脂质立方液晶设计制备与表征评价研究进展

目的：研究伊拉地平（ ISR ）对1-甲基-4-苯基吡啶离子（ MPP＋）损伤的PC12细胞的保护作用及可能机制。方法MPP ＋处理PC12细胞建立帕金森病细胞模型;4-甲基偶氮唑蓝（ MTT）比色法检测细胞存活率;双氯荧光黄乙酸乙酯（ DCFH-DA）染色流式细胞术检测细胞内活性氧（ ROS）的生成;JC-1染色流式细胞术检测细胞线粒体膜电位（ MMP）。结果1 mmol · L－1MPP＋处理PC12细胞24 h后能明显抑制细胞生长（P＜0．01）;降低线粒体膜电位;ROS含量增加。2μmol· L－1伊拉地平预处理后, PC12细胞存活率显著增加（ P＜0．01）;线粒体膜电位升高;ROS生成减少。结论伊拉地平对MPP＋损伤的PC12细胞具有保护作用,其作用机制可能与维持线粒体正常膜电位,稳定线粒体功能,阻止线粒体氧化应激发生有关。     

瓜子金皂苷丙对MPP~+诱导PC12细胞凋亡的抑制作用

目的观察瓜子金皂苷丙对MPP+诱导的PC12细胞凋亡的影响,并且探讨其作用机制。方法采用MTT法检测细胞存活率,碘化丙啶染色流式细胞术(FCM)检测PC12细胞凋亡,Western blotting检测Bax和Bcl-2蛋白的表达,罗丹明123染色FCM检测细胞线粒体膜电位(ΔΨm),荧光酶标仪检测细胞内活性氧(R0S)的含量。结果不同浓度MPP+作用PC12细胞24 h后,细胞存活率显著下降(P<0.01),细胞凋亡明显,凋亡相关蛋白Bcl-2/Bax比之下降,线粒体膜电位降低,细胞内ROS显著增加。与MPP+处理组相比,瓜子金皂苷丙10μmol·L-1组,细胞存活率显著升高(P<0.01);细胞凋亡率下降(P<0.01);Bcl-2/Bax比率增加(P<0.01),线粒体膜电位上升(P<0.01),ROS含量减少。结论瓜子金皂苷丙可以抑制MPP+诱导的PC12细胞凋亡,其作用机理可能与上调Bcl-2和下调Bax蛋白的表达,维持线粒体正常膜电位,稳定线粒体功能,清除R0S有关。     

MPP 诱导PC12细胞发生Ferroptosis（铁死亡）的实验观察

[摘要] 目的 1-甲基-4-苯基吡啶离子（1-methyl-4-phenyl pyridine,MPP ）诱导PC12细胞铁死亡（Ferroptosis）的研究。方法：采用MTT法检测细胞存活率和筛选Ferrostatin-1的最佳作用浓度;倒置显微镜观察Ferrostatin-1对PC12细胞的保护作用;MDC染色检测细胞自噬;ELISA法检测caspase-3的酶活性;流式细胞术检测活性氧ROS流式细胞术检测活性氧ROS。结果 研究结果表明1mmol/L MPP 对PC12细胞有明显抑制作用;5μmol/L Ferrostatin-1能够明显提高PC12细胞的存活率;倒置显微镜观察结果表明Ferrostatin-1预保护后,PC12细胞损伤明显减少;MDC染色检测细胞自噬和ELISA法检测caspase-3的酶活性结果显示Ferrostatin-1对MPP 诱导的PC12细胞自噬和凋亡的保护作用不明显;ROS活性氧检测结果显示ROS在胞内增加,可能发生氧化应激,加Ferrostatin-1后ROS荧光强度减弱。ROS活性氧检测结果显示ROS在胞内增加,可能发生氧化应激,加Ferrostatin-1后ROS荧光强度减弱。结论 铁死亡抑制剂Ferrostatin-1能够显著抑制MPP 诱导的PC12细胞损伤,且Ferrostatin-1对细胞的自噬和凋亡作用不明显,推测出MPP 损伤PC12细胞的帕金森病模型中可能有铁死亡的存在。     

