperm count and motility. In low doses, there is no significant change in arterial pressure. Infertile males with poor quality semen could benefit from a low dose of ACE inhibition. Where they are also hypertensive, ACE inhibition would be an appropriate first line treatment.     

EFFECTS OF CRYOPRESERVATION ON PENETRATION ABILITY OF HUMAN SPERMATOZOA INTO BOVINE CERVICAL MUCUS

Cryopreserved sperm exhibit lower fertilizing capacity in comparison to fresh sperm, partly due to effects of glycerol as the common cryoprotectant medium. Since standard semen analysis is not a good predictive method to assess sperm fertilizing capacity, functional tests like cervical mucus penetration may provide more useful information. A total of 24 semen samples were examined before and after cryopreservation for sperm parameters as well as number and motility of penetrated sperm into bovine cervical mucus (BCM) as an alternative for human cervical mucus. Freezing and thawing procedures have negative effects on sperm penetration into cervical mucus. No significant relation was noticed between sperm motility percentage or its penetration into BCM before and after cryopreservation, which denotes the variability in resistance of sperm to damaging effects of freezing.     

乙酰唑胺对大鼠精子发生和成熟的影响

目的:研究碳酸酐酶抑制剂对大鼠精子发生和成熟的影响。方法:乙酰唑胺混悬液(40mg/kg)给大鼠灌胃,分别于给药当天、1d和3d处死动物,进行睾丸、附睾称重、苏木素-伊红(HE)染色和光镜下观察分析;附睾尾剪碎计数精子密度、精子活力,染色后计数精子畸形比率;测定附睾管腔液α-葡糖苷酶水平;采用罗丹明(Rh)123荧光探针标记附睾精子,通过流式细胞仪检测线粒体膜电位。结果:给予乙酰唑胺的大鼠精子活力明显降低,畸形率显著增加,附睾精子线粒体膜电位明显降低,附睾管腔液α-葡糖苷酶水平有所下降,但睾丸和附睾无明显病理学改变。结论:碳酸酐酶抑制剂可能通过影响附睾精子成熟,对雄性生殖功能进行调控。     

EFFECT OF TYROSINE KINASE INHIBITORS ON TYROSINE PHOSPHORYLATION AND MOTILITY PARAMETERS IN HUMAN SPERM

Tyrosine phosphorylation has recently been associated with capacitation and suggested as a regulator of sperm movement, especially characterizing hyperactivation. The objective of this study was to verify if tyrosine phosphorylation of human sperm proteins was essentially required for the maintenance of motility as well as the development of hyperactivation. Washed sperm were incubated for 6 h in Ham's F10 + 0.35% HSA at 37°C in 5% CO 2, with and without the tyrosine kinase inhibitors genistein, tyrphostin, erbstatin, or herbimycin A and the wide-spectrum kinase inhibitor staurosporin. The concentrations of the inhibitors used in the experiments did not induce sperm toxicity, as measured by membrane integrity and mitochondrial function assays. Samples incubated without the inhibitors (control), increased their tyrosine kinase activity (ELISA), the number and intensity of tyrosine-phosphorylated (PY) protein bands (Western blot), the incidence of PY-immunoreactive sperm (immunofluorescence), and some of the sperm motion characteristics (CASA), such as velocity (VEL), amplitude of lateral head displacement (ALH), and hyperactivation. Among the selective protein tyrosine kinase inhibitors, genistein was the most active and consistent, inhibiting sperm tyrosine kinase activity, PY proteins, incidence of PY sperm, and sperm motility and motion parameters, such as VEL, ALH, and hyperactivation. The rest of the kinase inhibitors decreased motion characteristics to a varied extent and had different effects on phosphorylation parameters. In general, they decreased PY phosphorylation of 2 proteins (83 and 54 kDa) present in whole sperm extracts, and two sets of proteins of low (39-49 kDa) and medium (55-87 kDa) molecular weight present in the Triton X-100-solubilized sperm protein fraction. Thisinhibition was evident regardless of the total tyrosine kinase activity of the samples or the incidence of PY-immunoreactive sperm. The described findings further support the association between motility and protein tyrosine phosphorylation in human sperm and point to certain proteins as the main linkers.     

