人类睾丸精子和附睾精子的顶体蛋白酶活性

目的：探讨人类睾丸精子和附睾精子的顶体蛋白酶活性。方法：应用明胶薄层试验、穿卵试验和透射电镜观察顶体蛋白酶活性及其影响。结果：未经洗涤的睾丸精子和附睾精子不出现明显的晕轮，洗涤后则显示出顶体蛋白酶活性。附睾精子组的平均晕轮直径显著高于睾丸精子组（Ｐ＜００１）。附睾精子能与去透明带地鼠卵受精。老化和畸形的附睾精子降低或丢失酶活性。结论：睾丸精子已初步发育具功能活性的顶体蛋白酶，进入附睾顶体蛋白酶逐渐发育至功能成熟，而曲细精管和附睾管内液体存在着抑制酶活性的物质     

环磷酰胺下调大鼠睾丸OCT-4和GDNF表达

目的探讨抗肿瘤药物环磷酰胺对大鼠睾丸和精原干细胞相关因子OCT-4和GDNF表达的影响。方法 12只8周龄大鼠随机分为对照组和实验组,每组6只;实验组每天腹腔注射环磷酰胺50 mg/kg,连续3 d,用药4周后PAS染色法观察大鼠睾丸组织学变化,精原细胞,生精小管直径和附睾尾部精子数量。免疫组织化学法检测精原干细胞相关分子OCT-4和GDNF表达水平。结果与对照组比较,实验组大鼠体重、睾丸和附睾重量均显著减轻,睾丸组织学无明显改变,附睾精子活动度降低,附睾精子和精原细胞数量减少。实验组GDNF表达明显减少,而OCT-4表达水平无明显变化。结论环磷酰胺短期用药可造成精子发生的损害,其中精原细胞的减少可能与损害后GDNF表达下调有关。     

癫痫对雄性大鼠睾丸及附睾组织病理学和超微结构的影响

目的：探讨癫痫发作对雄性Wistar大鼠睾丸及附睾组织病理和超微结构的影响。方法：运用氯化锂一匹罗卡品建立Wistar雄性大鼠癫痫模型,取睾丸、附睾分别制片观察组织病理及超微结构的形态学改变。结果：光镜观察下,A组（造模成功组）及B组（造模不成功组）大鼠睾丸生精小管内各级生精细胞排列基本整齐,结构未见明显紊乱现象,但生精细胞层次呈不同程度的减少,睾丸生精小管管腔内精子数目减少,可见坏死脱落的生精细胞,睾丸间质结构缺如,间质细胞明显减少。附睾中,A组管壁柱状细胞,基细胞层次清晰,结构整齐,微绒毛排列整齐,但管腔中精子数目明显减少,有较多的非精子细胞成分。B组附睾管腔中精子数目未见明显下降,有时可见散在的生精细胞。电镜观察下,A组大鼠睾丸生精细胞细胞核明显畸形,线粒体肿胀,线粒体膜仍然完整,但脊消失,粗面内质网肿胀明显,精子头部细胞核清晰,顶体形态不规则,尾部“9＊2＋2”结构整齐,周围包绕的线粒体鞘明显肿胀,线粒体数目明显减少。B组中仍可见上述不同程度的损害表现,附睾中,A组及B组均可见处于同一层面的主细胞,细胞核未见明显异常,核周围可见大量溶酶体,同时核周内质网均处于明显肿胀状态。C组（正常对照组）鼠睾丸及附睾切片的光镜、电镜表现皆正常。结论：癫痫不同程度的发作可引起大鼠睾丸及附睾组织病理和超微结构不同程度的改变,进而造成了雄性Wistar大鼠生殖系统相关指标的改变。     

