微流控芯片及其在肿瘤研究中的应用进展

微流控芯片具有微型化、集成化、效率高、试剂消耗量小等特点,在细胞水平药物筛选及仿生器官方面的研究取得了一系列成果。本文着重介绍了微流控芯片的加工材料和制作技术,以及在肿瘤研究领域的应用和最新研究进展,并提出了微流控芯片领域存在的问题及解决问题的方法。     

用于肿瘤细胞三维培养的微流控芯片构建及培养条件研究

目的设计制作用于肿瘤细胞三维培养的微流控芯片,以海藻酸钙凝胶为细胞支架,基于芯片研究适合的凝胶形成、溶解条件,利用该条件长期培养SMMC-7721细胞并检测其活力。方法利用CDR软件、软光刻技术、模塑法、等离子键合法等设计并制作出微流控芯片并验证其适用性;在芯片中三维培养肝肿瘤细胞SMMC-7721,观察细胞形态,以IPP软件辅助计算细胞在72h的存活率。结果设计制作的微流控芯片适合于肿瘤细胞三维培养;芯片中培养的肝癌细胞72 h内生长状态良好,存活率达(96.1±4.5)%,细胞堆积排列紧密,出现肿瘤细胞聚集体(tumor cell spheroid,TCS)。结论该文设计构建用于肿瘤细胞三维培养的微流控芯片,并用于肝癌细胞SMMC-7721培养,细胞生长状态良好,且细胞生长与聚集状态表现出一定特殊性。利用该芯片进行肿瘤细胞三维培养,可以为肿瘤细胞生长状态研究及抗肿瘤药物筛选提供更有利的方式。     

药物研发中的微流控:从靶标筛选到产品生命周期管理

微流控技术因具有分析微型化和实验通量化的特点,在药物研发领域已引起广泛关注。微流控系统已成为药物研发应用中具有潜在价值的工具。本文综述了药物研发中微流控芯片的近年来进展,着重介绍其在药物研发过程中不同阶段,包括靶标筛选、先导化合物确定、临床前研究、临床试验、化学合成及产品管理诸方面的实际应用。     

基于微流控芯片的体外肠道吸收模型构建及其应用进展

肠道是口服药物吸收的主要部位，肠道的上皮细胞上含有绒毛和微绒毛，它们通过增加表面积等因素促进分泌、细胞黏附和吸收。传统的二维/三维（2D/3D）细胞培养模型、动物模型在研究药物吸收方面发挥了重要作用，但是由于缺乏足够的人体药动学的可预测性或伦理问题等，其应用存在一定局限性。因此，以体外活细胞为基础，模拟人肠道的核心结构和关键功能是构建基于微流控芯片的肠道模型的研究重点。该模型是利用微加工技术制备出的模拟人体肠道的复杂微结构、微环境和生理功能的微流控芯片仿生系统。与2D细胞培养和动物实验相比，肠芯片模型能有效地模拟人体内环境，在药物筛选方面更具特异性。本综述概括了国内外肠芯片模型以及与肠道相关的多器官耦合芯片模型的研究进展，及其在疾病建模、药物吸收和转运方面的应用，并总结了当前肠芯片模拟肠道稳态和疾病面临的挑战，为进一步建立更可靠的体外肠芯片模型提供参考。     

药物筛选微流控芯片的构建及其在花蕊石抗肝癌研究中的应用

目的构建一种集成有微阀结构的双层PDMS-玻璃复合芯片,考察其适用性并研究花蕊石生品、制品水煎液对肝肿瘤细胞Hep G2凋亡坏死的影响。方法制作双层微流控芯片并考察微阀的性能,同时进行了Hep G2细胞的培养,死活试剂盒检测细胞活力。将预制备的不同浓度花蕊石生品、制品水煎液通入微流控芯片,分别作用于Hep G2细胞24、48和72 h,凋亡坏死试剂盒检测Hep G2细胞凋亡坏死率。结果芯片微阀性能良好,可灵活控制液阀的开关且未表现出明显的延迟现象。细胞在芯片中生长状态良好,培养72 h期间细胞存活率≥97%。药物作用细胞后,发生凋亡坏死的细胞数量随给药浓度的升高、给药时间的延长而增加,呈现出一定的时间与浓度依赖性,且制品水煎液的效果好于生品。结论实验证明了花蕊石生品、制品水煎液对肝肿瘤细胞Hep G2具有促凋亡作用,也进一步验证了本实验室设计的芯片在细胞研究和药物筛选方面的可行性,为微流控芯片在中药抗肿瘤新药开发领域中的推广与应用提供理论依据。     

