己酮可可碱对人精子获能和顶体反应的影响

目的　探讨不同浓度的己酮可可碱对人精子获能和顶体反应的影响。方法 　在体外培养的精子悬液中加入不同浓度的己酮可可碱和孕酮处理 ,通过 CTC染色 ,镜检计数各型精子的百分率。结果 　与空白组比较 ,己酮可可碱处理组的获能型精子的百率明显增加 ,终浓度 8mmol·L- 1各处理组顶体反应型的精子百分率不增加 ,而终浓度为 16mmol·L- 1的处理组顶体反应型的精子百分率不明显增加。 结论 　己酮可可碱可以促进精子获能 ,在终浓度 8mmol· L- 1 时 PF不诱导精子发生顶体反应 ,但终浓度为 16mmol· L- 1 可促进精子发生顶体反应 (AcrosomeReaction,AR)     

司帕沙星、联苯乙酸对大鼠海马神经元GABA-A和离子型谷氨酸受体的作用(英文)

目的 :观察司帕沙星单用和与联苯乙酸合用时 ,对γ 氨基丁酸 (GABA) ,NMDA ,AMPA和海人藻酸诱导电流的作用。方法 :应用全细胞式膜片钳技术在急性打散的新生大鼠海马锥体神经元上记录GABA ,NMDA ,AMPA和海人藻酸的诱导电流。结果 :司帕沙星 1mmol·L- 1对GABA诱导的电流有迅速可逆的抑制作用 ,抑制率为 (4 7.6±s2 .9) % ,而联苯乙酸 10 μmol·L- 1无抑制作用。但联苯乙酸 (10 μmol·L- 1)与司帕沙星 1mmol·L- 1合用时 ,稍稍增强抑制作用 ,司帕沙星单用或合用联苯乙酸 (10 μmol·L- 1)对GABA诱导电流的IC50 分别为 (2 38± 2 1) μmol·L- 1和 (89± 5 ) μmol·L- 1。司帕沙星、联苯乙酸或司帕沙星合用联苯乙酸对NM DA ,AMPA和海人藻酸诱导的电流无明显作用。结论 :提示司帕沙星 ,或与联苯乙酸合用时所产生的中枢神经毒性可能与抑制GABA诱导的电流有关 ,而与NMDA ,AMPA和海人藻酸诱导的电流无关。     

槲皮素抑制血管生成作用的实验研究

目的　研究槲皮素 (Quercetin)对血管生成和培养的人脐静脉内皮细胞 (HUVEC)的影响。方法　采用生长因子 (血管内皮细胞生长因子VEGF、碱性成纤维细胞生长因子bFGF)诱导的鸡胚绒毛尿囊膜 (CAM)血管增生模型观察槲皮素对血管生成的影响 ;利用培养的HUVEC ,用MTT法观察槲皮素抑制内皮细胞增殖的作用 ;流式细胞仪观察槲皮素对HUVEC细胞周期的影响。结果　槲皮素 (0 1、0 0 5和 0 0 2 5mmol·L-1)能明显抑制VEGF诱导的CAM小血管生成 ;槲皮素 (0 1和 0 0 5mmol·L-1)能明显抑制bFGF诱导的CAM小血管生成 ;槲皮素 (2 4 0、12 0 μmol·L-1和 6 0 μmol·L-1)对内皮细胞增殖有抑制作用 ,抑制率分别为 6 7 0 %、5 8 1%和39 7% ;槲皮素 (2 4 0、12 0 μmol·L-1)能显著导致HUVEC的S、G2 期阻滞。结论　槲皮素能抑制VEGF和bFGF诱导的血管生成 ,且对内皮细胞增殖具有抑制作用。     

