甘精胰岛素抗胰岛β细胞脂性凋亡的信号转导通路

目的研究甘精胰岛素(glargine)是否通过胰岛素特有的信号途径(磷脂酰肌醇3激酶—蛋白激酶B通路,PI3K-PKB/Akt)激活核因子κB(NF-κB),揭示其完整的抗胰岛β细胞凋亡途径,寻找保护β细胞药物靶点。方法Western blot检测glargine和普通胰岛素(RI)抗β细胞脂性凋亡时PKB/Akt活性的变化,以及PI3K抑制剂wortmmanin对NF-κB活性的影响;流式细胞仪测定wortmmanin对glargine和RI抗凋亡作用的影响。结果Glargine或者RI增加PKB/Akt活性,PI3K抑制剂wortmmanin可阻断它们的抗凋亡作用,从而证实glargine和RI的抗凋亡作用通过PI3K-PKB/Akt途径;wortmmanin可阻断glargine和RI诱导的NF-κB激活,证明它们抗脂性凋亡时PI3K-PKB/Akt通路处于NF-κB的上游。结论glargine和RI激活PI3K-PKB/Akt通路导致NF-κB途径激活,从而发挥抗凋亡作用。     

厚朴提取物、厚朴酚及和厚朴酚的抗炎作用及其机制研究进展

厚朴系木兰科植物厚朴Magnolia officinalis或凹叶厚朴M.officinalis var.biloba的干燥干皮、根皮及枝皮,具有燥湿消痰、下气除满的功效。厚朴提取物、厚朴酚及和厚朴酚是厚朴抗炎的主要活性成分,其抗炎机制有：阻滞磷脂酰肌醇-3激酶/蛋白激酶B（PI3K/Akt）、细胞外调节蛋白激酶/丝裂原活化蛋白激酶（ERK/MAPK）和把关受体-2/丝裂原活化蛋白激酶（TLR/MAPK）信号通路,抑制炎性细胞因子表达;还可通过直接抑制诱导型一氧化氮合酶（iNOS）、环氧化酶-2（COX-2）、5-脂氧化酶（5-LO）的酶活性,阻滞炎症介质一氧化氮、前列腺素（PGs）、白三烯（LTs）的合成和释放,以及抑制组织胺释放等,产生广谱的抗炎作用。厚朴提取物、厚朴酚及和厚朴酚的抗氧化活性也是其抗炎作用的主要机制之一。厚朴为常用的非毒性中药,可以把研究重点放在防治慢性的或退行性疾病的微炎症方面。     

转化生长因子β1对人脐静脉血管内皮细胞活性的影响

目的观察转化生长因子β1（TGF—β1）刺激增生性瘢痕成纤维细胞上清液对人脐静脉血管内皮细胞（HUVEC）增殖和趋化能力的影响。方法以增生性瘢痕成纤维细胞为实验组,正常皮肤为对照组,添加TGF—β1（10μg/L）为TGF—β1组,向TGF—β1组内添加丝氨酸-苏氨酸蛋白激酶（Akt）阻断剂Tricirlbine（5Ixmol／L）为Akt阻断剂组,二者皆未添加的瘢痕成纤维细胞为非干扰组,利用各组细胞上清液（条件培养基）培养HUVEC,使用四甲基偶氮唑盐（MTT）法观察各条件培养基对HUVEC增殖的影响,并利用细胞划痕实验和Transwell细胞迁移实验对HUVEC的趋化情况进行评估。结果对照组吸光度值为0．91±0．34,非干扰组1．75±0．15,TGF—B,组3．26±0．58,Akt阻断剂组1．14±0．81。非干扰组吸光度值与对照组和TGF-β1组比较,差异均有统计学意义（均P〈0．01）;而Akt阻断剂组成纤维细胞增殖吸光度值明显低于TGF—β1组（P〈0．01）。细胞划痕24h后,TGF—β1组促使HUVEC划痕的闭合率较非干扰组高（P〈0．01）;而Akt阻断剂组则明显减弱HUVEC划痕闭合的速度（P〈0．01）。Transwell细胞迁移实验中,TGF—β1组迁移的细胞较非干扰组多[TGF—β1组细胞迁移数目：（1718±15）个,非干扰组：（829±2）个,P〈0．01];而Akt阻断剂组阳性细胞迁移率又明显降低[Akt阻断剂组：（935±11）个,P〈0．01]。对照组血管内皮生长因子（VEGF）（65．2±0．3）ng／L,非干扰组（87．0±0．5）ng／L,TGF—β1组（132．7±0．4）ng／L,Akt阻断剂组（70．5±0．6）ng／L。非干扰组与对照组、TGF-β1组与非干扰组、Akt阻断剂组与TGF—β1组比较,差异均有统计学意义（均P〈0．01）。结论TGF—β1可以通过刺激增生性瘢痕成纤维细胞旁分泌VEGF等细胞因子对血管内皮细胞增殖、趋化等能力产生影响。     