胚胎期暴露手机辐射对雄性仔鼠生殖功能影响

目的:探讨胚胎期暴露手机辐射对雄性仔鼠生殖功能影响。方法:建立胚胎期暴露手机辐射动物模型,将雄性仔鼠随机分为3组。饲养2个月后处死各组小鼠,分别测定各组实验动物睾丸体重系数、精子数量、精子存活率和精子活动率。结果:与对照组比较,胚胎期暴露不同制式手机辐射雄鼠体重系数和精子存活率之间差异不具有统计学意义;而精子数量和精子活动率低于对照组,且暴露于两种不同制式手机辐射组间差异不具有统计学意义。结论:胚胎期暴露手机辐射可致仔鼠精子数量减少,精子活动率降低。     

EFFECTS OF SEMEN CHARACTERISTICS ON IUI COMBINED WITH MILD OVARIAN STIMULATION

To determine the influence of sperm parameters inseminated on the outcome of intrauterine insemination (IUI) in patients undergoing ovarian stimulation with clomiphen citrat (CC) or human menopausal gonadotropin (HMG) therapy, a retrospective review was performed for 2 years on data from the IUI program. 190 couples underwent a total of 268 IUI cycles in which CC or HMG was used for ovulation induction. The initial sperm concentration (mil/ml), motility (percent), preprocessing total motile sperm (TMS) count (million), fast motile sperm (percent) and postprocessing sperm concentration (mil/ml), motility (percent), TMS count, fast motile sperm (percent), sperm morphology, hypoosmotic swelling (HOS) scores, semen leuocytes, and bacteria were analyzed. 268 inseminations were followed by a pregnancy rate of 12% and couple pregnancy rate of 17%. On multivariable logistic regression analysis, total motile sperm (TMS) count, percent motility, and percent of fast motile sperm were independent prognostic factors of fertility. The impact of the preprocessing and postprocessing sperm parameters on pregnancy outcome after IUI was evaluated. There was a trend toward an increasing percent of conception with increasing TMS count, motility, and percent of fast motile sperm. The TMS count, motility and percent of fast motile sperm independently predict success with IUI. Patients with original sperm motility ≥30% had a higher cumulative pregnancy rate (74%) than patient with motility <30% (p < 0.005). Pregnancy rate increased 4 times with motility of ≥30%.     

Immunolocalization of aquaporin 7 in human sperm and its relationship with semen parameters

Aquaporins (AQPs) are a family of 13 small hydrophobic trans-membrane proteins expressed in numerous tissues and cells. Some AQPs work as strict water channels, others are permeable to a range of substances, including glycerol. In the male reproductive system their localization in testis, efferent ducts, epididymis, and spermatozoa has been described. We studied the distribution of AQP7 in ejaculated human sperm and the relationship between AQP7 labeling and sperm characteristics. Semen samples from 33 men were examined by light and transmission electron microscopy (TEM). TEM data were quantified using a mathematical formula that calculates a fertility index (FI) and the percentages of sperm apoptosis, immaturity, and necrosis. Immunocytochemistry with a polyclonal antibody anti-AQP7 was performed on the sperm samples. Normal sperm were labeled in the pericentriolar area, midpiece, equatorial segment, and weakly in the tail (grade 1). Abnormal sperm showed a diffuse low intensity of fluorescence evident in the cytoplasmic residues, coiled tails, in the entire head, and acrosome (grade 2). A high number of motile sperm obtained by swim up were labeled in a dotted manner in the mitochondria. A significant positive correlation was found between the spermatozoa with AQP7 grade 1 labeling and the percentage of normal form (P<0.008), progressive motility and FI (P<0.005); a negative correlation was noted with the percentages of cytoplasmic residues (P<0.010) and immaturity (P<0.006) and coiled tails (P<0.012). The link between AQP7 distribution and sperm morphology and the particular dotted labeling in swim up selected motile sperm are novel and deserve additional studies.     

