雷公藤内酯醇与人乳腺癌细胞大分子DNA作用膜式的探讨

采用药物与ＤＮＡ作用的体外检测方法，以ＤＮＡ嵌入剂表阿霉素（ＥＤＲ）作对照，从分子药理水平研究雷公藤内酯醇（ＴＬ）对乳腺癌细胞ＤＮＡ的作用模式。实验发现ＴＬ在１２０μｇ／ｍｌ浓度，３７℃作用２４ｈ可与ＤＮＡ模板结合，同ＥＤＲ呈嵌合效应，但嵌入ＤＡＷ尚不牢固，易发生解离；ＥＤＲ在较低浓度（５μｇ／ｍｌ）作用３０ｍｉｎ即可显示明显的嵌合效率且不是可逆的。说明ＴＬ对乳腺癌细胞ＤＮＡ的作用模式为嵌合逆行模     

三尖杉酯碱和高三尖杉酯碱与白血病细胞DNA的结合实验

本文用琼脂糖电泳观察国产三尖杉酯碱(H)及高三尖杉酯碱(H.H)与白血病细胞DNA的作用模式。实验发现H及H.H在高浓度(800μg/ml),37℃作用16h,可与DNA发生嵌合效应,48h这种现象更为明显,同时该过程也伴有一定程度的DNA断链效应。阿霉素在低浓度(50μg/ml)作用30min即可显示嵌合效应。本实验表明H.H对白血病细胞DNA的嵌合能力较H有所增强,但两者所见嵌合效应与阿霉素比较相距甚远。     

5种抗肿瘤抗生素对人白血病细胞DNA嵌合能力的观察

采用琼脂糖电泳法观察５种抗癌药对白血病细胞ＤＮＡ的嵌合能力。结果发现表阿霉素同进口阿霉素在５０μｇ／ｍｌ浓度时可与ＤＮＡ发生嵌合效应；国产阿霉素在较高浓度（１００μｇ／ｍｌ）和柔红霉素需高浓度（２００μｇ／ｍｌ）才显示嵌合效应；而米托蒽醌在较低浓度（２５μｇ／ｍｌ）即可发生此作用。本实验体现了作用机制类同的抗肿瘤药物对ＤＮＡ嵌合能力的差异。     

大蒜素对人视网膜母细胞瘤Rb细胞中Bcl-2表达影响的研究

目的探讨大蒜素对人视网膜母细胞瘤Rb细胞的抑制作用，及其对Bcl-2表达的影响。方法人视网膜母细胞瘤Rb细胞株，经复苏，孵育，传代分为10个实验组。分别用不同浓度大蒜素（5μg／ml、10μg／ml、15μg／ml、20μg／ml、25μg／ml、30μg／ml）、顺铂（3μg／ml）及大蒜素（15μg／ml）和顺铂（3μg／ml）联合处理人视网膜母细胞瘤Rb细胞，在不同时间（24h、48h、72h）观察Rb细胞的形态，并应用免疫细胞化学SP法检测不同处理条件下和各时间点Bcl-2的表达情况。结果人视网膜母细胞瘤Rb细胞在不同浓度大蒜素、顺铂以及大蒜素和顺铂联合作用下，细胞生长增殖受到抑制，与大蒜素空白液、培养液对照组的Rb细胞相比形态学变化明显。不同浓度的大蒜素均可以下调人视网膜母细胞瘤Rb细胞中Bcl-2的表达，大蒜素作用组与对照组两两比较，差异均有统计学意义（P〈0．05）。大蒜素下调Bcl-2的表达具有时间和剂量依赖性，随着大蒜素作用时间的延长，Bcl-2的表达依次下降。大蒜素（15μg／ml）和顺铂（3μg／ml）联合用药可增强对人视网膜母细胞瘤Rb细胞的生长抑制，与大蒜素（15μg／ml）、顺铂（3μg／ml）单独处理Rb细胞比较，下调Bcl-2的表达作用具有显著性差异（P〈0．05）。结论人视网膜母细胞瘤Rb细胞中存在与细胞凋亡有关的Bcl-2蛋白低表达。大蒜素可抑制人视网膜母细胞瘤Rb细胞的生长，且抑制作用随药物作用浓度的增高和作用时间的延长而增强，具有时间和浓度依赖性。采用免疫细胞化学的方法检测人视网膜母细胞瘤Rb细胞中Bcl-2的表达，发现大蒜素作用组Bcl-2的表达较空白和溶剂对照组低，提示大蒜素抑制人视网膜母细胞瘤Rb细胞的生长与Bcl-2介导的细胞凋亡有关，大赫素和顺铂联合用药作用强于单独用药，可进一步提高药物对肿瘤细胞的生长抑制。     

