TLR3在poly（I：C）-LMW预处理后缺糖缺氧诱导的原代皮质神经元的表达意义

目的观察TRL3激动剂poly（I：C）-LMW预处理对原代皮质神经元Toll样受体3（TLR3）表达的影响,探讨TLR3在缺糖缺氧诱导的原代皮质神经元损伤中的作用。方法取体外培养7d的大鼠原代皮质神经元细胞,对细胞分别予以正常条件下培养（空白对照组）;缺糖缺氧2h,复糖复氧24h（OGD组）;TRL3激动剂预处理12h（激动剂组）;TRL3激动剂预处理12h后缺糖缺氧2h,复糖复氧24h（激动剂＋OGD组）。免疫荧光法观察各组细胞生长状态,分别以Western blot法、RT-PCR法测定TLR3蛋白及TLR3mRNA的表达情况。结果 与空白对照组相比,激动剂及缺氧复氧处理方法均诱导原代皮质神经元TLR3、TLR3mRNA表达增强（P〈0.05）;激动剂预处理后,与OGD组相比,激动剂＋OGD组神经元细胞状态较好,数目较多（P〈0.05）。结论 TLR3参与缺糖缺氧诱导的原代皮质神经元的损伤过程,激动剂可减轻神经元缺糖缺氧后损伤。     

低氧预适应小鼠脑匀浆液提取液对大鼠鼠胚海马神经元缺氧复氧后神经细胞的影响

目的 探讨低氧预适应小鼠脑匀浆液提取液对大鼠鼠胚海马神经元缺氧复氧后神经细胞活性和凋亡的影响. 方法在96孔培养板中将大鼠鼠胚海马神经细胞原代培养至8 d,将培养细胞按照处理的不同分为以下5组:(1)正常对照组(仅加入PBS)、(2)H<,4>R<,48>组(缺氧4 h/复氧48h后加PBS)、(3)H0组(缺氧4 h/复氧48 h前加正常小鼠脑匀浆提取液)、(4)H1组(缺氧4h/复氧48h前加急性低氧对照小鼠脑匀浆提取液)、(5)H4组(缺氧4 h/复氧48 h前加低氧预适应小鼠脑匀浆提取液),分别用酶标仪和流式细胞仪测定神经细胞活性和凋亡情况.结果 正常对照组细胞活性明显高于H<,4>R<,48>组,H0、H1和H4组分别与H<,4>R<,48>组相比,细胞活性明显增加,且H4组细胞活性又明显高于H0、H1组;正常对照组仅有极少量的凋亡细胞,而H<,4>R<,48>组凋亡细胞显著增多,H0、H1和H4组分别与H<,4>R<,48>组相比凋亡细胞明显减少,且H4组又分别明显少于H0、H1组.结论低氧预适应小鼠脑匀浆液提取液可能通过增加大鼠鼠胚海马神经元缺氧复氧后神经细胞活性和减少神经细胞凋亡起到抗缺氧性损伤作用.     

黄体酮对大鼠脑微血管内皮细胞氧糖剥夺后的保护作用

目的研究黄体酮(progesterone,PROG)对大鼠脑微血管内皮细胞氧糖剥夺后的保护作用机制。方法体外培养大鼠脑微血管内皮细胞(brain microvascular endothelial cell,BMEC),并构建氧糖剥夺(oxygen and glucose deprivation,OGD)模型。以3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐[3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide,MTT]法检测BMEC存活率,比色法测定乳酸脱氢酶(lactate dehydrogenase,LDH)漏出率反映细胞损伤程度。为反映BMEC的氧化应激损伤水平,利用生物化学法测定细胞培养液中及细胞内丙二醛(malondialdelyde,MDA)水平及细胞内超氧化物歧化酶(superoxide dismutase,SOD)和谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)活性。结果 OGD 6h明显降低BMEC的存活率,并增加LDH释放率。OGD 6h可明显增加BMEC培养基中及细胞内MDA含量,减少BMEC内SOD、GSH-Px活性。与氧糖剥夺组相比,黄体酮可明显降低培养基中和细胞内的MDA含量,提高BMEC内SOD、GSH-Px活性。结论 OGD可通过诱导氧化应激等途径损伤BMEC,黄体酮能减少OGD对BMEC造成的损伤。     