高盐对血管内皮生长因子受体-3+巨噬细胞表型及功能的影响

目的 观察高盐对血管内皮生长因子受体（VEGFR）-3+巨噬细胞表型、淋巴管内皮细胞特性及功能的影响。方法 利用流式分选将小鼠RAW264.7巨噬细胞中VEGFR-3+亚群分选出来，分为Control组、低盐组（LS组，20mmol/L NaCl）和高盐组（HS组，40 mmol/L NaCl）。利用CCK-8法观察不同组VEGFR-3+细胞活性；Real-time PCR检测NaCl干预后VEGFR-3+巨噬细胞表型变化及淋巴管内皮细胞标志物mRNA表达水平；Transwell实验检测各组细胞的迁移功能，流式细胞术检测不同组细胞的吞噬能力。结果 与Control组相比，高盐干预可以使VEGFR-3+巨噬细 胞的白细胞介素（IL）-1β、肿瘤坏死因子（TNF）-α、CC类趋化因子配体（CCL）2、血管内皮生长因子（VEGF）-C及张力应答性增强子结合蛋白（TonEBP）mRNA表达水平上调（P＜0.05）；HS组在高盐干预24、48 h后细胞活性均显著低于LS组（P＜0.05）；同时，HS组细胞迁移能力及细胞的吞噬能力与Control组相比显著增强，差异均有统计学意义（P＜0.05）。结论 高盐可使VEGFR-3+巨噬细胞向M1型巨噬细胞偏移并表现出促淋巴管生成的特性，其迁移及吞噬能力显著增强，为进一步研究该亚群与淋巴管生成及心血管疾病的关系提供了依据。     

Akt抑制剂的研究进展

Akt是一类丝/苏氨酸蛋白激酶,是磷脂酰肌醇-3激酶(PI3K)的下游靶点,通过磷酸化其下游一系列底物来促进细胞生长、代谢、凋亡、血管新生等重要的细胞生物学过程,其在多种恶性肿瘤中都存在过度表达或活性失调的现象。研究表明,通过抑制Akt的活性能有效地遏制肿瘤的生长,因此研发新型的Akt抑制剂已经成为抗肿瘤药物开发的热点之一。Akt抑制剂主要分为PH结构域抑制剂、ATP竞争性抑制剂、变构抑制剂等,目前已有多个Akt的抑制剂进入临床研究。本文综述了不同类型的Akt抑制剂的Akt抑制活性及特点,为开发新型抗肿瘤药物提供参考。     

木犀草素抗肿瘤血管生成作用机制的研究进展

血管生成是恶性肿瘤的标志之一,涉及血管内皮细胞的增殖、迁移和细胞外基质分解等多种途径。血管内皮生长因子（VEGF）靶向抗肿瘤血管生成是临床肿瘤治疗的有效方法。木犀草素为黄酮类化合物,具有抗肿瘤活性,可通过抑制VEGF及相关信号通路、抑制磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路、抑制血管新生分子、抑制孕激素活性、靶向刺激Wnt信号通路、阻断生长停滞特异性蛋白6(Gas6)/受体酪氨酸激酶（Axl）信号通路而发挥抗肿瘤的血管生成作用。归纳了木犀草素抗肿瘤血管生成的作用机制,以期为木犀草素的临床应用提供参考。     

