硒对大鼠肝细胞凋亡和原癌基因c-myc、c-fos和c-jun表达的影响

目的　研究亚硒酸钠对大鼠肝细胞凋亡和原癌基因c myc、c fos和c jun表达的影响。方法　选用大鼠 ,每组 5只 ,用 5、10和 2 0 μmol/kg亚硒酸钠腹腔注射染毒。用末端标记法 (TUNEL)、流式细胞术检测细胞凋亡 ,用Northern斑点杂交方法研究原癌基因c myc、c fos和c jun的表达。结果　 5、10和 2 0 μmol/kg的亚硒酸钠染毒大鼠 ,不仅能诱导大鼠肝细胞凋亡 ,细胞凋亡率均较对照组 ( 2 2 2± 0 43 ) %显著增高 ,分别为 ( 3 72± 1 76) % (P <0 0 5 )、( 5 82± 1 42 ) % (P <0 0 1)和 ( 11 76± 1 87) % (P <0 0 1) ,而且还存在着明显的剂量 反应关系。 5、10和 2 0 μmol/kg的亚硒酸钠也能引起大鼠肝细胞c myc、c fos和c junmRNA明显增多 ,免疫组化分析可以检测到表达产物c Myc ,c Fos和c Jun蛋白 ,说明这 3种原癌基因表达增强。结论　一定剂量的亚硒酸钠可以诱导大鼠肝细胞凋亡和原癌基因c myc、c fos和c jun的表达增强     

低剂量的硒与镉联合作用对大鼠肝细胞DNA损伤的影响

目的 研究低剂量的亚硒酸钠和氯化镉联合作用对离体大鼠肝细胞DNA损伤的影响。方法 在2．185、4．375和8．750μmol／L的剂量条件下，亚硒酸钠和氯化镉联合作用，用单细胞凝胶电泳检测大鼠肝细胞DNA损伤。结果 2．185μmol／L的亚硒酸钠对2．185μmol／L的氯化镉引起的DNA损伤拮抗作用不明显，但有拮抗作用的趋势；4．375μmol／L亚硒酸钠对4．375μmol／L氯化镉引起的DNA损伤有明显的拈抗作用；而在8．750μmol／L的剂量下，亚硒酸钠与氯化镉存在相互拈抗的关系。结论 在硒和镉对大鼠肝细胞DNA损伤的联合作用中，较低剂量的亚硒酸钠对氯化镉没有拮抗作用，而在一定的剂量条件下，亚硒酸钠和氯化镉存在着相互拮抗的关系。     

蔓荆子黄素调控miR-378/PRRX1轴对胃癌生物学行为的影响

目的 探究蔓荆子黄素（Cas）对胃癌（GC）SGC-7901细胞增殖、侵袭、迁移及微小RNA-378(miR-378)/配对相关同源框1(PRRX1)轴的影响。方法 不同浓度Cas(0、5、10、20、40、80μmol/L)处理SGC-7901细胞48 h。将对数生长期的SGC-7901细胞分为：对照组、Cas低浓度组（10μmol/L）、Cas中浓度组（20μmol/L）、Cas高浓度组（40μmol/L）、Cas高浓度+miR-378小干扰RNA(siRNA)组（40μmol/L Cas+miR-378 siRNA）、Cas高浓度+PRRX1组（40μmol/L Cas+PRRX1过表达质粒）、Cas高浓度+miR-378 siRNA+PRRX1组（40μmol/L Cas+miR-378 siRNA+PRRX1过表达质粒）。MTT法、集落形成实验、流式细胞术、Transwell小室、划痕实验分别测定各组SGC-7901细胞活力、集落形成数量、凋亡率、侵袭及迁移能力;q PCR、蛋白印迹实验分别检测SGC-7901细胞miR-378、PRRX1蛋白表达水平;双萤光素酶实验验证mi...     

铝和苯并[a]芘联合染毒对神经细胞凋亡的影响北大核心CSCD

目的通过研究铝和苯并[a]芘联合染毒对大鼠神经元细胞凋亡的影响,探索两种毒物共同作用下致神经细胞凋亡的联合类型。方法采用新生的SD大鼠进行原代神经元细胞培养,培养5 d后,选取生长良好的同批次细胞,分为对照组(DMSO+S9+maltol)、染苯并[a]芘组(10μmol/L B[a]P)、染铝组(50μmol/L Al(mal)3)和联合作用组(50μmol/L Al(mal)3+10μmol/L B[a]P)4组,继续培养72 h后电子显微镜观察不同实验组神经细胞凋亡形态学变化,AO-EB荧光染色法观察神经元细胞凋亡,流式细胞仪Annexin V-PI双染法定量检测神经元凋亡率的变化。结果透射电子显微镜和荧光倒置显微镜下观察,可以看到染苯并[a]芘组、染铝组神经元细胞出现核膜皱缩,胞质致密,核染色质边集等细胞凋亡的经典形态学变化;联合作用组出现了细胞核裂解甚至凋亡小体。细胞凋亡率测定结果显示,与对照组相比,染苯并[a]芘组早期凋亡率、晚期凋亡率、总凋亡率差异均无统计学意义(P>0.05);染铝组,早期凋亡率、晚期凋亡率差异无统计学意义(P>0.05),总凋亡率差异有统计学意义(P<0.05);联合作用组各指标差异均有统计学意义(P<0.05)。结论铝和苯并[a]芘联合染毒对神经细胞凋亡具有联合作用,作用类型为协同作用。     