低剂量棉酚与激素联合应用的抗生育作用位点的研究

目的 探讨低剂量棉酚与激素联合应用在8周内使大鼠达到不育效果的作用位点.方法 本实验采用大鼠喂服棉酚12.5mg/(kg·d) 去氧孕烯125μg/(kg·d) 炔雌醇25μg/(kg·d) 十一酸睾丸酮100mg/(kg·d)的联合用药方式,与单独喂服相同剂量的棉酚或激素的大鼠及喂服载体溶剂的大鼠相对照,通过一系列形态学观察和半定量检测,探讨联合用药的具体作用环节.结果 无论在睾丸组织形态还是睾丸精子数量,联合用药组的变化趋势都与单独应用激素组的相似,而不同于单独应用棉酚组和对照组.而在附睾中,3个用药组在精子活力和精子形态方面,都与对照组有显著差异.结论 联合用药组中激素主要使睾丸精子数量迅速下降;而在精子数量大大下降的基础上,棉酚和激素又对已成熟精子的结构和运动能力进一步破坏,从而使雄性大鼠受精能力迅速下降,联合用药组比单独应用其中一种成分更快、更有效地达到使大鼠不育的目的.     

胺碘酮对大鼠睾丸形态和功能的影响及其作用机制

目的:探讨胺碘酮对大鼠睾丸质量、形态、酶活性及精子活力的影响.方法:健康雄性SD大鼠随机分为对照组、低剂量组和高剂量组,连续灌胃方式给予胺碘酮4周.处死大鼠后,分离大鼠双侧睾丸,并称重;光学显微镜观察精子活力,H-E染色观察睾丸形态;采用乳酸脱氢酶(LDH)、琥珀酸脱氢酶(SDH)、一氧化氮合酶(NOS)及一氧化氮(N...     

Decline in fertility of mouse sperm with abnormal chromatin during epididymal passage as revealed by ICSI

BACKGROUND: Recent studies showed that ICSI with cauda epididymal or ejaculated sperm of infertile mice or men, respectively, was less effective in fertilization and normal embryo development than ICSI using sperm from the testes. These studies suggested that sperm nuclear quality declined after release from the testis, but the site where this loss of fertility occurs has not been localized. METHODS: We performed ICSI with testicular, caput, and cauda epididymal sperm from infertile Tnp1-/-Tnp2+/- mutant mice, which have a minimal level of transition nuclear proteins and are sterile by natural mating. RESULTS: When the heads of motile sperm from the testis or caput epididymis of Tnp1-/-Tnp2+/- males were injected into enucleated mouse oocytes, sperm chromosomes showed no difference from those of wild-type mice, but the chromosomes from sperm taken from the cauda epididymis of mutant males showed increased abnormalities. Injection of testicular or caput epididymal sperm from Tnp1-/-Tnp2+/- males into intact oocytes resulted in normal embryonic and fetal development and yields of liveborn equivalent to wild-type, but cauda sperm from Tnp1-/-Tnp2-/- mice produced lower implantation rates and yields of liveborn than did those from wild-type mice. CONCLUSIONS: These results demonstrate that in mice with sperm chromatin abnormalities, the decline in fertility of sperm with ICSI occurs after the caput epididymis. The advantage of using caput epididymal sperm for ICSI in certain situations may be considered as an approach to be tested in human assisted reproduction.     

Assisted reproduction for the treatment of azoospermia

Azoospermia, the most severe form of male infertility, is caused by obstructions in the genital tract or by testicular failure. Microsurgical techniques are available for the correction of some of these obstructions but no effective treatment is available for testicular failure. In recent years, methods have been developed for direct surgical sperm sampling from either the epididymis or the testis to be used by intracytoplasmic sperm injection. The main approach proven to be effective for the retrieval of spermatozoa from the epididymis in patients with obstructive azoospermia is microsurgical epididymal sperm aspiration, although recently the retrieval of spermatozoa by fine needle aspiration was shown to be equally effective. Recovery of spermatozoa is also now performed in patients with severely deficient spermatogenesis using testicular open biopsy as well as aspiration by fine needle. The ultimate choice of sperm retrieval method in these patients will depend not only on sperm availability, but also on the physiological consequences of the different techniques on testicular function. This article summarizes the recent advances achieved in the treatment of azoospermic patients using these assisted reproduction surgical techniques.     