瓜子金皂苷己对MPP~+诱导PC12细胞凋亡的保护作用

目的观察瓜子金皂苷己(polygalasaponin F,PS-F)对1-甲基-4-苯基-吡啶离子(1-methyl-4-phenylpyridinium,MPP+)诱导的PC12细胞损伤的影响,并且探讨其作用机制。方法 MTT法检测细胞存活率,Annexin V/PI染色流式细胞术检测PC12细胞凋亡,JC-1染色倒置显微镜检测细胞线粒体膜电位(mitochondrial membrane potential,MMP),Western blot检测Caspase-3蛋白的水平。结果 500μmol.L-1MPP+作用PC12细胞48 h,能明显抑制细胞生长(P<0.01),诱导细胞发生凋亡,同时降低MMP,增加活性Caspase-3的蛋白水平。同时给予不同浓度PS-F处理,PC12细胞存活率增加(P<0.01);凋亡细胞量减少;MMP增高;活性Caspase-3蛋白水平降低(P<0.01)。结论 PS-F能抑制MPP+诱导的PC12细胞的凋亡,其作用机制可能与维持线粒体正常膜电位,稳定线粒体功能,降低活性Caspase-3蛋白水平有关。     

1-甲基-4-苯基吡啶离子诱导大鼠肾上腺嗜铬细胞瘤PC12细胞发生铁死亡的实验观察

目的 进行1-甲基-4-苯基吡啶离子(1-methyl-4-phenyl pyridine,MPP+)诱导大鼠肾上腺嗜铬细胞瘤PC12细胞铁死亡(Ferroptosis)的研究.方法 采用噻唑蓝法(MTT)检测细胞存活率和筛选铁死亡抑制剂(Ferrostatin-1)的最佳作用浓度;倒置显微镜观察Ferrostatin-1对PC12细胞的保护作用;MDC染色检测细胞自噬;ELISA法检测caspase-3的酶活性;流式细胞术检测活性氧ROS.结果 1 mmol·L-1 MPP+对PC12细胞有明显抑制作用;5 μmol·L-1 Ferrostatin-1能够明显提高PC12细胞的存活率;倒置显微镜观察结果表明Ferrostatin-1预保护后,PC12细胞损伤明显减少;MDC染色检测细胞自噬和ELISA法检测caspase-3的酶活性结果显示Ferrostatin-1对MPP+诱导的PC12细胞自噬和凋亡的保护作用不明显;ROS活性氧检测结果显示ROS在胞内增加,可能发生氧化应激,加Ferrostatin-1后ROS荧光强度减弱.结论 Ferroptosis能够显著抑制MPP+诱导的PC12细胞损伤,且Ferrostatin-1对细胞的自噬和凋亡作用不明显,推测出MPP+诱导PC12细胞的损伤可能有Ferroptosis的存在.     

7,8-二羟基-4-甲基香豆素对谷氨酸诱导的PC12细胞损伤的神经保护作用

目的探讨7,8-二羟基-4-甲基香豆素(Dhmc)对谷氨酸诱导的PC12细胞损伤的神经保护作用。方法采用谷氨酸诱导PC12细胞损伤凋亡模型,给予Dhmc处理后,MTT法检测细胞增殖;化学检测法测定乳酸脱氢酶(LDH)漏出量、细胞上清液中丙二醛(MDA)含量及超氧化物歧化酶(SOD)水平;采用Annexin-V-FITC双染法检测细胞凋亡率;采用荧光法检测活性氧(ROS)的生成;采用流式细胞仪检测线粒体膜电位(MMP);Western blot法检测Bcl-2、Bax、Cyt C、cleaved caspase-3蛋白表达。结果 Dhmc可显著降低由谷氨酸诱导的细胞损伤,提高了细胞存活率,减少LDH漏出量,降低MDA含量,增加SOD活性,抑制ROS的生成,维持线粒体膜电位水平,下调Bcl-2/Bax比率,减少Cyt C的释放,降低Caspase-3活性。结论 Dhmc具有减轻由谷氨酸诱导的PC12细胞损伤的作用,其机制可能与抗氧化应激和减轻线粒体损伤有关。     

α-硫辛酸注射液对过氧化氢致PC12细胞损伤的保护作用

目的 研究α-硫辛酸注射液(抗氧化剂)对H2O2致PC12细胞损伤的保护作用.方法 600 μmol·L-1H2O2与细胞共同孵育4 h,建立PC12细胞氧化应激损伤模型,用MTT法,测定细胞生长抑制率;培养介质中的LDH测定,用紫外分光光度法,用流式细胞术测定细胞凋亡率和线粒体膜电位.结果 与模型组相比,3个剂量(10,1 μmol·L-1)α-硫辛酸注射液能提高H2O2损伤的PC12细胞的存活率,减少LDH的漏出,降低细胞凋亡率,抑制线粒体膜电位的下降(P＜0.05,P＜0.01).结论 α-硫辛酸注射液对H2O2致PC12细胞损伤模型具有较好的保护作用.     