Corrected Seminal Fructose Levels: Index of Secretory Activity of Seminal Vesicles

Seminal fructose, sperm count, and sperm motility were measured in 340 men attending an infertility clinic. Seminal fructose correlates negatively with sperm count but not with sperm motility. The best correlation between sperm count and seminal fructose was obtained using the logarithm (log) of sperm count. When seminal fructose was multiplied by the log of sperm count obtaining a value named “corrected fructose,” the correlation with sperm count disappeared, and there was positive correlation between corrected seminal fructose and sperm motility. Therefore, corrected seminal fructose level was significantly lower in asthenospermic than in normomotile subjects, irrespective of the sperm count. Corrected seminal fructose, but not seminal fructose levels, was lowered in subjects with either low levels of serum testosterone (< 3 ng/ml) or evidence of an obstructive process in the reproductive tract. In both situations sperm motility was also reduced. The corrected fructose level was not affected by the presence of varicocele, hyperprolactinemia, or hyperserotoninemia. Measurement of corrected seminal fructose rather than seminal fructose may be a useful marker of the secretory activity of the seminal vesicles.     

PHOSPHOLIPID COMPOSITION OF HUMAN SPERM AND SEMINAL PLASMA IN RELATION TO SPERM FERTILITY

The phospholipid and fatty acid composition of sperm was studied in 8 healthy and 16 infertile men. Infertile men randomly formed from the patients with normal semen parameters according to WHO criterion. Therefore, all semen parameters of infertile patients were similar to the same characteristics of the semen of healthy men, except the abnormal forms. The amount of abnormal forms in infertile men was significantly higher than in healthy men. Sperm from infertile men show a drastic loss of phosphatidyl ethanolamine. At the same time, the significant increase of phosphatidyl serine in the sperm and seminal plasma of sterile patients was found. Lysophosphatidyl serine in the sperm of the infertile men was detected. Fatty acid composition of the semen of infertile men was altered. The levels of stearic and n-3 polyunsaturated fatty acids (eicosopentaenoic and docosahexaenoic acids) was dramatically lowered, but the values of some n-6 polyunsaturated fatty acids (linolenic and docosatetraenoic) acids increased. There was significant positive correlation between docosahexaenoic acid and sperm motility (r =. 82, p &lt;. 001) and negative correlation between linolenic acid and spermatozoa motility (r=-0.58, p&lt;.05). Infertility of men with normal semen quality can originate from the disorder of sperm lipid metabolism. The drastic loss of phosphatidyl ethanolamine and n-3 polyunsaturated fatty acids with simultaneous enhancement of phosphatidyl serine and some n - 6 polyunsaturated fatty acids in sperm could be an important cause of male infertility.     

二甲基甲酰胺对雄性小鼠精子质量及其性激素的影响

目的 研究二甲基甲酰胺(DMF)对雄性小鼠精子质量及其血清性激素的影响,探讨:DMF对雄性生殖系统影响的作用机制.方法 将SPF级健康成年雄性昆明小鼠40只随机分为对照组(蒸馏水)和低(0.5 g/kg)、中(1 g/kg)、高(2g/kg)剂量DMF染毒组,每组10只.采用灌胃方式进行染毒,灌胃容积10 ml/kg,每天1次,连续灌胃30 d.染毒后,称重,迅速分离双侧附睾,测定精子活动度、精子计数和精子畸形数,并计算精子畸形率.摘除眼球取血,采用放射免疫分析方法测定血清睾酮(T)、黄体生成素(LH)、卵泡刺激素(FSH)和睾丸T水平.结果 与对照组相比,从染毒第9天开始各剂量DMF染毒组小鼠体重明显降低,差异均有统计学意义(P＜0.05或P＜0.01).与对照组相比,中、高剂量DMF染毒组小鼠附睾精子计数较少,精子活动度较低,差异均有统计学意义(P＜0.05或P＜0.01),且随着DMF染毒剂量的增高,精子计数及精子活动度均呈下降趋势,呈现一定的剂量-效应关系.与对照组相比,中、高剂量DMF染毒组小鼠血清FSH含量较高,高剂量DMF染毒组小鼠血清T和睾丸T含量较低,差异均有统计学意义(P＜0.05或P＜0.01).各染毒组小鼠的精子畸形率、血清LH含量间比较,差异均无统计学意义(P＞0.05).结论 DMF可能是通过睾丸轴调节性激素的分泌,最终导致精子质量的降低,从而造成雄性小鼠生殖功能的损害.