Epididymis-specific pathologic disorders in rats exposed to gossypol from weaning through puberty

Previous work in our laboratory revealed that the pubertal period of reproductive development in the male rat was particularly vulnerable to gossypol exposure, with a higher frequency of round structures in the lumen of the cauda epididymidis in the treated rats. Herein, we utilized hemicastration and electron microscopy to confirm that the epididymis is a definitive target of gossypol. Although exposure to gossypol from weaning through puberty caused a significant decrease in daily sperm production, as well as in the concentration of sperm in the epididymis, serum testosterone levels and reproductive organ weights were not altered. In gossypol treated rats, sperm morphology was compromised severely, but the epithelium in testis and epididymis appeared morphologically normal. Ultrastructural examination revealed that round structures, present only in gossypol exposed males, represented: (1) principal cells exfoliated from the epididymal epithelium; (2) epididymal epithelial cell cytoplasm containing degenerating sperm; and (3) degenerating epithelial cells, consisting of vesicles and particles of different sizes, forms and densities. Taken together, the data confirm that gossypol targets the epididymis, disturbing both the structure and function of this organ, and presumably disrupts sperm maturation.     

Tetracycline-induced reproductive toxicity in male rats: Effects of vitamin C and N-acetylcysteine

Tetracycline, a broad-spectrum antibiotic employed clinically in the treatment of bacteria infections, is known to cause a number of biochemical dysfunctions and suspected to induce testicular damage to animals and humans, but there is paucity of data on its effect and mechanism of action on the male reproductive system. The present study therefore evaluates its spermatotoxic and testicular toxicity in male rats and the chemoprotective effects of Vitamin C (Vit C) and N-acetylcysteine (NAC). Tetracycline was administered orally at the dose level of 28.6 mg/kg body weight per day in two equal divided doses (12h interval). Vit C and NAC were also administered orally to the rats at doses of 200 and 50 mg/kg body weight per day, respectively, for the 14 days of the experiment. While there was no change in the body weights of rats, tetracycline administration caused significant decrease in the relative weights of testis, epididymis and seminal vesicles (P<0.05). Administration of tetracycline caused a reduction in the epididymal sperm motility, percentage of live spermatozoa, sperm count, and an increase in abnormal sperm morphology, as well as induction of adverse histopathologic changes in the testes. While Vit C and NAC significantly mitigated the toxic effect of tetracycline on sperm parameters, the antioxidants did not improve the adverse histopathologic changes induced by antibiotic. Treatment of rats with tetracycline significantly decreased the activities of superoxide dismutase, catalase (CAT), glucose-6-phosphate dehydrogenase, glutathione-S-transferase (GST) and the levels of GSH and serum testosterone, while the activity of gamma-glutamyltranspeptidase and the formation of malondialdehyde (MDA) increased. Both Vit C and NAC significantly attenuated the toxic effects of tetracycline to the antioxidant and testicular marker enzymes as well as markers of oxidative stress. Collectively, the results suggest that therapeutic dose of tetracycline elicits spermatotoxic and testicular toxicity in male rats through induction of oxidative stress. The chemoprotective effects of Vit C and NAC during tetracycline treatment suggest that these antioxidants may find clinical application in cellular damage involving reactive oxygen species (ROS).     

Effects on antioxidant status of liver following atrazine exposure and its attenuation by vitamin E

In the present investigation, the effect of atrazine on antioxidant enzymes and body weight was studied in male Wistar rats. Atrazine (300 mg/kg bw) was administered by gavage for 7, 14 and 21 days. A significant increase in hepatic lipid peroxidation (LPO) was observed following atrazine administration. Vitamin E treatment (100 mg/kg bw), on the otherhand, attenuated atrazine-induced LPO in liver. In addition, vitamin E treatment restored the GSH content and glucose-6-phosophate dehydrogenase activity that was found to be lowered after atrazine administration. The activities of antioxidant enzymes: superoxide dismutase, catalase, glutathione peroxidase and glutathione-s-transferase were significantly increased following atrazine administration and vitamin E treatment could restore these activities. In conclusion, the results of the study demonstrate that atrazine induces oxidative stress in terms of enhanced lipid peroxidation. However, vitamin E treatment ameliorated the effects of atrazine suggesting it as potential antioxidant against atrazine-induced oxidative stress.     