右美托咪定对抗化学性低氧引起的PC12细胞损伤

目的探讨α2肾上腺素受体激动剂右美托咪定能否保护PC12细胞对抗化学性低氧引起的损伤。方法应用化学性低氧模拟剂氯化钴(CoCl2)处理PC12细胞以建立化学性缺氧损伤模型。应用CCK-8比色法检测细胞存活率;Ho-chest 33258核染色法观察细胞凋亡的形态学和数量的改变;双氯荧光素(DCFH-DA)染色荧光显微镜检测细胞内的活性氧(ROS)水平;罗丹明123(Rh123)染色荧光显微镜检测线粒体膜电位(MMP)。结果在浓度为100～600μmol.L-1的范围内,右美托咪定浓度依赖性地对抗CoCl2引起的细胞毒性,使细胞存活率明显增加。400μmol.L-1右美托咪定能抑制CoCl2的致凋亡作用,使凋亡细胞数量减少。右美托咪定也能抑制CoCl2引起的ROS过度生成及MMP降低的作用。结论右美托咪定能保护PC12细胞对抗CoCl2诱导的损伤,此作用可能与其抑制ROS过度生成及保护MMP有关。     

二氢石蒜碱对过氧化氢损伤的PC12细胞的保护作用

目的:探讨二氢石蒜碱(DL)对H2O2诱导的PC12细胞氧化损伤的影响及其可能机制。方法:用H2O2(200μmol.L-1)处理PC12细胞建立氧化应激模型,并加入二氢石蒜碱预处理作为保护。噻唑蓝(MTT)法和乳酸脱氢酶(LDH)检测细胞存活率和细胞损伤程度,利用荧光探针DCFH-DA和JC-1分别检测细胞内活性氧(ROS)和线粒体膜电位。结果:H2O2作用于PC12细胞后,细胞存活率下降,LDH活性和ROS含量增高,线粒体膜电位降低,与正常对照组比较具有显著性差异(P<0.01);10-7～10-5mol.L-1DL预处理后,细胞存活率提高,LDH和ROS变低,线粒体膜电位回升,且在一定范围呈剂量依赖性。结论:DL对H2O2诱导PC12细胞氧化损伤具有保护作用,其作用机制可能与减少ROS产生和稳定线粒体膜电位有关。     

新型孕激素衍生物3α-羟基-3β-甲氧基甲基-16,17-亚甲基-孕甾-20-酮对硝普钠损伤PC12细胞的保护作用

目的探讨新型孕激素衍生物3α-羟基-3β-甲氧基甲基-16,17-亚甲基-孕甾-20-酮(CPU)对硝普钠(SNP)诱导的PC12细胞损伤的神经保护作用及其机制。方法用终浓度为0.01,0.10,1.00μmol·L-1的CPU预处理30 min,然后加入SNP共同作用24 h。采用MTT法检测细胞存活率,分光光度计法检测乳酸脱氢酶(LDH)漏出率、丙二醛(MDA)含量和超氧化物歧化酶(SOD)活性的变化;流式细胞仪检测活性氧(ROS)、线粒体膜电位和细胞凋亡率;Western蛋白质印迹法检测Bcl-2、Bax和活化的胱天蛋白酶3蛋白表达水平。结果与细胞对照组比较,SNP模型组PC12细胞的存活率显著降低,LDH的释放明显增加(P<0.01),MDA生成增多,SOD活性降低,ROS水平升高,线粒体膜电位水平降低,细胞凋亡率由细胞对照组(1.20±0.14)%增加至(55.52±3.56)%(P<0.01);Bcl-2蛋白表达降低,而Bax表达升高,Bax/Bcl-2比值升高,同时胱天蛋白酶3被激活(P<0.01)。与模型组比较,CPU 0.01,0.10和1.00μmol·L-1作用24 h后,细胞存活率显著升高,LDH的释放减少(P<0.01),MDA的生成量减少,SOD活性增加,ROS累积减少,线粒体膜电位水平升高(P<0.01);凋亡细胞分别降至(37.79±4.85)%,(22.31±4.25)%和(10.39±2.65)%;Bcl-2蛋白表达增加,Bax和活化的胱天蛋白酶3表达降低。结论 CPU对SNP所致损伤的PC12细胞有一定的保护作用,其作用机制可能与其抗氧化和抗凋亡作用有关。     

Efficacy of gut lavage,hemodialysis, and hemoperfusion in the therapy of paraquat or diquat intoxication