天麻素通过抑制细胞凋亡保护谷氨酸诱导的PC12细胞损伤

目的：研究天麻素（Gastrodin）对谷氨酸诱导的大鼠肾上腺嗜铬细胞瘤PC12细胞损伤的影响及可能机制。方法：以谷氨酸建立体外培养PC12细胞损伤模型并采用MTT比色法测定细胞存活率;AO/EB双染法经荧光显微镜观察细胞凋亡形态;采用流式细胞术检测细胞内活性氧含量以及Annexin V/PI染色后的细胞凋亡率;Western blot法检测细胞内Caspase-3蛋白表达。结果：天麻素可明显抑制谷氨酸诱导的PC12细胞凋亡,在0.1～10μmol/L剂量呈一定的量效关系;同时,天麻素可明显抑制谷氨酸引起的活性氧（ROS）的累积,降低谷氨酸诱导的活性Caspase-3蛋白的表达,降低PC12细胞的凋亡率,在0.1～10μmol/L剂量呈量效相关性。结论：在一定剂量范围内,天麻素对谷氨酸损伤的PC12细胞具有保护作用,其机制可能与减少ROS的生成,阻止氧化损伤的发生,抑制Caspase-3途径依赖的细胞凋亡相关。     

亚硒酸钠通过Keap1/Nrf2/ARE信号通路诱导人肺腺癌A549细胞凋亡

目的探究亚硒酸钠诱导人肺腺癌A549细胞凋亡的作用及可能的分子机制。方法 MTT法检测不同浓度亚硒酸钠对人肺腺癌A549细胞的抑制增殖率;倒置荧光显微镜观察细胞经亚硒酸钠处理后的形态学改变;流式细胞术检测细胞凋亡率;激光共聚焦观察活性氧荧光强度;多功能酶标仪测定氧化应激参数含量;Western blot检测Keap1/Nrf2/ARE信号通路蛋白的表达,以及Nrf2在细胞质和细胞核中的蛋白表达情况。结果亚硒酸钠剂量依赖性地抑制人肺腺癌A549细胞增殖,亚硒酸钠作用于人肺腺癌A549细胞24 h后,通过Hoechst 33342固定和流式细胞术分析,细胞凋亡率明显增加;亚硒酸钠可使活性氧和丙二醛水平明显升高,还原型谷胱甘肽和超氧化物歧化酶含量明显下降;同时亚硒酸钠能够下调Keap1、Nrf2和HO-1蛋白的表达,并且抑制Nrf2发生核位移。结论亚硒酸钠通过抑制人肺腺癌A549细胞增殖,诱导细胞凋亡,调节细胞内的氧化应激反应,调控Keap1/Nrf2/ARE抗氧化信号通路,从而促进肿瘤细胞凋亡。     

甲胺鸢尾素对H2O2损伤大鼠心肌细胞线粒体膜电位的影响

目的：研究新型化合物甲胺鸢尾素（methyl-amine irisolidone,MMI）预处理对H2O2损伤大鼠心肌细胞线粒体膜电位变化的影响。方法：甲胺鸢尾素预处理心肌细胞12 h后,采用H2O2诱导细胞氧化应激损伤,瑞氏-吉姆萨染色观察细胞形态,MTT法检测细胞存活率,流式细胞仪检测细胞线粒体膜电位变化。结果：与正常对照组比较,模型组心肌细胞存活率明显降低（P〈0.01）。MMI0.1μmol/L组与模型组相比细胞存活率相近（P〉0.05）,0.5、1、5、10μmol/L组显著增高（P〈0.05,P〈0.01）。与模型组比较,10μmol/L MMI减轻细胞形态变化,1、5、10μmol/L MMI显著抑制线粒体膜电位下降（P〈0.05,P〈0.01）。结论：甲胺鸢尾素可改善H2O2所致心肌细胞氧化应激损伤,其机制与稳定细胞膜、抑制线粒体膜电位下降,进而抑制心肌细胞凋亡有关。     