Abstract：

Objective To study the effects of DMF on the quality of spermatozoa and sex hormone levels in male mice, and the possible mechanism. Methods Forty male KM mice were randomly divided into four groups and then treated with DMF dissolved in distilled water by gavage (10 ml/kg) at the doses of 0,0.5,1 and 2 g/kg,once a day for 30 consecutive days. The mice were weighted and then sacrificed on day 31. The epididymises were collected for determination of sperm motility, sperm counting and determination of abnormal sperm rate. The T,LH,and FSH levels in serum were determined. Results Compared with the control group,mice in all treated groups showed significant decrease in body weight (P<0.05). With the increase of administrating dosage,sperm counting and sperm motility showed decreasing tendency. Moreover,compared with the control group, sperm counts and sperm motility in 0.1 g/ml group and 0.2 g/ml group decreased significantly. FSH levels in serum in all treated groups were higher than those in the control group. T levels in serum and testis in all treated groups were lower than those in the control group. Conclusion Probably, DMF regulates the excretion of sex hormone by testicle axle and then lower the quality of spermatozoa.     

地塞米松对Wistar大鼠涎腺基因表达的影响

目的：观察腺病毒介导荧光素酶基因转导至大鼠颌下腺后地塞米松对基因表达的影响及IgG的变化.方法：将40只Wistar大鼠随机分为5组,经颌下腺导管转导腺病毒荧光素酶基因重组体即AdCMVLuc,3 d、1周、2周、4周、8周后观察肌注地塞米松对基因表达的影响.结果颌下腺转基因表达3 d最高,在注射地塞米松组明显大于未注射地塞米松组;以后逐渐下降,至4周、8周时仍可测到基因表达,但两组无明显差异.结论：肌注地塞米松能有效地减轻腺体的急性炎症反应,增加腺病毒介导涎腺短期基因表达,说明免疫及炎症反应直接影响转导基因的表达.     

1-溴丙烷吸入暴露对雄性大鼠生殖系统的影响

目的研究1-溴丙烷亚急性暴露对雄性大鼠的生殖毒性.方法将48只雄性大鼠分为对照组、1-溴丙烷1 000、2000、4000 mg/m3暴露组.暴露组给予7 d,每天8 h的吸入暴露.结果1-溴丙烷高浓度暴露组大鼠的体重、前列腺和精囊质量显著下降,附睾精子运动率降低,形态异常精子增多.血浆肌酸激酶活力和肌型肌酸激酶含量降低.睾丸病理切片观察到曲细精管的精子释放延迟.结论1-溴丙烷亚急性暴露对雄性大鼠的生殖系统产生毒性影响,附睾精子运动率的下降是本次实验的最敏感指标.     

CONCENTRATION OF GRANULOCYTE ELASTASE IN SEMINAL PLASMA IS NOT ASSOCIATED WITH SPERM MOTILITY

Although elastase could affect sperm motility in vitro, secretory leukocytes protease inhibitor (SLPI) prevents sperm from being attacked by elastase. The authors investigated the correlations of elastase level with sperm motility and SLPI level in vivo. Semen samples (n