Clinical and in vitro investigations were carried out to test the efficacy of gut lavage, hemodialysis, and hemoperfusion in the treatment of poisoning with paraquat or diquat. In a patient suffering from diquat intoxication 130 times more diquat was removed by gut lavage 30 h after ingestion than was removed by complete aspiration of the gastric contents.Determination of in vitro clearances for paraquat and diquat by hemodialysis showed that, at serum concentrations of 1–2 ppm, such as are frequently encountered in poisoning in man, toxicologically relevant quantities of herbicide cannot be removed from the body. At a concentration of 20 ppm, on the other hand, hemodialysis proved to be effective, the clearance being 70 ml/min at a blood flow rate of 100 ml/min. The efficacy of hemoperfusion with coated activated charcoal was on the whole better. Especially at concentrations around 1–2 ppm, the clearance values for hemoperfusion were some 5–7 times higher than those for hemodialysis.In a patient suffering from paraquat poisoning, both hemodialysis as well as hemoperfusion were carried out. The in vitro results could be confirmed: At serum concentrations of paraquat less than 1 ppm no clearance could be obtained by hemodialysis while by hemoperfusion with activated charcoal quite high clearance values were measured and the serum level dropped down to zero.

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Zusammenfassung Klinische Untersuchungen und Laboratoriumsversuche wurden durchgeführt, um die Wirksamkeit von Darmspülung, Hämodialyse und Hämoperfusion bei Paraquat- und Deiquat-Vergiftungen zu prüfen.Bei einem Patienten wurde 30 Std nach Deiquat-Aufnahme durch Darmspülung 130mal mehr Deiquat entfernt als durch vollständige Aspiration des Mageninhaltes. In vitro-Versuche ergaben, daß bei Blutserumkonzentrationen von 1–2 ppm, die bei Vergiftungen oft gemessen werden, durch Hämodialyse keine toxikologisch relevanten Paraquat- oder Deiquat-Mengen entfernt werden können. Dagegen erwies sich die Hämodialyse bei 20 ppm und einer Blutumlaufgeschwindigkeit von 100 ml/min mit einer Clearance von 70 ml/min als wirksam. Die Hämoperfusion mit beschicheter Aktivkohle war in diesen Versuchen aber eindeutig überlegen, denn insbesondere bei Konzentrationen um 1–2 ppm waren die Clearance-Werte 5–7mal höher als bei der Hämodialyse.Die in vitro-Ergebnisse wurden bei einem Patienten mit einer Paraquat-Vergiftung bestätigt: Bei Konzentrationen unter 1 ppm war die Hämodialyse wirkungslos, während durch Hämoperfusion relativ hohe Clearance-Werte erreicht wurden, so daß der Serumspiegel rasch unter die Nachweisgrenze abfiel.

     

Aquatic Insects and Trace Metals: Bioavailability,Bioaccumulation, and Toxicity

Abstract

The uptake of metals from food and water sources by insects is thought to be additive. For a given metal, the proportions taken up from water and food will depend both on the bioavailable concentration of the metal associated with each source and the mechanism and rate by which the metal enters the insect. Attempts to correlate insect trace metal concentrations with the trophic level of insects should be made with a knowledge of the feeding relationships of the individual taxa concerned. Pathways for the uptake of essential metals, such as copper and zinc, exist at the cellular level, and other nonessential metals, such as cadmium, also appear to enter via these routes. Within cells, trace metals can be bound to proteins or stored in granules. The internal distribution of metals among body tissues is very heterogeneous, and distribution patterns tend to be both metal and taxon specific. Trace metals associated with insects can be both bound on the surface of their chitinous exoskeleton and incorporated into body tissues. The quantities of trace meals accumulated by an individual reflect the net balance between the rate of metal influx from both dissolved and particulate sources and the rate of metal efflux from the organism. The toxicity of metals has been demonstrated at all levels of biological organization: cell, tissue, individual, population, and community. Much of the literature pertaining to the toxic effects of metals on aquatic insects is based on laboratory observations and, as such, it is difficult to extrapolate the data to insects in nature. The few experimental studies in nature suggest that trace metal contaminants can affect both the distribution and the abundance of aquatic insects. Insects have a largely unexploited potential as biomonitors of metal contamination in nature. A better understanding of the physico-chemical and biological mechanisms mediating trace metal bioavailability and exchange will facilitate the development of general predictive models relating trace metal concentrations in insects to those in their environment. Such models will facilitate the use of insects as contaminant biomonitors.     