Cpd5对人卵巢癌SKOV3细胞增殖抑制和凋亡诱导作用

目的：观察Cpd5[2-（2-巯基乙醇）-3-甲基-1,4-萘醌,Compound5]对人卵巢癌SKOV3细胞增殖和凋亡的影响。方法：MTT法观察Cpd5对SK-OV3细胞的生长抑制作用,AnnexinV/PI双标记流式细胞术检测Cpd5对SKOV3细胞的凋亡诱导,Hoechst-33258染色荧光显微镜观察细胞凋亡形态。结果：5、10、20、30、40、50、60μmol/LCpd5处理SKOV3细胞48h,生长抑制率分别为2.77%、5.19%、10.61%、41.15%、71.37%、82.90%、89.81%。不同时间点（12、24、48和72h）检测,细胞生长抑制率存在剂量-时间依赖关系。流式细胞术检测,30μmol/LCpd5处理12、24和48h后,细胞凋亡率分别达9.25%、20.07%、56.16%;50μmol/L组的细胞凋亡率明显高于30μmol/L,且随时间增加而显著增加。用40、50和60μmol/LCpd5处理细胞12h,光镜观察即可见明显的形态学改变。荧光显微镜观察：AnnexinV-EGFP/PI双染显示,实验组比阴性对照组细胞凋亡率明显增多;Cpd5作用24、48h后,Hoechst-33258染色可见细胞出现明显的凋亡形态,20μmol/L浓度组凋亡率增加,30、40、50、60μmol/L各组细胞凋亡率显著增加。结论：Cpd5能以剂量-时间依赖的方式抑制SKOV3细胞增殖并诱导凋亡,是潜在的抗卵巢癌新化合物。     

亚硒酸钠诱导人急性早幼粒细胞白血病细胞株NB4细胞氧化应激和细胞凋亡

目的研究亚硒酸钠诱导NB4细胞的氧化应激和细胞凋亡。方法MTT比色法检测亚硒酸钠对NB4细胞的生长抑制;用形态学、DNA琼脂糖电泳和流式细胞术研究亚硒酸钠诱导NB4细胞凋亡的作用;用化学发光法和比色法研究亚硒酸钠对NB4细胞内氧化应激的影响。 结果亚硒酸钠可以时间和剂量依赖性地抑制NB4细胞生长和诱导NB4细胞凋亡。亚硒酸钠(≥5 μmol·L^-1)提高了NB4细胞内ROS水平,同时伴有细胞内还原型谷胱甘肽含量下降,而抗氧化剂NAC可抑制亚硒酸钠诱导的NB4细胞氧化应激和细胞凋亡。结论亚硒酸钠诱导NB4细胞氧化应激可能是其诱导NB4细胞凋亡的重要机理。     

H2O2预处理对大鼠心肌细胞缺氧/复氧损伤的影响

摘要 目的 探讨H2O2预处理对大鼠心肌细胞缺氧/复氧引起H9C2细胞凋亡的影响。方法 大鼠心肌细胞系（H9C2）经体外培养扩增,使用对数生长期细胞做实验处理。分别使用0、5、10、20、50、100μmol/L浓度 H2O2处理H9C2细胞,流式细胞仪（FCM）、MTT法和免疫印迹法（Western blot法）检测不同浓度H2O2处理对H9C2细胞凋亡率、增殖活性和STAT3磷酸化水平的影响,以确定最佳H2O2预处理浓度。观察低浓度H2O2预处理对缺氧/复氧所引起细胞损伤的影响,细胞随机分为4组,①对照组：常规培养;②缺氧／复氧组：预先在 5％CO2,95%N2培养箱中培养 120min,复氧30min ;③H2O2预处理组：给予H2O2处理90min换液,24h后缺氧/复氧处理;④AG490+ H202组：在H2O2预处理前10min给予10μmol/L AG490处理,其余处理同H2O2预处理组。AnnexinV-FITC/PI双染法及流式细胞仪（FCM）检测细胞凋亡,MTT法检测H9C2细胞增殖活性,免疫印迹法（Western blot法）检测STAT3磷酸化水平。结果20μmol/L组的STAT3磷酸化水平明显高于其他各浓度组（P<0.01）,心肌细胞凋亡率亦明显低于50μmol/L及100μmol/L浓度组（P<0.01）,而其心肌细胞增殖活性及细胞凋亡率与5μmol/L及10μmol/L浓度组无明显差异; H2O2预处理能降低大鼠心肌细胞凋亡率（P<0.05）,增加心肌细胞活性（P<0.05）上调P- STAT3水平（P<0.05）。缺氧损伤后给予AG490处理可使H2O2预处理保护作用消失。结论20μmol/L H2O2预处理对心肌细胞缺氧/复氧损伤具有适应性保护作用。可能机制是激活 JAK2-STAT3通路发挥抑制细胞凋亡作用。     

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.     

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.     

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.