药物干预蛋白激酶B靶点改善糖代谢的研究进展

蛋白激酶B（PKB）属于丝氨酸／苏氨酸蛋白激酶家族成员，可被多种生长因子和细胞因子激活。PKB的表达及活性受损，可致胰岛素受体后信号转导缺陷，改变胰岛素生物学效应，进而形成胰岛素抵抗。研发以PKB为靶点的药物，干预PKB的表达或活性，可为糖代谢相关疾病的防治提供重要线索。     

黄芪注射液对肺结核大鼠免疫功能、自噬和凋亡的影响

目的 研究黄芪注射液对肺结核大鼠免疫功能、自噬和凋亡的影响及机制。方法 将SD大鼠分成对照组、模型组（注射结核杆菌）、实验组(注射结核杆菌,8 mL·kg^-1黄芪注射液处理)、实验+LY294002组[注射结核杆菌,8 mL·kg^-1黄芪注射液和磷脂酰肌醇-3激酶蛋白激酶B（PI3K/Akt）信号抑制药LY294002处理]。以蛋白质印迹（Western blot）法检测肺组织中磷脂酰肌醇-3激酶（PI3K）、蛋白激酶B（Akt）、磷酸化Akt（p-Akt）、Beclin1蛋白表达,用酶联免疫吸附（ELISA）法检测血清中炎症因子肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、白细胞介素-6（IL-6）水平,以流式细胞术检测T细胞亚群,以脱氧核糖核苷酸末端转移酶介导的缺口末端标记法（TUNEL）检测细胞凋亡。结果 对照组、模型组、实验组、实验+LY294002组PI3K蛋白表达水平为0.59±0.05,0.23±0.03,0.46±0.05和0.33±0.02,p-Akt/Akt表达水平为0.40±0.04,0.22±0.02...     

黄芪注射液对肺结核大鼠免疫功能、自噬和凋亡的影响

目的 研究黄芪注射液对肺结核大鼠免疫功能、自噬和凋亡的影响及机制。方法 将SD大鼠分成对照组、模型组（注射结核杆菌）、实验组(注射结核杆菌,8 mL·kg^-1黄芪注射液处理)、实验+LY294002组[注射结核杆菌,8 mL·kg^-1黄芪注射液和磷脂酰肌醇-3激酶蛋白激酶B（PI3K/Akt）信号抑制药LY294002处理]。以蛋白质印迹（Western blot）法检测肺组织中磷脂酰肌醇-3激酶（PI3K）、蛋白激酶B（Akt）、磷酸化Akt（p-Akt）、Beclin1蛋白表达,用酶联免疫吸附（ELISA）法检测血清中炎症因子肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、白细胞介素-6（IL-6）水平,以流式细胞术检测T细胞亚群,以脱氧核糖核苷酸末端转移酶介导的缺口末端标记法（TUNEL）检测细胞凋亡。结果 对照组、模型组、实验组、实验+LY294002组PI3K蛋白表达水平为0.59±0.05,0.23±0.03,0.46±0.05和0.33±0.02,p-Akt/Akt表达水平为0.40±0.04,0.22±0.02...     

糖尿病大鼠蛋白激酶B信号传导系统表达的实验研究

目的探讨在糖尿病大鼠胰腺中磷酸化蛋白激酶B（PKB）表达的变化及胰腺β细胞增殖及凋亡的变化。方法向糖尿病大鼠腹腔内注射链脲佐菌素（STZ）,实验中监测血糖,用免疫组化染色法观察胰腺中胰岛素、胰离血糖素、磷酸化PKB的表达,以及胰腺β细胞的增生及凋亡情况。结果糖尿病大鼠胰腺中胰岛素（46．62±4．35）％、磷酸化PKB（22．21±2．73）％的表达水平较对照组[（55．54±4．39）％与（27．37±2．93）％]均降低,而胰高血糖素（22．08±2．72）％的表达水平较对照组（17．27±1．92）％升高,糖尿病组β细胞凋亡率（31．17±2．19）％明显高于对照组（18．90±2．09％）（均P〈0．01）,2组大鼠胰腺β细胞均无明显增生。结论磷酸化PKB在糖尿病大鼠胰腺中的表达水平下降,PKB介导的信号传导系统能促进胰岛素的分泌,减少胰高血糖素的分泌,抑制胰腺β细胞的凋亡。     

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.