2-DG增强乳腺癌细胞对阿霉素化疗敏感性的作用

目的探讨糖基化抑制剂(2-deoxy-D-glucose,2-DG)对阿霉素(adriamycin,ADM)抗肿瘤作用的影响,以期为乳腺癌的治疗提供新的靶点。方法 MTT法检测不同浓度(0、1.25、2.5、5、10、20mmol·L-1)2-DG、不同浓度(0、0.625、1.25、2.5、5、10mg·L-1)ADM以及ADM与2-DG(10mmol·L-1)合用对乳腺癌细胞Sk-Br-3的增殖抑制作用。溴化丙啶(propidium iodide,PI)单染检测2-DG(10mmol·L-1)对ADM诱导乳腺癌细胞Sk-Br-3凋亡的影响;Western blot检测2-DG(10mmol·L-1)处理Sk-Br-3细胞不同时间(0、9、24、36h)葡萄糖调节蛋白78(glucose-regulated protein78,GRP-78)、Caspase-3的表达以及联合ADM处理不同时间(0、9、24、36h)GRP-78的表达;2-DG(10mmol·L-1)对ADM(0.4mg·L-1)诱导乳腺癌细胞Sk-Br-3所致Caspase-3活性的变化;实验中检测了2-DG及ADM合用对集落克隆形成的影响。结果 10mmol·L-12-DG对乳腺癌细胞Sk-Br-3的24、48、72h增殖抑制率分别为80.73%、75.16%、70.13%,而诱导Sk-Br-3细胞凋亡率仅为5.8%。10mmol·L-12-DG与ADM联合刺激乳腺癌细胞Sk-Br-348h的凋亡率为58.11%,高于ADM本身诱导凋亡率35.12%,且2-DG可增强ADM抑制乳腺癌细胞Sk-Br-3的集落克隆形成的作用。2-DG能上调GRP-78以及Caspase-3的活性。结论 2-DG能增加ADM诱导乳腺癌细胞的凋亡,其机制可能通过引起过度的内质网应激反应以及增加Caspase-3的活性。     

外源性腺苷三磷酸和腺苷对食管癌TE-13细胞株凋亡的影响

目的　探讨腺苷三磷酸 (ATP)和腺苷 (ADO)是否具有诱导人食管低分化鳞癌细胞株TE 13凋亡的作用。方法　MTT法 ,AO/EB双荧光染色法 ,琼脂糖凝胶电泳 ,流式细胞仪方法。结果　ATP在0 .1～ 1.0mmol·L- 1或ADO在 0 .0 1～ 1mmol·L- 1浓度范围内 ,可不同程度地抑制TE 13细胞的增殖 ,ATP和ADO作用 72h后 ,对TE 13细胞的最大抑制率分别达 (80 .5± 6 .2 ) %和 (74 .0± 5 .3) %。细胞经0 .3mmol·L- 1的ATP或ADO处理 4 8h后 ,细胞形态出现了凋亡特征 ;电泳可见明显的“梯状”条带 ;ATP和ADO可浓度依赖性地诱导细胞的凋亡 ,1mmol·L- 1的ATP和ADO诱导细胞的凋亡率分别为 (16 .6± 1.1) %和 (16 .9± 1.2 ) %。结论　ATP和ADO均有诱导食管癌TE 13细胞凋亡的作用     

小檗胺对ROCC介导的血管平滑肌细胞内游离钙的影响

目的　研究小檗胺 (BA)对受体调控性Ca2 +通道介导的家兔胸主动脉血管平滑肌细胞内游离钙 ([Ca2 +]i)的影响。方法　家兔主动脉血管平滑肌以Fluo 3/AM负载 ,通过激光扫描共聚焦显微镜 (LSCM )测定 [Ca2 +]i。结果　在细胞外Ca2 +存在的条件下 ,BA 30 μmol·L-1不影响静息[Ca2 +]i;但对去甲肾上腺素 (NE) 30mmol·L-1、5 羟色胺 (5 HT) 1μmol·L-1诱导的 [Ca2 +]i 升高有明显的抑制作用。在无胞外钙时 ,对咖啡因 40mmol·L-1诱导的 [Ca2 +]i 升高没有作用。结论　BA对ROCC激活后的外钙内流有明显的抑制作用 ,对内钙释放没有影响。其作用与维拉帕米相似     