一氧化氮在缺氧复氧性神经元凋亡中的作用

目的观察神经元缺氧复氧损伤时NO的动态变化及其与神经元凋亡的关系。方法取孕13~15d ICR小鼠,无菌条件下对胎鼠大脑皮质神经元进行原代分离培养,建立缺氧复氧诱导的皮质神经元凋亡模型。用流式细胞仪检测细胞凋亡率,用硝酸还原酶法测定细胞培养上清液中NO的含量,用MTT测定细胞活性,并在电镜下观察细胞形态学变化。结果经缺氧复氧处理的神经元,随着缺氧时间延长细胞存活率逐渐下降,神经元凋亡率呈时间依赖性升高,NO含量逐渐升高,至缺氧8h复氧24h达到高峰,与正常对照组比较有显著性差异(P<0.01)。细胞超微结构呈现凋亡样改变。结论NO介导了缺氧复氧性神经元凋亡过程。     

氢气预处理对乳鼠海马神经元细胞缺氧/复氧损伤后细胞活力的影响

目的探讨氢气对乳鼠海马神经元细胞缺氧/复氧损伤后细胞活力的影响。方法原代培养的SD乳鼠海马神经元细胞,随机分成对照组,缺氧/复氧组和氢气预处理组。对照组常规培养;缺氧/复氧组在100%N2中培养15min后复氧30min;氢气预处理组在2%H2+98%N2中培养15min后复氧30min。采用MTT法检测乳鼠海马神经元细胞活力。结果氢气预处理能够增强细胞活力(P<0.05)。结论氢气预处理对海马神经元细胞缺氧/复氧损伤具有保护作用,可能与提高细胞活力有关。     

PC12细胞化学缺氧复氧损伤与缺氧诱导因子1表达的关系

目的 研究缺氧诱导因子1(HIF1)在PC12细胞化学缺氧复氧损伤中的作用。方法 在培养液中加入和去除氯化钴模拟化学缺氧和复氧，以乳酸脱氢酶(LDH)漏出和细胞超微结构改变作为细胞损伤指标，观察化学缺氧和复氧后不同时间细胞损伤和HIF1 α蛋白变化。结果 在氯化钴模拟化学缺氧实验中，LDH漏出明显增加，8h达高峰，随后逐渐下降，电镜结果与LDH改变相一致，HIF1 α蛋白表达在化学缺氧后2，4，8和12h均明显增加，8h达高峰，提示化学缺氧8h后细胞损伤逐渐减轻可能与HIF1 α蛋白水平升高有关。在模拟复氧实验中，LDH和细胞形态学改变都显示化学复氧8h细胞损伤最为严重，而HIF1 α蛋白表达在化学复氧4和8h均明显下降，提示细胞化学复氧损伤可能与HIF1 α蛋白水平下降有关。结论 HIF1对神经细胞化学缺氧复氧损伤具有保护作用。     

缺氧预适应对体外培养神经元的神经保护作用存在一个时间窗

目的本实验采用体外培养皮质神经元研究缺氧预适应对谷氨酸损伤模型的保护作用,同时探索一定缺氧浓度下最佳保护作用的缺氧时间。方法原代培养神经元,确立谷氨酸损伤模型,对培养至第7天的细胞进行8%氧浓度和不同时间的缺氧预适应及谷氨酸损伤,用MTT法检测细胞的存活率。应用SPSS 17.0进行统计学分析。结果 8%O2缺氧4~8 h,皮质神经元谷氨酸损伤模型实验组与对照组相比,存活率提高,差异有统计学意义(P<0.05)。结论 8%O2缺氧4~8 h对谷氨酸损伤模型中的原代体外培养皮质神经元具有保护作用,该保护作用至少可持续6 h。     