环孢菌素A联合干扰素对白血病细胞株的多药耐药逆转的实验研究

目的：观察联合逆转剂对白血病细胞株多药耐药（MDR）的逆转作用，提高化疗的敏感性，方法：用环孢菌素A（CsA)和干扰素－α（INF-α）单独或联合逆转多药耐药细胞株K562／A02对柔红霉素（DNR）的耐药，药敏试验采用MTT法，同时用流式细胞仪检测逆转前后P糖蛋白（PgP)表达的变化及细胞内DNR浓度分布情况。结果：DNR对K562／A02及K562／S细胞的半数细胞抑制剂量（IC50）分别为7．3μg/ml和0．2μg/ml，CsA联合INF－α后明显增强DNR对K562／A02的细胞毒作用，IC60由7．3μg/ml降低到0．7μg/ml，而对K562／S细胞无影响，且逆转后细胞内DNR浓度明显增加，PgP表达无明显变化，结论：CsA和IFN－α联合能使白血病细胞株K562／A02对DNA的敏感性增加，具有逆转多药耐药的作用。     

三氧化二砷与顺铂联用对人乳腺癌细胞杀伤作用的研究

目的探讨三氧化二砷与顺铂联合应用对人乳腺癌细胞的杀伤作用。方法将三氧化二砷与顺铂单独及联合应用于体外培养的乳腺癌MCF-7细胞,采用MTT法测定细胞抑制率,于透射电镜下观察细胞超微结构变化;用流式细胞仪观察细胞凋亡情况。结果1μg/ml三氧化二砷与1μg/ml顺铂联合应用于乳腺癌MCF-7细胞48h后,细胞抑制率为27.01%,明显高于单用三氧化二砷（1μg/ml）时的12.12%及单用顺铂（1μg/ml）时的10.93%。细胞超微结构及流式细胞仪观察均显示,联合应用比单用促凋亡效应更显著（P〈0.01）。结论三氧化二砷与顺铂联合应用,可高效杀伤人乳腺癌细胞,此协同作用主要是通过诱导肿瘤细胞凋亡来实现的。     

MTT分析法检测沙立度胺对乳腺癌细胞增殖的作用

目的观察沙立度胺对乳腺癌细胞MCF-7和MDA—MB-231增殖的抑制作用。方法设置溶剂对照及沙立度胺不同浓度组、时间组，采用四甲基偶氮唑蓝（MTT）法检测各组细胞存活和生长情况，分析量效关系、时效关系。结果①在10—200μg／ml浓度范围，沙立度胺对MCF-7和MDA—MB-231细胞增殖均有抑制作用。在药物作用48h后各组的细胞抑制率均达到高峰，48h时又以100μg／ml浓度组抑制率最高。结论在一定浓度范围，沙立度胺对MCF-7和MDA—MB-231细胞增殖具有抑制作用。     

游离钙信号通路介导西兰花中异硫氰酸盐诱导细胞凋亡

目的：探讨了西兰花中异硫氰酸盐（ITCS）对体外培养的肿瘤细胞的抑制特点，并初步探讨了游离钙、活性氧水平和线粒体膜电位的变化与西兰花中ITCS诱导肿瘤细胞凋广的关系。方法：MTT法、SRB法、流式细胞仪技术结合激光共聚焦显微镜技术分析诱导凋亡作用。结果：在体外研究中，西兰花中ITCS可抑制不同来源的肿瘤细胞的生长，如人胃腺癌细胞（SGC-7901）和人肝癌细胞（HepG-2）。MTT法测得其在体外的半数抑制浓度IC50约为22．40μg／ml和31．429μg／ml。从SRB结果可看出，低浓度的H13通过抑制肿瘤细胞生长而起抗癌作用，SGC-7901和HepG-2的GL。分别为7．252μg／ml和15．824μg／ml，而高浓度的ITCS则主要通过杀死癌细胞来起到抗癌作用，对上述两种细胞的TGl50分别为382．825μg／ml和783．610μg／ml。西兰花中ITCS能够诱导细胞凋亡，并且随着ITCS浓度的加大，凋亡细胞的比例逐渐增加。采用流式细胞仪进行细胞DNA含量测定，可见在G1峰前出现一个“G1亚峰”，凋亡细胞的DNA含量随ITCS浓度的升高而增多，即呈一定的剂量依赖性。同时分析了对细胞周期的影响，结果显示ITCS可使G1期细胞的比例降低，S期细胞的比例增加，提示西兰花中ITCS阻断肿瘤细胞由S期向G2期的移行。从以上三方面的阳性结果可以认为药物作用后，细胞已发生凋亡。不同浓度的西兰花中ITCS作用于SGC-7901和HepG-2细胞24h后，发现ITCS能提高肿瘤细胞内钙离子浓度，且呈一定的剂量依赖关系。通过流式细胞仪分析，西兰花中ITCS能明显提高肿瘤细胞内活性氧的水平，并且能能明显降低肿瘤细胞内线粒体膜电位。结论：西兰花中ITCS对SGC-7901和HepG-2细胞具有明显的抗肿瘤活性。其作用机制可能与提高肿瘤细胞内钙离子浓度和增加肿瘤细胞内活性氧的产生，引起线粒体膜电位下降，进而启动细胞凋亡机制诱导肿瘤细胞凋亡有关。     