胍丁胺对异丙肾上腺素诱发人心房纤维迟后除极的影响(英文)

用标准微电极技术研究胍丁胺对异丙肾上腺素 ( Iso)诱发人心房纤维迟后除极的影响 .结果如下 :( 1 )胍丁胺 ( 1 - 1 0 mmol· L-1)以浓度依赖地方式明显抑制 Iso( 2 0 nmol· L-1)诱发人心房纤维的迟后除极 ;( 2 )预先应用咪唑啉受体和 α2 肾上腺素受体拮抗剂咪唑克生 ( 0 .1 mmol· L-1)可阻断胍丁胺 ( 1 0 mmol· L-1)对 Iso( 2 0 nmol· L-1)诱发迟后除极的抑制作用 ;( 3)预先应用一氧化氮合酶抑制剂硝基 - L-精氨酸甲酯 ( 0 .5mmol· L-1) ,不影响胍丁胺 ( 1 0 mmol· L-1)对 Iso( 2 0 nmol· L-1)诱发迟后除极的抑制作用 .结果表明 ,胍丁胺对 Iso诱发人心房纤维迟后除极的抑制作用可能是由于咪唑啉受体和 α2 肾上腺素受体介导钙内流减少所致 .     

南蛇藤素抑制豚鼠体外精子的受精能力

南蛇藤素(Cel)对豚鼠精子前向运动(FM)、获能(Cap)、顶体反应(AR)和穿透去透明带仓鼠卵(SPA)均有明显的抑制作用,其作用强度随剂量而增加;对Cel的敏感性依次为精子Cap>FM>SPA>AR。Cel对豚鼠精子AR,FM和Cap的抑制作用比乙酸棉酚(GA)明显为强。     

辛伐他汀对高胆固醇血症小鼠血管内皮粘附单核细胞的抑制作用

目的 :观察辛伐他汀对高胆固醇血症小鼠离体胸主动脉内皮粘附单核细胞的抑制效应。方法 :用不同剂量 (1,5 ,2 0mg·kg- 1)的辛伐他汀每日1次给高胆固醇血症小鼠皮下注射共 2wk或 6wk ,届时剪取小鼠胸主动脉 ,用于单核细胞粘附试验。结果 :(1)只有 2 0mg·kg- 1× 6wk组小鼠血浆胆固醇水平显著降低 ,从 6 .6± 0 .7mmol·L- 1降为 4 .6± 0 .7mmol·L- 1,(P <0 .0 5 ) ;(2 )高胆固醇血症小鼠胸主动脉内皮对单核细胞的粘附率明显高于正常小鼠 ,辛伐他汀 2 0mg·kg- 12个组对高胆固醇血症小鼠胸主动脉内皮粘附单核细胞有明显的抑制作用 ,粘附率 (4.1± 2 .5 ) %vs (1.9± 0 .1) % (和 )或(1.4± 0 .3) %均 (P <0 .0 5 )。结论 :高胆固醇血症小鼠胸主动脉内皮对单核细胞粘附增强 ,辛伐他汀对高胆固醇血症小鼠血管内皮粘附单核细胞有明显的抑制作用     