低剂量棉酚与甾体激素联合用药抗男性生育协同作用

目的 探讨低剂量棉酚和甾体激素联合用药发挥抗男性生育的药物协同作用.方法 采用大鼠喂服棉酚与甾体激素联合用药方式[棉酚12.5mg/(kg·d)+去氧孕烯125μg(kg·d)+炔雌醇25μg/(kg·d)+十一酸睾丸酮100mg/(kg·d)],与单独喂服相同剂量的棉酚或激素的大鼠及喂服载体溶剂的大鼠相对照,应用形态学观察、半定量检测和体视学睾丸组织结构定量研究的方法,探讨低剂量棉酚和甾体激素抗生育的药物协同作用. 结果 在睾丸重量、附睾重量、睾丸精子产量和睾丸组织结构定量方面,低剂量棉酚和甾体激素无药物协同作用;在降低附睾精子计数和精子活动度方面,联合用药组的效果强于其他3组,低剂量棉酚和甾体激素具有药物协同作用.结论与单用低剂量棉酚和甾体激素比较,两者联合用药可以更快地减少附睾精子的数量和活动度,从而快速达到避孕效果.     

Alterations in the spermatic function generated by obesity in rats

In order to assess the effect of obesity on epididymal and germinal epithelia in control rats and obese rats induced by a high fat diet, we evaluated the epididymal and testicular morphologies, lipid peroxidation in the epididymis, leptin serum levels, steroid hormones, insulin, cholesterol, triglycerides, glycemia and some spermatobioscopic parameters. No significant difference was observed in the levels of insulin, glucose, cholesterol and triglycerides between the two groups. Nonetheless, in the obese rats, circulating leptin and estradiol levels showed a significant increase and there was a decline in the testosterone levels. The same group showed an increase in the lipid peroxidation of the epididymis and reduced spermatobioscopic parameters. The heads of the epididymis showed morphological differences in obese rats. No significant difference was observed between the testes of both groups. There is a clear evidence of an effect on sperm in obese rats and this seems to occur in the epididymis.     

The relative viability of human spermatozoa from the vas deferens, epididymis and testis before and after cryopreservation

Testicular and epididymal spermatozoa are routinely used with in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) to achieve pregnancies. In addition, excess cryopreserved spermatozoa can be thawed and used for ICSI. However, information on the recovery of epididymal and testicular spermatozoa after freeze-thaw is lacking. This is important to determine the feasibility of using previously cryopreserved aspirated spermatozoa for ICSI. We prospectively compared the viability of fresh and frozen-thawed spermatozoa from the vas deferens, epididymis and testicle by several measures. Testis spermatozoa were obtained from men with non-obstructive azoospermia (n = 5), epididymal spermatozoa from men with obstructive azoospermia (n = 8), and vasal spermatozoa from fertile men by vasal irrigation at vasectomy (n = 5). The viability of fresh spermatozoa was assessed by motility, two vital stains (carboxyfluorescein, 0.08 mg/ml and propidium iodide, 20 mg/ml) and the hypo-osmotic swelling assay (HOS; 100 mmol/l citrate and fructose). After cryopreservation, spermatozoa were thawed and all viability measures repeated. Although fresh vasal spermatozoa were the most motile, testicular spermatozoa exhibited similar, high viability (91 and 86% respectively) by vital stain. Spermatozoa from testis, epididymis and vas deferens survived cryopreservation equally well by vital stain, but not by motility. As a selection measure, the HOS assay identified significantly more viable epididymal and testicular spermatozoa than did motility in both fresh and frozen-thawed populations. It appears feasible to use frozen-thawed extracted spermatozoa for ICSI when motility and a selection measure such as the HOS assay are used. With fresh testis spermatozoa, selection methods may not be necessary prior to ICSI, as cell viability is high.     

Rats with metabolic syndrome resist the protective effects of N-acetyl l-cystein against impaired spermatogenesis induced by high-phosphorus/zinc-free diet