阿霉素和阿克拉霉素A对HeLa细胞DNA的嵌合作用

本文用琼脂糖电泳观察国产阿霉素和阿克拉霉素A对HeLa细胞DNA的嵌合作用.DNA经琼脂糖电泳及溴乙锭染色后,紫外荧光摄影,以药物与双链DNA发生嵌合作用时DNA的溴乙锭染色性丧失为嵌合指标.实验发现阿霉素在低浓度时(25μg/ml)即可与HeLa细胞DNA发生嵌合作用,而阿克拉霉素A仅在高浓度(400～800μg/ml)才发生此作用.本实验实现了这两个蒽环类抗生素对DNA嵌合能力的差异.     

二烯丙基二硫对Raji细胞化疗增敏的研究

目的研究二烯丙基二硫（dimlyldisulfide,DADS）对长春新碱（VCR）作用于人淋巴瘤Raji细胞化疗增敏的作用及机制。方法DADS（0．625μg／ml、1．25μg／ml）及VCR（6．25,12．5,25．0μg／ml）单用及联合应用作用于Raji细胞,采用CCK-8（cellcountingkit）法检测各自的抑制率,应用金氏公式求q值。评价联合用药效果;DADS（0．625μg／ml）、VCR（6．25,12．5,25．0μg／ml）及联合应用作用于Raii细胞,采用流失细胞术检测细胞凋亡率,采用细胞免疫化学法检测细胞Bax及Bcl-2的表达情况。结果无毒剂量的DADSfo．6251Ag／ml、1．25μg／ml）分别与VCR联合应用,可提高VCR对Raji细胞增殖抑制率。DADS（0．625μg／m1）提高VCR（6．25,12．5,25．0μg/ml）诱导Raji细胞凋亡率（P〈0．05）。联合组Raji细胞Bcl-2蛋白表达水平比VCR单药组低（P〈0．05）;Bax蛋白表达水平在联合组Raji细胞VCR单药组高（P〈0．05）。结论DADS能增强VCR对Raji细胞增殖抑制和诱导凋亡作用．起化疗增敏作用：可能机制与其下调Bcl-2蛋白表达水平,上调Bax蛋白表达水平有关。     

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.

     

In vitro studies on sterically stabilized liposomes (SL) as enzyme carriers in organophosphorus (OP) antagonism

This study describes a new approach for organophosphorous (OP) antidotal treatment by encapsulating an OP hydrolyzing enzyme, OPA anhydrolase (OPAA), within sterically stabilized liposomes. The recombinant OPAA enzyme was derived from Alteromonas strain JD6. It has broad substrate specificity to a wide range of OP compounds: DFP and the nerve agents, soman and sarin. Liposomes encapsulating OPAA (SL)* were made by mechanical dispersion method. Hydrolysis of DFP by (SL)* was measured by following an increase of fluoride ion concentration using a fluoride ion selective electrode. OPAA entrapped in the carrier liposomes rapidly hydrolyze DFP, with the rate of DFP hydrolysis directly proportional to the amount of (SL)* added to the solution. Liposomal carriers containing no enzyme did not hydrolyze DFP. The reaction was linear and the rate of hydrolysis was first order in the substrate. This enzyme carrier system serves as a biodegradable protective environment for the recombinant OP-metabolizing enzyme, OPAA, resulting in prolongation of enzymatic concentration in the body. These studies suggest that the protection of OP intoxication can be strikingly enhanced by adding OPAA encapsulated within (SL)* to pralidoxime and atropine.     

Steroidal sapogenin contents in some domestic plants

In order to find out the values of the steroid resources for the future use. the compositions and contents of steroidal sapogenins from 13 domestic plants have been investigated. As a result,Dioscorea nipponica, D. quinqueloba andSmilax china were found to have large amount of diosgenin. And pennogenin inTrillium kamtschaticum andParis verticillata, yuccagenin inAllium fistulosum, hecogenin inAgave americana and neochlorogenin inSolanum nigum were appeared to be major steroidal sapogenins.     

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.