肿瘤坏死因子相关凋亡诱导配体及扇贝多肽在UVA诱导HaCaT细胞凋亡中的作用

目的研究UVA对HaCaT细胞肿瘤坏死因子相关凋亡诱导配体(tumour necrosis factor related apoptosis inducing ligand,TRAIL)表达的影响;复制UVA诱导的HaCaT细胞凋亡模型,探究TRAIL在UVA诱导的HaCaT细胞凋亡中的作用及扇贝多肽(Polypeptide from Chlamys farreri,PCF)对UVA诱导的TRAIL凋亡通路的影响。方法实验设计分为5组:对照组、UVA模型组、UVA+5.69mmol·L-1PCF组、UVA+2.84mmol·L-1PCF组、UVA+1.42mmol·L-1PCF组。Real-TimePCR检测TRAIL mRNA表达;蛋白质印迹法检测TRAIL蛋白表达及caspase-8活性;琼脂糖凝胶电泳分析TRAIL中和性抗体对UVA诱导的HaCaT细胞凋亡的影响。结果8J·cm-2UVA照射HaCaT细胞后TRAIL mR-NA及蛋白表达增加,与对照组相比差异有显著性(P<0.01);TRAIL中和性抗体对UVA诱导的HaCaT细胞凋亡有抑制作用;1.42~5.69mmol·L-1剂量范围内的PCF可剂量依赖性抑制UVA引起的HaCaT细胞TRAIL mRNA及蛋白表达(P<0.05,P<0.01);PCF对UVA引起的HaCaT细胞caspase-8的活化有抑制作用,且呈量效关系。结论UVA可增强HaCaT细胞TRAIL表达;TRAIL参与了UVA诱导的HaCaT细胞凋亡;PCF对UVA诱导的HaCaT细胞TRAIL表达有抑制作用,也可减弱UVA诱导的caspase-8活化,以其抗氧化活性抑制TRAIL凋亡通路而发挥抗凋亡作用。     

Xenoestrogenic chemicals effectively alter sperm functional behavior in mice

Xenoestrogenic compounds (XCs) can disrupt endogenous hormone function and affect sperm function by binding to receptors on sperm membrane. Albeit spermatozoa are potentially a useful model for screening estrogenic activities of endocrine disruptors, high-quality in vitro test system that examination of the XCs effects on sperm function is required. The objective of this study was to compare the effects of XCs (genistein and 4-tert-octylphenol) to those of steroids (estrogen and progesterone) and heparin on in vitro capacitation and acrosome reaction (AR) in mouse spermatozoa. Mouse spermatozoa were incubated with various concentrations (0.001–100 μM) of each chemical for 15 or 30 min, and then capacitation and AR were assessed using chlortetracycline. All chemicals studied effectively alter capacitation and/or AR in mouse spermatozoa with different manner. Therefore, we believed that our system will provide a good in vitro model system to characterize the physiological effect of XCs especially when compared with steroids.     

Changes in protein phosphorylation during acrosome reaction of mouse sperm

The changes of protein phosphorylation were analyzed during mouse sperm capacitation and acrosome reaction. Sperm adenosine 3',5'-cyclic monophosphate (cAMP) levels were gradually elevated during capacitation period. Although 32P uptake by sperm and protein phosphorylation increased during capacitation, the phosphorylation of a 45-kDa protein remained to be relatively constant. It was also demonstrated that acrosome reaction was induced by the treatment of capacitated sperm with 0.1 mM dibutyryl cAMP, 10 mM dibutyrylguanosine 3',5'-cyclic monophosphate, 20 microM calcium ionophore A23187 or acid-solubilized zonae pellucidae. The phosphorylation of a 45-kDa protein was augmented during the acrosome reaction induced by such agents. These results suggest that changes in protein phosphorylation might be involved in the regulation of mouse sperm capacitation and acrosome reaction.     

Role of estradiol in the capacitation and acrosome reaction of hamster epididymal spermatozoa in the isolated uterus of mice incubated in vitro.

The site of sperm capacitation, the agents and mechanisms causing capacitation and acrosome reaction (AR) in vivo are not well understood. The female reproductive tract has been reported to play a key role during capacitation and AR. Some experiments were carried out on the capacitation and AR of hamster epididymal spermatozoa in the estrogen and progesterone dominated uterus (estrous and diestrous respectively) albino mice, incubated in TALP without calcium and BSA. Also the effect of estradiol (200 micrograms/ml) supplemented to TALP, on capacitation and AR was examined. Capacitation and AR of hamster spermatozoa incubated in the isolated uterus of both estrous and diestrous mice were significantly (P < 0.05) higher in the presence of exogenous estradiol than that in its absence. Acrosome shedding occurred earlier i.e. at 3rd hour as compared to the in vitro studies where it occurred at 5th hour. The present study thus reveals that uterus of both estrogen and progesterone dominated mice play an important role in the induction of capacitation and AR. The addition of estradiol might have the influence on the synthesis of uterine proteins of mice which might be important for capacitation and AR.     