基于微流控芯片分析气滞胃痛片中不同药效组分对炎症细胞GES-1的影响

目的 采用微流控芯片技术,从细胞凋亡坏死和炎症因子等角度研究气滞胃痛片及其不同药效组分对炎症细胞GES-1的影响,明确气滞胃痛片药效物质基础。方法 首先考查脂多糖（LPS）和不同药效组分对 GES-1 细胞存活的影响,筛选出造模剂量和最佳给药剂量,考察炎症因子的表达;同时构建了一种集成有微阀结构的高通量药物筛选芯片,分析炎症细胞GES-1凋亡和坏死情况,结合炎症因子表达和微流控芯片中的细胞死亡结果共同探讨气滞胃痛片及其不同药效组分对炎症细胞GES-1的影响。结果 根据抑制细胞存活的药物浓度梯度考察,得到脂多糖（LPS）IC₅₀值和最佳给药浓度以及最佳造模浓度,结合TNF-α 试剂盒获得各组炎症表达;与此同时对于微流控芯片的适用性进行考察,芯片中细胞生长状态良好,符合实验要求,可进行后续相关实验;通过微流控芯片以及 TNF-α 试剂盒考察气滞胃痛片中不同药效组分对炎症细胞GES-1的作用,发现给药24 h 后,甘草黄酮、延胡索生物碱等6种有效组分均能使促炎症因子明显下降,并且均有明显的抑制炎症细胞GES-1凋亡或坏死的作用。与模型组相比,细胞凋亡坏死率和炎症因子含量差异均有统计学意义（P<0.05）,其中甘草黄酮、延胡索生物碱的差异性最为明显（P<0.01）,药效显著。结论 设计、制作微流控炎症细胞芯片,初步明确了气滞胃痛片对炎症细胞GES-1的药效物质基础;为中药复方中药效组分抗炎作用的快速筛选提供了一种高效低耗的方法。     

微流控芯片技术在药物分析领域的研究进展

目的:为微流控芯片技术在药物分析领域的研究与应用提供参考。方法:以"微芯片""微流控芯片""药物分析""紫外-可见光吸收检测""激光诱导荧光检测""化学发光检测""电导检测""安培检测""质谱检测""Microchip""Micro-fluidic chip""Medicine analysis""UV detection""Laser induced fluorescence detection""Chemiluminescence detection""Conductivity detection""Amperometric detect""Mass spectrometry"等为关键词,在中国知网、万方、维普、PubMed、ScienceDirect、Wiley Online Library、Web of Science等数据库中组合查询2000年1月-2019年3月发表的相关文献,对微流控芯片分析检测药物的相关研究进行分析和总结。结果与结论:共检索到相关文献121篇,其中有效文献40篇。微流控芯片检测作为一种先进的现代分析方法,近年来在药物分析领域取得了飞速的进展。微流控芯片具有独特的分析特点,可以与紫外-可见光吸收检测、激光诱导荧光检测、化学发光检测、电化学检测、质谱检测等不同的检测方法结合,在药物制剂主成分分析、手性药物分析、药动学分析、组织样本中的药物浓度分析、尿药浓度分析、血药浓度分析等方向展现了不同的分析检测优势,在科研与实际应用中均具有良好的前景。     

微流控芯片技术研究进展

微流控芯片分析技术近年来迅速发展,在生化分析中应用广泛。本文介绍了微流控芯片技术的加工方法、分离技术及检测技术,并从DNA检测、酶联免疫分和细胞信号通路研究等方面综述了其在生化分析中的应用,对其应用中的不足进行了分析,对其应用前景做了展望。     

毛细管电泳和微流控芯片在药物筛选中的应用

药物筛选是现代药物开发流程中测试和获取特定生理活性化合物的一个步骤.毛细管电泳技术由于具有样品消耗量小、速度快、柱效高以及所用溶液体系较接近生物体液组成等特点,已经成为一种非常具有潜力的药物及先导化合物的高效筛选工具.文中就毛细管电泳技术在药物筛选中的最新应用情况进行综述,具体从测定药物解离常数pKa值、药物脂水分配系数(logP)、药物与蛋白质的结合常数的测定以及手性药物筛选等方面进行论述,同时也探讨了微流控芯片技术在高通量药物筛选方面的最新研究进展.     

微流控芯片快速检测技术在铁皮石斛农药残留检测中的应用研究

目的 探究微流控芯片快速检测技术在铁皮石斛农药残留检测应用的可行性。方法 采用有机溶剂提取样本,并用QuEChERS dispersive SPE净化,通过离心式微流控芯片高通量筛选不同厂家的农药残留检测用酶,考察阴性样品基质干扰的情况,采用不同水平加标的方式,考察微流控芯片农药残留检测方法的灵敏度、重复性、准确性,并和质谱法测定的样品结果进行比较。结果 快速检测方法显著差异率：0.25;灵敏度：99.6%;特异性：96.4%;假阴性率：0.39%;假阳性率：3.57%;准确度：98.81%。采用微流控芯片农药残留快速检测方法与质谱法2种方法检测40批次样品,测定结果表明,2种测定方法结果高度一致,检测结果符合率均为100%。结论 开发的微流控农药残留芯片快速检测技术可用于铁皮石斛农药残留的检测,可以满足基层非专业人员针对大批量农药残留的筛查需求。     

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.     

Pharmacological treatment of COVID-19: an opinion paper

The precocity and efficacy of the vaccines developed so far against COVID-19 has been the most significant and saving advance against the pandemic. The development of vaccines has not prevented, during the whole period of the pandemic, the constant search for therapeutic medicines, both among existing drugs with different indications and in the development of new drugs. The Scientific Committee of the COVID-19 of the Illustrious College of Physicians of Madrid wanted to offer an early, simplified and critical approach to these new drugs, to new developments in immunotherapy and to what has been learned from the immune response modulators already known and which have proven effective against the virus, in order to help understand the current situation.     

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.     

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.