P~(38)激酶在PC12细胞缺氧/再给氧损伤中的作用

目的:探讨PC12细胞缺氧/再给氧损伤的信号转导机理。方法:培养的PC12细胞先缺氧(95％N2/5％CO2)6h,然后重新给氧,观测不同时间点细胞的存活率和caspase-3的活性;用MTT法测存活率,caspase-3检测试剂盒测caspase-3活性。用p38拮抗剂SB203580孵育细胞2h,之后缺氧/再给氧,观察SB203580对细胞存活率和caspase-3活性的影响。结果:PC12细胞缺氧/再给氧后caspase-3活性明显增加并使细胞存活率下降,SB203580明显降低缺氧/复氧后caspase-3的活性并使细胞死亡减少。结论:PC12细胞缺氧/再给氧后至少可以通过激活p38、caspase-3信号分子诱导PC12细胞死亡。     

依达拉奉对培养乳鼠海马神经元缺氧复氧损伤的保护作用

目的:探讨依达拉奉对海马神经元缺氧复氧损伤的保护作用及其机制。方法:制备培养乳鼠海马神经元缺氧复氧损伤模型,分别用依达拉奉1、10、100、300μmol·L-1干预后研究依达拉奉对脂质过氧化及细胞凋亡的作用。结果:海马神经元在缺氧复氧损伤后,依达拉奉干预能降低脂质过氧化产物丙二醛(MDA)含量,增加细胞内超氧化物歧化酶(SOD)活性,减少神经元凋亡。结论:缺氧复氧损伤后,依达拉奉具有清除氧自由基、抑制脂质过氧化及降低细胞凋亡率的作用。     

凋亡敏感基因在短暂性缺氧再复氧复注血清后的大鼠星形胶质细胞中的表达及其意义

目的研究一种新发现的抗氧化蛋白质--凋亡敏感基因(SAG)在短暂性缺氧再复氧复注血清诱导的细胞坏死和凋亡中的作用.方法用短暂性缺氧再复氧复注血清来诱导原代培养的大鼠大脑皮质星形胶质细胞损伤,用免疫细胞化学方法检测凋亡敏感基因的表达,并作图像分析;用流式细胞仪检测胶质细胞在短暂缺氧再复氧复注血清后不同时间点的凋亡率.结果凋亡敏感基因在缺氧15 min再复氧复注血清5 h后表达最高,在复氧复注血清16 h后恢复至对照组水平;细胞凋亡率在缺氧15 min再复氧复注血清1h时达到最高,而在再复氧复注血清5 h后降至对照组水平.结论凋亡敏感基因具有抗凋亡的作用,可减轻星形胶质细胞的缺血再灌注损伤.     

Death of motoneurons induced by trophic deprivation or by excitotoxicity is not prevented by overexpression of SMN

The telomeric copy of the survival motor neuron gene (SMN1) is deleted or mutated in all spinal muscular atrophy (SMA) patients and these patients present mainly a loss in spinal motoneurons. Although studies performed in HeLa cells suggest that SMN may be involved in the biogenesis and possibly in recycling of spliceosomal small nuclear ribonucleoproteins (snRNPs), no link has been established between this function and the consequence of the absence of SMN in the specific loss of motoneurons. We attempted to answer the question of whether SMN plays a direct role in motoneuron survival by transducing cultured motoneurons with lentiviral vectors coding either for an antisense Smn mRNA or for full-length or truncated forms of SMN. We studied their effect on survival under different anti- or proapoptotic culture conditions. Our results show that increased levels of SMN are unable to protect motoneurons from death induced by trophic deprivation or by excitotoxicity. These results suggest that SMN is not a survival factor per se for motoneurons. In addition, overexpression of a truncated form of SMN shown to induce a modified subcellular localization and to exert a dominant-negative effect on snRNP biogenesis and RNA splicing in HeLa cells was ineffective in modifying both localization and survival in motoneurons.     