Consumption of relatively high amounts of processed food can result in abnormal nutritional status, such as zinc deficiency or phosphorus excess. Moreover, hyperphosphatemia and hypozincemia are found in some patients with diabetic nephropathy and metabolic syndrome. The present study investigated the effects of high-phosphorus/zinc-free diet on the reproductive function of spontaneously hypertensive rats/NDmcr-cp (SHR/cp), a model of the metabolic syndrome. We also investigated the effects of antioxidant, N-acetyl-l-cysteine (NAC), on testicular dysfunction under such conditions. Male SHR/cp and control rats (Wistar Kyoto rats, WKY) were divided into three groups; rats fed control diet (P 0.3%, w/w; Zn 0.2%, w/w), high-phosphorus and zinc-deficient diet (P 1.2%, w/w; Zn 0.0%, w/w) with vehicle, or high-phosphorus and zinc-deficient diet with NAC (1.5 mg/g/day) for 12 weeks (n = 6 or 8 rats/group). The weights of testis and epididymis were significantly reduced by high-phosphate/zinc-free diet in both SHR/cp and WKY. The same diet significantly reduced caudal epididymal sperm count and motility and induced histopathological changes in the testis in both strains. Treatment with NAC provided significant protection against the toxic effects of the diet on testicular function in WKY, but not in SHR/cp. The lack of the protective effects of NAC on impaired spermatogenesis in SHR/cp could be due to the more pronounced state of oxidative stress observed in these rats compared with WKY.     

Temporal patterns of DNA adduct formation and glutathione S-transferase activity in the testes of rats fed aflatoxin B1: a comparison with patterns in the liver.

Fisher-344 male rats were fed 1.6 ppm of aflatoxin B1 (AFB1) continuously and intermittently for several weeks. At various time periods, DNA was isolated from the testes and livers and analyzed for AFB1-DNA adducts. The ability of the testis to detoxify AFB1 was also investigated by the glutathione S-transferase (GST) activity assay and compared with that of the liver. The levels of testicular AFB1-DNA adducts were 2.4 to 8.1 times lower than those of the liver after 4 to 16 weeks of continuous treatment and 2.2 to 46.2 times lower after 8 to 20 weeks of intermittent treatment. The testicular DNA adducts markedly decreased over time. By 16 weeks of continuous and 20 weeks of intermittent exposure, they had decreased 37 and 91%, respectively. In contrast, hepatic AFB1-DNA adducts increased four-fold from 4 to 16 weeks of continuous treatment but increased at a much slower rate after intermittent exposure. In both the liver and testis, significant levels of AFB1-DNA adducts persisted for at least 1 month after ending the treatment, suggesting that this type of lesion was poorly repaired. In control rats, the testis showed significantly higher GST activity than the liver. In treated rats, these differences were significant during the first 12 weeks of continuous treatment but not at later times. Tissue-specific differences such as germ-cell depletion and increased testicular detoxification may play an important role in the observed differential pattern of DNA adduct formation between the testis and liver.     

Identification of epididymis-specific antigens using neonatal tolerization

PROBLEM: Conventional immunization using whole sperm containing multiple antigens as the immunogen followed by hybridoma technology usually gives antibodies to antigens invariably of testicular origin, probably because of the strong immunogenic nature of these antigens. Therefore, an alternate approach of neonatal tolerization or subtractive immunization has been utilized to raise antibodies specific to epididymis by suppressing immune response to testicular antigens. METHOD OF STUDY: Neonatal mice were tolerized with testicular sperm proteins on days 0 and 5. These animals were then immunized with epididymal sperm proteins on day 21, followed by two boosters at biweekly intervals. Sera from these mice were used to localize epididymis-specific antigens. RESULTS: Sera from mice that were tolerized to testicular sperm proteins and later immunized with epididymal sperm proteins reacted only with epididymal proteins. CONCLUSION: The results of this study demonstrate that neonatal tolerization with testicular sperm proteins, followed by immunization with epididymal sperm proteins, enhances the production of antibodies to proteins exclusively of epididymal origin.     

巴戟天对微波损伤的雄鼠睾丸生精功能的影响

目的探讨微波辐射对雄鼠的生精功能的影响及巴戟天对损伤后雄鼠的生精功能的修复作用。方法 24只SD大鼠随机均分为空白组、辐射组和给药组。用对讲机微波辐照辐射组和给药组;给药组在辐射后巴戟天灌胃。三组均在相同条件下饲养。1周后观察各组雄鼠的睾丸指数、精子活性及精子密度等指标的变化。结果辐射组相对空白组睾丸指数、精子活性以及精子密度降低(P0.05),附睾指数变化没有统计学意义(P0.05);给药组相对辐射组附睾指数、睾丸指数、精子活性、精子活动率均增高(P0.05);而给药组相对空白组睾丸指数、精子活性以及精子密度增高(P0.05),附睾指数变化没有统计学意义(P0.05)。结论巴戟天对微波损伤的雄鼠睾丸生精功能有修复作用。     