Alzheimer’s disease – current trends in basic and clinical research. Highlights from the 16th ECNP

Advances in the molecular biological knowledge of neuronal nicotinic acetylcholine receptors (nAChRs) have led to a growing interest by the pharmaceutical industry in the development of novel compounds that selectively modulate nAChR function. The ability of (-)-nicotine, an activator of nAChRs, to enhance attentional aspects of cognition in animals and humans, to exert neuroprotective and anxiolytic-like effects, and presumably to mediate the negative correlation between smoking and Alzheimer's (and Parkinson's) Disease, has focused interest on the potential therapeutic utility of modulators of nAChR function for treatment of some of the deficits associated with these progressive, neurodegenerative conditions. Numerous compounds are known which activate nAChRs and which might serve as lead compounds toward the development of such agents. The pharmacologic diversity of neuronal nAChR subtypes suggests the possibility of developing selective compounds which would have more favourable side-effect profiles than existing agents. This broader class of agents, collectively called cholinergic channel modulators (ChCMs), is anticipated to encompass compounds which would have more favourable side-effect profiles than existing agents, which generally exhibit low selectivity. This selectivity may be achieved by preferentially activating some subtypes of nAChRs (i.e., Cholinergic Channel Activators, ChCAs) or inhibiting the function of other subtypes (Cholinergic Channel Inhibitors, ChCIs). An overview of the biology of nAChRs and the rationale for the use of ChCMs for the treatment of dementia related to neurodegenerative diseases are presented, followed by a discussion of lead compounds and compounds under consideration for clinical evaluation.     

2-(Acetoxyphenyl)-(Z)-styryl sulfides as cyclooxygenase inhibitors

2-(Acetoxyphenyl)-(Z)-styryl sulfides are described as selective cyclooxygenase-2 (COX-2) inhibitors, useful for treating inflammation and COX-2-mediated disorders including neoplasia. 2-(Acetoxyphenyl)-(Z)-styryl sulfide is claimed to be the most potent COX inhibitor in the series with a COX-2 selectivity ratio of 33. This compound is also claimed to be superior to celecoxib (Celebrex^®, Pfizer) in inhibiting cell growth of colorectal carcinoma cells. In this evaluation, the COX inhibitory activity of this compound is compared to that previously disclosed for diarylheterocycles and 2-(acetoxyphenyl)alkyl sulfides. The validity of the DLD-1 cell line in the growth inhibition studies is questioned based on recent literature reports indicating the lack of COX-2 expression in this cell line.     

Sleep-disordered breathing with chronic opioid use

Chronic opioid use for pain relief or as substitution therapy for illicit drug abuse is prevalent in our societies. In the US, retail distribution of methadone and oxycodone has increased by 824 and 660%, respectively, between 1997 and 2003. μ-Opioids depress respiration and deaths related to illicit and non illicit chronic opioid use are not uncommon. Since 2001 there has been an emerging literature that suggests that chronic opioid use is related to central sleep apnoea of both periodic and non-periodic breathing types, and occurs in ～ 30% of these subjects. The clinical significance of these sleep-related abnormalities are unknown. This review addresses the present knowledge of control of ventilation mechanisms during wakefulness and sleep, the effects of opioids on ventilatory control mechanisms, the sleep-disordered breathing found with chronic opioid use and a discussion regarding the future research directions in this area.     

Drug targets for cognitive enhancement in neuropsychiatric disorders

The investigation of novel drug targets for treating cognitive impairments associated with neurological and psychiatric disorders remains a primary focus of study in central nervous system (CNS) research. Many promising new therapies are progressing through preclinical and clinical development, and offer the potential of improved treatment options for neurodegenerative diseases such as Alzheimer's disease (AD) as well as other disorders that have not been particularly well treated to date like the cognitive impairments associated with schizophrenia (CIAS). Among targets under investigation, cholinergic receptors have received much attention with several nicotinic agonists (α7 and α4β2) actively in clinical trials for the treatment of AD, CIAS and attention deficit hyperactivity disorder (ADHD). Both glutamatergic and serotonergic (5-HT) agonists and antagonists have profound effects on neurotransmission and improve cognitive function in preclinical experiments with animals; some of these compounds are now in proof-of-concept studies in humans. Several histamine H3 receptor antagonists are in clinical development not only for cognitive enhancement, but also for the treatment of narcolepsy and cognitive deficits due to sleep deprivation because of their expression in brain sleep centers. Compounds that dampen inhibitory tone (e.g., GABA_A α5 inverse agonists) or elevate excitatory tone (e.g., glycine transporter inhibitors) offer novel approaches for treating diseases such as schizophrenia, AD and Down syndrome. In addition to cell surface receptors, intracellular drug targets such as the phosphodiesterases (PDEs) are known to impact signaling pathways that affect long-term memory formation and working memory. Overall, there is a genuine need to treat cognitive deficits associated with many neuropsychiatric conditions as well as an increasingly aging population.