Stimulation of glucose analogue uptake by cerebral microvessel endothelial cells by a product released by astrocytes

Astroglial cells, both normal and neoplastic, secreted a product that stimulated glucose uptake by cerebral microvessel endothelial cells by 23% and 50%, respectively. Neither cerebral microvessel smooth muscle cells nor oligodendrocytes affected endothelial cell glucose uptake. The astrocytic product(s) did not affect glucose uptake by aortic endothelial cells. The effect on the cerebral microvessel endothelial cells increased with increasing time of exposure of the cells to the astroglial product(s), and required the constant presence of the astrocytic product to be maintained. The presence of a protein synthesis inhibitor during endothelial cell exposure to the astroglial conditioned medium blocked the stimulation of glucose uptake. Treatment of the astrocytic product with a protease destroyed its effectiveness. These results support the hypothesis that astrocytes induce the expression of at least one blood-brain barrier property by the cerebral microvasculature, and suggest that this induction may be produced by a protein released by the astrocytes.     

Antagonism by haloperidol of locomotor depression induced by small doses of apomorphine

Summary Administration of 0.025–0.1 mg/kg of apomorphine i.p. to mice produced a dose-dependent locomotor depression. Haloperidol, 0.025 mg/kg, produced locomotor stimulation, whereas 0.1 mg/kg caused locomotor depression. Pretreatment with haloperidol also reversed the depression caused by apomorphine. The functional antagonism is discussed in terms of a possible agonist-antagonist interaction on dopaminergic autoreceptors.     

Infertility by choice or by nature

Sexually active women have only recently had the ability to make a conscious decision to delay or refrain from bearing children. This is not only the result of the availability of effective contraceptive methods but also due to attitudinal changes in society and individuals. These reproductive choices may result, for some women, in conflict over the use of contraceptives, and the decision or timing of pregnancy. However, infertility imposed by nature in the form of inability to conceive, miscarriage or stillbirth removes the woman's sense of control over this important aspect of her life and frequently results in severe distress. The psychological issues surrounding these reproductive choices and events are reviewed and discussed.     

Activation of human platelets by 2-arachidonoylglycerol is enhanced by serotonin

The endocannabinoid 2-arachidonoylglycerol (2-AG) has been shown to activate human platelets in platelet-rich plasma, by binding to a "platelet-type" cannabinoid receptor (CB(PT)). Here, washed human platelets were used to characterize the binding of [(3)H]2-AG to CB(PT), showing a dissociation constant (Kd) of 140 +/- 31 nM and a maximum binding (Bmax) of 122 +/- 10 pmol.mg protein(-1). Selective antagonists of both CB1 and CB2 cannabinoid receptors inhibited this binding, which was enhanced up to approximately 230% over the controls by 1 micro M serotonin (5-hydroxytryptamine, 5-HT). Human platelets were also able to bind [(3)H]5-HT (Kd = 79 +/- 17 nM, Bmax = 14.6 +/- 1.3 pmol.mg protein(-1)), and 1 micro M 2-AG enhanced this binding up to approximately 150%. Moreover, they were able to take up [(3)H]5-HT through a high affinity transporter (Michaelis-Menten constant = 22 +/- 2 nM, maximum velocity = 344 +/- 15 pmol.min(-1).mg protein(-1)), which was not affected by 2-AG. Interestingly, 5-HT did not affect the activity of the 2-AG transporter of human platelets. Treatment of washed platelets with 1 micro M 2-AG led to increased intracellular inositol-1,4,5-trisphosphate (up to approximately 300%) and decreased cyclic AMP (down to approximately 50%). Furthermore, treatment of pre-loaded platelets with 1 micro M 2-AG induced a approximately 300% increase in [(3)H]2-AG release, according to a CBPT-dependent mechanism. Also, 1 micro M 5-HT enhanced the effect of 2-AG on inositol-1,4,5-trisphosphate ( approximately 500% of the controls), cyclic AMP ( approximately 20%) and [(3)H]2-AG release ( approximately 570%), and the latter process was shown to be partly ( approximately 50%) involved in the 5-HT-dependent platelet activation. Taken together, reported findings represent the first demonstration that 2-AG and 5-HT can mutually reinforce their receptor binding on platelet surface, which might have therapeutic implications.