Aspermia and chronic testicular pain after imperforate anus correction. Cryopreservation of sperm cells extracted from whole orchiectomized testis: case report

This paper describes an unusual association of aspermia and untreatable, chronic testicular pain in a young man who underwent 14 surgical interventions for an imperforate anus. Physical examination and ultrasonography revealed left epididymal and vas enlargement, normal-sized testes, tubular ectasia of the left rete testis and a small intraprostatic paramedian left cyst. Retrograde ejaculation and urogenital infections were excluded, and the FSH and karyotype results were normal. The patient gave his consent to an exploratory intervention with possible radical left orchiectomy. The patency of the left distal seminal duct was unexpectedly normal, and no sperm were found in the epididymis or vas deferens despite their obstructive appearance. Sperm were only found in a 'testicular touch' preparation. The removed testis was immediately opened and most of the testicular lobules were removed, thus allowing the extraction of 25 x 10(6) sperm, which were cryopreserved in 35 straws. An 8-month follow-up examination documented the complete absence of pain and, during the next few months, it is planned to use the thawed sperm for ICSI. Radical orchiectomy plus the cryopreservation of sperm extracted from the whole testis must be considered in the case of the co-existence of chronic unilateral testicular pain and aspermia.     

Effects of selenium on histopathological and enzymatic changes in experimental liver injury of rats

The most important antioxidant aspect of selenium is its function in the active site of selenoenzyme glutathione peroxidase. Glutathione peroxidase not only allows the removal of the toxic radicals but also permits the regeneration of lipid molecules through reacylation in the cellular membrane. Thus, GSHPx may prevent the harmful effects of free radicals and may reduce the formation of the reactive metabolites of carbon tetrachloride. Carbon tetrachloride is a hepatotoxic agent which generates haloalkane radicals during its biotransformation in the liver and is widely used to make the experimental model of hepatic damage. Therefore, the aim of the present study is to investigate the possible protective role of selenium on the experimental liver cirrhosis and some enzyme activities in blood plasma from rats. While the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP) were significantly increased (p < 0.05, p < 0.05 and p < 0.01, respectively), gamma-glutamyle transferase (GGT) activity was not statistically affected (p < 0.05) with carbon tetrachloride-injection. The levels of AST, ALT and GGT in carbon tetrachloride-group decreased to nearly the enzyme values in control-group after the selenium-injection but the ALP was increased (p<0.01). On the other hand, it was noticed that selenium significantly decreased the hepatic injury. In conclusion, our results showed that carbon tetrachloride caused an increase in the activities of liver enzymes in plasma and selenium application decreased the hepatic injury. Plasma levels of the liver enzymes were decreased after selenium-injections. Based upon these results, selenium may play an important role in the preventive indication of hepatic cellular injury inducted by carbon tetrachloride.     

Polymyxin B Protects Against Hepatic Ischemia/Reperfusion Injury in a Rat Model of Obstructive Jaundice

This study was conducted in order to investigate the effects of polymyxin B (PMB) against hepatic ischemia/reperfusion (I/R) injury in rats with obstructive jaundice. Thirty-six Wistar rats (eighteen each) with induced hepatic I/R injury by biliary tract ligation and recanalization were assigned to a control group (reperfused with normal saline) and a PMB group (reperfused with PMB). Indicators involving liver function, oxidation resistance, pro-inflammatory state, and anti-apoptosis effect were determined following the instructions. Compared with normal saline, PMB reperfusion resulted in a significant improvement of liver function (increase of glutathione and reduction of aspartate aminotransferase, alanine aminotransferase, and lactate dehydrogenase), oxidation resistance (decreased malondialdehyde and myeloperoxidase activity), alleviation of pro-inflammatory state (less tumor necrosis factor (TNF)-α, interleukin-1 beta (IL-1β), nuclear factor kappa B (NF-κB) mRNA, and intercellular adhesion molecule (ICAM)-1), and anti-apoptosis effect (more Bcl-2 and less Bax). PMB protects the liver from I/R injury mainly through reducing cellular oncosis and apoptosis and regulating the expression of NF-κB, TNF-α, IL-1β, and ICAM-1.