Emodin inhibits human sperm functions by reducing sperm [Ca2+]i and tyrosine phosphorylation

Emodin, a bioactive anthraquinone widely used in Chinese traditional medicine, disrupts mouse testicular gene expression in vivo. In this study, we investigated the toxicity of emodin to human sperm in vitro. Different doses of emodin (25, 50, 100, 200 and 400 μM) were applied to ejaculated human sperm. The results indicated that 100, 200 and 400 μM emodin significantly inhibited the total motility, progressive motility and linear velocity of human sperm. In addition, sperm's ability to penetrate viscous medium together with progesterone induced capacitation and acrosome reaction was also adversely affected by emodin. In contrast, emodin did not affect sperm viability. Furthermore, intracellular Ca²⁺ concentration ([Ca²⁺]_i) and tyrosine phosphorylation, which serve as key regulators of sperm function, were dose-dependently reduced by emodin (50–400 μM). These results suggest that emodin inhibits human sperm functions by reducing sperm [Ca²⁺]_i and suppressing tyrosine phosphorylation in vitro.     

Matrine compromises mouse sperm functions by a [Ca2+]i-related mechanism

Matrine, a bioactive alkaloid widely used in Chinese medicine, inhibits mouse sperm functions in vitro. In this study, we investigated the reproductive toxicity of matrine to male mice in vivo. C57BL/6J mice were administered with daily doses of 0, 1, 10 and 50 mg/kg matrine by intraperitoneal injection for 30 days. The results showed that matrine did not affect testis size, testis weight, sperm count and sperm viability, but it significantly inhibited total motility, progressive motility, linear velocity, capacitation and the progesterone-induced acrosome reaction of mouse sperm. Furthermore, the intracellular Ca²⁺ concentration ([Ca²⁺]_i), a key regulator of sperm function, was reduced in sperm of matrine-exposed mice. The current and gene expression of the sperm specific Ca²⁺ channel, CatSper, which modulates Ca²⁺ influx in sperm, were decreased in testes of matrine-exposed mice. These results indicate that matrine inhibits mouse sperm functions by a [Ca²⁺]_i-related mechanism via CatSper channel.     

氯丙烯对神经细胞内 Ca2+, 游离钙调蛋白, 环腺苷酸含量和钙/钙调蛋白依赖性蛋白激酶Ⅱ活性的影响

用培养的鸡胚脑神经细胞研究了1.02和2.04 mmol·L^-1氯丙烯作用24 h后细胞内Ca²⁺, 游离钙调蛋白(CaM)和环腺苷酸(cAMP)含量及钙/钙调蛋白依赖性蛋白激酶Ⅱ(Ca²⁺/CaM-PK Ⅱ)活性的改变. 结果显示, 随着氯丙烯浓度的增加, 细胞内Ca²⁺分别增加了4.2和6.2倍, cAMP含量增加了22%和39%, Ca²⁺/CaM-PK Ⅱ活性增加了2.8和5.0倍(P<0.01); 而游离CaM含量则分别减少了55%和69%(P<0.01). 结果提示氯丙烯引起病理性神经轴浆内微管和神经微丝堆积与细胞内Ca²⁺增加有关. 这可能由于细胞内Ca²⁺增加, 激活CaM, 继而激活Ca²⁺/CaM-PK Ⅱ, 催化微管相关蛋白2和tau-蛋白磷酸化, 抑制微管组装, 使解聚的微管堆积在轴浆内.     

Antimicrobial drug ornidazole inhibits hamster sperm capacitation,in vitro

To be fertilization competent, spermatozoa undergo a series of changes in the female reproductive tract collectively referred to as capacitation. In an attempt to understand, if ornidazole, a known anti-fertility drug, adversely affects sperm functions by targeting capacitation, we designed experiments to study the influence of this drug on hyperactivation (HA), capacitation-associated protein tyrosine phosphorylation (pY) and the acrosome reaction (AR). Addition of ornidazole at 0 h, inhibited the onset of HA and total pY in a dose dependent manner. However, when ornidazole was added at 3.5 h, severe effects were still seen on HA and pY of high molecular weight proteins but, pY of lower M_r proteins (50–56 kDa) was affected only marginally. Further, lower doses of ornidazole (5 and 10 mM) had greater inhibitory effect when added at 0 h, while addition of ornidazole at 3.5 h required higher doses of ornidazole (25 mM) to cause significant inhibition of acrosome reaction. Collectively, through in vitro studies, we demonstrate that ornidazole affects the onset and progression of hamster sperm hyperactivation, capacitation associated protein tyrosine phosphorylation and acrosome reaction, and the severity depends on the dose (5, 10 or 25 mM) and the time of addition (0 or 3.5 h) of the drug to the spermatozoa.     

Antimicrobial drug ornidazole inhibits hamster sperm capacitation, in vitro

To be fertilization competent, spermatozoa undergo a series of changes in the female reproductive tract collectively referred to as capacitation. In an attempt to understand, if ornidazole, a known anti-fertility drug, adversely affects sperm functions by targeting capacitation, we designed experiments to study the influence of this drug on hyperactivation (HA), capacitation-associated protein tyrosine phosphorylation (pY) and the acrosome reaction (AR). Addition of ornidazole at 0 h, inhibited the onset of HA and total pY in a dose dependent manner. However, when ornidazole was added at 3.5 h, severe effects were still seen on HA and pY of high molecular weight proteins but, pY of lower M_r proteins (50–56 kDa) was affected only marginally. Further, lower doses of ornidazole (5 and 10 mM) had greater inhibitory effect when added at 0 h, while addition of ornidazole at 3.5 h required higher doses of ornidazole (25 mM) to cause significant inhibition of acrosome reaction. Collectively, through in vitro studies, we demonstrate that ornidazole affects the onset and progression of hamster sperm hyperactivation, capacitation associated protein tyrosine phosphorylation and acrosome reaction, and the severity depends on the dose (5, 10 or 25 mM) and the time of addition (0 or 3.5 h) of the drug to the spermatozoa.     

Hydrogen peroxide induces premature acrosome reaction in rat sperm and reduces their penetration of the zona pellucida

Recent studies have demonstrated that mammalian sperm are capable of generating reactive oxygen species (ROS) and that this activity is significantly accelerated in subfertile subjects. The observed decrease in penetration of zona-intact oocyte might be explained by chemical-induced ROS-related early onset of capacitation and premature acrosome reaction, but the mechanism is not clear. We determine whether zona-intact oocyte penetration capability in rat epididymal sperm was affected by premature acrosome reaction in rat sperm treated with hydrogen peroxide (H2O2) and calcium ionophore A23187 or H2O2 and lysophosphatidyl choline. Chlortetracycline fluorescence assay was used to study the status of acrosome reaction on epididymal sperm. The sperm-oocyte binding and penetration assay was used to evaluate the capability for zona pellucida penetration. There was a positive linear correlation between the frequency of acrosome-reacted sperm and capability of sperm-oocyte binding and penetration in zona-free oocytes. In the zona-intact oocytes, the sperm-oocyte penetration rate was suppressed as the proportions of acrosome-reacted sperm increased. In summary, this study showed that premature acrosome reaction reduced rat sperm's capability of penetrating zona-intact oocytes. However, this reduction is not seen in zona-free oocytes. These findings may provide a basis for understanding the effects of sperm ROS generation on zona pellucida penetration in male reproductive toxicology.